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Studies in Mycology 58 (2007)<br />
<strong>The</strong> <strong>genus</strong> <strong>Cladosporium</strong> <strong>and</strong> <strong>similar</strong><br />
<strong>dematiaceous</strong> hyphomycetes<br />
Pedro W. Crous, Uwe Braun, Konstanze Schubert<br />
<strong>and</strong> Johannes Z. Groenewald<br />
Centraalbureau voor Schimmelcultures,<br />
Utrecht, <strong>The</strong> Netherl<strong>and</strong>s<br />
An institute of the Royal Netherl<strong>and</strong>s Academy of Arts <strong>and</strong> Sciences
<strong>The</strong> <strong>genus</strong> <strong>Cladosporium</strong> <strong>and</strong> <strong>similar</strong> <strong>dematiaceous</strong> hyphomycetes<br />
Studies in Mycology 58, 2007
Studies in Mycology<br />
<strong>The</strong> Studies in Mycology is an international journal which publishes systematic monographs of filamentous fungi <strong>and</strong> yeasts, <strong>and</strong> in rare<br />
occasions the proceedings of special meetings related to all fields of mycology, biotechnology, ecology, molecular biology, pathology <strong>and</strong><br />
systematics. For instructions for authors see www.cbs.knaw.nl.<br />
Executive Editor<br />
Prof. dr Robert A. Samson, <strong>CBS</strong> Fungal Biodiversity Centre, P.O. Box 85167, 3508 AD Utrecht, <strong>The</strong> Netherl<strong>and</strong>s.<br />
E-mail: r.samson@cbs.knaw.nl<br />
Layout Editor<br />
Manon van den Hoeven-Verweij, <strong>CBS</strong> Fungal Biodiversity Centre, P.O. Box 85167, 3508 AD Utrecht, <strong>The</strong> Netherl<strong>and</strong>s.<br />
E-mail: m.verweij@cbs.knaw.nl<br />
Scientific Editors<br />
Prof. dr Uwe Braun, Martin-Luther-Universität, Institut für Biologie, Geobotanik und Botanischer Garten, Neuwerk 21,<br />
D-06099 Halle, Germany.<br />
E-mail: uwe.braun@botanik.uni-halle.de<br />
Prof. dr Pedro W. Crous, <strong>CBS</strong> Fungal Biodiversity Centre, P.O. Box 85167, 3508 AD Utrecht, <strong>The</strong> Netherl<strong>and</strong>s.<br />
E-mail: p.crous@cbs.knaw.nl<br />
Prof. dr David M. Geiser, Department of Plant Pathology, 121 Buckhout Laboratory, Pennsylvania State University, University Park, PA, U.S.A. 16802.<br />
E-mail: dgeiser@psu.edu<br />
Dr Lorelei L. Norvell, Pacific Northwest Mycology Service, 6720 NW Skyline Blvd, Portl<strong>and</strong>, OR, U.S.A. 97229-1309.<br />
E-mail: llnorvell@pnw-ms.com<br />
Dr Erast Parmasto, Institute of Zoology & Botany, 181 Riia Street, Tartu, Estonia EE-51014.<br />
E-mail: e.parmasto@zbi.ee<br />
Prof. dr Alan J.L. Philips, Faculdade de Ciências e Tecnologia, Universidade Nova de Lisboa, Quinta de Torre, 2829-516 Caparica, Portugal.<br />
E-mail: alp@mail.fct.unl.pt<br />
Dr Amy Y. Rossman, Rm 304, Bldg 011A, Systematic Botany & Mycology Laboratory, Beltsville, MD, U.S.A. 20705.<br />
E-mail: amy@nt.ars-grin.gov<br />
Dr Keith A. Seifert, Research Scientist / Biodiversity (Mycology <strong>and</strong> Botany), Agriculture & Agri-Food Canada, KW Neatby Bldg, 960 Carling Avenue,<br />
Ottawa, ON, Canada K1A OC6.<br />
E-mail: seifertk@agr.gc.ca<br />
Prof. dr Jeffrey K. Stone, Department of Botany & Plant Pathology, Cordley 2082, Oregon State University, Corvallis, OR, U.S.A. 97331-2902.<br />
E-mail: stonej@bcc.orst.edu<br />
Dr Richard C. Summerbell, 27 Hillcrest Park, Toronto, Ont. M4X 1E8, Canada.<br />
E-mail: summerbell@aol.com<br />
Copyright 2007 Centraalbureau voor Schimmelcultures, Fungal Biodiversity Centre, P.O. Box 85167, 3508 AD Utrecht, <strong>The</strong> Netherl<strong>and</strong>s.<br />
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Publication date: 31 August 2007<br />
Published <strong>and</strong> distributed by <strong>CBS</strong> Fungal Biodiversity Centre, P.O. Box 85167, 3508 AD Utrecht, <strong>The</strong> Netherl<strong>and</strong>s. Internet: www.cbs.knaw.nl.<br />
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ISBN/EAN : 978-90-70351-61-0<br />
Online ISSN : 1872-9797<br />
Print ISSN : 0166-0616<br />
Cover: Top: page 137, Fig. 30C <strong>Cladosporium</strong> pseudiridis (<strong>CBS</strong> 116463), conidiophores <strong>and</strong> conidia; page 40, Fig. 4D Toxicocladosporium irritans (type<br />
material), macroconidiophores; page 70, Fig. 11C Ramichloridium musae (<strong>CBS</strong> 365.36), sympodially proliferating conidiogenous cells, resulting in a long<br />
conidium-bearing rachis; Bottom: page 19, Fig. 8F Penidiella columbiana (type material), conidiophores with chains of disarticulating conidia; page 120, Fig.<br />
12C <strong>Cladosporium</strong> bruhnei (CPC 12211), conidial chains; page 126, Fig. 18L Davidiella tassiana (CPC 12181), asci in culture.
<strong>The</strong> <strong>genus</strong> <strong>Cladosporium</strong> <strong>and</strong> <strong>similar</strong><br />
<strong>dematiaceous</strong> hyphomycetes<br />
edited by<br />
Pedro W. Crous<br />
<strong>CBS</strong> Fungal Biodiversity Centre, P.O. Box 85167, 3508 AD Utrecht, <strong>The</strong> Netherl<strong>and</strong>s<br />
Uwe Braun<br />
Martin-Luther-Universität Institut für Biologie, Geobotanik und Botanischer Garten, Neuwerk 21, D-06099 Halle (Saale), Germany<br />
Konstanze Schubert<br />
Botanische Staatssammlung München, Menzinger Strasse 67, D-80638 München, Germany<br />
<strong>and</strong><br />
Johannes Z. Groenewald<br />
<strong>CBS</strong> Fungal Biodiversity Centre, P.O. Box 85167, 3508 AD Utrecht, <strong>The</strong> Netherl<strong>and</strong>s<br />
Centraalbureau voor Schimmelcultures,<br />
Utrecht, <strong>The</strong> Netherl<strong>and</strong>s<br />
An institute of the Royal Netherl<strong>and</strong>s Academy of Arts <strong>and</strong> Sciences
CONTENTS<br />
P.W. Crous, U. Braun <strong>and</strong> J.Z. Groenewald: Mycosphaerella is polyphyletic ...................................................................................................1<br />
P.W. Crous, U. Braun, K. Schubert <strong>and</strong> J.Z. Groenewald: Delimiting <strong>Cladosporium</strong> from morphologically <strong>similar</strong> genera ............................33<br />
M. Arzanlou, J.Z. Groenewald, W. Gams, U. Braun, H.-D Shin <strong>and</strong> P.W. Crous: Phylogenetic <strong>and</strong> morphotaxonomic revision of Ramichloridium<br />
<strong>and</strong> allied genera............................................................................................................................................................................................. 57<br />
K. Schubert, U. Braun, J.Z. Groenewald <strong>and</strong> P.W. Crous: <strong>Cladosporium</strong> leaf-blotch <strong>and</strong> stem rot of Paeonia spp. caused by Dichocladosporium<br />
chlorocephalum gen. nov.................................................................................................................................................................................95<br />
K. Schubert, J.Z. Groenewald, U. Braun, J. Dijksterhuis, M. Starink, C.F. Hill, P. Zalar, G.S. de Hoog <strong>and</strong> P.W. Crous: Biodiversity in<br />
the <strong>Cladosporium</strong> herbarum complex (Davidiellaceae, Capnodiales), with st<strong>and</strong>ardisation of methods for <strong>Cladosporium</strong> taxonomy<br />
<strong>and</strong> diagnostics............................................................................................................................................................................................. 105<br />
P. Zalar, G.S. de Hoog, H.-J. Schroers, P.W. Crous, J.Z. Groenewald <strong>and</strong> N. Gunde-Cimerman: Phylogeny <strong>and</strong> ecology of the<br />
ubiquitous saprobe <strong>Cladosporium</strong> sphaerospermum, with descriptions of seven new species from hypersaline environments<br />
...................................................................................................................................................................................................................... 157<br />
P.W. Crous, K. Schubert, U. Braun, G.S. de Hoog, A.D. Hocking, H.-D. Shin <strong>and</strong> J.Z. Groenewald: Opportunistic, humanpathogenic<br />
species in the Herpotrichiellaceae are phenotypically <strong>similar</strong> to saprobic or phytopathogenic species in the<br />
Venturiaceae ..............................................................................................................................................................................................185<br />
G.S. de Hoog, A.S. Nishikaku, G. Fern<strong>and</strong>ez Zeppenfeldt, C. Padín-González, E. Burger, H. Badali <strong>and</strong> A.H.G. Gerrits van den Ende:<br />
Molecular analysis <strong>and</strong> pathogenicity of the Cladophialophora carrionii complex, with the description of a novel species.......................... 219<br />
K.A. Seifert, S.J. Hughes, H. Boulay <strong>and</strong> G. Louis-Seize: Taxonomy, nomenclature <strong>and</strong> phylogeny of three cladosporiumlike<br />
hyphomycetes, Sorocybe resinae, Seifertia azalea <strong>and</strong> the Hormoconis anamorph of Amorphotheca resinae<br />
...................................................................................................................................................................................................................... 235
PREFACE<br />
DEDICATION<br />
This volume of the Studies in Mycology is dedicated to the memory<br />
of Gerardus Albertus de Vries (1919–2005), who spent his scientific<br />
career as mycologist at the <strong>CBS</strong>, where he was appointed as<br />
medical mycologist.<br />
of loyal friends <strong>and</strong> fellow mycologists dating back to the “Baarnera”<br />
of <strong>CBS</strong>. To the very end the <strong>CBS</strong> received a yearly Christmas<br />
card, which was always a water colour painting depicting some<br />
fascinating birds that he happened to be studying at the time. <strong>The</strong><br />
<strong>Cladosporium</strong> notebooks, annotations <strong>and</strong> live cultures are still at<br />
the <strong>CBS</strong>. It is thus with great joy that we dedicate this volume to<br />
Gerard de Vries, <strong>and</strong> build on his <strong>Cladosporium</strong> legacy, most of<br />
which is still sporulating, <strong>and</strong> will be available for scientific debate<br />
for generations to come.<br />
<strong>The</strong> <strong>genus</strong> <strong>Cladosporium</strong> <strong>and</strong> <strong>similar</strong> <strong>dematiaceous</strong><br />
hyphomycetes<br />
INTRODUCTION<br />
Fig. 1. Gerard de Vries studying <strong>Cladosporium</strong> spp. on different cultural growth<br />
media.<br />
On the 10 th of July 1952, Gerard de Vries graduated from the<br />
University of Utrecht, where he completed his Ph.D. on the topic<br />
“Contribution to the knowledge of the <strong>genus</strong> <strong>Cladosporium</strong> Link<br />
ex Fr.” under the guidance of the then director of the <strong>CBS</strong>, Prof.<br />
dr Johanna Westerdijk. At this time, his thesis (de Vries 1952)<br />
represented a benchmark <strong>and</strong> synthesis of our knowledge <strong>and</strong><br />
underst<strong>and</strong>ing of <strong>Cladosporium</strong> spp. studied in culture (Fig. 1).<br />
Gerard de Vries learned the basics of mycology by working during<br />
the 1930’s with mushroom taxonomy under the guidance of<br />
Abraham van Luyk. Via van Luyk he was also introduced to the<br />
Dutch Mycological Society, who organised excursions, also around<br />
Baarn, which is where the de Vries family lived. For the rest of his<br />
career, Gerard would retain his love for studying <strong>and</strong> collecting<br />
mushrooms. In 1948 he was employed at <strong>CBS</strong> under Johanna<br />
Westerdijk, <strong>and</strong> given the task of establishing <strong>and</strong> heading a<br />
new division called Medical Mycology. This he did, right up to his<br />
retirement in 1984. For several years de Vries played an important<br />
role in this discipline, <strong>and</strong> attended numerous medical congresses<br />
<strong>and</strong> workshops, <strong>and</strong> also published extensively on the topic. De<br />
Vries loved the outdoors <strong>and</strong> traveling, <strong>and</strong> combined this with his<br />
other passion, which was ornithology (van der Aa 2005). In 1952<br />
he graduated from the University of Utrecht, producing a revision<br />
of major species in the <strong>genus</strong> <strong>Cladosporium</strong> of importance to the<br />
medical, industrial <strong>and</strong> plant pathology disciplines. His book soon<br />
became highly popular, <strong>and</strong> with a strong dem<strong>and</strong> for additional<br />
copies, resulting in it being reprinted by J. Cramer. His contribution<br />
to <strong>Cladosporium</strong> was also recently acknowledged with the<br />
introduction of the cladosporium-like heat-tolerant <strong>genus</strong>, Devriesia<br />
by Seifert et al. (2004). Additional books published by de Vries dealt<br />
with mushrooms for amateur mycologists (1955) <strong>and</strong> a treatment of<br />
Hypogaea <strong>and</strong> truffels published in 1971 as part 3 of the “Fungi of<br />
the Netherl<strong>and</strong>s” (van der Aa 2005).<br />
Gerard de Vries was a well tempered, softly spoken man,<br />
who avoided conflicts, except when it dealt with scientific issues.<br />
He never married, <strong>and</strong> died a bachelor, surrounded by a circle<br />
Species of <strong>Cladosporium</strong> are common <strong>and</strong> widespread, <strong>and</strong> interact<br />
with humans in every phase of life, from growing behind your bed<br />
or bedroom cupboard <strong>and</strong> producing allergens, or growing on the<br />
bathroom ceiling, to the fruit decay happening in the fruit basket in<br />
the kitchen, to colonising the debris lying outside your house, <strong>and</strong><br />
even the plant diseases observed on some of the shrubs, trees, or<br />
flowers cultivated in your garden. However, scientists generally shy<br />
away from trying to identify these <strong>similar</strong> looking organisms, <strong>and</strong><br />
therefore the main aims of this volume were to:<br />
1) Establish st<strong>and</strong>ardised conditions <strong>and</strong> protocols for studying<br />
cladosporioid species <strong>and</strong> their teleomorphs in culture;<br />
2) Determine how they can morphologically be distinguished in<br />
culture, <strong>and</strong> highlight important diagnostic features;<br />
3) Circumscribe the <strong>genus</strong>, <strong>and</strong> delineate it from morphologically<br />
<strong>similar</strong> <strong>dematiaceous</strong> hyphomycetes;<br />
4) Determine which DNA gene loci are informative to accurately<br />
distinguish species of <strong>Cladosporium</strong>, <strong>and</strong> initiate a database of<br />
<strong>Cladosporium</strong> sequences that can in future be used to set up an<br />
online polyphasic identification key.<br />
Although <strong>Cladosporium</strong> is one of the largest <strong>and</strong> most<br />
heterogeneous genera of hyphomycetes, currently comprising<br />
more than 772 names (Dugan et al. 2004), only a mere fraction of<br />
these species are known from culture, <strong>and</strong> thus the real number<br />
of taxa that exist remains unknown. Species of <strong>Cladosporium</strong> are<br />
commonly encountered on plant <strong>and</strong> other kinds of debris, frequently<br />
colonising lesions of plant pathogenic fungi, <strong>and</strong> are also isolated<br />
from air, soil, food, paint, textiles <strong>and</strong> other organic matters (Ellis<br />
1971, 1976; Schubert 2005a), they are also common endophytes<br />
(Brown et al. 1998, El-Morsy 2000) as well as phylloplane fungi<br />
(Islam & Hasin 2000, de Jager et al. 2001, Inacio et al. 2002,<br />
Stohr & Dighton 2004, Levetin & Dorsey 2006). Some species of<br />
<strong>Cladosporium</strong> have a medical relevance in clinical laboratories,<br />
<strong>and</strong> also cause allergic lung mycoses (de Hoog et al. 2000). In<br />
spite of its obvious importance, species of <strong>Cladosporium</strong> are still<br />
poorly understood.<br />
Taxonomy of the anamorph<br />
<strong>The</strong> first binominal introduced for this group of fungi was that of<br />
Dematium herbarum Pers. (Persoon 1794) (Fig. 2). <strong>Cladosporium</strong><br />
herbarum (Pers.) Link was subsequently selected to serve as
lectotype for the <strong>genus</strong> by Clements & Shear (1931), a proposal<br />
which was accepted by de Vries (1952), Hughes (1958), <strong>and</strong> others<br />
(Prasil & de Hoog 1988). In subsequent years the number of taxa<br />
described in the <strong>genus</strong> grew rapidly, though the generic concept<br />
was rather vague. As a consequence, numerous morphologically<br />
<strong>similar</strong> <strong>dematiaceous</strong> hyphomycetes with catenulate conidia were<br />
incorrectly assigned to <strong>Cladosporium</strong>, making this one of the largest<br />
genera of hyphomycetes.<br />
In an attempt to circumscribe some of the more well-known taxa,<br />
de Vries (1952) published a revision of nine <strong>Cladosporium</strong> species in<br />
vivo <strong>and</strong> in vitro, <strong>and</strong> 13 additional taxa in an appendix. Ellis (1971,<br />
1976) described <strong>and</strong> illustrated 43 species, while Morgan-Jones<br />
<strong>and</strong> McKemy dealt with selected species in the series “Studies in<br />
the <strong>genus</strong> <strong>Cladosporium</strong> s. lat.” (Morgan-Jones & McKemy 1990,<br />
McKemy & Morgan-Jones 1990, 1991a–c). Other significant works<br />
that also treated <strong>Cladosporium</strong> species include Ho et al. (1999),<br />
<strong>and</strong> Zhang et al. (2003), though these authors still followed a<br />
wider generic concept. David (1997) followed the taxonomy of de<br />
Vries (1952), who first considered Heterosporium as a synonym<br />
of <strong>Cladosporium</strong>, <strong>and</strong> introduced the combination <strong>Cladosporium</strong><br />
subgen. Heterosporium. Subsequent to this publication, further<br />
monographic studies on the <strong>genus</strong> <strong>Cladosporium</strong> s. lat. were<br />
initiated by Braun <strong>and</strong> co-workers (Braun et al. 2003, 2006, Dugan<br />
et al. 2004, Schubert & Braun 2004, 2005a, b, 2006, 2007, Schubert<br />
2005a, b, Heuchert et al. 2005).<br />
Treatments of human pathogenic <strong>Cladosporium</strong> species<br />
(Masclaux et al. 1995, Untereiner 1997, Gerrits van den Ende & de<br />
Hoog 1999, Untereiner & Naveau 1999, Untereiner et al. 1999; de<br />
Hoog et al. 2000), concluded that they represent species belonging<br />
to the Herpotrichiellaceae (Capronia Sacc./Cladophialophora<br />
Borelli). Saprobic species, which appear morphologically <strong>similar</strong>,<br />
were found to belong to the Venturiaceae (Caproventuria U. Braun/<br />
Pseudocladosporium U. Braun; Braun et al. 2003, Schubert et al.<br />
2003, Beck et al. 2005) (see Crous et al. 2007 – this volume). Further<br />
genera that were separated from <strong>Cladosporium</strong> include Sorocybe<br />
resinae (Fr.) Fr. [≡ <strong>Cladosporium</strong> resinae (Lindau) G.A. de Vries,<br />
teleomorph: Amorphotheca resinae Parbery; Partridge & Morgan-<br />
Jones 2002] (see Seifert et al. 2007 – this volume), Devriesia Seifert<br />
& N.L. Nickerson, erected for heat tolerant species (Seifert et al.<br />
2004), Cladoriella Crous, erected for saprobic species (Crous et<br />
al. 2006b), Metulocladosporiella Crous, Schroers, Groenewald, U.<br />
Braun & K. Schub., erected for the causal agent of banana speckle<br />
disease (Crous et al. 2006a), Digitopodium U. Braun, Heuchert &<br />
K. Schub. <strong>and</strong> Parapericoniella U. Braun, Heuchert & K. Schub.,<br />
Fig. 2. Type specimen of Dematium herbarum Pers. (1794), preserved in the<br />
National Herbarium of the Netherl<strong>and</strong>s in Leiden.<br />
representing two genera of hyperparasitic hyphomycetes (Heuchert<br />
et al. 2005).<br />
Taxonomy of the teleomorph<br />
Teleomorphs of <strong>Cladosporium</strong> have traditionally been described in<br />
Mycosphaerella Johanson. <strong>The</strong> first indication that this may not be<br />
the case was the rDNA ITS sequence data presented by Crous et<br />
al. (2001), which revealed cladosporium-like taxa to cluster basal<br />
to Mycosphaerella s. str. This finding was further strengthened by<br />
adding 18S rDNA data, which clearly distinguished the <strong>Cladosporium</strong><br />
clade from Mycosphaerella (Braun et al. 2003). <strong>The</strong>se results<br />
lead to the erection of the <strong>genus</strong> Davidiella Crous & U. Braun for<br />
teleomorphs of <strong>Cladosporium</strong>, though it was largely established<br />
based on its unique anamorphs, rather than distinct teleomorph<br />
features. In a revision of the <strong>genus</strong> Mycosphaerella, Aptroot (2006)<br />
provided the first clear morphological characteristics to distinguish<br />
Davidiella from Mycosphaerella, referring to their sole-shaped<br />
ascospores, <strong>and</strong> angular lumina that are to be seen in Davidiella<br />
ascospores. Further phylogenetic evidence for the distinction was<br />
found by Schoch et al. (2006), which led to the erection of the<br />
family Davidiellaceae (Capnodiales). Detailed cultural studies of<br />
Davidiella teleomorphs, however, were still lacking (see Schubert<br />
et al. 2007 – this volume).<br />
What is <strong>Cladosporium</strong>?<br />
David (1997) provided the first modern concept of <strong>Cladosporium</strong> by<br />
conducting comprehensive scanning electron microscopic (SEM)<br />
examinations of the scar <strong>and</strong> hilum structure in <strong>Cladosporium</strong> <strong>and</strong><br />
Heterosporium, thereby confirming the observations of Roquebert<br />
(1981). He introduced the term “coronate” for the <strong>Cladosporium</strong><br />
scar type, which is characterised by having a central convex part<br />
(dome), surrounded by a raised periclinal rim (Fig. 3), <strong>and</strong> proved<br />
that these anamorphs are linked to teleomorphs now placed in<br />
Davidiella (see David 1997, fig. 12).<br />
This new concept of <strong>Cladosporium</strong> s. str. <strong>and</strong> Davidiella<br />
(David 1997, Braun et al. 2003, Aptroot 2006), supported by<br />
morphological <strong>and</strong> molecular data, rendered it possible to initiate<br />
a comprehensive revision of the <strong>genus</strong>. <strong>The</strong> first step was the<br />
preparation of a general, annotated check-list of <strong>Cladosporium</strong><br />
names (Dugan et al. 2004), followed by revisions of fungicolous<br />
(Heuchert et al. 2005) <strong>and</strong> foliicolous species of <strong>Cladosporium</strong> s.<br />
lat. (Schubert 2005b, Braun et al. 2006, Schubert & Braun 2004,<br />
2005a, b, 2006, 2007). <strong>The</strong> present study is the first to integrate<br />
these concepts on cladosporioid species in culture, in an attempt to<br />
further elucidate species of <strong>Cladosporium</strong>, <strong>and</strong> delineate the <strong>genus</strong><br />
from other, morphologically <strong>similar</strong> <strong>dematiaceous</strong> genera that have<br />
traditionally been confused with <strong>Cladosporium</strong> s. str.<br />
How natural should anamorph genera be?<br />
Article 59 of the International Code of Botanical Nomenclature<br />
was introduced to enable mycologists to name the asexual states<br />
of fungi that they encountered, <strong>and</strong> for which no teleomorph<br />
association was known. It was <strong>and</strong> remains a completely artificial<br />
system, complicated further by the evolution of the same anamorph<br />
morphology in different families, <strong>and</strong> even orders. In 1995 Gams<br />
discussed “How natural should anamorph genera be”, concluding<br />
that paraphyletic genera should be an acceptable option, <strong>and</strong> that<br />
anamorphs cannot reflect natural relationships. In a special volume<br />
dedicated at integrating molecular data <strong>and</strong> morphology, Seifert et<br />
al. (2000) proposed using anamorph names as adjectives, e.g.,<br />
acremonium-like, when they clustered in different clades, or were<br />
linked to different teleomorphs than the type species of the <strong>genus</strong>
Fig. 3. Coronate scar structure of <strong>Cladosporium</strong> herbaroides, visible by means of<br />
Scanning Electron Microscopy. Scale bar = 5 µm (Photo: Jan Dijksterhuis).<br />
Acremonium. In a phylogenetic study of the Herpotrichiellaceae,<br />
Haase et al. (1999) proposed to accept anamorphs as poly- <strong>and</strong><br />
paraphyletic within the order Chaetothyriales, as their taxonomy<br />
was unsupported by phylogeny, <strong>and</strong> Cook et al. (1997) as well as<br />
Braun et al. (2002) followed this methodology in naming anamorphs<br />
of the Erysiphaceae (Erysiphales). After much debate, we have<br />
chosen to use the same approach in this volume, <strong>and</strong> will refrain<br />
from introducing different anamorph genera for the same phenotype<br />
clustering within different clades of the same order. It is hoped that<br />
this approach will stop the unnecessary proliferation of names, until<br />
we can move to a single nomenclature for ascomycetous fungi.<br />
Anamorphs are just form taxa, established for the sole purpose<br />
of enabling mycologists to name asexual states that occur in<br />
the absence of their teleomorphs. Anamorph genera are simply<br />
phenotypic concepts that lack phylogenetic relevance within the<br />
order.<br />
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297.<br />
<strong>The</strong> Editors 1 July 2007<br />
Propositions for this volume:<br />
“Evolution gives rise to lineages, which we try to recognise as genera <strong>and</strong> species”<br />
“<strong>The</strong> most interesting fungi are those isolated by accident”
available online at www.studiesinmycology.org<br />
doi:10.3114/sim.2007.58.01<br />
Studies in Mycology 58: 1–32. 2007.<br />
Mycosphaerella is polyphyletic<br />
P.W. Crous 1* , U. Braun 2 <strong>and</strong> J.Z. Groenewald 1<br />
1<br />
<strong>CBS</strong> Fungal Biodiversity Centre, P.O. Box 85167, 3508 AD, Utrecht, <strong>The</strong> Netherl<strong>and</strong>s; 2 Martin-Luther-Universität, Institut für Biologie, Geobotanik und Botanischer Garten,<br />
Herbarium, Neuwerk 21, D-06099 Halle, Germany<br />
*Correspondence: Pedro W. Crous, p.crous@cbs.knaw.nl<br />
Abstract: Mycosphaerella, one of the largest genera of ascomycetes, encompasses several thous<strong>and</strong> species <strong>and</strong> has anamorphs residing in more than 30 form genera.<br />
Although previous phylogenetic studies based on the ITS rDNA locus supported the monophyly of the <strong>genus</strong>, DNA sequence data derived from the LSU gene distinguish several<br />
clades <strong>and</strong> families in what has hitherto been considered to represent the Mycosphaerellaceae. Several important leaf spotting <strong>and</strong> extremotolerant species need to be disposed<br />
to the <strong>genus</strong> Teratosphaeria, for which a new family, the Teratosphaeriaceae, is introduced. Other distinct clades represent the Schizothyriaceae, Davidiellaceae, Capnodiaceae,<br />
<strong>and</strong> the Mycosphaerellaceae. Within the two major clades, namely Teratosphaeriaceae <strong>and</strong> Mycosphaerellaceae, most anamorph genera are polyphyletic, <strong>and</strong> new anamorph<br />
concepts need to be derived to cope with dual nomenclature within the Mycosphaerella complex.<br />
Taxonomic novelties: Batcheloromyces eucalypti (Alcorn) Crous & U. Braun, comb. nov., Catenulostroma Crous & U. Braun, gen. nov., Catenulostroma abietis (Butin &<br />
Pehl) Crous & U. Braun, comb. nov., Catenulostroma chromoblastomycosum Crous & U. Braun, sp. nov., Catenulostroma elginense (Joanne E. Taylor & Crous) Crous & U.<br />
Braun, comb. nov., Catenulostroma excentricum (B. Sutton & Ganap.) Crous & U. Braun, comb. nov., Catenulostroma germanicum Crous & U. Braun, sp. nov., Catenulostroma<br />
macowanii (Sacc.) Crous & U. Braun, comb. nov., Catenulostroma microsporum (Joanne E. Taylor & Crous) Crous & U. Braun, comb. nov., Catenulostroma protearum (Crous<br />
& M.E. Palm) Crous & U. Braun, comb. nov., Penidiella Crous & U. Braun, gen. nov., Penidiella columbiana Crous & U. Braun, sp. nov., Penidiella cubensis (R.F. Castañeda) U.<br />
Braun, Crous & R.F. Castañeda, comb. nov., Penidiella nect<strong>and</strong>rae Crous, U. Braun & R.F. Castañeda, nom. nov., Penidiella rigidophora Crous, R.F. Castañeda & U. Braun, sp.<br />
nov., Penidiella strumelloidea (Milko & Dunaev) Crous & U. Braun, comb. nov., Penidiella venezuelensis Crous & U. Braun, sp. nov., Readeriella blakelyi (Crous & Summerell)<br />
Crous & U. Braun, comb. nov., Readeriella brunneotingens Crous & Summerell, sp. nov., Readeriella considenianae (Crous & Summerell) Crous & U. Braun, comb. nov.,<br />
Readeriella destructans (M.J. Wingf. & Crous) Crous & U. Braun, comb. nov., Readeriella dimorpha (Crous & Carnegie) Crous & U. Braun, comb. nov., Readeriella epicoccoides<br />
(Cooke & Massee) Crous & U. Braun, comb. nov., Readeriella gauchensis (M.-N. Cortinas, Crous & M.J. Wingf.) Crous & U. Braun, comb. nov., Readeriella molleriana (Crous<br />
& M.J. Wingf.) Crous & U. Braun, comb. nov., Readeriella nubilosa (Ganap. & Corbin) Crous & U. Braun, comb. nov., Readeriella pulcherrima (Gadgil & M. Dick) Crous &<br />
U. Braun, comb. nov., Readeriella stellenboschiana (Crous) Crous & U. Braun, comb. nov., Readeriella toledana (Crous & Bills) Crous & U. Braun, comb. nov., Readeriella<br />
zuluensis (M.J. Wingf., Crous & T.A. Cout.) Crous & U. Braun, comb. nov., Teratosphaeria africana (Crous & M.J. Wingf.) Crous & U. Braun, comb. nov., Teratosphaeria alistairii<br />
(Crous) Crous & U. Braun, comb. nov., Teratosphaeria associata (Crous & Carnegie) Crous & U. Braun, comb. nov., Teratosphaeria bellula (Crous & M.J. Wingf.) Crous & U.<br />
Braun, comb. nov., Teratosphaeria cryptica (Cooke) Crous & U. Braun, comb. nov., Teratosphaeria dentritica (Crous & Summerell) Crous & U. Braun, comb. nov., Teratosphaeria<br />
excentrica (Crous & Carnegie) Crous & U. Braun, comb. nov., Teratosphaeria fimbriata (Crous & Summerell) Crous & U. Braun, comb. nov., Teratosphaeria flexuosa (Crous<br />
& M.J. Wingf.) Crous & U. Braun, comb. nov., Teratosphaeria gamsii (Crous) Crous & U. Braun, comb. nov., Teratosphaeria jonkershoekensis (P.S. van Wyk, Marasas &<br />
Knox-Dav.) Crous & U. Braun, comb. nov., Teratosphaeria maxii (Crous) Crous & U. Braun, comb. nov., Teratosphaeria mexicana (Crous) Crous & U. Braun, comb. nov.,<br />
Teratosphaeria molleriana (Thüm.) Crous & U. Braun, comb. nov., Teratosphaeria nubilosa (Cooke) Crous & U. Braun, comb. nov., Teratosphaeria ohnowa (Crous & M.J. Wingf.)<br />
Crous & U. Braun, comb. nov., Teratosphaeria parkiiaffinis (Crous & M.J. Wingf.) Crous & U. Braun, comb. nov., Teratosphaeria parva (R.F. Park & Keane) Crous & U. Braun,<br />
comb. nov., Teratosphaeria perpendicularis (Crous & M.J. Wingf.) Crous & U. Braun, comb. nov., Teratosphaeria pluritubularis (Crous & Mansilla) Crous & U. Braun, comb.<br />
nov., Teratosphaeria pseudafricana (Crous & T.A. Cout.) Crous & U. Braun, comb. nov., Teratosphaeria pseudocryptica (Crous) Crous & U. Braun, comb. nov., Teratosphaeria<br />
pseudosuberosa (Crous & M.J. Wingf.) Crous & U. Braun, comb. nov., Teratosphaeria quasicercospora (Crous & T.A. Cout.) Crous & U. Braun, comb. nov., Teratosphaeria<br />
readeriellophora (Crous & Mansilla) Crous & U. Braun, comb. nov., Teratosphaeria secundaria (Crous & Alfenas) Crous & U. Braun, comb. nov., Teratosphaeria stramenticola<br />
(Crous & Alfenas) Crous & U. Braun, comb. nov., Teratosphaeria suberosa (Crous, F.A. Ferreira, Alfenas & M.J. Wingf.) Crous & U. Braun, comb. nov., Teratosphaeria suttonii<br />
(Crous & M.J. Wingf.) Crous & U. Braun, comb. nov., Teratosphaeria toledana (Crous & Bills) Crous & U. Braun, comb. nov., Teratosphaeriaceae Crous & U. Braun, fam. nov.<br />
Key words: Ascomycetes, Batcheloromyces, Colletogloeopsis, Readeriella, Teratosphaeria, Trimmatostroma, DNA sequence comparisons, systematics.<br />
Introduction<br />
<strong>The</strong> <strong>genus</strong> Mycosphaerella Johanson as presently circumscribed<br />
contains close to 3 000 species (Aptroot 2006), excluding its<br />
anamorphs, which represent thous<strong>and</strong>s of additional species<br />
(Crous et al. 2000, 2001, 2004a, b, 2006a, b, 2007b, Crous &<br />
Braun 2003). Crous (1998) predicted that Mycosphaerella would<br />
eventually be split according to its anamorph genera, <strong>and</strong> Crous<br />
et al. (2000) recognised six sections, as originally defined by Barr<br />
(1972). This was followed by a set of papers (Crous et al. 2001,<br />
Goodwin et al. 2001), where it was concluded, based on ITS DNA<br />
sequence data, that Mycosphaerella was monophyletic. A revision<br />
of the various coelomycete <strong>and</strong> hyphomycete anamorph concepts<br />
led Crous & Braun (2003) to propose a system whereby the asexual<br />
morphs could be allocated to various form genera affiliated with<br />
Mycosphaerella holomorphs.<br />
In a recent study that formed part of the US “Assembling<br />
the Fungal Tree of Life” project, Schoch et al. (2006) were able<br />
to show that the Mycosphaerellaceae represents a family within<br />
Capnodiales. Furthermore, some variation was also depicted within<br />
the family, which supported <strong>similar</strong> findings in other recent papers<br />
employing LSU sequence data, such as Hunter et al. (2006), <strong>and</strong><br />
Batzer et al. (2007). To further elucidate the phylogenetic variation<br />
observed within the Mycosphaerellaceae in these studies, a<br />
subset of isolates was selected for the present study, representing<br />
the various species recognised as morphologically distinct from<br />
Mycosphaerella s. str.<br />
<strong>The</strong> <strong>genus</strong> Mycosphaerella has in recent years been linked<br />
to approximately 30 anamorph genera (Crous & Braun 2003,<br />
Crous et al. 2007b). Many of these anamorph genera resulted<br />
from a reassessment of cercosporoid forms. Chupp (1954) was<br />
of the opinion that they all represented species of the <strong>genus</strong><br />
Cercospora Fresen., although he clearly recognised differences in<br />
their morphology. In a series of papers by Deighton, as well as<br />
others such as Sutton, Braun <strong>and</strong> Crous, the <strong>genus</strong> Cercospora<br />
was delimited based on its type species, Cercospora penicillata<br />
(Ces.) Fresen., while taxa formerly included in the <strong>genus</strong> by Chupp<br />
(1954) but differing in conidiophore arrangement, conidiogenesis,<br />
pigmentation, conidial catenulation, septation, <strong>and</strong> scar/hilum<br />
structure were allocated to other genera. Similar studies in which<br />
the type species were recollected <strong>and</strong> subjected to DNA sequence
Crous et al.<br />
Table 1. Isolates for which new sequences were generated.<br />
Anamorph Teleomorph Accession number 1 Host Country Collector GenBank Accession number<br />
Batcheloromyces eucalypti <strong>CBS</strong> 313.76; CPC 3632 Eucalyptus tessellaris Australia J.L. Alcorn EU019245<br />
Batcheloromyces leucadendri <strong>CBS</strong> 110892; CPC 1837 Leucadendron sp. South Africa L. Swart EU019246<br />
Batcheloromyces proteae <strong>CBS</strong> 110696; CPC 1518 Protea cynaroides South Africa L. Viljoen EU019247<br />
Capnobotryella renispora <strong>CBS</strong> 214.90*; <strong>CBS</strong> 176.88; IAM 13014; JCM 6932 Capnobotrys neessii Japan J. Sugiyama EU019248<br />
Catenulostroma abietis <strong>CBS</strong> 290.90 Man, skin lesion Netherl<strong>and</strong>s R.G.F. Wintermans EU019249<br />
Catenulostroma castellanii <strong>CBS</strong> 105.75*; ATCC 24788 Man, tinea nigra Venezuela — EU019250<br />
Catenulostroma chromoblastomycosum <strong>CBS</strong> 597.97 Man, chromoblastomycosis Zaire V. de Brouwere EU019251<br />
Catenulostroma elginense <strong>CBS</strong> 111030; CPC 1958 Protea gr<strong>and</strong>iceps South Africa J.E. Taylor EU019252<br />
Catenulostroma germanicum <strong>CBS</strong> 539.88 Stone Germany — EU019253<br />
Catenulostroma macowanii <strong>CBS</strong> 110756; CPC 1872 Protea nitida South Africa J.E. Taylor EU019254<br />
Catenulostroma microsporum Teratosphaeria microspora <strong>CBS</strong> 110890; CPC 1832 Protea cynaroides South Africa L. Swart EU019255<br />
Catenulostroma sp. Teratosphaeria<br />
<strong>CBS</strong> 118911; CPC 12085 Eucalyptus sp. Uruguay M.J. Wingfield EU019256<br />
pseudosuberosa<br />
Cercosporella centaureicola <strong>CBS</strong> 120253 Centaurea solstitiales Greece D. Berner EU019257<br />
Cibiessia dimorphospora <strong>CBS</strong> 120034; CPC 12636 Eucalyptus nitens Australia — EU019258<br />
Cibiessia minutispora CPC 13071* Eucalyptus henryii Australia A.J. Carnegie EU019259<br />
Cibiessia nontingens Teratosphaeria sp. <strong>CBS</strong> 120725*; CPC 13217 Eucalyptus tereticornis Australia B. Summerell EU019260<br />
<strong>Cladosporium</strong> bruhnei Davidiella allicina <strong>CBS</strong> 115683; ATCC 66670; CPC 5101 CCA-treated Douglas-fire pole U.S.A., New York C.J. Wang EU019261<br />
<strong>Cladosporium</strong> cladosporioides <strong>CBS</strong> 109.21; ATCC 11277; ATCC 200940; IFO 6368; IMI 049625 Sooty mould on Hedera helix U.K. — EU019262<br />
<strong>Cladosporium</strong> sphaerospermum <strong>CBS</strong> 188.54; ATCC 11290; IMI 049638 — — — EU019263<br />
<strong>Cladosporium</strong> uredinicola ATCC 46649 Hyperparasite on Cronartium<br />
fusiforme f. sp. quercum<br />
U.S.A., Alabama — EU019264<br />
Coccodinium bartschii <strong>CBS</strong> 121708; CPC 13861–13863 Sooty mould on unidentified tree Canada K.A. Seifert EU019265<br />
Dissoconium aciculare <strong>CBS</strong> 342.82*; CPC 1534 Erysiphe, on Medicago lupulina Germany T. Hijwegen EU019266<br />
Dissoconium commune “Mycosphaerella” communis <strong>CBS</strong> 114238*; CPC 10440 Eucalyptus globulus Spain J.P.M. Vazquez EU019267<br />
Dissoconium dekkeri “Mycosphaerella” lateralis <strong>CBS</strong> 567.89*; CPC 1535 Juniperus chinensis Netherl<strong>and</strong>s T. Hijwegen EU019268<br />
Fumagospora capnodioides Capnodium salicinum <strong>CBS</strong> 131.34 Sooty mould on Bursaria spinosa Indonesia — EU019269<br />
Hortaea werneckii <strong>CBS</strong> 107.67* Man, tinea nigra Portugal — EU019270<br />
Nothostrasseria dendritica Teratosphaeria dendritica CPC 12820 Eucalyptus nitens Australia A.J. Carnegie EU019271<br />
“Passalora” zambiae <strong>CBS</strong> 112970*; CPC 1228 Eucalyptus globulus Zambia T. Coutinho EU019272<br />
<strong>CBS</strong> 112971*; CMW 14782; CPC 1227 Eucalyptus globulus Zambia T. Coutinho EU019273<br />
Penidiella columbiana <strong>CBS</strong> 486.80 Paepalanthus columbianus Colombia W. Gams EU019274
Phylogenetic lineages in the Capnodiales<br />
Anamorph Teleomorph Accession number 1 Host Country Collector GenBank Accession number<br />
Penidiella nect<strong>and</strong>rae <strong>CBS</strong> 734.87*; ATCC 200932; INIFAT 87/45 Nect<strong>and</strong>ra coriacea Cuba R.F. Castañeda & EU019275<br />
G. Arnold<br />
Penidiella rigidophora <strong>CBS</strong> 314.95* Leaf litter of Smilax sp. Cuba R.F. Castañeda EU019276<br />
Penidiella strumelloidea <strong>CBS</strong> 114484*; VKM F-2534 Carex leaf, from stagnant water Russia S. Ozerskaya EU019277<br />
Penidiella venezuelensis <strong>CBS</strong> 106.75* Man, tinea nigra Venezuela D. Borelli EU019278<br />
Phaeotheca triangularis <strong>CBS</strong> 471.90* Wet surface of humidifier of<br />
airconditioning<br />
Belgium H. Beguin EU019279<br />
Phaeothecoidea eucalypti CPC 13010 Corymbia henryii Australia B. Summerell EU019280<br />
CPC 12918* Eucalyptus botryoides Australia B. Summerell EU019281<br />
Pleurophoma sp. Teratosphaeria fibrillosa CPC 1876 Protea nitida South Africa J.E. Taylor EU019282<br />
Pseudotaeniolina globosa <strong>CBS</strong> 109889* Rock Italy C. Urzi EU019283<br />
Ramularia pratensis var. pratensis CPC 11294 Rumex crispus Korea H.D. Shin EU019284<br />
Ramularia sp. <strong>CBS</strong> 324.87 On Mycosphaerella sp., leaf spot<br />
on Brassica sp.<br />
Netherl<strong>and</strong>s — EU019285<br />
Readeriella brunneotingens CPC 13303 Eucalyptus tereticornis Australia P.W. Crous EU019286<br />
Readeriella destructans <strong>CBS</strong> 111369*; CPC 1366 Eucalyptus gr<strong>and</strong>is Indonesia M.J. Wingfield EU019287<br />
Readeriella epicoccoides Teratosphaeria suttonii CPC 12352 Eucalyptus sp. U.S.A.,Hawaii W. Gams EU019288<br />
Readeriella eucalypti CPC 11186 Eucalyptus globulus Spain M.J. Wingfield EU019289<br />
Readeriella gauchensis <strong>CBS</strong> 120303*; CMW 17331 Eucalyptus gr<strong>and</strong>is Uruguay M.J. Wingfield EU019290<br />
Readeriella mirabilis <strong>CBS</strong> 116293; CPC 10506 Eucalyptus fastigata New Zeal<strong>and</strong> W. Gams EU019291<br />
Readeriella molleriana Teratosphaeria molleriana <strong>CBS</strong> 111164*; CMW 4940; CPC 1214 Eucalyptus globulus Portugal M.J. Wingfield EU019292<br />
Readeriella ovata complex CPC 18 Eucalyptus cladocalyx South Africa P.W. Crous EU019293<br />
<strong>CBS</strong> 111149; CPC 23 Eucalyptus cladocalyx South Africa P.W. Crous EU019294<br />
Readeriella stellenboschiana <strong>CBS</strong> 116428; CPC 10886 Eucalyptus sp. South Africa P.W. Crous EU019295<br />
Readeriella zuluensis <strong>CBS</strong> 120301*; CMW 17321 Eucalyptus gr<strong>and</strong>is South Africa M.J. Wingfield EU019296<br />
Septoria tritici Mycosphaerella graminicola <strong>CBS</strong> 100335; IPO 69001.61 Triticum aestivum — G.H.J. Kema EU019297<br />
<strong>CBS</strong> 110744; CPC 658 Triticum sp. South Africa P.W. Crous EU019298<br />
Trimmatostroma betulinum <strong>CBS</strong> 282.74 Betula verrucosa Netherl<strong>and</strong>s W.M. Loerakker EU019299<br />
Trimmatostroma salicis CPC 13571 Salix alba Germany U. Braun EU019300<br />
Teratosphaeria bellula <strong>CBS</strong> 111700; CPC 1821 Protea eximia South Africa J.E. Taylor EU019301<br />
Teratosphaeria mexicana CPC 12349 Eucalyptus sp. U.S.A.,Hawaii W. Gams EU019302<br />
Teratosphaeria nubilosa <strong>CBS</strong> 114419; CPC 10497 Eucalyptus globulus New Zeal<strong>and</strong> — EU019303<br />
<strong>CBS</strong> 116005*; CMW 3282; CPC 937 Eucalyptus globulus Australia A. Carnegie EU019304<br />
www.studiesinmycology.org
Crous et al.<br />
Table 1. (Continued).<br />
Anamorph Teleomorph Accession number 1 Host Country Collector GenBank Accession number<br />
Teratosphaeria ohnowa <strong>CBS</strong> 112896*; CMW 4937; CPC 1004 Eucalyptus gr<strong>and</strong>is South Africa M.J. Wingfield EU019305<br />
Teratosphaeria secundaria <strong>CBS</strong> 115608; CPC 504 Eucalyptus gr<strong>and</strong>is Brazil A.C. Alfenas EU019306<br />
Teratosphaeria sp. <strong>CBS</strong> 208.94; CPC 727 Eucalyptus gr<strong>and</strong>is Indonesia A.C. Alfenas EU019307<br />
1 ATCC: American Type Culture Collection, Virginia, U.S.A.; <strong>CBS</strong>: Centraalbureau voor Schimmelcultures, Utrecht, <strong>The</strong> Netherl<strong>and</strong>s; CPC: Culture collection of Pedro Crous, housed at <strong>CBS</strong>; CMW: Culture collection of Mike Wingfield, housed at FABI,<br />
Pretoria, South Africa; IAM: Institute of Applied Microbiology, University of Tokyo, Institute of molecular <strong>and</strong> cellular bioscience, Tokyo, Japan; IFO: Institute For Fermentation, Osaka, Japan; IMI: International Mycological Institute, CABI-Bioscience,<br />
Egham, Bakeham Lane, U.K.; INIFAT: Alex<strong>and</strong>er Humboldt Institute for Basic Research in Tropical Agriculture, Ciudad de La Habana, Cuba; JCM: Japan Collection Of Microorganisms, RIKEN BioResource Center, Japan; VKM: All-Russian Collection of<br />
Microorganisms, Institute of Biochemistry <strong>and</strong> Physiology of Microorganisms, Russian Academy of Sciences, Pushchino, Russia.<br />
*Ex-type cultures.<br />
analysis were undertaken to characterise Mycosphaerella (Verkley<br />
et al. 2004), <strong>and</strong> anamorph genera such as Pseudocercospora<br />
Speg., Stigmina Sacc., Phaeoisariopsis Ferraris (Crous et al.<br />
2006a), Ramulispora Miura (Crous et al. 2003), Batcheloromyces<br />
Marasas, P.S. van Wyk & Knox-Dav. (Taylor et al. 2003),<br />
Phaeophleospora Rangel <strong>and</strong> Dothistroma Hulbary (Crous et al.<br />
2000, 2001, Barnes et al. 2004).<br />
To assess the phylogeny of the species selected for the present<br />
study, DNA sequences were generated of the 28S rRNA (LSU)<br />
gene. In a further attempt to address monophyletic groups within<br />
this complex, these data were integrated with their morphological<br />
characteristics. To further resolve pleomorphism among the species<br />
studied, isolates were examined on a range of cultural media to<br />
induce possible synanamorphs.<br />
Materials <strong>and</strong> methods<br />
Isolates<br />
Chosen isolates represent various species previously observed to<br />
be morphologically distinct from Mycosphaerella s. str. (Crous 1998,<br />
Crous et al. 2004a, b, 2006a, b, 2007b). In a few cases, specifically<br />
Teratosphaeria fibrillosa Syd. & P. Syd. <strong>and</strong> Coccodinium bartschii<br />
A. Massal., fresh material had to be collected from South Africa<br />
<strong>and</strong> Canada, respectively. Excised tissue pieces bearing ascomata<br />
were soaked in water for approximately 2 h, after which they were<br />
placed in the bottom of Petri dish lids, with the top half of the<br />
dish containing 2 % malt extract agar (MEA) (Gams et al. 2007).<br />
Ascospore germination patterns were examined after 24 h, <strong>and</strong><br />
single-ascospore <strong>and</strong> conidial cultures established as described by<br />
Crous (1998). Colonies were sub-cultured onto synthetic nutrientpoor<br />
agar (SNA), potato-dextrose agar (PDA), oatmeal agar (OA),<br />
MEA (Gams et al. 2007), <strong>and</strong> incubated at 25 °C under continuous<br />
near-ultraviolet light to promote sporulation.<br />
DNA phylogeny<br />
Fungal colonies were established on agar plates, <strong>and</strong> genomic<br />
DNA was isolated following the CTAB-based protocol described<br />
in Gams et al. (2007). <strong>The</strong> primers V9G (de Hoog & Gerrits van<br />
den Ende 1998) <strong>and</strong> LR5 (Vilgalys & Hester 1990) were used to<br />
amplify part of the nuclear rDNA operon spanning the 3’ end of<br />
the 18S rRNA gene (SSU), the first internal transcribed spacer<br />
(ITS1), the 5.8S rRNA gene, the second ITS region (ITS2) <strong>and</strong> the<br />
5’ end of the 28S rRNA gene (LSU). <strong>The</strong> primers ITS4 (White et<br />
al. 1990), LR0R (Rehner & Samuels 1994), LR3R (www.biology.<br />
duke.edu/fungi/mycolab/primers.htm), <strong>and</strong> LR16 (Moncalvo et al.<br />
1993), were used as internal sequence primers to ensure good<br />
quality sequences over the entire length of the amplicon. <strong>The</strong><br />
ITS1, ITS2 <strong>and</strong> 5.8S rRNA gene (ITS) were only sequenced for<br />
isolates of which these data were not available. <strong>The</strong> ITS data<br />
were not included in the analyses but deposited in GenBank<br />
where applicable. <strong>The</strong> PCR conditions, sequence alignment <strong>and</strong><br />
subsequent phylogenetic analysis using parsimony, distance <strong>and</strong><br />
Bayesian analyses followed the methods of Crous et al. (2006c).<br />
Gaps longer than 10 bases were coded as single events for the<br />
phylogenetic analyses; the remaining gaps were treated as new<br />
character states. Sequence data were deposited in GenBank<br />
(Table 1) <strong>and</strong> alignments in TreeBASE (www.treebase.org).<br />
Taxonomy<br />
Wherever possible, 30 measurements (× 1 000 magnification)
Phylogenetic lineages in the Capnodiales<br />
were made of structures mounted in lactic acid, with the extremes<br />
of spore measurements given in parentheses. Ascospores were<br />
frequently also mounted in water to observe mucoid appendages<br />
<strong>and</strong> sheaths. Colony colours (surface <strong>and</strong> reverse) were assessed<br />
after 1–2 mo on MEA at 25 °C in the dark, using the colour charts of<br />
Rayner (1970). All cultures obtained in this study are maintained in<br />
the culture collection of the Centraalbureau voor Schimmelcultures<br />
(<strong>CBS</strong>) in Utrecht, the Netherl<strong>and</strong>s (Table 1). Nomenclatural<br />
novelties <strong>and</strong> descriptions were deposited in MycoBank (www.<br />
MycoBank.org).<br />
Results<br />
DNA phylogeny<br />
Amplification products of approximately 1 700 bases were obtained<br />
for the isolates listed in Table 1. <strong>The</strong> LSU region of the sequences<br />
was used to obtain additional sequences from GenBank which<br />
were added to the alignment. <strong>The</strong> manually adjusted alignment<br />
contained 97 sequences (including the two outgroup sequences)<br />
<strong>and</strong> 844 characters including alignment gaps. Of the 844 characters<br />
used in the phylogenetic analysis, 308 were parsimony-informative,<br />
105 were variable <strong>and</strong> parsimony-uninformative, <strong>and</strong> 431 were<br />
constant.<br />
<strong>The</strong> parsimony analysis of the LSU region yielded 1 135 equally<br />
most parsimonious trees (TL = 1 502 steps; CI = 0.446; RI = 0.787;<br />
RC = 0.351), one of which is shown in Fig. 1. Three orders are<br />
represented by the ingroup isolates, namely Chaetothyriales (100<br />
% bootstrap support), Helotiales (100 % bootstrap support) <strong>and</strong><br />
Capnodiales (100 % bootstrap support). <strong>The</strong>se are discussed in<br />
detail in the Taxonomy <strong>and</strong> Discussion sections. A new collection of<br />
Coccodinium bartschii A. Massal clusters (100 % bootstrap support)<br />
with members of the Herpotrichiellaceae (Chaetothyriales), whereas<br />
the type species of the <strong>genus</strong> Trimmatostroma Corda, namely<br />
T. salicis Corda, as well as T. betulinum (Corda) S. Hughes, are<br />
allied (99 % bootstrap support) with the Dermateaceae (Helotiales).<br />
<strong>The</strong> Capnodiales encompasses members of the Capnodiaceae,<br />
Trichosphaeriaceae, Davidiellaceae, Schizothyriaceae <strong>and</strong> taxa<br />
traditionally placed in the Mycosphaerellaceae, which is divided<br />
here into the Teratosphaeriaceae, (65 % bootstrap support), <strong>and</strong><br />
the Mycosphaerellaceae (76 % bootstrap support), which contains<br />
several subclades. Also included in the Capnodiales are Devriesia<br />
staurophora (W.B. Kendr.) Seifert & N.L. Nick., Staninwardia<br />
suttonii Crous & Summerell <strong>and</strong> Capnobotryella renispora Sugiy.<br />
as sister taxa to Teratosphaeriaceae s. str. Neighbour-joining<br />
analysis using three substitution models on the sequence data<br />
yielded trees supporting the same topologies, but differed from<br />
the parsimony tree presented with regard to the order of the<br />
families <strong>and</strong> orders at the deeper nodes, e.g., the Helotiales <strong>and</strong><br />
Chaetothyriales are swapped around, as are the Capnodiaceae <strong>and</strong><br />
the Trichosphaeriaceae / Davidiellaceae (data not shown). Using<br />
neighbour-joining analyses, the Mycosphaerellaceae s. str. clade<br />
obtained 71 %, 70 % <strong>and</strong> 70 % bootstrap support respectively with<br />
the uncorrected “p”, Kimura 2-parameter <strong>and</strong> HKY85 substitution<br />
models wherease the Teratosphaeriaceae clade obtained 74 %, 79<br />
% <strong>and</strong> 78 % bootstrap support respectively with the same models.<br />
<strong>The</strong> Schizothyriaceae clade appeared basal in the Capnodiales,<br />
irrespective of which substitution model was used.<br />
Bayesian analysis was conducted on the same aligned LSU<br />
dataset using a general time-reversible (GTR) substitution model<br />
with inverse gamma rates <strong>and</strong> dirichlet base frequencies. <strong>The</strong><br />
Markov Chain Monte Carlo (MCMC) analysis of 4 chains started<br />
from a r<strong>and</strong>om tree topology <strong>and</strong> lasted 23 881 500 generations.<br />
Trees were saved each 100 generations, resulting in 238 815 saved<br />
trees. Burn-in was set at 22 000 000 generations after which the<br />
likelihood values were stationary, leaving 18 815 trees from which<br />
the consensus tree (Fig. 2) <strong>and</strong> posterior probabilities (PP’s) were<br />
calculated. <strong>The</strong> average st<strong>and</strong>ard deviation of split frequencies<br />
was 0.011508 at the end of the run. <strong>The</strong> same overall topology as<br />
that observed using parsimony was obtained, with the exception of<br />
the inclusion of Staninwardia suttonii in the Mycosphaerellaceae<br />
(PP value of 0.74) <strong>and</strong> not in the Teratosphaeriaceae. <strong>The</strong><br />
Mycosphaerellaceae s. str. clade, as well as the Teratosphaeriaceae<br />
clade, obtained a PP value of 1.00.<br />
Taxonomy<br />
Based on the dataset generated in this study, several well-supported<br />
genera could be distinguished in the Mycosphaerella complex (Figs<br />
1–2), for which we have identified morphological characters. <strong>The</strong>se<br />
genera, <strong>and</strong> a selection of their species, are treated below.<br />
Key to Mycosphaerella, <strong>and</strong> Mycosphaerella-like genera treated<br />
1. Ascomata thyrothecial; anamorph Zygosporium ................................................................................................................. Schizothyrium<br />
1. Ascomata pseudothecial ............................................................................................................................................................................ 2<br />
2. Ascospores with irregular, angular lumens typical of Davidiella; anamorph <strong>Cladosporium</strong> s. str. .............................................. Davidiella<br />
2. Ascospores guttulate or not, lacking angular lumens; anamorph other than <strong>Cladosporium</strong> ...................................................................... 3<br />
3. Ascomata frequently linked by superficial stroma; hamathecial tissue, ascospore sheath, multi-layered endotunica, prominent<br />
periphysoids, <strong>and</strong> ascospores turning brown in asci frequently observed ......................................................................... Teratosphaeria<br />
3. Ascomata not linked by superficial stroma; hamathecial tissue, ascospore sheath, multi-layered endotunica, prominent periphysoids,<br />
ascospores turning brown in asci not observed ......................................................................................................................................... 4<br />
4. Conidiophores solitary, pale brown, giving rise to primary <strong>and</strong> secondary, actively discharged conidia; anamorph Dissoconium<br />
.................................................................................................................................................................... teleomorph Mycosphaerella-like<br />
4. Conidiomata variable from solitary conidiophores to sporodochia, fascicles to pycnidia, but conidia not actively discharged .....................<br />
.................................................................................................................................................................................. Mycosphaerella s. str.<br />
www.studiesinmycology.org
Crous et al.<br />
100<br />
10 changes<br />
65<br />
100<br />
Athelia epiphylla AY586633<br />
Paullicorticium ansatum AY586693<br />
65<br />
63<br />
68<br />
100<br />
99<br />
100<br />
91<br />
67<br />
94<br />
Coccodinium bartschii CPC 13861<br />
Exophiala oligosperma AF050289<br />
Exophiala jeanselmei AF050271<br />
Fonsecaea pedrosoi AF050276<br />
Capronia mansonii AY004338<br />
Exophiala dermatitidis DQ823100<br />
Vibrissea flavovirens AY789426<br />
Vibrissea truncorum AY789402<br />
Trimmatostroma salicis CPC 13571<br />
Mollisia cinerea DQ470942<br />
Trimmatostroma betulinum <strong>CBS</strong> 282.74<br />
Phaeotheca triangularis <strong>CBS</strong> 471.90<br />
Capnodium coffeae DQ247800<br />
Capnodium salicinum <strong>CBS</strong> 131.34<br />
Scorias spongiosa DQ678075<br />
63<br />
68<br />
73<br />
92<br />
65<br />
98<br />
100<br />
56<br />
58<br />
100 87<br />
100<br />
100<br />
76<br />
Trichosphaeria pilosa AY590297 Trichosphaeriaceae<br />
<strong>Cladosporium</strong> cladosporioides <strong>CBS</strong> 109.21<br />
<strong>Cladosporium</strong> uredinicola ATCC 46649<br />
<strong>Cladosporium</strong> bruhnei <strong>CBS</strong> 115683<br />
<strong>Cladosporium</strong> sphaerospermum <strong>CBS</strong> 188.54<br />
64<br />
Schizothyrium pomi EF134947<br />
Schizothyrium pomi EF134948<br />
80 100<br />
100<br />
99<br />
100<br />
100<br />
100<br />
63<br />
64<br />
61<br />
100<br />
Herpotrichiellaceae,<br />
Chaetothyriales,<br />
Chaetothyriomycetes<br />
Vibrisseaceae<br />
Dermateaceae<br />
Capnodiaceae<br />
Schizothyriaceae<br />
“Passalora” zambiae <strong>CBS</strong> 112970<br />
“Passalora” zambiae <strong>CBS</strong> 112971<br />
Dissoconium aciculare <strong>CBS</strong> 342.82<br />
“Mycosphaerella” communis <strong>CBS</strong> 114238<br />
“Mycosphaerella” lateralis <strong>CBS</strong> 567.89<br />
Mycosphaerella graminicola <strong>CBS</strong> 100335<br />
Septoria tritici <strong>CBS</strong> 110744<br />
Cercosporella centaureicola <strong>CBS</strong> 120253<br />
Mycosphaerella punctiformis AY490776<br />
Ramularia sp. <strong>CBS</strong> 324.87<br />
Ramularia miae DQ885902<br />
Ramularia pratensis var. pratensis CPC 11294<br />
Capnobotryella renispora <strong>CBS</strong> 214.90<br />
100<br />
94<br />
88<br />
80<br />
99<br />
81<br />
Staninwardia suttonii DQ923535<br />
Devriesia staurophora DQ008150<br />
Devriesia staurophora DQ008151<br />
Pseudotaeniolina globosa <strong>CBS</strong> 109889<br />
Batcheloromyces proteae <strong>CBS</strong> 110696<br />
Batcheloromyces leucadendri <strong>CBS</strong> 110892<br />
Teratosphaeria alistairii DQ885901<br />
Penidiella columbiana <strong>CBS</strong> 486.80<br />
Teratosphaeria sp. <strong>CBS</strong> 208.94<br />
Teratosphaeria ohnowa <strong>CBS</strong> 112896<br />
Teratosphaeria secundaria <strong>CBS</strong> 115608<br />
Teratosphaeria flexuosa DQ246232<br />
Hortaea werneckii <strong>CBS</strong> 107.67<br />
Catenulostroma castellanii <strong>CBS</strong> 105.75<br />
Teratosphaeria bellula <strong>CBS</strong> 111700<br />
Catenulostroma elginense <strong>CBS</strong> 111030<br />
Teratosphaeria parva DQ246240<br />
100 Teratosphaeria parva DQ246243<br />
Catenulostroma germanicum <strong>CBS</strong> 539.88<br />
100<br />
Catenulostroma microsporum <strong>CBS</strong> 110890<br />
Catenulostroma abietis <strong>CBS</strong> 290.90<br />
Catenulostroma chromoblastomycosum <strong>CBS</strong> 597.97<br />
Phaeothecoidea eucalypti CPC 13010<br />
Phaeothecoidea eucalypti CPC 12918<br />
Teratosphaeria suberosa DQ246235<br />
Teratosphaeria pseudosuberosa <strong>CBS</strong> 118911<br />
93<br />
Teratosphaeria mexicana CPC 12349<br />
97 Teratosphaeria mexicana DQ246237<br />
Teratosphaeria readeriellophora DQ246238<br />
100<br />
93 Readeriella eucalypti CPC 11186<br />
Cibiessia minutispora CPC 13071<br />
Cibiessia dimorphospora <strong>CBS</strong> 120034<br />
Readeriella mirabilis <strong>CBS</strong> 116293<br />
Readeriella novaezel<strong>and</strong>iae DQ246239<br />
57 Nothostrasseria dendritica CPC 12820<br />
Batcheloromyces eucalypti <strong>CBS</strong> 313.76<br />
99 Readeriella ovata complex <strong>CBS</strong> 111149<br />
Readeriella ovata complex CPC 18<br />
Readeriella destructans <strong>CBS</strong> 111369<br />
Readeriella pulcherrima DQ246224<br />
Readeriella brunneotingens CPC 13303<br />
Readeriella dimorpha DQ923528<br />
Teratosphaeria molleriana DQ246219<br />
Teratosphaeria molleriana DQ246221<br />
Teratosphaeria molleriana <strong>CBS</strong> 111164<br />
Teratosphaeria fibrillosa CPC 1876<br />
Teratosphaeria maxii DQ885899<br />
Catenulostroma macowanii <strong>CBS</strong> 110756<br />
Teratosphaeria nubilosa <strong>CBS</strong> 114419<br />
Teratosphaeria nubilosa <strong>CBS</strong> 116005<br />
Readeriella stellenboschiana <strong>CBS</strong> 116428<br />
Readeriella gauchensis <strong>CBS</strong> 120303<br />
Teratosphaeria cryptica DQ246222<br />
Readeriella considenianae DQ923527<br />
Cibiessia nontingens <strong>CBS</strong> 120725<br />
Readeriella zuluensis <strong>CBS</strong> 120301<br />
Readeriella blakelyi DQ923526<br />
Teratosphaeria toledana DQ246230<br />
100<br />
Teratosphaeria suttonii DQ246227<br />
Readeriella epicoccoides CPC 12352<br />
Helotiales, Leotiomycetes<br />
Davidiellaceae<br />
Mycosphaerellaceae<br />
Teratosphaeriaceae<br />
Capnodiales, Dothideomycetes<br />
Fig. 1. One of 1 135 equally most parsimonious trees obtained from a heuristic search with 100 r<strong>and</strong>om taxon additions of the LSU sequence alignment using PAUP v. 4.0b10.<br />
<strong>The</strong> scale bar shows 10 changes, <strong>and</strong> bootstrap support values from 1 000 replicates are shown at the nodes. Thickened lines indicate the strict consensus branches <strong>and</strong> extype<br />
sequences are printed in bold face. <strong>The</strong> tree was rooted to two sequences obtained from GenBank (Athelia epiphylla AY586633 <strong>and</strong> Paullicorticium ansatum AY586693).
Phylogenetic lineages in the Capnodiales<br />
1.00<br />
0.62<br />
0.1 expected changes per site<br />
Athelia epiphylla AY586633<br />
Paullicorticium ansatum AY586693<br />
0.62<br />
1.00<br />
0.75<br />
1.00<br />
1.00<br />
0.99<br />
0.99<br />
0.85<br />
0.97<br />
Capronia mansonii AY004338<br />
Exophiala dermatitidis DQ823100<br />
Fonsecaea pedrosoi AF050276<br />
Coccodinium bartschii CPC 13861<br />
Exophiala oligosperma AF050289<br />
Exophiala jeanselmei AF050271<br />
Vibrisseaceae<br />
0.97<br />
Vibrissea flavovirens AY789426<br />
Vibrissea truncorum AY789402<br />
1.00<br />
Trimmatostroma salicis CPC 13571<br />
Mollisia cinerea DQ470942<br />
Trimmatostroma betulinum <strong>CBS</strong> 282.74<br />
Phaeotheca triangularis <strong>CBS</strong> 471.90<br />
Capnodium coffeae DQ247800<br />
Capnodium salicinum <strong>CBS</strong> 131.34<br />
1.00<br />
1.00<br />
1.00<br />
0.80<br />
0.74<br />
1.00<br />
0.55<br />
1.00<br />
1.00<br />
0.91<br />
Dermateaceae<br />
Herpotrichiellaceae,<br />
Chaetothyriales,<br />
Chaetothyriomycetes<br />
Helotiales,<br />
Leotiomycetes<br />
Capnodiaceae<br />
Scorias spongiosa DQ678075<br />
1.00<br />
Trichosphaeria pilosa AY590297 Trichosphaeriaceae<br />
<strong>Cladosporium</strong> cladosporioides <strong>CBS</strong> 109.21<br />
<strong>Cladosporium</strong> uredinicola ATCC 46649<br />
1.00 <strong>Cladosporium</strong> bruhnei <strong>CBS</strong> 115683<br />
0.95 <strong>Cladosporium</strong> sphaerospermum <strong>CBS</strong> 188.54<br />
Staninwardia suttonii DQ923535<br />
1.00 Schizothyrium pomi EF134947<br />
Schizothyrium pomi EF134948<br />
Schizothyriaceae<br />
1.00<br />
1.00<br />
0.83 1.00<br />
0.72 1.00<br />
1.00 0.58<br />
0.79<br />
0.89 0.62 1.00<br />
0.99<br />
1.00<br />
0.58<br />
0.94<br />
0.56<br />
0.83<br />
0.94<br />
0.77<br />
0.51<br />
1.00<br />
1.00<br />
0.77<br />
0.85<br />
Dissoconium aciculare <strong>CBS</strong> 342.82<br />
“Mycosphaerella” communis <strong>CBS</strong> 114238<br />
“Mycosphaerella” lateralis <strong>CBS</strong> 567.89<br />
Passalora zambiae <strong>CBS</strong> 112970<br />
Passalora zambiae <strong>CBS</strong> 112971<br />
Cercosporella centaureicola <strong>CBS</strong> 120253<br />
Mycosphaerella punctiformis AY490776<br />
Ramularia sp. <strong>CBS</strong> 324.87<br />
Ramularia miae DQ885902<br />
Ramularia pratensis var. pratensis CPC 11294<br />
0.57 Mycosphaerella graminicola <strong>CBS</strong> 100335<br />
1.00 Septoria tritici <strong>CBS</strong> 110744<br />
Capnobotryella renispora <strong>CBS</strong> 214.90<br />
Pseudotaeniolina globosa <strong>CBS</strong> 109889<br />
Catenulostroma chromoblastomycosum <strong>CBS</strong> 597.97<br />
Teratosphaeria bellula <strong>CBS</strong> 111700<br />
Hortaea werneckii <strong>CBS</strong> 107.67<br />
Catenulostroma castellanii <strong>CBS</strong> 105.75<br />
Batcheloromyces proteae <strong>CBS</strong> 110696<br />
Batcheloromyces leucadendri <strong>CBS</strong> 110892<br />
Teratosphaeria alistairii DQ885901<br />
Penidiella columbiana <strong>CBS</strong> 486.80<br />
Teratosphaeria sp. <strong>CBS</strong> 208.94<br />
Teratosphaeria ohnowa <strong>CBS</strong> 112896<br />
Teratosphaeria secundaria <strong>CBS</strong> 115608<br />
Teratosphaeria flexuosa DQ246232<br />
Catenulostroma elginense <strong>CBS</strong> 111030<br />
Teratosphaeria parva DQ246240<br />
1.00<br />
1.00<br />
1.00<br />
1.00<br />
1.00<br />
1.00<br />
0.66<br />
0.76<br />
Teratosphaeria parva DQ246243<br />
Devriesia staurophora DQ008150<br />
Devriesia staurophora DQ008151<br />
Catenulostroma germanicum <strong>CBS</strong> 539.88<br />
Catenulostroma microsporum <strong>CBS</strong> 110890<br />
Catenulostroma abietis <strong>CBS</strong> 290.90<br />
Phaeothecoidea eucalypti CPC 13010<br />
Phaeothecoidea eucalypti CPC 12918<br />
Teratosphaeria pseudosuberosa <strong>CBS</strong> 118911<br />
Teratosphaeria suberosa DQ246235<br />
0.97<br />
Teratosphaeria mexicana CPC 12349<br />
Teratosphaeria mexicana <strong>CBS</strong> 110502<br />
Readeriella novaezel<strong>and</strong>iae DQ246239<br />
Nothostrasseria dendritica CPC 12820<br />
Readeriella mirabilis <strong>CBS</strong> 116293<br />
Cibiessia dimorphospora <strong>CBS</strong> 120034<br />
Cibiessia minutispora CPC 13071<br />
Teratosphaeria readeriellophora DQ246238<br />
Readeriella eucalypti CPC 11186<br />
1.00<br />
Readeriella brunneotingens CPC 13303<br />
Readeriella stellenboschiana <strong>CBS</strong> 116428<br />
Readeriella gauchensis <strong>CBS</strong> 120303<br />
Readeriella considenianae DQ923527<br />
Teratosphaeria cryptica DQ246222<br />
Cibiessia nontingens <strong>CBS</strong> 120725<br />
Readeriella zuluensis <strong>CBS</strong> 120301<br />
Teratosphaeria toledana DQ246230<br />
Teratosphaeria suttonii DQ246227<br />
Readeriella epicoccoides CPC 12352<br />
1.00<br />
Readeriella blakelyi DQ923526<br />
Readeriella ovata complex <strong>CBS</strong> 111149<br />
Readeriella ovata complex CPC 18<br />
Batcheloromyces eucalypti <strong>CBS</strong> 313.76<br />
Readeriella destructans <strong>CBS</strong> 111369<br />
Readeriella pulcherrima DQ246224<br />
Teratosphaeria nubilosa <strong>CBS</strong> 114419<br />
Teratosphaeria nubilosa <strong>CBS</strong> 116005<br />
Readeriella dimorpha DQ923528<br />
Teratosphaeria molleriana DQ246219<br />
Teratosphaeria molleriana DQ246221<br />
Teratosphaeria molleriana <strong>CBS</strong> 111164<br />
Teratosphaeria fibrillosa CPC 1876<br />
Catenulostroma macowanii <strong>CBS</strong> 110756<br />
Teratosphaeria maxii DQ885899<br />
Davidiellaceae<br />
Mycosphaerellaceae Teratosphaeriaceae<br />
Capnodiales, Dothideomycetes<br />
Fig. 2. Consensus phylogram (50 % majority rule) of 18 815 trees resulting from a Bayesian analysis of the LSU sequence alignment using MrBayes v. 3.1.2. Bayesian posterior<br />
probabilities are indicated at the nodes. Ex-type sequences are printed in bold face. <strong>The</strong> tree was rooted to two sequences obtained from GenBank (Athelia epiphylla AY586633<br />
<strong>and</strong> Paullicorticium ansatum AY586693).<br />
www.studiesinmycology.org
Crous et al.<br />
Treatment of phylogenetic clades<br />
Davidiellaceae clade<br />
Davidiella Crous & U. Braun, Mycol. Progr. 2: 8. 2003.<br />
Type species: Davidiella tassiana (De Not.) Crous & U. Braun,<br />
Mycol. Progr. 2: 8. 2003.<br />
Basionym: Sphaerella tassiana De Not., Sferiacei Italici 1: 87.<br />
1863.<br />
Description: Schubert et al. (2007 – this volume).<br />
Anamorph: <strong>Cladosporium</strong> Link, Ges. Naturf. Freunde Berlin Mag.<br />
Neuesten Entdeck. Gesammten Naturk. 7: 37. 1816.<br />
Type species: <strong>Cladosporium</strong> herbarum (Pers. : Fr.) Link, Ges.<br />
Naturf. Freunde Berlin Mag. Neuesten Entdeck. Gesammten<br />
Naturk. 7: 37. 1816.<br />
Basionym: Dematium herbarum Pers., Ann. Bot. (Usteri), 11 Stück:<br />
32. 1794: Fr., Syst. Mycol. 3: 370. 1832.<br />
Description: Schubert et al. (2007 – this volume).<br />
Notes: <strong>The</strong> <strong>genus</strong> Davidiella (Davidiellaceae) was recently<br />
introduced for teleomorphs of <strong>Cladosporium</strong> s. str. (Braun et al.<br />
2003). <strong>The</strong> <strong>genus</strong> <strong>Cladosporium</strong> is well-established, <strong>and</strong> contains<br />
around 772 names (Dugan et al. 2004), while Davidiella presently<br />
has 33 names (www.MycoBank.org), of which only around five<br />
have acknowledged <strong>Cladosporium</strong> states.<br />
Teratosphaeriaceae clade<br />
Teratosphaeria Syd. & P. Syd., Ann. Mycol. 10: 39. 1912.<br />
Type species: Teratosphaeria fibrillosa Syd. & P. Syd., Ann. Mycol.<br />
10: 40. 1912. Fig. 3.<br />
Description: Crous et al. (2004a; figs 182–185).<br />
Notes: Although <strong>similar</strong> in morphology, the <strong>genus</strong> Teratosphaeria<br />
was separated from Mycosphaerella based on its ascomatal<br />
arrangement, <strong>and</strong> periphysate ostioles (Müller & Oehrens 1982).<br />
It was later synonymised under Mycosphaerella by Taylor et<br />
al. (2003), who showed that the type species clustered within<br />
Mycosphaerella based on ITS DNA sequence data. <strong>The</strong> LSU<br />
sequence data generated in the present study, has clearly shown<br />
that Mycosphaerella is polyphyletic, thus contradicting earlier<br />
reports of monophyly by Crous et al. (2000) <strong>and</strong> Goodwin et al.<br />
(2001), which were based on ITS data.<br />
A re-examination of T. fibrillosa, the type species of<br />
Teratosphaeria, revealed several morphological features that<br />
characterise the majority of the taxa clustering in the clade, though<br />
several characters have been lost in some of the small-spored<br />
species. <strong>The</strong>se characters are discussed below:<br />
1. Teratosphaeria fibrillosa has a superficial stroma linking<br />
ascomata together, almost appearing like a spider’s web on the leaf<br />
surface. Although this feature is not seen in other taxa in this clade,<br />
some species, such as M. suberosa Crous, F.A. Ferreira, Alfenas<br />
& M.J. Wingf. <strong>and</strong> M. pseudosuberosa Crous & M.J. Wingf. have<br />
a superficial stroma, into which the ascomata are inbedded (Crous<br />
1998, Crous et al. 2006b).<br />
2. Ascospores of Teratosphaeria become brown <strong>and</strong><br />
verruculose while still in their asci. This feature is commonly<br />
observed in species such as M. jonkershoekensis P.S. van Wyk,<br />
Marasas & Knox-Dav., M. alistairii Crous, M. mexicana Crous, M.<br />
maxii Crous <strong>and</strong> M. excentricum Crous & Carnegie (Crous 1998,<br />
Crous & Groenewald 2006a, b, Crous et al. 2007b).<br />
3. A few ascomata of T. fibrillosa were found to have<br />
some pseudoparaphysoidal remnants (cells to distinguish<br />
pseudoparaphyses), though they mostly disappear with age. This<br />
feature is rather uncommon, though pseudoparaphyses were<br />
observed in ascomata of M. eucalypti (Wakef.) Hansf.<br />
4. Ascospores of Teratosphaeria were found to be covered in<br />
a mucous sheath, which is commonly observed in other taxa in this<br />
clade, such as M. bellula Crous & M.J. Wingf., M. pseudocryptica<br />
Crous, M. suberosa, M. pseudosuberosa, M. associata Crous &<br />
Carnegie, M. dendritica Crous & Summerell <strong>and</strong> M. fimbriata Crous<br />
& Summerell (Crous et al. 2004b, 2006b, 2007b). Re-examination<br />
of fresh collections also revealed ascospores of M. cryptica (Cooke)<br />
Hansf. <strong>and</strong> M. nubilosa (Cooke) Hansf. to have a weakly definable<br />
sheath. Germinating ascospores of species in this clade all exhibit<br />
a prominent mucoid sheath.<br />
5. Asci of T. fibrillosa were observed to have a multi-layered<br />
endotunica, which, although not common, can be seen in species<br />
such as M. excentrica, M. maxii, M. alistairii, M. pseudosuberosa,<br />
M. fimbriata (Crous et al. 2006b, 2007b, Crous & Groenewald<br />
2006a, b), <strong>and</strong> also M. nubilosa.<br />
6. Finally, ascomata of T. fibrillosa <strong>and</strong> T. proteae-arboreae<br />
P.S. van Wyk, Marasas & Knox-Dav. have well-developed<br />
ostiolar periphyses, which are also present in species such as M.<br />
suberosa, M. pseudosuberosa, M. maxii <strong>and</strong> T. microspora Joanne<br />
E. Taylor & Crous (Crous 1998, Crous et al. 2004a, b, 2006b).<br />
Morphologically thus, the Teratosphaeria clade is distinguishable<br />
from Mycosphaerella s. str., though these differences are less<br />
pronounced in some of the smaller-spored species. Based on<br />
these distinct morphological features, as well as its phylogenetic<br />
position within the Capnodiales, a new family is herewith proposed<br />
to accommodate species of Teratosphaeria:<br />
Teratosphaeriaceae Crous & U. Braun, fam. nov. MycoBank<br />
MB504464.<br />
Ascomata pseudotheciales, superficiales vel immersa, saepe in stromate ex cellulis<br />
brunneis pseudoparenchymatibus disposita, globulares, uniloculares, papillata,<br />
apice ostiolato, periphysata, saepe cum periphysoidibus; tunica multistratosa, ex<br />
cellulis brunneis angularibus composita, strato interiore ex cellulis applanatis hyalinis;<br />
saepe cum pseudoparaphysibus subcylindricis, ramosis, septatis, anastomosibus.<br />
Asci fasciculati, octospori, bitunicati, saepe cum endotunica multistratosa.<br />
Ascosporae ellipsoideae-fusiformes vel obovoideae, 1-septatae, hyalinae, deinde<br />
pallide brunneae et verruculosae, saepe mucosae.<br />
Ascomata pseudothecial, superficial to immersed, frequently<br />
situated in a stroma of brown pseudoparenchymatal cells,<br />
globose, unilocular, papillate, ostiolate, canal periphysate, with<br />
periphysoids frequently present; wall consisting of several layers<br />
of brown textura angularis; inner layer of flattened, hyaline cells.<br />
Pseudoparaphyses frequently present, subcylindrical, branched,<br />
septate, anastomosing. Asci fasciculate, 8-spored, bitunicate,<br />
frequently with multi-layered endotunica. Ascospores ellipsoidfusoid<br />
to obovoid, 1-septate, hyaline, but becoming pale brown <strong>and</strong><br />
verruculose, frequently covered in mucoid sheath.<br />
Typus: Teratosphaeria Syd. & P. Syd., Ann. Mycol. 10: 39. 1912.<br />
Teratosphaeria africana (Crous & M.J. Wingf.) Crous & U. Braun,<br />
comb. nov. MycoBank MB504466.
Phylogenetic lineages in the Capnodiales<br />
Fig. 3. Teratosphaeria fibrillosa (epitype material). A. Leaf spots. B. Subepidermal ascomata linked by means of stromatic tissue. C. Paraphyses among asci. D. Periphysoids. E.<br />
Ascospores becoming brown in asci. F–G. Multi-layered endotunica. H–K. Ascospores, becoming brown <strong>and</strong> verruculose. L–M. Germinating ascospores. Scale bars = 10 µm.<br />
Basionym: Mycosphaerella africana Crous & M.J. Wingf., Mycologia<br />
88: 450. 1996.<br />
Teratosphaeria associata (Crous & Carnegie) Crous & U. Braun,<br />
comb. nov. MycoBank MB504467.<br />
Basionym: Mycosphaerella associata Crous & Carnegie, Fungal<br />
Diversity 26: 159. 2007.<br />
Teratosphaeria alistairii (Crous) Crous & U. Braun, comb. nov.<br />
MycoBank MB504468.<br />
Basionym: Mycosphaerella alistairii Crous, in Crous & Groenewald,<br />
Fungal Planet, No. 4. 2006.<br />
Anamorph: Batcheloromyces sp.<br />
www.studiesinmycology.org
Crous et al.<br />
Teratosphaeria bellula (Crous & M.J. Wingf.) Crous & U. Braun,<br />
comb. nov. MycoBank MB504469.<br />
Basionym: Mycosphaerella bellula Crous & M.J. Wingf., Mycotaxon<br />
46: 20. 1993.<br />
Teratosphaeria cryptica (Cooke) Crous & U. Braun, comb. nov.<br />
MycoBank MB504470.<br />
Basionym: Sphaerella cryptica Cooke, Grevillea 20: 5. 1891.<br />
≡ Mycosphaerella cryptica (Cooke) Hansf., Proc. Linn. Soc. New South<br />
Wales 81: 35. 1956.<br />
Anamorph: Readeriella nubilosa (Ganap. & Corbin) Crous & U.<br />
Braun, comb. nov. MycoBank MB504471.<br />
Basionym: Colletogloeum nubilosum Ganap. & Corbin, Trans. Brit.<br />
Mycol. Soc. 72: 237. 1979.<br />
≡ Colletogloeopsis nubilosum (Ganap. & Corbin) Crous & M.J. Wingf.,<br />
Canad. J. Bot. 75: 668. 1997.<br />
Teratosphaeria dendritica (Crous & Summerell) Crous & U.<br />
Braun, comb. nov. MycoBank MB504472.<br />
Basionym: Mycosphaerella dendritica Crous & Summerell, Fungal<br />
Diversity 26: 161. 2007.<br />
Anamorph: Nothostrasseria dendritica (Hansf.) Nag Raj, Canad.<br />
J. Bot. 61: 25. 1983.<br />
Basionym: Spilomyces dendriticus Hansf., Proc. Linn. Soc. New<br />
South Wales 81: 32. 1956.<br />
Teratosphaeria excentrica (Crous & Carnegie) Crous & U. Braun,<br />
comb. nov. MycoBank MB504473.<br />
Basionym: Mycosphaerella excentrica Crous & Carnegie, Fungal<br />
Diversity 26: 164. 2007.<br />
Anamorph: Catenulostroma excentricum (B. Sutton & Ganap.)<br />
Crous & U. Braun, comb. nov. MycoBank MB504475.<br />
Basionym: Trimmatostroma excentricum B. Sutton & Ganap., New<br />
Zeal<strong>and</strong> J. Bot. 16: 529. 1978.<br />
Teratosphaeria fibrillosa Syd. & P. Syd., Ann. Mycol. 10: 40.<br />
1912.<br />
≡ Mycosphaerella fibrillosa (Syd. & P. Syd.) Joanne E. Taylor & Crous,<br />
Mycol. Res. 107: 657. 2003.<br />
Specimens examined: South Africa, Western Cape Province, Bains Kloof near<br />
Wellington, on living leaves of Protea gr<strong>and</strong>iflora, 26 Feb. 1911, E.M. Doidge,<br />
holotype PREM; Stellenbosch, Jonkershoek valley, S33° 59’ 44.7”, E18° 58’ 50.6”,<br />
1 Apr. 2007, on leaves of Protea sp., P.W. Crous & L. Mostert, epitype designated<br />
here <strong>CBS</strong> H-19913, culture ex-epitype <strong>CBS</strong> 121707 = CPC 13960.<br />
Teratosphaeria fimbriata (Crous & Summerell) Crous & U. Braun,<br />
comb. nov. MycoBank MB504476.<br />
Basionym: Mycosphaerella fimbriata Crous & Summerell, Fungal<br />
Diversity 26: 166. 2007.<br />
Teratosphaeria flexuosa (Crous & M.J. Wingf.) Crous & U. Braun,<br />
comb. nov. MycoBank MB504477.<br />
Basionym: Mycosphaerella flexuosa Crous & M.J. Wingf., Mycol.<br />
Mem. 21: 58. 1998.<br />
Teratosphaeria gamsii (Crous) Crous & U. Braun, comb. nov.<br />
MycoBank MB504478.<br />
Basionym: Mycosphaerella gamsii Crous, Stud. Mycol. 55: 113.<br />
2006.<br />
Teratosphaeria jonkershoekensis (P.S. van Wyk, Marasas &<br />
Knox-Dav.) Crous & U. Braun, comb. nov. MycoBank MB504479.<br />
Basionym: Mycosphaerella jonkershoekensis P.S. van Wyk,<br />
Marasas & Knox-Dav., J. S. African Bot. 41: 234. 1975.<br />
Teratosphaeria maxii (Crous) Crous & U. Braun, comb. nov.<br />
MycoBank MB504480.<br />
Basionym: Mycosphaerella maxii Crous, in Crous & Groenewald,<br />
Fungal Planet No. 6. 2006.<br />
Teratosphaeria mexicana (Crous) Crous & U. Braun, comb. nov.<br />
MycoBank MB504481.<br />
Basionym: Mycosphaerella mexicana Crous, Mycol. Mem. 21: 81.<br />
1998.<br />
Teratosphaeria microspora Joanne E. Taylor & Crous, Mycol.<br />
Res. 104: 631. 2000.<br />
≡ Mycosphaerella microspora (Joanne E. Taylor & Crous) Joanne E.<br />
Taylor & Crous, Mycol. Res. 107: 657. 2003.<br />
Anamorph: Catenulostroma microsporum (Joanne E. Taylor &<br />
Crous) Crous & U. Braun, comb. nov. MycoBank MB504482.<br />
Basionym: Trimmatostroma microsporum Joanne E. Taylor &<br />
Crous, Mycol. Res. 104: 631. 2000.<br />
Teratosphaeria molleriana (Thüm.) Crous & U. Braun, comb.<br />
nov. MycoBank MB504483.<br />
Basionym: Sphaerella molleriana Thüm., Revista Inst. Sci. Lit.<br />
Coimbra 28: 31. 1881.<br />
≡ Mycosphaerella molleriana (Thüm) Lindau, Nat. Pfanzenfam. 1: 424.<br />
1897.<br />
= Mycosphaerella vespa Carnegie & Keane, Mycol. Res. 102: 1275. 1998.<br />
= Mycosphaerella ambiphylla A. Maxwell, Mycol. Res. 107: 354. 2003.<br />
Anamorph: Readeriella molleriana (Crous & M.J. Wingf.) Crous &<br />
U. Braun, comb. nov. MycoBank MB504484.<br />
Basionym: Colletogloeopsis molleriana Crous & M.J. Wingf.,<br />
Canad. J. Bot. 75: 670. 1997.<br />
Teratosphaeria nubilosa (Cooke) Crous & U. Braun, comb. nov.<br />
MycoBank MB504485.<br />
Basionym: Sphaerella nubilosa Cooke, Grevillea 19: 61. 1892.<br />
≡ Mycosphaerella nubilosa (Cooke) Hansf., Proc. Linn. Soc. New South<br />
Wales 81: 36. 1965.<br />
= Mycosphaerella juvenis Crous & M.J. Wingf., Mycologia 88: 453. 1996.<br />
Teratosphaeria ohnowa (Crous & M.J. Wingf.) Crous & U. Braun,<br />
comb. nov. MycoBank MB504486.<br />
Basionym: Mycosphaerella ohnowa Crous & M.J. Wingf., Stud.<br />
Mycol. 50: 206. 2004.<br />
Teratosphaeria parkiiaffinis (Crous & M.J. Wingf.) Crous & U.<br />
Braun, comb. nov. MycoBank MB504487.<br />
Basionym: Mycosphaerella parkiiaffinis Crous & M.J. Wingf., Fungal<br />
Diversity 26: 168. 2007.<br />
Teratosphaeria parva (R.F. Park & Keane) Crous & U. Braun,<br />
comb. nov. MycoBank MB504488.<br />
Basionym: Mycosphaerella parva R.F. Park & Keane, Trans. Brit.<br />
Mycol. Soc. 79: 99. 1982.<br />
= Mycosphaerella gr<strong>and</strong>is Carnegie & Keane, Mycol. Res. 98: 414. 1994.<br />
Teratosphaeria perpendicularis (Crous & M.J. Wingf.) Crous & U.<br />
Braun, comb. nov. MycoBank MB504489.<br />
Basionym: Mycosphaerella perpendicularis Crous & M.J. Wingf.,<br />
Stud. Mycol. 55: 113. 2006.<br />
Teratosphaeria pluritubularis (Crous & Mansilla) Crous & U.<br />
Braun, comb. nov. MycoBank MB504490.<br />
Basionym: Mycosphaerella pluritubularis Crous & Mansilla, Stud.<br />
Mycol. 55: 114. 2006.<br />
10
Phylogenetic lineages in the Capnodiales<br />
Teratosphaeria pseudafricana (Crous & T.A. Cout.) Crous & U.<br />
Braun, comb. nov. MycoBank MB504491.<br />
Basionym: Mycosphaerella pseudafricana Crous & T.A. Cout.,<br />
Stud. Mycol. 55: 115. 2006.<br />
Teratosphaeria pseudocryptica (Crous) Crous & U. Braun, comb.<br />
nov. MycoBank MB504492.<br />
Basionym: Mycosphaerella pseudocryptica Crous, Stud. Mycol. 55:<br />
6. 2006.<br />
Anamorph: Readeriella sp.<br />
Teratosphaeria pseudosuberosa (Crous & M.J. Wingf.) Crous &<br />
U. Braun, comb. nov. MycoBank MB504493.<br />
Basionym: Mycosphaerella pseudosuberosa Crous & M.J. Wingf.,<br />
Stud. Mycol. 55: 118. 2006.<br />
Anamorph: Catenulostroma sp.<br />
Teratosphaeria quasicercospora (Crous & T.A. Cout.) Crous & U.<br />
Braun, comb. nov. MycoBank MB504494.<br />
Basionym: Mycosphaerella quasicercospora Crous & T.A. Cout.,<br />
Stud. Mycol. 55: 119. 2006.<br />
Teratosphaeria readeriellophora (Crous & Mansilla) Crous & U.<br />
Braun, comb. nov. MycoBank MB504495.<br />
Basionym: Mycosphaerella readeriellophora Crous & Mansilla,<br />
Stud. Mycol. 50: 207. 2004.<br />
Anamorph: Readeriella readeriellophora Crous & Mansilla, Stud.<br />
Mycol. 50: 207. 2004. Fig. 18.<br />
Teratosphaeria secundaria (Crous & Alfenas) Crous & U. Braun,<br />
comb. nov. MycoBank MB504496.<br />
Basionym: Mycosphaerella secundaria Crous & Alfenas, Stud.<br />
Mycol. 55: 122. 2006.<br />
Teratosphaeria stramenticola (Crous & Alfenas) Crous & U.<br />
Braun, comb. nov. MycoBank MB504497.<br />
Basionym: Mycosphaerella stramenticola Crous & Alfenas, Stud.<br />
Mycol. 55: 123. 2006.<br />
Teratosphaeria suberosa (Crous, F.A. Ferreira, Alfenas & M.J.<br />
Wingf.) Crous & U. Braun, comb. nov. MycoBank MB504498.<br />
Basionym: Mycosphaerella suberosa Crous, F.A. Ferreira, Alfenas<br />
& M.J. Wingf., Mycologia 85: 707. 1993.<br />
Teratosphaeria suttonii (Crous & M.J. Wingf.) Crous & U. Braun,<br />
comb. nov. MycoBank MB504499.<br />
Basionym: Mycosphaerella suttonii Crous & M.J. Wingf. (suttoniae),<br />
Canad. J. Bot. 75: 783. 1997.<br />
Anamorph: Readeriella epicoccoides (Cooke & Massee) Crous &<br />
U. Braun, comb. nov. MycoBank MB504500.<br />
Basionym: Cercospora epicoccoides Cooke & Massee apud Cooke,<br />
Grevillea 19: 91. 1891.<br />
≡ Phaeophleospora epicoccoides (Cooke & Massee) Crous, F.A. Ferreira<br />
& B. Sutton, S. African J. Bot. 63: 113. 1997.<br />
≡ Kirramyces epicoccoides (Cooke & Massee) J. Walker, B. Sutton &<br />
Pascoe, Mycol. Res. 96: 919. 1992.<br />
= Hendersonia gr<strong>and</strong>ispora McAlp., Proc. Linn. Soc. New South Wales 28:<br />
99. 1903.<br />
= Phaeoseptoria eucalypti Hansf., Proc. Linn. Soc. New South Wales 82: 225.<br />
1957.<br />
= Phaeoseptoria luzonensis T. Kobayashi, Trans. Mycol. Soc. Japan 19: 377.<br />
1978.<br />
Synanamorph: Pseudocercospora sp.<br />
Teratosphaeria toledana (Crous & Bills) Crous & U. Braun, comb.<br />
nov. MycoBank MB504501.<br />
Basionym: Mycosphaerella toledana Crous & Bills, Stud. Mycol. 50:<br />
208. 2004.<br />
Anamorph: Readeriella toledana (Crous & Bills) Crous & U. Braun,<br />
comb. nov. MycoBank MB504502.<br />
Basionym: Phaeophleospora toledana Crous & Bills, Stud. Mycol.<br />
50: 208. 2004.<br />
Key to treated anamorph genera of Teratosphaeria (Teratosphaeriaceae)<br />
1. Hyphae submerged to superficial, disarticulating into arthroconidia .......................................................................................................... 2<br />
1. Hyphae not disarticulating into arthroconidia ............................................................................................................................................. 3<br />
2. Mature, brown hyphae disarticulating into thick-walled, spherical, smooth to verruculose 0(–2) transversely septate,<br />
brown conidia ............................................................................................................................ Pseudotaeniolina (= Friedmanniomyces)<br />
2. Hyphae superficial, brown to green-brown, smooth, disarticulating to form pale brown, cylindrical, 0–3-septate conidia with subtruncate<br />
ends, frequently with a Readeriella synanamorph ....................................................................................................................... Cibiessia<br />
3. Hyphal ends forming endoconidia; hyphae pale to medium brown, verruculose, end cells dividing into several brown, verruculose,<br />
thick-walled, ellipsoid to obovoid endoconidia ................................................................................................................. Phaeothecoidea<br />
3. Endoconidia absent.................................................................................................................................................................................... 4<br />
4. Conidiogenous cells integrated in hyphae; well-developed conidiomata or long, solitary, macronematous, terminally penicillate<br />
conidiophores absent ................................................................................................................................................................................. 5<br />
4. Conidiomata well-developed or with long, solitary, terminally penicillate conidiophores ........................................................................... 7<br />
5. Conidia in chains, holoblastic, pseudocladosporium-like in morphology, but scars <strong>and</strong> hila not excessively thickened, nor refractive,<br />
producing chlamydospores in culture; species are mostly heat resistant .................................................................................... Devriesia<br />
5. Conidia solitary on indistict to well defined phialides on hyphae ............................................................................................................... 6<br />
6. Conidiogenous cells integrated in the distal ends of hyphae; conidia thick-walled, brown, smooth,<br />
-septate ......................................................................................................................................................................... Capnobotryella<br />
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Crous et al.<br />
6. Conidiophores short <strong>and</strong> frequently reduced to conidiogenous cells that proliferate percurrently via wide necks, giving rise to hyaline, 0(–2)-<br />
septate, broadly ellipsoidal conidia ................................................................................................................................................ Hortaea<br />
7. Conidia brown, with hyaline basal appendages; conidiomata pycnidial, conidiogenous cells phialidic, but also percurrent,<br />
subhyaline ....................................................................................................................................................................... Nothostrasseria<br />
7. Conidia brown, but basal appendages lacking, amero- to scolecospores ................................................................................................. 8<br />
8. Conidiomata pycnidial to acervular ............................................................................................................................................................ 9<br />
8. Conidiomata not enclosed by host tissue, fasciculate to sporodochial or solitary, hyphomycetous ........................................................ 10<br />
9. Conidia solitary, dry, without mucilaginous sheath .................................................................................................................... Readeriella<br />
9. Conidia catenulate, with persistant mucilaginous sheath ..................................................................................................... Staninwardia<br />
10. Conidiophores usually solitary, rarely densely fasciculate to synnematous (in vivo), penicillate, with a branched, apical conidiogenous<br />
apparatus giving rise to ramoconidia <strong>and</strong> branched chains of secondary conidia; scars not to slightly thickened <strong>and</strong> darkened-refractive<br />
..................................................................................................................................................................................................... Penidiella<br />
10. Conidiophores not penicillate, without a branched conidiogenous apparatus, in vivo fasciculate to sporodochial .................................. 11<br />
11. Biotrophic; fruiting composed of sporodochia <strong>and</strong> radiating layers of hyphae arising from the stromata, conidiophores arising from<br />
superficial sporodochia <strong>and</strong> radiating hyphae, conidiogenous cells unilocal, with conspicuous annellations, conidia solitary or in fragile<br />
disarticulating chains, aseptate or transversely 1–3-septate, usually with distinct frills, secession rhexolytic ............... Batcheloromyces<br />
. Biotrophic, leaf-inhabiting, with distinct, subepidermal to erumpent, well-developed sporodochia, or saxicolous, saprobic, sometimes<br />
causing opportunistic human infections; radiating layers of hyphae arising from sporodochia; conidiogenous cells without<br />
annellations; conidia in true simple or branched basipetal chains, transversely 1- to pluriseptate or with longitudinal <strong>and</strong> oblique septa<br />
(dictyosporous), occasionally distoseptate ...................................................................................................................... Catenulostroma<br />
To explain the arguments behind the selection <strong>and</strong> synonymies<br />
of some of these anamorphic genera, they are briefly discussed<br />
below:<br />
Acidomyces Baker et al., Appl. Environ. Microbiol. 70: 6270. 2004.<br />
(nom. inval.)<br />
Type species: Acidomyces richmondensis Baker et al., Appl.<br />
Environ. Microbiol. 70: 6270. 2004. (nom. inval.)<br />
Notes: <strong>The</strong> <strong>genus</strong> presently clusters among isolates in the<br />
Teratosphaeria clade based on sequences deposited in GenBank.<br />
Acidomyces lacks a Latin description <strong>and</strong> holotype specimen, <strong>and</strong><br />
is thus invalidly described. <strong>The</strong> <strong>genus</strong>, which was distinguished<br />
from other taxa based on its DNA phylogeny (Dothideomycetes),<br />
forms filamentous hyphae with disarticulating cells. It is unclear<br />
how it differs from Friedmanniomyces Onofri <strong>and</strong> Pseudotaeniolina<br />
J.L. Crane & Schokn.<br />
Batcheloromyces Marasas, P.S. van Wyk & Knox-Dav., J. S.<br />
African Bot. 41: 41. 1975.<br />
Type species: Batcheloromyces proteae Marasas, P.S. van Wyk &<br />
Knox-Dav., J. S. African Bot. 41: 43. 1975.<br />
Description: Crous et al. (2004a; figs 4–26).<br />
Notes: Batcheloromyces is presently circumscribed as a <strong>genus</strong><br />
that forms emergent hyphae, giving rise to superficial sporodochial<br />
plates, forming brown, verrucose, erect conidiophores that<br />
proliferate holoblastically, with ragged percurrent proliferations that<br />
become visible with age. Conidia are produced singly or in fragile,<br />
disarticulating chains, are brown, thick-walled, 0–3 transversely<br />
euseptate (though at times they appear as distoseptate). <strong>The</strong> <strong>genus</strong><br />
Batcheloromyces has in recent years been confused with Stigmina<br />
(Sutton & Pascoe 1989) on the basis that some collections showed<br />
conidiophores to give rise to solitary conidia only, though conidial<br />
catenulation was clearly illustrated by Taylor et al. (1999). In culture<br />
colonies tend to sporulate in a slimy mass (on OA), though a<br />
synanamorph can be seen (in B. leucadendri, Fig. 4) to sporulate<br />
via holoblastic conidiogenesis on hyphal tips of the aerial mycelium,<br />
forming elongate-globose to ellipsoid, muriformly septate, thickwalled<br />
conidia, that occur in clusters.<br />
<strong>The</strong> finding that Stigmina s. str. [based on S. platani (Fuckel)<br />
Sacc., the type species] is a generic synonym of Pseudocercospora<br />
Speg. (Crous et al. 2006a), <strong>and</strong> that the type species of<br />
Trimmatostroma (T. salicis, Fig. 5) belongs to the Helotiales<br />
(Fig. 1), raises the question of where to place stigmina- <strong>and</strong><br />
trimmatostroma-like anamorphs that reside in the Teratosphaeria<br />
clade. Although the stigmina-like species can be accommodated<br />
in Batcheloromyces (see Sutton & Pascoe 1989), a new <strong>genus</strong> is<br />
required for Teratosphaeria anamorphs that have a trimmatostromalike<br />
morphology. <strong>The</strong> recognition of Batcheloromyces <strong>and</strong> the<br />
introduction of a new anamorph <strong>genus</strong> for trimmatostroma-like<br />
anamorphs of Teratosphaeria are also morphologically justified.<br />
Batcheloromyces is easily distinguishable from Stigmina s.<br />
str. by its special structure of the fruiting body, composed of<br />
sporodochia <strong>and</strong> radiating layers of hyphae arising from the<br />
sporodochia <strong>and</strong> the conidia often formed in delicate disarticulating<br />
chains. Trimmatostroma-like anamorphs of Teratosphaeria are<br />
morphologically also sufficently distinct from Trimmatostroma s. str.<br />
(see notes under Catenulostroma Crous & U. Braun) as well as<br />
Batcheloromyces (see key above).<br />
Batcheloromyces eucalypti (Alcorn) Crous & U. Braun, comb.<br />
nov. MycoBank MB504503.<br />
Basionym: Stigmina eucalypti Alcorn, Trans. Brit. Mycol. Soc. 60:<br />
151. 1973.<br />
Capnobotryella Sugiy., in Sugiyama, Pleomorphic Fungi: <strong>The</strong><br />
Diversity <strong>and</strong> its Taxonomic Implications (Tokyo): 148. 1987.<br />
Type species: Capnobotryella renispora Sugiy., in Sugiyama,<br />
Pleomorphic Fungi: <strong>The</strong> Diversity <strong>and</strong> its Taxonomic Implications<br />
(Tokyo): 148. 1987.<br />
Description: Sugiyama & Amano (1987, figs 7.5–7.8).<br />
12
Phylogenetic lineages in the Capnodiales<br />
Fig. 4. Batcheloromyces leucadendri in vitro. A–B. Batcheloromyces state with synanamorph (arrows). C–D. Conidia occurring solitary or in short chains. Scale bar = 10 µm.<br />
Fig. 5. Trimmatostroma salicis. A. Sporodochia on twig. B–E. Chains of disarticulating conidia. Scale bars = 10 µm.<br />
Notes: <strong>The</strong> <strong>genus</strong> forms brown, septate, thick-walled hyphae, with<br />
ellipsoidal, 0–1-septate conidia forming directly on the hyphae, via<br />
minute phialides. Hambleton et al. (2003) also noted the occurrence<br />
of endoconidiation.<br />
Catenulostroma Crous & U. Braun, gen. nov. MycoBank<br />
MB504474.<br />
Etymology: Named after its catenulate conidia, <strong>and</strong> stromata giving<br />
rise to sporodochia.<br />
Hyphomycetes. Differt a Trimmatostromate habitu phytoparasitico, maculis<br />
formantibus, conidiophoris saepe fasciculatis, per stoma emergentibus vel habitu<br />
saxiphilo-saprophytico, interdum sejunctis ex mycosibus humanis.<br />
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Crous et al.<br />
Habit plant pathogenic, leaf-spotting or saxicolous-saprobic,<br />
occasionally isolated from opportunistic human mycoses.<br />
Mycelium internal <strong>and</strong> external; hyphae dark brown, septate,<br />
branched. Conidiomata in vivo vary from acervuli to sporodochia or<br />
fascicles of conidiophores arising from well-developed or reduced,<br />
pseudoparenchymatal stromata. Setae <strong>and</strong> hyphopodia absent.<br />
Conidiophores arising from hyphae or stromata, solitary, fasciculate<br />
to sporodochial, in biotrophic, plant pathogenic species emerging<br />
through stomata, little differentiated, semimacronematous, branched<br />
or not, continuous to septate, brown, smooth to verruculose.<br />
Conidiogenous cells integrated, terminal or conidiophores reduced<br />
to conidiogenous cells, holoblastic-thalloblastic, meristematic,<br />
unilocal, delimitation of conidium by a single septum with<br />
retrogressive delimitation of next conidium giving an unconnected<br />
chain of conidia, brown, smooth to verruculose, conidiogenous<br />
scars (conidiogenous loci) inconspicuous, truncate, neither<br />
thickened nor darkened. Conidia solitary or usually forming simple<br />
to branched basipetal chains of transversely to muriformly eu- or<br />
distoseptate, 1– to multiseptate, brown, smooth, verruculose to<br />
verrucose conidia, conidial secession schizolytic.<br />
Type species: Catenulostroma protearum (Crous & M.E. Palm)<br />
Crous & U. Braun, comb. nov.<br />
Description: Crous & Palm (1999), Crous et al. (2004a; figs 364–<br />
365).<br />
Notes: Catenulostroma contains several plant pathogenic species<br />
previously placed in Trimmatostroma, a morphologically <strong>similar</strong> but,<br />
based on its type species, phylogenetically distinct <strong>genus</strong> belonging<br />
to Helotiales (Fig. 1). Trimmatostroma s. str. is well-distinguished<br />
from most Catenulostroma species by being saprobic, living on<br />
twigs <strong>and</strong> branches of woody plants, or occasionally isolated<br />
from leaf litter, i.e., they are not associated with leaf spots. <strong>The</strong><br />
conidiomata of Trimmatostroma species are subepidermal,<br />
acervular-sporodochial with a well defined wall of textura angularis,<br />
little differentiated, micronematous conidiophores giving rise to long<br />
chains of conidia that disarticulate at the surface to form a greyblack<br />
to brown powdery mass. <strong>The</strong> generic affinity of other species<br />
assigned to Trimmatostroma, e.g. those having a lichenicolous<br />
habit, is unresolved.<br />
Trimmatostroma abietis Butin & Pehl (Butin et al. 1996) clusters<br />
together with the plant pathogenic Catenulostroma species, but<br />
differs from these species in having a more complex ecology.<br />
Trimmatostroma abietis is usually foliicolous on living or necrotic<br />
conifer needles on which characteristic acervuli to sporodochia<br />
with densely arranged, fasciculate fertile hyphae are formed,<br />
comparable to the fasciculate conidiomata of the plant pathogenic<br />
species of Catenulostroma (Butin et al. 1996: 205, fig. 1). Although<br />
not discussed by Butin et al. (1996), T. abietis needs to be compared<br />
to T. abietina Doherty, which was orginally described from Abies<br />
balsamea needles collected in Guelph, Canada (Doherty 1900).<br />
Morphologically the two species appear to be synonymous, except<br />
for reference to muriformly septate conidia, which is a feature not<br />
seen in vivo in the type of T. abietis. Furthermore, as this is clearly<br />
a species complex, this matter can only be resolved once fresh<br />
Canadian material has been collected to serve as epitype for T.<br />
abietina.<br />
Isolates from stone, agreeing with T. abietis in cultural,<br />
morphological <strong>and</strong> physiological characteristics, have frequently<br />
been found (Wollenzien et al. 1995, Butin et al. 1996, Gorbushina<br />
et al. 1996, Kogej et al. 2006, Krumbein et al. 1996). Furthermore,<br />
isolates from humans (ex skin lesions <strong>and</strong> ex chronic osteomycelitis<br />
of human patients) <strong>and</strong> Ilex leaves are known (Butin et al. 1996). De<br />
Hoog et al. (1999) included strains of T. abietis from stone, man <strong>and</strong><br />
Ilex leaves in molecular sequence analyses <strong>and</strong> demonstrated their<br />
genetical identity based on 5.8S rDNA <strong>and</strong> ITS2 data, but strains<br />
from conifer needles were not included. Furthermore, we consider<br />
T. abietis, as presently defined, to represent a species complex,<br />
with Dutch isolates from Pinus again appearing distinct from<br />
German Abies isolates, suggesting that different conifer genera<br />
could harbour different Catenulostroma species. Isolates from<br />
stone form stromatic, durable microcolonies, which are able to grow<br />
under extreme xerophilic environmental conditions. Cultural growth<br />
resembles that of other meristematic black yeasts (Butin et al. 1996,<br />
Kogej et al. 2006). Another fungus isolated from stone in Germany<br />
is in vitro morphologically close to C. abietis, but differs in forming<br />
conidia with oblique septa. Furthermore, a human pathogenic<br />
isolate from Africa clusters together with other Catenulostroma<br />
species. <strong>The</strong> habit <strong>and</strong> origin of this human pathogenic fungus in<br />
nature <strong>and</strong> its potential morphology on “natural” substrates, which<br />
typically deviates strongly from the growth in vitro, are still unknown.<br />
However, C. abietis, usually growing as a foliicolous <strong>and</strong> saxicolous<br />
fungus, has already shown the potential ability of Catenulostroma<br />
species to cause opportunistic human infections.<br />
Key to Catenulostroma species<br />
1. Conidia formed in basipetal chains, smooth, 4-celled, consisting of two basal cells with truncate lateral sides, each giving rise to a<br />
secondary globose apical cell, that can extend <strong>and</strong> develop additional septa, appearing as two lateral arms ................. C. excentricum<br />
1. Conidia variable in shape, but without two basal cells giving rise to two lateral arms ............................................................................... 2<br />
2. Conidia smooth or almost so, at most very faintly rough-walled; usually foliicolous on conifer needles or saxicolous, forming stromatic,<br />
xerophilic durable microcolonies on stone, occasionally causing opportunistic human infections ............................................................ 3<br />
2. Conidia distinctly verruculose to verrucose; plant pathogenic, forming leaf spots ..................................................................................... 5<br />
3. Conidia (8–)20–35(–60) × 4–5(–7) µm, 1–10-septate .................................................................................... C. chromoblastomycosum<br />
3. Conidia much shorter, 8–20 µm long, 0–5-septate .................................................................................................................................... 4<br />
4. Conidia 0–5 times transversely septate, mostly two-celled; usually foliicolous on conifer needles or saxicolous ....................... C. abietis<br />
4. Conidia 2–4 times transversely septate <strong>and</strong> often with 1–2 oblique septa; isolated from stone ........................................ C. germanicum<br />
5. Conidia rather broad, usually wider than 10 µm ........................................................................................................................................ 6<br />
14
Phylogenetic lineages in the Capnodiales<br />
Fig. 6. Catenulostroma chromoblastomycosum (type material). A. Sporodochium on pine needle in vitro. B–H. Chains of disarticulating conidia. Scale bars: A = 350, B, E, G,<br />
H = 10 µm.<br />
5. Conidia narrower, width below 10 µm ....................................................................................................................................................... 7<br />
6. Conidia distoseptate, rather long, (12–)25–35(–45) × (7–)10–15(–25) µm; conidiomata large, up to 250 µm diam, on Protea anceps<br />
............................................................................................................................................................................................... C. protearum<br />
6. Conidia euseptate, shorter, (9–)16–20(–36) × (10–)14–18(–27) µm; sporodochia 90–100 × 40–80 µm; on Protea gr<strong>and</strong>iceps<br />
................................................................................................................................................................................................. C. elginense<br />
7. Conidia 1- to multiseptate, (10–)15–17(–23) × (5–)6.5–7(–9) µm; on various Proteaceae .................................................. C. macowanii<br />
7. Conidia in vivo predominantly 1-septate, (8–)13–15(–21) × (3.5–)5.5–6(–8) µm; on Protea cynaroides<br />
......................................................................................................................................... C. microsporum (Teratosphaeria microspora)<br />
Catenulostroma abietis (Butin & Pehl) Crous & U. Braun, comb.<br />
nov. MycoBank MB504504.<br />
Basionym: Trimmatostroma abietis Butin & Pehl, Antonie van<br />
Leeuwenhoek 69: 204. 1996.<br />
Notes: Catenulostroma abietis needs to be compared to<br />
Trimmatostroma abietina Doherty (Abies balsamea needles<br />
Canada), which is either an older name for this species, or a closely<br />
related taxon. Presently T. abietina is not known from culture, <strong>and</strong><br />
needs to be recollected.<br />
Catenulostroma chromoblastomycosum Crous & U. Braun, sp.<br />
nov. MycoBank MB504505. Fig. 6.<br />
Etymology: Named after the disease symptoms observed due to<br />
opportunistic human infection.<br />
Differt a C. abieti et C. germanico conidiis longioribus, (8–)20–35(–60) × 4–5(–7)<br />
µm, 1–10-septatis.<br />
Description based on cultures sporulating on WA supplemented<br />
with sterile pine needles. Mycelium consisting of branched,<br />
septate, smooth to finely verruculose, medium to dark brown,<br />
thick-walled, 3–4 µm wide hyphae. Conidiomata brown, superficial,<br />
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Crous et al.<br />
Fig. 7. Catenulostroma germanicum (type material). A–D. Chains of disarticulating conidia in vitro. Scale bars = 10 µm.<br />
sporodochial, up to 350 µm diam. Conidiophores reduced to<br />
inconspicuous conidiogenous loci on hyphae, 2–4 µm wide, neither<br />
darkened nor thickened or refractive. Conidia occurring in branched<br />
chains, that tend to remain attached to each other, subcylindrical<br />
with subtruncate ends, straight to slightly curved, (8–)20–35(–60)<br />
× 4–5(–7) µm, 1–10-septate, medium brown, smooth to finely<br />
verruculose.<br />
Cultural characteristics: Colonies on PDA erumpent, spreading,<br />
slow growing, with sparse to moderate aerial mycelium <strong>and</strong> smooth,<br />
irregular, submerged margins; greenish black (surface).<br />
Specimen examined: Zaire, Pawa, isolated from man with chromoblastomycosis,<br />
Mar. 1997, V. de Brouwere, holotype <strong>CBS</strong> H-19935, culture ex-type <strong>CBS</strong> 597.97.<br />
Notes: Catenulostroma chromoblastomycosum was originally<br />
identified as an isolate of Stenella araguata Syd. <strong>The</strong> latter fungus<br />
is morphologically distinct, however, having much shorter <strong>and</strong><br />
narrower conidia, formed in acropetal chains, as well as quite<br />
different conidiogenous loci <strong>and</strong> conidial hila which are small,<br />
thickened <strong>and</strong> darkened.<br />
Catenulostroma elginense (Joanne E. Taylor & Crous) Crous &<br />
U. Braun, comb. nov. MycoBank MB504506.<br />
Basionym: Trimmatostroma elginense Joanne E. Taylor & Crous,<br />
Mycol. Res. 104: 633. 2000.<br />
Catenulostroma excentricum, see Teratosphaeria excentrica.<br />
Catenulostroma germanicum Crous & U. Braun, sp. nov.<br />
MycoBank MB504507. Fig. 7.<br />
Etymology: Named after the geographic location of its type strain<br />
in Germany.<br />
Differt a C. abieti conidiis 1–2 oblique septatis.<br />
Mycelium consisting of branched, septate, smooth, pale to<br />
medium brown, 2–4 µm wide hyphae, giving rise to conidial<br />
chains. Conidiophores integrated, subcylindrical, branched or not,<br />
septate, little differentiated, micronematous, 3–5 µm wide, 3- to<br />
multiseptate, medium brown, thick-walled; conidiogenous cells<br />
integrated, terminal, inconspicuous, unilocal, conidiogenous loci<br />
16
Phylogenetic lineages in the Capnodiales<br />
inconspicuous. Conidia in simple or branched basipetal chains,<br />
subcylindrical, straight to flexuous, (8–)10–15(–20) × 4–5(–6) µm,<br />
2–4 transversely septate or with 1–2 oblique septa, medium to dark<br />
brown, thick-walled, smooth.<br />
Cultural characteristics: Colonies on OA erumpent, spreading, with<br />
even, smooth margins <strong>and</strong> sparse to moderate aerial mycelium;<br />
olivaceous-grey, with iron-grey margins (surface). Colonies reaching<br />
12 mm diam after 1 mo at 25 °C in the dark; colonies fertile.<br />
Specimen examined: Germany (former West-Germany), isolated from stone, Oct.<br />
1988, J. Kuroczkin, holotype <strong>CBS</strong> H-19936, culture ex-type <strong>CBS</strong> 539.88.<br />
Notes: Catenulostroma germanicum was originally deposited as<br />
Taeniolina scripta (P. Karst.) P.M. Kirk. It is clearly distinct, however,<br />
as the latter fungus forms intricate, branched, brown conidia<br />
(Kirk 1981), unlike those of C. germanicum. Phylogenetically C.<br />
germanicum forms part of the C. abietis species complex.<br />
Catenulostroma macowanii (Sacc.) Crous & U. Braun, comb.<br />
nov. MycoBank MB504508.<br />
Basionym: Coniothecium macowanii Sacc., Syll. Fung. 4: 512.<br />
1886.<br />
≡ Coniothecium punctiforme G. Winter, Hedwigia 24: 33. 1885, non C.<br />
punctiforme Corda, Icones Fungorum (Prague) 1: 2. 1837.<br />
≡ Trimmatostroma macowanii (Sacc.) M.B. Ellis, More Dematiacous<br />
Hyphomycetes: 29. 1976.<br />
Catenulostroma microsporum, see Teratosphaeria microspora.<br />
Catenulostroma protearum (Crous & M.E. Palm) Crous & U.<br />
Braun, comb. nov. MycoBank MB504509.<br />
Basionym: Trimmatostroma protearum Crous & M.E. Palm, Mycol.<br />
Res. 103: 1303. 1999.<br />
Cibiessia Crous, Fungal Diversity 26: 151. 2007.<br />
Type species: Cibiessia dimorphospora Crous & C. Mohammed,<br />
Fungal Diversity 26: 151. 2007.<br />
Description: Crous et al. (2007b; figs 3–5).<br />
Notes: <strong>The</strong> <strong>genus</strong> Cibiessia was introduced to accommodate<br />
species with chains of disarticulating conidia (arthroconidia). Some<br />
species have been shown to have a Readeriella synanamorph.<br />
Devriesia Seifert & N.L. Nick., Can. J. Bot. 82: 919. 2004.<br />
Type species: Devriesia staurophora (W.B. Kendr.) Seifert & N.L.<br />
Nick., Canad. J. Bot. 82: 919. 2004.<br />
Description: Seifert et al. (2004; figs 2–42).<br />
Notes: <strong>The</strong> <strong>genus</strong> is characterised by producing chains of pale<br />
brown, subcylindrical to fusiform, 0–1-septate conidia with<br />
somewhat thickened, darkened hila, forming chlamydospores in<br />
culture, <strong>and</strong> being heat resistant. Morphologically they resemble<br />
taxa placed in Pseudocladosporium U. Braun (= Fusicladium<br />
Bonord.; Venturiaceae), though phylogenetically Devriesia is not<br />
allied to this family.<br />
Hortaea Nishim. & Miyaji, Jap. J. Med. Mycol. 26: 145. 1984.<br />
Type species: Hortaea werneckii (Horta) Nishim. & Miyaji, Jap. J.<br />
Med. Mycol. 26: 145. 1984.<br />
Description: de Hoog et al. (2000, illust. p. 721).<br />
www.studiesinmycology.org<br />
Notes: <strong>The</strong> <strong>genus</strong> forms brown, septate, thick-walled hyphae, with<br />
ellipsoidal, 0–1-septate (becoming muriformly septate), hyaline to<br />
pale brown conidia forming directly on the hyphae, via phialides<br />
with percurrent proliferation. Isolates of H. werneckii are restricted<br />
to tropical or subtropical areas, where they occur as halophilic<br />
saprobes, frequently being associated with tinea nigra of humans<br />
(de Hoog et al. 2000). <strong>The</strong> generic distinction with Capnobotryella<br />
is less clear, except that the latter tends to have darker, thick-walled<br />
conidia, <strong>and</strong> reduced, less prominent phialides.<br />
Penidiella Crous & U. Braun, gen. nov. MycoBank MB504463.<br />
Etymology: Named after its penicillate conidiophores.<br />
Differt a Periconiellae conidiophoris apice penicillato ex cellulis conidiogenis et<br />
ramoconidiis compositis, cellulis conidiogenis saepe 1–3(–4) locis conidiogenis,<br />
terminalibus vel subterminalibus, subdenticulatis, non vel subincrassatis, non vel<br />
leviter fuscatis-refractivis, ramoconidiis praesentibus, saepe numerosis, conidiis<br />
ramicatenatis.<br />
Mycelium consisting of branched, septate, smooth to verruculose,<br />
subhyaline to pale brown hyphae. Conidiophores macronematous,<br />
occasionally also with some micronematous conidiophores;<br />
macronematous conidiophores arising from superficial mycelium<br />
or stromata, solitary, fasciculate or in synnemata, erect, brown,<br />
thin- to thick-walled, smooth to finely verruculose; terminally<br />
penicillate, branched terminal part consisting of a conidiogenous<br />
apparatus composed of a series of conidiogenous cells <strong>and</strong>/or<br />
ramoconidia. Conidiogenous cells integrated, terminal, intercalary<br />
or pleurogenous, unbranched, pale to medium brown, smooth<br />
to finely verruculose, tapering to a flattened or rounded apical<br />
region or tips slightly inflated, polyblastic, sympodial, giving rise<br />
to a single or several sets of ramoconidia on different levels; with<br />
relatively few conidiogenous loci, 1–3(–4), terminal or subterminal,<br />
subdenticulate, denticle-like loci usually conical, terminally<br />
truncate, usually unthickened or at most very slightly thickened, not<br />
to slightly darkened or somewhat refractive. Conidia in branched<br />
acropetal chains. Ramoconidia 0–1-septate, pale to medium brown,<br />
smooth to verruculose, thin-walled, ellipsoidal, obovoid, fusiform,<br />
subcylindrical to obclavate; conidia subcylindrical, fusiform to<br />
ellipsoid-ovoid, 0–1-septate, pale olivaceous to brown, smooth to<br />
verruculose, thin-walled, catenate; hila truncate, unthickened or<br />
almost so, barely to somewhat darkened-refractive.<br />
Type species: Penidiella columbiana Crous & U. Braun, sp. nov.<br />
Notes: Three ramichloridium-like genera cluster within Capnodiales,<br />
namely Periconiella Sacc. [type: P. velutina (G. Winter) Sacc.],<br />
Ramichloridium Stahel ex de Hoog [type: R. apiculatum (J.H.<br />
Mill., Giddens & A.A. Foster) de Hoog] <strong>and</strong> Penidiella [type: P.<br />
columbiana Crous & U. Braun]. All three genera have brown,<br />
macronematous conidiophores with <strong>similar</strong> conidial scars. Within<br />
this complex, Ramichloridium is distinct in having a prominent<br />
rachis giving rise to solitary conidia. Periconiella <strong>and</strong> Penidiella<br />
are branched in the apical part of their conidiophores, <strong>and</strong> lack<br />
a rachis. In Periconiella conidia are solitary or formed in short,<br />
mostly simple chains, ramoconidia are lacking. <strong>The</strong> apical<br />
conidiogenous apparatus is composed of conidiogenous cells or<br />
branches with integrated, usually terminal conidiogenous cells,<br />
which are persistent. <strong>The</strong> conidiogenous cells are subcylindrical<br />
to somewhat clavate, usually not distinctly attenuated towards the<br />
tip, <strong>and</strong> have several, often numerous loci, aggregated or spread<br />
over the whole cell, terminal to usually lateral, flat, non-protuberant,<br />
not denticle-like, usually distinctly thickened <strong>and</strong> darkened, at least<br />
at the rim. In contrast, Penidiella has a quite distinct branching<br />
17
Crous et al.<br />
system, consisting of a single terminal conidiogenous cell giving<br />
rise to several ramoconidia that form secondary ramoconidia, etc.,<br />
or the branched apparatus is composed of several terminal <strong>and</strong><br />
sometimes lateral conidiogenous cells giving rise to sequences of<br />
ramoconidia (conidiogenous cells <strong>and</strong> ramoconidia are often barely<br />
distinguishable, with conidiogenous cells disarticulating, becoming<br />
ramoconidia). <strong>The</strong> branched apparatus may be loose to dense,<br />
metula-like. <strong>The</strong> conidiogenous cells have only few, usually 1–3<br />
(–4), terminal or subterminal subdenticulate loci, <strong>and</strong> ramoconidia<br />
are prominent <strong>and</strong> numerous, giving rise to branched chains of<br />
secondary conidia with flat-tipped hila. Some species of Penidiella<br />
with compact, metula-like branched apices are morphologically<br />
close to Metulocladosporiella Crous, Schroers, J.Z. Groenew., U.<br />
Braun & K. Schub. (Crous et al. 2006d). This <strong>genus</strong> encompasses<br />
two species of banana diseases belonging to Herpotrichiellaceae<br />
(Chaetothyriales), characterised by having conidiophore bases<br />
with rhizoid hyphal appendages <strong>and</strong> abundant micronematous<br />
conidiophores. Penidiella species with less pronounced penicillate<br />
apices, e.g. P. strumelloidea (Milko & Dunaev) Crous & U. Braun, are<br />
comparable with species of the <strong>genus</strong> Pleurotheciopsis B. Sutton<br />
(see Ellis 1976). <strong>The</strong> latter <strong>genus</strong> is distinct in having unbranched,<br />
often percurrently proliferating conidiophores, lacking ramoconidia<br />
<strong>and</strong> colourless conidia formed in simple chains.<br />
<strong>Cladosporium</strong> helicosporum R.F. Castañeda & W.B. Kendr.<br />
(Castañeda et al. 1997) is another penidiella-like fungus with<br />
terminally branched conidiophores, subdenticulate conidiogenous<br />
loci <strong>and</strong> conidia in long acropetal chains, but its affinity to Penidiella<br />
has still to be proven.<br />
Key to Penidiella species<br />
1. Conidiophores in vivo in well-developed, dense fascicles <strong>and</strong> distinct synnemata arising from a basal stroma; on fallen leaves of Ficus<br />
sp., Cuba ............................................................................................................................................................................... P. cubensis<br />
1. Conidiophores solitary, at most loosely aggregated .................................................................................................................................. 2<br />
2. Conidiophores with a terminal conidiogenous cell, often somewhat swollen, giving rise to several ramoconidia (on one level) that form<br />
chains of straight to distinctly curved conidia; isolated from leaf of Carex sp., Russia ..................................................... P. strumelloidea<br />
2. Penicillate apex of the conidiophores composed of a system of true branchlets, conidiogenous cells <strong>and</strong> ramoconidia or at least a sequence<br />
of ramoconidia on several levels; conidia usually straight ......................................................................................................................... 3<br />
3. Mycelium verruculose; long filiform conidiophores ending with a subdenticulate cell giving rise to sets of penicillate conidiogenous cells or<br />
ramoconidia which are barely distinguishable <strong>and</strong> turn into each other; ramoconidia <strong>and</strong> conidia consistently narrow, (1.5–)2(–2.5) µm<br />
wide, <strong>and</strong> aseptate, ramoconidia sometimes heterochromous; on living leaves of Nect<strong>and</strong>ra coriacea, Cuba ................... P. nect<strong>and</strong>rae<br />
3. Mycelium more or less smooth; penicillate apex at least partly with true branchlets; conidia wider, 2–5 µm, at least partly septate, uniformly<br />
pigmented ................................................................................................................................................................................................. 4<br />
4. Hyphae, conidiophores <strong>and</strong> conidia frequently distinctly constricted at the septa; penicillate apex of the conidiophores sparingly developed,<br />
branchlets more or less divergent; isolated from leaf litter of Smilax sp., Cuba .................................................................. P. rigidophora<br />
4. Hyphae <strong>and</strong> conidia without distinct constrictions at the septa; penicillate apex of the conidiophores usually well-developed, with abundant<br />
branchings ................................................................................................................................................................................................. 5<br />
5. Conidiophores short, up to 120 × 3–4 µm, frequently with intercalary conidiogenous cell, swollen at the conidiogenous portion just below<br />
the upper septum which render the conidiophores subnodulose to distinctly nodulose, apex ± loosely penicillate; conidia (4–)5–7(–8) µm<br />
long; occasionally with micronematous conidiophores; isolated from man with tinea nigra, Venezuela .......................... P. venezuelensis<br />
5. Conidiophores much longer, up to 800 µm, 7–9 µm wide at the base, not distinctly nodulose, penicillate apex loose to often more<br />
compact, tight, metula-like; conidia longer, 7–25 × 2–5 µm; micronematous conidiophores lacking; isolated from dead leaf of Paepalanthus<br />
columbianus, Colombia ........................................................................................................................................................ P. columbiana<br />
Penidiella columbiana Crous & U. Braun, sp. nov. MycoBank<br />
MB504510. Figs 8–9.<br />
Etymology: Named after its country of origin, Colombia.<br />
Mycelium ex hyphis ramosis, septatis, levibus, pallide brunneis, 2–3 µm latis<br />
compositum. Conidiophora ex hyphis superficialibus oriunda, penicillata, erecta,<br />
brunnea, crassitunicata, minute verruculosa, ad 800 µm longa, ad basim 7–9<br />
µm lata, ad apicem pluriramosa, ex ramibus diversibus et cellulis conidiogenis<br />
composita, ramibus primariis (–2) subcylindraceis, 1–7-septatis, 50–120 × 4–6 µm;<br />
ramibus secundariis (–2) subcylindraceis, 1–5-septatis, 40–60 × 4–6 µm; ramibus<br />
tertiariis et subsequentibus 1–4-septatis, 10–30 × 3–5 µm. Cellulae conidiogenae<br />
terminales vel laterales, non ramosae, 5–15 × 3–5 µm, modice brunneae, minute<br />
verruculosae, apicem versus attenuatae, truncatae vel rotundatae, polyblasticae,<br />
sympodiales, cicatrices conidiales incrassatae, sed leviter fuscatae et non<br />
refractivae. Ramoconidia 0–1-septata, modice brunnea, levia, ellipsoidea, obclavata<br />
vel obovoidea, cum 1–3 hilis terminalibus, 10–20 × 3–5 µm; conidia subcylindrica<br />
vel ellipsoidea, 0–1-septata, pallide brunnea, catenata (–10), hila truncata, non<br />
incrassata, vix vel leviter fuscata.<br />
Mycelium consisting of branched, septate, smooth, pale brown, 2–3<br />
µm wide hyphae. Conidiophores arising from superficial mycelium,<br />
terminally penicillate, erect, brown, wall up to 1 µm wide, almost<br />
smooth to finely verruculose, up to 800 µm tall, 7–9 µm wide at<br />
the base; conidiogenous region consisting of a series of branches<br />
composed of true branchlets, conidiogenous cells <strong>and</strong> ramoconidia,<br />
branched portion usually rather compact, even metula-like, but<br />
also looser, with divergent branches; primary branches (–2),<br />
subcylindrical, 1–7-septate, 50–120 × 4–6 µm; secondary branches<br />
(–2), subcylindrical, 1–5-septate, 40–60 × 4–6 µm; tertiary <strong>and</strong><br />
additional branches 1–4-septate, 10–30 × 3–5 µm. Conidiogenous<br />
cells terminal, intercalary or lateral, unbranched, 5–15 × 3–5<br />
µm, medium brown, finely verruculose, tapering to a flattened or<br />
rounded (frequently swollen) apical region, scars thickened, but<br />
only somewhat darkened, not refractive. Ramoconidia 0–1-septate,<br />
18
Phylogenetic lineages in the Capnodiales<br />
Fig. 8. Penidiella columbiana (type material). A. Conidiophores on pine needle in vitro. B–H. Conidiophores with chains of disarticulating conidia. Scale bars: A = 450, B–C =<br />
10 µm.<br />
medium brown, smooth, wall ≤ 1 µm wide, ellipsoidal to obclavate<br />
or obovoid, with 1–3 apical hila, 10–25 × 3–5 µm, ramoconidia<br />
with broadly truncate base, not or barely attenuated, up to 4 µm<br />
wide, or at least somewhat attenuated at the base, hila 1.5–3 µm<br />
wide. Conidia subcylindrical to ellipsoid, 0(–1)-septate, pale brown,<br />
in chains of up to 10, 7–15 × 2–3 µm, hila truncate, unthickened,<br />
barely to somewhat darkened, 1–2 µm wide.<br />
Cultural characteristics: Colonies on PDA erumpent, spreading,<br />
with moderate aerial mycelium <strong>and</strong> smooth, even, submerged<br />
margins; olivaceous-grey in central part, iron-grey in outer region<br />
(surface); colonies fertile.<br />
Specimen examined: Colombia, Páramo de Guasca, 3400 m alt., isolated from<br />
dead leaf of Paepalanthus columbianus (Eriocaulaceae), Aug. 1980, W. Gams,<br />
holotype <strong>CBS</strong> H-19937, culture ex-type <strong>CBS</strong> 486.80.<br />
Notes: This isolate was originally identified as belonging to the<br />
Stenella araguata species complex. <strong>The</strong> latter name has been<br />
somewhat confused in the literature, <strong>and</strong> has been incorrectly<br />
applied to isolates associated with opportunistic human infections<br />
(de Hoog et al. 2000). <strong>The</strong> “araguata” species complex is treated<br />
elsewhere in the volume (see Crous et al. 2007a – this volume).<br />
www.studiesinmycology.org<br />
Penidiella cubensis (R.F. Castañeda) U. Braun, Crous & R.F.<br />
Castañeda, comb. nov. MycoBank MB504511. Fig. 10.<br />
Basionym: <strong>Cladosporium</strong> cubense R.F. Castañeda, Fungi Cubenses<br />
II (La Habana): 4. 1987.<br />
In vivo: Colonies on fallen leaves, amphigenous, effuse, pilose,<br />
brown. Mycelium usually external, superficial, but also internal,<br />
composed of branched, septate, brown, thin-walled, smooth to<br />
rough-walled hyphae, 2–3 µm wide. Stromata present, 40–80 µm<br />
diam, brown, immersed. Conidiophores densely fasciculate or in<br />
distinct synnemata, arising from stromata, erect, synnemata up<br />
to about 1000 µm long <strong>and</strong> (10–)20–40(–50) µm wide, individual<br />
threads filiform, pluriseptate throughout, brown, thin-walled (≤<br />
0.5 µm), smooth or almost so to distinctly verruculose, apically<br />
penicillate. Conidiogenous cells integrated, terminal <strong>and</strong> intercalary,<br />
10–30 µm long, subcylindrical, terminal conidiogenous cells often<br />
slightly enlarged at the tip, with (1–)2–3(–4) terminal or subterminal<br />
subdenticulate conidiogenous loci, short conically truncate, 1–2<br />
µm diam, unthickened or almost so, but often slightly refractive or<br />
darkened-refractive, intercalary conidiogenous cells usually with a<br />
single lateral locus just below the upper septum, conidiogenous cells<br />
giving rise to a single set of primary ramoconidia, or a sequence<br />
of ramoconidia at different levels. Ramoconidia cylindrical to<br />
ellipsoid-fusoid, 8–18(–25) × 2–3 µm, aseptate, pale olivaceous,<br />
olivaceous-brown to brown, thin-walled, smooth or almost so to<br />
19
Crous et al.<br />
Fig. 9. Penidiella columbiana (type material). A.<br />
Conidiophores. B. Ramoconidia. C. Secondary conidia.<br />
Scale bar = 10 µm. U. Braun del.<br />
faintly verruculose, ramoconidia with broadly truncate base, barely<br />
narrowed, or ramoconidia more or less attenuated at the base, hila<br />
1–2 µm wide, unthickened or almost so, but often slightly refractive<br />
or darkened-refractive. Conidia in long acropetal chains, narrowly<br />
ellipsoid-ovoid, fusiform, 5–12(–15) × (1–)1.5–3 µm, aseptate, pale<br />
olivaceous to brownish, thin-walled, smooth to faintly rough-walled,<br />
ends attenuated, hila 1–1.5 µm wide, unthickened, not darkened,<br />
at most somewhat refractive.<br />
Specimen examined: Cuba, Guantánamo, Maisí, on fallen leaves of Ficus sp., 24<br />
Apr. 1986, M. Camino, holotype INIFAT C86/134 (HAL 2019 F, ex holotype).<br />
Notes: <strong>Cladosporium</strong> cubense was not available in culture <strong>and</strong><br />
molecular sequence data are not available, but type material could<br />
be re-examined <strong>and</strong> revealed that this species is quite distinct from<br />
<strong>Cladosporium</strong> s. str., but agreeing with the concept of the <strong>genus</strong><br />
Penidiella. Penidiella cubensis differs from all other species of this<br />
<strong>genus</strong> in having densely fasciculate conidiophores to synnematous<br />
conidiomata, arising from stromata.<br />
Penidiella nect<strong>and</strong>rae Crous, U. Braun & R.F. Castañeda, nom.<br />
nov. MycoBank MB504512. Fig. 11.<br />
Basionym: <strong>Cladosporium</strong> ferrugineum R.F. Castañeda, Fungi<br />
Cubenses II (La Habana): 4. 1987, homonym, non C. ferrugineum<br />
Allesch., 1895.<br />
In vivo: Colonies amphigenous, brown. Mycelium internal <strong>and</strong><br />
external, superficial, composed of sparingly branched hyphae,<br />
septate, 1–3 µm wide, pale olivaceous-brown or brown, thinwalled<br />
(≤ 0.5 µm), smooth or almost so to distinctly verruculose,<br />
fertile cells giving rise to conidiophores somewhat swollen at the<br />
branching point, up to 5 µm diam, <strong>and</strong> somewhat darker. Stromata<br />
lacking. Conidiophores erect, straight, filiform, up to 350 µm long,<br />
2.5–4 µm wide, pluriseptate throughout, brown, darker below <strong>and</strong><br />
paler above, thin-walled, smooth, apex penicillate, terminal cell of<br />
the conidiophore with 2–4 short denticle-like loci giving rise to sets<br />
of conidiogenous cells or ramoconidia that then form a sequence of<br />
new sets of ramoconidia on different levels, i.e., the loose to dense,<br />
metula-like branching system is composed of conidiogenous cells<br />
<strong>and</strong> ramoconidia which are often barely distinguishable <strong>and</strong> turn<br />
into each other; conidiogenous loci terminal or subterminal, usually<br />
1–3(–4), subdenticulate, 1–2 µm diam, conical, apically truncate,<br />
unthickened or almost so, not to somewhat darkened-refractive.<br />
Ramoconidia with truncate base, barely attenuated, or ramoconidia<br />
distinctly attenuated at the truncate base, up to 20 × 2 µm, aseptate,<br />
at the apex with 2–3(–4) subdenticulate hila, subcylindrical,<br />
20
Phylogenetic lineages in the Capnodiales<br />
very pale olivaceous, olivaceous-brown to brown, sometimes<br />
with different shades of brown (heterochromatous), thin-walled<br />
(≤ 0.5 µm), smooth to faintly verruculose. Conidia in long acropetal<br />
chains, narrowly ellipsoid-ovoid, fusiform to cylindrical, 5–16 ×<br />
(1.5–)2(–2.5) µm, aseptate, very pale olivaceous, olivaceousbrown<br />
to brown, thin-walled, smooth to very faintly rough-walled,<br />
primary conidia with rounded apex <strong>and</strong> trunacte base, somewhat<br />
attenuated, secondary conidia truncate at both ends, hila 1–1.5<br />
µm diam, unthickened or almost so, at most slightly darkenedrefractive.<br />
Cultural characteristics: Colonies on PDA slimy, smooth, spreading;<br />
aerial mycelium absent, margins smooth, irregular; surface black<br />
with patches of cream. Colonies reaching 20 mm diam after 1 mo<br />
at 25 °C in the dark; colonies sterile on PDA, SNA <strong>and</strong> OA.<br />
Specimen examined: Cuba, Matanzas, San Miguel de los Baños, isolated from<br />
living leaves of Nect<strong>and</strong>ra coriacea (Lauraceae), 24 Jan. 1987, R.F. Castañeda <strong>and</strong><br />
G. Arnold, holotype INIFAT C87/45, culture ex-type <strong>CBS</strong> 734.87, <strong>and</strong> HAL 2018 F<br />
(ex-holotype).<br />
Notes: Although the ex-type strain of <strong>Cladosporium</strong> ferrugineum<br />
is sterile, its LSU DNA phylogeny reveals it to be unrelated to<br />
<strong>Cladosporium</strong> s. str. (see Fig. 1 in Crous et al. 2007a – this volume).<br />
Based on a re-examination of the type material it could clearly be<br />
shown that the morphology of this species fully agrees with the<br />
concept of the new <strong>genus</strong> Penidiella, which is supported by its<br />
phylogenetic position within Capnodiales.<br />
Fig. 10. Penidiella cubensis (type material). A. Swollen stromatic base of synnema.<br />
B. Conidiophores. C. Ramoconidia. D. Secondary conidia. Scale bar = 10 µm.<br />
U. Braun del.<br />
Penidiella rigidophora Crous, R.F. Castañeda & U. Braun, sp.<br />
nov. MycoBank MB504513. Figs 12–13.<br />
≡ <strong>Cladosporium</strong> rigidophorum R.F. Castañeda, nom. nud. (herbarium<br />
name).<br />
Differt a specibus Penidiellae conidiophoris dimorphosis, hyphis et conidiis ad septa<br />
saepe distincte constrictis.<br />
Mycelium consisting of strongly branched, septate, smooth or<br />
almost so, pale olivaceous to medium brown, guttulate, commonly<br />
constricted at septa, 2–6 µm wide hyphae, swollen cells up to 8<br />
µm wide, wall up to 1(–1.5) µm wide. Conidiophores dimorphic.<br />
Macronematous conidiophores separate, erect, subcylindrical,<br />
predominantly straight to slightly curved, terminally loosely<br />
penicillate, up to 120 µm long, <strong>and</strong> 4–5 µm wide at the base, which<br />
is neither lobed nor swollen, <strong>and</strong> lacks rhizoids, up to 10-septate,<br />
medium to dark brown, wall up to 1(–1.5) µm wide. Micronematous<br />
conidiophores erect, subcylindrical, up to 40 µm tall, 3–4 µm wide,<br />
1–3-septate, pale to medium brown (concolorous with hyphae).<br />
Conidiogenous cells predominantly terminal, rarely intercalary,<br />
medium brown, smooth, subcylindrical, but frequently swollen at<br />
apex, 10–20 × 5–6 µm, loci (predominantly single in micronematous<br />
conidiophores, but up to 4 in macronematous conidiophores) flattipped,<br />
sub-denticulate or not, 1–1.5 µm wide, barely to slightly<br />
darkened <strong>and</strong> thickened-refractive. Conidia in branched chains,<br />
medium brown, verruculose, (appearing like small spines under<br />
light microscope), ellipsoid to cylindrical-oblong, up to 1(–1.5)<br />
µm wide, frequently constricted at septa, which turn dark with<br />
age; ramoconidia (10–)13–17(–25) × 3–4(–5) µm, 1(–3)-septate;<br />
secondary conidia (7–)8–10(–12) × 3–4(–5); hila unthickened to<br />
very slightly thickened <strong>and</strong> darkened, not refractive, (0.5–)1(–1.5)<br />
µm.<br />
Fig. 11. Penidiella nect<strong>and</strong>rae (type material). A. Conidiophores. B. Ramoconidia.<br />
C. Secondary conidia. Scale bar = 10 µm. U. Braun del.<br />
Cultural characteristics: Colonies on PDA erumpent, spreading,<br />
with lobate margins <strong>and</strong> moderate aerial mycelium; iron-grey<br />
(surface), with a greenish black margin; reverse greenish black.<br />
Colonies reaching 20 mm diam after 1 mo at 25 °C in the dark;<br />
colonies fertile.<br />
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21
Crous et al.<br />
Fig. 12. Penidiella rigidophora (type material). A–F. Micronematous conidiophores giving rise to chains of conidia. G–H. Macronematous conidiophores (note base in G, <strong>and</strong><br />
apex in H). I. Conidia. Scale bars = 10 µm.<br />
Specimen examined: Cuba, isolated from leaf litter of Smilax sp. (Smilacaceae), 6.<br />
Nov. 1994, R.F. Castañeda, holotype <strong>CBS</strong> H-19938, culture ex-type <strong>CBS</strong> 314.95.<br />
Notes: <strong>Cladosporium</strong> rigidophorum is a herbarium name, which was<br />
never validly published. <strong>The</strong> ex-type strain, however, represents a<br />
new species of Penidiella, for which a valid name with Latin diagnosis<br />
is herewith provided. This species is easily distinguishable from all<br />
other taxa of Penidiella by forming distinct constrictions at hyphal<br />
<strong>and</strong> conidial septa as well as micronematous conidiophores (except<br />
for P. venezuelensis in which a few micronematous conidiophores<br />
have been observed). It is also phylogenetically distinct from the<br />
other taxa of Penidiella (see Fig. 1 in Crous et al. 2007a – this<br />
volume).<br />
22
Phylogenetic lineages in the Capnodiales<br />
Penidiella strumelloidea (Milko & Dunaev) Crous & U. Braun,<br />
comb. nov. MycoBank MB504514. Figs 14–15.<br />
Basionym: <strong>Cladosporium</strong> strumelloideum Milko & Dunaev, Novosti<br />
Sist. Nizsh. Rast. 23: 134. 1986.<br />
Mycelium consisting of branched, septate, smooth, hyaline to<br />
pale olivaceous, 1–4 µm wide hyphae, sometimes constricted at<br />
somewhat darker septa. Conidiophores solitary, erect, arising from<br />
superficial mycelium, micronematous, i.e., reduced to conidiogenous<br />
cells, or macronematous, subcylindrical, straight to slightly curved,<br />
subcylindrical throughout or often somewhat attenuated towards the<br />
apex, 12–80 × (2–)2.5–4 µm, 0–6-septate, medium brown, smooth,<br />
wall ≤ 0.75 µm, penicillate apex formed by a terminal conidiogenous<br />
cell giving rise to a single set of ramoconidia. Conidiogenous cells<br />
terminal, integrated, subcylindrical, straight, 8–12 × 1.5–2(–2.5)<br />
µm, pale brown, thin-walled, smooth, apex obtusely rounded to<br />
somewhat clavate; loci terminal, occasionally subterminal or lateral,<br />
unthickened or almost so to slightly thickened <strong>and</strong> darkened, not<br />
refractive, 1–1.5 µm wide. Conidia in branched chains; ramoconidia<br />
subcylindrical, with 1–3 terminal loci, olivaceous-brown, smooth;<br />
secondary conidia ellipsoidal, with one side frequently straight <strong>and</strong><br />
the other convex, straight to slightly curved, (8–)10–12(–20) × 2(–<br />
3) µm, subhyaline to olivaceous-brown, smooth, thin-walled; hila<br />
unthickened or almost so to somewhat thickened <strong>and</strong> darkened,<br />
not refractive, 1 µm wide.<br />
Cultural characteristics: Colonies on PDA erumpent, spreading,<br />
with abundant, dense to woolly aerial mycelium, <strong>and</strong> uneven,<br />
feathery margins; surface pale olivaceous grey, reverse iron-grey.<br />
Colonies reaching 25 mm diam after 1 mo at 25 °C in the dark;<br />
colonies fertile.<br />
Fig. 13. Penidiella rigidophora (type material). A. Hyphae. B. Conidiophores. C.<br />
Ramoconidia. D. Secondary conidia. Scale bar = 10 µm. U. Braun del.<br />
Specimen examined: Russia, Yaroslavl Region, Rybinsk Reservoir, mouth of<br />
Sutka River, isolated from leaf of Carex sp. (Cyperaceae), from stagnant water, S.<br />
Ozerskaya, holotype BKMF-2534, culture ex-type <strong>CBS</strong> 114484.<br />
Fig. 14. Penidiella strumelloidea (type<br />
material). A–B. Micronematous conidiophores.<br />
C–D. Macronematous conidiophores. E–G.<br />
Conidia. Scale bars = 10 µm.<br />
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Crous et al.<br />
conidiophores, about 10–15 × 2–3 µm. Conidiogenous cells<br />
terminal <strong>and</strong> intercalary, unbranched, subcylindrical, 5–12 × 3–4<br />
µm, medium brown, smooth or almost so to finely verruculose,<br />
apex of conidiogenous cells frequently swollen, up to 6 µm diam,<br />
with 1–3(–4) flat-tipped, non to slightly thickened, non to slightly<br />
darkened-refractive loci, 1–1.5 µm wide, frequently appearing<br />
subdenticulate, up to 1.5 µm long, intercalary conidiogenous cells<br />
also slightly swollen at the conidiogenous portion just below the<br />
upper septum, which render the conidiophores subnodulose to<br />
nodulose, swellings round about the conidiophore axis or unilateral.<br />
Conidia ellipsoid-ovoid, subcylindrical, pale to medium olivaceousbrown<br />
or brown, finely verruculose, wall ≤ 0.5 µm wide, guttulate or<br />
not, occurring in branched chains. Ramoconidia 0–1(–3)-septate,<br />
5–15(–22) × 3–4(–5) µm, with 1–3 subdenticulate apical hila;<br />
secondary conidia 0(–1)-septate, ellipsoid, obovoid to irregular,<br />
(4–)5–7(–8) × (2–)2.5–3(–4) µm; hila non to slightly thickened, non<br />
to slightly darkened-refractive, (0.5–)1(–1.5) µm wide.<br />
Cultural characteristics: Colonies on OA erumpent, spreading,<br />
with dense, compact aerial mycelium, <strong>and</strong> even, smooth margins;<br />
olivaceous-grey (surface), margins iron-grey. Colonies reaching 22<br />
mm diam after 1 mo at 25 °C in the dark.<br />
Specimen examined: Venezuela, isolated from man with tinea nigra, Jan. 1975, D.<br />
Borelli, holotype <strong>CBS</strong> H-19934, culture ex-type <strong>CBS</strong> 106.75.<br />
Fig. 15. Penidiella strumelloidea (type material). A. Hyphae. B. Conidiophores. C.<br />
Ramoconidia. D. Secondary conidia. Scale bars = 10 µm. U. Braun del.<br />
Notes: Penidiella strumelloidea resembles other species of<br />
Penidiella by having penicillate conidiophores with a conidiogenous<br />
apparatus giving rise to branched conidial chains. It differs, however,<br />
from all other species of this <strong>genus</strong> in having a rather simple<br />
penicillate apex composed of a single terminal conidiogenous<br />
cell giving rise to one set of ramoconidia which form frequently<br />
somewhat curved conidia. It is also phylogenetically distinct from<br />
the other taxa of Penidiella (see Fig. 1 in Crous et al. 2007a – this<br />
volume).<br />
Penidiella venezuelensis Crous & U. Braun, sp. nov. MycoBank<br />
MB504515. Figs 16–17.<br />
Etymology: Named after the geographic location of its type strain,<br />
Venezuela.<br />
Differt a P. columbiana conidiophoris bevioribus et angustioribus, ad 120 × 3–4 µm,<br />
subnodulosis, apice plus minusve laxe penicillatis et conidiis brevioribus, (4–)5–7(–<br />
8) µm longis.<br />
Mycelium consisting of branched, septate, smooth to faintly<br />
rough-walled, thin-walled, subhyaline, pale olivaceous to medium<br />
brown, (1.5–)2–3 µm wide hyphae. Conidiophores solitary, erect,<br />
macronematous, subcylindrical, straight to flexuous to once<br />
geniculate, up to 120 µm long, 3–4 µm wide, 1–12-septate, pale to<br />
medium olivaceous-brown or brown, thin-walled (up to about 1 µm),<br />
terminally penicillate, branched portion composed of true branchlets<br />
<strong>and</strong>/or a single set or several sets of ramoconidia, branchlets up<br />
to 50 µm long; occasionally with a few additional micronematous<br />
Notes: <strong>The</strong> type culture of Penidiella venezuelensis was originally<br />
determined as Stenella araguata from which it is, however,<br />
quite distinct by having smooth mycelium, long penicillate<br />
conidiophores with subdenticulate conidiogenous loci, smaller<br />
conidia, <strong>and</strong> agreeing with the concept of the <strong>genus</strong> Penidiella. It is<br />
phylogenetically distinct from the other taxa of Penidiella (see Fig.<br />
1 in Crous et al. 2007a – this volume).<br />
Pseudotaeniolina J.L. Crane & Schokn., Mycologia 78: 88. 1986.<br />
? = Friedmanniomyces Onofri, Nova Hedwigia 68: 176. 1999.<br />
Type species: Pseudotaeniolina convolvuli (Esf<strong>and</strong>.) J.L. Crane &<br />
Schokn., Mycologia 78: 88. 1986.<br />
Description: Crane & Schoknecht (1986, figs 3–19).<br />
Notes: No cultures or sequence data are available of the type<br />
species, <strong>and</strong> Pseudotaeniolina globosa De Leo, Urzì & de Hoog was<br />
placed in Pseudotaeniolina based on its morphology <strong>and</strong> ecology.<br />
<strong>The</strong> <strong>genus</strong> Friedmanniomyces is presently known from two species<br />
(Selbmann et al. 2005). Morphologically Friedmanniomyces is<br />
<strong>similar</strong> to Pseudotaeniolina, but fresh material of Pseudotaeniolina<br />
convolvuli needs to be recollected before this can be clarified.<br />
Readeriella Syd. & P. Syd., Ann. Mycol. 6: 484. 1908.<br />
= Kirramyces J. Walker, B. Sutton & Pascoe, Mycol. Res. 96: 919. 1992.<br />
= Colletogloeopsis Crous & M.J. Wingf., Canad. J. Bot. 75: 668. 1997.<br />
Synanamorphs: Cibiessia Crous, Fungal Diversity 26: 151. 2007;<br />
also pseudocercospora-like, see Crous (1998).<br />
Type species: Readeriella mirabilis Syd. & P. Syd., Ann. Mycol. 6:<br />
484. 1908.<br />
Description: Crous et al. (2004b; figs 36–38).<br />
Notes: Several coelomycete genera are presently available<br />
to accommodate anamorphs of Capnodiales that reside in<br />
Teratosphaeriaceae, for which Readeriella is the oldest name. Other<br />
genera such as Phaeophleospora Rangel, Sonderhenia H.J. Swart<br />
& J. Walker <strong>and</strong> Lecanosticta Syd. belong to Mycosphaerellaceae.<br />
24
Phylogenetic lineages in the Capnodiales<br />
Fig. 16. Penidiella venezuelensis (type material). A. Microconidiophore. B. Apical part of macroconidiophore. C–F. Chains of conidia. Scale bars = 10 µm.<br />
Readeriella is polyphyletic within Teratosphaeriaceae. <strong>The</strong><br />
recognition <strong>and</strong> circumscription (synonymy) of this <strong>genus</strong> follows the<br />
principles for anamorph genera within Capnodiales as outlined in the<br />
introduction to this volume. <strong>The</strong> only unifying character is conidial<br />
pigmentation, <strong>and</strong> the mode of conidiogenesis. Conidiogenous<br />
cells range from mono- to polyphialides with periclinal thickening,<br />
to phialides with percurrent proliferation, as observed in the type<br />
species, R. mirabilis (Fig. 18). Within the form <strong>genus</strong> conidia vary<br />
from aseptate to multiseptate, smooth to rough, <strong>and</strong> have a range<br />
of synanamorphs. Readeriella mirabilis has a synanamorph with<br />
cylindrical, aseptate conidia, while other species of Readeriella<br />
again have Cibiessia synanamorphs (scytalidium-like, with chains<br />
of dry, disarticulating conidia), suggesting the conidial morphology<br />
to be quite plastic. A re-examination of R. readeriellophora Crous &<br />
Mansilla revealed pycnidia to form a central cushion on which the<br />
conidiogenous cells are arranged (Fig. 18). This unique feature is<br />
commonly known in genera such as Coniella Höhn. <strong>and</strong> Pilidiella<br />
Petr. & Syd. (Diaporthales) (Van Niekerk et al. 2004), <strong>and</strong> has never<br />
been observed among anamorphs of the Capnodiales. Another<br />
species of Readeriella, namely “Phaeophleospora” toledana Crous<br />
& Bills, again forms paraphyses interspersed among conidiogenous<br />
cells, a rare feature in this group of fungi, while several species<br />
Fig. 17. Penidiella venezuelensis (type material). A. Hypha. B. Micronematous<br />
conidiophores. C. Macronematous conidiophores. D–E. Ramoconidia. F. Secondary<br />
conidia. Scale bar = 10 µm. U. Braun del.<br />
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25
Crous et al.<br />
have conidiomata ranging from acervuli to pycnidia (Cortinas et<br />
al. 2006). Phylogenetically this coelomycete morphology, with<br />
its characteristic conidiogenesis, has evolved several times in<br />
Teratosphaeriaceae.<br />
Readeriella blakelyi (Crous & Summerell) Crous & U. Braun,<br />
comb. nov. MycoBank MB504516.<br />
Basionym: Colletogloeopsis blakelyi Crous & Summerell, Fungal<br />
Diversity 23: 342. 2006.<br />
Readeriella brunneotingens Crous & Summerell, sp. nov.<br />
MycoBank MB504517. Fig. 19.<br />
Etymology: Named after the diffuse brown pigment visible in agar<br />
when cultivated on MEA.<br />
Readeriellae gauchensi similis, sed coloniis viridi-atris et pigmento brunneo in agaro<br />
diffundente distinguenda.<br />
Leaf spots amphigenous, irregular specks up to 3 mm diam,<br />
medium brown with a thin, raised, concolorous border. Conidiomata<br />
amphigenous, substomatal, exuding conidia in black masses;<br />
conidiomata pycnidial in vivo <strong>and</strong> in vitro, globose, brown to black,<br />
up to 120 µm diam; wall consisting of 3–4 cell layers of brown<br />
cells of textura angularis. Conidiogenous cells brown, verruculose,<br />
aseptate, doliiform to ampulliform, or reduced to inconspicuous loci<br />
on hyphae (in vitro), proliferating percurrently near the apex, 5–7 ×<br />
3–5 µm; sympodial proliferation also observed in culture. Conidia<br />
brown, smooth to finely verruculose, ellipsoidal to subcylindrical,<br />
apex obtuse to subobtuse, tapering to a subtruncate or truncate<br />
base (1–1.5 µm wide) with inconspicuous, minute marginal frill,<br />
(5–)6–7(–8) × 2–3(–3.5) µm in vitro, becoming 1-septate; in older<br />
cultures becoming swollen, <strong>and</strong> up to 2-septate, 15 µm long <strong>and</strong><br />
5 µm wide.<br />
Cultural characteristics: Colonies on MEA reaching 20 mm diam<br />
after 2 mo at 25 °C; colonies erumpent, aerial mycelium sparse to<br />
absent, margins smooth but irregularly lobate; surface irregularly<br />
folded, greenish black, with profuse sporulation, visible as oozing<br />
black conidial masses; a diffuse dark-brown pigment is also<br />
produced, resulting in inoculated MEA plates appearing darkbrown.<br />
Specimen examined: Australia, Queensl<strong>and</strong>, Cairns, Eureka Creek, 48 km from<br />
Mareeba, S 17° 11’ 13.2”, E 145° 02’ 27.4”, 468 m, on leaves of Eucalyptus<br />
tereticornis, 26 Aug. 2006, P.W. Crous, <strong>CBS</strong>-H 19838 holotype, culture ex-type<br />
CPC 13303 = <strong>CBS</strong> 120747.<br />
Notes: Conidial dimensions of R. brunneotingens closely<br />
match those of Readeriella gauchensis (M.-N. Cortinas, Crous<br />
& M.J. Wingf.) Crous (Cortinas et al. 2006). <strong>The</strong> two species<br />
can be distinguished in culture, however, in that colonies of R.<br />
brunneotingens are greenish black in colour, sporulate profusely,<br />
<strong>and</strong> exude a diffuse, brown pigment into the agar, whereas colonies<br />
of R. gauchensis are more greenish olivaceous, <strong>and</strong> exude a yellow<br />
pigment into the agar (Cortinas et al. 2006).<br />
Readeriella considenianae (Crous & Summerell) Crous & U.<br />
Braun, comb. nov. MycoBank MB504518.<br />
Basionym: Colletogloeopsis considenianae Crous & Summerell,<br />
Fungal Diversity 23: 343. 2006.<br />
≡ Phaeophleospora destructans (M.J. Wingf. & Crous) Crous, F.A. Ferreira<br />
& B. Sutton, S. African J. Bot. 63: 113. 1997.<br />
Readeriella dimorpha (Crous & Carnegie) Crous & U. Braun,<br />
comb. nov. MycoBank MB504520.<br />
Basionym: Colletogloeopsis dimorpha Crous & Carnegie, Fungal<br />
Diversity 23: 345. 2006.<br />
Readeriella gauchensis (M.-N. Cortinas, Crous & M.J. Wingf.)<br />
Crous & U. Braun, comb. nov. MycoBank MB504521.<br />
Basionym: Colletogloeopsis gauchensis M.-N. Cortinas, Crous &<br />
M.J. Wingf., Stud. Mycol. 55: 143. 2006.<br />
Readeriella pulcherrima (Gadgil & M. Dick) Crous & U. Braun,<br />
comb. nov. MycoBank MB504522.<br />
Basionym: Septoria pulcherrima Gadgil & M. Dick, New Zeal<strong>and</strong> J.<br />
Bot. 21: 49. 1983.<br />
≡ Stagonospora pulcherrima (Gadgil & M. Dick) H.J. Swart, Trans. Brit.<br />
Mycol. Soc. 90: 285. 1988.<br />
= Cercospora eucalypti Cooke & Massee, Grevillea 18: 7. 1889.<br />
≡ Kirramyces eucalypti (Cooke & Massee) J. Walker, B. Sutton & Pascoe,<br />
Mycol. Res. 96: 920. 1992.<br />
≡ Phaeophleospora eucalypti (Cooke & Massee) Crous, F.A. Ferreira & B.<br />
Sutton, S. African J. Bot. 63: 113. 1997.<br />
Notes: <strong>The</strong> epithet “eucalypti” is preoccupied by Readeriella<br />
eucalypti (Gonz. Frag.) Crous (Summerell et al., 2006), <strong>and</strong> thus<br />
the synonym “pulcherrima” becomes the next available name for<br />
this species.<br />
Readeriella readeriellophora, see Teratosphaeria readeriellophora.<br />
Fig. 18.<br />
Readeriella stellenboschiana (Crous) Crous & U. Braun, comb.<br />
nov. MycoBank MB504523.<br />
Basionym: Colletogloeopsis stellenboschiana Crous, Stud. Mycol.<br />
55: 110. 2006.<br />
Readeriella zuluensis (M.J. Wingf., Crous & T.A. Cout.) Crous &<br />
U. Braun, comb. nov. MycoBank MB504524.<br />
Basionym: Coniothyrium zuluense M.J. Wingf., Crous & T.A. Cout.,<br />
Mycopathologia 136: 142. 1997.<br />
≡ Colletogloeopsis zuluensis (M.J. Wingf., Crous & T.A. Cout.) M.-N.<br />
Cortinas, M.J. Wingf. & Crous (zuluense), Mycol. Res. 110: 235. 2006.<br />
Staninwardia B. Sutton, Trans. Br. Mycol. Soc. 57: 540. 1971.<br />
Type species: Staninwardia breviuscula B. Sutton, Trans. Br. Mycol.<br />
Soc. 57: 540. 1971.<br />
Description: Sutton (1971; fig. 1).<br />
Notes: <strong>The</strong> <strong>genus</strong> Staninwardia presently contains two species,<br />
namely S. breviuscula <strong>and</strong> Staninwardia suttonii Crous & Summerell<br />
(Summerell et al. 2006), though its placement in Capnodiales was<br />
less well resolved. <strong>The</strong> <strong>genus</strong> forms acervuli on brown leaf spots,<br />
with brown, catenulate conidia covered in a mucilaginous sheath.<br />
Readeriella destructans (M.J. Wingf. & Crous) Crous & U. Braun,<br />
comb. nov. MycoBank MB504519.<br />
Basionym: Kirramyces destructans M.J. Wingf. & Crous, S. African<br />
J. Bot. 62: 325. 1996.<br />
26
Phylogenetic lineages in the Capnodiales<br />
Fig. 18. A–E. Readeriella mirabilis. A. Conidium with conidial cirrus. B. Conidiogenous cells with percurrent proliferation. C. Macroconidia. D. Slightly pigmented, verruculose<br />
conidiogenous cell. E. Macro- <strong>and</strong> microconidia. F–I. Readeriella readeriellophora (type material). F. Colony on OA. G. Central stromatal tissue giving rise to conidiophores. H.<br />
Conidiogenous cells. I. Conidia. Scale bars = 10 µm.<br />
Fig. 19. Readeriella brunneotingens (type material). A. Leaf spot. B. Colony on MEA. C–D. Conidia. Scale bar = 10 µm.<br />
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27
Crous et al.<br />
Schizothyriaceae clade<br />
Schizothyrium Desm., Ann. Sci. Nat., Bot., sér. 3: 11. 1849.<br />
Type species: Schizothyrium acerinum Desm., Ann. Sci. Nat., Bot.,<br />
sér. 3: 11. 1849.<br />
Description: Batzer et al. (2007; figs 3–7).<br />
Notes: Species of Schizothyrium (Schizothyriaceae) have<br />
Zygophiala E.W. Mason anamorphs, <strong>and</strong> were recently shown to<br />
be allied to Mycosphaerellaceae (Batzer et al. 2007). Although<br />
species of Schizothyrium have thyrothecia, they cluster among<br />
genera with pseudothecial ascomata, questioning the value of this<br />
character at the family level. Based on its bitunicate asci <strong>and</strong> 1-<br />
septate ascospores, the teleomorph is comparable to others in the<br />
Capnodiales.<br />
Mycosphaerellaceae clade<br />
Mycosphaerella subclade<br />
Mycosphaerella Johanson, Öfvers. Förh. Kongl. Svenska<br />
Vetensk.-Akad. 41(9): 163. 1884.<br />
Type species: Mycosphaerella punctiformis (Pers. : Fr.) Starbäck,<br />
Bih. Kongl. Svenska Vetensk.-Akad. H<strong>and</strong>l. 15(3, 2): 9. 1889.<br />
Anamorph: Ramularia endophylla Verkley & U. Braun, Mycol. Res.<br />
108: 1276. 2004.<br />
Description: Verkley et al. (2004; figs 3–16).<br />
Notes: <strong>The</strong> <strong>genus</strong> Mycosphaerella has in the past been linked to 23<br />
anamorph genera (Crous et al. 2000), while additional genera have<br />
been linked via DNA-based studies, bringing the total to at least 30<br />
genera (Crous & Braun 2003, Crous et al. 2007b). However, based<br />
on ITS <strong>and</strong> SSU DNA phylogenetic studies <strong>and</strong> a reassessment of<br />
morphological characters <strong>and</strong> conidiogenesis, several anamorph<br />
genera have recently been reduced to synonymy (Crous & Braun<br />
2003, Crous et al. 2006a). Furthermore, the DNA sequence data<br />
generated to date clearly illustrate that the anamorph genera in<br />
Mycosphaerella are polyphyletic, residing in several clades within<br />
Mycosphaerella. If future collections not known from culture or DNA<br />
sequences are to be described in form genera, we recommend that<br />
the concepts as explained in Crous & Braun (2003) be used until<br />
such stage as they can be placed in Mycosphaerella, pending<br />
a modification of Art. 59 of the International Code of Botanical<br />
Nomenclature. <strong>The</strong> <strong>genus</strong> Mycosphaerella <strong>and</strong> its anamorphs<br />
represent a future topical issue of the Studies in Mycology, <strong>and</strong> will<br />
thus be treated separately.<br />
Dissoconium subclade<br />
Dissoconium de Hoog, Oorschot & Hijwegen, Proc. K. Ned. Akad.<br />
Wet., Ser. C, Biol. Med. Sci. 86(2): 198. 1983.<br />
Type species: Dissoconium aciculare de Hoog, Oorschot &<br />
Hijwegen, Proc. K. Ned. Akad. Wet., Ser. C, Biol. Med. Sci. 86(2):<br />
198. 1983.<br />
? = Uwebraunia Crous & M.J. Wingf., Mycologia 88: 446. 1996.<br />
Teleomorph: Mycosphaerella-like.<br />
Description: de Hoog et al. (1983), Crous (1998), Crous et al.<br />
(2004b; figs 3–10).<br />
Notes: <strong>The</strong> <strong>genus</strong> Dissoconium presently encompasses six<br />
species (Crous et al. 2007b), of which two, M. lateralis Crous &<br />
M.J. Wingf. (D. dekkeri de Hoog & Hijwegen), <strong>and</strong> M. communis<br />
Crous & Mansilla (D. commune Crous & Mansilla) are also known<br />
from their Mycosphaerella-like teleomorphs. No teleomorph <strong>genus</strong><br />
will be introduced for this clade, however, until more sexual species<br />
have been collected to help clarify the morphological features of<br />
this <strong>genus</strong>. A further complication lies in the fact that yet other<br />
species, morphologically distinct from Dissoconium, also cluster in<br />
this clade (Crous, unpubl. data).<br />
“Passalora” zambiae subclade<br />
“Passalora” zambiae Crous & T.A. Cout., Stud. Mycol. 50: 209.<br />
2004.<br />
Description: Crous et al. (2004b; figs 32–33).<br />
Notes: This fungus was placed in the form <strong>genus</strong> “Passalora”<br />
based on its smooth mycelium, giving rise to conidiophores forming<br />
branched chains of brown conidia with thickened, darkened,<br />
refractive hila. Although derived from single ascospores, the<br />
teleomorph material was lost, <strong>and</strong> thus it needs to be recollected<br />
before the relavance of its phylogenetic position can be fully<br />
understood.<br />
Additional teleomorph genera considered<br />
Coccodinium A. Massal., Atti Inst. Veneto Sci. Lett. Arti, Série 2,<br />
5: 336. 1860. (Fig. 20).<br />
Type species: Coccodinium bartschii A. Massal., Atti Inst. Veneto<br />
Sci. Lett. Arti, Série 2, 5: 337. 1860.<br />
Description: Eriksson (1981, figs 34–35).<br />
Notes: <strong>The</strong> <strong>genus</strong> Coccodinium (Coccodiniaceae) is characterised<br />
by having ascomata that are sessile on a subiculum, or somewhat<br />
immersed, semiglobose, collapsed when dry, brownish, uniloculate,<br />
with a centrum that stains blue in IKI (iodine potassium iodide).<br />
Asci are bitunicate, stalked, 8-spored, saccate, <strong>and</strong> have a thick,<br />
undifferentiated endotunica. Periphyses <strong>and</strong> periphysoids are<br />
well-developed <strong>and</strong> numerous. Ascospores are elongate, fusiform,<br />
ellipsoidal or clavate, transversely septate or muriform, hyaline or<br />
brownish (Eriksson 1981), <strong>and</strong> lack a mucous sheath. Based on a<br />
SSU sequence (GenBank accession U77668) derived from a strain<br />
identified as C. bartschii (Winka et al. 1998), Coccodinium appears<br />
to be allied to the taxa treated here in Teratosphaeria. Freshly<br />
collected cultures are relatively slow growing, <strong>and</strong> on MEA they<br />
form erumpent round, black colonies with sparse hyphal growth. On<br />
the surface of these colonies hyphal str<strong>and</strong>s, consisting of brown,<br />
globose cells, give rise to conidia. Older cells (up to 15 µm diam)<br />
become fertile, giving rise to 1–3 conidia via inconspicuous phialidic<br />
loci. Conidia are fusoid-ellipsoidal to clavate, 3–5-septate, becoming<br />
constricted at the transverse septa, apex obtuse, base subtruncate,<br />
guttulate, smooth, widest in the upper third of the conidium, 15–<br />
40 × 4–7 µm. Phylogenetically Coccodinium is thus allied to the<br />
Chaetothyriales (Fig. 1), <strong>and</strong> not the Teratosphaeriaceae.<br />
28
Phylogenetic lineages in the Capnodiales<br />
Fig. 20. Coccodinium bartschii. A. Ascomata on host. B. Ostiolar area. C. Periphysoids. D–E. Ascospores shot onto agar. F–I. Asci with thick ectotunica. J–K. Young ascospores.<br />
L–M. Mature ascospores. N. Colony on MEA. O–Q. Conidiogenous cells giving rise to conidia. R–S. Conidia. Scale bars: A, N = 250, B, D, F–G, I, L–M, O = 10 µm.<br />
www.studiesinmycology.org<br />
29
Crous et al.<br />
Fig. 21. Stigmidium schaereri. A. Lichenicolous habit on Dacampia hookeri. B. Vertical section through an ascoma. C–D. Asci. E–G. Ascospores. H. Older, brown ascospores.<br />
Scale bars = 10 µm.<br />
Stigmidium Trevis., Consp. Verruc.: 17. 1860. (Fig. 21).<br />
Type species: Stigmidium schaereri (A. Massal.) Trevis., Consp.<br />
Verruc.: 17. 1860.<br />
Description: Roux & Triebel (1994, figs 47–50).<br />
Notes: <strong>The</strong> type species of the <strong>genus</strong> is lichenicolous, characterised<br />
by semi-immersed, black, globose ascomata with ostiolar<br />
periphyses <strong>and</strong> periphysoids. Asci are 8-spored, fasciculate,<br />
bitunicate, (endotunica not giving a special reaction in Congo red<br />
or toluidine blue). Ascospores are fusoid-ellipsoidal, medianly 1-<br />
septate, guttulate, thin-walled, lacking a sheath. Presently no<br />
culture is available, <strong>and</strong> thus the placement of Stigmidium remains<br />
unresolved.<br />
Discussion<br />
From the LSU sequence data presented here, it is clear that<br />
Mycosphaerella is not monophyletic as previously suggested (Crous<br />
et al. 2001, Goodwin et al. 2001). <strong>The</strong> first step to circumscribe<br />
natural genera within this complex was taken by Braun et al. (2003),<br />
who separated <strong>Cladosporium</strong> anamorphs from this complex,<br />
<strong>and</strong> erected Davidiella (Davidiellaceae; Schoch et al. 2006) to<br />
accommodate their teleomorphs. <strong>The</strong> present study reinstates<br />
the <strong>genus</strong> Teratosphaeria for a clade of largely extremotolerant<br />
fungi (Selbmann et al. 2005) <strong>and</strong> foliar pathogens of Myrtaceae<br />
<strong>and</strong> Proteaceae (Crous et al. 2004a, b, 2006b, 2007b), <strong>and</strong> further<br />
separates generic subclades within the Mycosphaerellaceae,<br />
while Batzer et al. (2007) again revealed Schizothyrium Desm.<br />
(Schizothyriaceae) to cluster within the Mycosphaerellaceae. Our<br />
results, however, provide support for recognition of Schizothyrium as<br />
a distinct <strong>genus</strong>, although Schizothyriaceae was less well supported<br />
as being separate from Mycosphaerellaceae (Capnodiales).<br />
Although pleomorphism represents a rather unstudied<br />
phenomenon in this group of fungi, it has been observed in several<br />
species. Within the Teratosphaeria clade, Crous et al. (2007b)<br />
recently demonstrated teleomorphs to have Readeriella <strong>and</strong><br />
Cibiessia synanamorphs, while the black yeast genera that belong<br />
to this clade, commonly have more than one anamorph state in<br />
culture. <strong>The</strong> present study also revealed Readeriella mirabilis to<br />
have two conidial types in culture, <strong>and</strong> to be highly plastic regarding<br />
its mode of conidiogenesis, <strong>and</strong> Readeriella to be the oldest generic<br />
name available for a large group of leaf-spotting coelomycetes in<br />
the Teratosphaeriaceae (Capnodiales).<br />
Although not commonly documented, there are ample<br />
examples of synanamorphs in Capnodiales. Within Mycosphaerella,<br />
Beilharz et al. (2004) described Passalora perplexa Beilharz,<br />
Pascoe, M.J. Wingf. & Crous as a species with a coelomycete<br />
<strong>and</strong> yeast synanamorph, while Crous & Corlett (1998) described<br />
Mycosphaerella stigmina-platani F.A. Wolf to have a Cercostigmina<br />
U. Braun <strong>and</strong> Xenostigmina Crous synanamorph, <strong>and</strong> recent<br />
collections also revealed the presence of a <strong>similar</strong> species that has<br />
typical “Stigmina” (distoseptate conidia) <strong>and</strong> Pseudocercospora<br />
(euseptate conidia) synanamorphs (Crous, unpubl. data), <strong>and</strong><br />
Crous (1998) reported Readeriella epicoccoides (coelomycete) to<br />
30
Phylogenetic lineages in the Capnodiales<br />
have a Cercostigmina (hyphomycete) synanamorph in culture.<br />
Although the Mycosphaerella complex encompasses<br />
thous<strong>and</strong>s of names, it may appear strange that it is only now that<br />
more clarity is obtained regarding the phylogenetic relationships<br />
among taxa in this group. This is partly due to the fact that these<br />
organisms are cultivated with difficulty, <strong>and</strong> also that the first paper<br />
to address the taxonomy of this complex based on DNA sequence<br />
data was only relatively recently published (Stewart et al. 1999).<br />
In the latter study, the <strong>genus</strong> Paracercospora Deighton (scars<br />
minutely thickened along the rim), was shown to be synonymous<br />
with the older <strong>genus</strong> Pseudocercospora. Similarily, Crous et al.<br />
(2001) showed that Cercostigmina (rough, irregular percurrent<br />
proliferations) was also synonymous with Pseudocercospora.<br />
This led Crous & Braun (2003) to conclude that conidiomatal type,<br />
conidial catenulation, septation <strong>and</strong> proliferation of conidiogenous<br />
cells were of less importance in separating species at the generic<br />
level. Mycovellosiella Rangel <strong>and</strong> Phaeoramularia Munt.-Cvetk.<br />
were subsequently reduced to synonymy with the older name,<br />
Passalora Fr., <strong>and</strong> characters identified as significant at the generic<br />
level were pigmentation (Cercospora vs. Passalora), scar structure<br />
(Passalora vs. Pseudocercospora), <strong>and</strong> verruculose superficial<br />
hyphae (Stenella vs. Passalora). Due to the unavailability of<br />
cultures, no decision was made regarding Stenella (verrucose<br />
conidia <strong>and</strong> mycelium), Stigmina (distoseptate conidia), <strong>and</strong><br />
several other, less well-known genera such as Asperisporium<br />
Maubl., Denticularia Deighton, Distocercospora N. Pons & B.<br />
Sutton, Prathigada Subram., Ramulispora, Pseudocercosporidium<br />
Deighton, Stenellopsis B. Huguenin <strong>and</strong> Verrucisporota D.E. Shaw<br />
& Alcorn. In a recent study, however, Crous et al. (2006a) were<br />
able to show that Phaeoisariopsis (synnemata, conidia with slightly<br />
thickened hila) <strong>and</strong> Stigmina (distoseptate conidia) were also<br />
synonyms of Pseudocercospora.<br />
<strong>The</strong> present study shows that most anamorph genera are<br />
polyphyletic within Teratosphaeria, <strong>and</strong> paraphyletic within<br />
Capnodiales. In some cases, generic concepts of anamorphs<br />
based on morphology <strong>and</strong> conidium ontogeny conform well with<br />
phylogenetic relationships, though this is not true in all cases due<br />
to convergence. Nevertheless, anamorphs still convey valuable<br />
morphological information that is contained in the anamorph name,<br />
<strong>and</strong> naming anamorphs continue to provide a practical system to<br />
identify the various asexual taxa encountered.<br />
Acknowledgements<br />
We gratefully acknowledge several colleagues in different countries who have<br />
collected the material studied here, without which this work would not have been<br />
possible. We thank M. Vermaas for preparing the photographic plates. M. Grube is<br />
gratefully acknowledged for sending us fresh material of Stigmidium to include in<br />
this study, <strong>and</strong> D. Triebel <strong>and</strong> M. Grube are thanked for advice regarding the strain of<br />
hamathecial tissue in Stigmidium. K.A. Seifert is thanked for numerous discussions<br />
about anamorph concepts <strong>and</strong> advice pertaining to this paper, <strong>and</strong> for recollecting<br />
Coccodinium bartschii to enable us to clarify its phylogeny. J.K. Stone is thanked for<br />
commenting on an earlier draft version of this paper.<br />
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(2006). A multigene phylogeny of the Dothideomycetes using four nuclear loci.<br />
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Schubert K, Groenewald JZ, Braun U, Dijksterhuis J, Starink M, Hill CF, Zalar P,<br />
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Summerell BA, Groenewald JZ, Carnegie AJ, Summerbell RC, Crous PW (2006).<br />
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32
available online at www.studiesinmycology.org<br />
doi:10.3114/sim.2007.58.02<br />
Studies in Mycology 58: 33–56. 2007.<br />
Delimiting <strong>Cladosporium</strong> from morphologically <strong>similar</strong> genera<br />
P.W. Crous 1* , U. Braun 2 , K. Schubert 3 <strong>and</strong> J.Z. Groenewald 1<br />
1<br />
<strong>CBS</strong> Fungal Biodiversity Centre, P.O. Box 85167, 3508 AD Utrecht, <strong>The</strong> Netherl<strong>and</strong>s; 2 Martin-Luther-Universität, Institut für Biologie, Geobotanik und Botanischer Garten,<br />
Herbarium, Neuwerk 21, D-06099 Halle (Saale), Germany; 3 Botanische Staatssammlung München, Menzinger Straße 67, D-80638 München, Germany<br />
*Correspondence: Pedro Crous, p.crous@cbs.knaw.nl<br />
Abstract: <strong>The</strong> <strong>genus</strong> <strong>Cladosporium</strong> is restricted to <strong>dematiaceous</strong> hyphomycetes with a coronate scar type, <strong>and</strong> Davidiella teleomorphs. In the present study numerous<br />
cladosporium-like taxa are treated, <strong>and</strong> allocated to different genera based on their morphology <strong>and</strong> DNA phylogeny derived from the LSU nrRNA gene. Several species are<br />
introduced in new genera such as Hyalodendriella, Ochrocladosporium, Rachicladosporium, Rhizocladosporium, Toxicocladosporium <strong>and</strong> Verrucocladosporium. A further new<br />
taxon is described in Devriesia (Teratosphaeriaceae). Furthermore, <strong>Cladosporium</strong> castellanii, the etiological agent of tinea nigra in humans, is confirmed as synonym of Stenella<br />
araguata, while the type species of Stenella is shown to be linked to the Teratosphaeriaceae (Capnodiales), <strong>and</strong> not the Mycosphaerellaceae as formerly presumed.<br />
Taxonomic novelties: Devriesia americana Crous & Dugan, sp. nov., Hyalodendriella Crous, gen. nov., Hyalodendriella betulae Crous sp. nov., Ochrocladosporium Crous &<br />
U. Braun, gen. nov., Ochrocladosporium elatum (Harz) Crous & U. Braun, comb. nov., Ochrocladosporium frigidarii Crous & U. Braun, sp. nov., Rachicladosporium Crous, U.<br />
Braun & Hill, gen. nov., Rachicladosporium luculiae Crous, U. Braun & Hill, sp. nov., Rhizocladosporium Crous & U. Braun, gen. nov., Rhizocladosporium argillaceum (Minoura)<br />
Crous & U. Braun, comb. nov., Toxicocladosporium Crous & U. Braun, gen. nov., Toxicocladosporium irritans Crous & U. Braun, sp. nov., Verrucocladosporium K. Schub., Aptroot<br />
& Crous, gen. nov., Verrucocladosporium dirinae K. Schub., Aptroot & Crous, sp. nov.<br />
Key words: <strong>Cladosporium</strong>, Davidiella, food spoilage, hyphomycetes, indoor air, LSU phylogeny, taxonomy.<br />
Introduction<br />
Cladosporioid hyphomycetes are common, widespread fungi.<br />
<strong>The</strong> <strong>genus</strong> <strong>Cladosporium</strong> Link is based on the type species,<br />
<strong>Cladosporium</strong> herbarum (Pers. : Fr.) Link, which in turn has been<br />
linked to Davidiella Crous & U. Braun teleomorphs (Braun et al.<br />
2003, Schubert et al. 2007b – this volume). <strong>Cladosporium</strong> is one<br />
of the largest, most heterogeneous genera of hyphomycetes,<br />
comprising more than 772 names (Dugan et al. 2004), <strong>and</strong> including<br />
endophytic, fungicolous, human pathogenic, phytopathogenic <strong>and</strong><br />
saprobic species. Species of this <strong>genus</strong> affect daily human life in<br />
various ways. <strong>The</strong> common saprobic members of <strong>Cladosporium</strong><br />
occur on all kinds of senescing <strong>and</strong> dead leaves <strong>and</strong> stems of<br />
herbaceous <strong>and</strong> woody plants, as secondary invaders on necrotic<br />
leaf lesions caused by other fungi, are frequently isolated from air,<br />
soil, food stuffs, paint, textiles <strong>and</strong> other organic matters, are also<br />
known to be common endophytes (Riesen & Sieber 1985, Brown<br />
et al. 1998, El-Morsy 2000) as well as phylloplane fungi (Islam &<br />
Hasin 2000, De Jager et al. 2001, Inacio et al. 2002, Stohr & Dighton<br />
2004, Levetin & Dorsey 2006). Furthermore, some <strong>Cladosporium</strong><br />
species are known to be potential agents of medical relevance.<br />
<strong>Cladosporium</strong> herbarum is, for instance, a common contaminant in<br />
clinical laboratories <strong>and</strong> causes allergic lung mycoses (de Hoog et<br />
al. 2000, Schubert et al. 2007b – this volume).<br />
In spite of the enormous relevance of this <strong>genus</strong>, there is<br />
no comprehensive modern revision of <strong>Cladosporium</strong>, but some<br />
attempts to revise <strong>and</strong> monograph parts of it have been initiated<br />
during the last decade (David 1997, Partridge & Morgan-Jones<br />
2002, Wirsel et al. 2002, Braun et al. 2003, Dugan et al. 2004, Park<br />
et al. 2004, Seifert et al. 2004, Schubert & Braun 2004, 2005a,<br />
b, 2007, Heuchert et al. 2005, Schubert 2005a, b, Schubert et al.<br />
2006).<br />
Previous molecular studies employing rDNA ITS sequence<br />
data (Crous et al. 2001) have shown <strong>Cladosporium</strong> spp. to cluster<br />
adjacent to the main monophyletic Mycosphaerella Johanson<br />
cluster, suggesting a position apart from the latter <strong>genus</strong>. Braun<br />
et al. (2003) carried out more comprehensive sequence analyses,<br />
based on ITS (ITS-1, 5.8S <strong>and</strong> ITS-2) <strong>and</strong> 18S rDNA data, providing<br />
further evidence that <strong>Cladosporium</strong> s. str. represents a sister clade<br />
of Mycosphaerella.<br />
Various authors discussed the taxonomy <strong>and</strong> circumscription<br />
of <strong>Cladosporium</strong> (von Arx 1981, 1983, McKemy & Morgan-Jones<br />
1990, Braun 1995), reaching different conclusions. However, a<br />
first decisive revision of <strong>Cladosporium</strong>, leading to a more natural<br />
concept of this <strong>genus</strong>, was published by David (1997), who carried<br />
out comprehensive scanning electron microscopic examinations of<br />
the scar <strong>and</strong> hilum structure in <strong>Cladosporium</strong> <strong>and</strong> Heterosporium<br />
Klotzsch ex Cook. <strong>The</strong> first Scanning Electron Micrograph (SEM)<br />
studies of these structures, published by Roquebert (1981), indicated<br />
that the conidiogenous loci <strong>and</strong> conidial hila in <strong>Cladosporium</strong> are<br />
characterised by having a unique structure. David (1997) confirmed<br />
these observations, based on a wide range of <strong>Cladosporium</strong> <strong>and</strong><br />
Heterosporium species, <strong>and</strong> demonstrated that the structures of the<br />
conidiogenous loci <strong>and</strong> hila in the latter <strong>genus</strong> fully agree with those<br />
of <strong>Cladosporium</strong>, proving that Heterosporium was indeed a synonym<br />
of <strong>Cladosporium</strong> s. str. He introduced the term “coronate” for the<br />
<strong>Cladosporium</strong> scar type, which is characterised by having a central<br />
convex part (dome), surrounded by a raised periclinal rim (David<br />
1997), <strong>and</strong> showed that this type is confined to anamorphs, as far<br />
as experimentally proven, connected with teleomorphs belonging<br />
in “Mycosphaerella” s. lat. <strong>The</strong>se results were confirmed in a later<br />
phylogenetic study by Braun et al. (2003). <strong>Cladosporium</strong> s. str. was<br />
shown to be a sister clade to Mycosphaerella s. str., for which the<br />
new teleomorph <strong>genus</strong> Davidiella was proposed. Although no clear<br />
morphological differences were reported between Davidiella <strong>and</strong><br />
Mycosphaerella, a further study by Aptroot (2006) found ascospores<br />
of Davidiella to have characteristic irregular cellular inclusions<br />
(lumina), which are absent in species of Mycosphaerella, along with<br />
periphysoids <strong>and</strong> pseudoparaphyses (Schubert et al. 2007b – this<br />
volume). Furthermore, a higher order phylogeny study by Schoch<br />
et al. (2006), which employed DNA sequence data of four loci (SSU<br />
nrDNA, LSU nrDNA, EF-1α, RPB2), revealed species of Davidiella<br />
to cluster in a separate family (Davidiellaceae) from species of<br />
Mycosphaerella (Mycosphaerellaceae), with both families residing<br />
in the Capnodiales (Dothideomycetes), <strong>and</strong> not Dothideales as<br />
always presumed.<br />
33
Crous et al.<br />
Table 1. Isolates for which new sequences were generated.<br />
Anamorph Teleomorph Accession number 1 Host Country Collector GenBank numbers 2<br />
(ITS, LSU)<br />
Cladoriella eucalypti <strong>CBS</strong> 115899*; CPC 10954 Eucalyptus sp. South Africa P.W. Crous EU040224, EU040224<br />
Coniothyrium palmarum <strong>CBS</strong> 758.73; CMW 5283 Phoenix dactylifera Israel Y. Pinkas DQ240000, EU040225<br />
Devriesia acadiensis <strong>CBS</strong> 115874; DAOM 232211 Soil Canada N. Nickerson AY692095, EU040226<br />
Devriesia americana <strong>CBS</strong> 117726; ATCC 96545; CPC 5121 Air U.S.A. F.M. Dugan AY251068, EU040227<br />
Devriesia shelburniensis <strong>CBS</strong> 115876; DAOM 232217 Soil Canada N. Nickerson AY692093, EU040228<br />
Devriesia thermodurans <strong>CBS</strong> 115878*; DAOM 225330 Soil Canada N. Nickerson AY692087, EU040229<br />
Hormoconis resinae <strong>CBS</strong> 365.86 – – – EU040230, EU040230<br />
<strong>CBS</strong> 184.54; ATCC 11841; CPC 3692; IMI 089837; IFO 31706 Creosote-treated wooden pole U.S.A. – AY251067, EU040231<br />
Hyalodendriella betulae <strong>CBS</strong> 261.82* Alnus glutinosa Netherl<strong>and</strong>s W. Gams EU040232, EU040232<br />
Ochrocladosporium elatum <strong>CBS</strong> 146.33*; ATCC 11280; ATHUM 2862; IFO 6372; IMI 049629; Wood pulp Sweden E. Melin EU040233, EU040233<br />
MUCL 10094<br />
Ochrocladosporium frigidarii <strong>CBS</strong> 103.81* Cooled room Germany B. Ahlert EU040234, EU040234<br />
Parapleurotheciopsis inaequiseptata MUCL 41089; INIFAT C98/30-1 Rotten leaf Brazil R.F. Castañeda EU040235, EU040235<br />
Passalora daleae <strong>CBS</strong> 113031* Dalea spinosa Mexico L.B. Sparrius EU040236, EU040236<br />
Rachicladosporium luculiae CPC 11407* Luculia sp. New Zeal<strong>and</strong> F. Hill EU040237, EU040237<br />
Ramularia aplospora Mycosphaerella alchemillicola <strong>CBS</strong> 545.82* Powdery mildew on Alchemilla vulgaris Germany T. Hijwegen EU040238, EU040238<br />
Retroconis fusiformis <strong>CBS</strong> 330.81; IMI 170799 Gossypium sp. Pakistan – EU040239, EU040239<br />
Rhizocladosporium argillaceum <strong>CBS</strong> 241.67*; ATCC 38103; IFO 7055; OUT 4262 Decayed myxomycete Japan K. Tubaki EU040240, EU040240<br />
Subramaniomyces fusisaprophyticus <strong>CBS</strong> 418.95; INIFAT C94/134 Leaf litter Cuba R.F. Castañeda EU040241, EU040241<br />
<strong>The</strong>dgonia ligustrina W1877 Ligustrum sp. – H. Evans EU040242, EU040242<br />
Toxicocladosporium irritans <strong>CBS</strong> 185.58* Mouldy paint Suriname M.B. Schol-Schwarz EU040243, EU040243<br />
Verrucocladosporium dirinae <strong>CBS</strong> 112794* Dirina massiliensis U.K. A. Aptroot EU040244, EU040244<br />
1 ATCC: American Type Culture Collection, Virginia, U.S.A.; ATHUM: Culture Collection of Fungi, University of Athens, Department of Biology, Section of Ecology <strong>and</strong> Systematics, Athens, Greece; <strong>CBS</strong>: Centraalbureau voor Schimmelcultures, Utrecht,<br />
<strong>The</strong> Netherl<strong>and</strong>s; CMW: Culture collection of Mike Wingfield, housed at FABI, Pretoria, South Africa; CPC: Culture collection of Pedro Crous, housed at <strong>CBS</strong>; DAOM: Plant Research Institute, Department of Agriculture (Mycology), Ottawa, Canada; IFO:<br />
Institute For Fermentation, Osaka, Japan; IMI: International Mycological Institute, CABI-Bioscience, Egham, Bakeham Lane, U.K.; INIFAT: Alex<strong>and</strong>er Humboldt Institute for Basic Research in Tropical Agriculture, Ciudad de La Habana, Cuba; MUCL:<br />
Mycotheque de l’ Université Catholique de Louvain, Louvain-la-Neuve, Belgium; OUT: Department of Fermentation Technology, Faculty of Engineering, Osaka University, Yamadaue, Suita-shi, Osaka, Japan.<br />
2 ITS: internal transcribed spacer regions <strong>and</strong> 5.8S rRNA gene; LSU: partial 28S rRNA gene.<br />
*Ex-type cultures.<br />
34
<strong>Cladosporium</strong> <strong>and</strong> morphologically <strong>similar</strong> genera<br />
<strong>The</strong> current circumscription of <strong>Cladosporium</strong> emend. can be<br />
summarised as follows: Dematiaceous hyphomycetes; Davidiella<br />
anamorphs; mycelium internal <strong>and</strong> external; hyphae branched,<br />
septate, pigmented; stromata lacking or occasionally present;<br />
conidiophores mononematous, solitary to fasciculate, cylindrical,<br />
geniculate-sinuous to nodulose, simple to branched, subhyaline<br />
to usually distinctly pigmented, continuous to septate, smooth<br />
to verruculose; conidiogenous cells integrated, terminal <strong>and</strong><br />
intercalary, usually sympodial, with a single to several scars;<br />
conidiogenesis holoblastic; conidiogenous loci coronate, i.e.,<br />
more or less protuberant, composed of a central convex dome,<br />
surrounded by a raised periclinal rim, barely to distinctly darkened;<br />
conidia solitary or in short to long, simple to branched acropetal<br />
chains, amero- to phragmosporous, subhyaline to usually distinctly<br />
pigmented, smooth, verruculose, verrucose, echinulate, cristate,<br />
hila coronate, more or less protuberant.<br />
<strong>The</strong> new concept of <strong>Cladosporium</strong> s. str., supported by<br />
molecular data <strong>and</strong> typical coronate conidiogenous loci <strong>and</strong> conidial<br />
hila, rendered it possible to initiate a comprehensive revision<br />
of <strong>Cladosporium</strong> s. lat. <strong>The</strong> preparation of a general, annotated<br />
check-list of <strong>Cladosporium</strong> s. lat. was the first step in this direction<br />
(Dugan et al. 2004). <strong>The</strong> aim of the present study, therefore, was to<br />
delineate <strong>Cladosporium</strong> s. str. from other taxa that have in recent<br />
years been described in <strong>Cladosporium</strong> s. lat. To attain this goal<br />
isolates were studied under st<strong>and</strong>ardised conditions on a set of<br />
predescribed media (Schubert et al. 2007b – this volume), <strong>and</strong><br />
subjected to DNA sequence analysis of the LSU nrRNA gene.<br />
were only sequenced for isolates of which these data were not<br />
available. <strong>The</strong> ITS data were not included in the analyses but<br />
deposited in GenBank where applicable. Gaps longer than 10<br />
bases were coded as single events for the phylogenetic analyses;<br />
the remaining gaps were treated as missing data. Sequence data<br />
were deposited in GenBank (Table 1) <strong>and</strong> alignments in TreeBASE<br />
(www.treebase.org).<br />
Morphology<br />
Wherever possible, 30 measurements (× 1 000 magnification)<br />
were made of structures mounted in lactic acid or Shear’s solution<br />
(Gams et al. 2007), with the extremes of spore measurements<br />
given in parentheses. Microscopic observations were made from<br />
colonies cultivated for 7 d under continuous near-ultraviolet light<br />
at 25 °C on SNA as explained in Schubert et al. (2007b – this<br />
volume). Three classes of conidia are distinguished. Ramoconidia<br />
are defined as short apical branches (often conidiogenous cells)<br />
of a conidiophore which secede <strong>and</strong> function as conidia. <strong>The</strong>y are<br />
characterised by having a truncate, undifferentiated base, i.e.,<br />
they differ from true conidia by lacking characteristic basal hila<br />
caused by conidiogenesis. Ramoconidia give rise to branched or<br />
unbranched conidia. Secondary ramoconidia are branched conidia<br />
with a narrowed base, bearing a true hilum, that can occur in chains,<br />
giving rise to conidia, which differ from secondary ramoconidia<br />
with regards to shape, size <strong>and</strong> septation. In previous literature on<br />
<strong>Cladosporium</strong> <strong>and</strong> allied genera, the true ramoconidia have often<br />
been classified as “ramoconidia s. str.” whereas the secondary<br />
ramoconidia have been named “ramoconidia s. lat.”<br />
Materials <strong>and</strong> methods<br />
Isolates<br />
Isolates used were obtained from the Centraalbureau voor<br />
Schimmelcultures (<strong>CBS</strong>), or freshly isolated from various<br />
substrates (Table 1). Strains were cultured on 2 % malt extract<br />
plates (MEA; Gams et al. 2007), by obtaining single conidial<br />
colonies as explained in Crous (1998). Colonies were subcultured<br />
onto fresh MEA, oatmeal agar (OA), potato-dextrose agar (PDA)<br />
<strong>and</strong> synthetic nutrient-poor agar (SNA) (Gams et al. 2007), <strong>and</strong><br />
incubated under near-ultraviolet light to study their morphology.<br />
Cultural characteristics were assessed after 2–4 wk on OA <strong>and</strong><br />
PDA at 25 °C in the dark, using the colour charts of Rayner (1970).<br />
Nomenclatural novelties <strong>and</strong> descriptions were deposited in<br />
MycoBank (www.MycoBank.org).<br />
DNA isolation, sequencing <strong>and</strong> phylogeny<br />
Fungal colonies were established on agar plates, <strong>and</strong> genomic<br />
DNA was isolated following the CTAB-based protocol described in<br />
Gams et al. (2007). <strong>The</strong> primers V9G (de Hoog & Gerrits van den<br />
Ende 1998) <strong>and</strong> LR5 (Vilgalys & Hester 1990) were used to amplify<br />
part of the nuclear rDNA operon spanning the 3’ end of the 18S<br />
rRNA gene (SSU), the first internal transcribed spacer (ITS1), the<br />
5.8S rRNA gene, the second ITS region <strong>and</strong> the 5’ end of the 28S<br />
rRNA gene (LSU). Four internal primers, namely ITS4 (White et al.<br />
1990), LR0R (Rehner & Samuels 1994), LR3R (www.biology.duke.<br />
edu/fungi/mycolab/primers.htm), <strong>and</strong> LR16 (Moncalvo et al. 1993),<br />
were used for sequencing to ensure that good quality overlapping<br />
sequences are obtained. <strong>The</strong> PCR conditions, sequence alignment<br />
<strong>and</strong> subsequent phylogenetic analysis followed the methods of<br />
Crous et al. (2006d). <strong>The</strong> ITS1, ITS2 <strong>and</strong> 5.8S rRNA gene (ITS)<br />
www.studiesinmycology.org<br />
Results<br />
DNA extraction, amplification <strong>and</strong> phylogeny<br />
Amplicons of approximately 1 700 bases were obtained for the<br />
isolates listed in Table 1. <strong>The</strong> newly generated sequences were<br />
used to obtain additional sequences from GenBank, which were<br />
added to the alignment. <strong>The</strong> manually adjusted LSU alignment<br />
contained 73 sequences (including the two outgroup sequences)<br />
<strong>and</strong> 996 characters including alignment gaps. Of the 849 characters<br />
used in the phylogenetic analysis, 336 were parsimony-informative,<br />
77 were variable <strong>and</strong> parsimony-uninformative, <strong>and</strong> 436 were<br />
constant. Neighbour-joining analyses using three substitution<br />
models on the sequence data yielded trees with identical topologies<br />
to one another. <strong>The</strong> neighbour-joining trees support the same<br />
clades as obtained from the parsimony analysis, but with a different<br />
arrangement at the deeper nodes, which were poorly supported<br />
in the bootstrap analyses or not at all (for example, the Helotiales<br />
<strong>and</strong> Pleosporales are swapped around). Performing a parsimony<br />
analysis with gaps treated as new characters increases the number<br />
of equally parsimonious trees to 94; the same topology is observed<br />
but with less resolution for the taxa in the Helotiales (data not<br />
shown). Forty-four equally most parsimonious trees (TL = 1 572<br />
steps; CI = 0.436; RI = 0.789; RC = 0.344), one of which is shown<br />
in Fig. 1, were obtained from the parsimony analysis of the LSU<br />
sequence data. <strong>The</strong> cladosporium-like taxa were found to belong to<br />
the Helotiales, Pleosporales, Sordariales <strong>and</strong> as sister taxa to the<br />
Davidiellaceae in the Capnodiales.<br />
<strong>The</strong> LSU alignment used for parsimony <strong>and</strong> distance analysis<br />
was supplemented with sequences for Parapleurotheciopsis<br />
inaequiseptata (Matsush.) P.M. Kirk <strong>and</strong> Subramaniomyces<br />
fusisaprophyticus (Matsush.) P.M. Kirk, as well as related sequences<br />
35
Crous et al.<br />
100<br />
61<br />
100<br />
52<br />
100<br />
10 changes<br />
Athelia epiphylla AY586633<br />
Paullicorticium ansatum AY586693<br />
Rhizocladosporium argillaceum <strong>CBS</strong> 241.67<br />
Myxotrichum deflexum AY541491<br />
Hormoconis resinae <strong>CBS</strong> 365.86<br />
Hormoconis resinae <strong>CBS</strong> 184.54<br />
56<br />
99<br />
99<br />
67<br />
100 84<br />
90<br />
93<br />
81<br />
100<br />
65<br />
75<br />
100<br />
Bisporella citrina AY789385<br />
Sarcoleotia turficola AY789277<br />
Torrendiella eucalypti DQ195799<br />
Torrendiella eucalypti DQ195800<br />
Hyalodendriella betulae <strong>CBS</strong> 261.82<br />
<strong>The</strong>dgonia ligustrina W1877<br />
Blumeria graminis f. sp. bromi AB022362 Erysiphaceae<br />
Neofabraea alba AY064705 Dermateaceae<br />
Neofabraea malicorticis AY544662<br />
Phoma herbarum DQ678066<br />
Ascochyta pisi DQ678070<br />
Didymella bryoniae AB266850<br />
Didymella cucurbitacearum AY293792<br />
Leptospora rubella DQ195792<br />
Phaeosphaeria avenaria AY544684<br />
Coniothyrium palmarum <strong>CBS</strong> 758.73<br />
Ochrocladosporium frigidarii <strong>CBS</strong> 103.81<br />
Ochrocladosporium elatum <strong>CBS</strong> 146.33<br />
Cladoriella eucalypti DQ195790<br />
100 Cladoriella eucalypti <strong>CBS</strong> 115899<br />
100<br />
83<br />
87<br />
85<br />
69<br />
Helotiaceae<br />
Chaetomium homopilatum AF286404<br />
Retroconis fusiformis <strong>CBS</strong> 330.81<br />
Chaetomium globosum AF286403<br />
Aporothielavia leptoderma AF096186<br />
Rachicladosporium luculiae CPC 11407<br />
Dichocladosporium chlorocephalum EU009456<br />
Dichocladosporium chlorocephalum EU009458<br />
Dichocladosporium chlorocephalum EU009457<br />
Dichocladosporium chlorocephalum EU009455<br />
Verrucocladosporium dirinae <strong>CBS</strong> 112794<br />
Toxicocladosporium irritans <strong>CBS</strong> 185.58<br />
97<br />
93<br />
89<br />
100<br />
<strong>Cladosporium</strong> cladosporioides DQ008145<br />
<strong>Cladosporium</strong> uredinicola DQ008147<br />
<strong>Cladosporium</strong> bruhnei DQ008149<br />
<strong>Cladosporium</strong> iridis DQ008148<br />
<strong>Cladosporium</strong> cladosporioides DQ008146<br />
Mycosphaerella marksii AF309578<br />
Penidiella nect<strong>and</strong>rae EU019275<br />
“Stenella” cerophilum AF050286<br />
84<br />
55<br />
100<br />
75<br />
94<br />
100<br />
99<br />
100<br />
94<br />
58<br />
52<br />
58<br />
Incertae sedis<br />
Myxotrichaceae<br />
Amorphotecaceae<br />
Incertae sedis<br />
Incertae sedis<br />
“Mycosphaerella” lateralis AF309583<br />
Pseudocercospora paraguayensis AF309574<br />
Passalora eucalypti AF309575<br />
Mycosphaerella irregulariramosa AF309576<br />
Mycosphaerella punctiformis AY490776<br />
Ramularia aplospora <strong>CBS</strong> 545.82<br />
Passalora daleae <strong>CBS</strong> 113031<br />
Mycosphaerella africana AF309581<br />
Passalora fulva DQ008163<br />
Staninwardia suttonii DQ923535<br />
Devriesia americana <strong>CBS</strong> 117726<br />
Penidiella strumelloidea EU019277<br />
Devriesia thermodurans <strong>CBS</strong> 115878<br />
64<br />
66<br />
Devriesia shelburniensis <strong>CBS</strong> 115876<br />
Devriesia acadiensis <strong>CBS</strong> 115874<br />
Devriesia staurophora DQ008150<br />
Devriesia staurophora DQ008151<br />
Catenulostroma germanicum EU019253<br />
Catenulostroma chromoblastomycosum EU019251<br />
Penidiella rigidophora EU019276<br />
Teratosphaeria alistairii DQ885901<br />
Penidiella columbiana EU019274<br />
Penidiella venezuelensis EU019278<br />
Catenulostroma castellanii EU019250<br />
Teratosphaeria suttonii AF309587<br />
Teratosphaeria molleriana AF309584<br />
Teratosphaeria cryptica AF309585<br />
Teratosphaeria juvenis AF309586<br />
Phaeosphaeriaceae<br />
Leptosphaeriaceae<br />
Incertae sedis<br />
Incertae sedis<br />
Chaetomiaceae, Sordariales<br />
Incertae sedis<br />
Davidiellaceae<br />
Pleosporales<br />
Fig. 1. One of 44 equally most parsimonious trees obtained from a heuristic search with 100 r<strong>and</strong>om taxon additions of the LSU sequence alignment using PAUP v. 4.0b10. <strong>The</strong><br />
scale bar shows 10 changes, <strong>and</strong> bootstrap support values from 1 000 replicates are shown at the nodes. Thickened lines indicate the strict consensus branches <strong>and</strong> ex-type<br />
sequences are printed in bold face. <strong>The</strong> tree was rooted to two sequences obtained from GenBank (Athelia epiphylla AY586633 <strong>and</strong> Paullicorticium ansatum AY586693).<br />
Mycosphaerellaceae<br />
Teratosphaeriaceae<br />
Helotiales<br />
Capnodiales<br />
36
<strong>Cladosporium</strong> <strong>and</strong> morphologically <strong>similar</strong> genera<br />
Athelia epiphylla AY586633<br />
Paullicorticium ansatum AY586693<br />
Blumeria graminis f. sp. bromi AB022362 Erysiphaceae<br />
Bisporella citrina AY789385 Helotiaceae<br />
0.80<br />
Hyalodendriella betulae <strong>CBS</strong> 261.82<br />
<strong>The</strong>dgonia ligustrina W1877<br />
Incertae sedis<br />
1.00<br />
0.69 Neofabraea alba AY064705<br />
Dermateaceae<br />
1.00 Neofabraea malicorticis AY544662<br />
Helotiales<br />
0.60 Sarcoleotia turficola AY789277 Helotiaceae<br />
1.00 Torrendiella eucalypti DQ195799<br />
0.90<br />
Incertae sedis<br />
Torrendiella eucalypti DQ195800<br />
1.00 Hormoconis resinae <strong>CBS</strong> 365.86<br />
Amorphotecaceae<br />
Hormoconis resinae <strong>CBS</strong> 184.54<br />
0.94<br />
Rhizocladosporium argillaceum <strong>CBS</strong> 241.67 Incertae sedis<br />
Myxotrichum deflexum AY541491 Myxotrichaceae<br />
1.00 Chaetomium globosum AF286403<br />
0.88<br />
Aporothielavia leptoderma AF096186<br />
0.95<br />
Chaetomiaceae, Sordariales<br />
1.00 0.79<br />
Chaetomium homopilatum AF286404<br />
Retroconis fusiformis <strong>CBS</strong> 330.81<br />
1.00 Fasciatispora petrakii AY083828 Xylariaceae<br />
0.81<br />
Phlogicylindrium eucalypti DQ923534 Incertae sedis<br />
Plectosphaera eucalypti DQ923538 Phyllachoraceae<br />
0.72<br />
Xylariales<br />
0.74 Subramaniomyces fusisaprophyticus <strong>CBS</strong> 418.95 Incertae sedis<br />
0.68 Parapleurotheciopsis inaequiseptata MUCL 41089 Incertae sedis<br />
Pseudomassaria carolinensis DQ810233 Hyponectriaceae<br />
Phoma herbarum DQ678066<br />
0.63 1.00 Ascochyta pisi DQ678070<br />
1.00 0.97<br />
Incertae sedis<br />
Didymella bryoniae AB266850<br />
1.00 Didymella cucurbitacearum AY293792<br />
Leptospora rubella DQ195792<br />
Pleosporales<br />
1.00<br />
Phaeosphaeriaceae<br />
Phaeosphaeria avenaria AY544684<br />
0.78 Coniothyrium palmarum <strong>CBS</strong> 758.73 Leptosphaeriaceae<br />
0.93 Ochrocladosporium frigidarii <strong>CBS</strong> 103.81<br />
1.00 Ochrocladosporium elatum <strong>CBS</strong> 146.33<br />
Incertae sedis<br />
1.00 Cladoriella eucalypti <strong>CBS</strong> 115899<br />
Incertae sedis<br />
1.00 Cladoriella eucalypti DQ195790<br />
Rachicladosporium luculiae CPC 11407<br />
0.85 Dichocladosporium chlorocephalum EU009456<br />
Dichocladosporium chlorocephalum EU009458<br />
0.68 Dichocladosporium chlorocephalum EU009457 Incertae sedis<br />
Dichocladosporium chlorocephalum EU009455<br />
0.89 Verrucocladosporium dirinae <strong>CBS</strong> 112794<br />
Toxicocladosporium irritans <strong>CBS</strong> 185.58<br />
0.90 <strong>Cladosporium</strong> cladosporioides DQ008145<br />
1.00 <strong>Cladosporium</strong> uredinicola DQ008147<br />
0.97 1.00 <strong>Cladosporium</strong> bruhnei DQ008149<br />
Davidiellaceae<br />
0.61 <strong>Cladosporium</strong> iridis DQ008148<br />
1.00 <strong>Cladosporium</strong> cladosporioides DQ008146<br />
Mycosphaerella marksii AF309578<br />
1.00 Penidiella nect<strong>and</strong>rae EU019275<br />
“Stenella” cerophilum AF050286<br />
0.71 “Mycosphaerella” lateralis AF309583<br />
1.00 Pseudocercospora paraguayensis AF309574<br />
0.99 Passalora eucalypti AF309575<br />
0.51 Mycosphaerella irregulariramosa AF309576<br />
1.00 Mycosphaerella punctiformis AY490776<br />
0.86 Ramularia aplospora <strong>CBS</strong> 545.82<br />
Passalora daleae <strong>CBS</strong> 113031<br />
1.00 Mycosphaerella africana AF309581<br />
1.00 Passalora fulva DQ008163<br />
0.94 1.00 Staninwardia suttonii DQ923535<br />
Devriesia americana <strong>CBS</strong> 117726<br />
Penidiella strumelloidea EU019277<br />
1.00 Devriesia thermodurans <strong>CBS</strong> 115878<br />
1.00<br />
Devriesia shelburniensis <strong>CBS</strong> 115876<br />
0.99 Devriesia acadiensis <strong>CBS</strong> 115874<br />
0.86 Devriesia staurophora DQ008150<br />
Devriesia staurophora DQ008151<br />
0.67<br />
Catenulostroma germanicum EU019253<br />
0.65 Penidiella venezuelensis EU019278<br />
Catenulostroma castellanii EU019250<br />
0.1 expected changes per site 0.73 Teratosphaeria suttonii AF309587<br />
1.00 Teratosphaeria molleriana AF309584<br />
1.00 Teratosphaeria cryptica AF309585<br />
Teratosphaeria juvenis AF309586<br />
1.00 Catenulostroma chromoblastomycosum EU019251<br />
0.64<br />
1.00<br />
Penidiella rigidophora EU019276<br />
Teratosphaeria alistairii DQ885901<br />
Penidiella columbiana EU019274<br />
Mycosphaerellaceae<br />
Teratosphaeriaceae<br />
Capnodiales<br />
Fig. 2. Consensus phylogram (50 % majority rule) of 800 trees resulting from a Bayesian analysis of the LSU sequence alignment using MrBayes v. 3.1.2. Bayesian posterior<br />
probabilities are indicated at the nodes. Ex-type sequences are printed in bold face. <strong>The</strong> tree was rooted to two sequences obtained from GenBank (Athelia epiphylla AY586633<br />
<strong>and</strong> Paullicorticium ansatum AY586693).<br />
www.studiesinmycology.org<br />
37
Crous et al.<br />
Fig. 3. Rachicladosporium luculiae (type material). A–F. Conidiophores with conidial chains, <strong>and</strong> conidiogenous loci aggregated in the upper region. G. Conidia. Scale bar =<br />
10 µm.<br />
from GenBank. This alignment was subjected to a Bayesian<br />
analysis using a general time-reversible (GTR) substitution model<br />
with inverse gamma rates <strong>and</strong> dirichlet base frequencies <strong>and</strong> the<br />
temp value set to 0.5. <strong>The</strong> Markov Chain Monte Carlo (MCMC)<br />
analysis of 4 chains started from a r<strong>and</strong>om tree topology <strong>and</strong> lasted<br />
1 000 000 generations. Trees were saved each 1 000 generations,<br />
resulting in 1 000 trees. Burn-in was set at 200 000 generations<br />
after which the likelihood values were stationary, leaving 800 trees<br />
from which the consensus tree (Fig. 2) <strong>and</strong> posterior probabilities<br />
(PP’s) were calculated. <strong>The</strong> average st<strong>and</strong>ard deviation of split<br />
frequencies was 0.018459 at the end of the run. <strong>The</strong> same overall<br />
topology as that observed using parsimony was obtained, with<br />
the main exception that the Helotiales <strong>and</strong> Pleosporales swapped<br />
around, as observed with the distance analysis.<br />
Taxonomy<br />
<strong>The</strong> present study has delineated several cladosporium-like genera<br />
which are phylogenetically unrelated to, <strong>and</strong> morphologically distinct<br />
from <strong>Cladosporium</strong> s. str. (Davidiellaceae, Capnodiales). <strong>The</strong>se are<br />
treated below:<br />
Capnodiales, incertae sedis<br />
Rachicladosporium Crous, U. Braun & C.F. Hill, gen. nov.<br />
MycoBank MB504430.<br />
Etymology: Named after the apical rachis on conidiophores, <strong>and</strong> its<br />
cladosporium-like appearance.<br />
Differt a Cladosporio conidiophoris cum rachibus terminalibus, locis conidiogenis<br />
inconspicuis vel subconspicuis, margine leviter incrassatis, non fuscatis et non<br />
refractivis, hilis inconspicuis.<br />
Mycelium consisting of branched, septate, smooth, hyaline to<br />
pale brown, thin-walled hyphae. Conidiophores erect, solitary,<br />
macronematous, arising from superficial hyphae, subcylindrical,<br />
straight to somewhat geniculate-sinuous, medium brown,<br />
finely verruculose; basal foot cell without swelling or rhizoids.<br />
Conidiogenous cells integrated, terminal, subcylindrical or tips<br />
slightly swollen, forming an apical rachis, multilocal, loci terminal<br />
<strong>and</strong> lateral, without evident sympodial proliferation (non-geniculate);<br />
conidiogenous loci inconspicuous or subconspicuous by being very<br />
slightly thickened along the rim, but neither darkened nor refractive,<br />
giving rise to simple or branched chains or solitary conidia.<br />
Ramoconidia medium brown, finely verruculose, 0–1-septate,<br />
subcylindrical to narrowly ellipsoid; conidia ellipsoid, pale brown,<br />
0(–1)-septate, smooth to finely verruculose; hila inconspicuous;<br />
secession schizolytic.<br />
Type species: Rachicladosporium luculiae Crous, U. Braun & C.F.<br />
Hill, sp. nov.<br />
38
<strong>Cladosporium</strong> <strong>and</strong> morphologically <strong>similar</strong> genera<br />
Rachicladosporium luculiae Crous, U. Braun & C.F. Hill, sp. nov.<br />
MycoBank MB504431. Fig. 3.<br />
Etymology: Named after its host <strong>genus</strong>, Luculia.<br />
Mycelium ex hyphis ramosis, septatis, levibus, hyalinis vel pallide brunneis, 2–3<br />
µm latis compositum. Conidiophora erecta, solitaria, macronemata, ex hyphis<br />
superficialibis oriunda, subcylindrica, recta to geniculata-sinuosa, ad 60 µm longa et<br />
6 µm lata, 3–6-septata, modice brunnea, subtiliter verruculosa, non crassitunicata, ad<br />
basim non inflatae et non rhizoideae. Cellulae conidiogenae integratae, terminales,<br />
8–15 × 4–5 µm, subcylindricae, apicem versus attenuatae, apice obtuso, rachidi<br />
terminali, locis conidialibus numerosis, 1–2 µm latis, margine leviter incrassatis,<br />
non fuscatis et non refractivis. Conidia catenata vel solitaria. Ramoconidia modice<br />
brunnea, subtile verruculosa, 0–1-septata, subcylindrica vel anguste ellipsoidea,<br />
10–17 × 4–5 µm; conidia secundaria ellipsoidea, pallide brunnea, 0(–1)-septata,<br />
levia vel subtile verruculosa, interdum guttulata, (7–)9–12(–15) × 3(–4) µm; hila<br />
inconspicua.<br />
Mycelium consisting of branched, septate, smooth, thin-walled,<br />
hyaline to pale brown, 2–3 µm wide hyphae. Conidiophores<br />
erect, solitary, macronematous, arising from superficial hyphae,<br />
subcylindrical, straight to somewhat geniculate-sinuous, up to 60 µm<br />
long, <strong>and</strong> 6 µm wide, 3–6-septate, medium brown, finely verruculose,<br />
thin-walled (≤ 1 µm), rarely with a single percurrent proliferation;<br />
basal foot cell without swelling or rhizoids. Conidiogenous cells<br />
integrated, terminal, 8–15 × 4–5 µm, subcylindrical, tapering to an<br />
obtuse apex, occasionally slightly swollen at the tip, without distinct<br />
sympodial proliferation (non-geniculate), forming a rachis, with<br />
several conidiogenous loci, terminal <strong>and</strong> lateral, 1–2 µm wide, nonprotuberant,<br />
quite inconspicuous to subconspicuous, very slightly<br />
thickened along the rim, but not darkened <strong>and</strong> refractive; giving rise<br />
to simple or branched chains or solitary conidia, thin-walled (≤ 0.75<br />
µm). Ramoconidia medium brown, finely verruculose, 0–1-septate,<br />
subcylindrical to narrowly ellipsoid, 10–17 × 4–5 µm; conidia<br />
ellipsoid, pale brown, 0(–1)-septate, smooth to finely verruculose,<br />
at times guttulate, (7–)9–12(–15) × 3(–4) µm; hila inconspicuous,<br />
neither thickened nor darkened-refractive.<br />
Cultural characteristics: Colonies on PDA erumpent, spreading,<br />
with moderate aerial mycelium <strong>and</strong> smooth, even margins; irongrey<br />
in the centre, olivaceous-grey in the outer region (surface);<br />
iron-grey underneath. Colonies reaching 4 cm diam after 1 mo at<br />
25 °C in the dark.<br />
Specimen examined: New Zeal<strong>and</strong>, Auckl<strong>and</strong>, isolated from leaf spots on Luculia<br />
sp. (Rubiaceae), 25 Jul. 2004, F. Hill 1059, holotype <strong>CBS</strong> H-19891, culture ex-type<br />
<strong>CBS</strong> 121620 = CPC 11407.<br />
Notes: Rachicladosporium is morphologically quite distinct from<br />
<strong>Cladosporium</strong> s. str. <strong>and</strong> allied cladosporioid genera by having an<br />
apical conidiophore rachis with inconspicuous to subconspicuous<br />
scars <strong>and</strong> unthickened, not darkened-refractive conidial hila. Due<br />
to the structure of the conidiogenous cells, R. luculiae superficially<br />
resembles species of the tretic <strong>genus</strong> Diplococcium Grove (Ellis<br />
1971, 1976; Goh & Hyde 1998). However, there is no evidence<br />
for a tretic conidiogenesis in R. luculiae. <strong>The</strong> conidia are formed<br />
holoblastically <strong>and</strong> separated by a thin septum. Furthermore,<br />
in Diplococcium the conidiogenous cells are terminal as well as<br />
intercalary, the conidiophores are often branched, <strong>and</strong> branched<br />
conidial chains are lacking or at least less common. Molecular<br />
sequence data about Diplococcium species are not yet available,<br />
though taxa that have been analysed show affinities to the<br />
Pleosporaceae <strong>and</strong> Helotiales (Wang et al., unpubl. data), whereas<br />
Rachicladosporium is allied with the Capnodiales. <strong>The</strong> ecology<br />
of R. luculiae is still unclear, although it has been isolated from<br />
lesions on Luculia sp. Fruiting of this species in vivo has not yet<br />
been observed, <strong>and</strong> its pathogenicity remains unproven.<br />
www.studiesinmycology.org<br />
Toxicocladosporium Crous & U. Braun, gen. nov. MycoBank<br />
MB504426.<br />
Etymology: Named after ample volatile metabolites produced in<br />
culture, <strong>and</strong> cladosporium-like morphology.<br />
Differt a Cladosporio locis conidiogenis denticulatis, incrassatis et fuscatis-refractivis,<br />
sed non coronatis, conidiophoris et conidiis cum septis incrassatis et atrofuscis, et<br />
culturis cum metabolitis volaticis toxicis.<br />
Mycelium consisting of branched, septate, dark brown, finely<br />
verruculose hyphae. Conidiophores solitary, dimorphic, solitary,<br />
macronematous or micronematous, reduced to conidiogenous<br />
cells. Macronematous conidiophores subcylindrical, straight<br />
to geniculate-sinuous, or irregularly curved, unbranched or<br />
branched above, septate, dark brown, finely verruculose, walls<br />
thick, septa dark brown; micronematous conidiophores reduced<br />
to conidiogenous cells, erect, doliiform to subcylindrical, with slight<br />
taper towards the apex. Conidiogenous cells integrated, terminal or<br />
lateral, subcylindrical with slight taper towards apex; proliferating<br />
sympodially with apical loci protruding <strong>and</strong> denticle-like, thickened,<br />
darkened <strong>and</strong> refractive, but not coronate. Conidia catenulate in<br />
branched or unbranched chains, medium to dark brown, thickwalled,<br />
with dark, thick septa, smooth to finely verruculose;<br />
ramoconidia septate, prominently constricted at septa, broadly<br />
ellipsoid to subcylindrical; conidia ellipsoid to ovoid, pale to medium<br />
brown, 0(–1)-septate; hila not coronate, but protruding, thickened,<br />
darkened <strong>and</strong> refractive in ramoconidia, but less obvious in young<br />
conidia.<br />
Type species: Toxicocladosporium irritans Crous & U. Braun, sp.<br />
nov.<br />
Toxicocladosporium irritans Crous & U. Braun, sp. nov.<br />
MycoBank MB504427. Fig. 4.<br />
Etymology: Named after the skin irritation resulting from exposure<br />
to the fungus.<br />
Mycelium (in PDA) ex hyphis ramosis, septatis, atro-brunneis, minute verruculosis,<br />
(2–)3–4 µm latis, ultimo crassitunicatis et crassiseptatis. Conidiophora solitaria,<br />
dimorphosa, macronemata et solitaria vel micronemata. Conidiophora macronemata<br />
ex hyphis modice brunneis lateraliter oriunda, erecta, subcylindrica, recta,<br />
geniculata-sinuosa vel irregulariter curvata, non ramosa vel ad apicem ramosa,<br />
2–7-septata, atro-brunnea, leviter verruculosa, crassitunicata, septa atro-brunnea,<br />
30–60 × 4–6 µm; conidiophora micronemata saepe non septata, raro 1–2-septata,<br />
erecta, doliiformes vel subcylindrica, apicem versus leviter attenuata, 10–30 × 2.5–<br />
4 µm. Cellulae conidiogenae integratae, terminales vel laterales, subcylindricae,<br />
apicem versus leviter attenuatae, 7–12 × 3–4 µm, sympodiales, cum 1–3 locis<br />
conidiogenibus, denticulatis, 1–1.5 µm latis, incrassatis, fuscatis-refractivis. Conidia<br />
catenulata vel rami-catenulata, modice vel atro-brunnea, crassitunicata, septis<br />
incrassatis, fuscatis, levia vel subtile verruculosa; ramoconidia (0–)1(–3)-septata,<br />
constricta, late ellipsoidea vel subcylindrica, 7–15 × 3–5 µm; conidia secundaria<br />
ellipsoidea vel ovoidea, pallide vel modice brunnea, 0(–1)-septata, (5–)6–8(–10) ×<br />
(3–)4(–5) µm; hila protuberantes, 1–1.5 µm lata, hila ramoconidiorum incrassata<br />
et fuscata-refractiva, vel hila conidiorum secundariorum 0.5–1 µm lata et<br />
subconspicua.<br />
Mycelium on PDA consisting of branched, septate, dark brown, finely<br />
verruculose, (2–)3–4 µm wide hyphae; walls <strong>and</strong> septa becoming<br />
thickened <strong>and</strong> darkened with age. Conidiophores solitary, dimorphic,<br />
macronematous <strong>and</strong> solitary, or micronematous, reduced to<br />
conidiogenous cells. Macronematous conidiophores subcylindrical,<br />
straight to geniculate-sinuous, or irregularly curved, unbranched<br />
or branched above, 2–7-septate, dark brown, finely verruculose,<br />
walls thick, septa dark brown, 30–60 × 4–6 µm; medium brown<br />
hyphae giving rise to lateral, erect branches that become swollen,<br />
dark brown, <strong>and</strong> develop into macronematous conidiophores<br />
with thick-walled <strong>and</strong> dark septa; micronematous conidiophores<br />
39
Crous et al.<br />
Fig. 4. Toxicocladosporium irritans (type material). A–B, F. Microconidiophores. C–E. Macroconidiophores. G–H. Ramoconidia <strong>and</strong> conidia. Scale bars = 10 µm.<br />
aseptate, reduced to conidiogenous cells (rarely 1–2-septate, i.e.,<br />
with 1–2 supporting cells), erect, doliiform to subcylindrical, with<br />
slight taper towards the apex, 10–30 × 2.5–4 µm. Conidiogenous<br />
cells integrated, terminal or lateral, subcylindrical with slight taper<br />
towards apex, 7–12 × 3–4 µm; proliferating sympodially with 1–3<br />
apical loci that can be slightly protruding <strong>and</strong> denticle-like, 1–1.5<br />
µm wide, thickened, darkened <strong>and</strong> refractive. Conidia catenulate<br />
in branched or unbranched chains, medium to dark brown, thickwalled,<br />
with dark, thick septa, smooth to finely verruculose;<br />
ramoconidia (0–)1(–3)-septate, prominently constricted at septa,<br />
broadly ellipsoid to subcylindrical, 7–15 × 3–5 µm; conidia ellipsoid<br />
to ovoid, younger apical conidia pale to medium brown, 0(–1)-<br />
septate, (5–)6–8(–10) × (3–)4(–5) µm; hila protruding, 1–1.5 µm<br />
wide, thickened, darkened <strong>and</strong> refractive in ramoconidia, but less<br />
obvious in young conidia, where hila are 0.5–1 µm wide.<br />
Cultural characteristics: Colonies on PDA erumpent, spreading,<br />
with dense aerial mycelium <strong>and</strong> smooth, even margins; surface<br />
olivaceous-black (centre), olivaceous-grey in outer region; reverse<br />
olivaceous-black. Colonies reaching 35 mm diam after 1 mo at 25<br />
°C in the dark; colonies fertile.<br />
40
<strong>Cladosporium</strong> <strong>and</strong> morphologically <strong>similar</strong> genera<br />
Specimen examined: Suriname, Paramaribo, isolated from mouldy paint, Feb.<br />
1958, M.B. Schol-Schwarz, holotype <strong>CBS</strong>-H 19892, culture ex-type <strong>CBS</strong> 185.58.<br />
Notes: Toxicocladosporium irritans produces ample amounts<br />
of volatile metabolites, which cause a skin rash within minutes<br />
of opening an inoculated dish for microscopic examination.<br />
Morphologically <strong>and</strong> phylogenetically it is very <strong>similar</strong> to<br />
<strong>Cladosporium</strong> s. str., <strong>and</strong> produces dimorphic conidiophores, which<br />
is also commonly observed in <strong>Cladosporium</strong>. It is distinct by having<br />
dark, thick-walled conidial <strong>and</strong> conidiophore septa, <strong>and</strong> lacking the<br />
typical coronate <strong>Cladosporium</strong> scar type (David 1997).<br />
Verrucocladosporium K. Schub., Aptroot & Crous, gen. nov.<br />
MycoBank MB504432.<br />
Etymology: Named after its frequently coarsely verrucose to<br />
warted hyphae, conidiophores <strong>and</strong> conidia, <strong>and</strong> cladosporium-like<br />
morphology.<br />
Differt a Cladosporio hyphis saepe verrucosis, hyalinis, conidiophoris cylindraceisfiliformibus,<br />
rectis, non vel vix geniculatis, non nodulosis, locis conidiogenis leviter<br />
incrassatis, distincte fuscatis-refractivis, sed non coronatis, conidiis saepe valde<br />
variantibus, saepe irregulariter formatis, grosse verrucosis-rugosis.<br />
Mycelium sparingly branched, hyphae septate, not constricted at<br />
septa, hyaline, almost smooth to irregularly rough-walled, coarsely<br />
verrucose to warted. Conidiophores arising laterally from creeping<br />
hyphae, erect, straight, or somewhat flexuous, narrowly cylindrical<br />
to filiform, neither geniculate nor nodulose, unbranched, septate,<br />
pale brown, thin-walled, smooth to often irregularly rough-walled or<br />
verrucose. Conidiogenous cells integrated, terminal or intercalary,<br />
cylindrical, polyblastic, with sympodial proliferation, with loci often<br />
crowded at the apex, truncate, barely to slightly thickened, but<br />
distinctly darkened-refractive. Ramoconidia cylindrical, aseptate,<br />
concolorous with conidiophores, thin-walled, irregularly roughwalled,<br />
coarsely verruculose to verrucose-rugose; hila unthickened<br />
but somewhat refractive. Conidia in long unbranched or loosely<br />
branched chains, obovoid, ellipsoid, fusiform to subcylindrical, with<br />
swollen <strong>and</strong> constricted parts, often appearing irregular in shape<br />
<strong>and</strong> outline, 0–1-septate, pale brown, thin-walled <strong>and</strong> irregularly<br />
rough-walled, verruculose-rugose; hila truncate, barely to slightly<br />
thickened, but distinctly darkened-refractive.<br />
Type species: Verrucocladosporium dirinae K. Schub., Aptroot &<br />
Crous, sp. nov.<br />
Verrucocladosporium dirinae K. Schub., Aptroot & Crous, sp.<br />
nov. MycoBank MB504433. Fig. 5.<br />
Etymology: Named after its host, Dirina massiliensis.<br />
Mycelium sparse ramosum. Hyphae 1–3 µm latae, septatae, non constrictae,<br />
hyalinae, leviae, vel irregulariter verruculosae, interdum verrucosae, tuberculatae,<br />
tenuitunicatae. Conidiophora ex hyphis repentibus lateraliter oriunda, erecta,<br />
recta, interdum leviter flexuosa, anguste cylindrica vel filiformes, non geniculta,<br />
non nodulosa, non ramosa, ad 85 µm longa, 2–3 µm lata, septata, tenuitunicata<br />
(≤ 0.75 µm), pallide brunnea, levia vel saepe irregulariter verrucosa, leviter<br />
crassitunicata. Cellulae conidiogenae integratae, saepe terminales, interdum<br />
intercalares, cylindricae, angustae, 9–20 µm longae, holoblasticae, sympodiales,<br />
locis conidiogenibus 1–3, saepe ad apicem aggregatis, interdum protuberantibus,<br />
truncatis, 1–1.8(–2) µm latis, incrassatis et fuscatis-refractivis. Ramoconidia<br />
cylindrica, 16–21 × (2–)2.5–3 µm, non septata, pallide brunnea, tenuitunicata,<br />
irregulariter verruculosa vel crosse verrucosa-rugosa, ad 4 hilis terminalibus,<br />
ad basim late truncata, non attenuata, 2–2.5 µm lata, non incrassata, sed leviter<br />
refractiva. Conidia catenata, in catenis longis, non ramosis vel laxe ramosis, plus<br />
minusve recta, obovoidea, ellipsoidea, fusiformes vel subcylindricae, sed saepe<br />
irregulares, 4–18(–23) × (2–)2.5–3.5 µm, 0–1-septata, ad septa interdum constricta,<br />
pallide brunnea, tenuitunicata (≤ 0.5 µm), irregulariter verruculosa-rugosa, utrinque<br />
leviter attenuata, hila truncata, (0.5–)0.8–1.5(–2) µm lata, vix vel leniter incrassata,<br />
sed distincte fuscata-refractiva.<br />
Mycelium sparingly branched; hyphae 1–3 µm wide, septate, not<br />
constricted at septa, hyaline, smooth to irregularly rough-walled,<br />
sometimes coarsely verrucose, with small to large drop-like,<br />
tuberculate warts, walls unthickened. Conidiophores arising laterally<br />
from creeping hyphae, erect, straight, sometimes slightly flexuous,<br />
narrowly cylindrical to filiform, not geniculate, non nodulose,<br />
unbranched, up to 85 µm long, 2–3 µm wide, septate, thin-walled<br />
(≤ 0.75 µm), pale brown, smooth to often irregularly rough-walled,<br />
verrucose, walls slightly thickened. Conidiogenous cells integrated,<br />
mostly terminal, sometimes also intercalary, cylindrical, narrow,<br />
9–20 µm long, conidiogenesis holoblastic, proliferation sympodial,<br />
with a single or up to three conidiogenous loci, often crowded at<br />
the apex, sometimes situated on small lateral prolongations, loci<br />
truncate, 1–1.8(–2) µm wide, thickened <strong>and</strong> darkened-refractive.<br />
Ramoconidia cylindrical, 16–21 × (2–)2.5–3 µm, aseptate,<br />
concolorous with conidiophores, thin-walled, irregularly roughwalled,<br />
verruculose to coarsely verrucose-rugose, apically with up<br />
to 4 hila, with a broadly truncate, non-attenuated base, 2–2.5 µm<br />
wide, unthickened but somewhat refractive. Conidia catenate, in<br />
long unbranched or loosely branched chains, more or less straight,<br />
obovoid, ellipsoid, fusiform to subcylindrical, but often appearing<br />
to form b<strong>and</strong>-like structures, with swollen <strong>and</strong> constricted parts,<br />
accordion or fir tree-like <strong>and</strong> also due to ornamentation often<br />
appearing irregular in shape <strong>and</strong> outline, 4–18(–23) × (2–)2.5–<br />
3.5 µm, 0–1-septate, sometimes constricted at the more or less<br />
median septum, pale brown, thin-walled (≤ 0.5 µm), irregularly<br />
rough-walled, verruculose-rugose, somewhat attenuated towards<br />
apex <strong>and</strong> base, hila truncate, (0.5–)0.8–1.5(–2) µm wide, barely<br />
or slightly thickened, but distinctly darkened-refractive; microcyclic<br />
conidiogenesis not observed.<br />
Cultural characteristics: Colonies erumpent, spreading, with<br />
catenate, feathery margins <strong>and</strong> moderate aerial mycelium on PDA.<br />
Surface grey-olivaceous, reverse iron-grey. Colonies reaching 25<br />
mm after 1 mo at 25 °C.<br />
Specimen examined: U.K., Somerset, Kingsbury Episcopi, isolated from the lichen<br />
Dirina massiliensis (Roccelaceae, Arthoniales), Mar. 2003, A. Aptroot, holotype<br />
<strong>CBS</strong>-H 19883, culture ex-type <strong>CBS</strong> 112794.<br />
Notes: Verrucocladosporium dirinae was deposited as<br />
<strong>Cladosporium</strong> arthoniae M. Christ. & D. Hawksw., but the name<br />
was misapplied. <strong>The</strong> latter species, described from apothecia of<br />
Arthonia impolita on Quercus from Sweden, does not possess<br />
clearly visible, distinct conidiogenous loci <strong>and</strong> hila, <strong>and</strong> therefore<br />
has to be excluded from the <strong>genus</strong> <strong>Cladosporium</strong> s. str. <strong>and</strong> is also<br />
easily distinguishable from the newly introduced species above.<br />
Furthermore the conidiophores are apically frequently branched <strong>and</strong><br />
the catenate, ellipsoid conidia are smaller <strong>and</strong> wider, 6–10 × 4–5 µm<br />
(Hawksworth 1979). Due to the conidiogenesis <strong>and</strong> the structure of<br />
the conidiogenous loci <strong>and</strong> conidia, C. arthoniae is rather close to<br />
lichenicolous Taeniolella S. Hughes species. <strong>The</strong> unique feature<br />
of the new <strong>genus</strong> Verrucocladosporium is its unusual conidial <strong>and</strong><br />
hyphal ornamentation. Furthermore, it differs from <strong>Cladosporium</strong><br />
s. str. in having cylindrical-filiform conidiophores, which are neither<br />
geniculate nor nodulose, quite distinct, thickened <strong>and</strong> darkened,<br />
but non-coronate conidiogenous loci <strong>and</strong> often irregularly shaped<br />
conidia. Phylogenetially, it is also distinct as a sister taxon to<br />
<strong>Cladosporium</strong> s. str. Concerning differences to other cladosporioid<br />
genera, see “key to the genera”. Verrucocladosporium dirinae has<br />
been isolated from the lichen species Dirina massiliensis, i.e., this<br />
species is probably lichenicolous, although its ecology is not quite<br />
clear. Fruiting of this species in vivo has not yet been observed. A<br />
second unnamed, taeniolella-like, lichenicolous hyphomycete was<br />
also present on the thallus of this lichen.<br />
www.studiesinmycology.org<br />
41
Crous et al.<br />
Fig. 5. Verrucocladosporium dirinae (type material). A. Colonies on MEA. B–C. Conidial chains. D–H. Ramoconidia <strong>and</strong> conidia. Scale bars = 10 µm.<br />
Capnodiales, Teratosphaeriaceae<br />
Devriesia americana Crous & Dugan, sp. nov. MycoBank<br />
MB504434. Fig. 6.<br />
Etymology: Named after the geographic location of its type strain,<br />
New York, U.S.A.<br />
Differt a D. shelburniensi conidiophoris brevioribus (ad 30 µm longis), leviter<br />
latioribus (2–3 µm), ramoconidiis saepe nullis et conidiis 0–1-septatis.<br />
Mycelium consisting of branched, septate, 1.5–3 µm wide hyphae,<br />
irregular in width, predominantly guttulate, smooth, forming hyphal<br />
str<strong>and</strong>s <strong>and</strong> hyphal coils; hyphae frequently forming dark brown,<br />
thick-walled, intercalary, muriformly septate chlamydospores on<br />
PDA in culture. Conidiophores subcylindrical, medium brown,<br />
straight to irregularly curved, up to 7-septate <strong>and</strong> 30 µm tall, 2–3<br />
µm wide, or reduced to conidiogenous cells. Conidiogenous cells<br />
terminal or lateral on hyphae, 5–12 × 2–3 µm, medium brown,<br />
smooth, guttulate, subcylindrical, mono- to polyblastic; scars<br />
somewhat darkened <strong>and</strong> thickened, but not refractive. Conidia<br />
medium brown, guttulate, smooth, in mostly unbranched chains,<br />
subcylindrical to narrowly ellipsoidal, tapering towards subtruncate<br />
ends, 0–1-septate, (7–)8–12(–16) × 2(–2.5) µm; hila darkened,<br />
somewhat thickened, not refractive, 1–1.5 µm wide.<br />
42
<strong>Cladosporium</strong> <strong>and</strong> morphologically <strong>similar</strong> genera<br />
Fig. 6. Devriesia americana (type material). A–B. Chlamydospore-like structures formed in culture. C–F. Conidiophores giving rise to conidial chains. G–H. Conidia. Scale bars<br />
= 10 µm<br />
Cultural characteristics: Colonies erumpent, with sparse aerial<br />
mycelium on PDA, <strong>and</strong> smooth, uneven, wide margins, submerged<br />
under the agar surface; greenish-black (surface); reverse<br />
olivaceous-black; on OA iron-grey (surface). Colonies reaching<br />
8–15 mm diam on PDA after 14 d at 25 °C in the dark; colonies<br />
fertile, but sporulation sparse.<br />
Specimen examined: U.S.A., New York, Long Isl<strong>and</strong>, isolated from air, F.M. Dugan,<br />
holotype <strong>CBS</strong>-H 19894, culture ex-type <strong>CBS</strong> 117726 = ATCC 96545 = CPC 5121.<br />
Notes: Until recently, this species was treated as part of the<br />
“Phaeoramularia” hachijoensis species complex (Braun et al.<br />
2003). <strong>The</strong> present strain has conidia that are smaller than those<br />
of “Phaeoramularia” hachijoensis, which has ramoconidia that are<br />
1–3-septate, up to 30 µm long, <strong>and</strong> conidia that are predominantly<br />
1-septate, 10–21 × 2–4 µm (Matsushima 1975). From the illustration<br />
provided by Matsushima, it appears that “Phaeoramularia”<br />
hachijoensis is indeed a species of Pseudocladosporium U. Braun,<br />
a finding which is in agreement with the name Pseudocladosporium<br />
hachijoense (Matsush.) U. Braun proposed by Braun (1998).<br />
Devriesia americana is both morphologically <strong>and</strong> phylogenetically<br />
more allied to Teratosphaeria Syd. & P. Syd. than Venturia<br />
Sacc. Based on its pigmented conidiophores <strong>and</strong> catenulate<br />
conidia, <strong>and</strong> scars that are somewhat darkened <strong>and</strong> thickened,<br />
<strong>and</strong> the formation of chlamydospores in culture, it is allocated to<br />
Devriesia Seifert & N.L. Nick. Species of the <strong>genus</strong> Devriesia are<br />
ecologically different, however (Seifert et al. 2004), being soil-borne<br />
www.studiesinmycology.org<br />
<strong>and</strong> thermotolerant. It is possible, therefore, that further collections<br />
of this fungus may eventually indicate that it needs to be placed in a<br />
distinct <strong>genus</strong> within the Teratosphaeriaceae. Devriesia americana<br />
is the second species of Devriesia with muriform chlamydospores,<br />
beside D. shelburniensis N.L. Nick. & Seifert, but the latter species<br />
is easily distinguishable by its long <strong>and</strong> narrow conidiophores (ca<br />
100–200 × 1.5–2.5 µm) <strong>and</strong> abundant ramoconidia, up to 25.5<br />
µm long, with 0–3 septa. Furthermore, D. shelburniensis is a<br />
thermotolerant soil-borne hyphomycete.<br />
Stenella araguata Syd., Ann. Mycol. 28(1/2): 205. 1930. Figs 7–8.<br />
≡ <strong>Cladosporium</strong> araguatum (Syd.) Arx, Genera of Fungi Sporulating in<br />
pure Culture, Edn 2 (Vaduz): 224. 1974.<br />
= <strong>Cladosporium</strong> castellanii Borelli & Marcano, Castellania 1: 154. 1973.<br />
Leaf spots hypophyllous, irregular to subcircular, up to 8 mm<br />
diam, indistinct, yellow to pale brown with indistinct margins on<br />
IMI 15728(a); on IMI 34905 (Fig. 7) lesions are amphigenous, <strong>and</strong><br />
fascicles <strong>and</strong> sporodochia are rare, with superficial mycelium being<br />
predominant. Mycelium consisting of internal <strong>and</strong> external, medium<br />
brown, septate, branched, verruculose, 3–4 µm wide hyphae.<br />
Caespituli fasciculate to sporodochial, hypophyllous, medium brown,<br />
up to 120 µm wide <strong>and</strong> 60 µm high. Conidiophores arising singly<br />
from superficial mycelium, or aggregated in loose to dense fascicles<br />
arising from the upper cells of a brown stroma up to 70 µm wide <strong>and</strong><br />
30 µm high; conidiophores medium brown, finely verruculose, 1–5-<br />
septate, subcylindrical, straight to geniculate-sinuous, unbranched<br />
43
Crous et al.<br />
Fig. 7. Stenella araguata (syntype material, IMI 34905). A. Leaf spot. B. Conidiophore, conidia <strong>and</strong> verruculose hypha on leaf surface. C–D. Conidiophore with terminal<br />
conidiogenous cells. E–G. Ramoconidia <strong>and</strong> conidia. Scale bars = 10 µm.<br />
or branched, 20–40 × 3–4 µm. Conidiogenous cells terminal or<br />
lateral, unbranched, medium brown, finely verruculose, tapering<br />
to slightly or flat-tipped loci, proliferating sympodially, 5–20 × 3–4<br />
µm; scars thickened, darkened <strong>and</strong> refractive. Conidia solitary<br />
or catenulate, in simple chains, medium brown, verruculose,<br />
subcylindrical to narrowly obclavate, apex obtuse, base bluntly<br />
rounded with truncate hilum, straight, 0–3-septate, (7–)13–20(–25)<br />
× 3(–3.5) µm; hila thickened, darkened, refractive, 1–1.5 µm wide.<br />
Description based on <strong>CBS</strong> 105.75 (Fig. 8): Mycelium consisting<br />
of branched, septate, verruculose, medium brown, 2–4 µm wide<br />
hyphae. Conidiomata brown, superficial, sporodochial, up to 200 µm<br />
diam Conidiophores solitary, erect, micro- to macronematous, 1–12-<br />
septate, subcylindrical, straight to geniculate-sinuous or irregularly<br />
curved, 10–70 × 3–4 µm; frequently swollen <strong>and</strong> constricted at<br />
septa, thick-walled, medium brown, verruculose. Conidiogenous<br />
cells terminal <strong>and</strong> intercalary, subcylindrical, straight, but frequently<br />
branched laterally, 6–20 × 3–4 µm, with 1–3 flat-tipped loci that<br />
can be subdenticulate, 1.5–2 µm wide, somewhat darkened <strong>and</strong><br />
thickened, not prominently refractive. Conidia medium brown, thickwalled,<br />
verruculose, septa becoming darkly pigmented, occurring in<br />
branched chains. Ramoconidia subcylindrical to narrowly ellipsoid,<br />
12–25 × 3.5–4(–5) µm, 1(–4)-septate. Conidia occurring in short<br />
chains (–8), subcylindrical to narrowly ellipsoid, 0–1(–3)-septate,<br />
(7–)10–15(–20) × (2–)3–3.5(–4) µm; hila somewhat thickened,<br />
darkened but not refractive, 1.5–2(–2.5) µm wide.<br />
Cultural characteristics: Colonies on OA erumpent, spreading, with<br />
moderate aerial mycelium <strong>and</strong> smooth, even margins; olivaceousgrey<br />
(surface); on PDA olivaceous-black (surface), margins<br />
feathery, uneven, with moderate aerial mycelium; reverse iron-grey.<br />
Colonies reaching 20 mm diam after 1 mo at 25°C in the dark on<br />
OA.<br />
Specimens examined: Venezuela, Aragua, La Victoria, on leaf spots of<br />
Pithecellobium lanceolatum (Mimosaceae), Jan. 1928, H. Sydow, lectotype of S.<br />
araguata (selected here!) IMI 15728(a); 3 Feb. 1928, syntype of S. araguata, IMI<br />
34905. Venezuela, isolated from man with tinea nigra, 1973, D. Borelli, holotype of<br />
C. castellanii, IMI 183818, culture ex-type <strong>CBS</strong> 105.75.<br />
Notes: Stenella araguata is a leaf spot pathogen of Pithecellobium<br />
in Venezuela, <strong>and</strong> represents the type species of the <strong>genus</strong> Stenella<br />
[Two collections were cited, viz. no. 407, ‘La Victoria’, <strong>and</strong> no. 370,<br />
‘inter La Victoria et Suata’, both without any date, <strong>and</strong> without<br />
any specific type indication. Thus, the two collections have to be<br />
considered syntypes. <strong>The</strong> two IMI collections with different dates<br />
are parts of the syntypes, of which IMI 15728(a) is proposed here<br />
to serve as lectotype]. Stenella araguata was incorrectly seen as<br />
a species of <strong>Cladosporium</strong> by von Arx (1974), which has recently<br />
been morphologically circumscribed (Braun et al. 2003, Schubert et<br />
al. 2007b – this volume), <strong>and</strong> is linked to Davidiella teleomorphs.<br />
In a study by McGinnis & Padhye (1978), <strong>Cladosporium</strong><br />
castellanii (tinea nigra of human in Venezuela) was shown to be<br />
synonymous to Stenella araguata (leaf spots of Pithecellobium<br />
lanceolatum in Venezuela). In the present study we re-examined<br />
the ex-type strain of C. castellanii (<strong>CBS</strong> 105.75), <strong>and</strong> found conidia<br />
to be 0–1(–3)-septate, (7–)10–15(–20) × (2–)3–3.5(–4) µm, while<br />
those of the type specimen of S. araguata were <strong>similar</strong>, namely<br />
0–3-septate, (7–)13–20(–25) × 3(–3.5) µm. Furthermore, both<br />
collections have verruculose hyphae, which is the primary feature<br />
distinguishing Stenella from Passalora Fr. (Crous & Braun 2003).<br />
44
<strong>Cladosporium</strong> <strong>and</strong> morphologically <strong>similar</strong> genera<br />
Fig. 8. Stenella araguata (<strong>CBS</strong> 105.75). A–B. Conidiophore fascicles on a pine needle <strong>and</strong> tap-water agar, respectively. C–D, G. Conidiophores giving rise to conidial chains.<br />
E–F, H–J. Conidial chains with ramoconidia <strong>and</strong> conidia. Scale bars = 10 µm.<br />
Stenella has always been used for anamorphs of Mycosphaerella<br />
(Crous et al. 2004, 2006c), <strong>and</strong> the fact that it belongs to<br />
Teratosphaeria (Teratosphaeriaceae), <strong>and</strong> not Mycosphaerella<br />
(Mycosphaerellaceae), raised the question of how to treat stenellalike<br />
anamorphs in Mycosphaerella. Due to insufficient availability of<br />
cultures (Crous et al. 2000, 2001), the status of Stenella was left<br />
unresolved (Crous & Braun 2003). Presently (Crous & Groenewald,<br />
unpubl. data), it is clear that the stenella-like morphology type is<br />
polyphyletic within the Mycosphaerellaceae, <strong>and</strong> paraphyletic<br />
within the Capnodiales. Several species are known that represent<br />
morphological transitions between Stenella <strong>and</strong> Passalora. It<br />
seems logical, therefore, that future studies should favour using<br />
Passalora to also accommodate Mycosphaerella anamorphs with<br />
superficial, verruculose hyphae, which have traditionally been<br />
placed in Stenella. This is in spite of the fact that there are other<br />
generic names available within the Mycosphaerellaceae for taxa<br />
with a stenella-like morphology (pigmented structures, darkened,<br />
thickened, refractive scars, <strong>and</strong> superficial, verruculose mycelium),<br />
namely Zasmidium Fr. (1849) (see Arzanlou et al. 2007 – this<br />
volume), <strong>and</strong> Verrucisporota D.E. Shaw & Alcorn (1993). Based<br />
on the phylogenetic position of the type species, Stenella s. str.<br />
is an anamorph of Teratosphaeria (Teratosphaeriaceae). Using<br />
the generic concept as employed in this volume of the Studies in<br />
Mycology, however, the anamorph <strong>genus</strong> is accepted as being<br />
poly- <strong>and</strong> paraphyletic within the order Capnodiales.<br />
www.studiesinmycology.org<br />
45
Crous et al.<br />
Helotiales, incertae sedis<br />
Hyalodendriella Crous, gen. nov. MycoBank MB504435.<br />
Etymology: Morphologically <strong>similar</strong> to Hyalodendron Diddens.<br />
Differt a Hyalodendro et Retroconi conidiophoris dimorphis, cicatricibus incrassatis<br />
et conidiis ultimo brunneis.<br />
Morphologically <strong>similar</strong> to Hyalodendron <strong>and</strong> Retroconis, but<br />
distinct in that it has dimorphic conidiophores, conidia that turn<br />
brown with age, <strong>and</strong> have thickened scars. Microconidiophores<br />
forming as lateral branches on hyphae, subcylindrical, subhyaline<br />
to pale brown, smooth, septate, with terminal conidiogenous cells.<br />
Macroconidiophores septate, subcylindrical, straight to curved,<br />
subhyaline to pale brown, smooth, with an apical rachis that is pale<br />
brown, smooth, subcylindrical, with numerous, aggregated loci.<br />
Conidia limoniform to ellipsoid, aseptate, smooth, pale brown, in<br />
short chains, tapering towards ends that are prominently apiculate,<br />
prominently thickened <strong>and</strong> darkened, but not refractive.<br />
Type species: Hyalodendriella betulae Crous, sp. nov.<br />
Hyalodendriella betulae Crous sp. nov. MycoBank MB504436.<br />
Fig. 9.<br />
Mycelium ex hyphis ramosis, septatis, 1.5–2 µm latis, levibus, hyalinis vel pallide<br />
brunnei compositum. Conidiophora dimorphosa: (A) Conidiophora ex hyphis<br />
lateraliter oriunda, subcylindrical, subhyalina vel pallide brunnea, levia, 1–6-<br />
septata, ad 40 µm longa et 2–3 µm lata. Cellulae conidiogenae terminales, 5–15<br />
× 2–3 µm, loco conidiogeno singulare et terminale, cellula ellipsoidea (conidio ?),<br />
persistente, interdum cellulis catenulatis (ad 6), pallide brunneo, apice subacute<br />
rotundato, basi truncata, 5–7 × 3–4 µm. (B) Conidiophoris 10–20 × 2–3 µm,<br />
1–2-septatis, subcylindraceis, rectis vel curvatis, subhyalinis vel pallide brunneis,<br />
levibus. Cellulae conidiogenae pallide brunneae, leviae, subcylindraceae, locis<br />
numerosis, aggregatis, inconspicuis vel subdenticulatis, leviter protuberantes, 0.5<br />
µm diam, incrassatis et fuscatis. Conidia catenulata (2–3), (4–)5–6(–7) × 2.5–3 µm,<br />
limoniformes vel ellipsoidea, non septata, levia, pallide brunnea, utrinque attenuata,<br />
apiculata, 0.5–1 × 0.5 µm, incrassata et fuscata, non refractiva.<br />
Mycelium consisting of branched, septate, 1.5–2 µm wide<br />
hyphae, smooth, hyaline to pale brown. Conidiophores dimorphic.<br />
Type A: Conidiophores forming as lateral branches on hyphae,<br />
subcylindrical, subhyaline to pale brown, smooth, 1–6-septate, up<br />
to 40 µm long, <strong>and</strong> 2–3 µm wide. Conidiogenous cells terminal, 5–<br />
15 × 2–3 µm, with a single, apical locus, giving rise to an ellipsoidal<br />
cell (conidium?) which mostly remains attached, pale brown, with<br />
a subacutely rounded apex <strong>and</strong> truncate base, 5–7 × 3–4 µm, at<br />
times forming chains of up to 6 such cells. Type B: Conidiophores<br />
10–20 × 2–3 µm, 1–2-septate, subcylindrical, straight to curved,<br />
subhyaline to pale brown, smooth. Conidiogenous cells pale<br />
brown, smooth, subcylindrical with numerous, aggregated loci,<br />
inconspicuous to subdenticulate <strong>and</strong> somewhat protruding, 0.5 µm<br />
wide, somewhat thickened <strong>and</strong> darkened. Conidia in chains of 2–3,<br />
limoniform to ellipsoid, widest in the middle, aseptate, smooth, pale<br />
brown, tapering towards ends that are prominently apiculate, 0.5–1<br />
µm long, 0.5 µm wide, prominently thickened <strong>and</strong> darkened, but<br />
not refractive.<br />
Cultural characteristics: Colonies on PDA slimy, spreading,<br />
somewhat erumpent in the centre, with even, catenulate margins,<br />
lacking aerial mycelium; surface fuscous-black to olivaceous-black,<br />
with patches of cream; reverse fuscous-black with patches of<br />
cream. Colonies reaching 25 mm diam on PDA after 1 mo at 25 °C<br />
in the dark; colonies fertile with profuse sporulation on SNA.<br />
Specimen examined: Netherl<strong>and</strong>s, Oostelijk Flevol<strong>and</strong>, Jagersveld, isolated from<br />
Alnus glutinosa (Betulaceae), May 1982, W. Gams, holotype <strong>CBS</strong>-H 19895, culture<br />
ex-type <strong>CBS</strong> 261.82.<br />
Notes: Morphologically Hyalodendriella resembles the genera<br />
Hyalodendron <strong>and</strong> Retroconis de Hoog & Bat. Vegte (de Hoog<br />
& Batenburg van der Vegte 1989). It is distinct, however, in its<br />
pigmentation, dimorphic conidiophores <strong>and</strong> conidia. Furthermore,<br />
a strain of Retroconis fusiformis (S.M. Reddy & Bilgrami) de Hoog<br />
& Bat. Vegte (<strong>CBS</strong> 330.81) clusters apart from Hyalodendriella,<br />
namely in the Chaetomiaceae, Sordariales.<br />
Pleosporales, incertae sedis<br />
Ochrocladosporium Crous & U. Braun, gen. nov. MycoBank<br />
MB504437.<br />
Etymology: Named after its pale brown, cladosporium-like conidia.<br />
Differt a Cladosporio et generis cladosporioidibus diversis conidiophoris cum cellulis<br />
basalibus T-formibus et/vel cicatricibus non incrassatis, non vel leviter fuscatisrefractivis.<br />
Mycelium consisting of branched, septate hyphae, subhyaline to<br />
pale brown, smooth, giving rise to two types of conidiophores.<br />
Macronematous conidiophores solitary, erect, arising from<br />
superficial hyphae, composed of a subcylindrical stipe, without a<br />
swollen or lobed base or rhizoids, with or without a T-shaped foot cell,<br />
pale to dark brown; apical conidiogenous apparatus with or without<br />
additional branches, branched part, if present, with short branchlets<br />
composed of conidiogenous cells <strong>and</strong> ramoconidia, continuous to<br />
septate, wall thin or slightly thicked, pale brown. Conidiogenous cells<br />
integrated, terminal or intercalary, subcylindrical to doliiform, pale<br />
brown, thin-walled, smooth; unilocal or multilocal, determinate to<br />
sympodial, loci conically truncate, subdenticulate, neither thickened,<br />
nor darkened-refractive or only slightly darkened-refractive.<br />
Micronematous conidiophores integrated in hyphae, reduced to a<br />
lateral peg-like locus or erect, frequently reduced to conidiogenous<br />
cells, pale brown, smooth, subcylindrical. Conidia occurring in<br />
branched chains, fusiform, ellipsoid-ovoid to subcylindrical, 0(–1)-<br />
septate, ramoconidia present, pale brown, thin-walled, smooth to<br />
finely verruculose, ends attenuated, hila obconically truncate to<br />
almost pointed, neither thickened nor darkened-refractive.<br />
Type species: Ochrocladosporium elatum (Harz) Crous & U. Braun,<br />
comb. nov.<br />
Ochrocladosporium elatum (Harz) Crous & U. Braun, comb.<br />
nov. MycoBank MB504438. Fig. 10.<br />
Basionym: Hormodendrum elatum Harz, Bull. Soc. Imp. Naturalistes<br />
Moscou 44: 140. 1871.<br />
≡ <strong>Cladosporium</strong> elatum (Harz) Nannf., in Melin & Nannfeldt, Svenska<br />
Skogsvardsfoereren Tidskr. 32: 397. 1934.<br />
≡ Cadophora elatum (Harz) Nannf., in Melin & Nannfeldt, Svenska<br />
Skogsvardsfoereren Tidskr. 32: 422. 1934.<br />
Mycelium consisting of branched, septate, smooth, hyaline, 2–4<br />
µm wide, thin-walled, hyphae, becoming darker brown in places,<br />
giving rise to erect conidiophores. Conidiophores either reduced<br />
to conidiogenous cells, or well-differentiated, terminal <strong>and</strong> lateral<br />
on hyphae, erect, highly variable, arising from superficial <strong>and</strong><br />
submerged hyphae, reduced to subdenticulate loci, 1–1.5 µm<br />
wide, or well-differentiated, up to 60 µm long, 1–3-septate, 3–4<br />
µm wide, hyaline to medium brown, smooth, thin-walled (≤ 1 µm).<br />
Conidiogenous cells integrated as lateral peg-like loci on hyphal<br />
cells, or erect, subcylindrical, up to 25 µm long, 2.5–4 µm wide,<br />
with 1–3 terminal loci, occasionally also lateral, 1–1.5 µm wide,<br />
not thickened <strong>and</strong> darkened, but frequently somewhat refractive<br />
(mounted in Shear’s solution, not lactic acid). Ramoconidia<br />
46
<strong>Cladosporium</strong> <strong>and</strong> morphologically <strong>similar</strong> genera<br />
Fig. 9. Hyalodendriella betulae (type material). A. Conidiophores on PDA. B–C. Microconidiophores. D–H. Macroconidiophores with fascicles of conidiogenous cells. I. Conidia<br />
with darkened, thickened hila. Scale bars = 10 µm.<br />
subcylindrical to ellipsoid, hyaline to pale brown, smooth to finely<br />
verruculose, 10–40 × 3–5 µm, 0(–1)-septate, giving rise to branched<br />
chains of conidia (up to 20 per chain) that are subcylindrical to<br />
ellipsoid, aseptate, (7–)8–10(–14) × (3–)4(–4.5) µm, smooth to<br />
finely verruculose, olivaceous-brown, thin-walled (up to 0.5 µm),<br />
hila 0.5–1 µm wide, neither thickened nor, or barely, darkened<br />
refractive.<br />
Cultural characteristics: Colonies erumpent, spreading, fast<br />
growing, covering the plate within 1 mo at 25 °C; aerial mycelium<br />
abundant, margins smooth on PDA; surface isabelline in centre,<br />
umber in outer region; olivaceous-black in reverse.<br />
Specimen examined: Sweden, Iggesund, isolated from wood pulp, Jan. 1976, E.<br />
Melin, specimen <strong>CBS</strong>-H 19896, culture <strong>CBS</strong> 146.33.<br />
Notes: “Hormodendrum” elatum was originally described from<br />
a wooden stump in Germany. <strong>The</strong> culture examined here was<br />
deposited by Melin in 1933 as culture 389:14, isolated from wood<br />
chips in Sweden, <strong>and</strong> described by Nannfeldt, <strong>and</strong> has since<br />
been accepted as authentic for the species. Earlier publications<br />
(de Vries 1952, Ho et al. 1999, de Hoog et al. 2000), clearly state<br />
that this species does not belong in <strong>Cladosporium</strong> s. str., <strong>and</strong> this<br />
statement is supported by the phylogenetic analysis placing it in<br />
the Pleosporales.<br />
www.studiesinmycology.org<br />
Ochrocladosporium frigidarii Crous & U. Braun, sp. nov.<br />
MycoBank MB504439. Fig. 11.<br />
Etymology: Named after it collection site, within a cooled incubation<br />
room.<br />
Differt a O. elato conidiophoris distincte dimorphis, macroconidiophoris majoribus,<br />
ad 600 × 5–7 µm, septis incrassates, cellulis basalibus T-formibus et conidiis leniter<br />
brevioribus et latioribus, (6–)7–8(–10) × (4–)4.5–5(–6) µm.<br />
Mycelium consisting of branched, septate, 2–7 µm wide hyphae,<br />
occasionally constricted at septa with hyphal swellings, subhyaline<br />
to pale brown, smooth, thin-walled, giving rise to two types of<br />
conidiophores. Macronematous conidiophores solitary, erect,<br />
arising from superficial hyphae, up to 600 µm long, composed of<br />
a subcylindrical stipe, 5–7 µm wide, 10–15(–20)-septate, without<br />
a swollen or lobed base or rhizoids, but with a T-shaped foot cell,<br />
wall ≤ 1 µm wide, guttulate, with thick septa, dark brown, finely<br />
verruculose, apical 1–2 cells at times medium brown, giving rise<br />
to 1–2 primary branches, 0–1-septate, subcylindrical, thin-walled,<br />
pale brown, smooth to finely verruculose, 10–20 × 4–6 µm, giving<br />
rise to (1–)2–4 secondary branches, 0–1-septate, subcylindrical,<br />
8–13(–20) × 4–5 µm, or giving rise directly to conidiogenous cells.<br />
Conidiogenous cells subcylindrical to doliiform, pale brown, smooth,<br />
8–15 × 3–4 µm, loci somewhat protruding 1–2 µm wide, neither<br />
47
Crous et al.<br />
Fig. 10. Ochrocladosporium elatum (<strong>CBS</strong> 146.33). A–C, E. Microconidiophores. D. Macro- <strong>and</strong> microconidiophore. F–H. Macroconidiophores. I–J. Ramoconidia <strong>and</strong> conidia.<br />
Scale bars = 10 µm.<br />
thickened, darkened, nor refractive. Micronematous conidiophores<br />
erect, pale brown, smooth, subcylindrical, reduced to conidiogenous<br />
cells, or up to 4-septate, 15–90 × 2–3.5 µm, mostly unbranched,<br />
rarely branched below; conidiogenous cells subcylindrical,<br />
pale brown, smooth to finely verruculose, tapering at apex <strong>and</strong><br />
sometimes at base, proliferating sympodially via 1(–3) loci, 1–1.5<br />
µm wide, denticle-like, which can appear somewhat darkened;<br />
micronematous conidiophores frequently occurring at the base of<br />
macronematous conidiophores. Ramoconidia, if present, up to 30<br />
µm long, 0–1-septate. Conidia <strong>and</strong> ramoconidia ellipsoid to ovoid,<br />
aseptate, pale brown, thin-walled (≤ 0.75 µm), finely verruculose,<br />
occurring in branched chains; conidia (6–)7–8(–10) × (4–)4.5–5(–6)<br />
µm; hila 0.5–1 µm wide, not darkened, thickened or refractive.<br />
Cultural characteristics: Colonies on PDA erumpent, spreading, with<br />
profuse sporulation <strong>and</strong> moderate aerial mycelium, even margins,<br />
olivaceous-grey (surface); reverse olivaceous-black. Colonies<br />
covering the dish after 1 mo at 25 °C in the dark.<br />
Specimen examined: Germany, Hannover, isolated from a cooled room, Jan. 1981,<br />
B. Ahlert, holotype <strong>CBS</strong>-H 19897, culture ex-type <strong>CBS</strong> 103.81.<br />
Notes: Ochrocladosporium frigidarii is characterised by its<br />
dimorphic fruiting, <strong>and</strong> inconspicuous scars <strong>and</strong> conidial hila, which<br />
48
<strong>Cladosporium</strong> <strong>and</strong> morphologically <strong>similar</strong> genera<br />
Fig. 11. Ochrocladosporium frigidarii (type material). A. Macro- <strong>and</strong> microconidiophores. B. Foot cell of macroconidiophore. C–D. Microconidiophore. E–J. Macroconidiophores.<br />
K. Conidia. Scale bars = 10 µm.<br />
are distinct from <strong>Cladosporium</strong> s. str. <strong>The</strong> phylogenetic analysis of<br />
its LSU sequence places it in the Pleosporales, together with O.<br />
elatum.<br />
<strong>The</strong> dimorphic conidiophores seen in O. frigidarii (<strong>CBS</strong> 103.81)<br />
are less obvious in O. elatum (<strong>CBS</strong> 146.33), but the scars <strong>and</strong> hila<br />
are <strong>similar</strong>. <strong>The</strong> macronematous conidiophores of O. frigidarii are<br />
much longer <strong>and</strong> wider <strong>and</strong> the conidia are shorter <strong>and</strong> slightly<br />
wider, (6–)7–8(–10) × (4–)4.5–5(–6) µm, than those of O. elatum<br />
which are (7–)8–10(–14) × (3–)4(–4.5) µm.<br />
www.studiesinmycology.org<br />
49
Crous et al.<br />
Fig. 12. Rhizocladosporium argillaceum (type material). A–E. Conidiophores with pigmented ramoconidia <strong>and</strong> hyaline conidia. F. Rhizoids forming at the foot cells of<br />
macroconidiophores. Scale bar = 10 µm.<br />
Incertae sedis<br />
Rhizocladosporium Crous & U. Braun, gen. nov. MycoBank<br />
MB504440.<br />
Etymology: Named after the presence of rhizoids on its<br />
conidiophores, <strong>and</strong> cladosporium-like conidia.<br />
Differt a Cladosporio et generis cladosporioidibus diversis hyphis hyalinis,<br />
conidiophoris cum cellulis basalibus lobatis vel rhizoidibus, cellulis conidiogenis<br />
monoblasticis, determinatis, locis margine leviter incrassatis et fuscatis, non<br />
refractivis, non coronatis, ramoconidiis brunneis sed conidiis hyalinis, hilis non<br />
incrassatis, non fuscatis-refractivis.<br />
Mycelium consisting of branched, septate, smooth, hyaline hyphae.<br />
Conidiophores solitary, macronematous, subcylindrical, erect,<br />
arising from superficial mycelium, septate, pigmented, smooth;<br />
base somewhat inflated, lobed or with rhizoids. Conidiogenous<br />
cells integrated, terminal, monoblastic, determinate, subcylindrical,<br />
tapering towards a single flat-tipped locus, straight to once<br />
geniculate, occasionally with two loci, pigmented, smooth; locus<br />
flattened, undifferentiated to somewhat darkened <strong>and</strong> thickened<br />
along the rim, not refractive, giving rise to a single conidial chain<br />
or a single ramoconidium with several simple acropetal chains of<br />
secondary ramoconidia or conidia. Conidia occurring in branched<br />
chains; ramoconidia subcylindrical to narrowly ellipsoidal, straight<br />
to geniculate-sinuous, with apical <strong>and</strong> lateral conidial hila;<br />
ramoconidia with broadly truncate base medium brown; secondary<br />
ramoconidia with narrowed base subhyaline or hyaline, smooth;<br />
conidia aseptate, in chains, hyaline, guttulate, ellipsoidal with<br />
obtuse ends; hila inconspicuous, neither darkened nor refractive<br />
or thickened.<br />
Type species: Rhizocladosporium argillaceum (Minoura) Crous &<br />
U. Braun, comb. nov.<br />
Rhizocladosporium argillaceum (Minoura) Crous & U. Braun,<br />
comb. nov. MycoBank MB504441. Fig. 12.<br />
Basionym: <strong>Cladosporium</strong> argillaceum Minoura, J. Ferment. Technol.<br />
44: 140. 1966.<br />
Mycelium consisting of branched, septate, smooth, hyaline,<br />
thin-walled, 1.5–2 µm wide hyphae. Conidiophores solitary,<br />
macronematous, erect, arising from superficial mycelium; base<br />
somewhat inflated, lobed or with rhizoids, up to 10 µm wide;<br />
conidiophore stipe subcylindrical, straight to curved, rarely<br />
geniculate-sinuous, wall up to 1 µm wide, medium brown,<br />
sometimes paler towards the tip, smooth, 1–6-septate, 35–160<br />
50
<strong>Cladosporium</strong> <strong>and</strong> morphologically <strong>similar</strong> genera<br />
µm tall, 4–6 µm wide. Conidiogenous cells terminal, straight,<br />
subcylindrical, tapering towards a flat-tipped locus, occasionally<br />
once geniculate, with two loci, medium brown, smooth, 15–35 ×<br />
4–6 µm; locus flattened, undifferentiated or very slightly darkened<br />
<strong>and</strong> thickened along the rim, not refractive, 1.5–2 µm wide. Conidia<br />
occurring in branched chains. Ramoconidia subcylindrical to<br />
narrowly ellipsoidal, straight to geniculate-sinuous, 17–35 × 4–5<br />
µm, medium brown, smooth, thin-walled, frequently branching<br />
laterally, with apical <strong>and</strong> lateral subdenticulate conidial hila, 1.5–2.5<br />
µm wide; secondary ramoconidia hyaline or subhyaline. Conidia<br />
aseptate, (10–)12–17(–20) × (3.5–)4(–4.5) µm, in branched<br />
chains (–6), hyaline or subhyaline, guttulate, ellipsoidal-fusiform,<br />
with obtuse ends, or tapering to obconically subtruncate ends with<br />
hila that are inconspicuous (neither darkened nor refractive or<br />
thickened), 0.5–1 µm wide.<br />
Cultural characteristics: Colonies on PDA spreading, erumpent, with<br />
smooth, even margins <strong>and</strong> sparse to moderate aerial mycelium;<br />
hazel to fawn (surface); reverse hazel to fawn. Colonies reaching<br />
35 mm diam after 1 mo at 25 °C in the dark; colonies fertile.<br />
Specimen examined: Japan, Yoku Isl<strong>and</strong>, isolated from decayed myxomycete, 21<br />
Oct. 1961, K. Tubaki No. 4262 holotype, culture ex-type <strong>CBS</strong> 241.67 = IFO 7055.<br />
Notes: <strong>The</strong> lobed-rhizoid conidiophore base, <strong>and</strong> brown,<br />
disarticulating ramoconidia, with hyaline chains of conidia, are<br />
characteristic of Rhizocladosporium. Although Minoura (1966)<br />
illustrated some conidiophores that were micronematous (reduced<br />
to conidiogenous cells on superficial mycelium), these were<br />
not observed in the present study. Metulocladosporiella Crous,<br />
Schroers, J.Z. Groenew., U. Braun & K. Schub. (Crous et al.<br />
2006a) (Herpotrichiellaceae), comprising two banana leaf-spotting<br />
pathogens, is another cladosporioid hyphomycete <strong>genus</strong> having<br />
distinct rhizoid hyphae at the swollen base of conidiophores. It<br />
differs, however, in having conidiophores terminally branched in<br />
a metula-like manner <strong>and</strong> distinct conidiogenous loci <strong>and</strong> conidial<br />
hila. Pleurotheciopsis B. Sutton (Ellis 1976) is also characterised<br />
by having pigmented conidiophores <strong>and</strong> hyaline or pale, septate<br />
conidia formed in acropetal chains, but the conidiophores<br />
proliferate percurrently, the conidiogenous cells are polyblastic <strong>and</strong><br />
ramoconidia are lacking, i.e., the conidia are formed in unbranched<br />
chains. Parapleurotheciopsis P.M. Kirk (Kirk 1982) is very <strong>similar</strong> to<br />
Rhizocladosporium. <strong>The</strong> conidiophores possess a single terminal<br />
unilocal conidiogenous cell giving rise to a single ramoconidium<br />
which forms several chains of acropetal, aseptate, hyaline to pale<br />
olivaceous conidia. <strong>The</strong> base of the conidiophores is somewhat<br />
swollen <strong>and</strong> lobed [except for Parapleurotheciopsis coccolobae<br />
R.F. Castañeda & B. Kendr., Castañeda & Kendrick (1990), with at<br />
most slightly swollen, but unlobed base]. However, R. argillaceum<br />
occasionally has once-geniculate conidiogenous cells with two<br />
loci. Furthermore, it clusters in the Helotiales (Fig. 1), whereas a<br />
sequenced strain of Parapleurotheciopsis inaequiseptata (MUCL<br />
41089), belongs to the Xylariales (Fig. 2). <strong>The</strong> occasionally<br />
occurring conidiogenous cells with two loci <strong>and</strong> the aseptate conidia<br />
connect Rhizocladosporium with Subramaniomyces Varghese &<br />
V.G. Rao (Varghese & Rao 1979, Kirk 1982) in which, however,<br />
terminal ramoconidia are lacking. Furthermore, the type species,<br />
S. fusisaprophyticus (Matsush.) P.M. Kirk, frequently has branched<br />
conidiophores. Subramaniomyces simplex U. Braun & C.F. Hill<br />
(Braun & Hill 2002), a species with unbranched conidiophores is,<br />
however, morphologically <strong>similar</strong> to R. argillaceum, but the <strong>genus</strong><br />
Subramaniomyces is phylogenetically distinct <strong>and</strong> also belongs to<br />
the Xylariales (<strong>CBS</strong> 418.95, Fig. 2).<br />
Key to <strong>Cladosporium</strong> <strong>and</strong> morphologically <strong>similar</strong> genera<br />
(bearing simple or branched acropetal chains of amero- to phragmosporous blastoconidia)<br />
1. Conidiophores <strong>and</strong> conidia hyaline............................................................................................................................................................. 2<br />
1. At least conidiophores pigmented .............................................................................................................................................................. 4<br />
2. Conidia in simple chains ........................................................................................................................................................... Hormiactis<br />
2. Conidia in branched chains ....................................................................................................................................................................... 3<br />
3. Conidiogenous cells sympodial, with distinct conidiogenous loci (scars), thickened <strong>and</strong> darkened; conidia amero- to phragmosporous;<br />
plant pathogenic, leaf-spotting fungi (Mycosphaerella anamorphs; Mycosphaerellaceae) ......................................................... Ramularia<br />
3. Terminal conidiogenous cells with denticle-like loci, giving rise to ramoconidia which form simple or branched conidial chains;<br />
lignicolous ........................................................................................................................................................................... Hyalodendron<br />
[Conidiophores dimorphic; mycelium, conidiophores <strong>and</strong> conidia at first hyaline, later turning pale brown; conidia in short chains, see<br />
Hyalodendriella]<br />
4(1). Conidia distoseptate, in simple chains .............................................................................................................................................. Lylea<br />
4. Conidia aseptate or euseptate ................................................................................................................................................................... 5<br />
5. Conidiophores little differentiated, micronematous to semimacronematous; conidiogenous loci undifferentiated, truncate, neither<br />
distinctly thickened nor darkened or only very slightly so .......................................................................................................................... 6<br />
5. Conidiophores well-differentiated, semimacronematous (but multilocal <strong>and</strong>/or conidiogenous loci well-differentiated) to<br />
macronematous ....................................................................................................................................................................................... 12<br />
6. Conidiophores <strong>and</strong> conidia delicate, thin-walled, in long, easily disarticulating chains .............................................................................. 7<br />
6. Conidiophores <strong>and</strong> conidia robust, wall thickened, dark, conidial chains often seceding with difficulty .................................................... 9<br />
7. Conidiophores semimacronematous, simple to often irregularly branched; conidia delicate, narrow, 1–3 µm wide, hyaline to pale<br />
olivaceous ............................................................................................................................................................................ Polyscytalum<br />
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Crous et al.<br />
7. Conidiophores unbranched, micronematous or semimicronematous, integrated in ordinary hyphae, forming minute, lateral,<br />
monoblastic, determinate, peg-like protuberances to semimacronematous, forming short lateral branches (conidiophores) with several<br />
inconspicuous to denticle-like loci .............................................................................................................................................................. 8<br />
8. Phialidic synanamorphs often present, but sometimes also lacking; saprobic, rarely plant pathogenic, often human pathogenic<br />
(Herpotrichiellaceae, Chaetothyriales) .......................................................................................................................... Cladophialophora<br />
8. Without phialidic synanamorphs; saprobic or plant pathogenic (Venturia, Venturiaceae)..............................................................................<br />
.......................................................................................................................................... Fusicladium s. lat. (incl. Pseudocladosporium)<br />
9(6). Conidia aseptate, rarely 1-septate; lignicolous, on dead wood ............................................................................................... Xylohypha<br />
9. Conidia septate ........................................................................................................................................................................................ 10<br />
10. Conidia 1-septate, with a dark brown to blackish b<strong>and</strong> at the septum; on dead wood .................................................................. Bispora<br />
10. Conidia at least partly 2- to pluriseptate <strong>and</strong>/or without dark brown to blackish b<strong>and</strong> at the septum ...................................................... 11<br />
11. Conidia branched ....................................................................................................................................................................... Taeniolina<br />
11. Conidia unbranched ................................................................................................................................................................... Taeniolella<br />
12(5). Conidiogenous loci <strong>and</strong> conidial hila distinctly coronate, i.e., composed of a central convex dome surrounded by a periclinal raised rim,<br />
mostly at least somewhat protuberant (anamorphs of Davidiella, Davidiellaceae, Capnodiales) ............................. <strong>Cladosporium</strong> s. str.<br />
12. Conidiogenous loci non-coronate (either inconspicuous, thickened <strong>and</strong> darkened or denticle-like) ........................................................ 13<br />
13. Mycelium, conidiophores <strong>and</strong> conidia at first hyaline or subhyaline, later turning pale brown; conidiophores dimorphic, either<br />
conidiogenous cells with a single conidiogenous locus, giving rise to an ellipsoid cell (conidium?) which mostly remains attached,<br />
base truncate, apex subacutely rounded, at times forming chains of such cells; or conidiophores with numerous aggregated loci,<br />
inconspicuous to subdenticulate; conidia in short chains, of mostly 2–3 (isolated from Alnus in Europe) ........................ Hyalodendriella<br />
13. Fruiting different; at least conidiophores consistently pigmented or conidiophores uniform or loci distinct; conidia mostly in long, often<br />
branched chains ...................................................................................................................................................................................... 14<br />
14. Conidiophores with verruculose conidiogenous apices, otherwise smooth; conidia distinctly verruculose-verrucose; conidiogenous loci<br />
<strong>and</strong> conidial hila inconspicuous ............................................................................................................................................................... 15<br />
14. Conidiophores either smooth throughout or verruculose below <strong>and</strong> smooth above or verruculose throughout; <strong>and</strong>/or conidiogenous loci<br />
conspicuous, i.e., thickened <strong>and</strong> darkened or denticle-like ...................................................................................................................... 16<br />
15. Conidiophores macronematous, unbranched, base swollen, with percurrent regenerative proliferations, unrelated to conidiation;<br />
conidiogenous cells terminal, occasionally also subterminal; conidia terminally <strong>and</strong> laterally formed, aseptate (saprobic on leaves)<br />
................................................................................................................................................................................................ Castanedaea<br />
15. Conidiophores little differentiated, semimacronematous, unbranched or with short lateral branchlets, base undifferentiated,<br />
without percurrent proliferations; conidiogenous cells terminal <strong>and</strong> occasionally intercalary-pleurogenous; conidia terminally <strong>and</strong><br />
subterminally formed, 0–2-septate (lignicolous, on decorticated wood) .......................................................................... Websteromyces<br />
16(14). Conidiophores unbranched, with a simple terminal conidiogenous cell, non-geniculate-sinuous, subcylindrical to somewhat inflated at<br />
the tip; conidiogenous loci terminal <strong>and</strong> lateral, inconspicuous or subconspicuous, neither thickened nor darkened, non-protuberant;<br />
conidia attached with a very narrow, pointed hilum ................................................................................................................................. 17<br />
16. Conidiophores with a branched terminal conidiogenous apparatus, composed of conidiogenous cells <strong>and</strong>/or ramoconidia or conidiophores<br />
unbranched, with a single terminal conidiogenous cell or additional intercalary ones, but conidiogenous loci different, conspicuous,<br />
thickened <strong>and</strong> darkened or denticle-like .................................................................................................................................................. 18<br />
17. Conidiophores with distinct rhizoid-digitate base; tips of the conidiogenous cells somewhat swollen, usually unilaterally swollen or<br />
somewhat curved; conidia solitary or only in very short unbranched chains; hyperparasitic on rusts .................................. Digitopodium<br />
17. Conidiophores without rhizoid-digitate base; tips of the conidiogenous cells subcylindrical to somewhat swollen, but swellings not unilateral<br />
<strong>and</strong> not curved; conidia solitary <strong>and</strong> in simple or branched chains; associated with leaf spots ................................. Rachicladosporium<br />
18(16). Conidiophores in synnematous conidiomata ..................................................................................................................................... 19<br />
18. Synnemata lacking .................................................................................................................................................................................. 20<br />
19. Conidiogenous cells with a single or several truncate to subdenticulate, relatively broad conidiogenous loci; conidia with truncate, flat hila;<br />
on wood, resin ............................................................................................................................................................................ Sorocybe<br />
19. Conidiogenous loci with few, mostly 1–2 conidiogenous loci formed as minute spicules; conidia with narrow hila (shallowly apiculate);<br />
plant pathogenic, causing bud blast <strong>and</strong> twig blight ....................................................................................................................... Seifertia<br />
20(18). Conidiophores unbranched or occasionally branched; conidiogenous cells distinctly inflated, ampulliform, doliiform or clavate, nondenticulate;<br />
conidia at least partly globose, dark brown when mature; colonies effuse, dark; wood-inhabiting .......... Phaeoblastophora<br />
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<strong>Cladosporium</strong> <strong>and</strong> morphologically <strong>similar</strong> genera<br />
20. Conidiogenous cells not inflated, if somewhat inflated, loci denticle-like or conidia non-globose ............................................................ 21<br />
21. Conidiophores penicillate, i.e., with an unbranched stipe <strong>and</strong> distinct terminal branched “head” composed of branchlets, conidiogenous<br />
cells <strong>and</strong>/or ramoconidia .......................................................................................................................................................................... 22<br />
21. Conidiophores non-penicillate, i.e., irregularly <strong>and</strong> loosely branched, branchings not confined to the apical portion, sometimes only with<br />
short lateral branchlets, or unbranched ................................................................................................................................................... 27<br />
22. Penicillate apex simple, only composed of a single terminal conidiogenous cells giving rise to several ramoconidia which form secondary<br />
ramoconidia <strong>and</strong> conidia ......................................................................................................................... Penidiella p.p. [P. strumelloidea]<br />
22. Penicillate apex more complex, composed of true branchlets <strong>and</strong>/or conidiogenous cells <strong>and</strong> ramoconidia .......................................... 23<br />
23. Conidiophores with a compact, dense, subglobose to broadly ovoid head; conidiogenous loci <strong>and</strong> conidial hila unthickened or almost so,<br />
but distinct by being darkened-refractive [fruiting dimorphic, periconioid branched conidiophores formed on overwintered<br />
stem of Paeonia spp., unbranched cladosporioid conidiophores on leaf spots, biotrophic] (belonging to the Capnodiales)<br />
.................................................................................................................................................................................... Dichocladosporium<br />
23. Penicillate apex looser, neither compact nor subglobose ........................................................................................................................ 24<br />
24. Branched head composed of short branchlets <strong>and</strong> conidiogenous cells; ramoconidia lacking; conidiogenous cells subcylindrical to<br />
subclavate, non-geniculate; conidiogenous loci usually numerous <strong>and</strong> aggregated, terminal <strong>and</strong> lateral, non-protuberant, flat, conspicuous,<br />
thickened <strong>and</strong> darkened, at least around the rim; conidia solitary or in short chains ............................................................... Periconiella<br />
24. Ramoconidia often present; conidiogenous cells distinct, sympodial, somewhat geniculate or subdenticulate; conidiogenous loci<br />
inconspicuous or somewhat protruding, denticle-like, unthickened or almost so, not or somewhat darkened-refractive; conidia in long,<br />
often branched chains ............................................................................................................................................................................. 25<br />
25. Branched apex composed of short branchlets consisting of conidiogenous cells or ramoconidia, in pairs or whorls of 3–4, mostly distinctly<br />
constricted at the base; hyperparasitic on Asterina spp. ................................................................................................. Parapericoniella<br />
25. Branched apex distinct, composed of branchlets, conidiogenous cells <strong>and</strong>/or ramoconidia, if true branchlets lacking conidiogenous cells<br />
<strong>and</strong> ramoconidia not in whorls <strong>and</strong> not distinctly constricted at the base; saprobic or biotrophic ............................................................ 26<br />
26. Penicillate apex of the conidiophores loosely to densely branched, occasionally metula-like, base of the conidiophores simple,<br />
undifferentiated; saprobic or biotrophic (Teratosphaeriaceae, Capnodiales) .............................................................................. Penidiella<br />
26. Penicillate apex always dense, metula-like, base of the conidiophores swollen or lobed, often with rhizoid hyphae; plant pathogenic [on<br />
banana] (Chaetothyriales) ........................................................................................................................................ Metulocladosporiella<br />
27(21). Conidiophores simple or branched; septa of the conidiophores <strong>and</strong> conidia becoming thick-walled <strong>and</strong> dark; conidiogenous loci<br />
subdenticulate, somewhat thickened <strong>and</strong> conspicuously darkened-refractive; cultures producing ample amounts of volatile metabolites<br />
causing skin irritation after exposure to the fungus; saprobic (isolated from mouldy paint) ...................................... Toxicocladosporium<br />
27. Without conspicuously thickened-darkened septa; cultures without irritant, volatile metabolites ............................................................ 28<br />
28. Conidiogenous loci conspicuous, distinctly thickened <strong>and</strong> darkened (visible as small dark circles when viewed upon the scar), sometimes<br />
on small shoulders formed by sympodial proliferation, but not distinctly denticulate (Capnodiales) ....................................................... 29<br />
28. Conidiogenous loci inconspicuous or conspicuous by being denticle-like, not or barely thickened, not darkened or at most upper truncate<br />
end very slightly thickened <strong>and</strong> somewhat darkened-refractive .............................................................................................................. 31<br />
29. Mycelium smooth; conidiophores <strong>and</strong> conidia smooth or almost so, at most faintly rough-walled; conidiophores solitary, fasciculate,<br />
sporodochial to synnematous; biotrophic, usually leaf-spotting (Mycosphaerella anamorphs, Mycosphaerellaceae)<br />
................................................................................................................. Passalora emend. (incl. Mycovellosiella, Phaeoramularia, etc.)<br />
29. At least mycelium distinctly verruculose .................................................................................................................................................. 30<br />
30. Mycelium, conidiophores <strong>and</strong> conidia coarsely verruculose-verrucose; conidial shape variable, often irregular; isolated from a lichen<br />
(Dirina) .................................................................................................................................................................... Verrucocladosporium<br />
30. Mycelium verruculose; conidiophores mostly smooth, sometimes somewhat rough-walled, conidia smooth to distinctly verruculose;<br />
biotrophic, often leaf-spotting ......................................................................................................................................................... Stenella<br />
31(28). Conidiophores with swollen, often lobed base ................................................................................................................................... 32<br />
31. Conidiophores without swollen base, at most slightly swollen, but not lobed .......................................................................................... 35<br />
32. Conidia septate ........................................................................................................................................................................................ 33<br />
32. Conidia aseptate ...................................................................................................................................................................................... 34<br />
33. Conidiophores with a single, terminal, monoblastic, determinate conidiogenous cell giving rise to a single ramoconidium that forms<br />
simple or branched chains of conidia ...................................................................................................................... Parapleurotheciopsis<br />
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Crous et al.<br />
33. Terminal conidiogenous cells polyblastic, with several denticle-like conidiogenous loci ................. Anungitea p.p. (e.g. A. longicatenata)<br />
34(32). Conidiogenous cells terminal, monoblastic, with a single ramoconidium giving rise to conidial chains or occasionally with 2(–3)<br />
denticle-like loci; base of the conidiophores often with rhizoid hyphae ...................................................................... Rhizocladosporium<br />
34. Conidiogenous cells polyblastic, with two or several denticle-like loci; base of the conidiophores without rhizoid hyphae<br />
...................................................................................................................................................................................... Subramaniomyces<br />
35(31). Conidiophores unbranched, with a terminal monoblastic conidiogenous cell, determinate or percurrent ......................................... 36<br />
35. Conidiophores branched or unbranched, but conidiogenous cells at least partly polyblastic .................................................................. 38<br />
36. Conidiogenous cell giving rise to a single ramoconidium which forms simple or branched chains of 0(–1)-septate conidia<br />
................................................................................................................................................ Parapleurotheciopsis p.p. (P. coccolobae)<br />
36. Conidiogenous cells giving rise to simple conidial chains without ramoconidia; conidia septate ............................................................ 37<br />
37. Conidiophores sometimes with percurrent proliferations; conidiophores <strong>and</strong> conidia with somewhat thickened, dark walls; conidia 1–10-<br />
septate, width usually exceeding 4 µm ................................................................................................................................ Heteroconium<br />
37. Percurrent proliferations lacking; conidiophores <strong>and</strong> conidia delicate, thin-walled <strong>and</strong> paler; conidia usually 0–1(–3)-septate <strong>and</strong> narrow,<br />
usually below 4 µm wide (Chaetothyriales) ......................................................................... Cladophialophora p.p. (e.g. C. chaetospira)<br />
[<strong>similar</strong> anamorphs of the Venturiaceae, see Fusicladium (incl. Pseudocladosporium)]<br />
38(35). Conidiophores often branched; conidiogenous loci distinctly denticle-like or subdenticulate; conidia aseptate; lignicolous, on dead<br />
wood, resin or isolated from hydrocarbone-rich substrates (jet-fuel, cosmetics, etc.) ............................................................................. 39<br />
38. Either with unbranched conidiophores or conidiogenous loci not distinctly denticle-like, or conidia septate, or on other substrates<br />
.................................................................................................................................................................................................................. 42<br />
39. Conidiogenous cells distinctly denticulate; conidia rather broad, approx. 7–13 µm ............................................................. Haplotrichum<br />
39. Conidiogenous cells non-denticulate or at most subdenticulate; conidia narrower, approx. 3–6 µm ...................................................... 40<br />
40. Colonies effuse, dense, but felted, black, brittle <strong>and</strong> appearing carbonaceous when dry; conidiophores solitary, brown; conidiogenous cells<br />
terminal <strong>and</strong> pleurogenous; conidia pale to dark brown, lateral walls conspicuously thicker than the hila; on conifer resin<br />
......................................................................................................................... Sorocybe (mononematous form, Hormodendrum resinae)<br />
40. Colonies effuse, dense, resupinate, hypochnoid, powdery, chocolate-brown <strong>and</strong>/or conidiophores lightly pigmented; conidia subhyaline<br />
to lightly pigmented <strong>and</strong>/or lateral walls not thickener than poles; on dead wood or isolated from hydrocarbone-rich substrates (jet-fuel,<br />
cosmetics, etc.) ........................................................................................................................................................................................ 41<br />
41. Colonies effuse, dense, resupinate, hypochnoid, powdery, chocolate-brown; conidiophores smooth; conidia subhyaline to very pale<br />
yellowish, hila very thin; on dead wood ......................................................................................................................... Parahaplotrichum<br />
41. Colonies neither resupinate nor hypochnoid; conidiophores warty; lateral walls of the conidia not thicker than the hila; isolated from<br />
hydrocarbone-rich substrates (jet-fuel, cosmetics, etc.) ......................................................................................................... Hormoconis<br />
42(38). Conidiophores simple or branched; conidiogenous cells monoblastic or occasionally polyblastic; conidiogenous loci subdenticulate,<br />
neither thickened nor darkened, forming simple or branched chains of regular conidia, uniform in shape, size <strong>and</strong> septation<br />
................................................................................................................................................................................................... Septonema<br />
42. Conidia not uniform in shape, size <strong>and</strong> septation; conidiogenous loci flat-tipped, subdenticulate, unthickened or slightly so, not to<br />
somewhat darkened-refractive ................................................................................................................................................................. 43<br />
43. Conidiophores simple or branched; in culture forming abundant chlamydospores; mostly soil-borne <strong>and</strong> heat-resistant (Teratosphaeriaceae,<br />
Capnodiales) ................................................................................................................................................................................ Devriesia<br />
43. Without chlamydospores in culture; phylogenetically distinct .................................................................................................................. 44<br />
44. Conidiophores dimorphic; conidia mostly aseptate, hila inconspicuous, neither thickened nor darkened (Pleosporales)<br />
.................................................................................................................................................................................... Ochrocladosporium<br />
44. Conidiophores either uniform or conidia at least partly septate or hila more conspicuous by being slightly thickened or at least somewhat<br />
darkened or refractive; phylogenetically distinct ...................................................................................................................................... 45<br />
45. Phialidic synanamorphs often present, but sometimes also lacking; saprobic, rarely plant pathogenic, often human pathogenic<br />
(Herpotrichiellaceae, Chaetothyriales) .......................................................................................................................... Cladophialophora<br />
45. Without phialidic synanamorphs; saprobic or plant pathogenic; phylogenetically distinct ....................................................................... 46<br />
46. Conidiophores usually unbranched (Venturia, Venturiaceae) .......................................... Fusicladium s. lat. (incl. Pseudocladosporium)<br />
[<strong>similar</strong>, barely distinguishable taxa, also clustering in the Venturiaceae, but apart from the Venturia clade are tentatively referred to as<br />
Anungitea until this <strong>genus</strong> will be resolved by sequences of its type species]<br />
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<strong>Cladosporium</strong> <strong>and</strong> morphologically <strong>similar</strong> genera<br />
46. Conidiophores simple to often irregularly branched; conidia delicate, narrow, 1–3 µm wide, hyaline to pale olivaceous (not belonging to<br />
the Venturiaceae) .................................................................................................................................................................. Polyscytalum<br />
Discussion<br />
Phylogenetic studies conducted on species of <strong>Cladosporium</strong> s. lat.<br />
proved the <strong>genus</strong> to be highly heterogeneous (Braun et al. 2003). It<br />
could be demonstrated that various anamorphs, previously referred<br />
to as <strong>Cladosporium</strong>, e.g. <strong>Cladosporium</strong> fulvum Cooke [≡ Passalora<br />
fulva (Cooke) U. Braun & Crous], have to be excluded since they<br />
clustered in the Mycosphaerella clade (Mycosphaerellaceae).<br />
Previous re-examinations <strong>and</strong> reassessments of human pathogenic<br />
<strong>Cladosporium</strong> species, including morphology, biology/ecology,<br />
physiology <strong>and</strong> molecular data (Masclaux et al. 1995, Untereiner<br />
1997, Gerrits van den Ende & de Hoog 1999, Untereiner & Naveau<br />
1999, Untereiner et al. 1999; de Hoog et al. 2000), could also be<br />
confirmed. In all phylogenetic analyses, it could be shown that the<br />
human pathogenic fungi concerned formed a clade belonging to<br />
the Herpotrichiellaceae (Capronia Sacc./Cladophialophora Borelli).<br />
Venturia anamorphs with catenate conidia, previously often<br />
assigned to <strong>Cladosporium</strong> s. lat., clustered together with other<br />
anamorphs of the Venturiaceae, <strong>and</strong> formed a monophyletic clade<br />
(Braun et al. 2003, Schubert et al. 2003, Beck et al. 2005). Venturia<br />
has now also been shown to accommodate less well-known<br />
anamorph genera such as Pseudocladosporium, which represent<br />
an additional synonym of Fusicladium Bonord. (Crous et al. 2007<br />
– this volume).<br />
Seifert et al. (2004) examined morphological, ecological <strong>and</strong><br />
molecular characters of <strong>Cladosporium</strong> staurophorum (W.B. Kendr.)<br />
M.B. Ellis <strong>and</strong> three allied heat-resistant species <strong>and</strong> placed them<br />
in the new <strong>genus</strong> Devriesia, which formed a monophyletic group<br />
apart from the <strong>Cladosporium</strong> clade. Crous et al. (2006b) erected<br />
the <strong>genus</strong> Cladoriella Crous for a saprobic species (incertae sedis)<br />
characterised by having narrowly ellipsoidal to cylindrical or fusoid,<br />
0–1-septate, medium brown, thick-walled, finely verruculose<br />
conidia arranged in simple or branched chains, with thickened,<br />
darkened, refractive hila, with a minute central pore. <strong>Cladosporium</strong><br />
musae E.W. Mason, the causal agent of banana speckle disease,<br />
has recently been shown to be allied to the Chaetothyriales (Crous<br />
et al. 2006a), <strong>and</strong> was placed in a new <strong>genus</strong>, Metulocladosporiella<br />
with C. musae as type species. Digitopodium U. Braun, Heuchert<br />
& K. Schub. (type species: <strong>Cladosporium</strong> hemileiae Steyaert)<br />
<strong>and</strong> Parapericoniella U. Braun, Heuchert & K. Schub. (type<br />
species: <strong>Cladosporium</strong> asterinae Deighton) represent two new<br />
genera of hyperparasitic hyphomycetes, introduced due to unique<br />
morphological features <strong>and</strong> striking differences to <strong>Cladosporium</strong> s.<br />
str. (Heuchert et al. 2005), but have as yet been excluded from<br />
DNA-based studies due to the absence of cultures. Schubert et al.<br />
(2007a – this volume) introduced a new <strong>genus</strong>, Dichocladosporium<br />
K. Schub., U. Braun & Crous (allied to the Davidiellaceae, Capnodiales)<br />
to accommodate a fungus with dimorphic fruiting that is<br />
pathogenic to Paeonia spp. <strong>The</strong> present study introduced yet several<br />
additional cladosporium-like genera, which could be distinguished<br />
based on their morphology <strong>and</strong> distinct DNA phylogeny, namely<br />
Ochrocladosporium (Pleosporales), Rhizocladosporium<br />
(incertae sedis), Rachicladosporium, Toxicocladosporium <strong>and</strong><br />
Verrucocladosporium (Capnodiales).<br />
Although all these genera are cladosporium-like, <strong>and</strong> many<br />
have in the past been confused with <strong>Cladosporium</strong> s. str., the unique<br />
www.studiesinmycology.org<br />
coronate scar type of <strong>Cladosporium</strong> s. str. allows a critical revision<br />
of cladosporioid hyphomycetes, based on reliable, distinctive<br />
morphological characters. In all cases where cladosporium-like<br />
(<strong>Cladosporium</strong> s. lat.) hyphomycetes clearly clustered apart<br />
from <strong>Cladosporium</strong> s. str. in the phylogenetic analyses, it could<br />
be demonstrated that the fungal groups concerned were also<br />
morphologically unambiguously distinguished, above all with regard<br />
to the structure of the conidiogenous hila. Hence, the excluded<br />
groups of species, belonging in other genera, sometimes even<br />
in new genera, are genetically as well as morphologically clearly<br />
distinct from <strong>Cladosporium</strong> s. str.<br />
Acknowledgements<br />
We thank F. Hill, A. Aptroot, F.M. Dugan, R.F. Castañeda <strong>and</strong> W. Gams for providing<br />
collections <strong>and</strong> cultures of <strong>Cladosporium</strong> <strong>and</strong> cladosporium-like species over the<br />
past few years, without which this study would not have been possible. A research<br />
visit of K. Schubert to <strong>CBS</strong> was supported by a Synthesys grant (No. 2559), <strong>and</strong><br />
the Odo van Vloten Stichting. We thank M. Vermaas for preparing the photographic<br />
plates, <strong>and</strong> A. van Iperen for preparing all the fungal cultures for examination. H.-J.<br />
Schroers is thanked for generating some of the sequence data used in this paper.<br />
A.J.L. Phillips is thanked for providing comments on an earlier draft of the paper.<br />
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56
available online at www.studiesinmycology.org<br />
doi:10.3114/sim.2007.58.03<br />
Studies in Mycology 58: 57–93. 2007.<br />
Phylogenetic <strong>and</strong> morphotaxonomic revision of Ramichloridium <strong>and</strong> allied<br />
genera<br />
M. Arzanlou 1,2 , J.Z. Groenewald 1 , W. Gams 1 , U. Braun 3 , H.-D Shin 4 <strong>and</strong> P.W. Crous 1,2*<br />
1<br />
<strong>CBS</strong> Fungal Biodiversity Centre, Uppsalalaan 8, 3584 CT Utrecht, <strong>The</strong> Netherl<strong>and</strong>s; 2 Laboratory of Phytopathology, Wageningen University, Binnenhaven 5, 6709 PD<br />
Wageningen, <strong>The</strong> Netherl<strong>and</strong>s; 3 Martin-Luther-Universität, Institut für Biologie, Geobotanik und Botanischer Garten, Neuwerk 21, D-06099 Halle (Saale), Germany; 4 Division of<br />
Environmental Science & Ecological Engineering, Korea University, Seoul 136-701, Korea<br />
*Correspondence: Pedro W. Crous, p.crous@cbs.knaw.nl<br />
Abstract: <strong>The</strong> phylogeny of the genera Periconiella, Ramichloridium, Rhinocladiella <strong>and</strong> Veronaea was explored by means of partial sequences of the 28S (LSU) rRNA<br />
gene <strong>and</strong> the ITS region (ITS1, 5.8S rDNA <strong>and</strong> ITS2). Based on the LSU sequence data, ramichloridium-like species segregate into eight distinct clusters. <strong>The</strong>se include the<br />
Capnodiales (Mycosphaerellaceae <strong>and</strong> Teratosphaeriaceae), the Chaetothyriales (Herpotrichiellaceae), the Pleosporales, <strong>and</strong> five ascomycete clades with uncertain affinities.<br />
<strong>The</strong> type species of Ramichloridium, R. apiculatum, together with R. musae, R. biverticillatum, R. cerophilum, R. verrucosum, R. pini, <strong>and</strong> three new species isolated from<br />
Strelitzia, Musa <strong>and</strong> forest soil, respectively, reside in the Capnodiales clade. <strong>The</strong> human-pathogenic species R. mackenziei <strong>and</strong> R. basitonum, together with R. fasciculatum<br />
<strong>and</strong> R. anceps, cluster with Rhinocladiella (type species: Rh. atrovirens, Herpotrichiellaceae, Chaetothyriales), <strong>and</strong> are allocated to this <strong>genus</strong>. Veronaea botryosa, the type<br />
species of the <strong>genus</strong> Veronaea, also resides in the Chaetothyriales clade, whereas Veronaea simplex clusters as a sister taxon to the Venturiaceae (Pleosporales), <strong>and</strong> is placed<br />
in a new <strong>genus</strong>, Veronaeopsis. Ramichloridium obovoideum clusters with Carpoligna pleurothecii (anamorph: Pleurothecium sp., Chaetosphaeriales), <strong>and</strong> a new combination<br />
is proposed in Pleurothecium. Other ramichloridium-like clades include R. subulatum <strong>and</strong> R. epichloës (incertae sedis, Sordariomycetes), for which a new <strong>genus</strong>, Radulidium<br />
is erected. Ramichloridium schulzeri <strong>and</strong> its varieties are placed in a new <strong>genus</strong>, Myrmecridium (incertae sedis, Sordariomycetes). <strong>The</strong> <strong>genus</strong> Pseudovirgaria (incertae sedis)<br />
is introduced to accommodate ramichloridium-like isolates occurring on various species of rust fungi. A veronaea-like isolate from Bertia moriformis with phylogenetic affinity to<br />
the Annulatascaceae (Sordariomycetidae) is placed in a new <strong>genus</strong>, Rhodoveronaea. Besides Ramichloridium, Periconiella is also polyphyletic. Thysanorea is introduced to<br />
accommodate Periconiella papuana (Herpotrichiellaceae), which is unrelated to the type species, P. velutina (Mycosphaerellaceae).<br />
Taxonomic novelties: Myrmecridium Arzanlou, W. Gams & Crous, gen. nov., Myrmecridium flexuosum (de Hoog) Arzanlou, W. Gams & Crous, comb. et stat. nov., Myrmecridium<br />
schulzeri (Sacc.) Arzanlou, W. Gams & Crous var. schulzeri, comb. nov., Myrmecridium schulzeri var. tritici (M.B. Ellis) Arzanlou, W. Gams & Crous, comb. nov., Periconiella<br />
arcuata Arzanlou, S. Lee & Crous, sp. nov., Periconiella levispora Arzanlou, W. Gams & Crous, sp. nov., Pleurothecium obovoideum (Matsush.) Arzanlou & Crous, comb. nov.,<br />
Pseudovirgaria H.D. Shin, U. Braun, Arzanlou & Crous, gen. nov., Pseudovirgaria hyperparasitica H.D. Shin, U. Braun, Arzanlou & Crous, sp. nov., Radulidium Arzanlou, W.<br />
Gams & Crous, gen. nov., Radulidium epichloës (Ellis & Dearn.) Arzanlou, W. Gams & Crous, comb. nov., Radulidium subulatum (de Hoog) Arzanlou, W. Gams & Crous, comb.<br />
nov., Ramichloridium australiense Arzanlou & Crous, sp. nov., Ramichloridium biverticillatum Arzanlou & Crous, nom. nov., Ramichloridium brasilianum Arzanlou & Crous, sp.<br />
nov., Ramichloridium strelitziae Arzanlou, W. Gams & Crous, sp. nov., Rhinocladiella basitona (de Hoog) Arzanlou & Crous, comb. nov., Rhinocladiella fasciculata (V. Rao &<br />
de Hoog) Arzanlou & Crous, comb. nov., Rhinocladiella mackenziei (C.K. Campb. & Al-Hedaithy) Arzanlou & Crous, comb. nov., Rhodoveronaea Arzanlou, W. Gams & Crous,<br />
gen. nov., Rhodoveronaea varioseptata Arzanlou, W. Gams & Crous, sp. nov., Thysanorea Arzanlou, W. Gams & Crous, gen. nov., Thysanorea papuana (Aptroot) Arzanlou, W.<br />
Gams & Crous, comb. nov., Veronaea japonica Arzanlou, W. Gams & Crous, sp. nov., Veronaeopsis Arzanlou & Crous, gen. nov., Veronaeopsis simplex (Papendorf) Arzanlou<br />
& Crous, comb.nov.<br />
Key words: Capnodiales, Chaetothyriales, Mycosphaerella, Periconiella, phylogeny, Rhinocladiella, Veronaea.<br />
Introduction<br />
<strong>The</strong> anamorph <strong>genus</strong> Ramichloridium Stahel ex de Hoog 1977<br />
presently accommodates a wide range of species with erect, dark,<br />
more or less differentiated, branched or unbranched conidiophores<br />
<strong>and</strong> predominantly aseptate conidia produced on a sympodially<br />
proliferating rachis (de Hoog 1977). This heterogeneous group<br />
of anamorphic fungi includes species with diverse life styles, viz.<br />
saprobes, human <strong>and</strong> plant pathogens, most of which were classified<br />
by Schol-Schwarz (1968) in Rhinocladiella Nannf. according to a<br />
very broad generic concept. Ramichloridium was originally erected<br />
by Stahel (1937) with R. musae Stahel as type species. However,<br />
because his publication lacked a Latin diagnosis, the <strong>genus</strong> was<br />
invalid. Stahel also invalidly described Chloridium musae Stahel for<br />
a fungus causing leaf spots (tropical speckle disease) on banana.<br />
Ellis (1976) validated Chloridium musae as Veronaea musae M.B.<br />
Ellis, <strong>and</strong> Ramichloridium musae as Periconiella musae Stahel ex<br />
M.B. Ellis.<br />
Periconiella Sacc. (1885) [type species P. velutina (G. Winter)<br />
Sacc.] differs from Veronaea Cif. & Montemart. chiefly based<br />
on its dark brown, apically branched conidiophores. However,<br />
de Hoog (1977) observed numerous specimens of V. musae to<br />
exhibit branched conidiophores in culture, as did Stahel (1937)<br />
for Ramichloridium musae. De Hoog (1977) subsequently reintroduced<br />
Ramichloridium, but typified it with R. apiculatum (J.H.<br />
Mill., Giddens & A.A. Foster) de Hoog. He regarded V. musae <strong>and</strong><br />
P. musae to be conspecific, <strong>and</strong> applied the name R. musae (Stahel<br />
ex M.B. Ellis) de Hoog to both species, regarding Periconiella<br />
musae as basionym. <strong>The</strong> circumscription by de Hoog was based<br />
on their <strong>similar</strong> morphology <strong>and</strong> ecology. Central in his <strong>genus</strong><br />
concept was the observed presence of more or less differentiated<br />
<strong>and</strong> pigmented conidiophores, with predominantly aseptate conidia<br />
produced on a sympodially proliferating rachis. De Hoog (1977)<br />
also used some ecological features as additional characters to<br />
discriminate Ramichloridium from other genera, noting, for instance,<br />
that species in Ramichloridium were non-pathogenic to humans<br />
(de Hoog 1977, Campbell & Al-Hedaithy 1993). This delimitation,<br />
however, was not commonly accepted (McGinnis & Schell 1980).<br />
De Hoog et al. (1983) further discussed the problematic separation<br />
of Ramichloridium from genera such as Rhinocladiella, Veronaea<br />
<strong>and</strong> <strong>Cladosporium</strong> Link. It was further noted that the main feature to<br />
distinguish Ramichloridium from Rhinocladiella, was the presence<br />
of exophiala-type budding cells in species of Rhinocladiella (de<br />
Hoog 1977, de Hoog et al. 1983, Veerkamp & Gams 1983). <strong>The</strong><br />
separation of Veronaea from this complex is more problematic, as<br />
the circumscriptions provided by Ellis (1976) <strong>and</strong> Morgan-Jones<br />
(1979, 1982) overlap with that of Ramichloridium sensu de Hoog<br />
(1977). <strong>Cladosporium</strong> is more distinct, having very conspicuous,<br />
protuberant, darkened <strong>and</strong> thickened, coronate conidial scars, <strong>and</strong><br />
catenate conidia (David 1997, Braun et al. 2003, Schubert et al.<br />
2007 – this volume).<br />
57
Arzanlou et al.<br />
To date 26 species have been named in Ramichloridium; they<br />
not only differ in morphology, but also in life style. Ramichloridium<br />
mackenziei C.K. Campb. & Al-Hedaithy is a serious human<br />
pathogen, causing cerebral phaeohyphomycosis (Al-Hedaithy et al.<br />
1988, Campbell & Al-Hedaithy 1993), whereas R. musae causes<br />
tropical speckle disease on members of the Musaceae (Stahel 1937,<br />
Jones 2000). Another plant-pathogenic species, R. pini de Hoog &<br />
Rahman, causes a needle disease on Pinus contorta (de Hoog et<br />
al. 1983). Other clinically relevant species of Ramichloridium are R.<br />
basitonum de Hoog <strong>and</strong> occasionally R. schulzeri (Sacc.) de Hoog,<br />
while the remaining species tend to be common soil saprobes.<br />
No teleomorph has thus far been linked to species of<br />
Ramichloridium. <strong>The</strong> main question that remains is whether shared<br />
morphology among the species in this <strong>genus</strong> reflects common<br />
ancestry (Seifert 1993, Untereiner & Naveau 1999). To delineate<br />
anamorphic genera adequately, morphology <strong>and</strong> conidial ontogeny<br />
alone are no longer satisfactory (Crous et al. 2006a, b), <strong>and</strong> DNA<br />
data provide additional characters to help delineate species <strong>and</strong><br />
genera (Taylor et al. 2000, Mostert et al. 2006, Zipfel et al. 2006).<br />
<strong>The</strong> aim of the present study was to integrate morphological<br />
<strong>and</strong> cultural features with DNA sequence data to resolve the<br />
species concepts <strong>and</strong> generic limits of the taxa currently placed in<br />
Periconiella, Ramichloridium, Rhinocladiella <strong>and</strong> Veronaea, <strong>and</strong> to<br />
resolve the status of several new cultures that were isolated during<br />
the course of this study.<br />
Materials <strong>and</strong> Methods<br />
Isolates<br />
Species names, substrates, geographical origins <strong>and</strong> GenBank<br />
accession numbers of the isolates included in this study are listed<br />
in Table 1. Fungal isolates are maintained in the culture collection<br />
of the Centraalbureau voor Schimmelcultures (<strong>CBS</strong>) in Utrecht, the<br />
Netherl<strong>and</strong>s.<br />
DNA extraction, amplification <strong>and</strong> sequence analysis<br />
Genomic DNA was extracted from colonies grown on 2 % malt<br />
extract agar (MEA, Difco) (Gams et al. 2007) using the FastDNA kit<br />
(BIO101, Carlsbad, CA, U.S.A.). <strong>The</strong> primers ITS1 <strong>and</strong> ITS4 (White<br />
et al. 1990) were used to amplify the internal transcribed spacer<br />
region (ITS) of the nuclear ribosomal RNA operon, including: the<br />
3’ end of the 18S rRNA gene, the first internal transcribed spacer<br />
region (ITS1), the 5.8S rRNA gene, the second internal transcribed<br />
spacer region (ITS2) <strong>and</strong> the 5’ end of 28S rRNA gene. Part of<br />
the large subunit 28S rRNA (LSU) gene was amplified with primers<br />
LR0R (Rehner & Samuels 1994) <strong>and</strong> LR5 (Vilgalys & Hester 1990).<br />
<strong>The</strong> ITS region was sequenced only for those isolates for which<br />
these data were not available. <strong>The</strong> ITS analyses confirmed the<br />
proposed classification based on LSU analysis for each major<br />
clade <strong>and</strong> are not presented here in detail; but the sequences are<br />
deposited in GenBank where applicable. <strong>The</strong> PCR reaction was<br />
performed in a mixture with 0.5 units Taq polymerase (Bioline,<br />
London, U.K.), 1× PCR buffer, 0.5 mM MgCl 2<br />
, 0.2 mM of each<br />
dNTP, 5 pmol of each primer, approximately 10–15 ng of fungal<br />
genomic DNA, with the total volume adjusted to 25 µL with<br />
sterile water. Reactions were performed on a GeneAmp PCR<br />
System 9700 (Applied Biosystems, Foster City, CA) with cycling<br />
conditions consisting of 5 min at 96 °C for primary denaturation,<br />
followed by 36 cycles at 96 °C (30 s), 52 °C (30 s), <strong>and</strong> 72 °C<br />
(60 s), with a final 7 min extension step at 72 °C to complete the<br />
reaction. <strong>The</strong> amplicons were sequenced using BigDye Terminator<br />
v. 3.1 (Applied Biosystems, Foster City, CA) or DYEnamicET<br />
Terminator (Amersham Biosciences, Freiburg, Germany) Cycle<br />
Sequencing Kits <strong>and</strong> analysed on an ABI Prism 3700 (Applied<br />
Biosystems, Foster City, CA) under conditions recommended by<br />
the manufacturer. Newly generated sequences were subjected<br />
to a Blast search of the NCBI databases, sequences with high<br />
<strong>similar</strong>ity were downloaded from GenBank <strong>and</strong> comparisons<br />
were made based on the alignment of the obtained sequences.<br />
Sequences from GenBank were also selected for <strong>similar</strong> taxa. <strong>The</strong><br />
LSU tree was rooted using sequences of Athelia epiphylla Pers.<br />
<strong>and</strong> Paullicorticium ansatum Liberta as outgroups. Phylogenetic<br />
analysis was performed with PAUP (Phylogenetic Analysis Using<br />
Parsimony) v. 4.0b10 (Swofford 2003), using the neighbour-joining<br />
algorithm with the uncorrected (“p”), the Kimura 2-parameter <strong>and</strong><br />
the HKY85 substitution models. Alignment gaps longer than 10<br />
bases were coded as single events for the phylogenetic analyses;<br />
the remaining gaps were treated as missing data. Any ties were<br />
broken r<strong>and</strong>omly when encountered. Phylogenetic relationships<br />
were also inferred with the parsimony algorithm using the heuristic<br />
search option with simple taxon additions <strong>and</strong> tree bisection <strong>and</strong><br />
reconstruction (TBR) as the branch-swapping algorithm; alignment<br />
gaps were treated as a fifth character state <strong>and</strong> all characters were<br />
unordered <strong>and</strong> of equal weight. Branches of zero length were<br />
collapsed <strong>and</strong> all multiple, equally parsimonious trees were saved.<br />
Only the first 5 000 equally most parsimonious trees were saved.<br />
Other measures calculated included tree length, consistency<br />
index, retention index <strong>and</strong> rescaled consistency index (TL, CI, RI<br />
<strong>and</strong> RC, respectively). <strong>The</strong> robustness of the obtained trees was<br />
evaluated by 1 000 bootstrap replications. Bayesian analysis was<br />
performed following the methods of Crous et al. (2006c). <strong>The</strong> best<br />
nucleotide substitution model was determined using MrModeltest<br />
v. 2.2 (Nyl<strong>and</strong>er 2004). MrBayes v. 3.1.2 (Ronquist & Huelsenbeck<br />
2003) was used to perform phylogenetic analyses, using a general<br />
time-reversible (GTR) substitution model with inverse gamma<br />
rates, dirichlet base frequencies <strong>and</strong> the temp value set to 0.5. New<br />
sequences were lodged with NCBI’s GenBank (Table 1) <strong>and</strong> the<br />
alignment <strong>and</strong> trees with TreeBASE (www.treebase.org).<br />
Morphology<br />
Cultural growth rates <strong>and</strong> morphology were recorded from colonies<br />
grown on MEA for 2 wk at 24 ºC in the dark, <strong>and</strong> colony colours<br />
were determined by reference to the colour charts of Rayner (1970).<br />
Microscopic observations were made from colonies cultivated on<br />
MEA <strong>and</strong> OA (oatmeal agar, Gams et al. 2007), using a slide culture<br />
technique. Slide cultures were set up in Petri dishes containing 2<br />
mL of sterile water, into which a U-shaped glass rod was placed,<br />
extending above the water surface. A block of freshly growing<br />
fungal colony, approx. 1 cm square was placed onto a sterile<br />
microscope slide, covered with a somewhat larger, sterile glass<br />
cover slip, <strong>and</strong> incubated in the moist chamber. Fungal sporulation<br />
was monitored over time, <strong>and</strong> when optimal, images were captured<br />
by means of a Nikon camera system (Digital Sight DS-5M, Nikon<br />
Corporation, Japan). Structures were mounted in lactic acid, <strong>and</strong><br />
30 measurements (× 1 000 magnification) determined wherever<br />
possible, with the extremes of spore measurements given in<br />
parentheses.<br />
58
Ramichloridium <strong>and</strong> allied genera<br />
Table 1. Isolates of Ramichloridium <strong>and</strong> <strong>similar</strong> genera used for DNA analysis <strong>and</strong> morphological studies.<br />
Species Accession number 1 Source Origin GenBank numbers<br />
(LSU, ITS)<br />
Myrmecridium flexuosum <strong>CBS</strong> 398.76*; IMI 203547 Soil Suriname EU041825, EU041768<br />
Myrmecridium schulzeri <strong>CBS</strong> 100.54; JCM 6974 Soil Zaire EU041826, EU041769<br />
<strong>CBS</strong> 134.68; ATCC 16310 Soil Germany EU041827, EU041770<br />
<strong>CBS</strong> 156.63 Homo sapiens Netherl<strong>and</strong>s EU041828, EU041771<br />
<strong>CBS</strong> 188.96 Soil Papua New Guinea EU041829, EU041772<br />
<strong>CBS</strong> 304.73 Wheat straw South Africa EU041830, EU041773<br />
<strong>CBS</strong> 305.73; JCM 6967 Wheat straw South Africa EU041831, EU041774<br />
<strong>CBS</strong> 325.74; JCM 7234 Triticum aestivum Netherl<strong>and</strong>s EU041832, EU041775<br />
<strong>CBS</strong> 381.87 — Australia EU041833, EU041776<br />
<strong>CBS</strong> 642.76 Malus sylvestris Switzerl<strong>and</strong> EU041834, EU041777<br />
<strong>CBS</strong> 114996 Cannomois virgata South Africa EU041835, EU041778<br />
Periconiella arcuata <strong>CBS</strong> 113477* Ischyrolepsis subverticellata South Africa EU041836, EU041779<br />
Periconiella levispora <strong>CBS</strong> 873.73* Turpinia pomifera Sri Lanka EU041837, EU041780<br />
Periconiella velutina <strong>CBS</strong> 101948*; CPC 2262 Brabejum stellatifolium South Africa EU041838, EU041781<br />
<strong>CBS</strong> 101949; CPC 2263 Brabejum stellatifolium South Africa EU041839, EU041782<br />
<strong>CBS</strong> 101950; CPC 2264 Brabejum stellatifolium South Africa EU041840, EU041783<br />
Pleurothecium obovoideum <strong>CBS</strong> 209.95*; MFC 12477 Pasania edulis Japan EU041841, EU041784<br />
Pseudovirgaria hyperparasitica <strong>CBS</strong> 121735; CPC 10702 On Phragmidium sp. on Rubus coreanus Korea EU041822, EU041765<br />
<strong>CBS</strong> 121738; CPC 10704 On Phragmidium sp. on Rubus coreanus Korea EU041823, EU041766<br />
<strong>CBS</strong> 121739*; CPC 10753<br />
On Pucciniastrum agrimoniae on<br />
Korea<br />
EU041824, EU041767<br />
Agrimonia pilosa<br />
Radulidium epichloës <strong>CBS</strong> 361.63*; MUCL 3124 Epichloë typhina U.S.A. EU041842, EU041785<br />
Radulidium sp. <strong>CBS</strong> 115704 Poaceae Guyana EU041843, EU041786<br />
Radulidium subulatum <strong>CBS</strong> 287.84 Puccinia allii U.K. EU041844, EU041787<br />
Ramichloridium apiculatum<br />
<strong>CBS</strong> 405.76* Phragmites australis Czech Republic EU041845, EU041788<br />
<strong>CBS</strong> 912.96 Incubator for cell cultures Germany EU041846, EU041789<br />
<strong>CBS</strong> 101010 Lasioptera arundinis Czech Republic EU041847, EU041790<br />
<strong>CBS</strong> 156.59*; ATCC 13211; IMI Forest soil U.S.A. EU041848, EU041791<br />
100716; JCM 6972; MUCL 7991;<br />
MUCL 15753; QM 7716<br />
<strong>CBS</strong> 390.67 Cucumis sativus South Africa EU041849, EU041792<br />
<strong>CBS</strong> 391.67; JCM 6966 Aloe sp. South Africa EU041850, EU041793<br />
<strong>CBS</strong> 400.76; IMI 088021 Soil Pakistan EU041851, EU041794<br />
Ramichloridium australiense <strong>CBS</strong> 121710 Musa banksii Australia EU041852, EU041795<br />
Ramichloridium biverticillatum <strong>CBS</strong> 335.36 Musa sapientum — EU041853, EU041796<br />
Ramichloridium brasilianum <strong>CBS</strong> 283.92* Forest soil Brazil EU041854, EU041797<br />
Ramichloridium cerophilum <strong>CBS</strong> 103.59* Sasa sp. Japan EU041855, EU041798<br />
Ramichloridium indicum <strong>CBS</strong> 171.96 — — EU041856, EU041799<br />
Ramichloridium musae <strong>CBS</strong> 190.63; MUCL 9557 Musa sapientum — EU041857, EU041800<br />
<strong>CBS</strong> 365.36*; JCM 6973; MUCL 9556 Musa sapientum Surinam EU041858, EU041801<br />
Ramichloridium pini <strong>CBS</strong> 461.82*; MUCL 28942 Pinus contorta U.K. EU041859, EU041802<br />
Ramichloridium strelitziae <strong>CBS</strong> 121711 Strelitzia sp. South Africa EU041860, EU041803<br />
Rhinocladiella anceps<br />
<strong>CBS</strong> 157.54; ATCC 15680; MUCL Fagus sylvatica France EU041861, EU041804<br />
1081; MUCL 7992; MUCL 15756<br />
<strong>CBS</strong> 181.65*; ATCC 18655; DAOM Soil Canada EU041862, EU041805<br />
84422; IMI 134453; MUCL 8233; OAC<br />
10215<br />
Rhinocladiella basitona <strong>CBS</strong> 101460*; IFM 47593 Homo sapiens Japan EU041863, EU041806<br />
Rhinocladiella fasciculata <strong>CBS</strong> 132.86* Decayed wood India EU041864, EU041807<br />
Rhinocladiella mackenziei <strong>CBS</strong> 367.92; NCPF 2738; UTMB 3169 Homo sapiens Israel EU041865, EU041808<br />
www.studiesinmycology.org<br />
59
Arzanlou et al.<br />
Table 1. (Continued).<br />
Species Accession number 1 Source Origin GenBank numbers<br />
(LSU, ITS)<br />
<strong>CBS</strong> 368.92; UTMB 3170 Homo sapiens Israel EU041866, EU041809<br />
<strong>CBS</strong> 102590; NCPF 2853 Homo sapiens United Arab Emirates EU041867, EU041810<br />
Rhinocladiella phaeophora <strong>CBS</strong> 496.78*; IMI 287527 Soil Colombia EU041868, EU041811<br />
Rhinocladiella sp. <strong>CBS</strong> 264.49; MUCL 9904 Honey France EU041869, EU041812<br />
Rhodoveronaea varioseptata <strong>CBS</strong> 431.88* Bertia moriformis Germany EU041870, EU041813<br />
Thysanorea papuana <strong>CBS</strong> 212.96* — Papua New Guinea EU041871, EU041814<br />
Veronaea botryosa <strong>CBS</strong> 121.92 Xanthorrhoea preissii Australia EU041872, EU041815<br />
<strong>CBS</strong> 254.57*; IMI 070233; MUCL 9821 — Italy EU041873, EU041816<br />
<strong>CBS</strong> 350.65; IMI 115127; MUCL 7972 Goat dung India EU041874, EU041817<br />
Veronaea compacta <strong>CBS</strong> 268.75* — South Africa EU041876, EU041819<br />
Veronaea japonica <strong>CBS</strong> 776.83* On dead bamboo culm Japan EU041875, EU041818<br />
Veronaeopsis simplex <strong>CBS</strong> 588.66*; IMI 203547 Acacia karroo South Africa EU041877, EU041820<br />
Zasmidium cellare <strong>CBS</strong> 146.36 Wine cellar — EU041878, EU041821<br />
1<br />
ATCC: American Type Culture Collection, Virginia, U.S.A.; <strong>CBS</strong>: Centraalbureau voor Schimmelcultures, Utrecht, <strong>The</strong> Netherl<strong>and</strong>s; CPC: Culture collection of Pedro Crous,<br />
housed at <strong>CBS</strong>; DAOM: Plant Research Institute, Department of Agriculture (Mycology), Ottawa, Canada; IFM: Research Center for Pathogenic Fungi <strong>and</strong> Microbial Toxicoses,<br />
Chiba University, Chiba, Japan; IMI: International Mycological Institute, CABI-Bioscience, U.K.; JCM: Japan Collection of Microorganism, RIKEN BioResource Center, Japan;<br />
MFC: Matsushima Fungus Collection, Kobe, Japan; MUCL: Mycotheque de l’ Université Catholique de Louvain, Louvain-la-Neuve, Belgium; NCPF: <strong>The</strong> National Collection of<br />
Pathogenic Fungi, Holborn, London, U.K.; OAC: Department of Botany <strong>and</strong> Genetics, University of Guelph, Ont., Canada; QM: Quartermaster Research <strong>and</strong> Developement<br />
Center, U.S. Army, MA, U.S.A.; UTMB: University of Texas Medical Branch, Texas, U.S.A.<br />
*Ex-type cultures.<br />
Results<br />
Phylogeny<br />
<strong>The</strong> manually adjusted alignment of the 28S rDNA data contained<br />
137 sequences (including the two outgroups) <strong>and</strong> 995 characters<br />
including alignment gaps. Of the 748 characters used in the<br />
phylogenetic analysis, 373 were parsimony-informative, 61 were<br />
variable <strong>and</strong> parsimony-uninformative, <strong>and</strong> 314 were constant.<br />
Neighbour-joining analysis using the three substitution models on<br />
the LSU alignment yielded trees with <strong>similar</strong> topology <strong>and</strong> bootstrap<br />
values. Parsimony analysis of the alignment yielded 5 000 equally<br />
most parsimonious trees, one of which is shown in Fig. 1 (TL =<br />
2 157, CI = 0.377, RI = 0.875, RC = 0.330). <strong>The</strong> Markov Chain<br />
Monte Carlo (MCMC) analysis of four chains started from a r<strong>and</strong>om<br />
tree topology <strong>and</strong> lasted 2 000 000 generations. Trees were saved<br />
each 1 000 generations, resulting in 2 000 trees. Burn-in was set<br />
at 500 000 generations after which the likelihood values were<br />
stationary, leaving 1 500 trees from which the consensus tree (Fig.<br />
2) <strong>and</strong> posterior probabilities (PP’s) were calculated. <strong>The</strong> average<br />
st<strong>and</strong>ard deviation of split frequencies was 0.043910 at the end<br />
of the run. Among the neighbour-joining, Bayesian <strong>and</strong> parsimony<br />
analyses, the trees differed in the hierarchical order of the main<br />
families <strong>and</strong> the support values (data not shown; e.g. the support<br />
within of the Capnodiales in Figs 1–2).<br />
<strong>The</strong> phylogenetic trees (Figs 1–2) show that the Ramichloridium<br />
species segregate into eight distinct clades, residing in the<br />
Capnodiales (Mycosphaerellaceae <strong>and</strong> Teratosphaeriaceae), the<br />
Chaetothyriales (Herpotrichiellaceae), the Pleosporales, <strong>and</strong> five<br />
other clades of which the relationships remain to be elucidated. <strong>The</strong><br />
type species of Ramichloridium, R. apiculatum, together with R.<br />
musae, R. cerophilum (Tubaki) de Hoog, R. indicum (Subram.) de<br />
Hoog, R. pini <strong>and</strong> three new species respectively isolated from Musa<br />
banksii, Strelitzia nicolai, <strong>and</strong> forest soil, reside in different parts of<br />
the Capnodiales clade (all in the Mycosphaerellaceae, except for the<br />
species from forest soil which clusters in the Teratosphaeriaceae).<br />
<strong>The</strong> second clade (in the Chaetothyriomycetes clade), including<br />
the human-pathogenic species R. mackenziei <strong>and</strong> R. basitonum,<br />
together with R. fasciculatum V. Rao & de Hoog <strong>and</strong> R. anceps<br />
(Sacc. & Ellis) de Hoog, groups together with Rhinocladiella in the<br />
Herpotrichiellaceae. <strong>The</strong> third clade (in the Sordariomycetes clade)<br />
includes R. obovoideum (Matsush.) de Hoog, which in a Blast<br />
search was found to have affinity with Carpoligna pleurothecii F.A.<br />
Fernández & Huhndorf (Chaetosphaeriales). <strong>The</strong> fourth clade (in<br />
the Sordariomycetes clade) includes a veronaea-like isolate from<br />
Bertia moriformis, with phylogenetic affinity to the Annulatascaceae<br />
(Sordariomycetidae). <strong>The</strong> fifth clade (in the Sordariomycetes clade)<br />
includes R. schulzeri var. schulzeri <strong>and</strong> R. schulzeri var. flexuosum<br />
de Hoog, the closest relatives being Thyridium vestitum (Fr.)<br />
Fuckel in the Thyridiaceae <strong>and</strong> Magnaporthe grisea (T.T. Hebert)<br />
M.E. Barr in the Magnaporthaceae. <strong>The</strong> sixth clade (in the Incertae<br />
sedis clade) includes R. subulatum de Hoog, R. epichloës (Ellis &<br />
Dearn.) de Hoog <strong>and</strong> a species isolated from the Poaceae. Three<br />
ramichloridium-like isolates from Rubus coreanus <strong>and</strong> Agrimonia<br />
pilosa form another unique clade (in the Incertae sedis clade)<br />
with uncertain affinity. Veronaea simplex Papendorf clusters as<br />
sister taxon to the Venturiaceae representing the eighth clade<br />
(Dothideomycetes). <strong>The</strong> type species of Periconiella, P. velutina,<br />
clusters within the Mycosphaerellaceae (Capnodiales clade),<br />
whereas P. papuana Aptroot resides in the Herpotrichiellaceae<br />
(Chaetothyriales clade). Veronaea botryosa Cif. & Montemart., the<br />
type species of Veronaea, also resides in the Herpotrichiellaceae.<br />
Taxonomy<br />
<strong>The</strong> species previously described in Ramichloridium share<br />
some morphological features, including erect, pigmented, more<br />
or less differentiated conidiophores, sympodially proliferating<br />
conidiogenous cells <strong>and</strong> predominantly aseptate conidia. Other than<br />
conidial morphology, features of the conidiogenous apparatus that<br />
60
Ramichloridium <strong>and</strong> allied genera<br />
appear to be more phylogenetically informative include pigmentation<br />
of vegetative hyphae, conidiophores <strong>and</strong> conidia, denticle density<br />
on the rachis, <strong>and</strong> structure of the scars. By integrating these data<br />
with the molecular data set, more natural genera are delineated,<br />
which are discussed below.<br />
Key to ramichloridium-like genera<br />
1. Conidiogenous cells integrated, terminal <strong>and</strong> lateral on creeping or ascending hyphae (differentiation between branched vegetative<br />
hyphae <strong>and</strong> conidiophores barely possible); conidiogenous loci bulging, more or less umbonate, apex rounded; occurring on rust<br />
pustules ......................................................................................................................................................................... Pseudovirgaria<br />
1. Conidiogenous cells integrated in distinct conidiophores; conidiogenous loci non-umbonate (flat, not prominent; subcylindrical or conical<br />
denticles; or terminally flat-tipped; or thickened <strong>and</strong> darkened); rarely occurring on rust pustules, but if so, with a raduliform rachis <strong>and</strong><br />
distinct denticles......................................................................................................................................................................................... 2<br />
2. Conidia 0–2(–3)-septate, conidial base truncate, retaining a marginal frill after liberation [anamorphs of Sordariomycetes]<br />
........................................................................................................................................................................................... Rhodoveronaea<br />
2. Conidial base without marginal frill ............................................................................................................................................................ 3<br />
3. Conidiophores composed of a well-developed erect stalk <strong>and</strong> a terminal branched head ........................................................................ 4<br />
3. Conidiophores unbranched or, if branched, branches loose, irregular or dichotomous, but not distinctly separated into stalk <strong>and</strong> branched<br />
head ........................................................................................................................................................................................................... 5<br />
4. Conidiophores dimorphic, either macronematous, dark brown with a dense apical cluster of branches or micronematous, undifferentiated,<br />
resembling vegetative hyphae; both kinds with a denticulate rachis; conidia predominantly 1-septate [anamorph of Chaetothyriales]<br />
.................................................................................................................................................................................................. Thysanorea<br />
4. Conidiophores monomorphic; branched head with fewer branches <strong>and</strong> looser; conidiogenous loci usually flat, non-prominent, less<br />
denticle-like; conidia aseptate to pluriseptate [anamorphs of Capnodiales] ............................................................................ Periconiella<br />
5. Rachis with denticles 1–1.5 µm long, denticles almost cylindrical; conidia at least partly in short chains ......................... Pleurothecium<br />
5. Rachis with denticles less than 1 µm long, denticles not cylindrical or denticles lacking, rachis with flat, barely prominent scars ........... 6<br />
6. Conidia predominantly septate .................................................................................................................................................................. 7<br />
6. Conidia predominantly aseptate ................................................................................................................................................................ 8<br />
7. Conidiophores up to 200 µm long; rachis straight, not to slightly geniculate; conidiogenous loci more or less flat, barely prominent,<br />
unthickened, slightly darkened [anamorphs of Chaetothyriales, Herpotrichiellaceae] ................................................................. Veronaea<br />
7. Conidiophores up to 60 µm long; rachis distinctly geniculate; conidiogenous loci denticle-like, prominent, up to 0.5 µm high, slightly<br />
thickened <strong>and</strong> darkened [anamorph of Pleosporales, Venturiaceae] ................................................................................... Veronaeopsis<br />
8. Vegetative mycelium entirely hyaline; rachis long, hyaline, with widely scattered pimple-shaped, terminally pointed, unpigmented<br />
denticles .............................................................................................................................................................................. Myrmecridium<br />
8. Vegetative mycelium at least partly pigmented; conidiogenous loci distinct, non-denticulate, somewhat darkened-refractive, or denticles,<br />
if present, neither pimple-shaped nor pointed ............................................................................................................................................ 9<br />
9. Rachis distinctly raduliform, with distinct, prominent blunt denticles, 0.5–1 µm long; scars <strong>and</strong> hila unthickened, but pigmented<br />
.................................................................................................................................................................................................. Radulidium<br />
9. Rachis not distinctly raduliform, at most subdenticulate; scars flat or only slightly prominent (subdenticulate), shorter ......................... 10<br />
10. Conidiophores usually poorly differentiated from the vegetative hyphae; conidial apparatus often loosely branched; exophiala-like<br />
budding cells usually present in culture [anamorphs of Chaetothyriales, Herpotrichiellaceae] .......................................... Rhinocladiella<br />
10. Conidiophores usually well differentiated from the vegetative mycelium (macronematous), usually unbranched; without exophiala-like<br />
states [anamorphs of Capnodiales] .................................................................................................................................. Ramichloridium<br />
Capnodiales (Mycosphaerellaceae, Teratosphaeriaceae)<br />
<strong>The</strong> type species of Ramichloridium, R. apiculatum, together<br />
with R. indicum cluster as a sister group to the Dissoconium de<br />
Hoog, Oorschot & Hijwegen clade in the Mycosphaerellaceae.<br />
Some other Ramichloridium species, including R. musae, R.<br />
biverticillatum Arzanlou & Crous, R. pini <strong>and</strong> R. cerophilum, are<br />
also allied with members of the Mycosphaerellaceae. Three<br />
additional new species are introduced for Ramichloridium isolates<br />
www.studiesinmycology.org<br />
from Musa banksii, Strelitzia nicolai, <strong>and</strong> forest soil. Periconiella<br />
velutina, the type species of Periconiella, which also resides in the<br />
Mycosphaerellaceae, is morphologically sufficiently distinct to retain<br />
its generic status. Two new species of Periconiella are introduced<br />
for isolates obtained from Turpinia pomifera <strong>and</strong> Ischyrolepis<br />
subverticellata in South Africa. Zasmidium cellare (Pers.) Fr., the<br />
type species of Zasmidium (Pers.) Fr., is also shown to cluster<br />
within the Mycosphaerellaceae.<br />
61
Arzanlou et al.<br />
10 changes<br />
Athelia epiphylla AY586633<br />
Paullicorticium ansatum AY586693<br />
100 Conioscyphascus varius AY484512<br />
100<br />
Conioscypha lignicola AY484513<br />
100 Carpoligna pleurothecii AF064645<br />
78<br />
Carpoligna pleurothecii AF064646<br />
Carpoligna pleurothecii AY544685<br />
98<br />
74 Pleurothecium obovoideum <strong>CBS</strong> 209.95<br />
Ascotaiwania hughesii AY316357<br />
100 Ascotaiwania hughesii AY094189<br />
Ophiostoma stenoceras DQ836904<br />
96<br />
Magnaporthe grisea AB026819<br />
100 Cryptadelphia polyseptata AY281102<br />
Cryptadelphia groenendalensis AY281103<br />
100<br />
100 82<br />
86<br />
96<br />
89<br />
Rhodoveronaea varioseptata <strong>CBS</strong> 431.88<br />
Annulatascus triseptatus AY780049<br />
Annulatascus triseptatus AY346257<br />
Thyridium vestitum AY544671<br />
Capronia pulcherrima AF050256<br />
Exophiala dermatitidis AF050270<br />
Capronia mansonii AY004338<br />
Rhinocladiella sp. <strong>CBS</strong> 264.49<br />
100<br />
100<br />
Myrmecridium schulzeri <strong>CBS</strong> 114996<br />
Myrmecridium flexuosum <strong>CBS</strong> 398.76<br />
Myrmecridium schulzeri <strong>CBS</strong> 188.96<br />
Myrmecridium schulzeri <strong>CBS</strong> 381.87<br />
Myrmecridium schulzeri <strong>CBS</strong> 305.73<br />
Myrmecridium schulzeri <strong>CBS</strong> 304.73<br />
Myrmecridium schulzeri <strong>CBS</strong> 100.54<br />
Myrmecridium schulzeri <strong>CBS</strong> 642.76<br />
Myrmecridium schulzeri <strong>CBS</strong> 134.68<br />
Myrmecridium schulzeri <strong>CBS</strong> 156.63<br />
Myrmecridium schulzeri <strong>CBS</strong> 325.74<br />
Rhinocladiella mackenziei <strong>CBS</strong> 368.92<br />
Rhinocladiella mackenziei AF050288<br />
Rhinocladiella mackenziei <strong>CBS</strong> 367.92<br />
Rhinocladiella mackenziei <strong>CBS</strong>102590<br />
Capronia coronata AF050242<br />
Rhinocladiella fasciculata <strong>CBS</strong> 132.86<br />
Rhinocladiella phaeophora <strong>CBS</strong> 496.78<br />
Rhinocladiella anceps AF050284<br />
Rhinocladiella anceps <strong>CBS</strong> 181.65<br />
Rhinocladiella anceps AF050285<br />
Rhinocladiella anceps <strong>CBS</strong> 157.54<br />
Fonsecaea pedrosoi AF050276<br />
Veronaea botryosa <strong>CBS</strong> 350.65<br />
Veronaea botryosa <strong>CBS</strong> 121.92<br />
Veronaea botryosa <strong>CBS</strong> 254.57<br />
Thysanorea papuana <strong>CBS</strong> 212.96<br />
Veronaea japonica <strong>CBS</strong> 776.83<br />
Veronaea compacta <strong>CBS</strong> 268.75<br />
99<br />
80 60<br />
100<br />
95<br />
88<br />
Rhinocladiella basitona <strong>CBS</strong> 101460<br />
Rhinocladiella atrovirens AF050289<br />
Exophiala jeanselmei AF050271<br />
Veronaeopsis simplex <strong>CBS</strong> 588.66<br />
Repetophragma goidanichii DQ408574<br />
Venturia pyrina EF114715<br />
Metacoleroa dickiei DQ384100<br />
Venturia chlorospora DQ384101<br />
Venturia inaequalis EF114713<br />
Venturia hanliniana AF050290<br />
54<br />
Venturia asperata EF114711<br />
Venturia carpophila AY849967<br />
Sordariomycetes<br />
Herpotrichiellaceae,<br />
Chaetothyriales,<br />
Chaetothyriomycetes<br />
Venturiaceae,<br />
Pleosporales,<br />
Dothideomycetes<br />
Fig. 1. (Page 62–63). One of 5 000 equally most parsimonious trees obtained from a heuristic search with simple taxon additions of the LSU sequence alignment using PAUP v.<br />
4.0b10. <strong>The</strong> scale bar shows 10 changes; bootstrap support values from 1 000 replicates are shown at the nodes. Thickened lines indicate the strict consensus branches <strong>and</strong> extype<br />
sequences are printed in bold face. <strong>The</strong> tree was rooted to two sequences obtained from GenBank (Athelia epiphylla AY586633 <strong>and</strong> Paullicorticium ansatum AY586693).<br />
Periconiella Sacc., in Sacc. & Berlese, Atti Ist. Veneto Sci., Ser. 6,<br />
3: 727. 1885.<br />
In vitro: Colonies with entire margin; aerial mycelium rather<br />
compact, raised, velvety, olivaceous-grey; reverse olivaceousblack.<br />
Submerged hyphae verrucose, hyaline, thin-walled, 1–3 µm<br />
wide; aerial hyphae subhyaline, later becoming dark brown, thickwalled,<br />
smooth. Conidiophores arising vertically from creeping<br />
hyphae, straight or flexuose, up to 260 µm long, dark brown at<br />
the base, paler towards the apex, thick-walled; in the upper part<br />
bearing short branches. Conidiogenous cells terminally integrated,<br />
polyblastic, smooth or verrucose, subcylindrical, mostly not or barely<br />
geniculate-sinuous, variable in length, subhyaline, later becoming<br />
pale brown, fertile part as wide as the basal part, proliferating<br />
sympodially, sometimes becoming septate <strong>and</strong> forming a short,<br />
straight rachis with pigmented, slightly thickened <strong>and</strong> hardly<br />
prominent, more or less flat scars. Conidia solitary, occasionally in<br />
short chains, 0–multi-septate, subhyaline to rather pale olivaceous<br />
or olivaceous-brown, smooth to verrucose, globose, ellipsoidal to<br />
obovoid or obclavate, with a slightly darkened <strong>and</strong> thickened hilum;<br />
conidial secession schizolytic.<br />
Type species: P. velutina (G. Winter) Sacc., Miscell. mycol. 2: 17.<br />
1884.<br />
62
Ramichloridium <strong>and</strong> allied genera<br />
54<br />
10 changes<br />
Fig. 1. (Continued).<br />
52<br />
76<br />
100 100<br />
98<br />
94<br />
71<br />
100<br />
77<br />
99<br />
90<br />
98<br />
Radulidium sp. <strong>CBS</strong> 115704<br />
Pseudovirgaria hyperparasitica CPC 10702<br />
Pseudovirgaria hyperparasitica CPC 10753<br />
Pseudovirgaria hyperparasitica CPC 10704<br />
96 Radulidium subulatum <strong>CBS</strong> 287.84<br />
99 Radulidium subulatum <strong>CBS</strong> 912.96<br />
Radulidium epichloës <strong>CBS</strong> 361.63<br />
78<br />
Radulidium epichloës AF050287<br />
64<br />
Radulidium subulatum <strong>CBS</strong> 405.76<br />
Radulidium subulatum <strong>CBS</strong> 101010<br />
Sporidesmium pachyanthicola DQ408557<br />
Staninwardia suttonii DQ923535<br />
Ramichloridium brasilianum <strong>CBS</strong> 283.92<br />
Batcheloromyces proteae <strong>CBS</strong> 110696<br />
Teratosphaeria alistairii DQ885901<br />
Teratosphaeria toledana DQ246230<br />
Readeriella considenianae DQ923527<br />
Teratosphaeria molleriana EU019292<br />
Teratosphaeria fibrillosa EU019282<br />
Catenulostroma macowanii EU019254<br />
67<br />
58 100<br />
100<br />
87<br />
100<br />
56<br />
52<br />
99 54<br />
99<br />
87<br />
Teratosphaeria suberosa DQ246235<br />
Cibiessia dimorphospora EU019258<br />
Teratosphaeria readeriellophora DQ246238<br />
Xenomeris juniperi EF114709<br />
Catenulostroma abietis EU019249<br />
Devriesia staurophora DQ008150<br />
Teratosphaeria parva DQ246240<br />
69<br />
78<br />
64<br />
Teratosphaeria parva DQ246243<br />
<strong>Cladosporium</strong> cladosporioides EU019262<br />
<strong>Cladosporium</strong> uredinicola EU019264<br />
<strong>Cladosporium</strong> bruhnei EU019261<br />
<strong>Cladosporium</strong> sphaerospermum EU019263<br />
Ramichloridium indicum <strong>CBS</strong> 171.96<br />
Dissoconium aciculare EU019266<br />
“Mycosphaerella” communis EU019267<br />
“Mycosphaerella” lateralis EU019268<br />
Ramichloridium apiculatum <strong>CBS</strong> 390.67<br />
Ramichloridium apiculatum <strong>CBS</strong> 400.76<br />
Ramichloridium apiculatum <strong>CBS</strong> 391.67<br />
Ramichloridium apiculatum <strong>CBS</strong> 156.59<br />
100<br />
100<br />
95<br />
99<br />
56<br />
55<br />
80<br />
Ramichloridium pini <strong>CBS</strong> 461.82<br />
Mycosphaerella endophytica DQ246252<br />
Mycosphaerella endophytica DQ246255<br />
Mycosphaerella gregaria DQ246251<br />
Mycosphaerella graminicola EU019297<br />
Septoria tritici EU019298<br />
Cercosporella centaureicola EU019257<br />
Mycosphaerella punctiformis AY490776<br />
Ramularia sp. EU019285<br />
Ramularia miae DQ885902<br />
Ramularia pratensis var. pratensis EU019284<br />
Mycosphaerella walkeri DQ267574<br />
Mycosphaerella parkii DQ246245<br />
Mycosphaerella madeirae DQ204756<br />
Periconiella levispora <strong>CBS</strong> 873.73<br />
Mycosphaerella marksii DQ246249<br />
Pseudocercospora epispermogonia DQ204758<br />
Pseudocercospora epispermogonia DQ204757<br />
Mycosphaerella intermedia DQ246248<br />
Mycosphaerella marksii DQ246250<br />
Periconiella arcuata <strong>CBS</strong> 113477<br />
Rasutoria pseudotsugae EF114704<br />
Rasutoria tsugae EF114705<br />
Periconiella velutina <strong>CBS</strong> 101950<br />
Periconiella velutina <strong>CBS</strong> 101948<br />
Periconiella velutina <strong>CBS</strong> 101949<br />
Ramichloridium biverticillatum <strong>CBS</strong> 335.36<br />
Ramichloridium musae <strong>CBS</strong> 365.36<br />
Ramichloridium musae <strong>CBS</strong> 190.63<br />
56<br />
56<br />
77<br />
66<br />
97<br />
Ramichloridium australiense <strong>CBS</strong> 121710<br />
Ramichloridium strelitziae <strong>CBS</strong> 121711<br />
Zasmidium cellare <strong>CBS</strong> 146.36<br />
Ramichloridium cerophilum <strong>CBS</strong> 103.59<br />
Ramichloridium cerophilum AF050286<br />
Incertae sedis<br />
Teratosphaeriaceae<br />
Davidiellaceae<br />
Mycosphaerellaceae<br />
Capnodiales, Dothideomycetes<br />
Notes: Periconiella is distinct from other ramichloridium-like genera<br />
by its conidiophores that are prominently branched in the upper<br />
part, <strong>and</strong> by its darkened, thickened conidial scars, that are more<br />
or less flat <strong>and</strong> non-prominent. Although conidiophores are also<br />
branched in the upper part in Thysanorea Arzanlou, W. Gams &<br />
Crous, the branching pattern in the latter <strong>genus</strong> is different from<br />
that of Periconiella. Thysanorea has a complex head consisting of<br />
up to six levels of branches, while in Periconiella the branching is<br />
limited, with mainly primary <strong>and</strong> secondary branches. Furthermore,<br />
Thysanorea is characterised by having dimorphic conidiophores<br />
<strong>and</strong> more or less prominent denticle-like conidiogenous loci.<br />
www.studiesinmycology.org<br />
63
Arzanlou et al.<br />
Athelia epiphylla AY586633<br />
Paullicorticium ansatum AY586693<br />
Veronaeopsis simplex <strong>CBS</strong> 588.66<br />
0.98 Repetophragma goidanichii DQ408574<br />
Metacoleroa dickiei DQ384100<br />
0.79 Venturia pyrina EF114715<br />
1.00 Venturia hanliniana AF050290<br />
Venturia inaequalis EF114713<br />
0.84 Venturia chlorospora DQ384101<br />
Venturia asperata EF114711<br />
1.00<br />
0.1 expected changes per site<br />
0.99<br />
0.52<br />
0.74<br />
1.00<br />
1.00<br />
0.97<br />
1.00<br />
0.80<br />
0.63<br />
0.99<br />
0.50<br />
0.68<br />
0.69<br />
1.00<br />
0.58<br />
Venturia carpophila AY849967<br />
Capronia pulcherrima AF050256<br />
Capronia coronata AF050242<br />
Rhinocladiella phaeophora <strong>CBS</strong> 496.78<br />
Rhinocladiella fasciculata <strong>CBS</strong> 132.86<br />
1.00<br />
Exophiala dermatitidis AF050270<br />
Capronia mansonii AY004338<br />
Rhinocladiella mackenziei <strong>CBS</strong> 368.92<br />
Rhinocladiella mackenziei AF050288<br />
Rhinocladiella mackenziei <strong>CBS</strong> 367.92<br />
Rhinocladiella mackenziei <strong>CBS</strong> 102590<br />
1.00<br />
Rhinocladiella anceps AF050284<br />
Rhinocladiella anceps <strong>CBS</strong> 181.65<br />
Rhinocladiella anceps AF050285<br />
Rhinocladiella anceps <strong>CBS</strong> 157.54<br />
Rhinocladiella sp. <strong>CBS</strong> 264.49<br />
Exophiala jeanselmei AF050271<br />
Rhinocladiella atrovirens AF050289<br />
Rhinocladiella basitona <strong>CBS</strong> 101460<br />
Fonsecaea pedrosoi AF050276<br />
Thysanorea papuana <strong>CBS</strong> 212.96<br />
Veronaea japonica <strong>CBS</strong> 776.83<br />
Veronaea compacta <strong>CBS</strong> 268.75<br />
Veronaea botryosa <strong>CBS</strong> 350.65<br />
Veronaea botryosa <strong>CBS</strong> 121.92<br />
Veronaea botryosa <strong>CBS</strong> 254.57<br />
0.84<br />
0.99<br />
0.96<br />
0.99<br />
0.91<br />
1.00<br />
1.00 Conioscyphascus varius AY484512<br />
Conioscypha lignicola AY484513<br />
1.00 Carpoligna pleurothecii AF064646<br />
1.00<br />
Carpoligna pleurothecii AF064645<br />
Carpoligna pleurothecii AY544685<br />
0.99 1.00 Pleurothecium obovoideum <strong>CBS</strong> 209.95<br />
1.00 Ascotaiwania hughesii AY316357<br />
Ascotaiwania hughesii AY094189<br />
Magnaporthe grisea AB026819<br />
Ophiostoma stenoceras DQ836904<br />
Cryptadelphia polyseptata AY281102<br />
Cryptadelphia groenendalensis AY281103<br />
1.00<br />
1.00<br />
1.00<br />
0.61<br />
0.87<br />
0.71<br />
0.92<br />
1.00<br />
Venturiaceae,<br />
Pleosporales,<br />
Dothideomycetes<br />
Rhodoveronaea varioseptata <strong>CBS</strong> 431.88<br />
Annulatascus triseptatus AY780049<br />
Annulatascus triseptatus AY346257<br />
Thyridium vestitum AY544671<br />
Myrmecridium schulzeri <strong>CBS</strong> 114996<br />
Myrmecridium schulzeri <strong>CBS</strong> 188.96<br />
Myrmecridium schulzeri <strong>CBS</strong> 381.87<br />
Myrmecridium schulzeri <strong>CBS</strong> 305.73<br />
Myrmecridium schulzeri <strong>CBS</strong> 304.73<br />
Myrmecridium flexuosum <strong>CBS</strong> 398.76<br />
Myrmecridium schulzeri <strong>CBS</strong> 100.54<br />
Myrmecridium schulzeri <strong>CBS</strong> 642.76<br />
Myrmecridium schulzeri <strong>CBS</strong> 134.68<br />
Myrmecridium schulzeri <strong>CBS</strong> 156.63<br />
Myrmecridium schulzeri <strong>CBS</strong> 325.74<br />
Herpotrichiellaceae,<br />
Chaetothyriales,<br />
Chaetothyriomycetes<br />
Sordariomycetes<br />
Fig. 2. (Page 64–65). Consensus phylogram (50 % majority rule) of 1 500 trees resulting from a Bayesian analysis of the LSU sequence alignment using MrBayes v. 3.1.2.<br />
Bayesian posterior probabilities are indicated at the nodes. Ex-type sequences are printed in bold face. <strong>The</strong> tree was rooted to two sequences obtained from GenBank (Athelia<br />
epiphylla AY586633 <strong>and</strong> Paullicorticium ansatum AY586693).<br />
Periconiella velutina (G. Winter) Sacc., Miscell. mycol. 2: 17.<br />
1884. Fig. 3.<br />
Basionym: Periconia velutina G. Winter, Hedwigia 23: 174. 1884.<br />
In vitro: Submerged hyphae verrucose, hyaline, thin-walled, 1–3<br />
µm wide; aerial hyphae subhyaline, later becoming dark brown,<br />
thick-walled, smooth. Conidiophores arising vertically from creeping<br />
hyphae, straight or flexuose, up to 260 µm long, dark brown at the<br />
base, paler towards the apex, thick-walled; in the upper part bearing<br />
short branches, 10–35 µm long. Conidiogenous cells mostly<br />
terminally integrated, sometimes discrete, smooth or verrucose,<br />
cylindrical, variable in length, subhyaline, later becoming pale<br />
brown, fertile part as wide as the basal part, proliferating sympodially,<br />
sometimes becoming septate <strong>and</strong> forming a short, straight rachis<br />
with pigmented, slightly thickened <strong>and</strong> hardly prominent, more<br />
or less flat scars, less than 1 µm diam. Conidia 0(–1)-septate,<br />
subhyaline, thin-walled, verrucose or smooth, globose, ellipsoidal<br />
to obovoid, (7–)8–9(–11) × (2.5–)3(–4) µm, with a slightly darkened<br />
<strong>and</strong> thickened hilum, 1.5–2 µm diam.<br />
Cultural characteristics: Colonies on MEA slow-growing, reaching<br />
4 mm diam after 14 d at 24 °C, with entire margin; aerial mycelium<br />
rather compact, raised, velvety, olivaceous-grey; reverse<br />
olivaceous-black.<br />
Specimens examined: South Africa, Cape Town, on Brabejum stellatifolium, P.<br />
MacOwan, herb. G. Winter (B), lectotype selected here; Cape Town, on leaves<br />
of Brabejum stellatifolium (= B. stellatum), P. Mac-Owan, PAD, F42165, F462166,<br />
isolectotypes; Stellenbosch, Jonkershoek Nature Reserve, on Brabejum<br />
stellatifolium, 21 Jan. 1999, J.E. Taylor, epitype designated here <strong>CBS</strong> H-15612,<br />
cultures ex-epitype <strong>CBS</strong> 101948–101950.<br />
64
Ramichloridium <strong>and</strong> allied genera<br />
0.1 expected changes per site<br />
Fig. 2. (Continued).<br />
0.64<br />
0.61<br />
0.96<br />
0.99<br />
0.87<br />
1.00<br />
1.00<br />
1.00<br />
0.68<br />
0.99<br />
1.00<br />
1.00<br />
1.00<br />
1.00<br />
0.99<br />
0.86<br />
Pseudovirgaria hyperparasitica CPC 10702<br />
Pseudovirgaria hyperparasitica CPC 10753<br />
Pseudovirgaria hyperparasitica CPC 10704<br />
Radulidium sp. <strong>CBS</strong> 115704<br />
0.95 Radulidium subulatum <strong>CBS</strong> 287.84<br />
Radulidium subulatum <strong>CBS</strong> 912.96<br />
Radulidium epichloës <strong>CBS</strong> 361.63<br />
Radulidium epichloës AF050287<br />
Radulidium subulatum <strong>CBS</strong> 405.76<br />
Radulidium subulatum <strong>CBS</strong> 101010<br />
0.60<br />
0.57<br />
<strong>Cladosporium</strong> cladosporioides EU019262<br />
0.91<br />
<strong>Cladosporium</strong> uredinicola EU019264<br />
1.00 <strong>Cladosporium</strong> bruhnei EU019261<br />
0.81 <strong>Cladosporium</strong> sphaerospermum EU019263<br />
1.00 Sporidesmium pachyanthicola DQ408557<br />
Staninwardia suttonii DQ923535<br />
1.00 Ramichloridium brasilianum <strong>CBS</strong> 283.92<br />
1.00 Batcheloromyces proteae EU019247<br />
Teratosphaeria alistairii DQ885901<br />
Readeriella considenianae DQ923527<br />
Teratosphaeria toledana DQ246230<br />
Teratosphaeria molleriana EU019292<br />
Teratosphaeria fibrillosa EU019282<br />
Catenulostroma macowanii EU019254<br />
0.63<br />
0.73<br />
0.72<br />
0.95<br />
1.00<br />
1.00<br />
1.00<br />
1.00<br />
0.64<br />
1.00<br />
1.00<br />
Teratosphaeria suberosa DQ246235<br />
Cibiessia dimorphospora EU019258<br />
Teratosphaeria readeriellophora DQ246238<br />
Teratosphaeria parva DQ246240<br />
Teratosphaeria parva DQ246243<br />
Devriesia staurophora DQ008150<br />
Xenomeris juniperi EF114709<br />
Catenulostroma abietis EU019249<br />
Ramichloridium indicum <strong>CBS</strong> 171.96<br />
Dissoconium aciculare EU019266<br />
“Mycosphaerella” communis EU019267<br />
“Mycosphaerella” lateralis EU019268<br />
Ramichloridium apiculatum <strong>CBS</strong> 390.67<br />
Ramichloridium apiculatum <strong>CBS</strong> 400.76<br />
Ramichloridium apiculatum <strong>CBS</strong> 391.67<br />
Ramichloridium apiculatum <strong>CBS</strong> 156.59<br />
0.99<br />
0.72<br />
1.00<br />
Mycosphaerella endophytica DQ246252<br />
Mycosphaerella endophytica DQ246255<br />
Mycosphaerella gregaria DQ246251<br />
Cercosporella centaureicola EU019257<br />
Mycosphaerella punctiformis AY490776<br />
Ramularia sp. EU019285<br />
Ramularia miae DQ885902<br />
Ramularia pratensis var. pratensis EU019284<br />
Mycosphaerella graminicola EU019297<br />
Septoria tritici EU019298<br />
Ramichloridium pini <strong>CBS</strong> 461.82<br />
1.00 Mycosphaerella walkeri DQ267574<br />
Mycosphaerella parkii DQ246245<br />
Mycosphaerella madeirae DQ204756<br />
0.95 Periconiella levispora <strong>CBS</strong> 873.73<br />
0.76<br />
0.60<br />
0.97<br />
0.64<br />
0.80<br />
1.00<br />
0.90<br />
0.95<br />
0.97<br />
1.00<br />
0.64<br />
0.99<br />
0.94<br />
0.97<br />
0.57<br />
Mycosphaerella marksii DQ246249<br />
Pseudocercospora epispermogonia DQ204758<br />
Pseudocercospora epispermogonia DQ204757<br />
Mycosphaerella intermedia DQ246248<br />
Mycosphaerella marksii DQ246250<br />
Periconiella arcuata <strong>CBS</strong> 113477<br />
Rasutoria pseudotsugae EF114704<br />
Rasutoria tsugae EF114705<br />
Periconiella velutina <strong>CBS</strong> 101950<br />
Periconiella velutina <strong>CBS</strong> 101948<br />
Periconiella velutina <strong>CBS</strong> 101949<br />
0.96<br />
0.97<br />
0.97<br />
0.96<br />
Ramichloridium biverticillatum <strong>CBS</strong> 335.36<br />
Ramichloridium musae <strong>CBS</strong> 365.36<br />
Ramichloridium musae <strong>CBS</strong> 190.63<br />
Ramichloridium australiense <strong>CBS</strong> 121710<br />
Ramichloridium strelitziae <strong>CBS</strong> 121711<br />
Zasmidium cellare <strong>CBS</strong> 146.36<br />
Ramichloridium cerophilum <strong>CBS</strong> 103.59<br />
Ramichloridium cerophilum AF050286<br />
Incertae sedis<br />
Davidiellaceae<br />
Teratosphaeriaceae<br />
Mycosphaerellaceae<br />
Capnodiales, Dothideomycetes<br />
Periconiella arcuata Arzanlou, S. Lee & Crous, sp. nov. MycoBank<br />
MB504547. Figs 4, 7A.<br />
Etymology: Named after its curved conidia.<br />
Ab aliis speciebus Periconiellae conidiis obclavatis, rectis vel curvatis, (30–)53–61(–<br />
79) × (3–)5(–7) µm, distinguenda.<br />
Submerged hyphae smooth, hyaline, thin-walled, 2 µm wide;<br />
aerial hyphae pale brown, smooth or verrucose, slightly narrower.<br />
Conidiophores arising vertically from creeping hyphae, straight<br />
or flexuose, up to 300 µm long, dark brown at the base, paler<br />
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towards the apex, thick-walled; loosely branched in the upper part,<br />
bearing short branches. Conidiogenous cells integrated, cylindrical,<br />
variable in length, 20–50 µm long, subhyaline, later becoming<br />
pale brown, fertile part as wide as the basal part, proliferating<br />
sympodially, forming a geniculate conidium-bearing rachis with<br />
pigmented <strong>and</strong> thickened, prominent, cone-shaped scars, 1 µm<br />
diam. Conidia formed singly, obclavate, straight or mostly curved,<br />
0(–4)-septate, coarsely verrucose, pale olive, thin-walled, tapering<br />
towards the apex, (30–)53–61(–79) × (3–)5(–7) µm, with a narrowly<br />
truncate base <strong>and</strong> a darkened, hardly thickened hilum, 2 µm diam;<br />
microcyclic conidiation observed in culture.<br />
65
Arzanlou et al.<br />
Fig. 3. Periconiella velutina (<strong>CBS</strong> 101948). A–B. Macronematous conidiophores with short branches in the upper part. C. Sympodially proliferating conidiogenous cell with<br />
darkened <strong>and</strong> slightly thickened scars. D. Conidia. Scale bar = 10 µm.<br />
Fig. 4. Periconiella arcuata (<strong>CBS</strong> 113477). A–B. Sympodially proliferating conidiogenous cells with darkened, thickened <strong>and</strong> cone-shaped scars. C–E. Macronematous<br />
conidiophores with loose branches in the upper part. F–I. Conidia. Scale bar = 10 µm.<br />
66
Ramichloridium <strong>and</strong> allied genera<br />
Fig. 5. Periconiella levispora (<strong>CBS</strong> 873.73). A–C. Conidial apparatus at different stages of development, which gives rise to macronematous conidiophores with dense branches<br />
in the upper part. D. Sympodially proliferating conidiogenous cells with darkened <strong>and</strong> somewhat protruding scars. E–F. Conidia with truncate base <strong>and</strong> darkened hilum. Scale<br />
bar = 10 µm.<br />
Fig. 6. A. Pseudovirgaria hyperparasitica (<strong>CBS</strong> 121739 = CPC 10753). B. Periconiella levispora (<strong>CBS</strong> 873.73). Scale bar = 10 µm.<br />
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67
Arzanlou et al.<br />
Cultural characteristics: Colonies on MEA reaching 12 mm diam<br />
after 14 d at 24 °C, with entire, smooth, sharp margin; mycelium<br />
compacted, becoming hairy, colonies up to 1 mm high; surface<br />
olivaceous to olivaceous-grey, reverse dark grey-olivaceous to<br />
olivaceous-black.<br />
Specimen examined: South Africa, Western Cape Province, Kogelberg, on dead<br />
culms of Ischyrolepis subverticillata, May 2001, S. Lee, holotype <strong>CBS</strong> H-19927,<br />
culture ex-type <strong>CBS</strong> 113477.<br />
Periconiella levispora Arzanlou, W. Gams & Crous, sp. nov.<br />
MycoBank MB504546. Figs 5–6B.<br />
Etymology: (Latin) levis = smooth.<br />
A simili Periconiella velutina conidiis levibus et maioribus, ad 23 μm longis<br />
distinguenda.<br />
In vitro: Submerged hyphae smooth, hyaline, thin-walled, 2–2.5 µm<br />
wide; aerial hyphae subhyaline, later becoming dark brown, thickwalled,<br />
smooth. Conidiophores arising vertically from creeping<br />
aerial hyphae, dark brown at the base, paler towards the apex,<br />
thick-walled; in the upper part bearing several short branches,<br />
up to 120 µm long. Conidiogenous cells integrated, occasionally<br />
discrete, cylindrical, variable in length, 10–20 µm long, subhyaline,<br />
later becoming pale brown, fertile part as wide as the basal part,<br />
proliferating sympodially, forming a short rachis with pigmented<br />
<strong>and</strong> slightly thickened, somewhat protruding scars, less than 1<br />
µm diam. Conidia solitary, 0(–2)-septate, smooth, pale olivaceous,<br />
cylindrical, ellipsoidal, pyriform to clavate, (7–)11–14(–23) × (3–)4–<br />
5(–6) µm, with a truncate base <strong>and</strong> a darkened, slightly thickened<br />
hilum, 2 µm diam.<br />
Cultural characteristics: Colonies on MEA slow-growing, reaching<br />
5 mm diam after 14 d at 24 °C, with entire margin; aerial mycelium<br />
compact, raised, velvety, olivaceous-grey; reverse olivaceousblack.<br />
Basionym: Chloridium apiculatum J.H. Mill., Giddens & A.A. Foster,<br />
Mycologia 49: 789. 1957.<br />
≡ Veronaea apiculata (J.H. Mill., Giddens & A.A. Foster) M.B. Ellis, in Ellis,<br />
More Dematiaceous Hyphomycetes: 209. 1976.<br />
[non Rhinocladiella apiculata Matsush., in Matsushima, Icon.<br />
Microfung. Mats. lect.: 122. 1975].<br />
= Rhinocladiella indica Agarwal, Lloydia 32: 388. 1969.<br />
[non Chloridium indicum Subram., Proc. Indian Acad. Sci., Sect. B,<br />
42: 286. 1955].<br />
In vitro: Submerged hyphae hyaline to subhyaline, thin-walled,<br />
1–2.5 µm wide; aerial hyphae slightly darker, smooth-walled.<br />
Conidiophores generally arising at right angles from creeping aerial<br />
hyphae, straight, unbranched, thick-walled, dark brown, continuous<br />
or with 1–2(–3) additional thin septa, up to 100 µm long; intercalary<br />
cells 10–28 µm long. Conidiogenous cells integrated, terminal,<br />
smooth, thick-walled, golden-brown, straight, cylindrical, 25–37(–<br />
47) × 2–3.5 µm; proliferating sympodially, resulting in a straight<br />
rachis with conspicuous conidiogenous loci; scars prominent,<br />
crowded, slightly pigmented, less than 1 µm diam. Conidia solitary,<br />
obovate to obconical, pale brown, finely verrucose, (3–)5–5.5(–7.5)<br />
× (2–)2.5–3(–4) µm, hilum conspicuous, slightly pigmented, about<br />
1 µm diam.<br />
Cultural characteristics: Colonies on MEA reaching 35 mm diam<br />
after 14 d at 24 °C; minimum temperature for growth above 6 °C,<br />
optimum 24 °C, maximum 30 °C. Colonies raised, velvety, dense,<br />
with entire margin; surface olivaceous-green, reverse olivaceousblack,<br />
often with a diffusing citron-yellow pigment.<br />
Specimens examined: Pakistan, Lahore, from soil, A. Kamal, <strong>CBS</strong> 400.76 = IMI<br />
088021. South Africa, from preserved Cucumis sativus in 8-oxyquinoline sulphate,<br />
M.C. Papendorf, <strong>CBS</strong> 390.67; Potchefstroom, from Aloe sp., M.C. Papendorf, <strong>CBS</strong><br />
391.67. U.S.A., Georgia, isolated from forest soil, <strong>CBS</strong> 156.59 = ATCC 13211 = IMI<br />
100716 = QM 7716, ex-type culture.<br />
Specimen examined: Sri Lanka, Hakgala Botanic Gardens, on dead leaves of<br />
Turpinia pomifera, Jan. 1973, W. Gams, holotype <strong>CBS</strong> H-15611, culture ex-type<br />
<strong>CBS</strong> 873.73.<br />
Ramichloridium Stahel ex de Hoog, Stud. Mycol. 15: 59. 1977.<br />
In vitro: Colonies flat to raised, with entire margin; surface<br />
olivaceous-green to olivaceous-black. Mycelium consisting of<br />
submerged <strong>and</strong> aerial hyphae; submerged hyphae hyaline to<br />
subhyaline, thin-walled, aerial hyphae smooth or verrucose.<br />
Conidiophores straight, unbranched, rarely branched, thick-walled,<br />
dark brown (darker than the subtending hyphae), continuous or<br />
with several additional thin septa. Conidiogenous cells integrated,<br />
terminal, polyblastic, smooth, thick-walled, golden-brown, apical<br />
part subhyaline, with sympodial proliferation, straight or flexuose,<br />
geniculate or nodose, with conspicuous conidiogenous loci;<br />
scars crowded or scattered, unthickened, unpigmented to faintly<br />
pigmented, or slightly prominent denticles. Conidia solitary, 0–1-<br />
septate, subhyaline to pale brown, smooth to coarsely verrucose,<br />
rather thin-walled, obovate, obconical or globose to ellipsoidal,<br />
fusiform, with a somewhat prominent, slightly pigmented hilum;<br />
conidial secession schizolytic.<br />
Type species: R. apiculatum (J.H. Mill., Giddens & A.A. Foster) de<br />
Hoog, Stud. Mycol. 15: 69. 1977.<br />
Ramichloridium apiculatum (J.H. Mill., Giddens & A.A. Foster) de<br />
Hoog, Stud. Mycol. 15: 69. 1977. Fig. 8.<br />
Fig. 7. A. Periconiella arcuata (<strong>CBS</strong> 113477). B. Myrmecridium schulzeri (<strong>CBS</strong><br />
325.74). C. Thysanorea papuana (<strong>CBS</strong> 212.96). Scale bars = 10 µm.<br />
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Ramichloridium <strong>and</strong> allied genera<br />
Fig. 8. Ramichloridium apiculatum (<strong>CBS</strong> 156.59). A–C. Macronematous conidiophores with sympodially proliferating conidiogenous cells, which give rise to a conidium-bearing<br />
rachis with crowded <strong>and</strong> prominent scars. D. Conidia. Scale bar = 10 µm.<br />
Fig. 9. Ramichloridium australiense (<strong>CBS</strong> 121710). A–C. Macronematous conidiophores with thick-walled <strong>and</strong> warted subtending hyphae. D. Sympodially proliferating<br />
conidiogenous cell, which results in a short rachis with darkened <strong>and</strong> slightly thickened scars. E. Conidia. Scale bar = 10 µm.<br />
Ramichloridium australiense Arzanlou & Crous, sp. nov.<br />
MycoBank MB504548. Figs 9–10A.<br />
Etymology: Named after its country of origin, Australia.<br />
Ab aliis speciebus Ramichloridii conidiophoris ex hyphis verrucosis, crassitunicatis<br />
ortis distinguendum.<br />
In vitro: Submerged hyphae hyaline, smooth, thin-walled, 1–2 µm<br />
wide; aerial hyphae pale brown, warted. Conidiophores arising<br />
vertically <strong>and</strong> clearly differentiated from creeping aerial hyphae, up<br />
to 400 µm tall, with several additional thin septa; intercalary cells,<br />
8–40 × 2–5 µm, from the broadest part at the base tapering towards<br />
the apex, subhyaline, later becoming pale brown <strong>and</strong> warted in the<br />
lower part. Subtending hyphae thick-walled, warted. Conidiogenous<br />
cells integrated, terminal, 10–18 µm long, proliferating sympodially,<br />
www.studiesinmycology.org<br />
giving rise to a short rachis with conspicuous conidiogenous loci;<br />
scars slightly thickened <strong>and</strong> darkened, about 1 µm diam. Conidia<br />
solitary, aseptate, thin-walled, smooth, subhyaline, subcylindrical<br />
to obclavate, (10–)12–15(–23) × 2.5–3 µm, with a truncate base<br />
<strong>and</strong> a slightly darkened <strong>and</strong> thickened hilum,1.5–2 µm diam, rarely<br />
fusing at the basal part.<br />
Cultural characteristics: Colonies on MEA rather slow growing,<br />
reaching 8 mm diam after 14 d at 24 °C, with entire, smooth<br />
margin; mycelium flat, olivaceous-grey, becoming granular, with<br />
gelatinous droplets at the margin developing with aging; reverse<br />
pale olivaceous-grey.<br />
Specimen examined: Australia, Queensl<strong>and</strong>, Mount Lewis, Mount Lewis Road,<br />
16°34’47.2” S, 145°19’7” E, 538 m alt., on Musa banksii leaf, Aug. 2006, P.W. Crous<br />
<strong>and</strong> B. Summerell, holotype <strong>CBS</strong> H-19928, culture ex-type <strong>CBS</strong> 121710.<br />
69
Arzanlou et al.<br />
Fig. 10. A. Ramichloridium australiense (<strong>CBS</strong> 121710). B. Ramichloridium brasilianum (<strong>CBS</strong> 283.92). C. Radulidium subulatum (<strong>CBS</strong> 405.76). D. Rhodoveronaea varioseptata<br />
(<strong>CBS</strong> 431.88). Scale bar = 10 µm.<br />
Fig. 11. Ramichloridium musae (<strong>CBS</strong> 365.36). A. Conidiophores with loose branches. B–D. Sympodially proliferating conidiogenous cells, resulting in a long conidium-bearing<br />
rachis. E. Rachis with hardly prominent, slightly darkened scars. F. Conidia. Scale bars = 10 µm.<br />
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Ramichloridium <strong>and</strong> allied genera<br />
Fig. 12. Ramichloridium biverticillatum (<strong>CBS</strong> 335.36). A–B. Profusely branched <strong>and</strong> biverticillate conidiophores. C. Sympodially proliferating conidiogenous cells,<br />
which give rise to a conidium-bearing rachis with crowded, slightly pigmented <strong>and</strong> thickened scars. D. Conidia. Scale bar = 10 µm.<br />
Fig. 13. Ramichloridium brasilianum (<strong>CBS</strong> 283.92). A–B. Macronematous conidiophores with sympodially proliferating conidiogenous cells, resulting in a conidiumbearing<br />
rachis. C. Rachis with crowded <strong>and</strong> slightly pigmented scars. D. Conidia. Scale bar = 10 µm.<br />
Fig. 14. Ramichloridium cerophilum (<strong>CBS</strong> 103.59). A–C. Conidial apparatus at different stages of development, resulting in macronematous conidiophores <strong>and</strong><br />
sympodially proliferating conidiogenous cells. D–E. Formation of secondary conidia. F. Conidia. Scale bar = 10 µm.<br />
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71
Arzanlou et al.<br />
Ramichloridium musae (Stahel ex M.B. Ellis) de Hoog, Stud.<br />
Mycol. 15: 62. 1977. Fig. 11.<br />
Basionym: Veronaea musae Stahel ex M.B. Ellis, in Ellis, More<br />
Dematiaceous Hyphomycetes: 209. 1976.<br />
≡ Chloridium musae Stahel, Trop. Agric., Trinidad 14: 43. 1937 (nom.<br />
inval. Art. 36).<br />
Misapplied name: Chloridium indicum Subram., sensu Batista &<br />
Vital, Anais Soc. Biol. Pernambuco 15: 379. 1957.<br />
In vitro: Submerged hyphae smooth, hyaline, thin-walled, 1–2 µm<br />
wide; aerial hyphae subhyaline, smooth. Conidiophores arising<br />
vertically <strong>and</strong> mostly sharply differentiated from creeping aerial<br />
hyphae, golden-brown; unbranched, rarely branched in the upper<br />
part, up to 250 µm tall, with up to 6 additional thin septa, cells<br />
23–40 × 2–2.5 µm, basal cell occasionally inflated. Conidiogenous<br />
cells terminally integrated, cylindrical, variable in length, 10–40 µm<br />
long, golden-brown near the base, subhyaline to pale brown near<br />
the end, fertile part as wide as the basal part, later also becoming<br />
septate; rachis elongating sympodially, 2–2.5 µm wide, with hardly<br />
prominent, scattered, slightly pigmented scars, about 0.5 µm diam.<br />
Conidia solitary, aseptate, hyaline to subhyaline, ellipsoidal, (4–)7–<br />
8(–12) × 2–3 µm, smooth or verruculose, thin-walled, with slightly<br />
darkened hilum, about 1 µm diam.<br />
Cultural characteristics: Colonies on MEA slow-growing, reaching<br />
27 mm diam after 14 d at 24 °C, with entire, smooth, sharp margin;<br />
mycelium mostly submerged, some floccose to lanose aerial<br />
mycelium in the olivaceous-grey centre, becoming pale pinkish<br />
olivaceous towards the margin; reverse pale orange.<br />
Specimens examined: Cameroon, from Musa sapientum, J.E. Heron, <strong>CBS</strong> 169.61<br />
= ATCC 15681 = IMI 079492 = DAOM 84655 = MUCL 2689; from Musa sapientum,<br />
J. Brun, <strong>CBS</strong> 190.63 = MUCL 9557. Surinam, Paramaribo, from Musa sapientum<br />
leaf, G. Stahel, <strong>CBS</strong> 365.36 = JCM 6973 = MUCL 9556, ex-type strain of Chloridium<br />
musae; from Musa sapientum, G. Stahel, <strong>CBS</strong> 365.36; dried culture preserved as<br />
<strong>CBS</strong> H-19933.<br />
Ramichloridium biverticillatum Arzanlou & Crous, nom. nov.<br />
MycoBank MB504549. Fig. 12.<br />
Basionym: Periconiella musae Stahel ex M.B. Ellis, Mycol. Pap.<br />
111: 5. 1967.<br />
[non Ramichloridium musae (Stahel ex M.B. Ellis) de Hoog, 1977].<br />
≡ Ramichloridium musae Stahel, Trop. Agric., Trinidad 14: 43. 1937 (nom.<br />
inval. Art. 36).<br />
= Ramichloridium musae (Stahel ex M.B. Ellis) de Hoog, Stud. Mycol. 15: 62.<br />
1977, sensu de Hoog, p.p.<br />
Etymology: Named after its biverticillate conidiophores.<br />
In vitro: Submerged hyphae smooth, hyaline, thin-walled, 1–2<br />
µm wide; aerial hyphae subhyaline, smooth, slightly darker.<br />
Conidiophores arising vertically from creeping aerial hyphae, pale<br />
brown, profusely branched, biverticillate, with up to three levels<br />
of main branches; branches tapering distally, 2–3 µm wide at the<br />
base, approx. 2 µm wide in the upper part, up to 250 µm long.<br />
Conidiogenous cells terminally integrated, cylindrical, variable in<br />
length, 15–50 µm long, rachis straight or geniculate, pale brown,<br />
as wide as the basal part; elongating sympodially, forming a rachis<br />
with crowded, slightly darkened <strong>and</strong> thickened minute scars, less<br />
than 0.5 µm wide. Conidia solitary, aseptate, hyaline to subhyaline,<br />
dacryoid to pyriform, (2–)3–4(–6) × (1.5–)2(–2.5) µm, smooth, thinwalled,<br />
with an inconspicuous hilum.<br />
Cultural characteristics: Colonies on MEA slow-growing, reaching<br />
16 mm diam after 14 d at 24 °C, with entire, smooth, sharp margin,<br />
rather compact, velvety; surface vinaceous-buff to olivaceous-buff;<br />
reverse buff.<br />
Specimen examined: Surinam, from Musa sapientum, Aug. 1936, G. Stahel, <strong>CBS</strong><br />
335.36.<br />
Notes: Ramichloridium biverticillatum is a new name based on<br />
Periconiella musae. <strong>The</strong> species is distinct from R. musae because<br />
of its profusely branched conidiophores, <strong>and</strong> conidia that are smaller<br />
(2–5 × 1.5–2.5 µm) than those of R. musae (5–11 × 2–3 µm).<br />
Ramichloridium brasilianum Arzanlou & Crous, sp. nov.<br />
MycoBank MB504550. Figs 10B, 13.<br />
Etymology: Named after its country of origin, Brazil.<br />
A simili Ramichloridio cerophilo conidiis minoribus, ad 8 μm longis, et conidiis<br />
secundariis absentibus distinguendum.<br />
In vitro: Submerged hyphae pale olivaceous, smooth or slightly<br />
rough, 1.5–2 µm wide; aerial hyphae olivaceous, smooth or rough,<br />
narrower <strong>and</strong> darker than the submerged hyphae. Conidiophores<br />
unbranched, arising vertically from creeping aerial hyphae, straight<br />
or flexuose, dark brown, with up to 10 additional septa, thick-walled,<br />
cylindrical, 2–2.5 µm wide <strong>and</strong> up to 70 µm long. Conidiogenous<br />
cells integrated, terminal, 10–30 µm long, proliferating sympodially,<br />
giving rise to a long, straight rachis with crowded, slightly darkened<br />
minute scars, about 0.5 µm diam. Conidia solitary, obovoid<br />
to fusiform with the widest part below the middle, thin-walled,<br />
verruculose, aseptate, pale brown, slightly rounded at the apex,<br />
truncate at the base, (4–)5–6(–8.5) × 2–2.5(–3) µm, with a slightly<br />
thickened <strong>and</strong> darkened hilum, 1–1.5 µm diam.<br />
Cultural characteristics: Colonies on MEA slow-growing, reaching<br />
6 mm diam after 14 d at 24 °C, velvety to hairy, colonies with entire<br />
margin, surface dark olivaceous-grey; black gelatinous exudate<br />
droplets produced on OA.<br />
Specimen examined: Brazil, São Paulo, Peruibe, Jureia Ecological Reserve, forest<br />
soil, Jan. 1991, D. Attili, holotype <strong>CBS</strong> H-19929, culture ex-type <strong>CBS</strong> 283.92.<br />
Ramichloridium cerophilum (Tubaki) de Hoog, Stud. Mycol. 15:<br />
74. 1977. Fig. 14.<br />
Basionym: Acrotheca cerophila Tubaki, J. Hattori Bot. Lab. 20: 143.<br />
1958.<br />
≡ <strong>Cladosporium</strong> cerophilum (Tubaki) Matsush., in Matsushima, Icon.<br />
Microfung. Matsush. lect. (Kobe): 34. 1975.<br />
In vitro: Submerged hyphae pale olivaceous-brown, smooth or<br />
slightly rough, 1.5–3 µm wide; aerial hyphae olivaceous-brown,<br />
smooth or slightly rough, somewhat narrower <strong>and</strong> darker than the<br />
submerged hyphae. Conidiophores unbranched, arising vertically<br />
from creeping aerial hyphae, dark brown, thick-walled, smooth or<br />
verruculose, hardly tapering towards the apex, 2–3 µm wide, up<br />
to 50 µm long, with up to 3 additional septa. Conidiogenous cells<br />
integrated, terminal, proliferating sympodially, rachis short <strong>and</strong><br />
straight, with crowded, prominent, pigmented unthickened scars,<br />
minute, approx. 0.5 µm diam. Conidia solitary, fusiform to clavate,<br />
thin-walled, smooth, 0(–1)-septate, subhyaline, (4–)6–7(–11) × (2–)<br />
2.5(–3) µm, with a conspicuous hilum, about 0.5 µm diam, slightly<br />
raised with an inconspicuous marginal frill, somehow resembling<br />
those of <strong>Cladosporium</strong>. Conidia sometimes producing 1–3(–4)<br />
short secondary conidia.<br />
Cultural characteristics: Colonies on MEA rather slow-growing,<br />
reaching 12 mm diam after 14 d at 24 °C, velvety to hairy, with<br />
entire margin; surface dark olivaceous-grey, with black gelatinous<br />
exudate droplets on OA.<br />
Specimen examined: Japan, isolated from Sasa sp., K. Tubaki, <strong>CBS</strong> 103.59, extype.<br />
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Ramichloridium <strong>and</strong> allied genera<br />
Fig. 15. Ramichloridium indicum (<strong>CBS</strong> 171.96). A–B. Macronematous conidiophores. C–E. Sympodially proliferating conidiogenous cells, resulting in a conidium-bearing rachis<br />
with pigmented <strong>and</strong> thickened scars. F. Conidia. Scale bar = 10 µm.<br />
Fig. 16. Ramichloridium strelitziae (<strong>CBS</strong> 121711). A–C. Conidial apparatus at different stages of development, resulting in macronematous conidiophores <strong>and</strong> sympodially<br />
proliferating conidiogenous cells. D–E. Rachis with crowded, slightly pigmented, thickened, circular scars. F. Conidia. Scale bars = 10 µm.<br />
Notes: Phylogenetically, this species together with Ramichloridium<br />
apiculatum <strong>and</strong> R. musae cluster within the Mycosphaerellaceae<br />
clade. Ramichloridium cerophilum can be distinguished from its<br />
relatives by the production of secondary conidia <strong>and</strong> its distinct<br />
conidial hila.<br />
Ramichloridium indicum (Subram.) de Hoog, Stud. Mycol. 15:<br />
70. 1977. Fig. 15.<br />
Basionym: Chloridium indicum Subram., Proc. Indian Acad. Sci.,<br />
Sect. B, 42: 286. 1955 [non Rhinocladiella indica Agarwal, Lloydia<br />
32: 388. 1969].<br />
≡ Veronaea indica (Subram.) M.B. Ellis, in Ellis, More Dematiaceous<br />
Hyphomycetes: 209. 1976.<br />
= Veronaea verrucosa Geeson, Trans. Brit. Mycol. Soc. 64: 349. 1975.<br />
In vitro: Submerged hyphae smooth, thin-walled, hyaline, 1–2.5<br />
µm wide, with thin septa; aerial hyphae coarsely verrucose,<br />
olivaceous-green, rather thick-walled, 2–2.5 µm wide, with thin<br />
septa. Conidiophores arising vertically from creeping hyphae at right<br />
angles, straight, unbranched, thick-walled, smooth, dark brown,<br />
with up to 10 thin septa, up to 250 µm long, 2–4 µm wide, often<br />
with inflated basal cells. Conidiogenous cells terminally integrated,<br />
up to 165 µm long, smooth, dark brown, sympodially proliferating,<br />
rachis straight or flexuose, geniculate or nodose, subhyaline; scars<br />
thickened <strong>and</strong> darkened, clustered at nodes, approx. 0.5 µm diam.<br />
Microcyclic conidiation observed in culture. Conidia solitary, (0–)1-<br />
septate, not constricted at the septum, subhyaline to pale brown,<br />
smooth or coarsely verrucose, rather thin-walled, broadly ellipsoidal<br />
to globose, (5–)7–8(–10) × (4–)6–6.5(–9) µm, with truncate base;<br />
hilum conspicuous, slightly darkened, not thickened, about 1 µm<br />
diam.<br />
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73
Arzanlou et al.<br />
Cultural characteristics: Colonies on MEA reaching 35 mm diam<br />
after 14 d at 24 °C. Colonies velvety, rather compact, slightly<br />
elevated, with entire, smooth, whitish margin, dark olivaceousgreen<br />
in the central part.<br />
Specimen examined: Living culture, Feb. 1996, L. Marvanová, <strong>CBS</strong> 171.96.<br />
Ramichloridium pini de Hoog & Rahman, Trans. Brit. Mycol. Soc.<br />
81: 485. 1983.<br />
Specimen examined: U.K., Scotl<strong>and</strong>, Old Aberdeen, branch of Pinus contorta<br />
(Pinaceae), 1982, M.A. Rahman, ex-type strain, <strong>CBS</strong> 461.82 = MUCL 28942.<br />
Note: <strong>The</strong> culture examined (<strong>CBS</strong> 461.82) was sterile. For a full<br />
description see de Hoog et al. (1983).<br />
Ramichloridium strelitziae Arzanlou, W. Gams & Crous, sp. nov.<br />
MycoBank MB504551. Figs 16–17A.<br />
Etymology: Named after its host, Strelitzia.<br />
Ab aliis speciebus Ramichloridii conidiophoris brevibus, ad 40 μm longis, et<br />
cicatricibus rotundis, paulo protrudentibus distinguendum.<br />
In vitro: Submerged hyphae smooth, hyaline, thin-walled, 2–2.5 µm<br />
wide; aerial hyphae pale brown, verrucose. Conidiophores arising<br />
vertically from creeping aerial hyphae, clearly differentiated from the<br />
vegetative hyphae, subhyaline, later becoming pale brown, thickwalled,<br />
smooth or verruculose, with 1–3 additional septa; up to 40<br />
µm long <strong>and</strong> 2 µm wide. Conidiogenous cells integrated, terminal,<br />
cylindrical, variable in length, 10–35 µm long, subhyaline, later<br />
turning pale brown, fertile part as wide as the basal part, proliferating<br />
sympodially, forming a straight rachis with slightly thickened <strong>and</strong><br />
darkened, circular, somewhat protruding scars, approx. 0.5 µm<br />
diam. Conidia solitary, aseptate, smooth or verruculose, subhyaline,<br />
oblong, ellipsoidal to clavate, (3–)4–5(–5.5) × (1–)2(–2.5) µm, with<br />
truncate base <strong>and</strong> unthickened, non-pigmented hilum.<br />
Cultural characteristics: Colonies on MEA slow-growing, reaching<br />
5 mm diam after 14 d at 24 °C, with entire margin; aerial mycelium<br />
rather compact, raised, dense, olivaceous-grey; reverse olivaceousblack.<br />
Specimen examined: South Africa, KwaZulu-Natal, Durban, near Réunion,<br />
on leaves of Strelitzia nicolai, 5 Feb. 2005, W. Gams & H. Glen, <strong>CBS</strong>-H 19776,<br />
holotype, culture ex-type <strong>CBS</strong> 121711.<br />
Zasmidium Fr., Summa Veg. Sc<strong>and</strong>. 2: 407. 1849.<br />
Fig. 17. A. Ramichloridium strelitziae (<strong>CBS</strong> 121711). B. Veronaea japonica (<strong>CBS</strong><br />
776.83). C. Veronaeopsis simplex (<strong>CBS</strong> 588.66). Scale bar = 10 µm.<br />
In vitro: Submerged hyphae smooth, thin-walled, hyaline, with<br />
thin septa; aerial hyphae coarsely verrucose, olivaceous-green,<br />
thick-walled, with thin septa. Conidiophores not differentiated<br />
from vegetative hyphae, often reduced to conidiogenous<br />
Fig. 18. Zasmidium cellare (<strong>CBS</strong> 146.36). A–D. Micronematous conidiophores with terminal, integrated conidiogenous cells. E. Conidiogenous cell with pigmented, thickened<br />
<strong>and</strong> refractive scars. F–G. Primary <strong>and</strong> secondary conidia. Scale bar = 10 µm.<br />
74
Ramichloridium <strong>and</strong> allied genera<br />
Fig. 19. Rhinocladiella anceps (<strong>CBS</strong> 181.65). A. Macronematous conidiophores. B–D. Conidial apparatus at different stages of development, resulting in semi-micronematous<br />
conidiophores <strong>and</strong> sympodially proliferating conidiogenous cells. E. Conidiogenous loci. F. Conidia. Scale bars = 10 µm.<br />
cells. Conidiogenous cells integrated, predominantly terminal,<br />
sometimes lateral, arising from aerial hyphae, cylindrical, pale<br />
brown; polyblastic, proliferating sympodially producing crowded,<br />
conspicuously pigmented, almost flat, darkened, somewhat<br />
refractive scars. Conidia in short chains, cylindrical to fusiform,<br />
verrucose, obovate to obconical, pale brown, base truncate,<br />
with a conspicuous, slightly pigmented, thickened <strong>and</strong> refractive<br />
hilum. Primary conidia sometimes larger, subhyaline, verrucose<br />
or smooth-walled, 0–4-septate, variable in length, fusiform to<br />
cylindrical; conidial secession schizolytic.<br />
Type species: Zasmidium cellare (Pers. : Fr.) Fr., Summa Veg.<br />
Sc<strong>and</strong>. 2: 407. 1849.<br />
Zasmidium cellare (Pers. : Fr.) Fr., Summa Veg. Sc<strong>and</strong>. 2: 407.<br />
1849. Fig. 18.<br />
Basionym: Racodium cellare Pers., Neues Mag. Bot. 1: 123. 1794.<br />
≡ Antennaria cellaris (Pers. : Fr.) Fr., Syst. Mycol. 3: 229. 1829.<br />
≡ <strong>Cladosporium</strong> cellare (Pers. : Fr.) Sch<strong>and</strong>erl, Zentralbl. Bakteriol., 2.<br />
Abt., 94: 117. 1936.<br />
≡ Rhinocladiella cellaris (Pers. : Fr.) M.B. Ellis, in Ellis, Dematiaceous<br />
Hyphomycetes: 248. 1971.<br />
In vitro: Submerged hyphae smooth, thin-walled, hyaline, 2–3<br />
µm wide, with thin septa; aerial hyphae coarsely verrucose,<br />
olivaceous-green, rather thick-walled, 2–2.5 µm wide, with thin<br />
septa. Conidiophores not differentiated from vegetative hyphae,<br />
often reduced to conidiogenous cells. Conidiogenous cells<br />
integrated, predominantly terminal, sometimes lateral, arising from<br />
aerial hyphae, cylindrical, 20–60 µm long <strong>and</strong> 2–2.5 µm wide, pale<br />
brown, proliferating sympodially producing crowded, conspicuously<br />
pigmented scars that are thickened <strong>and</strong> refractive, about 1 µm diam.<br />
Conidia cylindrical to fusiform, verrucose, obovate to obconical,<br />
pale brown, with truncate base, (6–)9–14(–27) × 2–2.5 µm, with<br />
a conspicuous, slightly pigmented, refractive hilum, approx. 1 µm<br />
diam. Primary conidia sometimes subhyaline, verrucose or smoothwalled,<br />
thin-walled, 0–1(–4)-septate, variable in length, fusiform to<br />
cylindrical.<br />
www.studiesinmycology.org<br />
Cultural characteristics: Colonies reaching 7 mm diam after 14 d<br />
at 24 °C. Colonies velvety, rather compact, slightly elevated with<br />
entire margin; surface dark olivaceous-green in the central part,<br />
margin smooth, whitish.<br />
Specimen examined: Wall in wine cellar, Jun. 1936, H. Sch<strong>and</strong>erl, ATCC 36951<br />
= IFO 4862 = IMI 044943 = LCP 52.402 = LSHB BB274 = MUCL 10089 = <strong>CBS</strong><br />
146.36.<br />
Notes: <strong>The</strong> name Racodium Fr., typified by Ra. rupestre Pers. : Fr.,<br />
has been conserved over the older one by Persoon, with Ra. cellare<br />
as type species. De Hoog (1979) defended the use of Zasmidium in<br />
its place for the well-known wine-cellar fungus.<br />
Morphologically Zasmidium resembles Stenella Syd., <strong>and</strong><br />
both reside in the Capnodiales, though the type of Stenella, S.<br />
araguata Syd., clusters in the Teratosphaeriaceae, <strong>and</strong> the type of<br />
Zasmidium, Z. cellare, in the Mycosphaerellaceae. When accepting<br />
anamorph genera as polyphyletic within an order, preference would<br />
be given to the well-known name Stenella over the less known<br />
Zasmidium, even though the latter name is older. Further studies<br />
are required, however, to clarify if all stenella-like taxa should be<br />
accommodated in a single <strong>genus</strong>, Stenella. If this is indeed the<br />
case, a new combination for Zasmidium cellare will be proposed<br />
in Stenella, <strong>and</strong> the latter <strong>genus</strong> will have to be conserved over<br />
Zasmidium.<br />
Chaetothyriales (Herpotrichiellaceae)<br />
<strong>The</strong> four “Ramichloridium” species residing in the Chaetothyriales<br />
clade do not differ sufficiently in morphology to separate them from<br />
Rhinocladiella (type Rh. atrovirens). Because of the pale brown<br />
conidiophores, conidiogenous cells with crowded, slightly prominent<br />
scars <strong>and</strong> the occasional presence of an Exophiala J.W. Carmich.<br />
synanamorph, Rhinocladiella is a suitable <strong>genus</strong> to accommodate<br />
them. <strong>The</strong>se four species chiefly differ from Ramichloridium in the<br />
morphology of their conidial apparatus, which is clearly differentiated<br />
from the vegetative hyphae. <strong>The</strong> appropriate combinations are<br />
therefore introduced for Ramichloridium anceps, R. mackenziei, R.<br />
fasciculatum <strong>and</strong> R. basitonum.<br />
75
Arzanlou et al.<br />
Fig. 20. Rhinocladiella basitona (<strong>CBS</strong> 101460). A–B. Semi-micronematous conidiophores with verticillate branching pattern. C–D. Sympodially proliferating conidiogenous cells,<br />
giving rise to a long rachis with slightly prominent, truncate conidium-bearing denticles. E. Intercalary conidiogenous cell. F. Conidia. Scale bars = 10 µm.<br />
<strong>The</strong> <strong>genus</strong> Veronaea (type species: V. botryosa) also resides<br />
in the Chaetothyriales clade. Veronaea can be distinguished from<br />
Rhinocladiella by the absence of exophiala-type budding cells <strong>and</strong><br />
its predominantly 1-septate conidia. Furthermore, the conidiogenous<br />
loci in Veronaea are rather flat, barely prominent.<br />
Rhinocladiella Nannf., Svensk Skogsvårdsfören. Tidskr., Häfte 32:<br />
461. 1934.<br />
In vitro: Colonies dark olivaceous-brown, slow-growing, almost<br />
moist. Submerged hyphae hyaline to pale olivaceous, smooth;<br />
aerial hyphae, if present, more darkly pigmented. Exophialatype<br />
budding cells usually present in culture. Conidial apparatus<br />
usually branched, olivaceous-brown, consisting of either slightly<br />
differentiated tips of ascending hyphae or septate, markedly<br />
differentiated conidiophores. Conidiogenous cells intercalary or<br />
terminal, polyblastic, cylindrical to acicular, with a sympodially<br />
proliferating, subdenticulate rachis; scars unthickened, nonpigmented<br />
to somewhat darkened-refractive. Conidia solitary,<br />
hyaline to subhyaline, aseptate, thin-walled, smooth, subglobose,<br />
with a slightly pigmented hilum; conidial secession schizolytic.<br />
Type species: Rh. atrovirens Nannf., Svenska Skogsvårdsfören.<br />
Tidskr. 32: 461. 1934.<br />
Rhinocladiella anceps (Sacc. & Ellis) S. Hughes, Canad. J. Bot.<br />
36: 801. 1958. Fig. 19.<br />
Basionym: Sporotrichum anceps Sacc. & Ellis, Michelia 2: 576.<br />
1882.<br />
= Veronaea parvispora M.B. Ellis, in Ellis, More Dematiaceous Hyphomycetes:<br />
210. 1976.<br />
Misapplied name: Chloridium minus Corda sensu Mangenot, Rev.<br />
Mycol. (Paris) 18: 137. 1953.<br />
In vitro: Submerged hyphae subhyaline, smooth, thick-walled, 2–<br />
2.5 µm wide; aerial hyphae pale brown. Swollen germinating cells<br />
often present on MEA, giving rise to an Exophiala synanamorph.<br />
Conidiophores slightly differentiated from vegetative hyphae, arising<br />
from prostrate aerial hyphae, consisting of either unbranched or<br />
loosely branched stalks, thick-walled, golden to dark-brown, up<br />
to 350 µm tall, which may have up to 15 thin, additional septa,<br />
intercalary cells 9–14 µm long. Conidiogenous cells terminal,<br />
rarely lateral, cylindrical, occasionally intercalary, variable in length,<br />
smooth, golden to dark brown at the base, paler toward the apex,<br />
later becoming inconspicuously septate, fertile part as wide as the<br />
basal part, 15–40 × 1.5–2 µm; with crowded, slightly prominent,<br />
unpigmented, conidium-bearing denticles, about 0.5 µm diam.<br />
Conidia solitary, subhyaline, thin-walled, smooth, subglobose to<br />
ellipsoidal, 2.5–4 × 2–2.5 µm, with a less conspicuous, slightly<br />
darkened hilum, less than 0.5 µm diam.<br />
Cultural characteristics: Colonies on MEA reaching 6–12 mm diam<br />
after 14 d at 24 °C, with entire, smooth, sharp margin; mycelium<br />
powdery, becoming hairy at centre; olivaceous-green to brown,<br />
reverse dark-olivaceous.<br />
Specimens examined: Canada, Ontario, Campbellville, from soil under Thuja plicata,<br />
Apr. 1965, G. L. Barron, <strong>CBS</strong> H-7715 (isoneotype); <strong>CBS</strong> H-7716 (isoneotype); <strong>CBS</strong><br />
H-7717 (isoneotype); <strong>CBS</strong> H-7718 (isoneotype); <strong>CBS</strong> H-7719 (isoneotype), ex-type<br />
strain, <strong>CBS</strong> 181.65 = ATCC 18655 = DAOM 84422 = IMI 134453 = MUCL 8233 =<br />
OAC 10215. France, from stem of Fagus sylvatica, 1953, F. Mangenot, <strong>CBS</strong> 157.54<br />
= ATCC 15680= MUCL 1081= MUCL 7992 = MUCL 15756.<br />
Notes: Rhinocladiella anceps (conidia 2.5–4 µm long) resembles<br />
Rh. phaeophora Veerkamp & W. Gams (1983) (conidia 5.5–6 µm<br />
long), but has shorter conidia.<br />
76
Ramichloridium <strong>and</strong> allied genera<br />
Fig. 21. Rhinocladiella fasciculata (<strong>CBS</strong> 132.86). A. Conidiophores. B. Sympodially proliferating conidiogenous cells, which give rise to a long rachis with slightly prominent,<br />
unthickened scars. C. Conidia. D–E. Synanamorph consisting of conidiogenous cells with percurrent proliferation. F. Conidia. Scale bars = 10 µm.<br />
Rhinocladiella basitona (de Hoog) Arzanlou & Crous, comb. nov.<br />
MycoBank MB504552. Fig. 20.<br />
Basionym: Ramichloridium basitonum de Hoog, J. Clin. Microbiol.<br />
41: 4774. 2003.<br />
In vitro: Submerged hyphae hyaline, smooth, thin-walled, 2 µm<br />
wide; aerial hyphae rather thick-walled, pale brown. Conidiophores<br />
slightly differentiated from vegetative hyphae, profusely <strong>and</strong> mostly<br />
verticillately branched, straight or flexuose, pale-brown, 2–2.5 µm<br />
wide. Conidiogenous cells terminal, variable in length, 10–100 µm<br />
long, pale brown, straight or geniculate, proliferating sympodially,<br />
giving rise to a long, 2–2.5 µm wide rachis, with slightly prominent,<br />
truncate conidium-bearing denticles, slightly darkened. Conidia<br />
solitary, hyaline, thin-walled, smooth, pyriform to clavate, with a<br />
round apex, <strong>and</strong> slightly truncate base, (1–)3–4(–5) × 1–2 µm,<br />
hilum conspicuous, slightly darkened <strong>and</strong> thickened, less than 0.5<br />
µm diam.<br />
Cultural characteristics: Colonies on MEA reaching 19 mm diam<br />
after 14 d at 24 °C, with entire, smooth, sharp margin; mycelium<br />
rather flat <strong>and</strong> slightly elevated in the centre, pale olivaceous-grey<br />
to olivaceous-grey; reverse olivaceous-black.<br />
Specimen examined: Japan, Hamamatsu, from subcutaneous lesion with fistula on<br />
knee of 70-year-old male, Y. Suzuki, ex-type culture <strong>CBS</strong> 101460 = IFM 47593.<br />
Rhinocladiella fasciculata (V. Rao & de Hoog) Arzanlou & Crous,<br />
comb. nov. MycoBank MB504553. Fig. 21.<br />
Basionym: Ramichloridium fasciculatum V. Rao & de Hoog, Stud.<br />
Mycol. 28: 39. 1986.<br />
www.studiesinmycology.org<br />
In vitro: Submerged hyphae subhyaline, smooth, thick-walled,<br />
2–2.5 µm wide; aerial hyphae pale brown. Conidiophores arising<br />
vertically from ascending hyphae in loose fascicles, unbranched or<br />
loosely branched at acute angles, cylindrical, smooth, brown <strong>and</strong><br />
thick-walled at the base, up to 220 µm long <strong>and</strong> 2–3 µm wide, with<br />
0–5 thin additional septa. Conidiogenous cells terminal, cylindrical,<br />
30–100 µm long, thin-walled, smooth, pale brown, fertile part as<br />
wide as the basal part, up to 2 µm wide, proliferating sympodially,<br />
giving rise to a rachis with hardly prominent, slightly pigmented,<br />
not thickened scars, less than 0.5 µm diam. Conidia solitary,<br />
smooth, thin-walled, subhyaline, ellipsoidal, (2.5–)4–5(–6) × 2–3<br />
µm, with truncate, slightly pigmented hilum, about 0.5 µm diam.<br />
Synanamorph forming on torulose hyphae originating from giant<br />
cells; compact heads of densely branched hyphae forming thinwalled,<br />
lateral, subglobose cells, on which conidiogenous cells are<br />
formed; conidiogenous cells proliferating percurrently, giving rise to<br />
tubular annellated zones with inconspicuous annellations, up to 12<br />
µm long, 1–1.5 µm wide. Conidia smooth, thin-walled, aseptate,<br />
subhyaline, globose, 2–2.5 µm diam.<br />
Cultural characteristics: Colonies on MEA reaching 8 mm diam after<br />
14 d at 24 °C, with entire, smooth, sharp margin; mycelium velvety,<br />
becoming farinose in the centre due to abundant sporulation,<br />
olivaceous-green to brown, reverse dark olivaceous. Blackish<br />
droplets often produced at the centre, which contain masses of<br />
Exophiala conidia.<br />
Specimen examined: India, Karnataka, Thirathahalli, isolated by V. Rao from<br />
decayed wood, holotype <strong>CBS</strong>-H 3866, culture ex-type <strong>CBS</strong> 132.86.<br />
77
Arzanlou et al.<br />
Fig. 22. Rhinocladiella mackenziei (<strong>CBS</strong> 368.92). A. Intercalary conidiogenous cell. B–E. Semi-micronematous conidiophores <strong>and</strong> sympodially proliferating conidiogenous cells,<br />
resulting in a rachis with slightly prominent, unthickened scars. F. Conidia. Scale bar = 10 µm.<br />
78
Ramichloridium <strong>and</strong> allied genera<br />
Fig. 24. Thysanorea papuana (<strong>CBS</strong> 212.96), periconiella-like synanamorph. A. Macronematous conidiophores. B–C. Conidiophores with dense apical branches. D. Branches<br />
with different levels of branchlets. E–I. Conidiogenous cells at different stages of development; sympodially proliferating conidiogenous cells give rise to a denticulate rachis.<br />
J–K. Conidia. Scale bars = 10 µm.<br />
Fig. 23. (Page 78). Thysanorea papuana (<strong>CBS</strong> 212.96). A. Intercalary conidiogenous cell. B–I. Semi-micronematous conidiophores <strong>and</strong> sympodially proliferating conidiogenous<br />
cells, resulting in a rachis with prominent conidium bearing denticles. J–K. Microcyclic conidiation observed in slide cultures. L. Conidia. Scale bar = 10 µm.<br />
www.studiesinmycology.org<br />
79
Arzanlou et al.<br />
Rhinocladiella mackenziei (C.K. Campb. & Al-Hedaithy) Arzanlou<br />
& Crous, comb. nov. MycoBank MB504554. Fig. 22.<br />
Basionym: Ramichloridium mackenziei C.K. Campb. & Al-Hedaithy,<br />
J. Med. Veterin. Mycol. 31: 330. 1993.<br />
In vitro: Submerged hyphae subhyaline, smooth, thin-walled, 2–3<br />
µm wide; aerial hyphae pale brown, slightly narrower. Conidiophores<br />
slightly or not differentiated from vegetative hyphae, arising<br />
laterally from aerial hyphae, with one or two additional septa, often<br />
reduced to a discrete or intercalary conidiogenous cell, pale-brown,<br />
10–25 × 2.5–3.5 µm. Conidiogenous cells terminal or intercalary,<br />
variable in length, 5–15 µm long <strong>and</strong> 3–5 µm wide, occasionally<br />
slightly wider than the basal part, pale brown, rachis with slightly<br />
prominent, unpigmented, non-thickened scars, about 0.5 µm diam.<br />
Conidia golden-brown, thin-walled, smooth, ellipsoidal to obovate,<br />
subcylindical, (5–)8–9(–12) × (2–)3–3.5(–5) µm, with darkened,<br />
inconspicously thickened, protuberant or truncate hilum, less than<br />
1 µm diam.<br />
Cultural characteristics: Colonies on MEA reaching 5 mm diam<br />
after 14 d at 24 °C, with entire, smooth, sharp margin; mycelium<br />
densely lanose <strong>and</strong> elevated in the centre, olivaceous-green to<br />
brown; reverse dark olivaceous.<br />
Specimens examined: Israel, Haifa, isolated from brain abscess, <strong>CBS</strong> 368.92 =<br />
UTMB 3170; human brain abscess, E. Lefler, <strong>CBS</strong> 367.92 = NCPF 2738 = UTMB<br />
3169. Saudi Arabia, from phaeohyphomycosis of the brain, S.S.A. Al-Hedaithy,<br />
ex-type strain, <strong>CBS</strong> 650.93 = MUCL 40057 = NCPF 2808; from brain abscess,<br />
Pakistani male who travelled to Saudi Arabia, <strong>CBS</strong> 102592 = NCPF 7460. United<br />
Arab Emirates, from fatal brain abscess, <strong>CBS</strong> 102590 = NCPF 2853.<br />
Notes: Morphologically Rhinocladiella mackenziei is somewhat<br />
<strong>similar</strong> to Pleurothecium obovoideum (Matsush.) Arzanlou &<br />
Crous, which was originally isolated from dead wood. However, P.<br />
obovoideum has distinct conidiophores, <strong>and</strong> the ascending hyphae<br />
are thick-walled, <strong>and</strong> the denticles cylindrical, up to 1.5 µm long.<br />
In contrast, Rh. mackenziei has only slightly prominent denticles.<br />
Rhinocladiella mackenziei is a member of the Chaetothyriales,<br />
while P. obovoideum clusters in the Chaetosphaeriales.<br />
Thysanorea Arzanlou, W. Gams & Crous, gen. nov. MycoBank<br />
MB504555.<br />
Etymology: (Greek) thysano = brush, referring to the brush-like<br />
branching pattern, suffix derived from Veronaea.<br />
Veronaeae similis sed conidiophoris partim Periconiae similibus dense ramosis<br />
distinguenda.<br />
In vitro: Submerged hyphae subhyaline, smooth, thin-walled; aerial<br />
hyphae pale brown, smooth or verrucose. Conidiophores dimorphic;<br />
micronematous conidiophores slightly differentiated from vegetative<br />
hyphae, branched or simple, multiseptate. Conidiogenous cells<br />
terminal, polyblastic, variable in length, smooth, golden- to dark<br />
brown at the base, paler towards the apex, later sometimes<br />
inconspicuously septate; fertile part often wider than the basal<br />
part, clavate to doliiform, with crowded, more or less prominent<br />
conidium-bearing denticles, unpigmented, but slightly thickened.<br />
Macronematous conidiophores consisting of well-differentiated,<br />
thick-walled, dark brown stalks; apically repeatedly densely<br />
branched, forming a complex head, each branchlet giving rise<br />
to a conidium-bearing denticulate rachis with slightly pigmented,<br />
thickened scars. Conidia of both kinds of conidiophore formed<br />
singly, smooth, pale brown, obovoidal to pyriform, (0–)1-septate,<br />
with a truncate base <strong>and</strong> darkened hilum; conidial secession<br />
schizolytic.<br />
Type species: Thysanorea papuana (Aptroot) Arzanlou, W. Gams<br />
& Crous, comb. nov.<br />
Thysanorea papuana (Aptroot) Arzanlou, W. Gams & Crous,<br />
comb. nov. MycoBank MB504556. Figs 7C, 23–24.<br />
Basionym: Periconiella papuana Aptroot, Nova Hedwigia 67: 491.<br />
1998.<br />
In vitro: Submerged hyphae subhyaline, smooth, thin-walled, 1.5–3<br />
µm wide; aerial hyphae pale brown, smooth to verrucose, 1.5–2<br />
µm wide. Conidiophores dimorphic; micronematous conidiophores<br />
slightly differentiated from vegetative hyphae, branched or simple,<br />
up to 6-septate. Conidiogenous cells terminal or intercalary, variable<br />
in length, 5–20 µm long, thin-walled, smooth, golden- to dark brown<br />
at the base, paler toward the apex, later sometimes becoming<br />
inconspicuously septate, fertile part wider than basal part, often<br />
clavate, with crowded, more or less prominent conidium-bearing<br />
denticles, about 1 µm diam, unpigmented but slightly thickened.<br />
Conidia solitary, subhyaline, thin-walled, smooth, cylindrical to<br />
pyriform, rounded at the apex <strong>and</strong> truncate at the base, pale brown,<br />
(0–)1-septate, (5–)7–8(–11) × (2–)3(–4) µm, with a truncate base<br />
<strong>and</strong> darkened hilum, 1 µm diam. Macronematous conidiophores<br />
present in old cultures after 1 mo of incubation, consisting of welldifferentiated,<br />
thick-walled, dark brown stalks, up to 220 µm long,<br />
(4–)5–6(–7) µm wide, with up to 15 additional septa, often with<br />
inflated basal cells; apically densely branched, forming a complex<br />
head, with up to five levels of branchlets, 20–50 µm long, each<br />
branchlet giving rise to a denticulate conidium-bearing rachis; scars<br />
slightly pigmented, thickened, about 1 µm diam. Conidia solitary,<br />
thin-walled, smooth, pale brown, obovoidal to pyriform, (0–)1-<br />
septate, (4–)5–6(–8) × (2–)3(–4) µm, with a truncate base <strong>and</strong><br />
darkened hilum, 1–2 µm diam.<br />
Cultural characteristics: Colonies on MEA reaching 10 mm diam<br />
after 14 d at 24 °C, with entire, sharp margin; mycelium velvety,<br />
elevated, with colonies up to 2 mm high, surface olivaceous-grey to<br />
iron-grey; reverse greenish black.<br />
Specimen examined: Papua New Guinea, Madang Province, foothill of Finisterre<br />
range, 40.8 km along road Madang-Lae, alt. 200 m, isolated from unknown stipe, 2<br />
Nov. 1995, A. Aptroot, holotype <strong>CBS</strong>-H 6351, culture ex-type <strong>CBS</strong> 212.96.<br />
Veronaea Cif. & Montemart., Atti Ist. Bot. Lab. Crittog. Univ. Pavia,<br />
sér. 5, 15: 68. 1957.<br />
In vitro: Colonies velvety, pale olivaceous-brown, moderately<br />
fast-growing. Submerged hyphae hyaline to pale olivaceous,<br />
smooth; aerial hyphae, more darkly pigmented. Exophiala-type<br />
budding cells absent in culture. Conidiophores erect, straight or<br />
flexuose, unbranched or occasionally loosely branched, sometimes<br />
geniculate, smooth-walled, pale to medium- or olivaceous-brown.<br />
Conidiogenous cells terminally integrated, polyblastic, occasionally<br />
intercalary, cylindrical, pale brown, later often becoming septate,<br />
fertile part subhyaline, often as wide as the basal part, rachis with<br />
crowded, flat to slightly prominent, faintly pigmented, unthickened<br />
scars. Conidia solitary, smooth, cylindrical to pyriform, rounded<br />
at the apex <strong>and</strong> truncate at the base, pale brown, 1(–2)-septate;<br />
conidial secession schizolytic.<br />
Type species: Veronaea botryosa Cif. & Montemart., Atti Ist. Bot.<br />
Lab. Crittog. Univ. Pavia, sér. 5, 15: 68. 1957.<br />
Veronaea botryosa Cif. & Montemart., Atti Ist. Bot. Lab. Crittog.<br />
Univ. Pavia, sér. 5, 15: 68. 1957. Fig. 25.<br />
80
Ramichloridium <strong>and</strong> allied genera<br />
Fig. 25. Veronaea botryosa (<strong>CBS</strong> 254.57). A–C. Semi-micronematous conidiophores <strong>and</strong> sympodially proliferating conidiogenous cells. D–E. Rachis with crowded <strong>and</strong> flat<br />
scars. F–G. Microcyclic conidiation. H. Conidia. Scale bars = 10 µm.<br />
In vitro: Submerged hyphae hyaline to pale olivaceous, smooth;<br />
aerial hyphae more darkly pigmented. Conidiophores erect,<br />
straight or flexuose, unbranched or occasionally loosely branched,<br />
sometimes geniculate, smooth-walled, pale brown to olivaceousbrown,<br />
2–3 µm wide <strong>and</strong> up to 200 µm long. Conidiogenous cells<br />
terminal, occasionally intercalary, cylindrical, 10–100 µm long,<br />
pale brown, later often becoming septate, fertile part subhyaline,<br />
often as wide as the basal part, rachis with crowded, flat to slightly<br />
prominent, faintly pigmented, unthickened scars. Conidia solitary,<br />
smooth, cylindrical to pyriform, (3–)6.5–8.5(–12) × (1.5–)2–2.5(–3)<br />
µm, rounded at the apex <strong>and</strong> truncate at the base, pale brown,<br />
1(–2)-septate, with a faintly darkened, unthickened hilum, about 0.5<br />
µm diam.<br />
Cultural characteristics: Colonies on MEA reaching 30 mm diam<br />
after 14 d at 24 °C, with entire, sharp margin; mycelium velvety,<br />
slightly elevated in the centre, surface olivaceous-grey to greyishbrown;<br />
reverse greenish black.<br />
Specimens examined: India, Ramgarh, about 38 km from Jaipur, isolated from goat<br />
dung, 1 Sep. 1963, B.C. Lodha, <strong>CBS</strong> 350.65 = IMI 115127 = MUCL 7972. Italy,<br />
Tuscany, Pisa, isolated from Sansa olive slag, 1954, O. Verona, ex-type strain, <strong>CBS</strong><br />
254.57 = IMI 070233 = MUCL 9821.<br />
Veronaea compacta Papendorf, Bothalia 12: 119. 1976. Fig. 26.<br />
In vitro: Submerged hyphae subhyaline, smooth, thin-walled,<br />
1.5–3 µm wide; aerial hyphae rather thick-walled, pale brown.<br />
Conidiophores slightly differentiated from vegetative hyphae,<br />
lateral or occasionally terminal, often wider than the supporting<br />
hypha, up to 4 µm wide, unbranched or branched at acute angles,<br />
with 1–3 adititional septa, cells often inflated <strong>and</strong> flask-shaped,<br />
pale-brown, up to 60 µm long. Conidiogenous cells terminal,<br />
occasionally intercalary, variable in length, up to 10 µm long,<br />
pale brown, cylindrical to doliiform or flask-shaped, with hardly<br />
prominent denticles; scars flat, slightly pigmented, not thickened,<br />
about 0.5 µm diam. Conidia solitary, pale brown, smooth, thinwalled,<br />
ellipsoidal to ovoid, 0–1(–2)-septate, often constricted at<br />
the septa, (4–)6–7(–9) × 2–3 µm, with a round apex <strong>and</strong> truncate<br />
base; hilum prominent, slightly darkened, unthickened, about 0.5<br />
µm diam.<br />
Cultural characteristics: Colonies rather slow growing, reaching 15<br />
mm diam on MEA after 14 d at 24 °C; surface velvety to lanose,<br />
slightly raised in the centre, pale grey to pale brownish grey; reverse<br />
dark grey.<br />
Specimen examined: South Africa, soil, M.C. Papendorf, ex-type culture <strong>CBS</strong><br />
268.75.<br />
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81
Arzanlou et al.<br />
Fig. 26. Veronaea compacta (<strong>CBS</strong> 268.75). A–B. Semi-micronematous conidiophores <strong>and</strong> sympodially proliferating conidiogenous cells. C–D. Rachis with hardly prominent<br />
denticles. E. Conidia. Scale bar = 10 µm.<br />
Fig. 27. Veronaea japonica (<strong>CBS</strong> 776.83). A. Intercalary conidiogenous cells. B–D. Semi-micronematous conidiophores <strong>and</strong> sympodially proliferating conidiogenous cells. E.<br />
Conidia. F. Thick-walled, dark brown hyphal cells. Scale bar = 10 µm.<br />
Veronaea japonica Arzanlou, W. Gams & Crous, sp. nov.<br />
MycoBank MB504557. Figs 17B, 27.<br />
Etymology: Named after the country of origin, Japan.<br />
Veronaeae compactae similis, sed cellulis inflatis, aggregatis, crassitunicatis, fuscis<br />
in vitro formatis distinguenda.<br />
In vitro: Submerged hyphae subhyaline, smooth, thin-walled, 1.5–3<br />
µm wide; aerial hyphae slightly narrower, pale brown; hyphal cells<br />
later becoming swollen, thick-walled, dark brown, often aggregated.<br />
Conidiophores slightly differentiated from aerial vegetative hyphae,<br />
lateral, or terminal, often wider than the supporting hypha, 2–3 µm<br />
wide, up to 65 µm long, unbranched or occasionally branched,<br />
82
Ramichloridium <strong>and</strong> allied genera<br />
Fig. 28. Pleurothecium obovoideum (<strong>CBS</strong> 209.95). A–C. Conidial apparatus<br />
consisting of conidiophores with sympodially proliferating conidiogenous cells as<br />
seen in slide cultures of ca. 14 d. D. Short chain of conidia. E–G. Sympodially<br />
proliferating conidiogenous cells, resulting in a short rachis with subcylindrical to<br />
cylindrical denticles. H. Conidia. Scale bar = 10 µm.<br />
pale brown, thin-walled, smooth, with 1–3 additional septa.<br />
Conidiogenous cells terminal, occasionally intercalary, variable in<br />
length, up to 15 µm long, pale brown, cylindrical to clavate, with<br />
hardly prominent denticles; scars flat, slightly pigmented, not<br />
thickened, about 0.5 µm diam. Conidia solitary, pale brown, smooth,<br />
thin-walled, ellipsoidal to ovoid, (0–)1-septate, often constricted at<br />
the septum, (6–)7–8(–10) × 2–2.5(–4) µm, with a round apex <strong>and</strong><br />
truncate base; hilum unthickened but slightly darkened, about 1 µm<br />
diam.<br />
Cultural characteristics: Colonies rather slow growing, reaching 7.5<br />
mm diam on MEA after 14 d at 24 °C; surface velvety to lanose,<br />
slightly raised in the centre, olivaceous-brown, with entire margin;<br />
reverse dark-olivaceous.<br />
Specimen examined: Japan, Kyoto, Daitokuji Temple, Kyoto, inside dead bamboo<br />
culm, Dec. 1983, W. Gams, holotype <strong>CBS</strong>-H 3490, ex-type culture <strong>CBS</strong> 776.83.<br />
Note: This species is morphologically <strong>similar</strong> to V. compacta<br />
(Papendorf 1976), but can be distinguished based on the presence<br />
of dark brown, swollen hyphal cells in culture, which are absent in<br />
V. compacta.<br />
Pleurothecium obovoideum clade (Chaetosphaeriales)<br />
Ramichloridium obovoideum was regarded as <strong>similar</strong> to<br />
“Ramichloridium” (Rhinocladiella) mackenziei by some authors,<br />
<strong>and</strong> subsequently reduced to synonymy (Ur-Rahman et al. 1988).<br />
However, R. obovoideum clusters with Carpoligna pleurothecii,<br />
the teleomorph of Pleurothecium Höhn. Because it is also<br />
morphologically <strong>similar</strong> to other species of Pleurothecium, we<br />
herewith combine it into that <strong>genus</strong>.<br />
Pleurothecium obovoideum (Matsush.) Arzanlou & Crous, comb.<br />
nov. MycoBank MB504558. Fig. 28.<br />
Basionym: Rhinocladiella obovoidea Matsush., Icones Microfung.<br />
Mats. lect.: 123. 1975.<br />
≡ Ramichloridium obovoideum (Matsush.) de Hoog, Stud. Mycol. 15: 73.<br />
1977.<br />
In vitro: Submerged hyphae smooth, hyaline, thin-walled, 1–2 µm<br />
wide; aerial hyphae hyaline to subhyaline, smooth. Conidiophores<br />
arising vertically from creeping hyphae, ascending hyphae thickwalled<br />
<strong>and</strong> dark brown; conidiophores 10–35 µm long, 1–2-septate,<br />
often reduced to a conidiogenous cell, unbranched, thick-walled,<br />
smooth, tapering towards the apex, pale brown. Conidiogenous<br />
cells integrated, cylindrical to ampulliform, 5–20 µm long, pale<br />
brown, elongating sympodially, with a short rachis giving rise to<br />
denticles, 1 µm long, slightly pigmented. Conidia aseptate, solitary<br />
or in short chains of up to 3, smooth, pale brown, ellipsoidal to<br />
obovate, (9–)11–12(–14.5) × (3–)4(–5) µm, smooth, thin-walled,<br />
with a more or less rounded apex, a truncate base <strong>and</strong> a slightly<br />
darkened, unthickened hilum, 1.5 µm diam.<br />
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83
Arzanlou et al.<br />
Cultural characteristics: Colonies slow-growing, reaching 15 diam<br />
after 14 d at 24 °C, with entire, smooth margin; surface rather<br />
compact, mycelium mainly flat, submerged, some floccose to<br />
lanose aerial mycelium in the centre, buff; reverse honey.<br />
Specimen examined: Japan, Kobe Municipal Arboretum, T. Matsushima, from dead<br />
leaf of Pasania edulis, <strong>CBS</strong> 209.95 = MFC 12477.<br />
Incertae sedis (Sordariomycetes)<br />
Ramichloridium schulzeri clade<br />
Ramichloridium schulzeri, including its varieties, clusters<br />
near Thyridium Nitschke <strong>and</strong> the Magnaporthaceae, <strong>and</strong> is<br />
phylogenetically as well as morphologically distinct from the other<br />
genera in the Ramichloridium complex. To accommodate these<br />
taxa, a new <strong>genus</strong> is introduced below.<br />
Myrmecridium Arzanlou, W. Gams & Crous, gen. nov. MycoBank<br />
MB504559.<br />
Etymology: (Greek) myrmekia = wart, referring to the wart-like<br />
denticles on the rachis, suffix -ridium from Chloridium.<br />
Genus ab allis generibus Ramichloridii similibus rachide recta longa, subhyalina,<br />
denticulis distantibus, verruciformibus praedita distinguendum.<br />
In vitro: Colonies moderately fast-growing, flat, with mainly<br />
submerged mycelium, <strong>and</strong> entire margin, later becoming powdery<br />
to velvety, pale orange to orange. Mycelium rather compact, mainly<br />
submerged, in the centre velvety with fertile bundles of hyphae.<br />
Conidiophores arising vertically <strong>and</strong> clearly distinct from creeping<br />
hyphae, unbranched, straight or flexuose, brown, thick-walled.<br />
Conidiogenous cells terminally integrated, polyblastic, cylindrical,<br />
straight or flexuose, pale brown, sometimes secondarily septate,<br />
fertile part subhyaline, as wide as the basal part, with scattered<br />
pimple-shaped, apically pointed, unpigmented, conidium-bearing<br />
denticles. Conidia solitary, subhyaline, smooth or finely verrucose,<br />
rather thin-walled, with a wing-like gelatinous sheath, obovoidal or<br />
fusiform, tapering towards a narrowly truncate base with a slightly<br />
prominent, unpigmented hilum; conidial secession schizolytic.<br />
Type species: Myrmecridium schulzeri (Sacc.) Arzanlou, W. Gams<br />
& Crous, comb. nov.<br />
Notes: Myrmecridium schulzeri was fully described as Acrotheca<br />
acuta Grove by Hughes (1951). <strong>The</strong> author discussed several<br />
genera, none of which is suitable for the present fungus for various<br />
reasons as analysed by de Hoog (1977). Only Gomphinaria Preuss<br />
is not yet sufficiently documented. Our examination of G. amoena<br />
Preuss (B!) showed that this is an entirely different fungus, of which<br />
no fresh material is available to ascertain its position.<br />
Myrmecridium can be distinguished from other ramichloridiumlike<br />
fungi by having entirely hyaline vegetative hyphae, <strong>and</strong> widely<br />
scattered, pimple-shaped denticles on the long hyaline rachis.<br />
<strong>The</strong> conidial sheath is visible in lactic acid mounts with brightfield<br />
microscopy. <strong>The</strong> Myrmecridium clade consists of several<br />
subclusters, which are insufficiently resolved based on the ITS<br />
sequence data. However, two morphologically distinct varieties of<br />
Myrmecridium are treated here. <strong>The</strong> status of the other isolates in<br />
this clade will be dealt with in a future study incorporating more<br />
strains, <strong>and</strong> using a multi-gene phylogenetic approach.<br />
Myrmecridium schulzeri (Sacc.) Arzanlou, W. Gams & Crous,<br />
comb. nov. MycoBank MB504560. var. schulzeri Figs 7B, 29.<br />
Basionym: Psilobotrys schulzeri Sacc., Hedwigia 23: 126. 1884.<br />
≡ Chloridium schulzerii (Sacc.) Sacc., Syll. Fung. 4: 322. 1886.<br />
≡ Rhinocladiella schulzeri (Sacc.) Matsush., Icon. Microfung. Mats. lect.<br />
(Kobe): 124. 1975.<br />
≡ Ramichloridium schulzeri (Sacc.) de Hoog, Stud. Mycol. 15: 64. 1977<br />
var. schulzeri.<br />
= Acrotheca acuta Grove, J. Bot., Lond. 54: 222. 1916.<br />
≡ Pleurophragmium acutum (Grove) M.B. Ellis in Ellis, More Dematiaceous<br />
Hyphomycetes: 165. 1976.<br />
= Rhinotrichum multisporum Doguet, Rev. Mycol., Suppl. Colon. 17: 78. 1953<br />
(nom. inval. Art. 36) [non Acrotheca multispora (Preuss) Sacc., Syll. Fung. 4:<br />
277. 1886].<br />
[non Acrothecium (?) multisporum G. Arnaud, Bull. Trimestriel Soc.<br />
Mycol. France 69: 288. 1953 (nom. inval. Art. 36)].<br />
[non Acrothecium multisporum G. Arnaud sensu Tubaki, J. Hattori<br />
Bot. Lab. 20: 145. 1958].<br />
In vitro: Submerged hyphae hyaline, thin-walled, 1–2 µm wide;<br />
aerial hyphae, if present, pale olivaceous-brown. Conidiophores<br />
arising vertically from creeping aerial hyphae, unbranched, straight,<br />
reddish brown, thick-walled, septate, up to 250 µm tall, 2.5–3.5<br />
µm wide, with 2–7 additional septa, basal cell often inflated, 3.5–5<br />
µm wide. Conidiogenous cells integrated, cylindrical, variable in<br />
length, 15–110 µm long, subhyaline to pale brown, later becoming<br />
inconspicuously septate, fertile part subhyaline, as wide as the<br />
basal part, forming a straight rachis with scattered, pimple-shaped<br />
denticles less than 1 µm long <strong>and</strong> approx. 0.5 µm wide, apically<br />
pointed, unpigmented, slightly thickened scars. Conidia solitary,<br />
subhyaline, thin-walled, smooth or finely verrucose, surrounded<br />
by a wing-like, gelatinous conidial sheath, up to 0.5 µm thick,<br />
ellipsoid, obovoid or fusiform, (6–)9–10(–12) × 3–4 µm, tapering to<br />
a subtruncate base; hilum unpigmented, inconspicuous.<br />
Cultural characteristics: Colonies reaching 29 mm diam after 14<br />
d at 24 °C, pale orange to orange, with entire margin; mycelium<br />
flat, rather compact, later becoming farinose or powdery due to<br />
sporulation, which occurs in concentric zones when incubated on<br />
the laboratory bench.<br />
Specimens examined: Germany, Kiel-Kitzeberg, from wheat-field soil, W. Gams,<br />
<strong>CBS</strong> 134.68 = ATCC 16310. <strong>The</strong> Netherl<strong>and</strong>s, isolated from a man, bronchial<br />
secretion, A. Visser, <strong>CBS</strong> 156.63 = MUCL 1079; Lienden, isolated from Triticum<br />
aestivum root, C.L. de Graaff, <strong>CBS</strong> 325.74 = JCM 7234.<br />
Myrmecridium schulzeri var. tritici (M.B. Ellis) Arzanlou, W.<br />
Gams & Crous, comb. nov. MycoBank MB504562.<br />
Basionym: Pleurophragmium tritici M.B. Ellis, in Ellis, More<br />
Dematiaceous Hyphomycetes: 165. 1976.<br />
≡ Ramichloridium schulzeri var. tritici (M.B. Ellis) de Hoog, Stud. Mycol.<br />
15: 68. 1977.<br />
Specimen examined: Irel<strong>and</strong>, Dublin, on wheat stem, Oct. 1960, J.J. Brady,<br />
holotype IMI 83291.<br />
Notes: No reliable living culture is available of this variety. Based<br />
on a re-examination of the type specimen in this study, the variety<br />
appears sufficiently distinct from Myrmecridium schulzeri var.<br />
schulzeri based on the frequent production of septate conidia.<br />
Myrmecridium flexuosum (de Hoog) Arzanlou, W. Gams & Crous,<br />
comb. et stat. nov. MycoBank MB504563. Fig. 30.<br />
Basionym: Ramichloridium schulzeri var. flexuosum de Hoog, Stud.<br />
Mycol. 15: 67. 1977.<br />
In vitro: Submerged hyphae hyaline, thin-walled, 1–2 µm wide.<br />
Conidiophores unbranched, flexuose, arising from creeping aerial<br />
84
Ramichloridium <strong>and</strong> allied genera<br />
Fig. 29. Myrmecridium schulzeri (<strong>CBS</strong> 325.74). A. Macronematous conidiophores. B. Inflated basal cells visible in some conidiophores. C–E. Conidial apparatus at different<br />
stages of development, resulting in macronematous conidiophores <strong>and</strong> sympodially proliferating conidiogenous cells. F–G. Rachis with scattered, pimple-shaped denticles.<br />
H. Conidia. Scale bars: A =100 µm, B–H = 10 µm.<br />
hyphae, pale brown, up to 250 µm tall, 3–3.5 µm wide, thick-walled,<br />
smooth, with up to 24 thin septa, delimiting 8–12 µm long cells.<br />
Conidiogenous cells integrated, elongating sympodially, cylindrical,<br />
20–150 µm long, flexuose, brown at the base, subhyaline in the<br />
upper part, later becoming inconspicuously septate; rachis slightly<br />
flexuose, subhyaline, as wide as the basal part, thick-walled near<br />
the base, hyaline <strong>and</strong> thin-walled in the apical part, with scattered<br />
pimple-shaped, unpigmented, approx. 0.5 µm long denticles.<br />
Conidia solitary, subhyaline, thin-walled, finely verrucose, with<br />
a wing-like gelatinous sheath, approx. 0.5 µm wide, ellipsoid<br />
to obovoid, (5–)6–7(–9) × 3–4 µm; hilum slightly prominent,<br />
unpigmented, approx. 0.5 µm diam.<br />
Cultural characteristics: Colonies reaching 40 mm diam after 14 d<br />
at 24 °C; mycelium submerged, flat, smooth; centrally orange, later<br />
becoming powdery to velvety <strong>and</strong> greyish brown due to sporulation,<br />
with sharp, smooth, entire margin; reverse yellowish orange.<br />
Specimen examined: Surinam, isolated from soil, J.H. van Emden, ex-type culture<br />
<strong>CBS</strong> 398.76 = JCM 6968.<br />
www.studiesinmycology.org<br />
Note: This former variety is sufficiently distinguished from M.<br />
schulzeri s. str. by its flexuose conidiophores <strong>and</strong> conidia which<br />
lack an acuminate base, to be regarded as a separate species.<br />
Ramichloridium torvi (Ellis & Everh.) de Hoog, Stud. Mycol. 15:<br />
79. 1977.<br />
≡ Ramularia torvi Ellis & Everh., Rep. Missouri Bot. Gard. 9: 119. 1898.<br />
≡ Hansfordia torvi (Ellis & Everh.) Deighton & Piroz., Mycol. Pap. 101:<br />
39. 1965.<br />
= Acladium biophilum Cif., Sydowia 10: 164. 1956.<br />
≡ Hansfordia biophila (Cif.) M.B. Ellis, in Ellis, More Dematiaceous<br />
Hyphomycetes: 199. 1976.<br />
Specimen: Jamaica, Port Marant, Dec. 1890, on leaves of Solanum torvum,<br />
holotype of Ramularia torvi (NY) (specimen not examined).<br />
Notes: According to the description <strong>and</strong> illustration of R. torvi<br />
provided by de Hoog (1977), this appears to be an additional<br />
species of Myrmecridium. Although it is morphologically <strong>similar</strong> to<br />
M. flexuosum in having a flexuose rachis, it differs from the other<br />
species of the <strong>genus</strong> by having smooth, clavate conidia. Fresh<br />
collections <strong>and</strong> cultures would be required to resolve its status.<br />
85
Arzanlou et al.<br />
Fig. 30. Myrmecridium flexuosum (<strong>CBS</strong> 398.76). A–C. Conidial apparatus at different stages of development, resulting in macronematous conidiophores with sympodially<br />
proliferating conidiogenous cells. D–H. Sympodially proliferating conidiogenous cells giving rise to a flexuose conidium-bearing rachis with pimple-shaped denticles. I. Conidia.<br />
Scale bar = 10 µm.<br />
Fig. 31. Pseudovirgaria hyperparasitica (<strong>CBS</strong> 121739). A–D. Conidial apparatus at different stages of development; conidiogenous cells with geniculate proliferation. E. Conidia.<br />
Scale bar = 10 µm.<br />
86
Ramichloridium <strong>and</strong> allied genera<br />
Pseudovirgaria H.D. Shin, U. Braun, Arzanlou & Crous, gen. nov.<br />
MycoBank MB504564.<br />
Etymology: Named after its morphological <strong>similar</strong>ity to Virgaria.<br />
Hyphomycetes. Uredinicola. Coloniae in vivo pallide vel modice brunneae,<br />
ferrugineae vel cinnamomeae, in vitro lentissime crescentes, murinae. Mycelium<br />
immersum et praecipue externum, ex hyphis ramosis et cellulis conidiogenis<br />
integratis compositum, conidiophoris ab hyphis vegetativis vix distinguendis.<br />
Hyphae ramosae, septatae, leves, tenuitunicatae, hyalinae vel pallide brunneae.<br />
Cellulae conidiogenae integratae in hyphis repentibus, terminales et intercalares,<br />
polyblasticae, sympodialiter proliferentes, subcylindricae vel geniculatae, cicatricibus<br />
conspicuis, solitariis vel numerosis, dispersis vel aggregatis, subdenticulatis,<br />
prominentibus, umbonatis vel apicem versus paulo attenuatis, non inspissatis,<br />
non vel parce fuscatis-refringentibus. Conidia solitaria, holoblastica, plus minusve<br />
obovoidea, recta vel leniter curvata, asymmetrica, continua, hyalina, subhyalina<br />
vel pallidissime olivaceo-brunnea, hilo subconspicuo vel conspicuo, truncato<br />
vel rotundato, non inspissato, non vel lenissime fuscato-refringente; secessio<br />
schizolytica.<br />
Hyperparasitic on uredosori of rust fungi. Colonies in vivo pale<br />
to medium brown, rusty or cinnamom, in vitro slow-growing,<br />
pale to dark mouse-grey. Mycelium immersed <strong>and</strong> mainly aerial,<br />
composed of branched hyphae with integrated conidiogenous<br />
cells, differentiation between vegetative hyphae <strong>and</strong> conidiophores<br />
barely possible. Hyphae branched, septate, smooth, thin-walled,<br />
hyaline to pale brown. Conidiogenous cells <strong>similar</strong>ly hyaline to<br />
pale brown, integrated in creeping threads (hyphae), terminal <strong>and</strong><br />
intercalary, polyblastic, proliferation sympodial, rachis subcylindrical<br />
to geniculate, conidiogenous loci (scars) conspicuous, solitary to<br />
numerous, scattered to aggregated, subdenticulate, bulging out,<br />
umbonate or slightly attenuated towards a rounded apex, wall<br />
unthickened, not to slightly darkened-refractive. Conidia solitary,<br />
formation holoblastic, more or less obovoid, straight to somewhat<br />
curved, asymmetrical, aseptate, hyaline, subhyaline to very pale<br />
olivaceous-brown, with more or less conspicuous hilum, truncate to<br />
rounded, unthickened, not or slightly darkened-refractive; conidial<br />
secession schizolytic.<br />
Type species: Pseudovirgaria hyperparasitica H.D. Shin, U. Braun,<br />
Arzanlou & Crous, sp.nov.<br />
Notes: Other ramichloridium-like isolates from various rust species<br />
form another unique clade, sister to Radulidium subulatum (de<br />
Hoog) Arzanlou, W. Gams & Crous <strong>and</strong> Ra. epichloës (Ellis &<br />
Dearn.) Arzanlou, W. Gams & Crous in the Sordariomycetidae.<br />
Although Pseudovirgaria is morphologically <strong>similar</strong> to Virgaria Nees,<br />
it has hyaline to pale brown hyphae, conidia <strong>and</strong> conidiogenous<br />
cells. <strong>The</strong> conidiogenous cells are integrated in creeping threads<br />
(hyphae), terminal <strong>and</strong> intercalary, <strong>and</strong> the proliferation is distinctly<br />
sympodial. <strong>The</strong> subdenticulate conidiogenous loci are scattered,<br />
solitary, at small shoulders of geniculate conidiogenous cells,<br />
caused by sympodial proliferation, or aggregated, forming slight<br />
swellings of the rachis, i.e., a typical raduliform rachis as in Virgaria<br />
is lacking. Furthermore, the conidiogenous loci of Pseudovirgaria<br />
are bulging, convex, slightly attenuated towards the rouded apex,<br />
in contrast to more cylindrical denticles in Virgaria (Ellis 1971).<br />
<strong>The</strong> scar type of Pseudovirgaria is peculiar due to its convex,<br />
papilla-like shape <strong>and</strong> reminiscent of conidiogenous loci in plantpathogenic<br />
genera like Neoovularia U. Braun <strong>and</strong> Pseudodidymaria<br />
U. Braun (Braun 1998). <strong>The</strong> superficially <strong>similar</strong> <strong>genus</strong> Veronaea is<br />
quite distinct from Pseudovirgaria by having erect conidiophores<br />
with a typical rachis <strong>and</strong> crowded conidiogenous loci which are<br />
flat or only slightly prominent <strong>and</strong> darkened. Pseudovirgaria is<br />
characterised by its mycelium which is composed of branched<br />
hyphae with integrated, terminal <strong>and</strong> intercalary conidiogenous<br />
cells. A differentiation between branched hyphae <strong>and</strong> “branched<br />
www.studiesinmycology.org<br />
conidiophores” is difficult <strong>and</strong> barely possible. It remains unclear<br />
if the “creeping threads” <strong>and</strong> terminal branches of hyphae are<br />
to be interpreted as “creeping conidiophores”. In any case, the<br />
mycelium forms complex fertile branched hyphal structures in which<br />
individual conidiophores are barely discernable. <strong>The</strong>se structures<br />
<strong>and</strong> difficulties in discerning individual conidiophores remind one of<br />
some species of Pseudocercospora Speg. <strong>and</strong> other cercosporoid<br />
genera with abundant superficial mycelium in vivo.<br />
Pseudovirgaria hyperparasitica H.D. Shin, U. Braun, Arzanlou &<br />
Crous, sp. nov. MycoBank MB504565. Figs 6A, 31.<br />
Etymology: Named after its hyperparasitic habit on rust fungi.<br />
Hyphae 1.5–4 µm latae, tenuitunicatae, ≤ 0.5 µm crassae. Cellulae conidiogenae<br />
15–50 × 2–5 µm, tenuitunicatae (≤ 0.5 µm), cicatricibus (0.5–)1.0(–1.5) µm diam,<br />
0.5–1 µm altis. Conidia saepe obovoidea, interdum subclavata, 10–20 × 5–9 µm,<br />
apice rotundato vel paulo attenuato, basi truncata vel rotundata, hilo ca 1 µm<br />
diam.<br />
In vivo: Colonies on rust sori, thin to moderately thick, loose,<br />
cobwebby, to dense, tomentose, pale to medium brown, rusty<br />
or cinnamon. Mycelium partly immersed in the sori, but mainly<br />
superficial, composed of a system of branched hyphae with<br />
integrated conidiogenous cells (fertile threads), distinction between<br />
conidiophores <strong>and</strong> vegetative hyphae difficult <strong>and</strong> barely possible.<br />
Hyphae 1.5–4 µm wide, hyaline, subhyaline to pale yellowish,<br />
greenish or very pale olivaceous, light brownish in mass, thin-walled<br />
(≤ 0.5 µm), smooth, pluriseptate, occasionally slightly constricted<br />
at the septa. Conidiogenous cells integrated in creeping fertile<br />
threads, terminal or intercalary, 15–50 µm long, 2–5 µm wide,<br />
subcylindrical to geniculate, subhyaline to very pale brownish, wall<br />
thin, ≤ 0.5 µm, smooth, proliferation sympodial, with a single to<br />
usually several conidiogenous loci per cell, often crowded, causing<br />
slight swellings, up to 6 µm wide, subdenticulate loci, formed by the<br />
slightly bulging wall, convex, slightly narrowed towards the rounded<br />
apex, (0.5–)1.0(–1.5) µm diam <strong>and</strong> 0.5–1 µm high, wall of the loci<br />
unthickened, not or slightly darkened-refractive, in surface view<br />
visible as minute circle (only rim visible <strong>and</strong> dark). Conidia solitary,<br />
obovoid, often slightly curved with ± unequal sides, 10–20 × 5–9<br />
µm, aseptate, subhyaline, pale yellowish greenish to very pale<br />
olivaceous, wall ≤ 0.5 µm thick, smooth, apex slightly attenuated<br />
to usually broadly rounded, base rounded to somewhat attenuated<br />
towards a more or less conspicuous hilum, (0.5–)1(–1.5) µm<br />
diam, convex to truncate, unthickened, not to slightly darkenedrefractive.<br />
In vitro: Submerged hyphae hyaline to subhyaline, smooth; aerial<br />
hyphae smooth, subhyaline, up to 4 µm wide. Conidiogenous cells<br />
arising imperceptably from aerial vegetative hyphae, terminal,<br />
occasionally intercalary, holoblastic, proliferating sympodially in a<br />
geniculate pattern, with more or less long intervals between groups<br />
of scars; loci slightly darkened, unthickened, approx. 0.5 µm diam.<br />
Conidia hyaline to subhyaline, aseptate, ovoid, often somewhat<br />
curved, (10–)13–15(–17) × (5–)6–7(–8) µm, with truncate base<br />
<strong>and</strong> acutely rounded apex; hila unthickened, slightly darkenedrefractive.<br />
Cultural characteristics: Colonies on MEA rather slow-growing,<br />
reaching 11 mm diam after 14 d at 24 °C, pale to dark mouse-grey,<br />
velvety, compacted, with colonies being up to 1 mm high.<br />
Specimens examined: Korea, Seoul, on uredosori of Frommeëlla sp., on Duchesnea<br />
chrysantha, 17 Sep. 2003, H.D. Shin, paratype, 4/10, CPC 10702–10703 = <strong>CBS</strong><br />
121735–121736, HAL 2053 F; Chunchon, on Phragmidium griseum on Rubus<br />
crataegifolius, 20 Jul. 2004, H.D. Shin, paratype, 2/8, HAL 2057 F; Suwon,<br />
87
Arzanlou et al.<br />
Fig. 32. Radulidium subulatum (<strong>CBS</strong> 405.76). A–B. Macronematous conidiophores with sympodially proliferating conidiogenous cells, resulting in a conidium-bearing rachis.<br />
C–D. Rachis with crowded, blunt conidium-bearing denticles. E. Conidia. Scale bar = 10 µm.<br />
Fig. 33. Radulidium epichloës (<strong>CBS</strong> 361.63). A–C. Conidial apparatus at different stages of development, resulting in semi-micronematous conidiophores <strong>and</strong> sympodially<br />
proliferating conidiogenous cells. D. Rachis with crowded, blunt conidium-bearing denticles. E–F. Conidiophores with acute branches in the lower part. G. Conidia. Scale bar<br />
= 10 µm.<br />
88
Ramichloridium <strong>and</strong> allied genera<br />
on Phragmidium pauciloculare on Rubus parvifolius, 14 Oct. 2003, H.D. Shin,<br />
paratype, 23/10, HAL 2055 F; Hongchon, on Phragmidium rosae-multiflorae on<br />
Rosa multiflora, 11 Aug. 2004, H.D. Shin, paratype, 23/8, HAL 2056 F; Yangpyong,<br />
on Phragmidium sp. on Rubus coreanus, 30 Sep. 2003, H.D. Shin, paratype, 11/10-<br />
1, CPC 10704–10705 = <strong>CBS</strong> 121737–121738, HAL 2052 F, <strong>and</strong> the same locality,<br />
23 Jul. 2004, HAL 2058 F; Chunchon, on Pucciniastrum agrimoniae on Agrimonia<br />
pilosa, 7 Oct. 2002, H.D. Shin, holotype, HAL 2054 F, culture ex-type CPC 10753–<br />
10755 = <strong>CBS</strong> 121739–121741.<br />
Radulidium subulatum <strong>and</strong> Ra. epichloës clade<br />
Ramichloridium subulatum <strong>and</strong> R. epichloës form a distinct, wellsupported<br />
clade with uncertain affinity. This clade is morphologically<br />
distinct <strong>and</strong> a new <strong>genus</strong> is introduced below to accommodate it.<br />
Radulidium Arzanlou, W. Gams & Crous, gen. nov. MycoBank<br />
MB504566.<br />
Etymology: Latin radula = A flexible tongue-like organ in gastropods,<br />
referring to the radula-like denticles on the rachis.<br />
Genus ab aliis generibus Ramichloridii similibus denticulis densissimis, prominentibus,<br />
hebetibus in rachide e cellula conidiogena aculeata orta distinguendum.<br />
Type species: Radulidium subulatum (de Hoog) Arzanlou, W. Gams<br />
& Crous, comb. nov.<br />
In vitro: Colonies fast-growing, velvety, floccose near the margin,<br />
centrally with fertile hyphal bundles up to 10 mm high, about 2<br />
mm diam, with entire but vague margin; mycelium whitish, later<br />
becoming greyish brown. Submerged hyphae smooth, thin-walled.<br />
Conidiophores usually reduced to polyblastic conidiogenous cells<br />
arising from undifferentiated or slightly differentiated aerial hyphae,<br />
terminally integrated or lateral, rarely a branched conidiophore<br />
present, smooth, slightly thick-walled, pale brown, cylindrical to<br />
acicular, widest at the base <strong>and</strong> tapering towards the apex; apical<br />
part forming a pale brown, generally straight rachis, with crowded,<br />
prominent, blunt denticles, suggesting a gastropod radula; denticles<br />
0.5–1 µm long, apically pale brown. Conidia solitary, subhyaline,<br />
thin- or slightly thick-walled, smooth or verrucose, obovoidal,<br />
fusiform to subcylindrical, base subtruncate <strong>and</strong> with a slightly<br />
prominent, conspicuously pigmented hilum; conidial secession<br />
schizolytic.<br />
Notes: Radulidium can be distinguished from other ramichloridiumlike<br />
fungi by its slightly differentiated conidiophores <strong>and</strong> prominent,<br />
blunt, very dense conidium-bearing denticles. Although the<br />
Radulidium clade consists of several subclusters that correlate<br />
with differences in morphology, the ITS sequence data appear<br />
insufficient to resolve this species complex. <strong>The</strong>refore, only two<br />
species of Radulidium with clear morphological <strong>and</strong> molecular<br />
differences are treated here. <strong>The</strong> phylogenetic situation of other<br />
taxa in this clade will be treated in a further study employing a multigene<br />
approach.<br />
Radulidium subulatum (de Hoog) Arzanlou, W. Gams & Crous,<br />
comb. nov. MycoBank MB504567. Figs 10C, 32.<br />
Basionym: Ramichloridium subulatum de Hoog, Stud. Mycol. 15:<br />
83. 1977.<br />
Misapplied name: Rhinocladiella elatior Mangenot sensu dal Vesco<br />
& B. Peyronel, Allionia 14: 38. 1968.<br />
In vitro: Submerged hyphae hyaline, thin-walled, 1–2.5 µm wide;<br />
aerial hyphae brownish. Conidiogenous cells arising laterally from<br />
vegetative hyphae, pale brown, smooth, thick-walled, sometimes<br />
without a basal septum, cylindrical to aculeate, tapering gradually<br />
towards the apex, widest at the base, 25–40 × 2–3 µm; proliferating<br />
sympodially, forming a pale brown rachis, with densely crowded,<br />
prominent, blunt conidium-bearing denticles, with pale brown apex.<br />
Conidia solitary, subhyaline, thin-walled, smooth, ellipsoidal to<br />
almost clavate, 5–7 × 1.5–2 µm, with a slightly pigmented, nonrefractive<br />
hilum, about 1 µm diam.<br />
Cultural characteristics: Colonies on MEA rather fast growing,<br />
reaching 50 mm diam after 14 d at 24 °C, with entire but vague<br />
margin, velvety, floccose near the margin, centrally with fertile<br />
hyphal bundles up to 10 mm high, about 2 mm diam; mycelium<br />
whitish, later becoming greyish brown; reverse grey, zonate.<br />
Specimens examined: Czech Republic, on Phragmites australis, A. Samšiňáková,<br />
ex-type culture <strong>CBS</strong> 405.76; Opatovicky pond, from Lasioptera arundinis (gall<br />
midge) mycangia on Phragmites australis, M. Skuhravá, <strong>CBS</strong> 101010.<br />
Radulidium epichloës (Ellis & Dearn.) Arzanlou, W. Gams &<br />
Crous, comb. nov. MycoBank MB504568. Fig. 33.<br />
Basionym: Botrytis epichloës Ellis & Dearn., Canad. Record Sci.<br />
9: 272. 1893.<br />
≡ Ramichloridium epichloës (Ellis & Dearn.) de Hoog, Stud. Mycol. 15:<br />
81. 1977.<br />
In vitro: Submerged hyphae hyaline, thin-walled, 1–2.5 µm wide;<br />
aerial hyphae somewhat darker. Conidiogenous cells arising<br />
laterally or terminally from undifferentiated or slightly differentiated<br />
aerial hyphae, occasionally acutely branched in the lower part,<br />
smooth, thick-walled, pale brown, more or less cylindrical, later<br />
with thin septa, 25–47 µm long; proliferating sympodially, forming<br />
a rather short, pale brown, straight or somewhat geniculate<br />
rachis, with crowded, prominent, blunt denticles with pale brown<br />
apex. Conidia solitary, subhyaline, rather thin-walled, verruculose,<br />
obovoidal to fusiform, (4.5–)7–8(–11) × 2–3 µm, with a pigmented<br />
hilum, 1–1.5 µm diam.<br />
Cultural characteristics: Colonies reaching 45 mm diam after 14 d<br />
at 24 °C, with smooth, rather vague, entire margin; velvety, centrally<br />
floccose <strong>and</strong> elevated up to 2 mm high; surface mycelium whitish,<br />
later becoming greyish brown; reverse pale ochraceous.<br />
Specimen examined: U.S.A., Cranberry Lake, Michigan, isolated from Epichloë<br />
typhina on Glyceria striata, G.L. Hennebert, <strong>CBS</strong> 361.63 = MUCL 3124; specimen<br />
in MUCL designated here as epitype.<br />
Veronaea-like clade, allied to the Annulatascaceae<br />
A veronaea-like isolate from Bertia moriformis clusters near the<br />
Annulatascaceae, <strong>and</strong> is morphologically distinct from other known<br />
anamorph genera in the Ramichloridium complex, <strong>and</strong> therefore a<br />
new <strong>genus</strong> is introduced to accommodate it.<br />
Rhodoveronaea Arzanlou, W. Gams & Crous, gen. nov. MycoBank<br />
MB504569.<br />
Etymology: (Greek) rhodon = the rose, referring to the red-brown<br />
conidiophores, suffix -veronaea from Veronaea.<br />
Genus ab aliis generibus Ramichloridii similibus basi condiorum late truncata et<br />
marginata distinguenda.<br />
In vitro: Colonies slow-growing, velvety, floccose; surface<br />
olivaceous-grey to dark olivaceous-green; reverse olivaceousblack.<br />
Hyphae smooth, thin-walled, pale olivaceous. Conidiophores<br />
arising vertically from creeping hyphae, straight or flexuose, simple,<br />
thick-walled, red-brown, with inflated basal cell. Conidiogenous<br />
cells terminally integrated, polyblastic, sympodial, smooth, thick-<br />
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89
Arzanlou et al.<br />
Fig. 34. Rhodoveronaea varioseptata (<strong>CBS</strong> 431.88). A–D. Macronematous conidiophores with sympodially proliferating conidiogenous cells, resulting in conidium bearing rachis<br />
with slightly prominent conidium-bearing denticles. E–F. Conidia with minute marginal frill. Scale bar = 10 µm.<br />
Fig. 35. Veronaeopsis simplex (<strong>CBS</strong> 588.66). A–C. Conidial apparatus at different stages of development, resulting in semi-micronematous conidiophores <strong>and</strong> sympodially<br />
proliferating conidiogenous cells. D–E. Rachis with crowded, prominent denticles. F. Intercalary conidiogenous cells. G. Conidia. Scale bar = 10 µm.<br />
walled, pale brown, rachis straight, occasionally geniculate, with<br />
crowded, slightly prominent conidium-bearing denticles; denticles<br />
flat-tipped, slightly pigmented. Conidia solitary, pale brown, thinor<br />
slightly thick-walled, smooth, ellipsoidal to obovoidal, 0–multiseptate,<br />
with a protruding base <strong>and</strong> a marginal basal frill; conidial<br />
secession schizolytic.<br />
Type species: Rhodoveronaea varioseptata Arzanlou, W. Gams &<br />
Crous, sp. nov.<br />
Notes: Rhodoveronaea differs from other ramichloridium-like fungi<br />
by the presence of a basal, marginal conidial frill, <strong>and</strong> variably<br />
septate conidia.<br />
90
Ramichloridium <strong>and</strong> allied genera<br />
Rhodoveronaea varioseptata Arzanlou, W. Gams & Crous, sp.<br />
nov. MycoBank MB504570. Figs 10D, 34.<br />
Etymology: Named for its variably septate conidia.<br />
Hyphae 2–3 µm latae. Conidiophora ad 125 µm longa et 3–5 µm lata. Cellulae<br />
conidiogenae 30–70 µm longae et 3–5 µm latae. Conidia 0–2(–3)-septata, (8–)11–<br />
13(–15) × (2–)3–4(–6) µm.<br />
In vitro: Submerged hyphae smooth, thin-walled, pale olivaceous,<br />
2–3 µm wide; aerial hyphae smooth, brownish <strong>and</strong> slightly narrower.<br />
Conidiophores arising vertically from creeping hyphae, straight or<br />
flexuose, simple, smooth, thick-walled, red-brown, up to 125 µm<br />
long, 3–5 µm wide, often with inflated basal cell. Conidiogenous<br />
cells terminally integrated, smooth, thick-walled, pale brown at the<br />
base, paler towards the apex, straight, variable in length, 30–70<br />
µm long <strong>and</strong> 3–5 µm wide, rachis straight, occasionally geniculate;<br />
slightly prominent conidium-bearing denticles, crowded, with slightly<br />
pigmented apex, about 1 µm diam. Conidia solitary, pale brown,<br />
thin- or slightly thick-walled, smooth, ellipsoid to obovoid, 0–2(–3)-<br />
septate, (8–)11–13(–15) × (2–)3–4(–6) µm with a protruding base,<br />
1.5 µm wide, <strong>and</strong> marginal frill.<br />
Cultural characteristics: Colonies reaching 12 mm diam after 14<br />
d at 24 °C, velvety, floccose; surface olivaceous-grey to dark<br />
olivaceous-green; reverse olivaceous-black.<br />
Specimen examined: Germany, Eifel, Berndorf, on Bertia moriformis, Sep. 1987, W.<br />
Gams, holotype <strong>CBS</strong>-H 19932, culture ex-type <strong>CBS</strong> 431.88.<br />
Venturiaceae (Pleosporales)<br />
<strong>The</strong> ex-type strain of Veronaea simplex (Papendorf 1969) did not<br />
cluster with the <strong>genus</strong> Veronaea (Herpotrichiellaceae), but is allied<br />
to the Venturiaceae. Veronaea simplex is distinct from species<br />
of Fusicladium Bonord. by having a well-developed rachis with<br />
densely aggregated scars. A new <strong>genus</strong> is thus introduced to<br />
accommodate this taxon.<br />
Veronaeopsis Arzanlou & Crous, gen. nov. MycoBank<br />
MB504571.<br />
Etymology: <strong>The</strong> suffix -opsis refers to its <strong>similar</strong>ity with Veronaea.<br />
Genus Veronaeae simile sed conidiophoris brevioribus (ad 60 μm longis) et rachide<br />
dense denticulata distinguendum.<br />
In vitro: Colonies moderately fast-growing; surface velvety, floccose,<br />
greyish sepia to hazel, with smooth margin; reverse mouse-grey<br />
to dark mouse-grey. Conidiophores arising vertically from aerial<br />
hyphae, lateral or intercalary, simple or branched, occasionally<br />
reduced to conidiogenous cells, pale brown. Conidiogenous<br />
cells terminally integrated on simple or branched conidiophores,<br />
polyblastic, smooth, thin-walled, pale brown; rachis commonly<br />
straight, geniculate, with densely crowded, prominent denticles,<br />
<strong>and</strong> slightly pigmented scars. Conidia solitary, subhyaline to pale<br />
brown, thin- or slightly thick-walled, smooth, oblong-ellipsoidal to<br />
subcylindrical, (0–)1-septate, with a slightly darkened, thickened,<br />
hilum; conidial secession schizolytic.<br />
Type species: Veronaeopsis simplex (Papendorf) Arzanlou &<br />
Crous, comb. nov.<br />
Veronaeopsis simplex (Papendorf) Arzanlou & Crous, comb.<br />
nov. MycoBank MB504572. Figs 17C, 35.<br />
Basionym: Veronaea simplex Papendorf, Trans. Brit. Mycol. Soc.<br />
52: 486. 1969.<br />
www.studiesinmycology.org<br />
In vitro: Submerged hyphae smooth, thin-walled, pale brown; aerial<br />
hyphae aggregated in bundles. Conidiophores arising vertically<br />
from aerial hyphae, lateral or intercalary, simple or branched,<br />
occasionally reduced to conidiogenous cells, pale brown, rather<br />
short, up to 60 µm long, 1.5–2 µm wide. Conidiogenous cells<br />
terminally integrated in the conidiophores, smooth, thin-walled, pale<br />
brown, variable in length, 5–25 µm long, rachis generally straight<br />
or irregularly geniculate, with crowded, prominent denticles, about<br />
0.5 µm long, flat-tipped, with slightly pigmented apex. Conidia<br />
solitary, subhyaline to pale brown, thin- or slightly thick-walled,<br />
smooth, oblong-ellipsoidal to subcylindrical, (0–)1-septate, slightly<br />
constricted at the septum, (6–)10–12(–15) × (2–)2.5–3(–4) µm;<br />
hilum slightly darkened <strong>and</strong> thickened, not refractive, about 1 µm<br />
diam.<br />
Cultural characteristics: Colonies reaching 25 mm diam after 14<br />
d at 24 °C; surface velvety, floccose, greyish sepia to hazel, with<br />
smooth margin; reverse mouse-grey to dark mouse-grey.<br />
Specimen examined: South Africa, Potchefstroom, on leaf litter of Acacia karroo,<br />
1966, M.C. Papendorf, holotype, <strong>CBS</strong> H-7810; culture ex-type <strong>CBS</strong> 588.66 = IMI<br />
203547.<br />
Notes: <strong>The</strong> presence of 1-septate conidia in Veronaeopsis overlaps<br />
with Veronaea. However, Veronaeopsis differs from Veronaea<br />
based on its conidiophore <strong>and</strong> conidiogenous cell morphology.<br />
Veronaea has much longer, macronematous conidiophores than<br />
Veronaeopsis. Furthermore, Veronaea has a more or less straight<br />
rachis, whereas in Veronaeopsis the rachis is often geniculate.<br />
<strong>The</strong> conidiogenous loci in Veronaea are less prominent, i.e., less<br />
denticle-like.<br />
Discussion<br />
<strong>The</strong> present study was initiated chiefly to clarify the status of<br />
Ramichloridium musae, the causal organism of tropical speckle<br />
disease of banana (Jones 2000). Much confusion surrounded this<br />
name in the past, relating, respectively, to its validation, species<br />
<strong>and</strong> generic status. As was revealed in the present study, however,<br />
two species are involved in banana speckle disease, namely R.<br />
musae <strong>and</strong> R. biverticillatum. Even more surprising was the fact<br />
that Ramichloridium comprises anamorphs of Mycosphaerella<br />
Johanson (Mycosphaerellaceae), though no teleomorphs have<br />
thus far been conclusively linked to any species of Ramichloridium.<br />
By investigating the Ramichloridium generic complex as outlined<br />
by de Hoog (1977), another <strong>genus</strong> associated with leaf spots,<br />
namely Periconiella, was also shown to represent an anamorph<br />
of Mycosphaerella. Although no teleomorph connections have<br />
been proven for ramichloridium-like taxa, de Hoog et al. (1983)<br />
refer to the type specimen of Wentiomyces javanicus Koord.<br />
(Pseudoperisporiaceae), on the type specimen of which (PC)<br />
some ramichloridium-like conidiophores were seen. Without<br />
fresh material <strong>and</strong> an anamorph-teleomorph connection proven<br />
in culture, however, this matter cannot be investigated further. It<br />
is interesting to note, however, that Wentiomyces Koord. shows<br />
a strong resemblance to Mycosphaerella, except for the external<br />
perithecial appendages.<br />
<strong>The</strong> <strong>genus</strong> Mycosphaerella is presently one of the largest genera<br />
of ascomycetes, containing close to 3 000 names (Aptroot 2006), to<br />
which approximately 30 anamorph genera have already been linked<br />
(Crous et al. 2006a, b, 2007). By adding two additional anamorph<br />
genera, the Mycosphaerella complex appears to be exp<strong>and</strong>ing<br />
91
Arzanlou et al.<br />
even further, though some taxa have been shown to reside in other<br />
families in the Capnodiales, such as Davidiella Crous & U. Braun<br />
(Davidiellaceae) <strong>and</strong> Teratosphaeria (Teratosphaeriaceae) (Braun<br />
et al. 2003, Crous et al. 2007, Schubert et al. 2007 – this volume).<br />
Another family, which proved to accommodate several<br />
ramichloridium-like taxa, is the Herpotrichiellaceae (Chaetothyriales).<br />
Members of the Chaetothyriales are regularly encountered as<br />
causal agents of human mycoses (Haase et al. 1999, de Hoog<br />
et al. 2003), whereas species of the Capnodiales are common<br />
plant pathogens, or chiefly associated with plants. Species in the<br />
Chaetothyriales have consistently melanized thalli, which is a factor<br />
enabling them to invade humans, <strong>and</strong> cause a wide diversity of<br />
mycoses, such as chromoblastomycosis, mycetoma, brain infection<br />
<strong>and</strong> subcutaneous phaeohyphomycosis (de Hoog et al. 2003). <strong>The</strong><br />
only known teleomorph connection in this <strong>genus</strong> is Capronia Sacc.<br />
(Untereiner & Naveau 1999).<br />
Rhinocladiella <strong>and</strong> Veronaea were in the past frequently<br />
confused with the <strong>genus</strong> Ramichloridium. However, Rhinocladiella,<br />
as well as Veronaea <strong>and</strong> Thysanorea, were shown to cluster in the<br />
Chaetothyriales, while Ramichloridium clusters in the Capnodiales.<br />
Rhinocladiella mackenziei, which causes severe cerebral<br />
phaeohyphomycosis in humans (Sutton et al. 1998), has in the<br />
past been confused with Pleurothecium obovoideum (Ur-Rahman<br />
et al. 1988). Data presented here reveal, however, that although<br />
morphologically <strong>similar</strong>, these species are phylogenetically<br />
separate, with P. obovoideum belonging to the Sordariales, where<br />
it clusters with sexual species of Carpoligna F.A. Fernández &<br />
Huhndorf that have Pleurothecium anamorphs (Fernández et al.<br />
1999).<br />
In addition to the genera clustering in the Capnodiales <strong>and</strong><br />
Chaetothyriales, several ramichloridium-like genera are newly<br />
introduced to accommodate species that cluster elsewhere<br />
in the ascomycetes, namely Pseudovirgaria, Radulidium <strong>and</strong><br />
Myrmecridium, Veronaeopsis, <strong>and</strong> Rhodoveronaea. Although the<br />
ecological role of these taxa is much less known than that of taxa in<br />
the Capnodiales <strong>and</strong> Chaetothyriales, some exhibit an interesting<br />
ecology. For instance, the fungicolous habit of Pseudovirgaria, as<br />
well as some species in Radulidium, which are found on various<br />
rust species, suggests that these genera should be screened<br />
further to establish if they have any potential biocontrol properties.<br />
Furthermore, these two genera share a common ancestor, <strong>and</strong><br />
further work is required to determine whether speciation was shaped<br />
by co-evolution with the rusts. A further species of “Veronaea” that<br />
might belong to Pseudovirgaria is Veronaea harunganae (Hansf.)<br />
M.B. Ellis, which is known to occur on Hemileia harunganae<br />
Cummins on Harungana in Tanzania <strong>and</strong> Ug<strong>and</strong>a (Ellis 1976). <strong>The</strong><br />
latter species, however, is presently not known from culture, <strong>and</strong><br />
needs to be recollected to facilitate further study.<br />
<strong>The</strong> genera distinguished here represent homogeneous clades<br />
in the phylogenetic analysis. Only the species of Rhinocladiella are<br />
dispersed among others morphologically classified in Exophiala or<br />
other genera.<br />
By integrating the phylogenetic data generated here with<br />
the various morphological data sets, we were able to resolve<br />
eight clades for taxa formerly regarded as representative of the<br />
Ramichloridium complex. According to the phylogeny inferred<br />
from 28S rDNA sequence data, the genera Ramichloridium <strong>and</strong><br />
Periconiella were heterogeneous, requiring the introduction of<br />
several novel genera. Although the present 11 odd genera can<br />
still be distinguished based on their morphology, it is unlikely that<br />
morphological identifications without the supplement of molecular<br />
data would in the future be able to accurately identify all the novel<br />
isolates that undoubtably await description. <strong>The</strong> integration of<br />
morphology with phylogenetic data not only helps to resolve generic<br />
affinities, but it also assists in discriminating between the various<br />
cryptic species that surround many of these well-known names that<br />
are presently freely used in the literature. To that end it is interesting<br />
to note that for the majority of the taxa studied here, the ITS domain<br />
(Table 1) provided good species resolution. However, more genes<br />
will have to be screened in future studies aimed at characterising<br />
some of the species complexes where the ITS domain provided<br />
insufficient phylogenetic signal (data not shown) to resolve all of the<br />
observed morphological species.<br />
ACKNOWLEDGEMENTS<br />
<strong>The</strong> work of Mahdi Arzanlou was funded by the Ministry of Science, Research <strong>and</strong><br />
Technology of Iran, which we gratefully acknowledge. Several colleagues from<br />
different countries provided material without which this work would not have been<br />
possible. We thank Marjan Vermaas for preparing the photographic plates, <strong>and</strong><br />
Arien van Iperen for taking care of the cultures.<br />
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93
available online at www.studiesinmycology.org<br />
doi:10.3114/sim.2007.58.04<br />
Studies in Mycology 58: 95–104. 2007.<br />
<strong>Cladosporium</strong> leaf-blotch <strong>and</strong> stem rot of Paeonia spp. caused by Dichocladosporium<br />
chlorocephalum gen. nov.<br />
K. Schubert 1* , U. Braun 2 , J.Z. Groenewald 3 <strong>and</strong> P.W. Crous 3<br />
1<br />
Botanische Staatssammlung München, Menzinger Strasse 67, D-80638 München, Germany; 2 Martin-Luther-Universität, Institut für Biologie, Geobotanik und Botanischer Garten,<br />
Herbarium, Neuwerk 21, D-06099 Halle (Saale), Germany; 3 <strong>CBS</strong> Fungal Biodiversity Centre, P.O. Box 85167, 3508 AD Utrecht, <strong>The</strong> Netherl<strong>and</strong>s<br />
*Correspondence: Konstanze Schubert, konstanze.schubert@gmx.de<br />
Abstract: <strong>Cladosporium</strong> chlorocephalum (= C. paeoniae) is a common, widespread leaf-spotting hyphomycete of peony (Paeonia spp.), characterised by having dimorphic<br />
conidiophores. During the season, one stage of this fungus causes distinct, necrotic leaf-blotch symptoms on living leaves of Paeonia spp. In late autumn, winter or after<br />
overwintering, a second morphologically distinct conidiophore type occurs on dead, blackish, rotting stems. Conspecificity of the two morphs, previously proposed on the basis<br />
of observations in culture, was supported by DNA sequence data from the ITS <strong>and</strong> LSU gene regions, using cultures obtained from leaf-blotch symptoms on living leaves, as<br />
well as from dead stems of Paeonia spp. Sequence data were identical, indicating a single species with two morphs. On account of its distinct conidiogenous loci <strong>and</strong> conidial<br />
hila, as well as its sequence-based phylogenetic position separate from the Davidiella/<strong>Cladosporium</strong> clade, the peony fungus has to be excluded from <strong>Cladosporium</strong> s. str., but<br />
still belongs to the Davidiellaceae (Capnodiales). <strong>The</strong> leaf-blotching (cladosporioid) morph of this fungus morphologically resembles species of Fusicladium, but differs in having<br />
dimorphic fruiting, <strong>and</strong> is phylogenetically distant from the Venturiaceae. <strong>The</strong> macronematous (periconioid) morph resembles Metulocladosporiella (Chaetothyriales), but lacks<br />
rhizoid conidiophore hyphae, <strong>and</strong> has 0–5-septate conidia. Hence, C. chlorocephalum is assigned to the new <strong>genus</strong> Dichocladosporium.<br />
Taxonomic novelties: Dichocladosporium K. Schub., U. Braun & Crous, gen. nov., Dichocladosporium chlorocephalum (Fresen.) K. Schub., U. Braun & Crous, comb. nov.<br />
Key words: Anamorphic fungi, <strong>Cladosporium</strong> chlorocephalum, C. paeoniae, hyphomycetes, new <strong>genus</strong>, phylogeny, taxonomy.<br />
Introduction<br />
Fresenius (1850) described Periconia chlorocephala Fresen. from<br />
Germany on dead stems of Paeonia sp. Mason & Ellis (1953)<br />
examined this species in vitro <strong>and</strong> in vivo <strong>and</strong> stated that it only<br />
occurred on dead stems of Paeonia spp. <strong>The</strong>y described, illustrated<br />
<strong>and</strong> discussed this species in detail, <strong>and</strong> reallocated it to the <strong>genus</strong><br />
<strong>Cladosporium</strong> Link.<br />
A second, cladosporioid hyphomycete on Paeonia spp.,<br />
<strong>Cladosporium</strong> paeoniae Pass., was collected by Passerini on living<br />
leaves of P. albiflora (as P. edulis) in Italy, <strong>and</strong> distributed in Thümen,<br />
Herbarium mycologicum oeconomicum, Fasc. IX, No. 416 (1876),<br />
together with the first valid description, which was repeated by<br />
Passerini (1876). Later, Passerini collected this fungus on Paeonia<br />
officinalis at Parma in Italy <strong>and</strong> distributed it in Thümen, Mycotheca<br />
universalis, No. 670 (1877). Saccardo (1882) listed a collection of<br />
this species on Paeonia anomala from Russia, Siberia, which he<br />
later described as <strong>Cladosporium</strong> paeoniae var. paeoniae-anomalae<br />
Sacc. (Saccardo 1886). A first examination of C. paeoniae in culture<br />
was accomplished by Meuli (1937), followed by a treatment in vitro<br />
by de Vries (1952). Mason & Ellis (1953) described <strong>and</strong> illustrated in<br />
their treatment of C. chlorocephalum macroconidiophores, agreeing<br />
with those of the original diagnosis <strong>and</strong> illustration of Periconia<br />
chlorocephala, as well as semi-macronematous conidiophores<br />
concurring with those of C. paeoniae, although no mention was<br />
made of the latter name. McKemy & Morgan-Jones (1991) carried<br />
out comprehensive studies on <strong>Cladosporium</strong> on Paeonia spp. in<br />
vitro <strong>and</strong> in vivo, including detailed discussions of the history of<br />
the taxa concerned, taxonomic implications <strong>and</strong> comprehensive<br />
descriptions <strong>and</strong> illustrations. <strong>The</strong>y concluded that <strong>Cladosporium</strong><br />
paeoniae, found in culture together with C. chlorocephalum, was<br />
a semi-macronematous form (synanamorph) of the latter species,<br />
<strong>and</strong> reduced C. paeoniae to synonymy with the latter species.<br />
In the present study, re-examination <strong>and</strong> reassessment of<br />
morphological characters, conidiogenesis, <strong>and</strong> DNA sequence<br />
data of the ITS <strong>and</strong> 28S nrDNA were used to confirm the identity<br />
of <strong>Cladosporium</strong> chlorocephalum (the periconioid morph) <strong>and</strong> C.<br />
paeoniae (the cladosporioid morph), <strong>and</strong> clarify their relation to<br />
<strong>Cladosporium</strong> s. str. (Davidiellaceae) (emend. David 1997, Braun<br />
et al. 2003).<br />
Materials <strong>and</strong> methods<br />
Isolates<br />
Single-conidial isolates were obtained from symptomatic leaves<br />
<strong>and</strong> dead stems, <strong>and</strong> cultured as detailed in Crous (1998). Cultural<br />
characteristics <strong>and</strong> morphology of isolates (Table 1) were recorded<br />
from plates containing either 2 % potato-dextrose agar (PDA) or<br />
synthetic nutrient-poor agar (SNA) (Gams et al. 2007). Plates were<br />
incubated at 25 °C under continuous near-UV light to promote<br />
sporulation.<br />
DNA isolation, amplification <strong>and</strong> sequencing<br />
Fungal colonies were established on agar plates, <strong>and</strong> genomic<br />
DNA was isolated following the protocol of Lee & Taylor (1990).<br />
<strong>The</strong> primers V9G (de Hoog & Gerrits van den Ende 1998) <strong>and</strong> LR5<br />
(Vilgalys & Hester 1990) were used to amplify part (ITS) of the nuclear<br />
rDNA operon spanning the 3’ end of the 18S rRNA gene (SSU), the<br />
first internal transcribed spacer (ITS1), the 5.8S rRNA gene, the<br />
second ITS region <strong>and</strong> the 5’ end of the 28S rRNA gene (LSU).<br />
<strong>The</strong> primers ITS4 (White et al. 1990), LR0R (Rehner & Samuels<br />
1994), LR3R (www.biology.duke.edu/fungi/mycolab/primers.htm)<br />
<strong>and</strong> LR16 (Moncalvo et al. 1993) were used as internal sequence<br />
primers to ensure good quality sequences over the entire length<br />
of the amplicon. <strong>The</strong> PCR conditions, sequence alignment <strong>and</strong><br />
subsequent phylogenetic analysis followed the methods of Crous<br />
et al. (2006b). Gaps longer than 10 bases were coded as single<br />
events for the phylogenetic analyses; the remaining gaps were<br />
treated as new character states. Sequence data were deposited<br />
in GenBank (Table 1) <strong>and</strong> alignments in TreeBASE (www.treebase.<br />
org).<br />
95
Schubert et al.<br />
Morphology<br />
Morphological examinations were made from herbarium samples,<br />
fresh symptomatic leaves <strong>and</strong> stems, as well as cultures sporulating<br />
on SNA. Structures were mounted in water or Shear’s solution<br />
(Dhingra & Sinclair 1985), <strong>and</strong> 30 measurements at × 1 000<br />
magnification were made of each structure under an Olympus BX<br />
50 microscope (Hamburg, Germany). <strong>The</strong> 95 % confidence levels<br />
were determined <strong>and</strong> the extremes of spore measurements given<br />
in parentheses. Scanning electron microscopic examinations were<br />
conducted at the Institute of Zoology, Martin-Luther-University,<br />
Halle (Saale), Germany, using a Hitachi S-2400. Samples were<br />
coated with a thin layer of gold applied with a sputter coater SCD<br />
004 (200 s in an argon atmosphere of 20 mA, 30 mm distant<br />
from the electrode). Colony colours were noted after 2 wk growth<br />
on PDA at 25 °C in the dark, using the colour charts of Rayner<br />
(1970). All cultures studied were deposited in the culture collection<br />
of the Centraalbureau voor Schimmelcultures (<strong>CBS</strong>), Utrecht,<br />
the Netherl<strong>and</strong>s (Table 1). Taxonomic novelties were lodged with<br />
MycoBank (www.MycoBank.org).<br />
Results<br />
DNA phylogeny<br />
Amplification products of approximately 1 700 bases were obtained<br />
for the isolates listed in Table 1. <strong>The</strong> ITS region of the sequences<br />
was used to obtain additional sequences from GenBank which were<br />
added to the alignment. <strong>The</strong> manually adjusted alignment contained<br />
26 sequences (including the two outgroup sequences) <strong>and</strong> 518<br />
characters including alignment gaps. Of the 518 characters used<br />
in the phylogenetic analysis, 226 were parsimony-informative, 33<br />
were variable <strong>and</strong> parsimony-uninformative, <strong>and</strong> 259 were constant.<br />
Neighbour-joining analysis using three substitution models on the<br />
sequence data yielded trees supporting the same clades but with<br />
a different arrangement at the deeper nodes. <strong>The</strong>se nodes were<br />
supported poorly in the bootstrap analyses (the highest value<br />
observed for one of these nodes was 64 %; data not shown).<br />
Two equally most parsimonious trees (TL = 585 steps; CI =<br />
0.761; RI = 0.902; RC = 0.686), one of which is shown in Fig. 1,<br />
was obtained from the parsimony analysis of the ITS region. <strong>The</strong><br />
Dichocladosporium K. Schub., U. Braun & Crous isolates formed a<br />
well-supported clade (100 % bootstrap support), distinct from clades<br />
containing species of Davidiella Crous & U. Braun, Mycosphaerella<br />
Johanson <strong>and</strong> Teratosphaeria Syd. & P. Syd. This placement was<br />
also supported by analyses of the first part of the 28S rRNA gene<br />
(see Crous et al. 2007 – this volume).<br />
Taxonomy<br />
Because conidia formed holoblastically in simple or branched<br />
acropetal chains, <strong>Cladosporium</strong> chlorocephalum <strong>and</strong> C. paeoniae<br />
coincided with previous concepts of <strong>Cladosporium</strong> s. lat. (Braun<br />
et al. 2003, Schubert 2005), belonging to a wide assemblage of<br />
genera classified by Kiffer & Morelet (1999) as “Acroblastosporae”.<br />
Previous studies conducted in vitro concluded that <strong>Cladosporium</strong><br />
chlorocephalum <strong>and</strong> C. paeoniae represent two developmental<br />
stages (morphs) of a single species, a result confirmed here by<br />
DNA sequence analyses. A detailed analysis of conidiogenesis,<br />
structure of the conidiogenous loci <strong>and</strong> conidial hila, <strong>and</strong> a<br />
comparison with <strong>Cladosporium</strong> s. str., typified by C. herbarum<br />
(Pers. : Fr.) Link, revealed obvious differences: <strong>The</strong> conidiogenous<br />
loci <strong>and</strong> conidial hila of C. chlorocephalum are quite distinct from<br />
those of <strong>Cladosporium</strong> s. str. by being denticulate or subdenticulate,<br />
apically broadly truncate, unthickened or slightly thickened, but<br />
somewhat darkened-refractive. <strong>The</strong> scars in <strong>Cladosporium</strong> s.<br />
str. are, however, characteristically coronate, i.e., with a central<br />
convex dome surrounded by a raised periclinal rim (David 1997,<br />
Braun et al. 2003, Schubert 2005). Hence, the peony fungus<br />
has to be excluded from <strong>Cladosporium</strong> s. str. A comparison with<br />
phaeoblastic hyphomycetous genera revealed a close <strong>similar</strong>ity of<br />
this fungus with the <strong>genus</strong> Metulocladosporiella Crous, Schroers,<br />
J.Z. Groenew., U. Braun & K. Schub. recently introduced for the<br />
<strong>Cladosporium</strong> speckle disease of banana (Crous et al. 2006a).<br />
Both fungi have dimorphic fruiting, pigmented macronematous<br />
conidiophores often with distinct basal swellings <strong>and</strong> densely<br />
branched terminal heads composed of short branchlets <strong>and</strong><br />
ramoconidia, denticulate or subdenticulate unthickened, but<br />
somewhat darkened-refractive conidiogenous loci, as well as<br />
phaeoblastic conidia, formed in simple or branched acropetal<br />
chains. <strong>The</strong> semi-macronematous leaf-blotching morph is close<br />
to <strong>and</strong> barely distinguishable from Fusicladium Bonord. However,<br />
unlike Metulocladosporiella, the peony fungus does not form rhizoid<br />
hyphae at the base of conidiophore swellings <strong>and</strong> the conidia are<br />
amero- to phragmosporous [0–5-septate versus 0(–1)-septate<br />
in Metulocladosporiella]. Furthermore, the peony fungus neither<br />
clusters within the Chaetothyriales (with Metulocladosporiella)<br />
nor within the Venturiaceae (with Fusicladium), but clusters basal<br />
to the Davidiellaceae (see also Crous et al. 2007 – this volume).<br />
Hence, we propose to place C. chlorocephalum in the new <strong>genus</strong><br />
Dichocladosporium.<br />
Dichocladosporium K. Schub., U. Braun & Crous, gen. nov.<br />
MycoBank MB504428. Figs 2–5.<br />
Etymology: dicha in Greek = twofold.<br />
Differt a Metulocladosporiella conidiophoris cum cellulis basalibus saepe inflatis,<br />
sed sine hyphis rhizoidibus, conidiis amero- ad phragmosporis (0–5-septatis).<br />
Type species: Dichocladosporium chlorocephalum (Fresen.) K.<br />
Schub., U. Braun & Crous, comb. nov.<br />
Dichocladosporium chlorocephalum (Fresen.) K. Schub., U.<br />
Braun & Crous, comb. nov. MycoBank MB504429. Figs 2–5.<br />
Basionym: Periconia chlorocephala Fresen., Beiträge zur Mykologie<br />
1: 21. 1850.<br />
≡ Haplographium chlorocephalum (Fresen.) Grove, Sci. Gossip 21: 198.<br />
1885.<br />
≡ Graphiopsis chlorocephala (Fresen.) Trail, Scott. Naturalist (Perth) 10:<br />
75. 1889.<br />
≡ <strong>Cladosporium</strong> chlorocephalum (Fresen.) E.W. Mason & M.B. Ellis,<br />
Mycol. Pap. 56: 123. 1953.<br />
= <strong>Cladosporium</strong> paeoniae Pass., in Thümen, Herb. Mycol. Oecon., Fasc. IX,<br />
No. 416. 1876, <strong>and</strong> in Just´s Bot. Jahresber. 4: 235. 1876.<br />
= Periconia ellipsospora Penz. & Sacc., Atti Reale Ist. Veneto Sci. Lett. Arti,<br />
Ser. 6, 2: 596. 1884.<br />
= <strong>Cladosporium</strong> paeoniae var. paeoniae-anomalae Sacc., Syll. Fung. 4: 351.<br />
1886.<br />
= Haplographium chlorocephalum var. ovalisporum Ferraris, Fl. Ital. Cryptog.,<br />
Hyphales: 875. 1914.<br />
Descriptions: Mason & Ellis (1953: 123–126), McKemy & Morgan-<br />
Jones (1991: 140–144), Schubert (2005: 216).<br />
Illustrations: Fresenius (1850: Pl. IV, figs 10–15), Mason & Ellis<br />
(1953: 124–125, figs 42–43), McKemy & Morgan-Jones (1991:<br />
137, fig. 1; 141, fig. 2; 139, pl. 1; 143, pl. 2), Schubert (2005: 217,<br />
fig. 113; 275, pl. 34).<br />
96
Dichocladosporium gen. nov.<br />
10 changes<br />
Botryosphaeria ribis DQ316076<br />
Botryosphaeria stevensii AY343484<br />
100<br />
97<br />
70<br />
100<br />
100<br />
100<br />
97<br />
100<br />
100<br />
69<br />
<strong>Cladosporium</strong> tenellum CPC 12053<br />
<strong>Cladosporium</strong> variabile CPC 12753<br />
<strong>Cladosporium</strong> herbarum CPC 11600<br />
<strong>Cladosporium</strong> macrocarpum <strong>CBS</strong> 299.67<br />
<strong>Cladosporium</strong> bruhnei CPC 12211<br />
<strong>Cladosporium</strong> ossifragi <strong>CBS</strong> 842.91<br />
<strong>Cladosporium</strong> iridis <strong>CBS</strong> 138.40<br />
CPC 11383<br />
CPC 11969<br />
<strong>CBS</strong> 213.73<br />
<strong>CBS</strong> 100405<br />
100<br />
91<br />
Mycosphaerella punctiformis AY490763<br />
Passalora arachidicola AF297224<br />
Mycosphaerella aurantia AY509744<br />
Passalora fulva AF393701<br />
Cercospora apii AY840512<br />
Cercospora beticola AY840527<br />
Readeriella mirabilis AY725529<br />
Readeriella novaezel<strong>and</strong>iae DQ267603<br />
Teratosphaeria readeriellophora AY725577<br />
Teratosphaeria microspora AY260098<br />
Trimmatostroma abietis AY559362<br />
Trimmatostroma abietina AJ244267<br />
Trimmatostroma salicis AJ244264<br />
Dichocladosporium chlorocephalum<br />
Fig. 1. One of two equally most parsimonious trees obtained from a heuristic search with 100 r<strong>and</strong>om taxon additions of the ITS sequence alignment. <strong>The</strong> scale bar shows<br />
10 changes, <strong>and</strong> bootstrap support values from 1 000 replicates are shown at the nodes. Thickened lines indicate the strict consensus branches. <strong>The</strong> tree was rooted to two<br />
Botryosphaeria species.<br />
Characters of the cladosporioid morph: Leaf-blotch symptoms on<br />
living leaves amphigenous, variable in shape <strong>and</strong> size, subcircularoval<br />
to irregular, broad, oblong to exp<strong>and</strong>ed, up to 30 mm long<br />
<strong>and</strong> 20 mm wide, at times covering the entire leaf surface, forming<br />
olivaceous-brown to blackish brown patches, rarely violet-brown,<br />
margin usually indefinite, attacked areas turning dry with age,<br />
also occurring on young, green stems. Colonies amphigenous,<br />
punctiform to effuse, loose to dense, caespitose, brown, villose.<br />
Mycelium immersed, subcuticular to intraepidermal; hyphae<br />
sparingly branched, 4–7(–10) µm wide, septate, sometimes with<br />
swellings <strong>and</strong> constrictions, swollen cells up to 13 µm diam,<br />
subhyaline to pale brown, smooth, walls thickened, hyphae<br />
sometimes aggregated; in vitro mycelium at first mainly immersed,<br />
later also superficial, branched, 1–5(–7) µm wide, pluriseptate,<br />
often constricted at septa <strong>and</strong> with swellings <strong>and</strong> constrictions,<br />
therefore irregular in outline, smooth to verruculose or irregularly<br />
rough-walled, loosely verruculose with distinct large warts. Semimacronematous<br />
conidiophores formed on leaf-blotches solitary<br />
or in small, loose groups, arising from internal hyphae or swollen<br />
hyphal cells, erumpent through the cuticle, occasionally emerging<br />
through stomata, erect, straight to somewhat flexuous, oblongcylindrical,<br />
usually unbranched or occasionally branched, 13–80<br />
(–120) × (4–)5–8(–10) µm, slightly attenuated towards the apex,<br />
septate, septa often dense, unconstricted, pale to medium brown,<br />
sometimes paler towards the apex, smooth, thick-walled, wall often<br />
with two distinct layers, often somewhat inflated at the very base,<br />
up to 14 µm diam, occasionally proliferating enteroblastically; in<br />
vitro conidiophores arising laterally from plagiotropous hyphae or<br />
terminally from ascending hyphae, the latter usually appearing<br />
more filiform than those arising laterally from plagiotropous hyphae,<br />
erect, straight to slightly flexuous, cylindrical-oblong, not geniculate,<br />
usually unbranched, rarely with a short lateral prolongation near<br />
the apex, 18–60(–100) × 3–6 µm, slightly attenuated towards<br />
the apex, septate, pale to medium brown or olivaceous-brown,<br />
smooth to asperulate, walls somewhat thickened. Conidiogenous<br />
cells integrated, terminal or intercalary, subcylindrical, 7–45 µm<br />
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97
Schubert et al.<br />
Fig. 2. Dichocladosporium chlorocephalum (HAL 1924 F), periconioid, stem rotting morph. Conidiophores <strong>and</strong> conidia. Scale bar = 10 µm.<br />
Table 1. Isolates subjected to DNA analysis <strong>and</strong> morphological examination.<br />
Species Accession number 1 Host Country Collector GenBank accession<br />
number<br />
Dichocladosporium chlorocephalum <strong>CBS</strong> 213.73; IMI 048108a Paeonia sp. United Kingdom F. Rilstone EU009455<br />
<strong>CBS</strong> 100405 Paeonia sp. New Zeal<strong>and</strong> M. Braithwaite EU009456<br />
<strong>CBS</strong> 121522; CPC 11383 Paeonia delavayi Germany K. Schubert EU009457<br />
<strong>CBS</strong> 121523*; CPC 11969 Paeonia officinalis Germany K. Schubert EU009458<br />
1<br />
<strong>CBS</strong>: Centraalbureau voor Schimmelcultures, Utrecht, <strong>The</strong> Netherl<strong>and</strong>s; CPC: Culture collection of Pedro Crous, housed at <strong>CBS</strong>; IMI: International<br />
Mycological Institute, CABI-Bioscience, Egham, Bakeham Lane, U.K.<br />
*Ex-type cultures.<br />
98
Dichocladosporium gen. nov.<br />
Fig. 3. Dichocladosporium chlorocephalum (<strong>CBS</strong> 121523 = CPC 11969). A–B. Symptoms of the periconioid, stem rotting morph. C–E, H. Macroconidiophores <strong>and</strong> conidia. F–G.<br />
Semi-macronematous conidiophores <strong>and</strong> conidia. I–J. Ramoconidia <strong>and</strong> conidia. K–M. Scanning electron microscopic photographs. K. Conidiophores. L. Conidial chain. M.<br />
Single conidium showing the surface ornamentation <strong>and</strong> scar structure. Scale bars: C–J, L = 10 µm; K = 100 µm; M = 2 µm.<br />
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99
Schubert et al.<br />
Fig. 4. Dichocladosporium chlorocephalum (HAL 2011 F), cladosporioid, leaf-spotting morph. Conidiophores <strong>and</strong> conidia. Scale bar = 10 µm.<br />
long, proliferation sympodial, with one to several conidiogenous<br />
loci, subdenticulate or denticulate, protuberant, terminally broadly<br />
truncate, 1.5–3 µm wide, unthickened or almost so, somewhat<br />
darkened-refractive. Conidia catenate, in simple or branched<br />
chains, polymorphous, small conidia globose, subglobose, broadly<br />
obovoid, 3–9 × 3–5 µm, aseptate, pale to medium brown, smooth,<br />
intercalary conidia limoniform, ellipsoid-fusiform, oblong, 5–23<br />
× 3.5–6.5 µm, 0–2-septate, medium brown, smooth to minutely<br />
verruculose or irregularly rough-walled, large conidia ellipsoid,<br />
oblong-cylindrical, ampulliform, 22–45(–52) × (4.5–)5–8 µm,<br />
0–5-septate, medium brown, smooth to minutely verruculose or<br />
irregularly rough-walled, walls somewhat thickened, hila truncate,<br />
1–3 µm wide, unthickened or almost so, somewhat darkenedrefractive;<br />
occasionally with microcyclic conidiogenesis; in vitro<br />
numerous, polymorphous, catenate, in loosely branched chains,<br />
small conidia globose, subglobose, or obovoid, 3–8 × 3–4 µm,<br />
aseptate, intercalary conidia limoniform to ellipsoid-fusiform, 9–18<br />
× 3.5–4.5 µm, 0–1-septate, large conidia ellipsoid to cylindricaloblong,<br />
14–30(–38) × 3–6(–7) µm, 0–3-septate, pale to medium<br />
brown, asperulate, minutely verruculose to irregularly roughwalled,<br />
walls thickened, hila usually short denticle-like, protuberant,<br />
truncate, in smaller conidia 0.5–1.8 µm wide, in larger conidia<br />
(1.5–)2–3 µm wide, unthickened or almost so but usually darkenedrefractive;<br />
with occasional microcyclic conidiogenesis.<br />
Characters of the periconioid morph: Macronematous conidiophores<br />
formed on faded or dead stems in late autumn, winter or after<br />
overwintering; colonies at first visible as reddish brown streaks, later<br />
turning olivaceous-brown to black, sometimes linear, sometimes<br />
encircling the stems, often occupying large stem segments, effuse,<br />
densely caespitose, velvety. Mycelium immersed, subcuticular<br />
100
Dichocladosporium gen. nov.<br />
to intraepidermal; hyphae at first sparsely branched, 3–7 µm<br />
wide, septate, not constricted at the septa, becoming swollen<br />
<strong>and</strong> wider, up to 11 µm wide, often branched, pale to medium<br />
olivaceous-brown, walls thickened, forming loose to dense hyphal<br />
aggregations; in vitro mycelium immersed to superficial, loosely<br />
branched, 2–6(–7) µm wide, pluriseptate, usually without swellings<br />
<strong>and</strong> constrictions, subhyaline to medium brown or olivaceousbrown,<br />
almost smooth to asperulate or irregularly rough-walled, in<br />
older colonies on PDA up to 10 µm wide, sometimes single hyphal<br />
cells distinctly swollen, up to 16(–20) µm wide, mainly at the base<br />
of conidiophores, sometimes covered by a slime coat or enveloped<br />
in a polysaccharide-like layer. Stromata well-developed, large <strong>and</strong><br />
exp<strong>and</strong>ed, up to about 50–320 µm in length, 15–30 µm deep,<br />
composed of a single to several layers of swollen pale to medium<br />
brown stromatic cells, 5–18 µm diam, thick-walled. Conidiophores<br />
solitary or in loose groups, arising from swollen hyphal cells or<br />
stromata, erumpent through the cuticle, erect, straight, rigid to<br />
slightly flexuous, 150–680 µm long, composed of a subcylindrical<br />
stipe, 13–24 µm wide at the base, slightly attenuated towards<br />
the apex, 5–15 µm just below the branched head, pluriseptate,<br />
not constricted at the septa, young conidiophores pale medium<br />
olivaceous-brown, later medium to usually dark brown, sometimes<br />
slightly paler at the distal end, smooth or almost so, often appearing<br />
somewhat granular, roughened, walls distinctly thickened, 1.5–3(–<br />
4) µm wide; apex with a roughly subglobose to ovoid head, about<br />
35–70 µm diam, composed of dense branchlets <strong>and</strong> ramoconidia,<br />
primary branchlets close to the apex <strong>and</strong> below the first <strong>and</strong><br />
sometimes second <strong>and</strong> third septa, solitary, in pairs or small verticils,<br />
appressed against the stipe or somewhat divergent, subcylindrical<br />
to ellipsoid-oval, aseptate, rarely 1-septate, pale olivaceous to dark<br />
brown, 10–20 × 5–8.5 µm; in vitro conidiophores initially micro<strong>and</strong><br />
semimacronematous, then progressively macronematous<br />
as colonies age, arising laterally from plagiotropous hyphae or<br />
terminally from ascending hyphae, sometimes also from swollen<br />
hyphal cells; micronematous conidiophores filiform, narrowly<br />
cylindrical-oblong, unbranched, up to 150 µm long, 2–3.5 µm wide,<br />
septate, septa often appear to be darkened, pale to pale medium<br />
olivaceous-brown, asperulate, walls slightly thickened; semimacronematous<br />
conidiophores often resembling those formed<br />
by the leaf-blotching (cladosporioid) morph on the natural host,<br />
subcylindrical to cylindrical-oblong, straight to slightly flexuous,<br />
unbranched, rarely branched, (10–)15–120 × 3–5(–6) µm, slightly<br />
attenuated towards the apex, septate, medium brown, minutely<br />
verruculose to irregularly rough-walled, walls more or less thickened;<br />
macronematous conidiophores formed in older cultures on SNA,<br />
PDA <strong>and</strong> also MEA (according to McKemy & Morgan-Jones 1991),<br />
but more prominent on PDA <strong>and</strong> MEA, resembling those formed<br />
by the stem-rotting morph (i.e., the periconioid morph, in planta),<br />
consisting of a long unbranched stipe <strong>and</strong> a subglobose head, but<br />
in culture the heads are often more loosely branched than on the<br />
natural substratum, not always forming a compact head, up to 580<br />
µm long, 5–13 µm wide, attenuated towards the apex, 4–8 µm<br />
just below the branched upper part, somewhat swollen at the base,<br />
septate, medium to very dark brown, minutely verruculose, walls<br />
distinctly thickened, two distinct wall layers visible, 1–2 µm thick.<br />
Conidiogenous cells holoblastic, integrated, terminal, intercalary or<br />
even discrete, ellipsoid to cylindrical or doliiform, subdenticulate,<br />
proliferation sympodial, multilocal, conidiogenous loci truncate,<br />
flat, unthickened, 1–3 µm wide, somewhat darkened-refractive; in<br />
culture conidiogenous loci appearing to be somewhat thickened<br />
<strong>and</strong> distinctly darkened-refractive, 1–2.5(–3) µm wide. Conidia<br />
catenate, in long, branched chains, straight, subglobose, aseptate,<br />
3.5–7 µm diam, or ellipsoid-ovoid, 6–15 × 4–9 µm, 0(–1)-septate,<br />
pale olivaceous to olivaceous-brown, smooth to verruculose (under<br />
the light microscope), hila flat, truncate, unthickened, (0.5–)1–<br />
2(–2.5) µm wide, not darkened, but somewhat refractive; in vitro<br />
conidia numerous, catenate, formed in long, branched chains, small<br />
conidia globose to subglobose, (2–)3–7 × (2–)3–4 µm, aseptate,<br />
intercalary ones ellipsoid-ovoid, 6–16 × 3.5–5 µm, 0(–1)-septate,<br />
secondary ramoconidia ellipsoid to cylindrical-oblong, (13–)<br />
15–34(–47) × (3–)4–6(–7) µm, 0–2-septate, sometimes slightly<br />
constricted at the septa, medium olivaceous-brown, verruculose or<br />
irregularly rough-walled, walls slightly to distinctly thickened, hila<br />
more or less protuberant, subdenticulate to denticulate, in small <strong>and</strong><br />
intercalary conidia 0.5–1(–1.5) µm, in secondary ramoconidia 1–2.5<br />
(–3) µm, unthickened or somewhat thickened, darkened-refractive;<br />
occasional microcyclic conidiogenesis.<br />
Cultural characteristics: Colonies on PDA at first whitish or smoke<br />
grey, reverse smoke-grey to olivaceous-grey, with age smoke-grey<br />
to olivaceous or olivaceous-grey, sometimes even dark mouse-grey,<br />
reverse iron-grey to dark mouse-grey or black, felty; margin white<br />
to smoke-grey, narrow to more or less broad, regular to slightly<br />
undulate, glabrous to somewhat feathery; aerial mycelium at first<br />
mainly in the colony centre, with age abundantly formed, covering<br />
almost the whole colony, whitish, smoke-grey to olivaceous, felty;<br />
growth low convex to raised; numerous small exudates formed,<br />
sometimes becoming prominent; fertile.<br />
Specimens examined: Czechoslovakia, Bohemia, Turnau, on leaves of Paeonia<br />
arborea, 15 Sep. 1905, J.E. Kabát, Kabát & Bubák, Fungi Imperf. Exs. 396, B 70-<br />
6669. France, on dead stems of Paeonia sp., 1901, ex Herbario Musei Parisiensis,<br />
ex herb. Magnus, exs. Desmazières, Pl. Crypt. N. France, Ed. 2, Ser. 1, 1621, HBG,<br />
as “Periconia atra”; Chailly-en-Biere, Seine-et-Marne, Feuilleaubois, on stems of<br />
P. officinalis, 27 Mar. 1881, Roumeguère, Fungi Sel. Gall. Exs. 1803, HBG, as<br />
“Periconia atra”. Germany, Baden-Würtemberg, Kreis Tübingen, Drusslingen, on<br />
leaves of P. officinalis, Jun. 1935, Raabe, B 70-6670; Bayern, Freising, on leaves<br />
of P. officinalis, Sep. 1918, Prof. Dr. J.E. Weiß, Herbarium pathologicum, B 70-<br />
6663; Br<strong>and</strong>enburg, Schloßpark zu Tamsel, on leaves of P. officinalis, 15 Aug.<br />
1924, P. Vogel, Sydow, Mycoth. Germ. 2447, M-57751, PH; Triglitz, on leaves of<br />
P. officinalis, 3 Oct. 1909, Jaap, B 70-6668; Hessen, Frankfurt am Main, botanical<br />
garden, on leaves of P. potaninii, 7 Oct. 2004, R. Kirschner, HAL, RoKi 2222; Kreis<br />
Kassel, Hofgeismar, Garten von Prof. Grupe, on leaves of P. officinalis, 3 Sep.<br />
1947, Schulz, B 70-6658; Mecklenburg-Vorpommern, Rostock, neuer botanischer<br />
Garten, on leaves of P. corallina (= P. mascula), 27 Aug. 1950, Becker, B 70-6662;<br />
Nordrhein-Westfalen, Duisburg, Dinslake, private garden, on leaves of P. anomala,<br />
9 Aug. 2005, N. Ale-Agha, HAL 2014 F; Hamborn, botanical garden, on leaves of P.<br />
obovata, 10 Aug. 2005, N. Ale-Agha, HAL 2017 F; Essen, botanical garden of the<br />
university of Essen, on leaves of P. mlokosewitschii, 10 Aug. 2005, N. Ale-Agha,<br />
HAL 2013 F; on leaves of P. officinalis <strong>and</strong> P. suffruticosa, 11 Aug. 2005, N. Ale-<br />
Agha, HAL 2016, 2017 F; Sachsen, Königstein, in Gärten, verbreitet, on leaves of<br />
P. officinalis, Aug. 1896, W. Krieger, Krieger, Fungi Saxon. Exs. 1545, M-57749;<br />
Aug., Sep. 1896, 1915, W. Krieger, Krieger, Schädliche Pilze, B 70-6666, 70-6667;<br />
Sachsen-Anhalt, Halle (Saale), Botanical Garden, on leaves of P. delavayi, 22<br />
Jun. 2004, K. Schubert, HAL 2011 F, culture deposited at the <strong>CBS</strong>, <strong>CBS</strong> 121522 =<br />
CPC 11383; on leaves of P. officinalis, 22 Jun. 2004, K. Schubert, HAL 2012 F; on<br />
stems of P. officinalis, 16 Mar. 2005, K. Schubert, neotype of Dichocladosporium<br />
chlorocephalum designated here HAL 1924 F, isoneotype <strong>CBS</strong>-H 19869, culture<br />
ex-neotype <strong>CBS</strong> 121523 = CPC 11969; on dead stems of Paeonia sp., Jan. 1873,<br />
G. Winter, Rabenhorst, Fungi Eur. Exs. 1661, HBG, as “Periconia chlorocephala”;<br />
Thüringen, Fürstlicher Park zu Sondershausen, on leaves of P. arborea, 20 Aug.<br />
1903, G. Oertel, Sydow, Mycoth. Germ. 196, PH. Italy, Thümen, Herb. Mycol.<br />
Oecon. 416, on living leaves of Paeonia lactiflora [= P. edulis] (M-57753), lectotype<br />
of “<strong>Cladosporium</strong> paeoniae” designated here; isolectotypes: Thümen, Herb.<br />
Mycol. Oecon. 416; Padova, on leaves of P. officinalis, Aug. 1902, P.A. Saccardo,<br />
Saccardo, Mycoth. Ital. 1186, B 70-6660, SIENA; Parma, on leaves of P. officinalis,<br />
Jul. 1876, Prof. Passerini, Thümen, Mycoth. Univ. 670, B 70-6654, 70-6655, M-<br />
57752; Pavia, botanical garden, on leaves of P. officinalis, summer 1889, Briosi &<br />
Cavara, Fung. Paras. Piante Colt. Utili Ess. 78, M-57748; F. Cavara, Roumeguère,<br />
Fungi Sel. Gall. Exs. 5193, mixed infection with <strong>Cladosporium</strong> herbarum, B 70-<br />
6656; Siena, Hort. Bot., on leaves of Paeonia sp., Nov. 1899, SIENA. Latvia, prov.<br />
Vidzeme, Kreis Riga, Riga, in a garden, on leaves of P. foemina [= P. officinalis], 28<br />
Aug. 1936, J. Smarods, Fungi Lat. Exs. 799, M-57747. New Zeal<strong>and</strong>, isolated from<br />
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Schubert et al.<br />
Fig. 5. Dichocladosporium chlorocephalum (<strong>CBS</strong> 121522 = CPC 11383). A–B. Symptoms on leaves of Paeonia officinalis <strong>and</strong> P. delavayi caused by the cladosporioid, leaf spotting<br />
morph. C–D. Conidiophores <strong>and</strong> conidia. E. Ramoconidia <strong>and</strong> conidia. F–G. Scanning electron microscopic photographs. F. Conidial chain still attached to a conidiophore.<br />
G. Conidia showing surface ornamentation <strong>and</strong> scar structure. Scale bars: C–E, G = 10 µm; F = 5 µm.<br />
red leaf <strong>and</strong> stem lesions on Paeonia sp., M. Braithwaite, <strong>CBS</strong> 100405. Romania,<br />
Râmnicu-Vâlcea, distr. Vâlcea, Oltenia, on leaves of P. officinalis, 17 Aug. 1930,<br />
Tr. Săvulescu & C. S<strong>and</strong>u, Săvulescu, Herb. Mycol. Roman. 298, M-57742. U.K.,<br />
Engl<strong>and</strong>, Cornwall, Lambounce Hill, Perranzubuloe, isolated from dead stems of<br />
Paeonia sp., isol. F. Rilstone, <strong>CBS</strong> 213.73 = IMI 048108a. U.S.A., on leaves of<br />
Paeonia sp., Sep. 1878, Ellis, N. Amer. Fungi 543, B 70-6659, M-57744, PH; Illinois,<br />
Cobden, on leaves of Paeonia sp., 8 Aug. 1882, F.S. Earle, No. 91, B 70-6657;<br />
Kansas, Topeka, on leaves of P. officinalis, 7 Jul. 1922, C.F. Menninger, US Dept.<br />
Agric., Pathol. Mycol. Coll. 60085, B 70-6661, F; Montana, Columbia, on leaves of<br />
P. officinalis, Aug. 1886, B.T. Galloway, Ellis & Everh., N. Amer. Fungi Ser. II, 1991,<br />
PH; on leaves of Paeonia sp., 18 Oct. 1931, W.E. Maneval, F.<br />
Host range <strong>and</strong> distribution: On Paeonia anomala, P. arborea, P.<br />
delavayi, P. hybrida, P. lactiflora, P. mascula, P. mlokosewitschii,<br />
P. moutan, P. obovata, P. obovata var. willmotiae, P. officinalis,<br />
P. potaninii, P. suffruticosa, Paeonia spp. (Paeoniaceae), Asia<br />
(Armenia, China, Georgia, Kazakhstan, Russia), Europe (Belgium,<br />
102
Dichocladosporium gen. nov.<br />
Czechoslovakia, Denmark, France, Germany, Italy, Latvia, Moldova,<br />
Pol<strong>and</strong>, Romania, Switzerl<strong>and</strong>, U.K., Ukraine), North America<br />
(Canada, U.S.A.), New Zeal<strong>and</strong>.<br />
Notes: Type material of Periconia chlorocephala is not preserved<br />
in the herbarium of G. Fresenius at FR (Forschungsinstitut<br />
Senkenberg, Frankfurt a. M., Germany). Hence, a new specimen<br />
collected in the Botanical Garden of the Martin-Luther-University<br />
Halle (Saale), Germany, is proposed to serve as neotype. A culture<br />
derived from this collection is deposited at the <strong>CBS</strong>, Utrecht, the<br />
Netherl<strong>and</strong>s as ex-neotype culture. A leaf-blotch sample, also<br />
collected in the Botanical Garden at Halle (Saale), from which we<br />
also derived a living culture, is designated as representative of the<br />
synanamorph, <strong>Cladosporium</strong> paeoniae. Both cultures have been<br />
used to generate DNA sequence data.<br />
<strong>The</strong> two stages (morphs) of this fungus are usually ecologically<br />
<strong>and</strong> seasonally separated, but sometimes conidiophores of the<br />
leaf-blotching (cladosporioid) morph also occur on dead stems of<br />
peony intermixed with the macronematous conidiophores of the<br />
periconioid morph. In culture conidiophore <strong>and</strong> conidial width tends<br />
to be narrower than on the natural substratum, <strong>and</strong> the conidia are<br />
not as frequently septate.<br />
DISCUSSION<br />
Cultural studies by ourselves <strong>and</strong> McKemy & Morgan-Jones<br />
(1991), <strong>and</strong> molecular sequence analyses documented herein<br />
clearly demonstrate that <strong>Cladosporium</strong> chlorocephalum, occurring<br />
on necrotic stems, <strong>and</strong> C. paeoniae, causing leaf-blotch symptoms<br />
on living leaves of Paeonia spp., are two synanamorphs of a single<br />
species, which has to be excluded from <strong>Cladosporium</strong> s. str. since<br />
the conidiogenous loci are quite distinct from the characteristically<br />
coronate scars in the latter <strong>genus</strong> <strong>and</strong> because ITS sequences<br />
indicate clear separation from <strong>Cladosporium</strong> s. str.<br />
Analysis of the morphology <strong>and</strong> conidiogenesis showed that<br />
the macronematous stage of this fungus (C. chlorocephalum, the<br />
periconioid morph) closely resembles Metulocladosporiella, recently<br />
introduced for the <strong>Cladosporium</strong> speckle disease of banana. <strong>The</strong>re<br />
are, however, some differences. In Metulocladosporiella musae<br />
(E.W. Mason) Crous et al., the type species, micronematous<br />
conidiophores occur in vitro <strong>and</strong> in vivo, <strong>and</strong> macronematous<br />
conidiophores occur on leaf-spots, whereas in C. chlorocephalum<br />
the semi-macronematous conidiophores usually accompany<br />
leaf-blotch symptoms on living leaves <strong>and</strong> the macronematous<br />
conidiophores occur in saprobic growth on old necrotic stems.<br />
Rhizoid hyphae arising from the swollen basal cells of the<br />
macronematous conidiophores are characteristic for M. musae, but<br />
lacking in C. chlorocephalum, <strong>and</strong> the conidia in the latter species<br />
are 0–5-septate, but only 0(–1)-septate in M. musae. <strong>The</strong> semimacronematous,<br />
leaf-blotching stage (the cladosporioid morph)<br />
is barely distinguishable from the present concept of Fusicladium,<br />
which includes species with catenate conidia (Schubert et al. 2003).<br />
However, the peony fungus does not cluster within the Venturiaceae.<br />
Since C. chlorocephalum clusters apart of the Chaetothyriales,<br />
the clade to which Metulocladosporiella belongs, the differences<br />
observed here seem to be sufficient to place this fungus in a new<br />
<strong>genus</strong> (also see Crous et al. 2007 – this volume). Crous et al.<br />
(2006a) discussed differences between Metulocladosporiella <strong>and</strong><br />
allied <strong>dematiaceous</strong> hyphomycete genera <strong>and</strong> provided a key to<br />
the latter <strong>genus</strong> <strong>and</strong> morphologically <strong>similar</strong> genera. Using this key,<br />
attempts to determine the macronematous morph of <strong>Cladosporium</strong><br />
www.studiesinmycology.org<br />
chlorocephalum lead to Metulocladosporiella. Differences between<br />
morphologically <strong>similar</strong> genera have been discussed in the paper<br />
by Crous et al. (2006a) <strong>and</strong> are also valid for the new <strong>genus</strong><br />
Dichocladosporium. Parapericoniella U. Braun, Heuchert & K.<br />
Schub., a fungicolous <strong>genus</strong> recently introduced to accommodate<br />
<strong>Cladosporium</strong> asterinae Deighton, is also morphologically <strong>similar</strong><br />
in having apically, densely branched conidiophores <strong>and</strong> truncate,<br />
unthickened conidiogenous loci <strong>and</strong> hila, but is quite distinct in not<br />
having micronematous conidiophores (Heuchert et al. 2005).<br />
ACKNOWLEDGEMENTS<br />
We are much obliged to the curators of B, F, HBG, M, PH <strong>and</strong> SIENA for the<br />
loans of the collections studied. R. Kirschner <strong>and</strong> N. Ale-Agha are thanked for<br />
sending collections <strong>and</strong> cultures on Paeonia spp. We are very grateful to the<br />
Institute of Zoology of the Martin-Luther-University, above all to G. Tschuch, for<br />
providing access to SEM facilities. This work was supported in part by a grant of<br />
the “Graduiertenförderung des L<strong>and</strong>es Sachsen-Anhalt” <strong>and</strong> a grant of Synthesys<br />
(No. 2559) awarded to K.S. We thank M. Vermaas for preparing the photographic<br />
plates.<br />
REFERENCES<br />
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146.<br />
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Passerini G (1876). <strong>Cladosporium</strong> paeoniae. Just’s Botanische Jahresberichte 4:<br />
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(hyphomycetes). Ph.D. dissertation. Martin-Luther-University Halle-Wittenberg.<br />
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Schubert K, Ritschel A, Braun U (2003). A monograph of Fusicladium s. lat.<br />
(hyphomycetes). Schlechtendalia 9: 1–132.<br />
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enzymatically amplified ribosomal DNA from several Cryptococcus species.<br />
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ex Fr. <strong>CBS</strong>, Baarn.<br />
White TJ, Bruns T, Lee S, Taylor J (1990). Amplification <strong>and</strong> direct sequencing of<br />
fungal ribosomal RNA genes for phylogenetics. In: PCR Protocols: a guide to<br />
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104
available online at www.studiesinmycology.org<br />
doi:10.3114/sim.2007.58.05<br />
Studies in Mycology 58: 105–156. 2007.<br />
Biodiversity in the <strong>Cladosporium</strong> herbarum complex (Davidiellaceae, Capnodiales),<br />
with st<strong>and</strong>ardisation of methods for <strong>Cladosporium</strong> taxonomy <strong>and</strong> diagnostics<br />
K. Schubert 1* , J. Z. Groenewald 2 , U. Braun 3 , J. Dijksterhuis 2 , M. Starink 2 , C.F. Hill 4 , P. Zalar 5 , G.S. de Hoog 2 <strong>and</strong> P.W. Crous 2<br />
1<br />
Botanische Staatssammlung München, Menzinger Strasse 67, D-80638 München, Germany; 2 <strong>CBS</strong> Fungal Biodiversity Centre, P.O. Box 85167, 3508 AD Utrecht, <strong>The</strong> Netherl<strong>and</strong>s;<br />
3<br />
Martin-Luther-Universität, Institut für Biologie, Geobotanik und Botanischer Garten, Herbarium, Neuwerk 21, D-06099 Halle (Saale), Germany; 4 Plant & Environment Laboratory<br />
Biosecurity NZ, Ministry of Agriculture & Forestry, P.O. Box 2095, Auckl<strong>and</strong> 1140, New Zeal<strong>and</strong>; 5 Biotechnical Faculty, Department of Biology, Večna pot 111, SI-1000 Ljubljana,<br />
Slovenia<br />
*Correspondence: Konstanze Schubert, konstanze.schubert@gmx.de<br />
Abstract: <strong>The</strong> <strong>Cladosporium</strong> herbarum complex comprises five species for which Davidiella teleomorphs are known. <strong>Cladosporium</strong> herbarum s. str. (D. tassiana), C.<br />
macrocarpum (D. macrocarpa) <strong>and</strong> C. bruhnei (D. allicina) are distinguishable by having conidia of different width, <strong>and</strong> by teleomorph characters. Davidiella variabile is<br />
introduced as teleomorph of C. variabile, a homothallic species occurring on Spinacia, <strong>and</strong> D. macrospora is known to be the teleomorph of C. iridis on Iris spp. <strong>The</strong> C. herbarum<br />
complex combines low molecular distance with a high degree of clonal or inbreeding diversity. Entities differ from each other by multilocus sequence data <strong>and</strong> by phenetic<br />
differences, <strong>and</strong> thus can be interpreted to represent individual taxa. Isolates of the C. herbarum complex that were formerly associated with opportunistic human infections,<br />
cluster with C. bruhnei. Several species are newly described from hypersaline water, namely C. ramotenellum, C. tenellum, C. subinflatum, <strong>and</strong> C. herbaroides. <strong>Cladosporium</strong><br />
pseudiridis collected from Iris sp. in New Zeal<strong>and</strong>, is also a member of this species complex <strong>and</strong> shown to be distinct from C. iridis that occurs on this host elsewhere in the<br />
world. A further new species from New Zeal<strong>and</strong> is C. sinuosum on Fuchsia excorticata. <strong>Cladosporium</strong> antarcticum is newly described from a lichen, Caloplaca regalis, collected<br />
in Antarctica, <strong>and</strong> C. subtilissimum from grape berries in the U.S.A., while the new combination C. ossifragi, the oldest valid name of the <strong>Cladosporium</strong> known from Narthecium<br />
in Europe, is proposed. St<strong>and</strong>ard protocols <strong>and</strong> media are herewith proposed to facilitate future morphological examination of <strong>Cladosporium</strong> spp. in culture, <strong>and</strong> neotypes or<br />
epitypes are proposed for all species treated.<br />
Taxonomic novelties: <strong>Cladosporium</strong> antarcticum K. Schub., Crous & U. Braun, sp. nov., C. herbaroides K. Schub., Zalar, Crous & U. Braun, sp. nov., C. ossifragi (Rostr.) U.<br />
Braun & K. Schub., comb. nov., C. pseudiridis K. Schub., C.F. Hill, Crous & U. Braun, sp. nov., C. ramotenellum K. Schub., Zalar, Crous & U. Braun, sp. nov., C. sinuosum K.<br />
Schub., C.F. Hill, Crous & U. Braun, sp. nov., C. subinflatum K. Schub., Zalar, Crous & U. Braun, sp. nov., C. subtilissimum K. Schub., Dugan, Crous & U. Braun, sp. nov., C.<br />
tenellum K. Schub., Zalar, Crous & U. Braun sp. nov., Davidiella macrocarpa Crous, K. Schub. & U. Braun, sp. nov., D. variabile Crous, K. Schub. & U. Braun, sp. nov.<br />
Key words: Clonality, Davidiella, homothallism, new species, phylogeny, recombination, taxonomy.<br />
Introduction<br />
<strong>Cladosporium</strong> herbarum (Pers. : Fr.) Link, type species of the <strong>genus</strong><br />
<strong>Cladosporium</strong> Link, is one of the most common environmental fungi<br />
to be isolated worldwide. It abundantly occurs on fading or dead<br />
leaves of herbaceous <strong>and</strong> woody plants, as secondary invader<br />
on necrotic leaf spots, <strong>and</strong> has frequently been isolated from<br />
air (Samson et al. 2000), soil (Domsch et al. 1980), foodstuffs,<br />
paints, textiles, humans (de Hoog et al. 2000) <strong>and</strong> numerous<br />
other substrates. It is also known to occur on old carpophores<br />
of mushrooms <strong>and</strong> other fungi (Heuchert et al. 2005) <strong>and</strong> to be<br />
a common endophyte (Riesen & Sieber 1985, Brown et al. 1998,<br />
El-Morsy 2000), especially in temperate regions. Under favourable<br />
climatic conditions C. herbarum also germinates <strong>and</strong> grows as an<br />
epiphyte on the surface of green, healthy leaves (Schubert 2005).<br />
Persoon (1794) introduced C. herbarum as Dematium<br />
herbarum Pers., which was later reclassified by Link (1809) as<br />
Acladium herbarum (Pers.) Link. In 1816, Link included C. herbarum<br />
together with three additional species in his newly described <strong>genus</strong><br />
<strong>Cladosporium</strong>. Clements & Shear (1931) proposed C. herbarum<br />
as lectotype species of the latter <strong>genus</strong>, a decision followed by de<br />
Vries (1952) <strong>and</strong> Hughes (1958). Several authors provided detailed<br />
treatments of C. herbarum (de Vries 1952, Ellis 1971, Domsch et al.<br />
1980, Prasil & de Hoog 1988), <strong>and</strong> there are literally thous<strong>and</strong>s of<br />
records of this species in the literature. McKemy & Morgan-Jones<br />
(1991) <strong>and</strong> Ho et al. (1999) examined C. herbarum in culture <strong>and</strong><br />
published detailed descriptions of its features in vitro.<br />
<strong>Cladosporium</strong> macrocarpum Preuss, a second component<br />
within the herbarum complex, has hitherto been known <strong>and</strong> treated<br />
as an allied, but morphologically distinct species on the basis of its<br />
wider <strong>and</strong> somewhat larger, frequently 2–3-septate, more regularly<br />
verrucose conidia, shorter conidial chains <strong>and</strong> more pronounced<br />
prolongations of the conidiophores. Dugan & Roberts (1994) carried<br />
out examinations of morphological <strong>and</strong> reproductive aspects of both<br />
species, <strong>and</strong> in so doing demonstrated a morphological continuum<br />
between C. macrocarpum <strong>and</strong> C. herbarum, concluding that the<br />
name herbarum should have preference. <strong>The</strong>refore, Ho et al. (1999)<br />
introduced the new combination C. herbarum var. macrocarpum<br />
(Preuss) M.H.M. Ho & Dugan. Although transitional forms have<br />
been discussed to occur between the two species, several authors<br />
still prefer to retain C. macrocarpum as a separate species.<br />
In an attempt to elucidate the species within the C. herbarum<br />
complex, therefore, a multilocus DNA sequence typing approach<br />
was used, employing five genes, namely the internal transcribed<br />
spacers of the rDNA genes (ITS), actin, calmodulin, translation<br />
elongation factor 1-α, <strong>and</strong> histone H3. <strong>The</strong>se data were<br />
supplemented with morphological examinations under st<strong>and</strong>ardised<br />
conditions, using light <strong>and</strong> scanning electron microscopy, as well as<br />
cultural characteristics <strong>and</strong> growth studies.<br />
Material <strong>and</strong> methods<br />
Isolates<br />
Isolates included in this study were obtained from the culture<br />
collection of the Centraalbureau voor Schimmelcultures (<strong>CBS</strong>),<br />
Utrecht, Netherl<strong>and</strong>s, or were freshly isolated from a range of<br />
different substrates. Single-conidial <strong>and</strong> ascospore isolates were<br />
obtained using the techniques as explained in Crous (1998) for<br />
species of Mycosphaerella Johanson <strong>and</strong> its anamorphs. Isolates<br />
105
Schubert et al.<br />
were inoculated onto 2 % potato-dextrose agar (PDA), synthetic<br />
nutrient-poor agar (SNA), 2 % malt extract agar (MEA) <strong>and</strong> oatmeal<br />
agar (OA) (Gams et al. 2007), <strong>and</strong> incubated under continuous<br />
near-ultraviolet light at 25 °C to promote sporulation. All cultures<br />
obtained in this study are maintained in the culture collection of<br />
the <strong>CBS</strong> (Table 1). Nomenclatural novelties <strong>and</strong> descriptions were<br />
deposited in MycoBank (www.MycoBank.org).<br />
DNA isolation, amplification <strong>and</strong> sequence analysis<br />
Fungal colonies were established on agar plates, <strong>and</strong> genomic<br />
DNA was isolated as described in Gams et al. (2007). Partial gene<br />
sequences were determined as described by Crous et al. (2006)<br />
for actin (ACT), calmodulin (CAL), translation elongation factor 1-<br />
alpha (EF), histone H3 (HIS) <strong>and</strong> part (ITS) of the nuclear rDNA<br />
operon spanning the 3’ end of the 18S rRNA gene (SSU), the<br />
first internal transcribed spacer, the 5.8S rRNA gene, the second<br />
internal transcribed spacer <strong>and</strong> the 5’ end of the 28S rRNA gene<br />
(LSU). <strong>The</strong> nucleotide sequences were generated using both PCR<br />
primers to ensure good quality sequences over the entire length of<br />
the amplicon. Subsequent sequence alignment <strong>and</strong> phylogenetic<br />
analysis followed the methods of Crous et al. (2006). Gaps longer<br />
than 10 bases were coded as single events for the phylogenetic<br />
analyses; the remaining gaps were treated as new character<br />
states. Sequence data were deposited in GenBank (Table 1) <strong>and</strong><br />
the alignment <strong>and</strong> tree in TreeBASE (www.treebase.org).<br />
Data analysis<br />
<strong>The</strong> number of entities in the dataset of 79 strains was inferred<br />
with Structure v. 2.2 software (Pritchard et al. 2000, Falush et al.<br />
2003) using an UPGMA tree of data of the ACT gene compared<br />
with CAL, EF <strong>and</strong> HIS with the exclusion of the nearly invariant ITS<br />
region. For this analysis group indications were derived from a tree<br />
produced with MrAic (Nyl<strong>and</strong>er 2004). <strong>The</strong> length of the burn-in<br />
period was set to 1 000 000, number of MCMC repeats after burn-in<br />
10 000, with admixture ancestry <strong>and</strong> allele frequencies correlated<br />
models, assuming that all groups diverged from a recent ancestral<br />
population <strong>and</strong> that allele frequencies are due to drift. Uniform prior<br />
for ALPHA was set to 1.0 (default) <strong>and</strong> allele frequencies with λ set to<br />
1.0 (default). <strong>The</strong> numbers of MCMC repetitions after burn-in were<br />
set as 10 000 <strong>and</strong> 100 000. <strong>The</strong> number of clusters (K) in Structure<br />
was assumed from 5 to 7. Population differentiation F ST<br />
(index: θ)<br />
was calculated with 1–6 runs using the same software. <strong>The</strong> null<br />
hypothesis for this analysis is no population differentiation. When<br />
observed theta (θ) is significantly different from those of r<strong>and</strong>om<br />
data sets (p < 0.05), population differentiation is considered.<br />
Association of multilocus genotypes was screened with the<br />
multilocus option in BioNumerics v. 4.5. To test for reproductive<br />
mode in each population, the st<strong>and</strong>ardised index of association<br />
(I S ; Haubold et al. 1998) was calculated with start2 software<br />
A<br />
(Jolley et al. 2001). <strong>The</strong> null hypothesis for this analysis is complete<br />
panmixia. <strong>The</strong> values of I S were compared between observed <strong>and</strong><br />
A<br />
r<strong>and</strong>omised datasets. <strong>The</strong> hypothesis would be rejected when p <<br />
0.05. Mean genetic diversity (H) <strong>and</strong> diversities of individual loci<br />
were calculated with lian v. 3.5 (Haubold & Hudson 2000). Degrees<br />
of recombination or horizontal gene transfer were also visualised<br />
using SplitsTree v. 4.8 software (Huson & Bryant 2006). Split<br />
decomposition was carried out with default settings, i.e., character<br />
transformation using uncorrected (observed, “P”) distances, splits<br />
transformation using “equal angle”, <strong>and</strong> optimise boxes iteration set<br />
to 2.<br />
Morphology<br />
As the present study represents the first in a series dealing<br />
with <strong>Cladosporium</strong> spp. <strong>and</strong> their Davidiella Crous & U. Braun<br />
teleomorphs in culture, a specific, st<strong>and</strong>ardised protocol was<br />
established by which all species complexes will be treated in<br />
future.<br />
Morphology of the anamorph: Microscopic observations were made<br />
from colonies cultivated for 7 d under continuous near-ultraviolet<br />
light at 25 °C on SNA. Preparations were mounted in Shear’s<br />
solution (Gams et al. 2007). To study conidial development <strong>and</strong><br />
branching patterns, squares of transparent adhesive tape (Titan<br />
Ultra Clear Tape, Conglom Inc., Toronto, Canada) were placed on<br />
conidiophores growing in the zone between the colony margin <strong>and</strong><br />
2 cm inwards, <strong>and</strong> mounted between two drops of Shear’s solution<br />
under a glass coverslip. Different types of conidia are formed by<br />
<strong>Cladosporium</strong> species for which different terms need to be adopted.<br />
Ramoconidia are conidia with usually more than one (mostly 2<br />
or 3) conidial hilum, which typically accumulate at the tip of these<br />
conidia. Conidiogenous cells with more than one conidiogenous<br />
locus are first formed as apical parts of conidiophores. Such apical<br />
Fig. 1. <strong>Cladosporium</strong> conidiophore with ramoconidia, secondary ramoconidia,<br />
intercalary conidia, <strong>and</strong> small, terminal conidia. Scale bar = 10 µm. K. Schubert<br />
del.<br />
106
<strong>Cladosporium</strong> herbarum species complex<br />
Aculeate<br />
Spinulose<br />
Digitate<br />
Muricate<br />
Granulate<br />
Colliculate<br />
Pustulate<br />
Pedicellate<br />
Fig. 2. Terms used to describe conidium wall ornamentation under the cryo-electron<br />
microscope. Adapted from David (1997).<br />
than the small terminal conidia. In older literature true ramoconidia<br />
were often cited as “ramoconidia s. str.”, whereas secondary<br />
ramoconidia have been referred to as “ramoconidia s. lat.”<br />
Morphology of the teleomorph: Teleomorphs were induced by<br />
inoculating plates of 2 % tap water agar onto which autoclaved<br />
stem pieces of Urtica dioica (European stinging nettle) were placed.<br />
Inoculated plates were incubated on the laboratory bench for 7 d,<br />
after that period they were further incubated at 10 °C in the dark for<br />
1–2 mo to stimulate teleomorph development. Wherever possible,<br />
30 measurements (× 1 000 magnification) were made of conidia<br />
<strong>and</strong> ascospores, with the extremes of spore measurements given<br />
in parentheses. Cultural characteristics: Colonies were cultivated<br />
on PDA, MEA <strong>and</strong> OA plates for 14 d at 25 °C in the dark, after<br />
which the surface <strong>and</strong> reverse colours were rated using the charts<br />
of Rayner (1970). Linear growth was determined on MEA, PDA <strong>and</strong><br />
OA plates by inoculating three plates per isolate for each medium,<br />
<strong>and</strong> incubating them for 14 d at 25 ºC, after that period colony<br />
diameters were determined.<br />
Low-temperature scanning electron microscopy<br />
Isolates of <strong>Cladosporium</strong> spp. were grown on SNA with 30 g agar/L<br />
for 3–4 d at room temperature under black light. Relevant parts of<br />
the small colonies with conidiophores <strong>and</strong> conidia were selected<br />
under a binocular, excised with a surgical blade as small agar (3<br />
× 3 mm) blocks, <strong>and</strong> transferred to a copper cup for snap-freezing<br />
in nitrogen slush. Agar blocks were glued to the copper surface<br />
with frozen tissue medium (KP-Cryoblock, Klinipath, Duiven,<br />
Netherl<strong>and</strong>s) mixed with 1 part colloidal graphite (Agar Scientific,<br />
Stansted, U.K.). Samples were examined in a JEOL 5600LV<br />
scanning electron microscope (JEOL, Tokyo, Japan) equipped<br />
with an Oxford CT1500 Cryostation for cryo-electron microscopy<br />
(cryoSEM). Electron micrographs were acquired from uncoated<br />
frozen samples, or after sputter-coating by means of a gold/<br />
palladium target for 3 times during 30 s (Fig. 2). Micrographs of<br />
uncoated samples were taken at an acceleration voltage of 3 kV,<br />
<strong>and</strong> consisted out of 30 averaged fast scans (SCAN 2 mode), <strong>and</strong><br />
at 5 kV in case of the coated samples (PHOTO mode).<br />
parts of conidiophores are called ramoconidia if they secede at<br />
a septum from the conidiophore (Kirk et al. 2001). <strong>The</strong> septum at<br />
which the ramoconidium secedes often appears to be somewhat<br />
refractive or darkened. Ramoconidia are characterised by having<br />
a truncate, undifferentiated base (thus they lack a differentiated,<br />
coronate basal hilum formed in the context of conidiogenesis)<br />
<strong>and</strong> they can be very long, aseptate to sometimes multi-septate.<br />
Although they were formed initially as part of the conidiophore,<br />
they function as propagules. Only few of the species known<br />
until now have the ability to form true ramoconidia. Secondary<br />
ramoconidia also have more than one distal conidial hilum but<br />
they always derive from a conidiogenous locus of an earlier formed<br />
cell, which can be either a conidiogenous cell or a ramoconidium.<br />
Secondary ramoconidia are often shorter but somewhat wider than<br />
ramoconidia; they are often septate, <strong>and</strong> typically have a narrowed<br />
base with a coronate hilum (Fig. 1). Conidia in <strong>Cladosporium</strong> are<br />
cells with a coronate basal hilum, which is formed in the context of<br />
conidiogenesis <strong>and</strong> with either a single (when formed as intercalary<br />
units in unbranched parts of chains) or without any distal conidial<br />
hilum (when formed at the tip of conidial chains). For the first, the<br />
term “intercalary conidium” <strong>and</strong> for the latter, “small terminal<br />
conidium” is used. Intercalary conidia typically are larger <strong>and</strong> more<br />
pigmented <strong>and</strong> have a more differentiated surface ornamentation<br />
RESULTS<br />
Phylogeny <strong>and</strong> differentiation<br />
<strong>The</strong> manually adjusted alignment contained 80 sequences (including<br />
the outgroup sequence) <strong>and</strong> the five loci were represented by a<br />
total of 1 516 characters including alignment gaps which were<br />
used in the analysis. Of the 1 516 characters, 369 were parsimonyinformative,<br />
259 were variable <strong>and</strong> parsimony-uninformative, <strong>and</strong><br />
888 were constant.<br />
Forty equally most parsimonious trees (TL = 1 933 steps; CI<br />
= 0.569; RI = 0.786; RC = 0.447), one of which is shown in Fig.<br />
3, were obtained from the parsimony analysis of the combined<br />
genes. Neighbour-joining analysis using three substitution models<br />
(uncorrected “p”, Kimura 2-parameter <strong>and</strong> HKY85) on the sequence<br />
data yielded trees with identical topologies. <strong>The</strong>se differed from<br />
the tree presented in Fig. 3 with regard to the placement of C.<br />
macrocarpum strain CPC 12054 which was placed as a sister<br />
branch to the C. bruhnei Linder clade in the distance analyses<br />
(results not shown) because it shares an identical CAL sequence.<br />
All cryptic species consisting of multiple strains are clustering in<br />
well-supported clades with bootstrap support values ranging from<br />
71 % (C. herbarum) to 100 % [e.g. C. ramotenellum K. Schub.,<br />
www.studiesinmycology.org<br />
107
Schubert et al.<br />
Table 1. Isolates subjected to DNA sequence analyses <strong>and</strong> morphological examinations.<br />
Anamorph Teleomorph Accession number 1 Host Country Collector GenBank numbers 2<br />
(ITS, EF, ACT, CAL, HIS)<br />
<strong>Cladosporium</strong> antarcticum — <strong>CBS</strong> 690.92* (ex-type) Caloplaca regalis Antarctica C. Möller EF679334, EF679405, EF679484, EF679560, EF679636<br />
<strong>Cladosporium</strong> bruhnei Davidiella allicina <strong>CBS</strong> 134.31 = ATCC 11283 — Germany — EF679335, EF679406, EF679485, EF679561, EF679637<br />
<strong>CBS</strong> 157.82 Quercus robur Belgium — EF679336, EF679407, EF679486, EF679562, EF679638<br />
<strong>CBS</strong> 159.54 = ATCC 36948 Man, skin <strong>The</strong> Netherl<strong>and</strong>s — EF679337, EF679408, EF679487, EF679563, EF679639<br />
<strong>CBS</strong> 161.55 Man, sputum <strong>The</strong> Netherl<strong>and</strong>s — EF679338, EF679409, EF679488, EF679564, EF679640<br />
<strong>CBS</strong> 177.71 Thuja tincture <strong>The</strong> Netherl<strong>and</strong>s — EF679339, EF679410, EF679489, EF679565, EF679641<br />
<strong>CBS</strong> 188.54 = ATCC 11290 = IMI 049638 = CPC — — — AY251077, EF679411, EF679490, EF679566, EF679642<br />
3686<br />
<strong>CBS</strong> 366.80 Man, skin <strong>The</strong> Netherl<strong>and</strong>s — EF679340, EF679412, EF679491, EF679567, EF679643<br />
<strong>CBS</strong> 399.80 Man, skin <strong>The</strong> Netherl<strong>and</strong>s — AJ244227, EF679413, EF679492, EF679568, EF679644<br />
<strong>CBS</strong> 521.68 Air <strong>The</strong> Netherl<strong>and</strong>s — EF679341, EF679414, EF679493, EF679569, EF679645<br />
<strong>CBS</strong> 572.78 Polyporus radiatus Russia V.K. Melnik DQ289799, EF679415, DQ289866, DQ289831, EF679646<br />
<strong>CBS</strong> 813.71 Polygonatum odoratum Czech Republic — EF679342, EF679416, EF679494, EF679570, EF679647<br />
<strong>CBS</strong> 110024 Industrial water Germany, Nordrhein-Westfalen — EF679343, EF679417, EF679495, EF679571, EF679648<br />
<strong>CBS</strong> 115683 = ATCC 66670 = CPC 5101 CCA-treated Douglas-fire pole U.S.A., New York C.J. Wang AY361959, EF679418, AY752193, AY752224, AY752255<br />
<strong>CBS</strong> 121624* = CPC 12211 (neotype) Hordeum vulgare Belgium J.Z. Groenewald EF679350, EF679425, EF679502, EF679578, EF679655<br />
CPC 11386 Tilia cordata Germany, Sachsen-Anhalt K. Schubert EF679344, EF679419, EF679496, EF679572, EF679649<br />
CPC 11840 Ourisia macrophylla New Zeal<strong>and</strong> A. Blouin EF679345, EF679420, EF679497, EF679573, EF679650<br />
CPC 12042 = EXF-389 Hypersaline water from salterns Slovenia P. Zalar EF679346, EF679421, EF679498, EF679574, EF679651<br />
CPC 12045 = EXF-594 Hypersaline water from salterns Spain P. Zalar EF679347, EF679422, EF679499, EF679575, EF679652<br />
CPC 12046 = EXF-680 Air conditioning system Slovenia P. Zalar EF679348, EF679423, EF679500, EF679576, EF679653<br />
CPC 12139 Hordeum vulgare <strong>The</strong> Netherl<strong>and</strong>s — EF679349, EF679424, EF679501, EF679577, EF679654<br />
CPC 12212 Hordeum vulgare Belgium J.Z. Groenewald EF679351, EF679426, EF679503, EF679579, EF679656<br />
CPC 12921 Eucalyptus sp. Australia — EF679352, EF679427, EF679504, EF679580, EF679657<br />
<strong>Cladosporium</strong> cladosporioides — <strong>CBS</strong> 673.69 Air <strong>The</strong> Netherl<strong>and</strong>s — EF679353, EF679428, EF679505, EF679581, EF679658<br />
complex<br />
Davidiella sp. <strong>CBS</strong> 109082 Silene maritima United Kingdom A. Aptroot EF679354, EF679429, EF679506, EF679582, EF679659<br />
— CPC 11606 Musa sp. India M. Arzanlou EF679355, EF679430, EF679507, EF679583, EF679660<br />
— CPC 11609 Musa sp. India M. Arzanlou EF679356, EF679431, EF679508, EF679584, EF679661<br />
<strong>Cladosporium</strong> herbaroides — <strong>CBS</strong> 121626* = CPC 12052 = EXF-1733 (ex-type) Hypersaline water from salterns Israel P. Zalar EF679357, EF679432, EF679509, EF679585, EF679662<br />
<strong>Cladosporium</strong> herbarum Davidiella tassiana <strong>CBS</strong> 111.82 Arctostaphylos uva-ursi Switzerl<strong>and</strong> E. Müller AJ238469, EF679433, EF679510, EF679586, EF679663<br />
<strong>CBS</strong> 300.49 Biscutella laevigata Switzerl<strong>and</strong> J.A. von Arx EF679358, EF679434, EF679511, EF679587, EF679664<br />
<strong>CBS</strong> 121621* = CPC 12177 (epitype) Hordeum vulgare <strong>The</strong> Netherl<strong>and</strong>s — EF679363, EF679440, EF679516, EF679592, EF679670<br />
CPC 11600 Delphinium barbeyi U.S.A., Colorado A. Ramalay DQ289800, EF679435, DQ289867, DQ289832, EF679665<br />
CPC 11601 Delphinium barbeyi U.S.A., Colorado A. Ramalay EF679359, EF679436, EF679512, EF679588, EF679666<br />
CPC 11602 Delphinium barbeyi U.S.A., Colorado A. Ramalay EF679360, EF679437, EF679513, EF679589, EF679667<br />
CPC 11603 Delphinium barbeyi U.S.A., Colorado A. Ramalay EF679361, EF679438, EF679514, EF679590, EF679668<br />
108
<strong>Cladosporium</strong> herbarum species complex<br />
<strong>Cladosporium</strong> iridis Davidiella<br />
macrospora<br />
<strong>Cladosporium</strong> macrocarpum Davidiella<br />
macrocarpa<br />
CPC 11604 Delphinium barbeyi U.S.A., Colorado A. Ramalay EF679362, EF679439, EF679515, EF679591, EF679669<br />
CPC 12178 Hordeum vulgare <strong>The</strong> Netherl<strong>and</strong>s — EF679364, EF679441, EF679517, EF679593, EF679671<br />
CPC 12179 Hordeum vulgare <strong>The</strong> Netherl<strong>and</strong>s — EF679365, EF679442, EF679518, EF679594, EF679672<br />
CPC 12180 Hordeum vulgare <strong>The</strong> Netherl<strong>and</strong>s — EF679366, EF679443, EF679519, EF679595, EF679673<br />
CPC 12181 Hordeum vulgare <strong>The</strong> Netherl<strong>and</strong>s — EF679367, EF679444, EF679520, EF679596, EF679674<br />
CPC 12183 Hordeum vulgare <strong>The</strong> Netherl<strong>and</strong>s — EF679368, EF679445, EF679521, EF679597, EF679675<br />
<strong>CBS</strong> 107.20 Iris sp. — — EF679369, EF679446, EF679522, EF679598, EF679676<br />
<strong>CBS</strong> 138.40* (epitype) Iris sp. <strong>The</strong> Netherl<strong>and</strong>s — EF679370, EF679447, EF679523, EF679599, EF679677<br />
<strong>CBS</strong> 175.82 Water Romania — EF679371, EF679448, EF679524, EF679600, EF679678<br />
<strong>CBS</strong> 223.31 = ATCC 11287 Mycosphaerella tulasnei — — AF222830, EF679449, EF679525, EF679601, EF679679<br />
<strong>CBS</strong> 299.67 Triticum aestivum Turkey — EF679372, EF679450, EF679526, EF679602, EF679680<br />
<strong>CBS</strong> 121811* = CPC 12755 (neotype) Spinacia oleracea U.S.A. — EF679376, EF679454, EF679530, EF679606, EF679684<br />
CPC 11817 Corylus sp. U.S.A. — EF679373, EF679451, EF679527, EF679603, EF679681<br />
CPC 12054 = EXF-2287 Hypersaline water from salterns Slovenia P. Zalar EF679374, EF679452, EF679528, EF679604, EF679682<br />
<strong>CBS</strong> H-19855 = CPC 12752 = <strong>CBS</strong> 121623 Spinacia oleracea U.S.A. — EF679375, EF679453, EF679529, EF679605, EF679683<br />
CPC 12756 Spinacia oleracea U.S.A. — EF679377, EF679455, EF679531, EF679607, EF679685<br />
CPC 12757 Spinacia oleracea U.S.A. — EF679378, EF679456, EF679532, EF679608, EF679686<br />
CPC 12758 Spinacia oleracea U.S.A. — EF679379, EF679457, EF679533, EF679609, EF679687<br />
CPC 12759 Spinacia oleracea U.S.A. — EF679380, EF679458, EF679534, EF679610, EF679688<br />
<strong>Cladosporium</strong> ossifragi — <strong>CBS</strong> 842.91* (epitype) Narthecium ossifragum Norway M. di Menna EF679381, EF679459, EF679535, EF679611, EF679689<br />
<strong>CBS</strong> 843.91 Narthecium ossifragum Norway M. di Menna EF679382, EF679460, EF679536, EF679612, EF679690<br />
<strong>Cladosporium</strong> pseudiridis — <strong>CBS</strong> 116463* = LYN 1065 = ICMP 15579 (ex-type) Iris sp. New Zeal<strong>and</strong> C.F. Hill EF679383, EF679461, EF679537, EF679613, EF679691<br />
<strong>Cladosporium</strong> ramotenellum — <strong>CBS</strong> 121628* = CPC 12043 = EXF-454 (ex-type) Hypersaline water from salterns Slovenia P. Zalar EF679384, EF679462, EF679538, EF679614, EF679692<br />
CPC 12047 = EXF-967 Air conditioning system Slovenia P. Zalar EF679385, EF679463, EF679539, EF679615, EF679693<br />
Fuchsia excorticata New Zeal<strong>and</strong> A. Blouin EF679386, EF679464, EF679540, EF679616, EF679694<br />
<strong>Cladosporium</strong> sinuosum — <strong>CBS</strong> 121629* = CPC 11839 = ICMP 15819 (extype)<br />
<strong>Cladosporium</strong> spinulosum — <strong>CBS</strong> 102044 Hypersaline water from salterns Slovenia S. Soujak EF679387, EF679465, EF679541, EF679617, EF679695<br />
<strong>CBS</strong> 119907* = CPC 12040 = EXF-334 (ex-type) Hypersaline water from salterns Slovenia P. Zalar EF679388, EF679466, EF679542, EF679618, EF679696<br />
<strong>Cladosporium</strong> subinflatum — <strong>CBS</strong> 121630* = CPC 12041 = EXF-343 (ex-type) Hypersaline water from salterns Slovenia P. Zalar EF679389, EF679467, EF679543, EF679619, EF679697<br />
<strong>Cladosporium</strong> sp. — <strong>CBS</strong> 172.52 = ATCC 11320 Carya illinoensis U.S.A. — EF679390, EF679468, EF679544, EF679620, EF679698<br />
<strong>CBS</strong> 113741 Grape berry U.S.A. — EF679391, EF679469, EF679545, EF679621, EF679699<br />
<strong>CBS</strong> 113742 Grape berry U.S.A. — EF679392, EF679470, EF679546, EF679622, EF679700<br />
<strong>CBS</strong> 113744 Grape bud U.S.A. — EF679393, EF679471, EF679547, EF679623, EF679701<br />
CPC 12484 Pinus ponderosa Argentina A. Greslebin EF679394, EF679472, EF679548, EF679624, EF679702<br />
CPC 12485 Pinus ponderosa Argentina A. Greslebin EF679395, EF679473, EF679549, EF679625, EF679703<br />
<strong>Cladosporium</strong> subtilissimum — <strong>CBS</strong> 113753 Bing cherry fruits U.S.A. — EF679396, EF679474, EF679550, EF679626, EF679704<br />
<strong>CBS</strong> 113754* Grape berry U.S.A. — EF679397, EF679475, EF679551, EF679627, EF679705<br />
CPC 12044 = EXF-462 Hypersaline water from salterns Slovenia P. Zalar EF679398, EF679476, EF679552, EF679628, EF679706<br />
<strong>Cladosporium</strong> tenellum — <strong>CBS</strong> 121634* = CPC 12053 = EXF-1735 (ex-type) Hypersaline water from salterns Israel P. Zalar EF679401, EF679479, EF679555, EF679631, EF679709<br />
www.studiesinmycology.org<br />
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Schubert et al.<br />
Table 1. (Continued).<br />
Anamorph Teleomorph Accession number 1 Host Country Collector GenBank numbers 2<br />
(ITS, EF, ACT, CAL, HIS)<br />
CPC 11813 Phyllactinia sp. on Corylus sp. U.S.A. D. Glawe EF679399, EF679477, EF679553, EF679629, EF679707<br />
CPC 12051 = EXF-1083 Hypersaline water from salterns Israel P. Zalar EF679400, EF679478, EF679554, EF679630, EF679708<br />
<strong>Cladosporium</strong> variabile Davidiella variabile <strong>CBS</strong> 121636* = CPC 12751 (epitype) Spinacia oleracea U.S.A. — EF679402, EF679480, EF679556, EF679632, EF679710<br />
CPC 12753 Spinacia oleracea U.S.A. — EF679403, EF679481, EF679557, EF679633, EF679711<br />
— Davidiella sp. <strong>CBS</strong> 289.49 Allium schoenoprasum Switzerl<strong>and</strong> E. Müller AY152552, EF679482, EF679558, EF679634, EF679712<br />
<strong>CBS</strong> 290.49 Trisetum distichophyllum Switzerl<strong>and</strong> E. Müller EF679404, EF679483, EF679559, EF679635, EF679713<br />
1 ATCC: American Type Culture Collection, Virginia, U.S.A.; <strong>CBS</strong>: Centraalbureau voor Schimmelcultures, Utrecht, <strong>The</strong> Netherl<strong>and</strong>s; CPC: Culture collection of Pedro Crous, housed at <strong>CBS</strong>; EXF: Extremophilic Fungi Culture Collection of the Department<br />
of Biology, Biotechnical Faculty, University of Ljubljana, Slovenia; ICMP: International Collection of Micro-organisms from Plants, L<strong>and</strong>care Research, Private Bag 92170, Auckl<strong>and</strong>, New Zeal<strong>and</strong>; IMI: International Mycological Institute, CABI-Bioscience,<br />
Egham, Bakeham Lane, U.K.<br />
2 ACT: partial actin gene, CAL: partial calmodulin gene, EF: partial elongation factor 1-alpha gene, HIS: partial histone H3 gene, ITS: internal transcribed spacer region.<br />
*Ex-type cultures.<br />
Zalar, Crous & U. Braun <strong>and</strong> C. ossifragi (Rostr.) U. Braun & K.<br />
Schub.]. <strong>The</strong> intraspecific variation in the C. bruhnei clade is due<br />
to genetic variation present in the sequence data of all loci except<br />
for ITS, those in the C. macrocarpum clade in all loci except for ITS<br />
<strong>and</strong> ACT, <strong>and</strong> those in the C. herbarum clade in all loci except for<br />
ITS <strong>and</strong> CAL (data not shown). However, none of the variation for<br />
these species could be linked to host specificity or morphological<br />
differences. In general, ITS data did not provide any resolution<br />
within the C. herbarum complex, whereas EF data provided species<br />
clades with very little intraspecific variation <strong>and</strong> ACT, CAL <strong>and</strong> HIS<br />
revealed increasing intraspecific variation (ACT the least <strong>and</strong> HIS<br />
the most).<br />
<strong>The</strong> mean genetic diversity (H) of the entire data set excluding<br />
the nearly invariant ITS region was 0.9307, with little difference<br />
between genes (ACT = 0.9257, CAL = 0.9289, EF = 0.9322, HIS<br />
= 0.9361). <strong>The</strong> loci showed different numbers of alleles (ACT: 22,<br />
CAL: 16, EF: 21, HIS: 20, ITS: 6). Differentiation of entities when<br />
calculated with Structure software using the admixture/correlated<br />
model showed highest value with K = 6. At this value F ST<br />
varied<br />
between 0.1362 <strong>and</strong> 0.3381. Linkage disequilibrium calculated<br />
using the st<strong>and</strong>ardised index of association (I S ) for the entire<br />
A<br />
dataset (observed variance V o<br />
= 0.5602, expected variance V e<br />
=<br />
0.2576) was 0.3914 (P = 0.0001), consistent with a small amount<br />
of recombination that did not destroy the linkage between alleles.<br />
Only few groups appeared to be separated for all alleles; degrees<br />
of gene flow are indicated in Fig. 4. SplitsTree software produced<br />
unresolved star-shaped structures for all genes, without any sign of<br />
reticulation (Fig. 5).<br />
110
<strong>Cladosporium</strong> herbarum species complex<br />
100<br />
74<br />
100<br />
10 changes<br />
Cercospora beticola CPC11557<br />
CPC 11609<br />
<strong>CBS</strong> 109082<br />
<strong>CBS</strong> 673.69<br />
<strong>Cladosporium</strong> cladosporioides complex<br />
98<br />
CPC 11606<br />
Davidiella sp. <strong>CBS</strong> 290.49<br />
100 CPC 12043<br />
CPC 12047 <strong>Cladosporium</strong> ramotenellum<br />
100 <strong>CBS</strong> 842.91<br />
80<br />
<strong>Cladosporium</strong> ossifragi<br />
<strong>CBS</strong> 843.91<br />
Davidiella sp. <strong>CBS</strong> 289.49<br />
CPC 12053<br />
91<br />
100<br />
CPC 11813 <strong>Cladosporium</strong> tenellum<br />
64<br />
CPC 12051<br />
<strong>Cladosporium</strong> sp. <strong>CBS</strong> 113744<br />
63<br />
54<br />
94<br />
100<br />
98<br />
72 62<br />
81<br />
66<br />
67<br />
54<br />
67<br />
91<br />
51<br />
<strong>CBS</strong> 113753<br />
<strong>CBS</strong> 113754<br />
CPC 12044<br />
CPC 12139<br />
<strong>CBS</strong> 159.54<br />
<strong>CBS</strong> 188.54<br />
<strong>CBS</strong> 177.71<br />
<strong>CBS</strong> 399.80<br />
<strong>CBS</strong> 521.68<br />
63<br />
<strong>CBS</strong> 115683<br />
<strong>CBS</strong> 366.80<br />
<strong>CBS</strong> 134.31<br />
<strong>CBS</strong> 813.71<br />
<strong>CBS</strong> 110024<br />
CPC 12921<br />
CPC 12046<br />
<strong>CBS</strong> 572.78<br />
CPC 12045<br />
CPC 11840<br />
<strong>CBS</strong> 157.82<br />
CPC 12212<br />
CPC 12211<br />
CPC 12042<br />
CPC 11386<br />
<strong>CBS</strong> 161.55<br />
<strong>Cladosporium</strong> sp. CPC 12485<br />
74<br />
83<br />
58<br />
85<br />
<strong>CBS</strong> 102044<br />
CPC 12040<br />
<strong>Cladosporium</strong> sp. CPC 12484<br />
<strong>Cladosporium</strong> sp. <strong>CBS</strong> 172.52<br />
<strong>Cladosporium</strong> sp. <strong>CBS</strong> 113741<br />
<strong>Cladosporium</strong> antarcticum <strong>CBS</strong> 690.92<br />
<strong>Cladosporium</strong> sp. <strong>CBS</strong> 113742<br />
<strong>Cladosporium</strong> subinflatum CPC 12041<br />
<strong>Cladosporium</strong> pseudiridis <strong>CBS</strong> 116463<br />
<strong>Cladosporium</strong> sinuosum CPC 11839<br />
93<br />
100<br />
51<br />
66<br />
81<br />
94<br />
65<br />
<strong>CBS</strong> 107.20 <strong>Cladosporium</strong> iridis<br />
<strong>CBS</strong> 138.40<br />
<strong>Cladosporium</strong> herbaroides CPC 12052<br />
100<br />
64<br />
95<br />
71<br />
61<br />
99<br />
CPC 12751<br />
CPC 12753<br />
CPC 11817<br />
CPC 12758<br />
CPC 12759<br />
CPC 12054<br />
<strong>CBS</strong> 175.82<br />
<strong>CBS</strong> 223.31<br />
51 CPC 12757<br />
56 <strong>CBS</strong> 299.67<br />
CPC 12756<br />
67 CPC 12755<br />
CPC 12752<br />
CPC 11603<br />
<strong>CBS</strong> 300.49<br />
CPC 11600<br />
CPC 11604<br />
CPC 11602<br />
CPC 11601<br />
<strong>CBS</strong> 111.82<br />
CPC 12177<br />
CPC 12183<br />
100<br />
CPC 12181<br />
CPC 12180<br />
CPC 12179<br />
CPC 12178<br />
<strong>Cladosporium</strong> subtilissimum<br />
<strong>Cladosporium</strong> bruhnei<br />
<strong>Cladosporium</strong> spinulosum<br />
<strong>Cladosporium</strong> variabile<br />
<strong>Cladosporium</strong> macrocarpum<br />
<strong>Cladosporium</strong> herbarum<br />
Fig. 3. One of 40 equally most parsimonious trees obtained from a heuristic search with 100 r<strong>and</strong>om taxon additions of the combined sequence alignment (ITS, ACT, CAL, EF,<br />
HIS). <strong>The</strong> scale bar shows ten changes, <strong>and</strong> bootstrap support values from 1 000 replicates are shown at the nodes. Thickened lines indicate the strict consensus branches<br />
<strong>and</strong> strain numbers in bold represent ex-type sequences. <strong>The</strong> tree was rooted to sequences of Cercospora beticola strain CPC 11557 (GenBank accession numbers AY840527,<br />
AY840458, AY840425, AY840494, AY840392, respectively).<br />
www.studiesinmycology.org<br />
111
Schubert et al.<br />
Davidiella sp.<br />
Global (Gapcost:0%)<br />
ACT<br />
80<br />
85<br />
D. macrosporum D. tassiana<br />
D. macrocarpa<br />
D. macrocarpa<br />
90<br />
D. macrospora<br />
Davidiella sp.<br />
D. variabile<br />
D. Variabile<br />
D. tassiana<br />
D. allicina<br />
D. alliacea<br />
95<br />
100<br />
• . <strong>CBS</strong> 673.69<br />
• . CPC 11606<br />
• . <strong>CBS</strong> 109082<br />
• . CPC 11609<br />
<br />
. <strong>CBS</strong> 110024<br />
<br />
. <strong>CBS</strong> 157.82<br />
<br />
. <strong>CBS</strong> 161.55<br />
<br />
. <strong>CBS</strong> 572.78<br />
<br />
. CPC 11386<br />
<br />
. CPC 12042<br />
<br />
. CPC 12046<br />
<br />
. CPC 12211<br />
<br />
. CPC 12212<br />
<br />
. CPC 11840<br />
<br />
. <strong>CBS</strong> 188.54<br />
<br />
. <strong>CBS</strong> 366.80<br />
<br />
. CPC 12921<br />
<br />
. <strong>CBS</strong> 159.54<br />
<br />
. <strong>CBS</strong> 177.71<br />
<br />
. <strong>CBS</strong> 399.80<br />
<br />
. <strong>CBS</strong> 521.68<br />
<br />
. CPC 12139<br />
<br />
. <strong>CBS</strong> 115683<br />
<br />
. CPC12041<br />
<br />
. CPC 11839<br />
<br />
. <strong>CBS</strong> 134.31<br />
<br />
<br />
. <strong>CBS</strong> 813.71<br />
<br />
. CPC 12045<br />
<br />
. <strong>CBS</strong> 113753<br />
<br />
. <strong>CBS</strong> 113754<br />
<br />
. CPC 12044<br />
<br />
. CPC 12485<br />
<br />
. <strong>CBS</strong> 690.92<br />
<br />
. <strong>CBS</strong> 113742<br />
<br />
. <strong>CBS</strong> 113741<br />
<br />
. <strong>CBS</strong> 172.52<br />
<br />
. <strong>CBS</strong> 116463<br />
<br />
. <strong>CBS</strong> 102044<br />
<br />
. CPC 12040<br />
<br />
. <strong>CBS</strong> 289.49<br />
<br />
. <strong>CBS</strong> 842.91<br />
<br />
. <strong>CBS</strong> 843.91<br />
<br />
. <strong>CBS</strong> 113744<br />
<br />
. CPC 12484<br />
<br />
. CPC 12751<br />
<br />
. CPC 12753<br />
•<br />
. <strong>CBS</strong> 107.20<br />
•<br />
. <strong>CBS</strong> 138.40<br />
<br />
. CPC 11600<br />
<br />
. CPC 11601<br />
<br />
. CPC 11602<br />
<br />
. CPC 11604<br />
<br />
. <strong>CBS</strong> 111.82<br />
<br />
. <strong>CBS</strong> 300.49<br />
<br />
. CPC 11603<br />
•<br />
. <strong>CBS</strong> 175.82<br />
•<br />
. <strong>CBS</strong> 223.31<br />
•<br />
. <strong>CBS</strong> 299.67<br />
•<br />
. CPC 11817<br />
•<br />
. CPC 12054<br />
<br />
. CPC 12752<br />
<br />
. CPC 12755<br />
<br />
. CPC 12756<br />
•<br />
. CPC 12758<br />
•<br />
. CPC 12759<br />
•<br />
. CPC 12757<br />
<br />
. CPC 12177<br />
<br />
. CPC 12178<br />
<br />
. CPC 12179<br />
<br />
. CPC 12180<br />
<br />
. CPC 12181<br />
<br />
. CPC 12183<br />
•<br />
. CPC 12052<br />
<br />
. CPC 11813<br />
<br />
. CPC 12051<br />
<br />
. CPC 12053<br />
<br />
. CPC 12043<br />
<br />
. CPC 12047<br />
<br />
. <strong>CBS</strong> 290.49<br />
<strong>Cladosporium</strong> cladosporioides complex<br />
<strong>Cladosporium</strong> cladosporioides complex<br />
<strong>Cladosporium</strong> cladosporioides complex<br />
<strong>Cladosporium</strong> cladosporioides complex<br />
<strong>Cladosporium</strong> bruhnei<br />
<strong>Cladosporium</strong> bruhnei<br />
<strong>Cladosporium</strong> bruhnei<br />
<strong>Cladosporium</strong> bruhnei<br />
<strong>Cladosporium</strong> bruhnei<br />
<strong>Cladosporium</strong> bruhnei<br />
<strong>Cladosporium</strong> bruhnei<br />
<strong>Cladosporium</strong> bruhnei<br />
<strong>Cladosporium</strong> bruhnei<br />
<strong>Cladosporium</strong> bruhnei<br />
<strong>Cladosporium</strong> bruhnei<br />
<strong>Cladosporium</strong> bruhnei<br />
<strong>Cladosporium</strong> bruhnei<br />
<strong>Cladosporium</strong> bruhnei<br />
<strong>Cladosporium</strong> bruhnei<br />
<strong>Cladosporium</strong> bruhnei<br />
<strong>Cladosporium</strong> bruhnei<br />
<strong>Cladosporium</strong> bruhnei<br />
<strong>Cladosporium</strong> bruhnei<br />
<strong>Cladosporium</strong> subinflatum<br />
<strong>Cladosporium</strong> sinuosum<br />
<strong>Cladosporium</strong> bruhnei<br />
<strong>Cladosporium</strong> bruhnei<br />
<strong>Cladosporium</strong> bruhnei<br />
<strong>Cladosporium</strong> subtilissimum s.str.<br />
<strong>Cladosporium</strong> subtilissimum s.str.<br />
<strong>Cladosporium</strong> subtilissimum s.str.<br />
<strong>Cladosporium</strong> subtilissimum s.lat.<br />
<strong>Cladosporium</strong> antarcticum<br />
<strong>Cladosporium</strong> subtilissimum s.lat.<br />
<strong>Cladosporium</strong> subtilissimum s.lat.<br />
<strong>Cladosporium</strong> subtilissimum s.lat.<br />
<strong>Cladosporium</strong> pseudiridis<br />
<strong>Cladosporium</strong> spinulosum<br />
<strong>Cladosporium</strong> spinulosum<br />
Davidiella sp.<br />
<strong>Cladosporium</strong> ossifragi<br />
<strong>Cladosporium</strong> ossifragi<br />
<strong>Cladosporium</strong> subtilissimum s.lat.<br />
<strong>Cladosporium</strong> subtilissimum s.lat.<br />
<strong>Cladosporium</strong> variabile<br />
<strong>Cladosporium</strong> variabile<br />
<strong>Cladosporium</strong> iridis<br />
<strong>Cladosporium</strong> iridis<br />
<strong>Cladosporium</strong> herbarum<br />
<strong>Cladosporium</strong> herbarum<br />
<strong>Cladosporium</strong> herbarum<br />
<strong>Cladosporium</strong> herbarum<br />
<strong>Cladosporium</strong> herbarum<br />
<strong>Cladosporium</strong> herbarum<br />
<strong>Cladosporium</strong> herbarum<br />
<strong>Cladosporium</strong> macrocarpum<br />
<strong>Cladosporium</strong> macrocarpum<br />
<strong>Cladosporium</strong> macrocarpum<br />
<strong>Cladosporium</strong> macrocarpum<br />
<strong>Cladosporium</strong> macrocarpum<br />
<strong>Cladosporium</strong> macrocarpum<br />
<strong>Cladosporium</strong> macrocarpum<br />
<strong>Cladosporium</strong> macrocarpum<br />
<strong>Cladosporium</strong> macrocarpum<br />
<strong>Cladosporium</strong> macrocarpum<br />
<strong>Cladosporium</strong> macrocarpum<br />
<strong>Cladosporium</strong> herbarum<br />
<strong>Cladosporium</strong> herbarum<br />
<strong>Cladosporium</strong> herbarum<br />
<strong>Cladosporium</strong> herbarum<br />
<strong>Cladosporium</strong> herbarum<br />
<strong>Cladosporium</strong> herbarum<br />
<strong>Cladosporium</strong> herbaroides<br />
<strong>Cladosporium</strong> tenellum<br />
<strong>Cladosporium</strong> tenellum<br />
<strong>Cladosporium</strong> tenellum<br />
<strong>Cladosporium</strong> ramotenellum<br />
<strong>Cladosporium</strong> ramotenellum<br />
Davidiella sp.<br />
Water<br />
Germany<br />
Wood<br />
Belgium<br />
Human<br />
Netherl<strong>and</strong>s<br />
Fungus<br />
Russia<br />
Tree<br />
Germany<br />
Hypersaline Slovenia<br />
Airco<br />
Slovenia<br />
Grass<br />
Belgium<br />
Grass<br />
Belgium<br />
Plant<br />
Australia<br />
? ?<br />
Human<br />
Netherl<strong>and</strong>s<br />
Tree<br />
Australia<br />
Human<br />
Netherl<strong>and</strong>s<br />
Chemical Netherl<strong>and</strong>s<br />
Human<br />
Netherl<strong>and</strong>s<br />
Air<br />
Netherl<strong>and</strong>s<br />
Grass<br />
Netherl<strong>and</strong>s<br />
Treated pole USA<br />
Hypersaline Slovenia<br />
Plant<br />
New Zeal<strong>and</strong><br />
? Germany<br />
Plant<br />
Czechia<br />
Hypersaline Spain<br />
Cherry<br />
USA .<br />
Grape<br />
USA<br />
Hypersaline Slovenia<br />
.<br />
Wood<br />
Argentina<br />
Lichen<br />
Antarctica<br />
Grape<br />
USA<br />
Grape<br />
USA<br />
Plant<br />
USA<br />
Plant<br />
New Zeal<strong>and</strong><br />
Hypersaline Slovenia<br />
Hypersaline Slovenia<br />
Plant<br />
Switzerl<strong>and</strong><br />
Plant<br />
Norway<br />
Plant<br />
Norway<br />
Grape<br />
USA<br />
Wood<br />
Argentina<br />
Spinach<br />
USA<br />
Spinach<br />
USA<br />
Plant<br />
Netherl<strong>and</strong>s<br />
Plant<br />
Netherl<strong>and</strong>s<br />
Plant<br />
USA<br />
Plant<br />
USA<br />
Plant<br />
USA<br />
Plant<br />
USA<br />
Plant<br />
Switzerl<strong>and</strong><br />
Plant<br />
Switzerl<strong>and</strong><br />
Plant<br />
USA<br />
Water<br />
Romania<br />
? ?<br />
Straw<br />
Turkey<br />
Branch<br />
USA<br />
Hypersaline Slovenia<br />
Spinach<br />
USA<br />
Spinach<br />
USA<br />
Spinach<br />
USA<br />
Spinach<br />
USA<br />
Spinach<br />
USA<br />
Spinach<br />
USA<br />
Grass<br />
Netherl<strong>and</strong>s<br />
Grass<br />
Netherl<strong>and</strong>s<br />
Grass<br />
Netherl<strong>and</strong>s<br />
Grass<br />
Netherl<strong>and</strong>s<br />
Grass<br />
Netherl<strong>and</strong>s<br />
Grass<br />
Netherl<strong>and</strong>s<br />
Saltern<br />
Israel<br />
Fungus<br />
USA<br />
Hypersaline Slovenia<br />
Hypersaline Slovenia<br />
Hypersaline Slovenia<br />
Airco<br />
Slovenia<br />
Plant<br />
Switzerl<strong>and</strong><br />
Fig. 4. Distance tree of the <strong>Cladosporium</strong> herbarum complex based on ACT sequence data generated with UPGMA, showing Structure analysis at K = 6 under admixture model<br />
with correlated allele frequencies. Group indications (18) are taken from a tree based on EF sequences with AIC under the HKYG model.<br />
112
<strong>Cladosporium</strong> herbarum species complex<br />
B<br />
A<br />
C<br />
D<br />
Fig. 5. Split decomposition of the <strong>Cladosporium</strong> herbarum complex using SplitsTree of 16–22 unique alleles obtained from 79 <strong>Cladosporium</strong> isolates for four loci. <strong>The</strong> star-like<br />
structures suggest clonal development. A = ACT, B = CAL, C = HIS, D = EF. Scale bars = 0.01 nucleotide substitutions per site.<br />
Taxonomy<br />
Key to the <strong>Cladosporium</strong> species treated<br />
Morphological features used in the key to distinguish the species treated in this study were determined after 7 d growth at 25 ºC on SNA using<br />
light microscopy, <strong>and</strong> cultural characteristics after 14 d incubation on PDA.<br />
1. Conidia usually smooth, rarely minutely verruculose ..................................................................... C. cladosporioides (species complex)<br />
1. Conidia with different surface ornamentation, minutely to distinctly verruculose, verrucose to echinulate or spiny<br />
.................................................................................................................................................................................................................... 2<br />
2. Conidiophores uniform, macronematous; conidia solitary, sometimes formed in short unbranched chains .............................................. 3<br />
2. Conidiophores both macronematous <strong>and</strong> micronematous; conidia always catenate, usually formed in branched chains<br />
.................................................................................................................................................................................................................... 5<br />
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Schubert et al.<br />
3. Conidiophores due to geniculations often growing zigzag-like, (4–)5–7 µm wide; conidia 9–21 × (5–)6–8 µm, 0–1-septate;<br />
conidiogenous loci <strong>and</strong> conidial hila 1.2–2(–2.2) µm diam .................................................................................................... C. sinuosum<br />
3. Conidiophores not growing zigzag-like, wider, 6–11 µm; conidia very large <strong>and</strong> wide, 15–75(–87) × (7–)10–19(–21) µm, often with more<br />
septa; conidiogenous loci <strong>and</strong> hila wider, (2–)2.5–4 µm diam ................................................................................................................... 4<br />
4. Conidia (18–)30–75(–87) × (7–)10–16(–18) µm, (0–)2–6(–7)-septate, walls thickened, especially in older conidia, up to 1 µm thick<br />
......................................................................................................................................................................................................... C. iridis<br />
4. Conidia shorter <strong>and</strong> wider, 15–55 × (9–)11–19(–21) µm, 0–3-septate, walls distinctly thickened, up to 2 µm, usually appearing zonate<br />
.............................................................................................................................................................................................. C. pseudiridis<br />
5(2) Macronematous conidiophores nodulose or nodose with conidiogenous loci usually confined to swellings ........................................... 6<br />
5. Macronematous conidiophores non-nodulose or only occasionally subnodulose due to geniculate proliferation, but conidiogenous loci not<br />
confined to swellings ................................................................................................................................................................................ 11<br />
6. Macronematous conidiophores 3–6 µm wide, swellings 5–11 µm wide .................................................................................................... 7<br />
6. Macronematous conidiophores somewhat narrower, (1.5–)2.5–5 µm wide, swellings 3–8 µm wide ........................................................ 8<br />
7. Aerial mycelium twisted; conidial septa often distinctly darkened, becoming sinuous with age, apex <strong>and</strong> base of the conidia often appear<br />
to be distinctly darkened; slower growing in culture (29 mm after 14 d on PDA) ...................................................................... C. variabile<br />
7. Aerial mycelium not twisted; conidial septa as well as apex <strong>and</strong> base not distinctly darkened, septa not sinuous with age; faster growing<br />
in culture (on average 38 mm after 14 d on PDA) .......................................................................................................... C. macrocarpum<br />
8. Macronematous conidiophores (1.5–)2.5–4.5(–5.5) µm wide, swellings 3–6.5 µm wide; conidia 4–17(–22) µm long, ornamentation<br />
variable, but usually densely echinulate, spines up to 0.8 µm long .................................................................................... C. subinflatum<br />
8. Macronematous conidiophores slightly wider, 3–5 µm, swellings (4–)5–8(–9) µm wide; conidia longer, up to 25(–35) µm, ornamentation<br />
minutely verruculose to verrucose, but not echinulate or spiny ................................................................................................................. 9<br />
9. Conidia formed by macronematous conidiophores 3–33 × (2–)3–6(–7) µm, with age becoming wider, (3.5–)5–9(–11) µm, darker <strong>and</strong><br />
more thick-walled ................................................................................................................................................................ C. herbaroides<br />
9. Conidia formed by macronematous conidiophores not becoming wider <strong>and</strong> darker with age, usually up to 7 µm wide<br />
.................................................................................................................................................................................................................. 10<br />
10. Conidiophores usually with small head-like swellings, sometimes also with a second intercalary nodule; small terminal conidia<br />
4–9 × 2.5–3.5 µm, secondary ramoconidia <strong>and</strong> occasionally formed ramoconidia 10–24(–31) × 3–5(–7) µm ........................ C. bruhnei<br />
10. Conidiophores with a single or often numerous swellings in short succession giving the stalk a knotty/gnarled appearance;<br />
conidia wider, small terminal conidia 4–10 × 3–5(–6) µm, intercalary conidia 6–16 × 4–6 µm, secondary ramoconidia 12–25(–35) ×<br />
(3–)5–7(–9) µm ....................................................................................................................................................................... C. herbarum<br />
11(5) Small terminal <strong>and</strong> intercalary conidia 4–15 × 3–5 µm, secondary ramoconidia 16–36(–40) × (4–)5–8 µm, 0–3(–4)-septate,<br />
ramoconidia absent ................................................................................................................................................................. C. ossifragi<br />
11. Small terminal conidia, ramoconidia <strong>and</strong> secondary ramoconidia distinctly narrower, 2–5(–6) µm wide, 0–2(–3)-septate<br />
.................................................................................................................................................................................................................. 12<br />
12. Mycelium dimorphic, narrow hyphae 1–3 µm wide, hyaline to subhyaline, thin-walled, hyphae of the second type wider, 3.5–8(–9)<br />
µm, pale to dark greyish olivaceous or olivaceous-brown, thick-walled, sometimes even two-layered, 1(–1.5) µm thick, hyphae<br />
appearing consistently enveloped in polysaccharide-like material or covered by a slime coat; conidiophores usually several times slightly<br />
to distinctly geniculate towards the apex, with numerous conidiogenous loci crowded towards the apex, up to 14 per conidiogenous cell<br />
............................................................................................................................................................................................. C. antarcticum<br />
12. Mycelium not dimorphic, neither enveloped in polysaccharide-like material nor covered by a slime coat; conidiophores usually not<br />
geniculate, occasionally only slightly so ................................................................................................................................................... 13<br />
13. Conidial ornamentation distinctly echinulate, spiny (baculate, digitate or capitate under SEM), spines 0.5–1.3 µm long, loose to moderately<br />
dense, conidial hila usually situated on small peg-like prolongations or denticles .............................................................. C. spinulosum<br />
13. Conidial ornamentation different, minutely verruculose to verruculose, conidial hila not situated on peg-like prolongations<br />
.................................................................................................................................................................................................................. 14<br />
14. Small terminal conidia narrowly obovoid, limoniform or fusiform, but neither globose nor subglobose; conidiogenous loci <strong>and</strong> conidial hila<br />
0.5–2(–2.5) µm diam ........................................................................................................................ C. subtilissimum (species complex)<br />
14. Numerous small globose or subglobose terminal conidia formed, also ovoid or limoniform; conidiogenous loci <strong>and</strong> conidial hila somewhat<br />
smaller, 0.5–1.5(–2) µm diam .................................................................................................................................................................. 15<br />
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<strong>Cladosporium</strong> herbarum species complex<br />
15. Conidiophores usually with numerous conidiogenous loci forming sympodial clusters of pronounced scars at the apex, sometimes up to<br />
10 or even more denticulate loci; conidia 3–20(–28) × 2.5–5(–6) µm, 0–1(–2)-septate, often with several apically crowded hila, up to 7(–9)<br />
.................................................................................................................................................................................................. C. tenellum<br />
15. Conidiophores usually only with few conidiogenous loci, mostly 1–3; conidia longer <strong>and</strong> narrower, 2.5–35 × 2–4(–5) µm, 0–3-septate,<br />
usually with up to three distal conidial hila ....................................................................................................................... C. ramotenellum<br />
Key to the Davidiella species treated<br />
1. Ascospores frequently wider than 7 µm when mounted in Shear’s solution or lactic acid, apical cell obtusely rounded<br />
.................................................................................................................................................................................................................... 2<br />
1. Ascospores not wider than 7 µm when mounted in Shear’s solution or lactic acid, apical cell acutely rounded, ascospores (20–)25–27<br />
(–30) × (5.5–)6–7 µm ................................................................................................................................................................. D. allicina<br />
2. Pseudoparaphyses prominent; asci frequently >95 µm; ascospores (22–)23–26(–28) × (6–)6.5–7(–8) µm...................... D. macrocarpa<br />
2. Pseudoparaphyses mostly absent in older ascomata; asci
Schubert et al.<br />
Fig. 6. <strong>Cladosporium</strong> antarcticum (<strong>CBS</strong> 690.92). Macro- <strong>and</strong> micronematous conidiophores <strong>and</strong> conidia. Scale bar = 10 µm. K. Schubert del.<br />
geniculate with several conidiogenous loci at the apex, 2–22 ×<br />
2–3 µm, pale greyish olivaceous, loci denticulate. Ramoconidia<br />
occasionally occurring, cylindrical, up to 30 µm long, 4–5 µm<br />
wide, 0–1-septate, concolorous with the tips of conidiophores, with<br />
a broadly truncate, unthickened <strong>and</strong> not darkened base, without<br />
dome <strong>and</strong> rim, 2.5 µm wide. Conidia catenate, in branched chains,<br />
straight, small terminal conidia obovoid, limoniform or narrowly<br />
ellipsoid, 4–14 × 2.5–4 µm [av. ± SD, 8.5 (± 3.3) × 3.5 (± 0.6)],<br />
0(–1)-septate, secondary ramoconidia ellipsoid to cylindrical, often<br />
with several or numerous conidial hila crowded at the distal end, up<br />
to 12, 13–30 × 4–5 µm [av. ± SD, 20.1 (± 5.8) × 4.3 (± 0.5) µm],<br />
0–3-septate, sometimes slightly constricted at the median septum,<br />
pale olivaceous-brown or greyish brown, minutely verruculose to<br />
verrucose (granulate under SEM), walls more or less thickened,<br />
rounded or slightly attenuated towards apex <strong>and</strong> base, hila<br />
protuberant, denticulate, 0.8–1.5(–2) µm diam, thickened <strong>and</strong><br />
darkened-refractive; microcyclic conidiogenesis occurring.<br />
Cultural characteristics: Colonies on PDA attaining 9 mm diam after<br />
14 d at 25 ºC, greenish olivaceous to grey-olivaceous, at the margin<br />
becoming dull green, reverse with a pale olivaceous-grey centre<br />
<strong>and</strong> a broad olivaceous-black margin, margin narrow, regular,<br />
entire edge, white, feathery, aerial mycelium sparse but colonies<br />
appearing felty, growth flat with somewhat elevated colony centre,<br />
prominent exudates not formed, sporulation dense, covering almost<br />
the whole colony. Colonies on MEA attaining 12 mm diam after 14<br />
d at 25 ºC, olivaceous-grey to iron-grey, iron-grey reverse, velvety<br />
to powdery, aerial mycelium sparse, sporulation profuse. Colonies<br />
on OA attaining 4 mm after 14 d at 25 ºC, olivaceous-grey, aerial<br />
mycelium sparse, diffuse, growth flat, without prominent exudates,<br />
sporulating.<br />
Specimen examined: Antarctica, King George, Arctowski, isolated from the lichen<br />
Caloplaca regalis (Teloschistaceae), C. Möller, No. 32/12, 1991, <strong>CBS</strong>-H 19857,<br />
holotype, isotype HAL 2024 F, culture ex-type <strong>CBS</strong> 690.92.<br />
Substrate <strong>and</strong> distribution: On the lichen Caloplaca regalis;<br />
Antarctica.<br />
Notes: This is the second genuine lichenicolous species of the<br />
<strong>genus</strong> <strong>Cladosporium</strong>. <strong>Cladosporium</strong> licheniphilum Heuchert & U.<br />
Braun, occurring on apothecia of Pertusaria alpina in Russia, is<br />
quite distinct from C. antarcticum by having subcylindrical or only<br />
slightly geniculate-sinuous, wider conidiophores, 5–8 µm, with<br />
numerous characteristic terminal branches <strong>and</strong> much shorter, 0–<br />
1-septate, smooth conidia, 3.5–13 × 3–7 µm (Heuchert & Braun<br />
2006). <strong>Cladosporium</strong> lichenicola Linds. was invalidly published <strong>and</strong><br />
C. arthoniae M.S. Christ. & D. Hawksw. as well as C. lichenum<br />
Keissl. are to be excluded from the <strong>genus</strong> <strong>Cladosporium</strong> since<br />
they do not possess the typical cladosporioid scar structure but<br />
inconspicuous, unthickened conidiogenous loci <strong>and</strong> conidial hila<br />
(Hawksworth 1979, Heuchert et al. 2005). <strong>The</strong> fungicolous species<br />
C. uredinicola Speg. <strong>and</strong> the foliicolous species C. alneum Pass.<br />
ex K. Schub. <strong>and</strong> C. psoraleae M.B. Ellis are morphologically<br />
superficially <strong>similar</strong>. However, C. uredinicola, a widespread fungus<br />
on rust fungi, downy mildews <strong>and</strong> powdery mildew fungi, differs<br />
in having somewhat longer <strong>and</strong> wider, smooth conidia, 3–39 × 2–<br />
6.5(–8) µm, <strong>and</strong> wider conidiogenous loci <strong>and</strong> conidial hila, 0.5–3<br />
116
<strong>Cladosporium</strong> herbarum species complex<br />
Fig. 7. <strong>Cladosporium</strong> antarcticum (<strong>CBS</strong> 690.92). A.<br />
Overview of the growth pattern on SNA. Note the very<br />
large bulbous cells formed at the base of different<br />
conidiophores. Other conidiophores sprout from the<br />
agar surface. B. Overview of conidiophores <strong>and</strong> conidia.<br />
Note the large distance of the scars on the conidiophore<br />
<strong>and</strong> the different stages of conidial formation on the<br />
tips of other conidia. <strong>The</strong> long secondary ramoconidia<br />
are also visible, <strong>and</strong> sparse aerial hyphae. C. Detail<br />
of B with details of the ornamentation <strong>and</strong> scars. <strong>The</strong><br />
absence of ornamentation at the apical (spore-forming)<br />
end of the secondary ramoconidium is clearly visible. D–<br />
E. Tubular structures on coniophore (D) <strong>and</strong> secondary<br />
ramoconidium (E). Scale bars: A–B = 10 μm, C–D = 5<br />
µm, E = 2 µm.<br />
Fig. 8. <strong>Cladosporium</strong> antarcticum (<strong>CBS</strong> 690.92). A–B. Macronematous conidiophores. C, G. Mycelium enveloped by a polysaccharide-like layer. D, F. Conidia. E. Micronematous<br />
conidiophore. H. Ramoconidium with numerous distal scars. Scale bars = 10 µm.<br />
µm (Heuchert et al. 2005); C. alneum, which causes leaf spots on<br />
Alnus glutinosa, possesses longer <strong>and</strong> wider conidiophores, 25–<br />
260 × (2–)3–7(–8.5) µm, <strong>and</strong> somewhat shorter, smooth conidia<br />
(Schubert 2005, Schubert et al. 2006); <strong>and</strong> C. psoraleae, known<br />
from Myanmar on Psoralea corylifolia, can easily be distinguished<br />
from C. antarcticum by its smooth <strong>and</strong> wider conidia, 3.5–7 µm,<br />
<strong>and</strong> wider conidiogenous loci <strong>and</strong> conidial hila, 1–3 µm diam (Ellis<br />
1972, Schubert 2005).<br />
www.studiesinmycology.org<br />
117
Schubert et al.<br />
Fig. 9. <strong>Cladosporium</strong> bruhnei (CPC 12211). Macro- <strong>and</strong> micronematous conidiophores <strong>and</strong> conidia. Scale bar = 10 µm. K. Schubert del.<br />
<strong>Cladosporium</strong> bruhnei Linder, Bull. Natl. Mus. Canada 97: 259.<br />
1947. Figs 9–12.<br />
≡ Hormodendrum hordei Bruhne, in W. Zopf, Beitr. Physiol. Morph. nied.<br />
Org. 4: 1. 1894, non C. hordei Pass., 1887.<br />
≡ <strong>Cladosporium</strong> herbarum (Pers.: Fr.) Link var. (δ) cerealium Sacc. f.<br />
hordei (Bruhne) Ferraris, Flora Ital. Crypt., Pars I, Fungi, Fasc.13: 882.<br />
1914.<br />
≡ <strong>Cladosporium</strong> hordei (Bruhne) Pidopl., Gribnaja Flora Grubych Kormov:<br />
268. 1953, nom. illeg., homonym, non C. hordei Pass., 1887.<br />
Teleomorph: Davidiella allicina (Fr. : Fr.) Crous & Aptroot, in<br />
Aptroot, Mycosphaerella <strong>and</strong> its anamorphs: 2. Conspectus of<br />
Mycosphaerella. <strong>CBS</strong> Biodiversity Ser. 5: 30. 2006.<br />
Basionym: Sphaeria allicina Fr., Kongl. Vetensk. Acad. H<strong>and</strong>l. 38:<br />
247. 1817, sactioned by Fr., Syst. Mycol. 2: 437. 1823.<br />
≡ Sphaerella allicina (Fr. : Fr.) Auersw., in Gonn. & Rabenh., Mycol.<br />
Europaea 5–6: 19. 1869.<br />
Ascomata pseudothecial, black, superficial, situated on a small<br />
stroma, globose, up to 250 µm diam; ostioles periphysate, with<br />
apical periphysoids present; wall consisting of 3–6 layers of<br />
reddish brown textura angularis. Asci fasciculate, bitunicate,<br />
subsessile, obovoid to broadly ellipsoid, straight to slightly curved,<br />
8-spored, 65–90 × 16–25 µm; with pseudoparenchymatal cells<br />
of the hamathecium persistent. Ascospores tri- to multiseriate,<br />
overlapping, hyaline, with irregular lumina, thick-walled, straight<br />
to slightly curved, fusoid-ellipsoidal with obtuse basal end, <strong>and</strong><br />
acutely rounded apical end, widest near the middle of the apical<br />
cell, medianly 1-septate, not to slightly constricted at the septum,<br />
(20–)25–27(–30) × (5.5–)6–7 µm.<br />
Mycelium superficial, hyphae branched, 1.5–8 µm wide,<br />
pluriseptate, broader hyphae usually slightly constricted at the<br />
septa <strong>and</strong> somewhat swollen, hyaline to subhyaline, almost smooth<br />
to somewhat verruculose or irregularly rough-walled, sometimes<br />
appearing to have a slime coat, walls unthickened. Conidiophores<br />
macronematous, sometimes also micronematous, arising as lateral<br />
or terminal branches from plagiotropous or ascending hyphae, erect,<br />
straight to more or less flexuous, sometimes geniculate, nodulose,<br />
usually with small head-like swellings, sometimes also with<br />
intercalary nodules, sometimes swellings protruding <strong>and</strong> elongated<br />
to one side, unbranched, occasionally branched, (7–)20–330 µm,<br />
sometimes even longer, (2–)3–5 µm wide, swellings (4–)5–8 µm<br />
wide, pluriseptate, not constricted at the septa, septa sometimes<br />
not very conspicuous, subhyaline to pale brown or pale olivaceous,<br />
smooth or somewhat verruculose, walls unthickened or almost so,<br />
more thickened with age. Conidiogenous cells integrated, usually<br />
terminal, cylindrical with a terminal head-like swelling, sometimes<br />
with a second swelling, 15–40 µm long, proliferation sympodial,<br />
with few conidiogenous loci confined to swellings, up to five per<br />
swelling, loci protuberant, conspicuous, 1–2 µm diam, thickened<br />
<strong>and</strong> darkened-refractive. Conidia catenate, formed in branched<br />
chains, straight to slightly curved, small terminal conidia subglobose,<br />
ovoid to obovoid or somewhat limoniform, 4–9 × 2.5–3.5 µm [av. ±<br />
SD, 6.5 (± 1.5) × 3.1 (± 0.5) µm], aseptate; secondary ramoconidia<br />
<strong>and</strong> occasionally formed ramoconidia ellipsoid to subcylindrical<br />
or cylindrical, 10–24(–31) × 3–5(–7) µm [av. ± SD, 16.1 (± 4.1)<br />
× 4.1 (± 0.8) µm], rarely up to 40 µm long, 0–1(–3)-septate, very<br />
rarely 5-septate, subhyaline to pale brown or pale olivaceous,<br />
minutely verruculose to verrucose (mostly granulate with some<br />
muricate projections under SEM), walls unthickened or almost so,<br />
apex rounded or slightly attenuated towards apex <strong>and</strong> base, hila<br />
protuberant, conspicuous, 1–2 µm wide, up to 1 µm high, thickened<br />
<strong>and</strong> darkened-refractive; microcyclic conidiogenesis occurring.<br />
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<strong>Cladosporium</strong> herbarum species complex<br />
Belgium, isolated from Quercus robur (Fagaceae), <strong>CBS</strong> 157.82; Kampenhout,<br />
isolated from Hordeum vulgare (Poaceae), 26 June 2005, J.Z. Groenewald, <strong>CBS</strong>-H<br />
19856, neotype designated here of C. bruhnei, isoneotype HAL 2023 F, cultures<br />
ex-type <strong>CBS</strong> 121624 = CPC 12211, CPC 12212. Czech Republic, Lisen, isolated<br />
from Polygonatum odoratum (Liliaceae), <strong>CBS</strong> 813.71, albino mutant of <strong>CBS</strong> 812.71.<br />
Germany, <strong>CBS</strong> 134.31 = ATCC 11283 = IMI 049632; Nordrhein-Westfalen, Mühlheim<br />
an der Ruhr, isolated from industrial water, IWW 727, <strong>CBS</strong> 110024; Sachsen-Anhalt,<br />
Halle (Saale), Robert-Franz-Ring, isolated from leaves of Tilia cordata (Tiliaceae),<br />
2004, K. Schubert, CPC 11386. Netherl<strong>and</strong>s, isolated from air, <strong>CBS</strong> 521.68; isolated<br />
from Hordeum vulgare, 1 Jan. 2005, P.W. Crous, CPC 12139; isolated from man,<br />
skin, <strong>CBS</strong> 159.54 = ATCC 36948; Amsterdam, isolated from Thuja tincture, <strong>CBS</strong><br />
177.71; Geleen, St. Barbara Ziekenhuis, isolated from man, skin, <strong>CBS</strong> 366.80, <strong>CBS</strong><br />
399.80; isolated from man, sputum, Aug. 1955, <strong>CBS</strong> 161.55. New Zeal<strong>and</strong>, Otago,<br />
Lake Harris, isolated from Ourisia macrophylla (Scrophulariaceae), 30 Jan. 2005,<br />
A. Blouin, Hill 1135, CPC 11840. Russia, Moscow region, isolated from Polyporus<br />
radiatus (Polyporaceae), Oct. 1978, <strong>CBS</strong> 572.78 = VKM F-405. Slovenia, Ljubljana,<br />
isolated from an air conditioning system, 2004, M. Butala, EXF-680 = CPC 12046;<br />
Sečovlje, isolated from hypersaline water from salterns (reserve pond), 2005, P.<br />
Zalar, EXF-389 = CPC 12042. Spain, Ebro Delta, isolated from hypersaline water<br />
from salterns (crystallisation pond), 2004, P. Zalar, EXF-594 = CPC 12045. Sweden,<br />
Skåne, on tip blight of living leaves of Allium sp. (Alliaceae), Fr. no. F-09810, UPS-<br />
FRIES, holotype of Davidiella allicina. U.S.A., New York, Geneva, isolated from<br />
CCA-treated Douglas-fir pole, <strong>CBS</strong> 115683 = ATCC 66670 = CPC 5101.<br />
Substrate <strong>and</strong> distribution: Living <strong>and</strong> decaying plant material, man,<br />
air, hypersaline <strong>and</strong> industrial water; widespread.<br />
Literature: Saccardo (1899: 1076), Linder (1947: 289).<br />
Fig. 10. <strong>Cladosporium</strong> bruhnei (CPC 12211). A. Conidiophore with characteristic<br />
long secondary ramoconidium <strong>and</strong> complex conidiophore. B. Detail of hila on<br />
secondary ramoconidia. C. Details of prominent ornamentation on conidia. Scale<br />
bars: A = 10 µm, B = 2 µm, C = 5 µm.<br />
Cultural characteristics: Colonies on PDA reaching 22–32 mm diam<br />
after 14 d at 25 ºC, olivaceous-grey to iron-grey, sometimes whitish,<br />
smoke-grey to pale olivaceous due to abundant aerial mycelium<br />
covering almost the whole colony, with age collapsing becoming<br />
olivaceous-grey, occasionally zonate, velvety to floccose, margin<br />
narrow, entire edge, white, glabrous to somewhat feathery, aerial<br />
mycelium sparse to abundant, white, fluffy, growth regular, flat to<br />
low convex, sometimes forming few exudates in the colony centre,<br />
sporulating. Colonies on MEA reaching 21–32 mm diam after 14<br />
d at 25 ºC, grey-olivaceous, olivaceous-grey to dull green or irongrey,<br />
sometimes whitish to pale smoke-grey due to abundant aerial<br />
mycelium, olivaceous-grey to iron-grey reverse, velvety, margin<br />
narrow, entire edge to slightly undulate, white, radially furrowed,<br />
glabrous to slightly feathery, aerial mycelium sparse to abundant,<br />
mainly in the centre, white, fluffy, growth convex to raised, radially<br />
furrowed, distinctly wrinkled in the colony centre, without prominent<br />
exudates, sporulating. Colonies on OA reaching 20–32 mm diam<br />
after 14 d at 25 ºC, smoke-grey, grey-olivaceous to olivaceous-grey,<br />
greenish black or iron-grey reverse, margin narrow, entire edge,<br />
colourless to white, glabrous, aerial mycelium sparse to abundant,<br />
dark smoke-grey, diffuse, high, later collapsed, felty, growth flat,<br />
without prominent exudates, sporulation profuse.<br />
Specimens examined: Sine loco et dato, <strong>CBS</strong> 188.54 = ATCC 11290 = IMI<br />
049638. Australia, N.S.W., Barrington Tops National Park, isolated from leaves<br />
of Eucalyptus stellulata (Myrtaceae), 3 Jan. 2006, B. Summerell, CPC 12921.<br />
Fig. 11. Davidiella allicina (F-09810, UPS-FRIES, holotype). Ascus <strong>and</strong> ascospores.<br />
Scale bar = 10 µm. P.W. Crous del.<br />
www.studiesinmycology.org<br />
119
Schubert et al.<br />
Fig. 12. <strong>Cladosporium</strong> bruhnei (CPC 12211) <strong>and</strong> its teleomorph Davidiella allicina. A–B. Macronematous conidiophores. C. Conidial chains. D. Micronematous conidiophore. E.<br />
Ascomata of the teleomorph formed on the host. F–G. Asci. Scale bars: A–B, D, F = 10 µm, E = 200 µm.<br />
Notes: <strong>Cladosporium</strong> bruhnei proved to be an additional component<br />
of the herbarum complex. <strong>The</strong> species resembles C. herbarum s.<br />
str. as already stated by Linder (1947), but possesses consistently<br />
narrower conidia, usually 2.5–5 µm wide, <strong>and</strong> the conidiophores<br />
often form only a single apical swelling. <strong>The</strong> species was described<br />
by Bruhne (l.c.) as Hormodendrum hordei from Germany but type<br />
material could not be located. Linder (1947) examined No. 1481a-5<br />
(Canada, N. Quebec, Sugluk, on Elymus arenarius var. villosus,<br />
31 Jul. 1936, E. Meyer), presumably in the National Museum, <strong>and</strong><br />
stated that this specimen agreed well with the description <strong>and</strong><br />
illustration given by Bruhne (l.c.). Although the species occurs<br />
on numerous substrates <strong>and</strong> is widely distributed, it has not yet<br />
been recognised as a distinct species since it has probably been<br />
interpreted as a narrow variant of C. herbarum.<br />
Based on morphology <strong>and</strong> DNA sequence data, the <strong>CBS</strong> strain<br />
<strong>CBS</strong> 177.71 chosen by Prasil & de Hoog (1988) as representative<br />
living strain of C. herbarum, rather clusters together with isolates<br />
of C. bruhnei. <strong>The</strong> strain <strong>CBS</strong> 813.71 is an albino mutant of the<br />
latter species as it does not appear to contain colour pigment.<br />
Furthermore, all isolates from humans treated until now as C.<br />
herbarum proved to be conspecific with the narrow-spored C.<br />
bruhnei.<br />
Although Davidiella tassiana (ascospores 17–25 × 6–8.5 µm,<br />
RO) was treated as synonymous to D. allicina (ascospores 20–27<br />
× 6–7 µm, UPS) in Aptroot (2006), they differ in apical ascospore<br />
taper, with ascospores of D. allicina being acutely rounded, while<br />
those of D. tassiana are obtusely rounded. <strong>The</strong> same ascospore<br />
taper was also observed in the teleomorph of C. bruhnei, <strong>and</strong> thus<br />
the name D. allicina is herewith linked to C. bruhnei, which is distinct<br />
from C. herbarum, having D. tassiana as teleomorph.<br />
<strong>Cladosporium</strong> herbaroides K. Schub., Zalar, Crous & U. Braun,<br />
sp. nov. MycoBank MB504574. Figs 13–15.<br />
Etymology: Refers to its morphological <strong>similar</strong>ity to <strong>Cladosporium</strong><br />
herbarum.<br />
Differt a Cladosporio herbaro conidiis polymorphis, 3–33 × (2–)3–6(–7) µm,<br />
postremo latioribus, (3.5–)5–9(–11) µm, fuscis et crassitunicatis; et a Cladosporio<br />
macrocarpo conidiophoris leniter angustioribus, 3–5 µm latis, nodulis angustioribus,<br />
5–8 µm latis.<br />
Mycelium branched, (1–)2–8 µm wide, septate, often with small<br />
swellings <strong>and</strong> constrictions, subhyaline to pale brown or pale<br />
olivaceous-brown, smooth or almost so to somewhat verruculose,<br />
walls unthickened or almost so. Conidiophores macronematous<br />
<strong>and</strong> micronematous, arising lateral from plagiotropous hyphae or<br />
terminally from ascending hyphae. Macronematous conidiophores<br />
erect, straight to slightly flexuous, often geniculate, nodulose, with<br />
unilateral or multilateral swellings, often numerous swellings in<br />
short succession giving them a gnarled appearance, often forming<br />
somewhat protruding or prolonged lateral swellings or a branch-<br />
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<strong>Cladosporium</strong> herbarum species complex<br />
Fig. 13. <strong>Cladosporium</strong> herbaroides (CPC 12052). Macro- <strong>and</strong> micronematous conidiophores <strong>and</strong> conidia. Scale bar = 10 µm. K. Schubert del.<br />
like prolongation below the terminal swelling (due to sympodial<br />
proliferation), unbranched or sometimes branched, 30–230 µm<br />
long or even longer, 3–5 µm wide, swellings 5–8 µm wide, septate,<br />
not constricted at septa, pale to medium olivaceous-brown,<br />
smooth or almost so, walls slightly thickened. Conidiogenous cells<br />
integrated, terminal or intercalary, cylindrical, usually nodulose to<br />
nodose forming distinct swellings, sometimes geniculate, 15–55<br />
µm long, with numerous conidiogenous loci usually confined to<br />
swellings or situated on small lateral shoulders, sometimes on the<br />
top of short peg-like prolongations or denticles, loci protuberant,<br />
1–2 µm diam, thickened <strong>and</strong> darkened-refractive. Micronematous<br />
conidiophores much shorter, narrower, paler, neither nodulose nor<br />
geniculate, arising laterally from plagiotropous hyphae, often only<br />
as short lateral denticles or branchlets of hyphae, erect, straight,<br />
conical to cylindrical, unbranched, 3–65 × 2–3 µm, mostly aseptate,<br />
sometimes up to five septa, subhyaline, smooth, walls unthickened.<br />
Conidiogenous cells integrated, terminal or conidiophores reduced<br />
to conidiogenous cells, conidiogenous loci solitary or sometimes<br />
as sympodial clusters of pronounced denticles, protuberant, 1–1.5<br />
µm diam, thickened <strong>and</strong> somewhat darkened-refractive. Conidia<br />
polymorphous, two main morphological types recognisable, formed<br />
www.studiesinmycology.org<br />
by the two different types of conidiophores, conidia formed by<br />
macronematous conidiophores catenate, in branched chains,<br />
straight to slightly curved, subglobose, obovoid, limoniform, ellipsoid<br />
to cylindrical, 3–33 × (2–)3–6(–7) µm [av. ± SD, 14.5 (± 7.9) ×<br />
5.2 (± 1.2) µm], 0–2(–3)-septate, sometimes slightly constricted at<br />
septa, septa median or somewhat in the lower half, pale to medium<br />
olivaceous-brown, verruculose to verrucose (granulate under<br />
SEM), walls slightly thickened, with up to three rarely four distal<br />
scars, with age becoming medium or even dark brown (chocolate<br />
brown), wider <strong>and</strong> more thick-walled, 5.5–33 × (3.5–)5–9(–11) µm<br />
[av. ± SD, 14.4 (± 6.9) × 7.2 (± 1.9) µm], walls up to 1 µm thick,<br />
hila protuberant, 0.8–2(–2.5) µm diam, thickened <strong>and</strong> darkenedrefractive;<br />
microcyclic conidiogenesis occurring. Conidia formed by<br />
micronematous conidiophores paler <strong>and</strong> narrower, mostly formed<br />
in unbranched chains, sometimes in branched chains with up to<br />
three distal hila, straight to slightly curved, limoniform, narrowly<br />
fusiform, almost filiform to subcylindrical, 10–26(–35) × 2–3.5<br />
µm [av. ± SD, 15.6 (± 6.2) × 2.9 (± 0.5) µm], 0–1(–3)-septate,<br />
subhyaline to pale brown, almost smooth to minutely verruculose,<br />
walls unthickened, hila protuberant, 1–1.5 µm diam, thickened <strong>and</strong><br />
somewhat darkened-refractive.<br />
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Fig. 14. <strong>Cladosporium</strong> herbaroides (CPC 12052). A–B, D. Macronematous conidiophores. C. Conidial chain. E. Micronematous conidiophore. F. Microcyclic conidiogenesis. G.<br />
Conidia formed by micronematous conidiophores. Scale bars = 10 µm.<br />
Cultural characteristics: Colonies on PDA attaining 23 mm diam<br />
after 14 d at 25 ºC, grey-olivaceous to olivaceous, olivaceous-grey<br />
reverse, velvety, margin regular, entire edge, narrow, feathery, aerial<br />
mycelium abundantly formed, loose, with age covering large parts<br />
of the colony, woolly, growth flat with somewhat elevated colony<br />
centre, folded, regular, deep into the agar, with few prominent<br />
exudates, sporulation profuse. Colonies on MEA attaining 24 mm<br />
diam after 14 d at 25 ºC, grey- to greenish olivaceous, olivaceousgrey<br />
or iron-grey reverse, velvety to powdery, margin narrow,<br />
colourless, entire edge, somewhat feathery, aerial mycelium pale<br />
olivaceous-grey, sparse, growth convex, radially furrowed, folded<br />
in the colony centre, without prominent exudates, sporulating.<br />
Colonies on OA attaining 23 mm diam after 14 d at 25 ºC, greyolivaceous,<br />
margin more or less regular, entire edge, colourless,<br />
somewhat feathery, aerial mycelium whitish to smoke grey, at first<br />
sparse, later more abundantly formed, growth flat, without exudates,<br />
sporulation profuse.<br />
Specimen examined: Israel, from hypersaline water of Eilat salterns, 2004, coll.<br />
N. Gunde-Cimerman, isol. M. Ota, <strong>CBS</strong>-H 19858, holotype, isotype HAL 2025 F,<br />
culture ex-type <strong>CBS</strong> 121626 = EXF-1733 = CPC 12052.<br />
Substrate <strong>and</strong> distribution: Hypersaline water; Israel.<br />
Notes: <strong>Cladosporium</strong> herbaroides is morphologically <strong>similar</strong> to C.<br />
herbarum but differs in having somewhat longer conidia becoming<br />
wider, darker <strong>and</strong> even more thick-walled with age [at first conidia<br />
3–33 × (2–)3–6(–7) µm, with age (3.5–)5–9(–11) µm wide]. Besides<br />
that, the species often produces a second conidial type formed on<br />
micronematous conidiophores, giving rise to unbranched conidial<br />
chains which are almost filiform, limoniform, narrowly fusiform to<br />
subcylindrical, much narrower <strong>and</strong> paler than the ones formed by<br />
macronematous conidiophores, 10–26(–35) × 2–3.5 µm. In C.<br />
herbarum, conidia formed by micronematous conidiophores do not<br />
occur as frequently as in C. herbaroides, <strong>and</strong> differ in being often<br />
clavate <strong>and</strong> somewhat wider, up to 4(–5) µm wide. <strong>Cladosporium</strong><br />
macrocarpum is easily distinguishable by having somewhat wider<br />
conidiophores (3–)4–6 µm, with distinctly wider swellings, 5–10 µm<br />
wide, <strong>and</strong> the conidia are usually (3–)5–9(–10) µm wide.<br />
<strong>Cladosporium</strong> herbarum (Pers. : Fr.) Link, Ges. Naturf. Freunde<br />
Berlin Mag. Neuesten Entdeck. Gesammten Naturk. 7: 37. 1816:<br />
Fr., Syst. mycol. 3(2): 370. 1832. Figs 16–19.<br />
Basionym: Dematium herbarum Pers., Ann. Bot. (Usteri) 11: 32.<br />
1794: Fr., Syst. mycol. 3(2): 370. 1832.<br />
= Dematium epiphyllum var. (β) chionanthi Pers., Mycol. eur. 1: 16. 1822, syn.<br />
nov.<br />
For additional synonyms see Dugan et al. (2004), Schubert<br />
(2005).<br />
Teleomorph: Davidiella tassiana (De Not.) Crous & U. Braun,<br />
Mycol. Progr. 2: 8. 2003.<br />
Basionym: Sphaerella tassiana De Not., Sferiacei Italici 1: 87.<br />
1863.<br />
≡ Mycosphaerella tassiana (De Not.) Johanson, Öfvers. Förh. Kongl.<br />
Svenska Vetensk.-Akad. 41: 167. 1884.<br />
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<strong>Cladosporium</strong> herbarum species complex<br />
Fig. 15. <strong>Cladosporium</strong> herbaroides (CPC 12052). A. Overview of the growth characteristics of this fungus. Broad hyphae run over the surface of the agar, <strong>and</strong> possibly give rise<br />
to conidiophore branches. <strong>The</strong> conidiophores of this fungus can be rather long, resembling aerial hyphae. Clusters of conidia are clearly visible in this micrograph. B. <strong>The</strong> very<br />
wide surface hyphae can anastomose. C. Conidiophore with secondary ramoconidia <strong>and</strong> conidia. Note the variation in scar size. D. A very elaborate, complex conidiophore with<br />
different scars of variable size, one being more than 2 μm wide! E. Details of secondary ramoconidia <strong>and</strong> hila. Note the rather strong ornamentation in which smaller “particles”<br />
are between larger ones. F. Three conidia in a row. Note the scar formation in the chain <strong>and</strong> the reduction of the size of the cells throughout the spore-chain. <strong>The</strong> inset shows<br />
the resemblance of the scars on a conidiophore <strong>and</strong> on a secondary ramoconidium. Scale bars: A = 50 µm, B–C, F (inset) = 10 µm, D–E = 5 µm, F = 2 µm.<br />
Ascomata pseudothecial, black, globose, erumpent to superficial,<br />
up to 200 µm diam, with 1(–3) short, periphysate ostiolar necks;<br />
wall consisting of 3–6 layers of medium red-brown textura<br />
angularis. Asci fasciculate, bitunicate, subsessile, obovoid to<br />
broadly ellipsoid, straight to slightly curved, 8-spored, 65–85 × 13–<br />
17 µm. Pseudoparaphyses absent in host material, but remnants<br />
observed when studied in culture, hyaline, septate, subcylindrical,<br />
anastomosing, 3–4 µm wide. Ascospores tri- to multiseriate,<br />
overlapping, hyaline, with irregular luminar inclusions, thickwalled,<br />
straight to slightly curved, fusoid-ellipsoidal with obtuse<br />
ends, widest near middle of apical cell, medianly 1-septate, not to<br />
slightly constricted at the septum, tapering towards both ends, but<br />
more prominently towards the lower end, (17–)20–23(–25) × (6–)<br />
7(–8) µm; becoming brown <strong>and</strong> verruculose in asci. Ascospores<br />
germinating after 24 h on MEA from both ends, with spore body<br />
becoming prominently constricted at the septum, but not distorting,<br />
up to 7 µm wide, hyaline to pale brown <strong>and</strong> appearing somewhat<br />
verruculose, enclosed in a mucoid sheath, with germ tubes being<br />
irregular, somewhat nodular.<br />
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Schubert et al.<br />
almost smooth to minutely verruculose or irregularly rough-walled,<br />
sometimes forming clavate conidia, up to 33 µm long, 0–2-septate.<br />
Conidiogenous cells integrated, terminal or conidiophores reduced<br />
to conidiogenous cells, narrowly cylindrical or filiform, with a single<br />
or two loci. Conidia catenate, in unbranched or loosely branched<br />
chains with branching mostly occurring in the lower part of the chain,<br />
straight to slightly curved, small terminal conidia without distal hilum<br />
obovoid, 4–10 × 3–5(–6) µm [av. ± SD, 7.8 (± 1.9) × 4.7 (± 0.9) µm],<br />
aseptate, intercalary conidia with a single or sometimes up to three<br />
distal hila limoniform, ellipsoid to subcylindrical, 6–16 × 4–6 µm<br />
[av. ± SD, 12.4 (± 1.6) × 5.3 (± 0.6) µm], 0–1-septate, secondary<br />
ramoconidia with up to four distal hila, ellipsoid to cylindrical-oblong,<br />
12–25(–35) × (3–)5–7(–9) µm [av. ± SD, 18.8 (± 4.5) × 6.2 (± 0.9)<br />
µm], 0–1(–2)-septate, rarely with up to three septa, sometimes<br />
distinctly constricted at the septum, septum median or somewhat<br />
in the upper or lower half, pale greyish brown or brown to medium<br />
brown or greyish brown, minutely verruculose to verrucose, walls<br />
slightly to distinctly thickened, guttulate to somewhat granular,<br />
usually only slightly attenuated towards apex <strong>and</strong> base, apex<br />
obtuse or slightly truncate, towards the base sometimes distinctly<br />
attenuated with hila situated on short stalk-like prolongations, hila<br />
slightly to distinctly protuberant, truncate to slightly convex, (0.8–)<br />
1–2.5(–3) µm wide, 0.5–1 µm high, somewhat thickened <strong>and</strong><br />
darkened-refractive; microcyclic conidiogenesis occurring, conidia<br />
forming micro- <strong>and</strong> macronematous secondary conidiophores.<br />
Fig. 16. Davidiella tassiana (RO, holotype). Ascus <strong>and</strong> ascospores. Scale bar = 10<br />
µm. P.W. Crous del.<br />
Mycelium superficial, loosely branched, (0.5–)1–5 µm wide, septate,<br />
sometimes constricted at septa, hyaline, subhyaline to pale brown,<br />
smooth or almost so to verruculose or irregularly rough-walled,<br />
sometimes appearing irregular in outline due to small swellings<br />
<strong>and</strong> constrictions, walls unthickened to somewhat thickened, cell<br />
lumen appearing to be granular. Conidiophores both macro- <strong>and</strong><br />
micronematous, arising laterally from plagiotropous hyphae or<br />
terminally from ascending hyphae. Macronematous conidiophores<br />
erect, straight to flexuous, somewhat geniculate-sinuous, nodulose<br />
to nodose with unilateral or multilateral swellings, with a single to<br />
numerous swellings in short succession giving the stalk a knotty/<br />
gnarled appearance, unbranched or occasionally branched, up to<br />
three times, sometimes with a lateral branch-like proliferation below<br />
or at the apex, 10–320 × 3.5–5 µm, swellings 5–8(–9) µm wide,<br />
pluriseptate, septa sometimes constricted when formed after a<br />
node, pale to medium brown, older ones almost dark brown, paler<br />
towards the apex, smooth or minutely verruculose, walls thickened,<br />
sometimes even two-layered. Conidiogenous cells integrated,<br />
terminal or intercalary, nodulose to nodose, with a single or up<br />
to five swellings per cell, 10–24 µm long, proliferation sympodial,<br />
with several conidiogenous loci confined to swellings, mostly<br />
situated on small lateral shoulders, more or less protuberant,<br />
broadly truncate to slightly convex, 1.5–2.5 µm diam, thickened<br />
<strong>and</strong> somewhat darkened-refractive. Micronematous conidiophores<br />
hardly distinguishable from hyphae, sometimes only as short lateral<br />
outgrowth with a single apical scar, short, conical to almost filiform<br />
or narrowly cylindrical, non-nodulose, not geniculate, unbranched,<br />
5–120 × 1.5–3(–4) µm, pluriseptate, not constricted at septa,<br />
cells usually very short, 5–15 µm long, subhyaline to pale brown,<br />
Cultural characteristics: Colonies on PDA reaching 19–37 mm diam<br />
after 14 d at 25 ºC, grey-olivaceous to olivaceous-grey, whitish<br />
to smoke-grey or pale olivaceous-grey due to abundant aerial<br />
mycelium, velvety, reverse olivaceous-grey or iron-grey, margin<br />
almost colourless, regular, entire edge, glabrous to feathery,<br />
aerial mycelium abundant mainly in the colony centre, dense,<br />
felty, woolly, sometimes becoming somewhat reddish brown,<br />
fawn coloured, growth regular, flat to low convex with an elevated<br />
colony centre, sometimes forming few large prominent exudates,<br />
sporulation profuse. Colonies on MEA reaching 17–37 mm diam<br />
after 14 d at 25 ºC, smoke-grey to pale olivaceous-grey towards<br />
margin, olivaceous-grey to iron-grey reverse, velvety, margin white,<br />
entire edge to slightly undulate, aerial mycelium abundant, dense,<br />
fluffy to felty, growth low convex or raised, radially furrowed, folded<br />
<strong>and</strong> wrinkled in the colony centre, without prominent exudates but<br />
sporulating. Colonies on OA reaching 12–28 mm diam after 14<br />
d at 25 ºC, olivaceous-grey to iron-grey, due to abundant aerial<br />
mycelium pale olivaceous-grey, olivaceous-grey reverse, margin<br />
narrow, more or less undulate, white, aerial mycelium white, loose to<br />
dense, high, fluffy to felty, covering large parts of the colony, growth<br />
flat to low convex, without prominent exudates, sporulating.<br />
Specimens examined: Sine loco, sine dato, L 910.225-733, lectotype of C.<br />
herbarum, selected by Prasil & de Hoog, 1988. Sine loco, on leaves of Chionanthus<br />
sp. (Oleaceae), L 910.255-872 = L-0115833, holotype of Dematium epiphyllum<br />
var. (β) chionanthi. Netherl<strong>and</strong>s, Wageningen, isolated from Hordeum vulgare<br />
(Poaceae), 2005, P.W. Crous, <strong>CBS</strong>-H 19853, epitype designated here of C.<br />
herbarum <strong>and</strong> D. tassiana, isoepitype HAL 2022 F, ex-type cultures, CPC 12177 =<br />
<strong>CBS</strong> 121621, CPC 12178–12179, 12181, 12183. Italy, on upper <strong>and</strong> lower surface<br />
of dead leaves of Carex nigra [“fusca”] (Cyperaceae), Tassi no. 862, RO, holotype<br />
of Davidiella tassiana. U.S.A., Colorado, San Juan Co., above Little Molas Lake,<br />
isolated from stems of Delphinium barbeyi (Ranunculaceae), 12 Sep. 2004, A.<br />
Ramaley, <strong>CBS</strong>-H 19868 (teleomorph), single ascospore isolates, <strong>CBS</strong> 121622 =<br />
CPC 11600, CPC 11601–11604.<br />
Substrate <strong>and</strong> distribution: On fading <strong>and</strong> decaying plant material,<br />
on living leaves (phylloplane fungus), as secondary invader, as an<br />
endophyte, isolated from air, soil, foodstuffs, paints, textiles <strong>and</strong><br />
numerous other materials; cosmopolitan.<br />
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<strong>Cladosporium</strong> herbarum species complex<br />
Fig. 17. <strong>Cladosporium</strong> herbarum (CPC 11600). Macro- <strong>and</strong> micronematous conidiophores <strong>and</strong> conidia. Scale bar = 10 µm. K. Schubert del.<br />
Literature: de Vries (1952: 71), Hughes (1958: 750), Ellis (1971:<br />
313), Domsch et al. (1980: 204), Sivanesan (1984: 225), Ellis &<br />
Ellis (1985: 290, 468, 1988: 168), Prasil & de Hoog (1988), Wang<br />
& Zabel (1990: 202), McKemy & Morgan-Jones (1991), Dugan &<br />
Roberts (1994), David (1997: 59), Ho et al. (1999: 129), de Hoog et<br />
al. (2000: 587), Samson et al. (2000: 110), Samson et al. (2001).<br />
Notes: De Vries (1952) incorrectly selected a specimen of Link’s<br />
herbarium at herb. B as lectotype. Prasil & de Hoog (1988) discussed<br />
this typification <strong>and</strong> designated one of Persoon’s original specimens<br />
as lectotype in which C. herbarum could be recognised. <strong>The</strong> latter<br />
material, which is in poor condition, could be re-examined within<br />
the course of these investigations <strong>and</strong> showed conidia agreeing<br />
with the current species concept of C. herbarum being (6–)9.5–<br />
14.5(–21) × (5–)6–7(–8) µm. Since the identity of the strain <strong>CBS</strong><br />
177.71 chosen by Prasil & de Hoog (1988) as representative living<br />
strain of C. herbarum could not be corroborated, an epitype with<br />
a living ex-epitype culture is designated. <strong>The</strong> holotype specimen<br />
of D. tassiana (RO) is morphologically <strong>similar</strong> to that observed on<br />
the epitype of C. herbarum, having ascospores which are (17–)21–<br />
www.studiesinmycology.org<br />
23(–25) × (6–)7–8(–8.5) µm, turning brown <strong>and</strong> verruculose in asci<br />
with age. However, no hamathecial remnants were observed in<br />
ascomata in vivo.<br />
<strong>The</strong> connection to the teleomorph D. tassiana could be<br />
confirmed, which is in agreement with the findings of von Arx (1950)<br />
<strong>and</strong> Barr (1958). Ascospore isolates formed the typical C. herbarum<br />
anamorph in culture, <strong>and</strong> these anamorph cultures developed<br />
some immature fruiting bodies within the agar. When inoculated<br />
onto water agar plates with nettle stems, numerous ascomata with<br />
viable ascospores were formed in culture.<br />
<strong>Cladosporium</strong> iridis (Fautrey & Roum.) G.A. de Vries, Contr.<br />
Knowl. Genus <strong>Cladosporium</strong>: 49. 1952. Figs 20–21.<br />
Basionym: Scolicotrichum iridis Fautrey & Roum., Rev. Mycol.<br />
(Toulouse) 13: 82. 1891.<br />
≡ Heterosporium iridis (Fautrey & Roum.) J.E. Jacques, Contr. Inst. Bot.<br />
Univ. Montréal 39: 18. 1941.<br />
For additional synonyms see Dugan et al. (2004).<br />
Teleomorph: Davidiella macrospora (Kleb.) Crous & U. Braun,<br />
Mycol. Progr. 2: 10. 2003.<br />
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Schubert et al.<br />
Fig. 18. <strong>Cladosporium</strong> herbarum (CPC 11600) <strong>and</strong> its teleomorph Davidiella tassiana (from the host <strong>and</strong> CPC 12181). A–B. Macronematous conidiophores. C. Micronematous<br />
conidiophore. D. Microcyclic conidiogenesis. E. Conidial chain. F. Ascomata on the leaf. G. Ascomata formed in culture on nettle stems. H–I. Asci on the host. J–K. Ascospores<br />
in culture. L. Asci in culture. Scale bars: A, E, H, J–L = 10 µm, F–G, I = 200 µm.<br />
Basionym: Didymellina macrospora Kleb., Ber. Deutsch. Bot. Ges.<br />
42: 60, 1924. 1925.<br />
≡ Mycosphaerella macrospora (Kleb.) Jørst., Meld. Stat. Plantepatol. Inst.<br />
1: 20. 1945.<br />
Mycelium branched, 2–8 µm wide, septate, not constricted at the<br />
septa, hyaline to pale brown, smooth, walls slightly thickened,<br />
sometimes guttulate. Conidiophores very long, usually terminally<br />
arising from ascending hyphae, erect to subdecumbent, slightly<br />
to distinctly flexuous, geniculate-sinuous, usually several times,<br />
subnodulose due to geniculate, sympodial proliferation forming<br />
swollen lateral shoulders, unbranched, rarely branched, up to<br />
720 µm long, 6–11 µm wide, swellings 8–11(–14) µm wide,<br />
pluriseptate, often very regularly septate, not constricted at<br />
the septa, pale to medium olivaceous-brown, somewhat paler<br />
towards the apex, smooth to minutely verruculose, walls only<br />
slightly thickened. Conidiogenous cells integrated, terminal as<br />
well as intercalary, cylindrical-oblong, 15–55 µm long, proliferation<br />
percurrent to sympodial, usually with a single geniculation forming<br />
laterally swollen shoulders often below a septum, conidiogenous<br />
loci confined to swellings, usually one locus per swelling, rarely<br />
two, protuberant, (2–)2.5–4 µm diam, somewhat thickened<br />
<strong>and</strong> darkened-refractive. Conidia solitary, sometimes in short,<br />
unbranched chains, straight to curved, young conidia pyriform to<br />
subcylindrical, connection between conidiophore <strong>and</strong> conidium<br />
being rather broad, subhyaline to pale olivaceous-brown, walls<br />
slightly thickened, then enlarging <strong>and</strong> becoming more thick-walled,<br />
cylindrical-oblong, soleiform with age, both ends rounded, usually<br />
with a slightly to distinctly bulbous base, visible from a very early<br />
stage, but broadest part often towards the apex not at the base, (18–)<br />
30–75(–87) × (7–)10–16(–18) µm [av. ± SD, 53.3 (± 17.8) × 12.6 (±<br />
2.2) µm], (0–)2–6(–7)-septate, usually not constricted at the septa,<br />
rarely slightly constricted, septa often becoming sinuous with age,<br />
pale to medium olivaceous-brown, sometimes darker, verrucose to<br />
echinulate, walls thickened, especially in older conidia, up to 1 µm<br />
thick, hila protuberant, often stalk-like or conically prolonged, up<br />
to 2 µm long, (2–)2.5–3.5(–4) µm diam, with age becoming more<br />
sessile, sometimes just visible as a thickened plate just below the<br />
outer wall layer, especially in distal scars of branched conidia,<br />
periclinal rim often distinctly visible, hila somewhat thickened <strong>and</strong><br />
darkened-refractive; microcyclic conidiogenesis not observed.<br />
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<strong>Cladosporium</strong> herbarum species complex<br />
Fig. 19. <strong>Cladosporium</strong> herbarum (CPC 11600). A. Overview of hyphal growth <strong>and</strong> conidiophore formation of a colony on SNA. Conidiophores are often formed on very wide<br />
(approx. 10 μm), septate hyphae that often grow near the agar surface. B. A more detailed view on colony organisation reveals the ornamented conidia. Note the septum near the<br />
conidiophore (arrow). C. Detail of spore ornamentation <strong>and</strong> hila on a secondary ramoconidium (arrow). Ornamentation is visible during early stages of spore formation (arrow).<br />
D. Structure of the conidiophore, illustrating the complex morphology of the spore-forming apparatus. In addition, secondary ramoconidia, conidia, <strong>and</strong> a hilum on the conidium<br />
are visible. E. Complex structure of the spore-forming apparatus. F. Details of secondary ramoconidia with complex scar-pattern on the right cell. G. Details of a secondary<br />
ramoconidium giving rise to conidia. Note the lack of ornamentation at the location of spore formation. Scale bars: A = 50 µm, B, F = 10 µm, C–E, G = 5 µm.<br />
Cultural characteristics: Colonies on PDA reaching 19–23 mm<br />
diam after 14 d at 25 ºC, pale greenish olivaceous, smoke-grey<br />
to olivaceous-grey due to abundant aerial mycelium, greenish<br />
olivaceous to olivaceous reverse, margin broad, regular, entire<br />
edge to slightly undulate, feathery, aerial mycelium abundantly<br />
formed, felty, fluffy, covering large parts of the colony, mainly in<br />
the central parts, high, growth low convex with a somewhat raised<br />
colony centre. Colonies on MEA reaching 9–23 mm diam after 14 d<br />
at 25 ºC, pale olivaceous-grey to olivaceous-grey, olivaceous-grey<br />
reverse, felty, margin slightly undulate, white, somewhat raised,<br />
aerial mycelium abundant, loose, diffuse, high, growth low convex,<br />
radially furrowed, slightly folded. Colonies on OA reaching 10–19<br />
mm diam after 14 d at 25 ºC, olivaceous, margin broad, undulate,<br />
white, aerial mycelium white, very high, loose, diffuse, hairy, growth<br />
flat, due to the mycelium low convex, without prominent exudates<br />
<strong>and</strong> sporulating on all media.<br />
Specimens examined: Isolated from Iris sp. (Iridaceae), <strong>CBS</strong> 107.20. France,<br />
Cote d’Or, Jardin de Noidan, on leaves of Iris germanica, Jul. 1880, F. Fautrey,<br />
Roumeguère, Fungi Sel. Gall. Exs. No. 5689, PC, lectotype of C. iridis, selected<br />
by David, 1997; K, isolectotype. Netherl<strong>and</strong>s, Boterenbrood, isolated from leaves<br />
of Iris sp., Aug. 1940, <strong>CBS</strong>-H 19859, epitype designated here of C. iridis, culture<br />
ex-epitype <strong>CBS</strong> 138.40.<br />
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Schubert et al.<br />
Fig. 20. <strong>Cladosporium</strong> iridis (<strong>CBS</strong> 138.40). Conidiophores <strong>and</strong> conidia. Scale bar = 10 µm. K. Schubert del.<br />
Fig. 21. <strong>Cladosporium</strong> iridis (teleomorph Davidiella macrospora) (<strong>CBS</strong> 138.40). A–C. Conidiophores with conidia. D. Conidium. Scale bar = 10 µm.<br />
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<strong>Cladosporium</strong> herbarum species complex<br />
Fig. 22. <strong>Cladosporium</strong> macrocarpum (<strong>CBS</strong> 299.67). Macro- <strong>and</strong> micronematous conidiophores <strong>and</strong> conidia. Scale bar = 10 µm. K. Schubert del.<br />
Substrates <strong>and</strong> distribution: Leaf spot <strong>and</strong> blotch of Iris spp.<br />
including I. crocea, I. florentina, I. foetidissima, I. germanica, I.<br />
gueldenstaedtiana, I. kamaonensis, I. pallida, I. plicata (= I. swertii<br />
Hort.), I. pseudacorus, I. pumila, I. spuria ssp. halophila, <strong>and</strong> other<br />
species, also on Belacam<strong>and</strong>a chinensis (= Gemmingia chinensis),<br />
Hemerocallis fulva, Gladiolus g<strong>and</strong>avensis; Africa (Algeria,<br />
Morocco, South Africa, Zambia, Zimbabwe), Asia (Armenia,<br />
Azerbaijan, China, Georgia, India, Iran, Israel, Japan, Kazakhstan,<br />
Kirgizstan, Korea, Russia, Turkey, Turkmenistan, Uzbekistan),<br />
Australasia (Australia, New Zeal<strong>and</strong>), Europe (Austria, Belgium,<br />
Belorussia, Cyprus, Czech Republic, Denmark, Estonia, France,<br />
Germany, Great Britain, Greece, Italy, Latvia, Lithuania, Malta,<br />
Moldavia, Montenegro, Netherl<strong>and</strong>s, Norway, Pol<strong>and</strong>, Romania,<br />
Russia, Serbia, Spain, Sweden, Ukraine), North America (Canada,<br />
U.S.A.), Central & South America (Argentina, Chile, Jamaica,<br />
Panama, Uruguay).<br />
Literature: Ellis (1971: 312), Ellis & Waller (1974), Sivanesan (1984:<br />
222), McKemy & Morgan-Jones (1990), David (1997: 43), Shin et<br />
al. (1999).<br />
Notes: <strong>The</strong> description of the morphological parameters in culture<br />
is based on the isolate sporulating on PDA, since sporulation on<br />
SNA was not observed. <strong>The</strong> conidiophores <strong>and</strong> conidia in vivo are<br />
usually wider than in culture [conidiophores (6–)9–15(–17) µm<br />
wide, conidia (11–)15–23(–28) µm].<br />
www.studiesinmycology.org<br />
<strong>Cladosporium</strong> macrocarpum Preuss, in Sturm, Deutsch. Fl.<br />
3(26): 27. 1848. Figs 22–25.<br />
≡ <strong>Cladosporium</strong> herbarum var. macrocarpum (Preuss) M.H.M. Ho &<br />
Dugan, in Ho et al., Mycotaxon 72: 131. 1999.<br />
= Dematium herbarum var. (β) brassicae Pers., Syn. meth. fung. 2: 699. 1801,<br />
syn. nov.<br />
= Dematium graminum Pers., Mycol. eur. 1: 16. 1822, syn. nov.<br />
= Dematium vulgare var. (δ) typharum Pers., Mycol. eur. 1: 14. 1822, syn.<br />
nov.<br />
= Dematium vulgare var. (β) foliorum Pers., Mycol. eur. 1: 14. 1822, syn. nov.<br />
For additional synonyms see Dugan et al. (2004), Schubert<br />
(2005).<br />
Teleomorph: Davidiella macrocarpa Crous, K. Schub. & U. Braun,<br />
sp. nov. MycoBank MB504582.<br />
Davidiellae tassianae similis, sed pseudoparaphysibus prominentibus et ascosporis<br />
maioribus, (22–)23–26(–28) × (6–)6.5–7(–8) µm.<br />
Ascomata superficial on a small stroma, black, up to 200 µm<br />
diam, globose, separate, but developing with 1–3 necks with age;<br />
ostioles consisting of pale brown to subhyaline cells, periphysate,<br />
with periphysoids growing into the cavity; wall consisting of 3–6<br />
layers of medium brown textura angularis. Pseudoparaphyses<br />
present, hyaline, subcylindrical, septate, anastomosing, 3–4<br />
µm diam; hamathecial cells persistent in cavity. Asci fasciculate,<br />
bitunicate, subsessile, broadly ellipsoid with a long tapered stalk,<br />
straight to curved, 8-spored, 70–110 × 15–20 µm. Ascospores<br />
tri- to multiseriate, overlapping, hyaline, guttulate, irregular lumina<br />
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Schubert et al.<br />
Fig. 23. <strong>Cladosporium</strong> macrocarpum (<strong>CBS</strong> 299.67) <strong>and</strong> its teleomorph Davidiella macrocarpa (CPC 12755). A–C. Macronematous conidiophores <strong>and</strong> conidia. D–G.<br />
Micronematous conidiophores. H. Microcyclic conidiogenesis. I. Ascomata formed on nettle stems in culture. J. Periphyses. K, M–N. Asci. L. Ostiole. Scale bars: A, D–H, J–N<br />
= 10 µm, I = 200 µm.<br />
rarely observed, thick-walled, straight to slightly curved, fusoidellipsoidal<br />
with obtuse ends, widest in the middle of the apical<br />
cell, medianly 1-septate, not to slightly constricted at the septum,<br />
tapering towards both ends, but more prominently towards lower<br />
end, (22–)23–26(–28) × (6–)6.5–7(–8) µm; mucoid sheath rarely<br />
observed, mostly absent.<br />
Mycelium unbranched or loosely branched, 1–4.5(–5) µm wide,<br />
septate, sometimes slightly constricted at septa, hyaline to pale<br />
brown, smooth to minutely verruculose, walls unthickened or slightly<br />
thickened. Conidiophores micronematous <strong>and</strong> macronematous,<br />
solitary, arising terminally from plagiotropous hyphae or terminally<br />
from ascending hyphae. Macronematous conidiophores erect,<br />
straight to somewhat flexuous, cylindrical-oblong, nodulose to<br />
nodose, with a single apical or usually several swellings either<br />
somewhat distinct from each other or often in short succession giving<br />
conidiophores a knotty appearance, swellings sometimes laterally<br />
elongated or formed at the top of a branch-like outgrowth below<br />
the apical swelling, sometimes distinctly geniculate, unbranched,<br />
sometimes branched, 12–260 × (3–)4–6 µm, swellings 5–10 µm<br />
wide, pluriseptate, sometimes slightly constricted at septa, pale to<br />
medium brown or olivaceous-brown, somewhat paler at apices,<br />
130
<strong>Cladosporium</strong> herbarum species complex<br />
Fig. 24. <strong>Cladosporium</strong> macrocarpum (<strong>CBS</strong> 299.67). A. Survey of a conidiophore that forms several secondary ramoconidia <strong>and</strong> conidia. Several aerial hyphae are also visible<br />
in this picture. B. Conidiophore with broadly ellipsoid secondary ramoconidia <strong>and</strong> obovoid conidia. Note the different scars on the conidiophore at the lower left. C. Ellipsoid<br />
or obovoid conidia with notable areas of scar formation. <strong>The</strong> ornamentation is relatively widely distributed over the body of the cell <strong>and</strong> <strong>similar</strong> to C. variabile. D. Detail of<br />
a conidiophore (see B) with scars. Note the relatively shallow rings of the scars. E. Details of conidia <strong>and</strong> a secondary ramoconidium. F. Conidiophore with a secondary<br />
ramoconidium <strong>and</strong> conidia. Note the hila on several spores <strong>and</strong> the lack of ornamentation at the site where spores are formed. Scale bars: A–C, = 10 µm, D, F = 5 µm, E = 2<br />
µm.<br />
smooth to minutely verruculose or verruculose, walls somewhat<br />
thickened, sometimes even two-layered. Conidiogenous cells<br />
integrated, terminal or intercalary, cylindrical, nodulose with lateral<br />
shoulders or nodose with swellings round about the stalk, with<br />
conidiogenous loci confined to swellings, 12–37 µm long, with up to<br />
12 loci per cell, usually with up to six, loci conspicuous, protuberant,<br />
(1–)1.5–2 µm diam, somewhat thickened <strong>and</strong> darkened-refractive.<br />
Micronematous conidiophores almost indistinguishable from<br />
hyphae, straight, narrowly filiform, non-nodulose or with a single or<br />
few swellings, mostly with small head-like swollen apices, usually<br />
www.studiesinmycology.org<br />
131
Schubert et al.<br />
or branched chains, subglobose, obovoid to limoniform, ellipsoid<br />
or fusiform, 2.5–16 × 1.5–5 µm, 0(–1)-septate, few longer conidia<br />
subcylindrical to clavate, up to 37(–43) µm long, 0–2(–3)-septate,<br />
occasionally with up to four septa, sometimes slightly constricted<br />
at the septa, subhyaline to pale brown, almost smooth to minutely<br />
verruculose, walls unthickened, hila 0.8–1.2 µm diam, thickened<br />
<strong>and</strong> darkened-refractive.<br />
Cultural characteristics: Colonies on PDA reaching 30–43 mm<br />
in diam after 14 d at 25 ºC, dark dull green to olivaceous-grey,<br />
olivaceous-grey, dark olivaceous- to iron-grey reverse, pulvinate,<br />
velvety, sometimes somewhat zonate, paler zones towards the<br />
margin, margin regular, entire edge, almost colourless to white,<br />
glabrous to feathery, aerial mycelium sparse to more abundant<br />
in the colony centre or covering large areas of the colony, hairy,<br />
fluffy or felty, whitish to smoke-grey, sometimes becoming reddish,<br />
livid red to vinaceous, growth flat, regular, sometimes forming<br />
few prominent exudates, exudates sometimes slightly reddish,<br />
sporulation profuse with two kinds of conidiophores, low <strong>and</strong> high.<br />
Colonies on MEA reaching 31–50 mm in diam after 14 d at 25<br />
ºC, grey-olivaceous to olivaceous-grey or iron-grey, sometimes<br />
pale olivaceous-grey to whitish due to abundant aerial mycelium,<br />
olivaceous-grey or iron-grey reverse, velvety or powdery, margin<br />
narrow, entire edge, colourless to white, glabrous, aerial mycelium<br />
sparse to abundant, hairy or felty, growth regular, flat to low convex,<br />
radially furrowed, without prominent exudates, sporulation profuse.<br />
Colonies on OA reaching 29–40 mm in diam after 14 d at 25 ºC,<br />
grey-olivaceous, olivaceous-grey to dark smoke-grey, olivaceousblack<br />
or iron grey reverse, margin entire edge, narrow, colourless<br />
or white, glabrous, aerial mycelium sparse, mainly in the colony<br />
centre, felty, white to smoke-grey or grey-olivaceous, felty, growth<br />
flat, regular, without exudates, sporulating.<br />
Fig. 25. Davidiella macrocarpa (CPC 12755). Ascus <strong>and</strong> ascospores. Scale bar =<br />
10 µm. P.W. Crous del.<br />
only few micrometer long, 1.5–3 µm wide, aseptate or with only<br />
few septa, subhyaline, smooth or almost so, walls unthickened,<br />
with a single or only few conidiogenous loci, narrow, 0.8–1.2 µm<br />
diam, thickened <strong>and</strong> somewhat darkened-refractive. Conidia<br />
catenate, in branched chains, small terminal conidia subglobose,<br />
obovoid, oval, limoniform, 4–11 × (3–)4–6 µm [av. ± SD, 7.6 (±<br />
1.9) × 5.0 (± 0.8) µm], aseptate, intercalary conidia broadly ovoidellipsoid,<br />
10–17 × (4.5–)5–9 µm [av. ± SD, 12.7 (± 2.1) × 6.8 (±<br />
0.8) µm], 0–1-septate; secondary ramoconidia broadly ellipsoid<br />
to subcylindrical, 14–25(–30) × (5–)6–9(–10) µm [av. ± SD, 19.4<br />
(± 3.5) × 7.6 (± 1.0) µm], 0–2(–3)-septate, sometimes slightly<br />
constricted at the septa, septa somewhat sinuous with age, pale<br />
brown to medium olivaceous-brown or brown, sometimes even<br />
dark brown, verruculose to echinulate (muricate under SEM),<br />
walls thickened, up to 1 µm thick, mostly broadly rounded at<br />
apex <strong>and</strong> base, sometimes attenuated, sometimes guttulate<br />
by oil drops, with up to three apical hila, mostly 1–2, hila sessile<br />
(apparently somewhat immersed) to somewhat protuberant, 1–<br />
2(–2.5) µm diam, thickened <strong>and</strong> darkened-refractive; microcyclic<br />
conidiogenesis occurring with conidia forming secondary micro<strong>and</strong><br />
macronematous conidiophores, conidia often germinating with<br />
long hyphae. Conidia formed by micronematous conidiophores<br />
usually smaller, narrower <strong>and</strong> paler, catenate, in short unbranched<br />
Specimens examined: Sine loco et dato, L 910.255-723 = L-0115836, lectotype<br />
designated here of Dematium graminum. Sine loco, on dead stems of Brassica sp.<br />
(Brassicaceae), No. 601, L 910.255-716 = L-0115849, holotype of D. herbarum var.<br />
(β) brassicae. Sine loco, on leaves of Iris (Iridaceae), Quercus (Fagaceae), Brassica<br />
etc., L 910.255-736 = L-0115871, holotype of D. vulgare var. (β) foliorum, isotype L<br />
910.255-718 = L-0115872. Sine loco et dato, L 910.255-698 = L-0115852, lectotype<br />
designated here for D. vulgare var. (δ) typharum. Isolated from “Mycosphaerella<br />
tulasnei”, <strong>CBS</strong> 223.32 = ATCC 11287 = IMI 049635. Romania, isolated from water,<br />
<strong>CBS</strong> 175.82. Slovenia, Sečovlje, isolated from hypersaline water from salterns<br />
(precrystalisation pond), 2004, P. Zalar, EXF-2287 = CPC 12054. Turkey, Ankara,<br />
Tekeli, isolated from Triticum aestivum (Poaceae), isol. S. Tahsin, ident. A.C. Stolk,<br />
<strong>CBS</strong> 299.67. U.S.A., Seattle, University of Washington Campus, 47.6263530, -<br />
122.3331440, isolated from cleistothecia of Phyllactinia guttata (Erysiphaceae)<br />
on leaves of Corylus sp. (Corylaceae), 16 Sep. 2004, D. Glawe, CPC 11817;<br />
Washington, isolated from Spinacia oleracea (Chenopodiaceae), 1 Jan. 2003,<br />
L. DuToit, <strong>CBS</strong>-H 19855, neotype designated here for C. macrocarpum, <strong>and</strong><br />
holotype of D. macrocarpa, isoneotype HAL 2020 F, isotype HAL 2021 F, culture<br />
ex-type CPC 12752, 12756–12759, CPC 12755 = <strong>CBS</strong> 121623.<br />
Substrate <strong>and</strong> distribution: Decaying plant material, human,<br />
hypersaline water, water; widespread.<br />
Literature: de Vries (1952: 76), Ellis (1971: 315), Domsch et al.<br />
(1980: 208), Ellis & Ellis (1985: 290, 468), Matsushima (1985: 5),<br />
McKemy & Morgan-Jones (1991), Dugan & Roberts (1994), David<br />
(1997: 71), Samson et al. (2000: 112).<br />
Notes: In the absence of Preuss’s type material (not preserved)<br />
de Vries (1952) “lectotypified” C. macrocarpum by a specimen in<br />
Saccardo’s herbarum (Herb. Myc. P.A. Saccardo no. 419, PAD).<br />
This material, subsequently distributed in Mycotheca Italica no.<br />
1396, should correctly be regarded as neotype (David 1997). A<br />
single collection of Saccardo’s Mycotheca Italica no. 1396 from<br />
herb. HBG, which can be considered as isoneotype material,<br />
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<strong>Cladosporium</strong> herbarum species complex<br />
Fig. 26. <strong>Cladosporium</strong> ossifragi (<strong>CBS</strong> 842.91). Conidiophores <strong>and</strong> conidia. Scale bar = 10 µm. K. Schubert del.<br />
was re-examined <strong>and</strong> proved to rather agree with the species<br />
concept of C. herbarum s. str. <strong>The</strong> conidia were formed in simple,<br />
rarely branched chains, 6–26 × (4–)5.5–8(–9) µm, 0–3-septate,<br />
almost smooth or minutely to densely verruculose or verrucose<br />
(Schubert 2005). However, since de Vries’ “lectotypification” was<br />
incorrect according to the code (ICBN, Art. 9.2, 9.17), a neotype is<br />
designated.<br />
<strong>The</strong> delimitation of C. macrocarpum as a morphologically<br />
distinct species from C. herbarum has been controversially<br />
discussed by several authors (McKemy & Morgan-Jones 1991,<br />
Dugan & Robert 1994, Ho et al. 1999). Based on molecular as well<br />
morphological studies, it can be shown that C. macrocarpum is a<br />
well-defined species distinguishable from C. herbarum s. str. by<br />
forming conidiophores with wider nodes, 5–10 µm, wider <strong>and</strong> more<br />
frequently septate conidia [small terminal conidia 4–11 × (3–)4–6<br />
µm versus 4–10 × 3–5(–6) µm in C. herbarum, intercalary conidia<br />
10–17 × (4.5–)5–9 µm versus 6–16 × 4–6 µm in C. herbarum,<br />
secondary ramoconidia 14–25(–30) × (5–)6–9(–10) µm versus 12–<br />
25(–35) × (3–)5–7(–9) µm in C. herbarum] <strong>and</strong> by being connected<br />
to Davidiella macrocarpa. On natural substrates the conidiophores<br />
are usually somewhat wider than in culture, 4–8(–10) µm wide, <strong>and</strong><br />
also the conidia can be somewhat wider, sometimes up to 13(–15) µm.<br />
<strong>Cladosporium</strong> graminum, described by Persoon (1822), as<br />
well as C. brunneum <strong>and</strong> C. gracile, introduced by Corda (1837),<br />
are older synonyms of C. macrocarpum <strong>and</strong>, according to the<br />
code, would have priority. However, since C. macrocarpum is a<br />
well established, currently used name with numerous records in<br />
literature, a proposal to conserve the name against these older<br />
names is in preparation for formal publication in Taxon.<br />
A characteristic difference between ascomata of C.<br />
macrocarpum in comparison to those of C. herbarum, are the<br />
smaller, globose pseudothecia, asci with longer stalks, prominence<br />
of pseudoparaphyses, <strong>and</strong> rather inconspicuous luminar ascospore<br />
inclusions.<br />
<strong>Cladosporium</strong> ossifragi (Rostr.) U. Braun & K. Schub., comb.<br />
nov. MycoBank MB504575. Figs 26–28.<br />
Basionym: Napicladium ossifragi Rostr., Bot. Fǽröes 1: 316.<br />
1901.<br />
≡ Heterosporium ossifragi (Rostr.) Lind, Dan. fung.: 531. 1913.<br />
= Heterosporium magnusianum Jaap, Schriften Naturwiss. Vereins Schleswig-<br />
Holstein 12: 346. 1902.<br />
≡ <strong>Cladosporium</strong> magnusianum (Jaap) M.B. Ellis in Ellis, More<br />
Dematiaceous Hyphomycetes: 337. 1976.<br />
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Schubert et al.<br />
Fig. 27. <strong>Cladosporium</strong> ossifragi (<strong>CBS</strong> 842.91). A. Macronematous conidiophore. B. Micronematous conidiophore. C–D. Conidia. E. Conidia <strong>and</strong> microcyclic conidiogenesis.<br />
Scale bars = 10 µm.<br />
Fig. 28. <strong>Cladosporium</strong> ossifragi (<strong>CBS</strong> 842.91). A. Survey on different secondary ramoconidia <strong>and</strong> conidia. B. Details of conidia <strong>and</strong> hila. Note the very pronounced ornamentation<br />
<strong>and</strong> the absence of ornamentation near the site of spore formation. C. Detail of the end of a secondary ramoconidium with pronounced hila. D. Formation of a new conidium.<br />
Note the broad scar behind it (> 1 μm). E. Formation of a new conidium from a smooth-walled stalk. F. Hila on a secondary ramoconidium. This micrograph is from the sample<br />
before coating with gold-palladium <strong>and</strong> shows <strong>similar</strong> features as the sample after sputter coating. Scale bars: A = 10 µm, B–D, F = 2 µm, E = 5 µm.<br />
134
<strong>Cladosporium</strong> herbarum species complex<br />
Mycelium abundantly formed, twisted, often somewhat aggregated,<br />
forming ropes, branched, 1–5 µm wide, septate, often irregularly<br />
swollen <strong>and</strong> constricted, hyaline or subhyaline to pale brown,<br />
smooth, walls unthickened or only slightly thickened. Conidiophores<br />
macronematous <strong>and</strong> micronematous, arising from plagiotropous<br />
hyphae, terminally or laterally, erect to subdecumbent, more<br />
or less straight to flexuous, cylindrical, sometimes geniculate,<br />
subnodulose with loci often situated on small lateral shoulders,<br />
unbranched, sometimes branched, often very long, up to 350 µm<br />
long, 3–4.5(–5) µm wide, pluriseptate, shorter ones aseptate, not<br />
constricted at septa, pale to pale medium brown, paler towards<br />
apices, sometimes subhyaline, smooth to minutely verruculose,<br />
especially towards apices, walls somewhat thickened, up to 0.5 µm,<br />
sometimes appearing two-layered. Conidiogenous cells integrated,<br />
terminal as well as intercalary, cylindrical, sometimes geniculate,<br />
subnodulose, 5–31 µm long, proliferation sympodial, with few<br />
loci (1–3) per cell, loci usually confined to small lateral shoulders,<br />
protuberant, conspicuous, short cylindrical, 1–2 µm wide, up to<br />
1 µm high, somewhat thickened, darkened-refractive. Conidia<br />
catenate, in short, unbranched or branched chains, straight, small<br />
terminal <strong>and</strong> intercalary conidia subglobose, obovoid to ellipsoid,<br />
4–15 × 3–5 µm [av. ± SD, 9.3 (± 3.7) × 4.0 (± 0.7) µm], 0–1-<br />
septate, not constricted at the septa, pale brown, hila 0.8–1 µm<br />
diam, secondary ramoconidia cylindrical, sometimes ellipsoid or<br />
subfusiform, 16–36(–40) × (4–)5–8 µm [av. ± SD, 26.6 (± 7.4) ×<br />
6.0 (± 1.2) µm], (0–)1–3(–4)-septate [in vivo wider, (6–)7–9(–11)<br />
µm, <strong>and</strong> with up to five, rarely seven septa], not constricted at<br />
the septa, septa sometimes slightly sinuous, pale brown to pale<br />
medium brown, densely verruculose, verrucose to echinulate<br />
(densely muricate under SEM), walls unthickened to somewhat<br />
thickened, rounded or somewhat attenuated at apex <strong>and</strong> base,<br />
hila protuberant, conspicuous, sometimes situated on short, small<br />
prolongations, 1–2.5 µm diam, somewhat thickened <strong>and</strong> darkenedrefractive;<br />
microcyclic conidiogenesis occasionally occurring.<br />
Cultural characteristics: Colonies on PDA reaching 53 mm diam<br />
after 14 d at 25 ºC, greenish olivaceous, grey-olivaceous to<br />
olivaceous-grey or iron-grey, appearing somewhat zonate, dull<br />
green to olivaceous-black reverse, margin colourless, regular,<br />
entire edge, aerial mycelium abundantly formed, covering at first<br />
the colony centre later most of the surface, dense, high, growth flat<br />
with elevated colony centre, somewhat folded. Colonies on MEA<br />
reaching 54 mm diam after 14 d at 25 ºC, pale olivaceous-grey<br />
to olivaceous-grey in the centre, iron-grey reverse, velvety, margin<br />
colourless to white, entire edge, radially furrowed, aerial mycelium<br />
abundantly formed, fluffy to felty, growth flat with somewhat raised,<br />
folded colony centre. Colonies on OA attaining 52 mm diam after<br />
14 d at 25 ºC, olivaceous-grey to iron-grey, iron-grey to greenish<br />
black reverse, margin white, entire edge, aerial mycelium diffuse,<br />
loose, growth flat, prominent exudates absent, sporulation profuse<br />
on all media.<br />
Specimens examined: Denmark, Undallslund, on leaves of Narthecium ossifragum<br />
(Melanthiaceae), 13 Sep. 1885, E. Rostrup, CP, neotype designated here of C.<br />
ossifragi; Tǿnder, Rǿmǿ near Twismark, 19 Aug. 1911, H. Sydow, Sydow, Mycoth.<br />
Germ. 1047, M. Germany, Hamburg, Eppendorfer Moor, on leaves of Narthecium<br />
ossifragum, 12 Sep. 1897, O. Jaap, HBG, lectotype selected here of C.<br />
magnusianum; 4 Sep. 1903, O. Jaap, Jaap, Fungi Sel. Exs. 49, M; Wernerwald near<br />
Cuxhaven, Aug. 1927, A. Ludwig, Petrak, Mycoth. Gen. 146, M. Norway, Bjerkreim<br />
County, isolated from leaves of Narthecium ossifragum, M. di Menna, <strong>CBS</strong>-H 19860,<br />
epitype designated here of C. ossifragi, culture ex-epitype <strong>CBS</strong> 842.91 = ATCC<br />
200946; Møre og Romsdal County, isolated from leaves of Narthecium ossifragum,<br />
M. di Menna, <strong>CBS</strong> 843.91.<br />
Substrate <strong>and</strong> distribution: Causing leaf spots on Narthecium<br />
ossifragum; Europe (Austria, Denmark, Germany, Great Britain,<br />
Irel<strong>and</strong>, Norway).<br />
Literature: Ellis & Ellis (1985: 390), David (1995a; 1997: 85–86,<br />
88), Ho et al. (1999: 132).<br />
Notes: Type material of Napicladium ossifragi is not preserved in<br />
Rostrup’s herbarium (on Narthecium ossifragum, Faeroe Isl<strong>and</strong>s,<br />
Viderö, Viderejde <strong>and</strong> Österö, Svinaa, sine dato, leg. Ostenfeld &<br />
Harz). However, other authentic collections seen <strong>and</strong> examined<br />
by Rostrup are deposited at CP. Lind (1913) re-examined these<br />
samples, synonymised N. ossifragi with H. magnusianum <strong>and</strong><br />
correctly introduced the combination H. ossifragi. Nevertheless,<br />
the correct oldest name for this fungus has been ignored by<br />
most authors. David (1997), who clearly stated that N. ossifragi is<br />
the earliest name for this species, preferred to use the name C.<br />
magnusianum because the typification of Rostrup’s name was still<br />
uncertain. Despite the lacking type material, there is no doubt about<br />
the correct identity of N. ossifragi since authentic material of this<br />
species, examined by <strong>and</strong> deposited in Rostrup’s herbarium (CP),<br />
is preserved. <strong>The</strong>refore, there is no reason to reject the oldest valid<br />
name for this species. <strong>The</strong> original collection of C. magnusianum<br />
cited by Jaap (1902) (on leaves of Narthecium ossifragum, Denmark,<br />
Tǿnder, Rǿmǿ, peatbog by Twismark, Jul.–Aug. 1901, Jaap), but<br />
not designated as type, is not preserved (David 1997). It is neither<br />
deposited at B, HBG nor S. However, in the protologue Jaap (1902)<br />
also referred to material of this species found near Hamburg, which<br />
is, hence, syntype material available for lectotypification.<br />
<strong>Cladosporium</strong> pseudiridis K. Schub., C.F. Hill, Crous & U. Braun,<br />
sp. nov. MycoBank MB504576. Figs 29–30.<br />
Etymology: Epithet derived from its <strong>similar</strong> morphology to<br />
<strong>Cladosporium</strong> iridis.<br />
Differt a Cladosporio iridis conidiis 0–3-septatis, brevioribus et latioribus, 15–55 ×<br />
(9–)11–19(–21) µm.<br />
Mycelium sparingly branched, 2–7 µm wide, septate, not constricted<br />
at the septa, subhyaline to pale brown, smooth or almost so, walls<br />
somewhat thickened, guttulate or protoplasm appearing granular,<br />
sometimes enveloped by a slime coat. Conidiophores arising<br />
mostly terminally from ascending hyphae, sometimes also laterally<br />
from plagiotropous hyphae, erect, more or less straight, broadly<br />
cylindrical-oblong, once or several times slightly to distinctly<br />
geniculate-sinuous, forming more or less pronounced lateral<br />
shoulders, nodulose, unbranched, 100–320(–500) × 7–11 µm,<br />
swellings 10–14 µm wide, becoming narrower <strong>and</strong> paler towards the<br />
apex, septate, not constricted at the septa, septa mainly basal, apical<br />
cell often very long, pale to medium olivaceous-brown, subhyaline<br />
at the apex, smooth or almost so, sometimes minutely verruculose,<br />
walls usually distinctly thickened, sometimes even two-layered, up<br />
to 1(–2) µm thick, protoplasm granular, often clearly contrasting<br />
from the outer wall. Conidiogenous cells integrated, terminal <strong>and</strong><br />
intercalary, cylindrical-oblong, slightly to distinctly geniculatesinuous,<br />
nodulose with conidiogenous loci confined to swellings<br />
or lateral shoulders, 30–110 µm long, proliferation percurrent to<br />
sympodial, with a single or three, sometimes up to five geniculations<br />
per cell, usually only a single locus per swelling, protuberant, very<br />
prominent, short cylindrical, peg-like, clearly composed of a dome<br />
<strong>and</strong> surrounding rim, dome often higher than the periclinal rim,<br />
broad, somewhat paler than rim, conically narrowed, (2–)2.5–4<br />
µm wide, up to 2 µm high, thickened <strong>and</strong> darkened-refractive.<br />
www.studiesinmycology.org<br />
135
Schubert et al.<br />
Fig. 29. <strong>Cladosporium</strong> pseudiridis (<strong>CBS</strong> 116463). Conidiophores <strong>and</strong> conidia. Scale bar = 10 µm. K. Schubert del.<br />
Conidia solitary, sometimes in short unbranched chains of two or<br />
three, straight to slightly curved, young conidia small, 0–1-septate,<br />
broadly ovoid to pyriform, 15–26 × (9–)11–16(–18) µm [av. ± SD,<br />
19.2 (± 4.3) × 14.2 (± 3) µm], first septum somewhat in the upper<br />
half, the upper cell is much smaller but gradually extending as the<br />
conidium matures, mature conidia 1–3-septate, broadly pyriform,<br />
cylindrical-oblong or soleiform, usually with a distinctly bulbous<br />
base, 30–55 × 12–19(–21) µm [av. ± SD, 41.5 (± 6.8) × 17.1 (± 2.1)<br />
µm], broadest part of conidia usually at the bulbous base, mostly<br />
attenuated towards the basal septum, septa becoming sinuous with<br />
age, pale to medium olivaceous-brown or brown, usually echinulate,<br />
sometimes coarsely verrucose, walls distinctly thickened, up to 2<br />
µm thick, often appearing layered with a large lumen in the centre<br />
of the cell, broadly rounded to flattened at apex <strong>and</strong> base, hila often<br />
very prominent, often peg-like elongated, up to 3 µm long, with age<br />
becoming less prominent, visible as a thickened flat plate just below<br />
the outer echinulate wall layer, slightly raised towards the middle,<br />
2–3.5 µm diam, thickened <strong>and</strong> darkened-refractive; microcyclic<br />
conidiogenesis not observed.<br />
Cultural characteristics: Colonies on PDA attaining 6 mm diam<br />
after 14 d at 25 ºC, whitish, smoke-grey to pale olivaceous-grey<br />
due to abundant aerial mycelium, olivaceous-black reverse, margin<br />
narrow, white, more or less crenate, aerial mycelium zonate, fluffy,<br />
covering most of the colony, mainly in the colony centre, growth<br />
136
<strong>Cladosporium</strong> herbarum species complex<br />
Fig. 30. <strong>Cladosporium</strong> pseudiridis (<strong>CBS</strong> 116463). A–C. Conidiophores <strong>and</strong> conidia. D. Part of a conidiogenous cell showing a protuberant cladosporioid conidiogenous locus.<br />
E–F. Conidia. Scale bars = 10 µm.<br />
convex to raised, deep into the agar, with age few large prominent<br />
exudates formed, sparingly sporulating. Colonies on MEA attaining<br />
7 mm diam after 14 d at 25 ºC, olivaceous-grey, pale olivaceousgrey<br />
to pale rosy-buff due to abundant aerial mycelium covering<br />
almost the whole colony, iron-grey reverse, margin colourless or<br />
white, broad, regular, more or less glabrous, aerial mycelium fluffy,<br />
dense, high, growth convex to umbonate, sometimes with elevated<br />
colony centre, prominent exudates lacking, sporulation sparse.<br />
Colonies on OA attaining 8 mm diam after 14 d at 25 ºC, white, pale<br />
buff to pale olivaceous-grey in the centre, margin grey-olivaceous,<br />
olivaceous- to iron-grey reverse, margin entire edge or somewhat<br />
undulate, somewhat feathery, growth raised with a somewhat<br />
depressed centre forming an elevated outer rim, without prominent<br />
exudates, sporulation more abundant.<br />
Specimen examined: New Zeal<strong>and</strong>, Auckl<strong>and</strong>, Mt. Albert, Carrington Road, Unitec<br />
Campus, isolated from large leaf lesions on Iris sp. (Iridaceae), 15 Aug. 2004, C.F.<br />
Hill, <strong>CBS</strong>-H 19861, holotype, culture ex-type <strong>CBS</strong> 116463 = LYN 1065 = ICMP<br />
15579.<br />
Substrate <strong>and</strong> distribution: On living leaves of Iris sp.; New<br />
Zeal<strong>and</strong>.<br />
Notes: <strong>Cladosporium</strong> pseudiridis closely resembles C. iridis, a<br />
common <strong>and</strong> widespread species causing leaf spots on numerous<br />
Iris spp. <strong>and</strong> a few additional hosts of the host family Iridaceae,<br />
but the latter species is easily distinguishable by having longer<br />
<strong>and</strong> narrower, more frequently septate conidia, (18–)30–75(–87) ×<br />
(7–)10–16(–18) µm, (0–)2–6(–7)-septate.<br />
www.studiesinmycology.org<br />
It is unlikely that C. pseudiridis is of New Zeal<strong>and</strong> origin since<br />
the <strong>genus</strong> Iris is not indigenous to New Zeal<strong>and</strong>. All Iris species<br />
that are found in this country have been introduced, mainly for<br />
horticultural purposes. <strong>The</strong> species is, therefore, probably more<br />
common than indicated above. However, within the course of the<br />
recent monographic studies in the <strong>genus</strong> <strong>Cladosporium</strong> numerous<br />
herbarium specimens, mainly of European origin, have been<br />
examined <strong>and</strong> proved to be correctly identified agreeing with the<br />
species concept of C. iridis. Additional collections <strong>and</strong> cultures are<br />
necessary to determine its distribution.<br />
<strong>Cladosporium</strong> ramotenellum K. Schub., Zalar, Crous & U. Braun,<br />
sp. nov. MycoBank MB504577. Figs 31–33.<br />
Etymology: Refers to the morphological <strong>similar</strong>ity with <strong>Cladosporium</strong><br />
tenellum.<br />
Differt a Cladosporio cladosporioide conidiophoris et conidiis leniter angustioribus,<br />
2–4(–5) µm latis, conidiis 0–2(–3)-septatis, semper verruculosis; et a Cladosporio<br />
tenello locis conidiogenis non numerosis et non aggregatos ad apicem, conidiis<br />
longioribus et angustioribus, 2.5–35 × 2–4(–5) µm, 0–3-septatis.<br />
Mycelium unbranched or only sparingly branched, 1.5–4 µm wide,<br />
septate, without swellings <strong>and</strong> constrictions, hyaline or subhyaline,<br />
smooth, sometimes irregularly rough-walled, walls unthickened.<br />
Conidiophores solitary, macronematous <strong>and</strong> micronematous,<br />
arising as lateral branches of plagiotropous hyphae or terminally<br />
from ascending hyphae, erect, straight or slightly flexuous,<br />
cylindrical, neither geniculate nor nodulose, without head-like<br />
swollen apices or intercalary swellings, unbranched, sometimes<br />
137
Schubert et al.<br />
Fig. 31. <strong>Cladosporium</strong> ramotenellum (CPC 12043). Conidiophores <strong>and</strong> conidia. Scale bar = 10 µm. K. Schubert del.<br />
branched, branches often only as short lateral prolongations,<br />
mainly formed below a septum, 14–110 × 2–4 µm, septate, not<br />
constricted at the septa, subhyaline to pale olivaceous or brown,<br />
smooth to minutely verruculose, walls unthickened, sometimes<br />
guttulate. Conidiogenous cells integrated, terminal, sometimes also<br />
intercalary, cylindrical, not geniculate, non-nodulose, 10–28(–50)<br />
µm long, proliferation sympodial, with few conidiogenous loci,<br />
mostly 1–3, loci sometimes situated on small lateral prolongations,<br />
protuberant, 0.5–1.5(–2) µm diam, thickened <strong>and</strong> somewhat<br />
darkened-refractive. Ramoconidia formed, cylindrical-oblong, up<br />
to 47 µm long, 2–4 µm wide, 0–1-septate, rarely up to 4-septate,<br />
subhyaline to very pale olivaceous, smooth or almost so, with a<br />
broadly truncate base, without any dome <strong>and</strong> raised rim, 2–3 µm<br />
wide, not thickened but somewhat refractive. Conidia numerous,<br />
polymorphous, catenate, in branched chains, straight, sometimes<br />
slightly curved, small terminal conidia numerous, globose,<br />
subglobose or ovoid, obovoid or limoniform, 2.5–7 × 2–4(–4.5) µm<br />
[av. ± SD, 5.1 (± 1.3) × 3.1 (± 0.6) µm], aseptate, without distal<br />
hilum or with a single apical scar, intercalary conidia ellipsoid to<br />
subcylindrical, 8–15 × 3–4(–4.5) µm [av. ± SD, 11.5 (± 2.4) × 3.6<br />
(± 0.5) µm], 0–1-septate; secondary ramoconidia subcylindrical<br />
to cylindrical-oblong, 17–35 × 3–4(–5) µm [av. ± SD, 22.5 (±<br />
5.6) × 3.7 (± 0.5) µm], 0–3-septate, not constricted at the septa,<br />
subhyaline to very pale olivaceous, minutely verruculose (granulate<br />
under SEM), walls unthickened or almost so, apex broadly rounded<br />
or slightly attenuated towards apex <strong>and</strong> base, sometimes guttulate,<br />
hila protuberant, conspicuous, 0.8–1.5(–2) µm diam, somewhat<br />
thickened <strong>and</strong> darkened-refractive; microcyclic conidiogenesis<br />
occurring.<br />
Cultural characteristics: Colonies on PDA reaching 46–49 mm diam<br />
after 14 d at 25 ºC, olivaceous to grey-olivaceous due to abundant<br />
sporulation, appearing zonate in forming concentric zones, margin<br />
entire edge to slightly undulate, white, glabrous, aerial mycelium<br />
absent or sparse, growth flat with a somewhat folded <strong>and</strong> wrinkled<br />
colony centre, without prominent exudates, sporulation profuse.<br />
Colonies on MEA reaching 48–49 mm diam after 14 d at 25 ºC,<br />
grey-olivaceous to olivaceous-grey, velvety, olivaceous-grey to<br />
138
<strong>Cladosporium</strong> herbarum species complex<br />
Fig. 32. <strong>Cladosporium</strong> ramotenellum (CPC 12043). A, C. Macronematous conidiophore. B. Conidial chain. D. Micronematous conidiophore. E. Ramoconidia <strong>and</strong> conidia. Scale<br />
bars = 10 µm.<br />
Fig. 33. <strong>Cladosporium</strong> ramotenellum (CPC 12043). A. Survey of colony development showing a large bulbous “foot cell” that gives rise to conidiophores, which can be branched.<br />
B. Details of conidiophores showing secondary ramoconidia <strong>and</strong> conidia. <strong>The</strong> inset shows scar formation on a conidiophore. C. Conidiophore <strong>and</strong> several conidia. D. Details<br />
of ornamentation on conidia. Note the wide, but relatively low ornamentation units. E. A micrograph illustrating the organisation within a conidiophore. Scale bars A–D = 5 µm,<br />
E = 10 µm.<br />
www.studiesinmycology.org<br />
139
Schubert et al.<br />
Fig. 34. <strong>Cladosporium</strong> sinuosum (CPC 11839). Conidiophores <strong>and</strong> conidia. Scale bar = 10 µm. K. Schubert del.<br />
iron-grey reverse, margin entire edge to undulate, radially furrowed,<br />
colourless, glabrous to feathery, aerial mycelium sparse, diffuse,<br />
growth flat with slightly elevated colony centre, distinctly wrinkled,<br />
prominent exudates not formed, abundantly sporulating. Colonies<br />
on OA attaining 40 mm diam after 14 d at 25 ºC, grey-olivaceous,<br />
margin entire edge, colourless or white, glabrous, aerial mycelium<br />
absent or sparse, growth flat, without exudates, sporulation<br />
profuse.<br />
Specimens examined: Slovenia, Ljubljana, isolated from an air conditioning system<br />
(bathroom), 2004, M. Butala, <strong>CBS</strong> 121627 = CPC 12047 = EXF-967; Sečovlje,<br />
isolated from hypersaline water from reverse ponds, salterns, 2005, P. Zalar, <strong>CBS</strong>-H<br />
19862, holotype, isotype HAL 2026 F, culture ex-type <strong>CBS</strong> 121628 = CPC 12043<br />
= EXF-454.<br />
Substrate <strong>and</strong> distribution: Hypersaline water, air; Slovenia.<br />
Notes: <strong>Cladosporium</strong> ramotenellum, which appears to be a<br />
saprobe in air <strong>and</strong> hypersaline water, morphologically resembles<br />
C. cladosporioides <strong>and</strong> C. tenellum K. Schub., Zalar, Crous &<br />
U. Braun, but is quite distinct from C. cladosporioides by having<br />
somewhat narrower conidiophores <strong>and</strong> conidia, 2–4(–5) µm<br />
wide, <strong>and</strong> 0–3-septate, always minutely verruculose conidia.<br />
<strong>Cladosporium</strong> tenellum, a newly introduced species (see below)<br />
isolated from hypersaline water <strong>and</strong> plant material, possesses<br />
conidiophores with numerous conidiogenous loci, usually crowded<br />
towards the apex forming sympodial clusters of pronounced scars,<br />
<strong>and</strong> shorter <strong>and</strong> somewhat wider, 0–1(–2)-septate conidia, 3–20(–<br />
28) × (2.5–)3–5(–6) µm. Besides these morphological differences,<br />
C. ramotenellum is faster growing in culture than C. tenellum.<br />
<strong>Cladosporium</strong> arthrinioides Thüm. & Beltr. <strong>and</strong> C. hypophyllum<br />
Fuckel are also close to C. ramotenellum, but C. arthrinioides,<br />
known from Italy on leaves of Bougainvillea spectabilis, deviates in<br />
having shorter <strong>and</strong> wider, 0–1(–2)-septate, mostly smooth conidia<br />
(2–18 × 2–6.5 µm) which become larger <strong>and</strong> more frequently<br />
septate with age (up to 32 µm long <strong>and</strong> with up to four septa);<br />
<strong>and</strong> C. hypophyllum occurring in Europe on leaves of Ulmus<br />
minor differs in having often mildly to distinctly geniculate-sinuous,<br />
sometimes subnodulose conidiophores <strong>and</strong> shorter <strong>and</strong> somewhat<br />
wider, 0–1(–3)-septate conidia, 4–17(–19) × 2–5 µm, becoming<br />
distinctly swollen, darker, longer <strong>and</strong> wider with age, 5–7 µm, with<br />
the septa often being constricted (Schubert 2005).<br />
140
<strong>Cladosporium</strong> herbarum species complex<br />
Fig. 35. <strong>Cladosporium</strong> sinuosum (CPC 11839). A–D. Conidiophores. E–F. Conidia. Scale bars = 10 µm.<br />
<strong>Cladosporium</strong> sinuosum K. Schub., C.F. Hill, Crous & U. Braun,<br />
sp. nov. MycoBank MB504578. Figs 34–35.<br />
Etymology: Refers to the usually distinctly sinuous conidiophores.<br />
Differt a Cladosporio herbaro conidiophoris distincte sinuosis, conidiis solitariis vel<br />
breve catenatis, catenis non ramosis, echinulatis.<br />
Mycelium sparingly branched, 1–7 µm wide, septate, not<br />
constricted at the septa, subhyaline to pale brown, smooth to<br />
minutely verruculose, walls unthickened or slightly thickened,<br />
sometimes with small swellings. Conidiophores arising laterally<br />
from plagiotropous hyphae or terminally from ascending hyphae,<br />
erect, more or less straight to flexuous, often once or several times<br />
slightly to distinctly geniculate-sinuous, sometimes even zigzag-like,<br />
nodulose with small to large lateral shoulders, shoulders somewhat<br />
distant from each other or in close succession giving them a knotty/<br />
gnarled appearance, unbranched or once branched, 25–260 × 5–7<br />
µm, shoulders up to 10 µm wide, pluriseptate, septa sometimes<br />
in short succession, not constricted at the septa, pale brown to<br />
medium brown, smooth to minutely verruculose, walls thickened,<br />
often distinctly two-layered, up to 1 µm thick. Conidiogenous<br />
cells integrated, terminal or intercalary, often slightly to distinctly<br />
geniculate-sinuous, nodulose with small to large laterally swollen<br />
shoulders, 8–30 µm long, proliferation sympodial, with a single<br />
or up to three conidiogenous loci, usually confined to lateral<br />
www.studiesinmycology.org<br />
shoulders, protuberant, often denticle-like or on the top of short<br />
cylindrical stalk-like prolongations, 1.2–2(–2.2) µm diam, mainly<br />
2 µm, somewhat thickened <strong>and</strong> darkened-refractive, dome often<br />
slightly higher than the surrounding rim. Conidia solitary or in short<br />
unbranched chains with up to three conidia, straight, obovoid, oval,<br />
broadly ellipsoid to subcylindrical or sometimes clavate (broader at<br />
the apex), 9–21 × (5–)6–8 µm [av. ± SD, 14.5 (± 2.5) × 6.6 (± 0.7)<br />
µm], 0–1-septate, not constricted at the septa, septum more or less<br />
median, pale greyish brown, densely echinulate, spines up to 1 µm<br />
long, walls thickened, apex mostly broadly rounded or sometimes<br />
attenuated, towards the base mostly distinctly attenuated forming a<br />
peg-like prolongation, up to 2 µm long, hila protuberant, 1.2–2 µm<br />
diam, mainly 2 µm, somewhat thickened <strong>and</strong> darkened-refractive;<br />
microcyclic conidiogenesis not observed.<br />
Cultural characteristics: Colonies on PDA attaining 20 mm<br />
diam after 14 d at 25 ºC, pale olivaceous-grey due to abundant<br />
aerial mycelium, olivaceous-grey towards margins, iron-grey<br />
to olivaceous-black reverse, margin regular, entire edge, aerial<br />
mycelium abundant, cottony, dense, high, growth regular, low<br />
convex, radially furrowed in the centre, growing deep into the agar,<br />
with age numerous small to large prominent exudates, sporulation<br />
sparse. Colonies on MEA attaining 16 mm diam after 14 d at 25 ºC,<br />
white to pale smoke-grey, fawn reverse, velvety, margin undulate,<br />
glabrous, aerial mycelium abundant, dense, high, fluffy, growth<br />
raised with elevated colony centre, laterally furrowed, without<br />
141
Schubert et al.<br />
Fig. 36. <strong>Cladosporium</strong> spinulosum (CPC 12040). A. Overview on agar surface with conidiophores arising from the surface. <strong>The</strong> spore clusters on the conidiophore are very<br />
compact. Note several simple, tubular conidiophore ends. <strong>The</strong> inset shows details of a conidium showing two pronounced hila <strong>and</strong> a unique, very distinct ornamentation on the<br />
cell wall. B. Conidiophore with globose or subsphaerical secondary ramoconidia <strong>and</strong> conidia. Note the newly forming cells <strong>and</strong> hila. C. Two conidiophores. D. Details of spores<br />
<strong>and</strong> spore formation. E. <strong>The</strong> end of a conidiophore <strong>and</strong> two scars. Scale bars: A = 20 µm, A (inset) = 1 µm, B, D–E = 5 µm, C = 10 µm.<br />
prominent exudates. Colonies on OA attaining 18 mm diam after 14<br />
d at 25 ºC, olivaceous, white to pale olivaceous-grey in the centre<br />
due to abundant aerial mycelium, olivaceous-grey reverse, margin<br />
white, entire edge, glabrous, aerial mycelium loose to dense, high,<br />
fluffy to felty, growth flat to low convex, regular, without prominent<br />
exudates, sporulating.<br />
Specimen examined: New Zeal<strong>and</strong>, Te Anau, isolated from leaves of Fuchsia<br />
excorticata (Onagraceae), 31 Jan. 2005, A. Blouin, Hill 1134A, <strong>CBS</strong>-H 19863,<br />
holotype, culture ex-type <strong>CBS</strong> 121629 = CPC 11839 = ICMP 15819.<br />
Substrate <strong>and</strong> distribution: On living leaves of Fuchsia excorticata;<br />
New Zeal<strong>and</strong>.<br />
Notes: This new species is well characterised by its slightly to<br />
distinctly geniculate-sinuous, often zigzag-like conidiophores <strong>and</strong><br />
its conidia formed solitary or rarely in short unbranched chains <strong>and</strong><br />
is therefore morphologically not comparable with any of the species<br />
described until now. Most <strong>Cladosporium</strong> species with conidia usually<br />
formed solitary or in short unbranched chains have previously<br />
been treated as species of the <strong>genus</strong> Heterosporium Klotzsch ex<br />
Cooke, now considered to be synonymous with <strong>Cladosporium</strong>. All<br />
of them, including the newly introduced C. arthropodii K. Schub.<br />
& C.F. Hill from New Zeal<strong>and</strong>, which also belongs to this species<br />
complex (Braun et al. 2006), possess very large <strong>and</strong> wide, often<br />
pluriseptate conidia quite distinct from those of C. sinuosum (David<br />
1997). <strong>Cladosporium</strong> alopecuri (Ellis & Everh.) U. Braun, known<br />
from the U.S.A. on Alopecurus geniculatus is also quite different by<br />
having larger <strong>and</strong> wider conidia, 20–40 × 7–13(–15) µm, <strong>and</strong> wider<br />
conidiogenous loci <strong>and</strong> conidial hila, 3.5–5 µm diam (Braun 2000).<br />
<strong>Cladosporium</strong> herbarum is superficially <strong>similar</strong> but the<br />
conidiophores of the latter species are sometimes only slightly<br />
geniculate-sinuous but never zigzag-like <strong>and</strong> the verruculose to<br />
verrucose conidia are frequently formed in unbranched or branched<br />
chains.<br />
<strong>Cladosporium</strong> spinulosum Zalar, de Hoog & Gunde-Cimerman,<br />
Studies in Mycology 58: 180. 2007 – this volume. Fig. 36.<br />
Note: This new species is described <strong>and</strong> illustrated in Zalar et al.<br />
(2007 – this volume).<br />
142
<strong>Cladosporium</strong> herbarum species complex<br />
Fig. 37. <strong>Cladosporium</strong> subinflatum (CPC 12041). Macro- <strong>and</strong> micronematous conidiophores <strong>and</strong> conidia. Scale bar = 10 µm. K. Schubert del.<br />
<strong>Cladosporium</strong> subinflatum K. Schub., Zalar, Crous & U. Braun,<br />
sp. nov. MycoBank MB504579. Figs 37–39.<br />
Etymology: Refers to its nodulose conidiophores.<br />
Differt a Cladosporio bruhnei conidiophoris cum nodulis angustioribus, 3–6.5 µm<br />
latis, conidiis brevioribus, 4–17(–22) µm longis, spinulosis, cum spinulis ad 0.8 µm<br />
longis; et a Cladosporio spinuloso conidiophoris nodulosis, conidiis spinulosis, cum<br />
spinulis brevioribus, ad 0.8 longis, locis conidiogenis et hilis latioribus, (0.5–)1–2<br />
µm latis.<br />
Mycelium unbranched or occasionally branched, 1.5–3 µm wide,<br />
later more frequently branched <strong>and</strong> wider, up to 7 µm wide,<br />
septate, not constricted at the septa, hyaline or subhyaline, almost<br />
smooth to somewhat verruculose or irregularly rough-walled, walls<br />
unthickened. Conidiophores mainly macronematous, sometimes<br />
also micronematous, arising terminally from ascending hyphae<br />
or laterally from plagiotropous hyphae, erect or subdecumbent,<br />
straight or flexuous, sometimes bent, cylindrical, nodulose, usually<br />
with small head-like swellings, sometimes swellings also on a<br />
lower level or intercalary, occasionally geniculate, unbranched,<br />
occasionally branched, (5–)10–270 × (1.5–)2.5–4.5(–5.5) µm,<br />
www.studiesinmycology.org<br />
swellings 3–6.5 µm wide, aseptate or with few septa, not constricted<br />
at the septa, pale brown, pale olivaceous-brown or somewhat<br />
reddish brown, smooth, usually verruculose or irregularly roughwalled<br />
<strong>and</strong> paler, subhyaline towards the base, walls thickened,<br />
sometimes appearing even two-layered, up to 1 µm thick.<br />
Conidiogenous cells integrated, usually terminal or conidiophores<br />
reduced to conidiogenous cells, cylindrical, nodulose, usually with<br />
small head-like swellings with loci confined to swellings, sometimes<br />
geniculate, 5–42 µm long, proliferation sympodial, with several loci,<br />
up to four situated at nodules or on lateral swellings, protuberant,<br />
conspicuous, denticulate, (0.8–)1–2 µm diam, thickened <strong>and</strong><br />
darkened-refractive. Conidia catenate, in branched chains, more<br />
or less straight, numerous globose <strong>and</strong> subglobose conidia, ovoid,<br />
obovoid, broadly ellipsoid to cylindrical, 4–17(–22) × (2.5–)3.5–<br />
5.5(–7) µm [av. ± SD, 11.7 (± 4.6) × 4.5 (± 0.8) µm], 0–1(–2)-<br />
septate, not constricted at septa, pale brown or pale olivaceousbrown,<br />
ornamentation variable, mainly densely verruculose to<br />
echinulate (loosely muricate under SEM), spines up to 0.8 µm<br />
high, sometimes irregularly verrucose with few scattered tubercles<br />
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Schubert et al.<br />
Fig. 38. <strong>Cladosporium</strong> subinflatum (CPC 12041). A–C. Macronematous conidiophores. D–E. Conidia. Scale bar = 10 µm.<br />
Fig. 39. <strong>Cladosporium</strong> subinflatum<br />
(CPC 12041). A–G. Images of an 11-<br />
d-old culture on SNA. A. Overview of<br />
colony with clusters of conidia <strong>and</strong><br />
aerial hyphae. Many of the hyphae<br />
have a collapsed appearance. B.<br />
Detail of colony with conidiophores,<br />
conidia <strong>and</strong> aerial hyphae that<br />
are partly collapsed. C. Detail of a<br />
conidiophore end <strong>and</strong> a secondary<br />
ramoconidium. Note the scars at the<br />
end of the conidiophore. D. Details<br />
of conidia <strong>and</strong> ornamentation.<br />
<strong>The</strong> ornamentation consists out of<br />
markedly defined units, which have<br />
a relatively large distance from each<br />
other. Note the hilum on the right<br />
conidium. E. Conidiophore with<br />
large scars <strong>and</strong> conidia. F. Different<br />
blastoconidia with very early stages<br />
of new spore formation in the middle<br />
of the picture. G. Pattern of spore<br />
development. Scale bars: A = 20<br />
µm, B, E–G = 5 µm, C = 10 µm, D<br />
= 2 µm.<br />
144
<strong>Cladosporium</strong> herbarum species complex<br />
or irregularly echinulate, walls unthickened or slightly thickened,<br />
apex rounded or slightly attenuated towards apex <strong>and</strong> base, hila<br />
conspicuous, protuberant, denticulate, 0.5–2 µm diam, thickened<br />
<strong>and</strong> darkened-refractive; microcyclic conidiogenesis observed.<br />
Cultural characteristics: Colonies on PDA attaining 29 mm diam<br />
after 14 d at 25 ºC, olivaceous-black to olivaceous-grey towards<br />
margin, margin regular, entire edge, narrow, colourless to white,<br />
glabrous to feathery, aerial mycelium formed, fluffy, mainly near<br />
margins, growth flat, somewhat folded in the colony centre, deep<br />
into the agar, few prominent exudates formed with age, sporulation<br />
profuse. Colonies on MEA attaining 25 mm diam after 14 d at 25<br />
ºC, olivaceous-grey to olivaceous due to abundant sporulation<br />
in the colony centre, pale greenish grey towards margin, irongrey<br />
reverse, velvety to powdery, margin crenate, narrow, white,<br />
glabrous, radially furrowed, aerial mycelium diffuse, growth convex<br />
with papillate surface, wrinkled colony centre, without prominent<br />
exudates, sporulation profuse. Colonies on OA attaining 26 mm<br />
diam after 14 d at 25 ºC, olivaceous, iron-grey to greenish black<br />
reverse, growth flat, deep into the agar, with a single exudate,<br />
abundantly sporulating.<br />
Specimen examined: Slovenia, Sečovlje, isolated from hypersaline water from<br />
crystallization ponds, salterns, 2005, S. Sonjak, <strong>CBS</strong>-H 19864, holotype, isotype<br />
HAL 2027 F, culture ex-type <strong>CBS</strong> 121630 = CPC 12041 = EXF-343.<br />
Substrate <strong>and</strong> distribution: Hypersaline water; Slovenia.<br />
Notes: <strong>Cladosporium</strong> subinflatum, an additional saprobic species<br />
isolated from hypersaline water, was at first identified as C.<br />
spinulosum, but proved to be both morphologically as well as<br />
phylogenetically distinct from the latter species in having somewhat<br />
wider [(1.5–)2.5–4.5(–5.5) µm], nodulose macronematous<br />
conidiophores with conidiogenous loci confined to swellings, wider<br />
conidiogenous loci <strong>and</strong> hila, (0.8–)1–2 µm, <strong>and</strong> spiny conidia<br />
with shorter spines than in C. spinulosum (up to 0.8 µm versus<br />
0.5–1.3 µm long) (Zalar et al. 2007). With its narrow, nodulose<br />
macronematous conidiophores <strong>and</strong> catenate conidia, C. bruhnei<br />
is morphologically also <strong>similar</strong> but differs by having conidiophores<br />
with wider swellings, (4–)5–8 µm, <strong>and</strong> longer conidia 4–24(–31)<br />
µm, rarely up to 40 µm long which are minutely verruculose to<br />
verrucose but not spiny.<br />
Fig. 40. <strong>Cladosporium</strong> subtilissimum (<strong>CBS</strong> 113754). Macro- <strong>and</strong> micronematous conidiophores <strong>and</strong> conidia. Scale bar = 10 µm. K. Schubert del.<br />
www.studiesinmycology.org<br />
145
Schubert et al.<br />
Fig. 41. <strong>Cladosporium</strong> subtilissimum (<strong>CBS</strong> 113754). A–C. Macronematous conidiophores. D. Conidial chain. E. Micronematous conidiophore. F–G. Conidia. Scale bars = 10<br />
µm.<br />
<strong>Cladosporium</strong> subtilissimum K. Schub., Dugan, Crous & U.<br />
Braun, sp. nov. MycoBank MB504580. Figs 40–42.<br />
Etymology: Refers to its narrow conidiophores <strong>and</strong> conidia.<br />
Differt a Cladosporio cladosporioide conidiophoris et conidiis semper asperulatis ad<br />
verruculosis, conidiis 0–1(–2)-septatis.<br />
Mycelium unbranched or sparingly branched, 1–5 µm wide, septate,<br />
without swellings <strong>and</strong> constrictions, hyaline to subhyaline or pale<br />
brown, smooth to minutely verruculose, walls unthickened or almost<br />
so, protoplasm somewhat guttulate or granular. Conidiophores<br />
macronematous <strong>and</strong> micronematous, arising laterally from<br />
plagiotropous hyphae or terminally from ascending hyphae, erect,<br />
straight to slightly flexuous, filiform to cylindrical-oblong, nonnodulose,<br />
sometimes geniculate towards the apex, unbranched or<br />
once branched, branches short to somewhat longer, usually formed<br />
below a septum, sometimes only short, denticle-like or conical,<br />
25–140 × 2–4 µm, 0–4-septate, not constricted at the septa,<br />
subhyaline to pale brown, almost smooth, minutely verruculose<br />
to verruculose, sometimes irregularly rough-walled in the lower<br />
part, walls unthickened or slightly thickened, protoplasm guttulate<br />
or somewhat granular. Conidiogenous cells integrated, terminal<br />
or pleurogenous, sometimes also intercalary, filiform to narrowly<br />
cylindrical, non-nodulose, sometimes geniculate, 14–57 µm long,<br />
with usually sympodial clusters of pronounced conidiogenous loci<br />
at the apex or on a lower level, denticle-like or situated on short<br />
lateral prolongations, up to five loci, intercalary conidiogenous<br />
cells usually with a short denticle-like lateral outgrowth below a<br />
septum, protuberant, denticulate, somewhat truncate, 1.2–2 µm<br />
diam, thickened <strong>and</strong> darkened-refractive. Ramoconidia sometimes<br />
occurring, conidiogenous cells seceding at one of the upper septa<br />
of the conidiophore <strong>and</strong> behaving like conidia, filiform or cylindrical,<br />
20–40(–55) µm long, 1.5–4 µm wide, 0–1-septate, concolorous<br />
with conidiophores, not attenuated towards apex <strong>and</strong> base, base<br />
broadly truncate, non-cladosporioid, without any dome <strong>and</strong> raised<br />
rim, 2–3.5 µm wide, neither thickened nor darkened, sometimes<br />
slightly refractive. Conidia catenate, in branched chains, up to 12<br />
or even more in a chain, straight, small terminal conidia numerous,<br />
subglobose, narrowly obovoid, limoniform or fusiform, 4–9 × 2–3.5<br />
µm [av. ± SD, 6.4 (± 1.5) × 2.8 (± 0.4) µm], with up to three distal<br />
scars, aseptate, hila (0.5–)0.8–1 µm diam, intercalary conidia<br />
narrowly ellipsoid, fusiform to subcylindrical, 9–18 × 3–4(–6) µm<br />
[av. ± SD, 13.0 (± 2.5) × 3.8 (± 0.3) µm], 0(–1)-septate, hila 1–1.2(–<br />
1.8) µm diam, with up to four distal scars, secondary ramoconidia<br />
ellipsoid, fusiform or subcylindrical, (13–)17–32(–37) × 3–5(–6) µm<br />
[av. ± SD, 21.4 (± 4.4) × 4.1 (± 0.5) µm], 0–1(–2)-septate, septum<br />
median or somewhat in the lower half, usually not constricted at<br />
the septa, with up to six distal hila crowded at the apex, hila (1.2–)<br />
1.5–2(–2.5) µm diam, apex often somewhat laterally enlarged<br />
or prolonged with hila crowded there, very pale or pale brown or<br />
olivaceous-brown, minutely verruculose to verruculose (granulate<br />
under SEM), walls unthickened or only slightly thickened, often<br />
slightly attenuated towards apex <strong>and</strong> base, protoplasm often<br />
guttulate or granular, hila protuberant, denticulate, (0.5–)0.8–2(–<br />
2.2) µm diam, thickened <strong>and</strong> darkened-refractive; microcyclic<br />
conidiogenesis occasionally observed.<br />
Cultural characteristics: Colonies on PDA attaining 24 mm diam<br />
after 14 d at 25 °C, grey-olivaceous to olivaceous, olivaceousgrey,<br />
iron-grey or olivaceous-black reverse, velvety, margin<br />
regular, entire edge, white or pale greenish olivaceous, glabrous<br />
to feathery, aerial mycelium sparse, only few areas with abundant<br />
146
<strong>Cladosporium</strong> herbarum species complex<br />
Fig. 42. <strong>Cladosporium</strong> subtilissimum (<strong>CBS</strong> 113754). A. Overview on the organisation of spore formation. <strong>The</strong> micrograph shows a large basal secondary ramoconidium which<br />
has chains of secondary ramoconidia, intercalary <strong>and</strong> small terminal conidia. <strong>The</strong> conidia are formed in rows of often three cells. Note the size difference in the different cells.<br />
B. Conidiophore showing very pronounced scars that almost appear as branches. C. Detail of (A), illustrating the scar formation between the cells. D. Conidia during different<br />
stages of formation. E. Details of pronounced hila, <strong>and</strong> prominent ornamentation on secondary ramoconidia with the central dome-formed area. F. Different conidia <strong>and</strong> hila.<br />
Scale bars: A = 10 µm, B–D, F = 5 µm, E = 2 µm.<br />
mycelium, diffuse, growth regular, flat or with a raised <strong>and</strong> wrinkled<br />
colony centre, radially furrowed, effuse, usually without prominent<br />
exudates, with age several exudates formed, sporulation profuse,<br />
colonies consisting of two kinds of conidiophores, short <strong>and</strong> a few<br />
longer ones. Colonies on MEA reaching 25 mm diam after 14 d<br />
at 25 °C, greenish olivaceous to grey-olivaceous in the centre,<br />
olivaceous-grey to iron-grey reverse, velvety, margin entire edge,<br />
crenate or umbonate, narrow, pale greenish olivaceous, sometimes<br />
radially furrowed, aerial mycelium absent or sparse, growth low<br />
convex with distinctly wrinkled colony centre, without prominent<br />
exudates, abundantly sporulating. Colonies on OA attaining 25 mm<br />
diam after 14 d at 25 °C, dark grey-olivaceous to olivaceous due to<br />
profuse sporulation, iron-grey reverse, sometimes releasing some<br />
olivaceous-buff pigments into the agar, velvety, margin regular,<br />
entire edge or crenate, narrow, colourless or white, glabrous or<br />
feathery, aerial mycelium sparse, growth flat with slightly raised<br />
colony centre, prominent exudates lacking, sporulation profuse.<br />
www.studiesinmycology.org<br />
Specimens examined: Slovenia, Sečovlje, isolated from hypersaline water from<br />
salterns (reserve pond), 2005, P. Zalar, CPC 12044 = EXF-462. U.S.A., isolated<br />
from bing cherry fruits, F. Dugan, <strong>CBS</strong> 113753; isolated from a grape berry, F.<br />
Dugan, wf 99-2-9 sci 1, <strong>CBS</strong>-H 19865, holotype, isotype HAL 2028 F, culture extype<br />
<strong>CBS</strong> 113754.<br />
Excluded strains within the subtilissimum complex: Argentina,<br />
isolated from Pinus ponderosa (Pinaceae), 2005, A. Greslebin,<br />
CPC 12484, CPC 12485. U.S.A., isolated from grape berry, F.<br />
Dugan, <strong>CBS</strong> 113741, <strong>CBS</strong> 113742; isolated from grape bud, F.<br />
Dugan, <strong>CBS</strong> 113744.<br />
Substrate <strong>and</strong> distribution: Plant material <strong>and</strong> hypersaline water;<br />
Slovenia, U.S.A.<br />
Notes: <strong>Cladosporium</strong> cladosporioides is morphologically comparable<br />
with the new species but deviates in having usually smooth<br />
conidiophores <strong>and</strong> conidia, with the conidia being mainly aseptate.<br />
C. subtilissimum is represented by three isolates of different origins<br />
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Schubert et al.<br />
Fig. 43. <strong>Cladosporium</strong> tenellum (CPC 12053). Macro- <strong>and</strong> micronematous conidiophores <strong>and</strong> conidia. Scale bar = 10 µm. K. Schubert del.<br />
Fig. 44. <strong>Cladosporium</strong> tenellum (CPC 12053). A–C, E. Macronematous conidiophore. D. Micronematous conidiophore. F. Ramoconidium <strong>and</strong> conidia. Scale bars = 10 µm.<br />
148
<strong>Cladosporium</strong> herbarum species complex<br />
Differt a Cladosporio cladosporioide conidiophoris et conidiis semper asperulatis,<br />
locis conidiogenis apicalibus, numerosis, hilis quoque numerosis, conidiophoris<br />
angustioribus, (1–)1.5–3.5(–4) µm latis; et a Cladosporio subtilissimo loci<br />
conidiogenis et hilis apicalibus, numerosis, angustioribus, saepe 1–1.5 µm latis,<br />
conidiis minutis numerosis, saepe globosis.<br />
Fig. 45. <strong>Cladosporium</strong> tenellum (CPC 12053). A. A bird’s eye view of a colony of<br />
C. tenellum with its very characteristic bundles of aerial hyphae. Numerous conidia<br />
are visible, formed on simple conidiophores. B. Hyphae that run on the agar surface<br />
give rise to conidiophores <strong>and</strong> numerous conidia, that are relatively rounded. C.<br />
Conidiophore ends are rather simple <strong>and</strong> have large scars. D. Hila on a secondary<br />
ramoconidium with non-ornamented area. E. Detail of the prominent ornamentation<br />
on a secondary ramoconidium. Scale bars: A = 20 µm, B = 10 µm, C, E = 2 µm,<br />
D = 5 µm.<br />
<strong>and</strong> substrates. Besides these strains, several additional isolates<br />
listed under excluded strains are morphologically indistinguishable<br />
from C. subtilissimum in culture, but genetically different, clustering<br />
in various subclades. <strong>The</strong>y are indicated as <strong>Cladosporium</strong> sp. in<br />
the tree (Fig. 3).<br />
<strong>Cladosporium</strong> tenellum K. Schub., Zalar, Crous & U. Braun, sp.<br />
nov. MycoBank MB504581. Figs 43–45.<br />
Etymology: Refers to its narrow conidiophores <strong>and</strong> conidia.<br />
www.studiesinmycology.org<br />
Mycelium sparingly branched, 1–3 µm wide, septate, septa often<br />
not very conspicuous, not constricted at the septa, sometimes<br />
slightly swollen, subhyaline, smooth, walls unthickened.<br />
Conidiophores macronematous <strong>and</strong> micronematous, solitary,<br />
arising terminally or laterally from plagiotropous or ascending<br />
hyphae, erect or subdecumbent, almost straight to more or less<br />
flexuous, cylindrical, sometimes geniculate towards the apex, but<br />
not nodulose, sometimes with short lateral prolongations at the<br />
apex, unbranched to once or twice branched (angle usually 30–45°<br />
degree, sometimes up to 90°), branches usually below a septum,<br />
6–200 × (1–)2–4(–5) µm, septate, septa not very conspicuous,<br />
not constricted at the septa, subhyaline to pale brown, almost<br />
smooth to usually asperulate, walls unthickened or almost so.<br />
Conidiogenous cells integrated, terminal or intercalary, sometimes<br />
conidiophores reduced to conidiogenous cells, cylindrical,<br />
sometimes geniculate, non-nodulose, 6–40 µm long, proliferation<br />
sympodial, with several conidiogenous loci often crowded at the<br />
apex <strong>and</strong> sometimes also at a lower level, situated on small lateral<br />
shoulders, unilateral swellings or prolongations, with up to 6(–10)<br />
denticulate loci, forming sympodial clusters of pronounced scars,<br />
intercalar conidiogenous cells with short or somewhat long lateral<br />
outgrowths, short denticle-like or long branches with several scars<br />
at the apex, usually below a septum, loci protuberant, 1–1.5(–2) µm<br />
diam, thickened <strong>and</strong> darkened-refractive. Ramoconidia sometimes<br />
occurring, cylindrical, up to 32 µm long, 2.5–4 µm wide, with a<br />
broadly truncate, unthickened base, about 2 µm wide. Conidia<br />
catenate, formed in branched chains, straight, small terminal<br />
conidia globose, subglobose, ovoid, oval, 3–6 × 2.5–3.5 µm [av.<br />
± SD, 4.5 (± 1.3) × 2.8 (± 0.4) µm], aseptate, asperulate, with 0–2<br />
distal hila, intercalary conidia <strong>and</strong> secondary ramoconidia ellipsoidovoid,<br />
ellipsoid to subcylindrical, 3.5–20(–28) × (2.5–)3–5(–6) µm<br />
[av. ± SD, 12.4 (± 5.4) × 4.1 (± 0.7) µm], 0–1-septate, rarely with<br />
up to three septa, sometimes slightly constricted at the septa,<br />
subhyaline, pale brown to medium olivaceous-brown, asperulate<br />
or verruculose (muricate, granulate or colliculate under SEM),<br />
walls unthickened or slightly thickened, apex rounded or slightly<br />
to distinctly attenuated towards apex <strong>and</strong> base, often forming<br />
several apical hila, up to 7(–9), crowded, situated on small lateral<br />
outgrowths giving them a somewhat irregular appearance, hila<br />
protuberant, 0.5–1.5 µm diam, thickened <strong>and</strong> darkened-refractive;<br />
microcyclic conidiogenesis sometimes occurring.<br />
Cultural characteristics: Colonies on PDA reaching 27–34 mm diam<br />
after 14 d at 25 ºC, smoke-grey, grey-olivaceous to olivaceous-grey,<br />
olivaceous-grey to iron-grey reverse, velvety to powdery, margin<br />
regular, entire edge, narrow, colourless to white, aerial mycelium<br />
absent or sparingly formed, felty, whitish, growth regular, flat,<br />
radially furrowed, with folded <strong>and</strong> elevated colony centre, deep into<br />
the agar, with age forming few to numerous prominent exudates,<br />
sporulation profuse, few high conidiophores formed. Colonies on<br />
MEA reaching 25–44 mm diam after 14 d at 25 ºC, olivaceous-grey<br />
to olivaceous- or iron-grey due to abundant sporulation in the colony<br />
centre, velvety, margin regular, entire edge, narrow, colourless,<br />
white to pale olivaceous-grey, aerial mycelium loose, diffuse,<br />
growth convex with papillate surface, radially furrowed, wrinkled,<br />
without prominent exudates, sporulating. Colonies on OA reaching<br />
23–32 mm diam after 14 d at 25 ºC, grey-olivaceous, olivaceous-<br />
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Schubert et al.<br />
Fig. 46. <strong>Cladosporium</strong> variabile (CPC 12751). Macro- <strong>and</strong> micronematous conidiophores <strong>and</strong> conidia. Scale bar = 10 µm. K. Schubert del.<br />
grey to olivaceous due to abundant sporulation in the colony centre,<br />
olivaceous- or iron-grey reverse, velvety, margin regular, entire<br />
edge, narrow, colourless or white, aerial mycelium sparse, diffuse,<br />
floccose, growth flat to low convex, radially furrowed, wrinkled,<br />
without prominent exudates, sporulation profuse.<br />
Specimens examined: Israel, Eilat, isolated from hypersaline water from salterns,<br />
2004, N. Gunde-Cimerman, <strong>CBS</strong> 121633 = CPC 12051 = EXF-1083; Ein Bokek,<br />
isolated from hypersaline water of the Dead Sea, 2004, M. Ota, <strong>CBS</strong>-H 19866,<br />
holotype, isotype HAL 2029 F, culture ex-type <strong>CBS</strong> 121634 = CPC 12053 = EXF-<br />
1735. U.S.A., Seattle, University of Washington campus, isolated from Phyllactinia<br />
sp. (Erysiphaceae) on leaves of Corylus sp. (Corylaceae), 16 Sep. 2004, D. Glawe,<br />
CPC 11813.<br />
Substrates <strong>and</strong> distribution: Hypersaline water <strong>and</strong> plant material;<br />
Israel, U.S.A.<br />
Notes: <strong>Cladosporium</strong> subtilissimum <strong>and</strong> C. cladosporioides are<br />
morphologically comparable with the new species C. tenellum, but<br />
C. cladosporioides deviates in having usually smooth conidiophores<br />
<strong>and</strong> conidia with only few conidiogenous loci <strong>and</strong> conidial hila crowded<br />
at the apex <strong>and</strong> somewhat wider conidiophores, 3–5(–6) µm; <strong>and</strong><br />
in C. subtilissimum the small terminal conidia are not globose but<br />
rather narrowly obovoid to limoniform, the conidiogenous loci <strong>and</strong><br />
conidial hila are somewhat wider, (0.5–)0.8–2(–2.2) µm, <strong>and</strong> at the<br />
apices of conidiophores <strong>and</strong> conidia only few scars are formed.<br />
<strong>Cladosporium</strong> ramotenellum, which morphologically also<br />
resembles C. tenellum, possesses longer <strong>and</strong> narrower, 0–3-septate<br />
conidia, 2.5–35 × 2–4(–5) µm, but forms only few conidiogenous<br />
loci <strong>and</strong> conidial hila at the apices of conidiophores <strong>and</strong> conidia.<br />
<strong>Cladosporium</strong> variabile (Cooke) G.A. de Vries, Contr. Knowl.<br />
Genus <strong>Cladosporium</strong>: 85. 1952. Figs 46–48.<br />
Basionym: Heterosporium variabile Cooke, Grevillea 5(35): 123.<br />
1877.<br />
≡ Helminthosporium variabile Cooke, Fungi Brit. Exs. Ser. 1, No. 360.<br />
1870, nom. inval.<br />
= <strong>Cladosporium</strong> subnodosum Cooke, Grevillea 17(83): 67. 1889.<br />
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<strong>Cladosporium</strong> herbarum species complex<br />
Fig. 47. <strong>Cladosporium</strong> variabile <strong>and</strong> its teleomorph Davidiella variabile (CPC 12751). A–C. Macronematous conidiophores. D, F. Micronematous conidiophores. E, G–H.<br />
Conidia. I. Twisted aerial mycelium. J. Ascomata formed on nettle stem in culture. K. Surface view of ascomal wall of textura epidermoidea. L–M. Asci. N–P. Ascospores. Q.<br />
Ascus with a sheath. Scale bars A, D, G–J, K–N = 10 µm, J = 250 µm.<br />
www.studiesinmycology.org<br />
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Schubert et al.<br />
Fig. 48. <strong>Cladosporium</strong> variabile (CPC 12753). A. Survey of hyphae that grow on the agar surface. Some of the fungal cells have a swollen appearance <strong>and</strong> could develop into<br />
a “foot cell” that gives rise to a conidiophore. B. A number of aerial hyphae obstruct the swollen, large structures on the agar surface, which give rise to conidiophores. Some<br />
of them appear ornamented. C. A series of conidia formed on a conidiophore (bottom of the micrograph). D. Detail of the ornamented conidia. <strong>The</strong> ornamentations are isolated<br />
<strong>and</strong> dispersed. Note also the ornamentation-free scar zone <strong>and</strong> the hilum of the left cell. E. Two conidia behind an aerial hypha. F. Two conidiophores forming secondary<br />
ramoconidia. Note the bulbous shape of the spore-forming apparatus. This micrograph is from an uncoated sample. Scale bars: A–C, F = 10 µm, D = 2 µm, E = 5 µm.<br />
Teleomorph: Davidiella variabile Crous, K. Schub. & U. Braun, sp.<br />
nov. MycoBank MB504583.<br />
Davidiellae tassianae similis, sed ascosporis maioribus, (22–)26–30(–35) × (7–)7.5–<br />
8(–9) µm, et ascis latioribus, plus quam 18 µm.<br />
Ascomata pseudothecial, black, superficial, situated on a small<br />
stroma, globose, up to 250 µm diam, with 1–3 ostiolate necks;<br />
ostioles periphysate, with apical periphysoids present; wall<br />
consisting of 3–6 layers of dark brown textura angularis, textura<br />
epidermoidea in surface view. Asci fasciculate, bitunicate,<br />
subsessile, obovoid to broadly ellipsoid, straight to slightly curved,<br />
8-spored, 70–95 × 18–28 µm; with pseudoparenchymatal cells<br />
of the hamathecium persistent. Ascospores tri- to multiseriate,<br />
overlapping, hyaline, with irregular lumina, thick-walled, straight<br />
to slightly curved, fusoid-ellipsoidal with obtuse ends, widest near<br />
the middle of the apical cell, medianly 1-septate, not to slightly<br />
constricted at the septum, at times developing a second septum<br />
in each cell, several ascospores with persistent, irregular mucoid<br />
sheath, (22–)26–30(–35) × (7–)7.5–8(–9) µm.<br />
Mycelium immersed <strong>and</strong> superficial, irregularly branched, aerial<br />
mycelium twisted <strong>and</strong> spirally coiled, 1–3 µm wide, septate,<br />
sometimes with swellings or small lateral outgrowths, hyaline<br />
to subhyaline, smooth, walls unthickened, hyphae which give<br />
rise to conidiophores somewhat wider, 3–4.5 µm, subhyaline to<br />
pale brown, almost smooth to minutely verruculose, sometimes<br />
enveloped by a polysaccharide-like cover. Conidiophores usually<br />
macronematous, but also micronematous, arising terminally<br />
from ascending hyphae or laterally from plagiotropous hyphae.<br />
Macronematous conidiophores erect, more or less straight to<br />
flexuous, often distinctly geniculate-sinuous forming lateral<br />
shoulders or unilateral swellings, sometimes zigzag-like or<br />
somewhat coralloid, nodulose, swellings at first terminal, then<br />
becoming lateral due to sympodial proliferation, often as distinct<br />
lateral shoulders, unbranched, sometimes once branched, 6–<br />
180 × (2.5–)3–6 µm, swellings (3–)6–11 µm wide, septate, not<br />
constricted at the septa, pale to medium olivaceous-brown or<br />
brown, usually verruculose, walls somewhat thickened, about 1 µm<br />
thick, sometimes appearing to be two-layered. Conidiogenous cells<br />
integrated, terminal <strong>and</strong> intercalary, cylindrical, nodulose to nodose,<br />
152
<strong>Cladosporium</strong> herbarum species complex<br />
with a single or two swellings per cell, swellings apart from each<br />
other or formed in short succession, loci confined to swellings, up to<br />
six per node, protuberant, 1–2 µm diam, thickened <strong>and</strong> darkenedrefractive.<br />
Micronematous conidiophores erect, straight to slightly<br />
flexuous, unbranched, usually without swellings, filiform to narrowly<br />
cylindrical, sometimes only as short lateral outgrowths of hyphae,<br />
often almost indistinguishable from hyphae, up to 50 µm long,<br />
1.5–2.5(–3) µm wide, longer ones pluriseptate, septa appear to be<br />
somewhat more darkened, with very short cells, 4–12 µm long,<br />
subhyaline to pale brown, smooth, walls unthickened or almost so.<br />
Conidiogenous cells integrated, usually terminal, rarely intercalary,<br />
cylindrical, non-nodulose, with a single, two or few conidiogenous<br />
loci at the distal end, protuberant, up to 2 µm diam, thickened<br />
<strong>and</strong> darkened-refractive. Conidia catenate, in branched chains,<br />
straight, subglobose, obovoid, oval, broadly ellipsoid to cylindrical,<br />
sometimes clavate, 4–26(–30) × (3.5–)5–9(–10) µm [av. ± SD,<br />
16.8 (± 6.9) × 6.5 (± 1.4) µm], 0–3-septate, usually not constricted<br />
at the septa, septa becoming sinuous with age, often appearing to<br />
be darkened, pale to medium or even dark brown or olivaceousbrown,<br />
verruculose to densely verrucose or echinulate (granulate<br />
under SEM), walls slightly to distinctly thickened in larger conidia,<br />
apex <strong>and</strong> base broadly rounded, sometimes broadly truncate or<br />
somewhat attenuated, apex <strong>and</strong> base often appear to be darkened<br />
or at least refractive, hila protuberant to somewhat sessile (within<br />
the outer wall ornamentation), (0.8–)1–2 µm diam, thickened <strong>and</strong><br />
darkened-refractive; microcyclic conidiogenesis occurring.<br />
Cultural characteristics: Colonies on PDA attaining 29 mm diam<br />
after 14 d at 25 ºC, olivaceous to olivaceous-grey or iron-grey, irongrey<br />
or olivaceous-grey reverse, velvety to powdery, margin regular,<br />
entire edge to fimbriate, almost colourless, aerial mycelium whitish<br />
turning olivaceous-grey, sometimes reddish, greyish rose, woollyfelty,<br />
growth flat with elevated colony centre, somewhat folded or<br />
radially furrowed, with age forming several very small but prominent<br />
exudates, sporulation profuse. Colonies on MEA attaining 27 mm<br />
diam after 14 d at 25 ºC, olivaceous-grey to iron-grey, white to pale<br />
olivaceous-grey in the centre due to abundant aerial mycelium,<br />
velvety, margin very narrow, colourless, more or less entire edge,<br />
radially furrowed, aerial mycelium fluffy to floccose, dense, growth<br />
low convex with wrinkled <strong>and</strong> folded centre, without exudates,<br />
sporulation profuse. Colonies on OA attaining 25 mm diam after<br />
14 d at 25 ºC, iron-grey or olivaceous, margin regular, entire edge,<br />
narrow, white, glabrous, aerial mycelium whitish, at first mainly in<br />
the colony centre, high, dense, floccose, growth flat, abundantly<br />
sporulating, no exudates.<br />
Specimens examined: Great Britain, Wales, Montgomeryshire, Welshpool, Forden<br />
Vicarage, on Spinacia oleracea (Chenopodiaceae), J.E. Vize, Cooke, Fungi Brit. Exs.<br />
Ser. I, No. 360, K, holotype. U.S.A., Washington, isolated from Spinacia oleracea, 1<br />
Jan. 2003, L. DuToit, <strong>CBS</strong>-H 19867, epitype designated here of C. variabile <strong>and</strong> D.<br />
variabile, cultures ex-epitype <strong>CBS</strong> 121635 = CPC 12753, CPC 12751.<br />
Substrate <strong>and</strong> distribution: Leaf-spotting fungus on Spinacia<br />
oleracea; Asia (China, India, Iraq, Pakistan), Europe (Austria,<br />
Belgium, Cyprus, Denmark, France, Germany, Great Britain,<br />
Hungary, Italy, Montenegro, Netherl<strong>and</strong>s, Norway, Romania, Spain,<br />
Turkey), North America (U.S.A.).<br />
Literature: de Vries (1952: 85–88), Ellis (1971: 315), Ellis & Ellis<br />
(1985: 429), David (1995b; 1997: 94, 96–98), Ho et al. (1999:<br />
144).<br />
Notes: In vivo the conidia are usually longer, somewhat wider <strong>and</strong><br />
more frequently septate, (6.5–)10–45(–55) × (4.5–)6–14(–17) µm,<br />
www.studiesinmycology.org<br />
0–4(–5)-septate (Schubert 2005). In culture the dimensions tend to<br />
be smaller, which was already mentioned by de Vries (1952).<br />
This leaf-spotting fungus superficially resembles C.<br />
macrocarpum, but besides its pathogenicity to Spinacia, C.<br />
variabile differs from the latter species in having distinctly larger<br />
<strong>and</strong> more frequently septate conidia on the natural host, forming<br />
twisted <strong>and</strong> spirally coiled aerial mycelium in culture <strong>and</strong> in having<br />
lower growth rates in culture (29 mm after 14 d on PDA versus<br />
38 mm on average in C. macrocarpum). Furthermore, the conidial<br />
septa of C. variabile are often distinctly darkened, become sinuous<br />
with age <strong>and</strong> the apex <strong>and</strong> base of the conidia often appear to<br />
be distinctly darkened. A Davidiella teleomorph has not previously<br />
been reported for this species.<br />
<strong>The</strong> cladosporioides complex<br />
This species complex will be treated in an additional paper in<br />
this series, dealing with the epitypification of this common <strong>and</strong><br />
widespread species, <strong>and</strong> with numerous isolates identified <strong>and</strong><br />
deposited as C. cladosporioides.<br />
DISCUSSION<br />
In the present study, a multilocus genealogy supported by light<br />
<strong>and</strong> SEM microscopy, <strong>and</strong> cultural characteristics was used to<br />
redefine species borders within <strong>Cladosporium</strong>, especially within<br />
the C. herbarum complex. Most of the diagnostic features used<br />
for species delimitation on host material (Heuchert et al. 2005,<br />
Schubert 2005), proved to be applicable in culture. However,<br />
morphological features were often more pronounced in vivo than<br />
in vitro. For instance, conidiophore arrangement is not applicable<br />
to cultures, conidiophore <strong>and</strong> conidium widths were often<br />
narrower in culture than on the natural host, <strong>and</strong> macro- as well<br />
as microconidiophores were often observed in culture, but not on<br />
host material. All species belonging to the C. herbarum complex<br />
are characterised by possessing conidia which are ornamentated,<br />
the ornamentation ranging from minutely verruculose to verrucose,<br />
echinulate or spiny whereas in the C. sphaerospermum complex<br />
species with both smooth-walled as well as ornamented conidia<br />
are included (Zalar et al. 2007). <strong>The</strong> surface ornamentation varies<br />
based on the length of surface protuberances <strong>and</strong> in the density<br />
of ornamentation. Furthermore, the conidia are mainly catenate,<br />
formed in unbranched or branched chains. However, species<br />
previously referred to the <strong>genus</strong> Heterosporium, which usually<br />
produce solitary conidia or unbranched chains of two or three<br />
conidia at the most on the natural host, also belong to this species<br />
complex (e.g., C. iridis). In vitro these chains can become longer<br />
<strong>and</strong> may even be branched. <strong>The</strong> conidiophores formed in culture<br />
are mostly macro- but may also be micronematous, sometimes<br />
forming different types of conidia that vary in shape <strong>and</strong> size from<br />
each other. Most of the species possess nodulose conidiophores<br />
with the conidiogenesis confined to the usually lateral swellings.<br />
However, this phenetic trend is not consistently expressed in all<br />
of the species belonging to the C. herbarum complex. <strong>The</strong> various<br />
<strong>Cladosporium</strong> species within the C. herbarum complex were<br />
observed to have subtle differences in their phenotype which were<br />
visible via cryo-electron microscopy (cryoSEM), <strong>and</strong> are discussed<br />
below.<br />
Fungal colonies: CryoSEM provides the opportunity to study the<br />
organisation of the fungal colony at relatively low magnifications.<br />
<strong>Cladosporium</strong> tenellum proved to be the only fungus able to form<br />
153
Schubert et al.<br />
aerial hyphal str<strong>and</strong>s under the conditions studied. <strong>Cladosporium</strong><br />
variabile formed abundant aerial hyphae, but in C. spinulosum<br />
these were sparse, <strong>and</strong> only conidiophores were observed on the<br />
agar surface. Three-day-old colonies of C. subinflatum formed<br />
numerous, long aerial hyphae, <strong>and</strong> no conidiophores could be<br />
discerned under the binocular. After 11 d the aerial hyphae seemed<br />
to have disappeared, giving rise to conidiophores. <strong>Cladosporium</strong><br />
antarcticum, C. variabile <strong>and</strong> C. ramotenellum showed very large,<br />
swollen (> 10 μm) cells which gave rise to conidiophores. With C.<br />
variabile possible earlier stages of these cells were visible (Fig. 48),<br />
which gave rise to conidiophores. More than one conidiophore could<br />
be formed on such a structure (C. variabile <strong>and</strong> C. ramotenellum).<br />
<strong>Cladosporium</strong> herbarum has very wide hyphae on the agar surface,<br />
which gave rise to conidiophores as lateral branches. <strong>The</strong>se wide<br />
hyphae were observed to anastomose, which may provide a firm<br />
interconnected supporting mycelium for these conidiophores. In<br />
C. herbaroides these wide hyphae could also be discerned, but<br />
conidiophore formation was less obvious. Similarily, C. tenellum<br />
has wide, parallel hyphae that gave rise to conidiophores.<br />
<strong>The</strong>se observations reveal fungal structures in <strong>Cladosporium</strong><br />
that have not previously been reported on, <strong>and</strong> that raise intriguing<br />
biological questions. For instance, why are hyphal str<strong>and</strong>s observed<br />
in some species (C. tenellum), <strong>and</strong> not in others, <strong>and</strong> what happens<br />
to the aerial hyphae during incubation in some species such as C.<br />
subinflatum? Furthermore, these preliminary results suggest that<br />
CryoSEM provide additional features that can be used to distinguish<br />
the different species in the C. herbarum complex.<br />
Fine details of morphological stuctures: CryoSEM provides the<br />
opportunity to study fine details of the conidiophore, (ramo)conidia<br />
<strong>and</strong> scars. Samples can be studied at magnification up to<br />
× 8 000, revealing details at a refinement far above what is<br />
possible under the light microscope (LM) (Fig. 2). However, the LM<br />
micrographs provide information about the different compartments<br />
of ramoconidia, as well as the thickness <strong>and</strong> pigmentation of the<br />
cell wall of different structures. With other words, the different<br />
techniques are complementary, <strong>and</strong> both reveal fungal details that<br />
build up the picture that defines a fungal species.<br />
Conidiophores can vary with respect to their width <strong>and</strong> the<br />
length. <strong>Cladosporium</strong> ramotenellum, C. antarcticum <strong>and</strong> C. variabile<br />
have tapered conidiophores formed on large globoid “foot cells”.<br />
<strong>The</strong> conidiophore itself can be branched. <strong>Cladosporium</strong> spinulosum<br />
has conidiophores that rise from the agar surface, but can have<br />
a common point of origin. <strong>The</strong>se conidiophores are not tapered,<br />
but parallel <strong>and</strong> slender. <strong>The</strong> conidiophores of C. bruhnei <strong>and</strong> C.<br />
herbaroides are rather long, <strong>and</strong> can appear as aerial hyphae.<br />
An important feature of the conidiophore is the location were the<br />
conidia are formed. Conidiophore ends can be simple <strong>and</strong> tubular,<br />
or rounded to more complex, several times geniculate, with several<br />
scars. Conidiophore ends become more elaborate over time.<br />
<strong>Cladosporium</strong> spinulosum <strong>and</strong> C. tenellum have nearly tubular<br />
conidiophore ends, with often very closely aggregated scars. <strong>The</strong><br />
conidiophore ends of C. subinflatum are also near tubular with a<br />
hint of bulbousness. <strong>Cladosporium</strong> subtilissimum is <strong>similar</strong>, but with<br />
somewhat more elevated scars that look denticulate. <strong>Cladosporium</strong><br />
variabile has nodulose, somewhat swollen apices with often sessile,<br />
almost inconspicuous scars. In the case of C. macrocarpum, these<br />
structures are also nodulose to nodose <strong>and</strong> somewhat bent, with<br />
only slightly protuberant loci. <strong>Cladosporium</strong> ramotenellum has<br />
tubular conidiophore ends with pronounced scars. <strong>Cladosporium</strong><br />
antarcticum has very characteristic, tapered ends, <strong>and</strong> widely<br />
dispersed (5 μm) scars. More complex conidiophore ends are<br />
more irregular in shape, <strong>and</strong> have scars dispersed over a longer<br />
distance, such as observed in C. bruhnei, C. herbaroides, <strong>and</strong> C.<br />
herbarum.<br />
Secondary ramoconidia are usually the first conidia formed on<br />
a conidiophore. <strong>The</strong>y are often multicellular, <strong>and</strong> have one basal<br />
cladosporioid hilum, <strong>and</strong> more at the apex. Few <strong>Cladosporium</strong><br />
species additionally form true ramoconidia representing apical<br />
parts of the conidiophore which secede at a septum resulting in<br />
an undifferentiated non-coronate base <strong>and</strong> function as conidia.<br />
Ramification of conidial chains is realised through these conidia.<br />
<strong>The</strong>y can occur in up to three stages, which results in elaborated<br />
spore structures. <strong>The</strong> basal secondary ramoconidium is invariably<br />
the largest, <strong>and</strong> cell size decreases through a series of additional<br />
secondary ramoconidia, intercalary conidia, <strong>and</strong> small, terminal<br />
conidia. <strong>The</strong> elongation of secondary ramoconidia varies among<br />
the different species. <strong>Cladosporium</strong> macrocarpum has broadly<br />
ellipsoid to cylindrical secondary ramoconidia usually with broadly<br />
rounded ends, like C. variabile, while C. spinulosum has secondary<br />
ramoconidia that can often hardly be discerned from the conidia<br />
that are formed at later stages. <strong>The</strong> conidia of the other species<br />
roughly fall between these species. <strong>The</strong> most notable structures on<br />
these conidia are their ornamentation, scar pattern <strong>and</strong> morphology.<br />
<strong>Cladosporium</strong> spinulosum forms numerous globose to subsphaerical<br />
spores with digitate, non-tapered surface ornamentation, which<br />
is unique for all the species discussed here. In his study on<br />
<strong>Cladosporium</strong> wall ornamentation, David (1997) recognised three<br />
classes of echinulate surfaces (aculeate, spinulose, digitate), <strong>and</strong><br />
five classes of verrucose surfaces (muricate, granulate, colliculate,<br />
pustulate <strong>and</strong> pedicellate) (Fig. 2). <strong>The</strong> ornamentation particles vary<br />
in shape, width, height <strong>and</strong> density. <strong>The</strong> most strongly ornamented<br />
conidia of the species examined by SEM are formed by C. ossifragi,<br />
with the ornamentation both large (up to 0.5 μm wide) <strong>and</strong> high,<br />
<strong>and</strong> can be regarded as densely muricately ornamented. Strong<br />
ornamentation is also seen in C. herbaroides, which is mostly<br />
granulate. <strong>Cladosporium</strong> tenellum (with muricate, granulate <strong>and</strong><br />
colliculate tendencies) <strong>and</strong> C. bruhnei (mostly granulate with some<br />
muricate projections) have relatively large ornamentation structures<br />
with slightly more space between the units than the other two<br />
species. <strong>Cladosporium</strong> antarcticum, C. ramotenellum, C. variabile<br />
<strong>and</strong> C. subtilissimum exhibit rather large granulate ornamentations<br />
that have a more irregular <strong>and</strong> variable shape. <strong>Cladosporium</strong><br />
subinflatum shows the widest dispersed structures of the series,<br />
being muricate. In contrast, C. macrocarpum has a very neat <strong>and</strong><br />
regular pattern of muricate ornamentation. <strong>The</strong> area of formation<br />
of new spores on conidia is invariably not ornamented, <strong>and</strong> hila all<br />
have the typical <strong>Cladosporium</strong> morphology with a central dome <strong>and</strong><br />
a ring-like structure around it.<br />
Branching patterns: Spores usually show a “line of weakness”<br />
between them where the coronate scars form. It seems that scars at<br />
both sides of the line of weakness have the central dome structure,<br />
which appears to play a major role in the effective mechanism<br />
<strong>Cladosporium</strong> employs for spore dispersal, with the dome actively<br />
pushing the conidia apart. This mechanism is also illustrated in<br />
David (1997, fig. 2E). Indeed, conidia of <strong>Cladosporium</strong> are very<br />
easily dislodged; even snap freezing or the electrical forces inside<br />
the SEM often result in dislodgement of the spores in a powdery<br />
“wave”. It is no surprise, therefore, that <strong>Cladosporium</strong> conidia are<br />
to be found in most air samples. In <strong>Cladosporium</strong>, conidia are<br />
mostly formed in chains, with the size invariably decreasing from<br />
the base to the apex of the row. Upon formation each conidium is<br />
separated from the conidiophore, or previously formed conidium,<br />
<strong>and</strong> hence from its nutrients. <strong>The</strong> basal ramoconidium or secondary<br />
ramoconidia have the nutrients <strong>and</strong> metabolic power to produce<br />
154
<strong>Cladosporium</strong> herbarum species complex<br />
a number of additional secondary ramoconidia that in turn could<br />
produce a chain of intercalary conidia, <strong>and</strong> finally, some small,<br />
single-celled, terminal conidia. Further research is still necessary<br />
to determine if specific branching patterns can be linked to different<br />
species.<br />
A surprising finding from the present study is the huge diversity<br />
in species <strong>and</strong> genotypes that exist in nature, be it in the indoor<br />
environment, on fruit surfaces, or in extreme ecological niches such<br />
as salterns, etc. It is clear that detailed studies would be required<br />
to find <strong>and</strong> characterise other species of <strong>Cladosporium</strong> <strong>and</strong> obtain<br />
a better underst<strong>and</strong>ing of their host ranges <strong>and</strong> ecology. A further<br />
surprise lay in the fact that several of these species are capable<br />
of sexual reproduction, <strong>and</strong> readily form Davidiella teleomorphs in<br />
culture. <strong>The</strong> Davidiella states induced here were all from homothallic<br />
species. Further attention now needs to be given to elucidating<br />
teleomorphs from other species which, as in Mycosphaerella<br />
(Groenewald et al. 2006, Ware et al. 2007) could be heterothallic,<br />
<strong>and</strong> experiencing cl<strong>and</strong>estine sex.<br />
Despite the occurrence of many different genotypes in<br />
variable genes, the degree of diversity in the entire data set was<br />
low. For the majority of the species ITS was almost invariant, with<br />
only six genotypes in the entire dataset. This suggests a very<br />
recent evolution. <strong>The</strong> st<strong>and</strong>ardised index of association (I S A ) was<br />
high (0.3914), indicating an overabundance of clonality <strong>and</strong> / or<br />
inbreeding, the latter possibly matching with observed homothallism<br />
of Davidiella teleomorphs. Clonality was visualised with SplitsTree<br />
software, where star-shaped representations without any sign of<br />
reticulation were obtained for all genes, though at different branch<br />
lengths (Fig. 5). With Structure software an optimal subdivision<br />
was achieved at six putative groups. Some of them were distinctly<br />
separated, yielding a theta (θ) around 0.14, but in most cases there<br />
was considerable overlap in representation of motifs, with θ at<br />
significantly higher values. Results are difficult to interpret due to<br />
the small size of the data set compared to the number of predicted<br />
groups, <strong>and</strong> due to unknown but probably large sampling effects.<br />
With optimal subdivision of the 79 strains at a hypothesised value<br />
of K = 6 (Fig. 4), still a large degree of inter-group <strong>similar</strong>ity was<br />
noted, as was the case at any other level of K. This was particularly<br />
obvious when data from the most variable genes (EF <strong>and</strong> ACT) are<br />
superimposed (Fig. 4). <strong>The</strong> ACT groups are further subdivided by EF<br />
data, but in many cases the same EF motif (indicated with arrows)<br />
was encountered in different (multilocus) species, for example in C.<br />
antarcticum, C. spinulosum, Davidiella sp., <strong>and</strong> the various clusters<br />
comprising <strong>Cladosporium</strong> strains which are phenotypically almost<br />
indistinguishable but genetically distinct from C. subtilissimum. A<br />
<strong>similar</strong> situation was found with the distribution of EF genotypes<br />
(indicated with doughnuts) in C. herbarum <strong>and</strong> C. macrocarpum.<br />
Nevertheless, the data set showed significant structuring, partly<br />
correlating with geography, e.g. the EF-determined cluster of C.<br />
bruhnei that contained isolates from different sources in <strong>The</strong><br />
Netherl<strong>and</strong>s. Differences may be over-accentuated by known<br />
sampling effects, particularly in C. herbarum <strong>and</strong> C. macrocarpum,<br />
where single-spore isolates from a single collection are included.<br />
Taken together the data suggest a recent, preponderantly clonal<br />
evolution, combined with limited natural selection at a low level of<br />
evolutionary pressure. As a result, many genotypes produced by<br />
hot spots in the genes analysed have survived, leading to nearly<br />
r<strong>and</strong>om variation in the data set. Many combinations of motifs that<br />
possibly could emerge have maintained in the course of time due<br />
to the absence of recombination. This indicates that the observed<br />
structure is that of populations within a single species, <strong>and</strong><br />
consequently a distinction of clonal “species” could be redundant.<br />
This conclusion is underlined by the fact that a single source in<br />
a single location can be colonised by various genotypes, such<br />
as grapes in the U.S.A. containing three different, closely related<br />
genotypes. However, the phenomenon of co-inhabitation by<br />
different Mycosphaerella species on the same lesion of Eucalyptus<br />
has been described before (Crous 1998, Crous et al. 2004) <strong>and</strong> it<br />
is therefore not surprising that different genotypes occurring close<br />
together are also observed for the related <strong>genus</strong> <strong>Cladosporium</strong>.<br />
<strong>The</strong>re is no obvious ecological difference between genotypes, <strong>and</strong><br />
hence isolates seem to have equal fitness.<br />
However, in general we noticed a remarkable concordance<br />
of genetic <strong>and</strong> phenetic characters. <strong>The</strong> morphological study was<br />
done prior to sequencing, <strong>and</strong> nearly all morphotypes clustered<br />
in separate molecular entities. <strong>The</strong>re are some exceptions, such<br />
as with C. antarcticum with striking morphology that was almost<br />
identical on the molecular level to <strong>Cladosporium</strong> spp. that resemble<br />
C. subtilissimum <strong>and</strong> would normally have been interpreted to<br />
be a mutant. Conversely, nearly all genetically distinguishable<br />
groups proved to be morphologically different, with the exception<br />
of members of the C. subtilissimum s. lat. complex (indicated as<br />
<strong>Cladosporium</strong> sp. in Fig. 3 <strong>and</strong> Table 1). <strong>The</strong> possibility remains<br />
that the found genetic parameters correlate with phenetic markers<br />
other than morphology, such as virulence, toxins or antifungal<br />
susceptibilities. For this reason we introduce the established<br />
entities here as formal species. <strong>The</strong>y can be diagnosed by ACT<br />
sequencing or by phenetic characters provided in the key. For<br />
simple routine purposes, however, they can be seen <strong>and</strong> treated<br />
as the “C. herbarum complex”, based on their close phylogenetic<br />
relationships.<br />
ACKNOWLEDGEMENTS<br />
Several colleagues from different countries provided material <strong>and</strong> valuable cultures<br />
without which this work would not have been possible. In this regard, we thank<br />
particularly Alina G. Greslebin (CIEFAP, Esquel, Chubut, Argentina), Frank Dugan<br />
<strong>and</strong> Lindsey DuToit (Washington State Univ., U.S.A.), <strong>and</strong> Annette Ramaley<br />
(Colorado, U.S.A.). Konstanze Schubert was financially supported by a Synthesys<br />
grant (No. 2559) which is gratefully acknowledged. We thank Marjan Vermaas for<br />
preparing the photographic plates, Rianne van Dijken for the growth studies, <strong>and</strong><br />
Arien van Iperen for maintaining the cultures studied. Bert Gerrits van den Ende is<br />
acknowledged for assisting with the molecular data analyses.<br />
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156
available online at www.studiesinmycology.org<br />
doi:10.3114/sim.2007.58.06<br />
Studies in Mycology 58: 157–183. 2007.<br />
Phylogeny <strong>and</strong> ecology of the ubiquitous saprobe <strong>Cladosporium</strong> sphaerospermum,<br />
with descriptions of seven new species from hypersaline environments<br />
P. Zalar 1* , G.S. de Hoog 2,3 , H.-J. Schroers 4 , P.W. Crous 2 , J.Z. Groenewald 2 <strong>and</strong> N. Gunde-Cimerman 1<br />
1<br />
Biotechnical Faculty, Department of Biology, Večna pot 111, SI-1000 Ljubljana, Slovenia; 2 <strong>CBS</strong> Fungal Biodiversity Centre, P.O. Box 85167, 3508 AD Utrecht, <strong>The</strong> Netherl<strong>and</strong>s;<br />
3<br />
Institute for Biodiversity <strong>and</strong> Ecosystem Dynamics, University of Amsterdam, Kruislaan 315, 1098 SM Amsterdam, <strong>The</strong> Netherl<strong>and</strong>s; 4 Agricultural Institute of Slovenia,<br />
Hacquetova 17, p.p. 2553, 1001 Ljubljana, Slovenia<br />
*Correspondence: Polona Zalar, polona.zalar@bf.uni-lj.si<br />
Abstract: Saprobic <strong>Cladosporium</strong> isolates morphologically <strong>similar</strong> to C. sphaerospermum are phylogenetically analysed on the basis of DNA sequences of the ribosomal RNA<br />
gene cluster, including the internal transcribed spacer regions ITS1 <strong>and</strong> ITS2, the 5.8S rDNA (ITS) <strong>and</strong> the small subunit (SSU) rDNA as well as β-tubulin <strong>and</strong> actin gene introns<br />
<strong>and</strong> exons. Most of the C. sphaerospermum-like species show halotolerance as a recurrent feature. <strong>Cladosporium</strong> sphaerospermum, which is characterised by almost globose<br />
conidia, is redefined on the basis of its ex-neotype culture. <strong>Cladosporium</strong> dominicanum, C. psychrotolerans, C. velox, C. spinulosum <strong>and</strong> C. halotolerans, all with globoid conidia,<br />
are newly described on the basis of phylogenetic analyses <strong>and</strong> cryptic morphological <strong>and</strong> physiological characters. <strong>Cladosporium</strong> halotolerans was isolated from hypersaline<br />
water <strong>and</strong> bathrooms <strong>and</strong> detected once on dolphin skin. <strong>Cladosporium</strong> dominicanum <strong>and</strong> C. velox were isolated from plant material <strong>and</strong> hypersaline water. <strong>Cladosporium</strong><br />
psychrotolerans, which grows well at 4 °C but not at 30 °C, <strong>and</strong> C. spinulosum, having conspicuously ornamented conidia with long digitate projections, are currently only known<br />
from hypersaline water. We also newly describe C. salinae from hypersaline water <strong>and</strong> C. fusiforme from hypersaline water <strong>and</strong> animal feed. Both species have ovoid to ellipsoid<br />
conidia <strong>and</strong> are therefore reminiscent of C. herbarum. <strong>Cladosporium</strong> langeronii (= Hormodendrum langeronii) previously described as a pathogen on human skin, is halotolerant<br />
but has not yet been recorded from hypersaline environments.<br />
Taxonomic novelties: <strong>Cladosporium</strong> dominicanum Zalar, de Hoog & Gunde-Cimerman, sp. nov., C. fusiforme Zalar, de Hoog & Gunde-Cimerman, sp. nov., C. halotolerans<br />
Zalar, de Hoog & Gunde-Cimerman, sp. nov., C. psychrotolerans Zalar, de Hoog & Gunde-Cimerman, sp. nov., C. salinae Zalar, de Hoog & Gunde-Cimerman, sp. nov.,<br />
C. spinulosum Zalar, de Hoog & Gunde-Cimerman, sp. nov., C. velox Zalar, de Hoog & Gunde-Cimerman, sp. nov.<br />
Key words: Actin, β-tubulin, halotolerance, ITS rDNA, phylogeny, SSU rDNA, taxonomy.<br />
INTRODUCTION<br />
<strong>The</strong> halophilic <strong>and</strong> halotolerant mycobiota from hypersaline<br />
aqueous habitats worldwide frequently contain <strong>Cladosporium</strong><br />
Link isolates (Gunde-Cimerman et al. 2000, Butinar et al. 2005).<br />
Initially, they were considered as airborne contaminants, but<br />
surprisingly, many of these <strong>Cladosporium</strong> isolates were identified<br />
as C. sphaerospermum Penz. because they formed globoid conidia<br />
(data unpublished). <strong>Cladosporium</strong> sphaerospermum, known as<br />
one of the most common air-borne, cosmopolitan <strong>Cladosporium</strong><br />
species, was frequently isolated from indoor <strong>and</strong> outdoor air (Park<br />
et al. 2004), dwellings (Aihara et al. 2001), <strong>and</strong> occasionally from<br />
humans (Badillet et al. 1982) <strong>and</strong> plants (Pereira et al. 2002).<br />
Strains morphologically identified as C. sphaerospermum were<br />
able to grow at a very low water activity (a w<br />
0.816), while other<br />
cladosporia clearly preferred a higher, less extreme water activity<br />
(Hocking et al. 1994). This pronounced osmotolerance suggests<br />
a predilection for osmotically stressed environments although<br />
C. sphaerospermum is reported from a wide range of habitats<br />
including osmotically non-stressed niches.<br />
We therefore hypothesised that C. sphaerospermum<br />
represents a complex of species having either narrow or wide<br />
ecological amplitudes. <strong>The</strong> molecular diversity of strains identified<br />
as C. sphaerospermum has not yet been determined <strong>and</strong> isolates<br />
from humans have not yet been critically compared with those from<br />
environmental samples. <strong>The</strong>refore, a taxonomic study was initiated<br />
with the aim to define phylogenetically <strong>and</strong> morphologically distinct<br />
entities <strong>and</strong> to describe their in vitro osmotolerance <strong>and</strong> their natural<br />
ecological preferences.<br />
MATERIALS AND METHODS<br />
Sampling<br />
Samples of hypersaline water were collected from salterns located<br />
at different sites of the Mediterranean basin (Slovenia, Bosnia <strong>and</strong><br />
Herzegovina, Spain), different coastal areas along the Atlantic<br />
Ocean (Monte Cristy, Dominican Republic; Swakopmund, Namibia),<br />
the Red Sea (Eilat, Israel), the Dead Sea (Ein Gedi, Israel), <strong>and</strong><br />
the salt Lake Enriquillio (Dominican Republic). Samples from the<br />
Sečovlje salterns (Slovenia) were collected once per month in 1999.<br />
Samples from the Santa Pola salterns <strong>and</strong> Ebre delta river saltern<br />
(Spain) were taken twice (July <strong>and</strong> November) in 2000. A saltern in<br />
Namibia <strong>and</strong> one in the Dominican Republic were sampled twice<br />
(August <strong>and</strong> October) in 2002. Various salinities, ranging from 15 to<br />
32 % NaCl were encountered in these ponds.<br />
Isolation <strong>and</strong> maintenance of fungi<br />
Strains were isolated from salterns using filtration of hypersaline<br />
water through membrane filters (pore diam 0.45 μm), followed by<br />
incubation of the membrane filters on different culture media with<br />
lowered water activity (Gunde-Cimerman et al. 2000). Only colonies<br />
of different morphology on one particular selective medium per<br />
sample were analysed further. Strains were carefully selected from<br />
different evaporation ponds, collected at different times, in order to<br />
avoid sampling of identical clones. Subcultures were maintained<br />
at the Culture Collection of Extremophilic Fungi (EXF, Biotechnical<br />
Faculty, Ljubljana, Slovenia), while a selection was deposited at<br />
the Centraalbureau voor Schimmelcultures (<strong>CBS</strong>, Utrecht, <strong>The</strong><br />
Netherl<strong>and</strong>s) <strong>and</strong> the Culture Collection of the National Institute<br />
of Chemistry (MZKI, Ljubljana, Slovenia). Reference strains were<br />
obtained from <strong>CBS</strong>, <strong>and</strong> were selected either on the basis of the<br />
strain history, name, or on the basis of their ITS rDNA sequence.<br />
Strains were maintained on oatmeal agar (OA; diluted OA, Difco:<br />
15 g of Difco 255210 OA medium, 12 g of agar, dissolved in 1 L<br />
of distilled water) with or without 5 % additional NaCl. <strong>The</strong>y were<br />
preserved in liquid nitrogen or by lyophilisation. Strains studied are<br />
listed in Table 1.<br />
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Zalar et al.<br />
Table 1. List of <strong>Cladosporium</strong> strains, with their current <strong>and</strong> original names, geography, GenBank accession numbers <strong>and</strong> references to earlier published sequences.<br />
Strain Nr. a Source Geography GenBank accession Nr. b<br />
ITS rDNA / 18S rDNA actin β-tubulin<br />
<strong>Cladosporium</strong> bruhnei<br />
<strong>CBS</strong> 177.71 Thuja tincture <strong>The</strong> Netherl<strong>and</strong>s, Amsterdam DQ780399 / DQ780938 EF101354 EF101451<br />
<strong>CBS</strong> 812.71 Polygonatum odoratum, leaf Czech Republic, Lisen DQ780401 / – – –<br />
<strong>Cladosporium</strong> cladosporioides<br />
<strong>CBS</strong> 170.54 NT Arundo, leaf U.K., Engl<strong>and</strong>, Kew AY213640 / DQ780940 EF101352 EF101453<br />
EXF-321 Hypersaline water Slovenia, Sečovlje saltern DQ780408 / – – –<br />
EXF-780 DQ780409 / – – –<br />
EXF-946 Hypersaline water Bosnia <strong>and</strong> Herzegovina, DQ780410 / – – –<br />
Ston saltern<br />
<strong>Cladosporium</strong> dominicanum<br />
CPC 11683 Citrus fruit (orange) Iran DQ780357 / – EF101369 EF101419<br />
EXF-696 Hypersaline water Dominican Republic, saltern DQ780358 / – EF101367 EF101420<br />
EXF-718 Hypersaline water Dominican Republic, salt lake DQ780356 / – EF101370 EF101418<br />
Enriquilio<br />
EXF-720 Hypersaline water Dominican Republic, saltern DQ780355 / – – EF101417<br />
EXF-727 Hypersaline water Dominican Republic, saltern DQ780354 / – – EF101416<br />
EXF-732 T; <strong>CBS</strong> 119415 Hypersaline water Dominican Republic, salt lake DQ780353 / – EF101368 EF101415<br />
Enriquilio<br />
<strong>Cladosporium</strong> fusiforme<br />
<strong>CBS</strong> 452.71 Chicken food Canada DQ780390 / – EF101371 EF101447<br />
EXF-397 Hypersaline water Slovenia, Sečovlje saltern DQ780389 / – EF101373 EF101445<br />
EXF-449 T; <strong>CBS</strong> 119414 Hypersaline water Slovenia, Sečovlje saltern DQ780388 / DQ780935 EF101372 EF101446<br />
<strong>Cladosporium</strong> herbarum<br />
ATCC 66670, as Davidiella tassiana CCA-treated Douglas-fir pole U.S.A., New York, Geneva AY361959 2 & DQ780400 / DQ780939 AY752193 11 EF101452<br />
<strong>Cladosporium</strong> halotolerans<br />
ATCC 26362 Liver <strong>and</strong> intestine of diseased frog U.S.A., New Jersey AY361982 2 / – – –<br />
ATCC 64726 Peanut cell suspension tissue culture U.S.A., Georgia AY361968 2 / – – –<br />
<strong>CBS</strong> 280.49<br />
Stem of Hypericum perforatum Switzerl<strong>and</strong>, Glarus,<br />
DQ780369 / – EF101402 EF101432<br />
identified as Mycosphaerella hyperici Mühlehorn<br />
<strong>CBS</strong> 191.54 Laboratory air Great Britain – / – – –<br />
<strong>CBS</strong> 573.78 Aureobasidium caulivorum Russia, Moscow region – / – – –<br />
<strong>CBS</strong> 626.82 – Sweden, Stockholm – / – – –<br />
dH 12862; EXF-2533 Culture contaminant Brazil DQ780371 / – EF101400 EF101422<br />
dH 12941; EXF-2534 Culture contaminant Turkey – / – EF101421<br />
dH 12991; EXF-2535 Brain Turkey DQ780372 / – EF101423<br />
dH 13911; EXF-2422 Ice Arctics DQ780370 / – EF101401 EF101430<br />
EXF-228; MZKI B-840 Hypersaline water Slovenia, Sečovlje saltern DQ780365 / DQ780930 EF101393 EF101425<br />
EXF-380 Hypersaline water Slovenia, Sečovlje saltern DQ780368 / – EF101394 EF101427<br />
EXF-564 Hypersaline water Namibia, saltern DQ780363 / – EF101395 EF101433<br />
EXF-565 Hypersaline water Namibia, saltern – / – – –<br />
EXF-567 Hypersaline water Namibia, saltern – / – – –<br />
EXF-571 Hypersaline water Namibia, saltern – / – – –<br />
EXF-572 T; <strong>CBS</strong> 119416 Hypersaline water Namibia, saltern DQ780364 / – EF101397 EF101424<br />
EXF-646 Hypersaline water Spain, Santa Pola saltern DQ780366 / – EF101398 EF101428<br />
EXF-698 Hypersaline water Dominican Republic, saltern – / – – –<br />
EXF-703 Hypersaline water Dominican Republic, salt lake DQ780367 / – EF101392 EF101426<br />
Enriquilio<br />
EXF-944 Hypersaline water Bosnia <strong>and</strong> Herzegovina, – / – – –<br />
Ston saltern<br />
EXF-972 Bathroom Slovenia – / – – –<br />
EXF-977 Bathroom Slovenia DQ780362 / – EF101396 EF101431<br />
158
<strong>Cladosporium</strong> sphaerospermum species complex<br />
Strain Nr. a Source Geography GenBank accession Nr. b<br />
ITS rDNA / 18S rDNA actin β-tubulin<br />
EXF-1072 Hypersaline water Israel, Dead Sea DQ780373 / – EF101399 EF101428<br />
EXF-2372 Hypersaline water Slovenia, Sečovlje saltern – / – – –<br />
UAMH 7686<br />
Indoor air ex RCS strip, from Apis<br />
mellifera overwintering facility<br />
U.S.A., Alta, Clyde Corner AY625063 5 / – – –<br />
– rDNA from bottlenose dolphin skin U.S.A., Texas AF035674 6 / – – –<br />
infected with Loboa loboi<br />
– Microcolony, on rock Turkey, Antalya AJ971409 7 / – – –<br />
– Microcolony, on rock Turkey, Antalya AJ971408 7 / – – –<br />
– Tomato leaves – L25433 8 / – – –<br />
<strong>Cladosporium</strong> langeronii<br />
<strong>CBS</strong> 189.54 NT Mycosis Brazil DQ780379 / DQ780932 EF101357 EF101435<br />
<strong>CBS</strong> 601.84 Picea abies, wood Germany, Göttingen DQ780382 / – EF101360 EF101438<br />
<strong>CBS</strong> 101880 Moist aluminium school window frame Belgium, Lichtervoorde DQ780380 / – EF101359 EF101440<br />
<strong>CBS</strong> 109868 Mortar of Muro Farnesiano Italy, Parma DQ780377 / – EF101362 EF101434<br />
dH 11736 Biomat in a lake Antarctics DQ780381 / – EF101363 EF101436<br />
dH 12459 Orig. face lesion Brazil DQ780378 / – EF101358 EF101439<br />
dH 13833 Ice Arctics DQ780383 / – EF101361 EF101437<br />
– Nasal mucus – AF455525 4 / – – –<br />
– Nasal mucus – AY345352 4 / – – –<br />
– Mycorrhizal roots – DQ068982 9 / – – –<br />
<strong>Cladosporium</strong> oxysporum<br />
ATCC 66669 Creosote-treated southern pine pole U.S.A., New York,<br />
AF393689 10 / DQ780395 AY752192 11 EF101454<br />
Binghamton<br />
ATCC 76499 Decayed leaf, Lespedeza bicolor – AF393720 – –<br />
<strong>CBS</strong> 125.80 Cirsium vulgare, seedcoat <strong>The</strong> Netherl<strong>and</strong>s AJ300332 12 / DQ780941 EF101351 EF101455<br />
EXF-697 Hypersaline water Dominican Republic, salt lake DQ780392 / – – –<br />
Enriquilio<br />
EXF-699 Hypersaline water Dominican Republic, saltern DQ780394 / – – –<br />
EXF-710 Hypersaline water Dominican Republic, saltern DQ780393 / – – –<br />
EXF-711 Hypersaline water Dominican Republic, saltern DQ780391 / – – –<br />
<strong>Cladosporium</strong> psychrotolerans<br />
EXF-326 Hypersaline water Slovenia, Sečovlje saltern DQ780387 / DQ780934 – EF101444<br />
EXF-332 Hypersaline water Slovenia, Sečovlje saltern DQ780385 / DQ780933 EF101364 EF101441<br />
EXF-391 T; <strong>CBS</strong> 119412 Hypersaline water Slovenia, Sečovlje saltern DQ780386 / – EF101365 EF101442<br />
EXF-714 Hypersaline water Dominican Republic DQ780384 / – EF101366 EF101443<br />
<strong>Cladosporium</strong> ramotenellum<br />
EXF-454 T; CPC 12043 Hypersaline water Slovenia, Sečovlje saltern DQ780403 / – – –<br />
<strong>Cladosporium</strong> salinae<br />
EXF-322 Hypersaline water Slovenia, Sečovlje DQ780375 / – EF101391 EF101403<br />
EXF-335 T; <strong>CBS</strong> 119413 Hypersaline water Slovenia, Sečovlje DQ780374 / DQ780931 EF101390 EF101405<br />
EXF-604 Hypersaline water Spain, Santa Pola DQ780376 / – EF101389 EF101404<br />
<strong>Cladosporium</strong> sp.<br />
<strong>CBS</strong> 300.96 Soil along coral reef coast Papua New Guinea, Madang, DQ780352 / – EF101385 –<br />
Jais Aben<br />
EXF-595 Hypersaline water Spain, Santa Pola saltern DQ780402 / – – –<br />
<strong>Cladosporium</strong> sphaerospermum<br />
ATCC 12092 Soil Canada AY361988 2 / – – –<br />
ATCC 200384 Compost biofilter <strong>The</strong> Netherl<strong>and</strong>s AY361991 2 / – – –<br />
<strong>CBS</strong> 109.14; ATCC 36950 Carya illinoensis leaf scale U.S.A. DQ780350 / – EF101384 EF101410<br />
<strong>CBS</strong> 122.47; IFO 6377; IMI 49640; Decaying stem of Begonia sp., <strong>The</strong> Netherl<strong>and</strong>s, Aalsmeer AJ244228 1 / – – –<br />
VKM F-772; ATCC 11292<br />
with Thielaviopsis basicola<br />
<strong>CBS</strong> 188.54; ATCC 11290; IMI de Vries (Engelhardt strain) – AY361990 2 & AY251077 3 / – – –<br />
049638<br />
<strong>CBS</strong> 190.54; ATCC 11293; IFO 6380; – – AY361992 2 / – – –<br />
IMI 49641<br />
<strong>CBS</strong> 192.54; ATCC 11288; IMI 49636 Nail of man – AY361989 2 / – – –<br />
www.studiesinmycology.org<br />
159
Zalar et al.<br />
Table 1. (Continued).<br />
Strain Nr. a Source Geography GenBank accession Nr. b<br />
<strong>CBS</strong> 193.54 NT; ATCC 11289; IMI<br />
49637<br />
ITS rDNA / 18S rDNA actin β-tubulin<br />
Human nails – DQ780343 & AY361958 2 / DQ780925 EF101380 EF101406<br />
<strong>CBS</strong> 122.63 Plywood of Betula sp. Finl<strong>and</strong>, Helsinki – / – – –<br />
<strong>CBS</strong> 102045; EXF-2524; MZKI B- Hypersaline water<br />
Spain, Barcelona, Salines de DQ780351 / – EF101378 EF101411<br />
1066<br />
la Trinitat<br />
<strong>CBS</strong> 114065 Outdoor air Germany, Stuttgart – / – – –<br />
CPC 10944 Gardening peat substrate Russia, Kaliningrad DQ780350 / – – –<br />
EXF-131; MZKI B-1005 Hypersaline water Slovenia, Sečovlje saltern AJ238670 1 / – – –<br />
EXF-328 Hypersaline water Slovenia, Sečovlje saltern – / – – –<br />
EXF-385 Hypersaline water Slovenia, Sečovlje saltern – / – – –<br />
EXF-446 Hypersaline water Slovenia, Sečovlje saltern – / – – –<br />
EXF-455 Hypersaline water Slovenia, Sečovlje saltern DQ780349 / – EF101375 EF101412<br />
EXF-458 Hypersaline water Slovenia, Sečovlje saltern DQ780345 / – EF101374 EF101409<br />
EXF-461 Hypersaline water Slovenia, Sečovlje saltern – / – – –<br />
EXF-464 Hypersaline water Slovenia, Sečovlje saltern – / DQ780927 – –<br />
EXF-465 Hypersaline water Slovenia, Sečovlje saltern – / – – –<br />
EXF-598 Hypersaline water Spain, Santa Pola – / – EF101377 –<br />
EXF-644 Hypersaline water Spain, Santa Pola – / – – –<br />
EXF-645 Hypersaline water Spain, Santa Pola – / – – –<br />
EXF-649 Hypersaline water Spain, Santa Pola – / – – –<br />
EXF-715 Hypersaline water Dominican Republic, saltern – / – – –<br />
EXF-738 Bathroom Slovenia DQ780348 / – EF101383 EF101414<br />
EXF-739 Bathroom Slovenia DQ780344 / – EF101381 EF101407<br />
EXF-781; MZKI B-899 Hypersaline water Slovenia, Sečovlje – / – – –<br />
EXF-788 Hypersaline water Slovenia, Sečovlje – / – – –<br />
EXF-962 Bathroom Slovenia DQ780347 / – EF101382 EF101413<br />
EXF-965 Bathroom Slovenia – / – – –<br />
EXF-1069 Hypersaline water Israel, Eilat saltern – / – EF101376 –<br />
EXF-1061 Hypersaline water Israel, Dead Sea DQ780346 / – EF101379 EF101408<br />
EXF-1726 Hypersaline water Israel, Dead Sea – / – – –<br />
EXF-1732 Hypersaline water Israel, Eilat saltern – / DQ780928 – –<br />
– Bryozoa sp. – AJ557744 / – – –<br />
– Nasal mucus – AF455481 4 / – – –<br />
<strong>Cladosporium</strong> spinulosum<br />
EXF-333 Hypersaline water Slovenia, Sečovlje saltern DQ780404 / – – –<br />
EXF-334 T Hypersaline water Slovenia, Sečovlje saltern DQ780406 / – EF101355 EF101450<br />
EXF-382 Hypersaline water Slovenia, Sečovlje saltern DQ780407 / DQ780936 EF101356 EF101449<br />
<strong>Cladosporium</strong> subinflatum<br />
EXF-343 T; CPC 12041 Hypersaline water Slovenia, Sečovlje saltern DQ780405 / – EF101353 EF101448<br />
<strong>Cladosporium</strong> tenuissimum<br />
ATCC 38027 Soil New Caledonia AF393724 / – – –<br />
EXF-324 Hypersaline water Slovenia, Sečovlje saltern – / DQ780926 – –<br />
EXF-371 Hypersaline water Slovenia, Sečovlje saltern DQ780396 / – – –<br />
EXF-452 Hypersaline water Slovenia, Sečovlje saltern DQ780397 / – – –<br />
EXF-563 Hypersaline water Namibia, saltern DQ780398 / – – –<br />
<strong>Cladosporium</strong> velox<br />
<strong>CBS</strong> 119417 T; CPC 11224 Bamboo sp. India, Charidij DQ780361 / DQ780937 EF101388 EF101456<br />
EXF-466 Hypersaline water Slovenia, Sečovlje saltern DQ780359 / – EF101386 –<br />
EXF-471 Hypersaline water Slovenia, Sečovlje saltern DQ780360 / – EF101387 –<br />
160
<strong>Cladosporium</strong> sphaerospermum species complex<br />
Cultivation <strong>and</strong> microscopy<br />
For growth rate determination <strong>and</strong> phenetic description of colonies,<br />
strains were point inoculated on potato-dextrose agar (PDA, Difco),<br />
OA <strong>and</strong> Blakeslee malt extract agar (MEA, Samson et al. 2002)<br />
<strong>and</strong> incubated at 25 °C for 14 d in darkness. Surface colours were<br />
rated using the colour charts of Kornerup & Wanscher (1967). For<br />
studies of microscopic morphology, strains were grown on synthetic<br />
nutrient agar (SNA, Gams et al. 2007) in slide cultures. SNA blocks<br />
of approximately 1 × 1 cm were cut out aseptically, placed upon<br />
sterile microscope slides, <strong>and</strong> inoculated at the upper four edges<br />
by means of a conidial suspension (Pitt 1979). Inoculated agar<br />
blocks were covered with sterile cover slips <strong>and</strong> incubated in moist<br />
chambers for 7 d at 25 °C in darkness. <strong>The</strong> structure <strong>and</strong> branching<br />
pattern of conidiophores were observed at magnifications × 100,<br />
× 200 <strong>and</strong> × 400 in intact slide cultures under the microscope<br />
without removing the cover slips from the agar blocks. For higher<br />
magnifications (× 400, × 1 000) cover slips were carefully removed<br />
<strong>and</strong> mounted in lactic acid with aniline blue.<br />
Morphological parameters<br />
Morphological terms follow David (1997), Kirk et al. (2001) <strong>and</strong><br />
Schubert et al. (2007 – this volume). Conidiophores in <strong>Cladosporium</strong><br />
are usually ascending <strong>and</strong> sometimes poorly differentiated. Though<br />
the initiation point of conidiophore stipes could sometimes be<br />
determined only approximately, their lengths were in some cases<br />
useful for distinguishing morphologically <strong>similar</strong> species when<br />
observed in slide cultures. <strong>The</strong> branching patterns can be rotationally<br />
symmetric or unilateral. Characters of conidial scars were studied<br />
by light <strong>and</strong> scanning electron microscopy (SEM). Conidial chains<br />
show different branching patterns, determined by the numbers of<br />
conidia in unbranched parts, the nature of ramoconidia as well as<br />
their distribution in conidial chains. Measurements are given as (i)<br />
n 1<br />
–n 2<br />
or (ii) (n 1<br />
–)n 3<br />
–n 4<br />
(–n 2<br />
), with n 1<br />
= minimum value observed; n 2<br />
= maximum value observed; n 3<br />
/n 4<br />
= first/third quartile. For conidia<br />
<strong>and</strong> ramoconidia also average values <strong>and</strong> st<strong>and</strong>ard deviations are<br />
listed. <strong>The</strong> values provided are based on at least 25 measurements<br />
for the conidiophores of each strain, <strong>and</strong> at least 50 measurements<br />
for conidia.<br />
Ecophysiology<br />
To determine the degree of halotolerance, strains were pointinoculated<br />
on MEA without <strong>and</strong> with additional NaCl at concentrations<br />
of 5, 10, 17 <strong>and</strong> 20 % NaCl (w/v) <strong>and</strong> incubated at 25 °C for 14 d.<br />
To determine cardinal temperature requirements for growth, plates<br />
were incubated at 4, 10, 25, 30 <strong>and</strong> 37 °C, <strong>and</strong> colony diameters<br />
measured after 14 d of incubation.<br />
DNA extraction, sequencing <strong>and</strong> analysis<br />
For DNA isolation strains were grown on MEA for 7 d. DNA was<br />
extracted according to Gerrits van den Ende & de Hoog (1999)<br />
by mechanical lysis of approx. 1 cm 2 of mycelium. A fragment of<br />
the rDNA including the Internal Transcribed Spacer region 1, 5.8S<br />
rDNA <strong>and</strong> the ITS 2 (ITS) was amplified using the primers V9G<br />
(de Hoog & Gerrits van den Ende 1998) <strong>and</strong> LS266 (Masclaux<br />
et al. 1995). Sequence reactions were done using primers ITS1<br />
<strong>and</strong> ITS4 (White et al. 1990). For amplification <strong>and</strong> sequencing<br />
of the partial actin gene, primers ACT-512F <strong>and</strong> ACT-783R were<br />
applied according to Carbone & Kohn (1999). For amplification <strong>and</strong><br />
sequencing of the β-tubulin gene primers T1 <strong>and</strong> T22 were used<br />
according to O’Donnell & Cigelnik (1997). A BigDye terminator<br />
cycle sequencing kit (Applied Biosystems, Foster City, CA, U.S.A.)<br />
was used in sequence reactions. Sequences were obtained with<br />
an ABI Prism 3700 DNA Analyzer (Applied Biosystems). <strong>The</strong>y<br />
were assembled <strong>and</strong> edited using SeqMan v. 3.61 (DNAStar,<br />
Inc., Madison, U.S.A.). Sequences downloaded from GenBank<br />
are indicated in the trees by their GenBank accession numbers;<br />
newly generated sequences are indicated by strain numbers<br />
(see also Table 1). Sequences were automatically aligned using<br />
ClustalX v. 1.81 (Jeanmougin et al. 1998). <strong>The</strong> alignments were<br />
adjusted manually using MEGA3 (Kumar et al. 2004). Phylogenetic<br />
relationships of the taxa were estimated from aligned sequences<br />
by the maximum parsimony criterion as implemented in PAUP v.<br />
4.0b10 (Swofford 2003). Data sets of the SSU rDNA, ITS rDNA<br />
<strong>and</strong> the β-tubulin <strong>and</strong> actin genes are analysed separately. Species<br />
of <strong>Cladosporium</strong> s. str. were compared with various taxa of the<br />
Mycosphaerellaceae using SSU rDNA sequences <strong>and</strong> Fusicladium<br />
effusum G. Winter (Venturiaceae) as outgroup. <strong>The</strong> other data sets<br />
focus on <strong>Cladosporium</strong> s. str., using <strong>Cladosporium</strong> salinae Zalar,<br />
de Hoog & Gunde-Cimerman as an outgroup, because this species<br />
was most deviant within <strong>Cladosporium</strong> in the SSU rDNA analysis<br />
(see below). Heuristic searches were performed on all characters,<br />
which were unordered <strong>and</strong> equally weighted. Gaps were treated<br />
as missing characters. Starting tree(s) were obtained via stepwise,<br />
r<strong>and</strong>om, 100 times repeated sequence addition. Other parameters<br />
included a “MaxTrees” setting to 9 000, the tree-bisectionreconnection<br />
as branch-swapping algorithm, <strong>and</strong> the “MulTrees”<br />
option set to active. Branch robustness was tested in the parsimony<br />
analysis by 10 000 search replications, each on bootstrapped data<br />
sets using a fast step-wise addition bootstrap analysis. Bootstrap<br />
values larger than 60 are noted near their respective branches.<br />
Newly generated sequences were deposited in GenBank (www.<br />
ncbi.nlm.nih.gov); their accession numbers are listed in Table 1.<br />
Alignments <strong>and</strong> trees were deposited in TreeBASE (www.treebase.<br />
org).<br />
Table 1. (Page 158–160).<br />
a<br />
Abbreviations used: ATCC = American Type Culture Collection, Virginia, U.S.A.; <strong>CBS</strong> = Centraalbureau voor Schimmelcultures, Utrecht, <strong>The</strong> Netherl<strong>and</strong>s; CPC = Culture<br />
Collection of Pedro Crous, housed at <strong>CBS</strong>, Utrecht, <strong>The</strong> Netherl<strong>and</strong>s; dH = de Hoog Culture Collection, housed at <strong>CBS</strong>, Utrecht, <strong>The</strong> Netherl<strong>and</strong>s; EXF = Culture Collection<br />
of Extremophilic Fungi, Ljubljana, Slovenia; IFO = Institute for Fermentation, Culture Collection of Microorganizms, Osaka, Japan; IMI = <strong>The</strong> International Mycological<br />
Institute, Egham, Surrey, U.K.; MZKI = Microbiological Culture Collection of the National Institute of Chemistry, Ljubljana, Slovenia; UAMH = University of Alberta Microfungus<br />
Collection, Alberta, Canada; VKM = All-Russian Collection of Microorganisms, Russian Academy of Sciences, Institute of Biochemistry <strong>and</strong> Physiology of Microorganisms,<br />
Pushchino, Russia; NT = ex-neotype strain; T = ex-type strain.<br />
b<br />
Reference: 1 de Hoog et al. 1999; 2 Park et al. 2004; 3 Braun et al. 2003; 4 Buzina et al. 2003; 5 Meklin et al. 2004; 6 Haubold et al. 1998; 7 Sert & Sterflinger, unpubl.; 8 Curtis et<br />
al. 1994; 9 Menkis et al. 2005; 10 Managbanag et al. unpubl.; 11 Crous et al. 2004; 12 Wirsel et al. 2002. All others are newly reported here.<br />
www.studiesinmycology.org<br />
161
Zalar et al.<br />
AY251097 <strong>Cladosporium</strong> uredinicola<br />
<strong>Cladosporium</strong> oxysporum <strong>CBS</strong> 125.80<br />
<strong>Cladosporium</strong> herbarum ATCC 66670<br />
<strong>Cladosporium</strong> bruhnei <strong>CBS</strong> 177.71<br />
<strong>Cladosporium</strong> spinulosum EXF-382<br />
<strong>Cladosporium</strong> psychrotolerans EXF-326<br />
<strong>Cladosporium</strong> psychrotolerans EXF-332<br />
<strong>Cladosporium</strong> langeronii <strong>CBS</strong> 189.54 T<br />
<strong>Cladosporium</strong> tenuissimum EXF-324<br />
<strong>Cladosporium</strong> cladosporioides <strong>CBS</strong> 170.54 NT<br />
AY251092 <strong>Cladosporium</strong> colocasiae<br />
AY251096 <strong>Cladosporium</strong> herbarum<br />
98<br />
<strong>Cladosporium</strong> halotolerans <strong>CBS</strong> 119416 T<br />
<strong>Cladosporium</strong> halotolerans EXF-228<br />
<strong>Cladosporium</strong> dominicanum <strong>CBS</strong> 119415 T<br />
<strong>Cladosporium</strong> velox <strong>CBS</strong> 119417 T<br />
66<br />
<strong>Cladosporium</strong> sphaerospermum EXF-464<br />
<strong>Cladosporium</strong> fusiforme <strong>CBS</strong> 119414 T<br />
<strong>Cladosporium</strong> sphaerospermum <strong>CBS</strong> 193.54 NT<br />
<strong>Cladosporium</strong> salinae <strong>CBS</strong> 119413 T<br />
AY251107 Pseudocercospora protearum<br />
80 AY251106 Pseudocercospora angolensis<br />
64<br />
AY251105 Cercospora cruenta<br />
AY251104 Cercospora zebrina<br />
AY251109 Passalora fulva<br />
79<br />
AY251108 Passalora dodonaeae<br />
AY251117 Mycosphaerella graminicola<br />
AY251110 Ramulispora sorghi<br />
100<br />
AY251114 Mycosphaerella latebrosa<br />
AY251102 Batcheloromyces proteae<br />
AY251101 Mycosphaerella lateralis<br />
76 AY251120 Teratosphaeria nubilosa<br />
AY251118 Catenulostroma macowanii<br />
AY251121 Devriesia staurophorum<br />
97 U42474 Dothidea insculpta<br />
93 AY016343 Dothidea ribesia<br />
AY016342 Discosphaerina fagi<br />
AY251126 Fusicladium effusum<br />
AY251122 Fusicladium amoenum Venturiaceae<br />
Dothioraceae & Dothideaceae<br />
10 steps<br />
Fig. 1. One of 30 equally most parsimonious <strong>and</strong> equally looking phylogenetic trees based on a heuristic tree search using aligned small subunit ribosomal DNA sequences.<br />
<strong>The</strong> tree was r<strong>and</strong>omly selected. Support based on 10 000 replicates of a fast step-wise addition bootstrap analysis is indicated near the branches. Species of <strong>Cladosporium</strong> s.<br />
str., including the seven newly described species, form a strongly supported monophyletic group among other taxa of the Mycosphaerellaceae (Dothideomycetes) (CI = 0.631,<br />
RI = 0.895, PIC = 50).<br />
Teratosphaeriaceae<br />
Davidiellaceae<br />
Mycosphaerellaceae<br />
Table 2. Statistical parameters describing phylogenetic analyses performed on sequence alignments of four different loci.<br />
Parameter SSU rDNA ITS rDNA 1 β-tubulin 2 Actin 3<br />
Number of alignment positions 1031 498 654 210<br />
Number of parsimony informative characters (PIC) 50 68 220 103<br />
Length of tree / number of steps 103 102 714 338<br />
Consistency Index (CI) 0.631 0.804 0.538 0.586<br />
Retention Index (RI) 0.895 0.975 0.883 0.885<br />
Rescaled Consistency Index (RC) 0.565 0.784 0.475 0.518<br />
Homoplasy index (HI) 0.369 0.196 0.462 0.414<br />
Number of equally parsimonious trees retained 30 600 90 32<br />
1<br />
Including the internal transcribed spacer region 1 <strong>and</strong> 2 <strong>and</strong> the 5.8S rDNA.<br />
2<br />
Including partial sequences of 4 exons <strong>and</strong> complete sequences of 3 introns.<br />
3<br />
Including partial sequences of 3 exons <strong>and</strong> 2 introns.<br />
162
<strong>Cladosporium</strong> sphaerospermum species complex<br />
69<br />
EXF-322<br />
<strong>CBS</strong> 119413<br />
EXF-604<br />
10 steps<br />
100<br />
85<br />
C. salinae<br />
62<br />
64<br />
79<br />
98<br />
EXF-780<br />
EXF-321<br />
EXF-946<br />
<strong>CBS</strong> 170.54<br />
ATCC 76499<br />
ATCC 66669<br />
EXF-699<br />
EXF-710<br />
EXF-697<br />
EXF-711<br />
<strong>CBS</strong> 125.80<br />
EXF-563<br />
EXF-452<br />
EXF-371<br />
Moricca et al.<br />
ATCC 38027<br />
EXF-714<br />
EXF-332<br />
EXF-391<br />
EXF-326<br />
87 <strong>CBS</strong> 109868<br />
dH 13833<br />
<strong>CBS</strong> 601.84<br />
dH 11736<br />
<strong>CBS</strong> 101880<br />
<strong>CBS</strong> 189.54<br />
dH 12459<br />
100 EXF-449<br />
<strong>CBS</strong> 452.71<br />
EXF-397<br />
EXF-466<br />
<strong>CBS</strong> 119417<br />
EXF-471<br />
EXF-739<br />
<strong>CBS</strong> 102045<br />
<strong>CBS</strong> 109.14<br />
EXF-455<br />
EXF-738<br />
EXF-962<br />
EXF-1061<br />
<strong>CBS</strong> 193.54<br />
EXF-458<br />
<strong>CBS</strong> 300.96 <strong>Cladosporium</strong> sp.<br />
EXF-732<br />
80<br />
90<br />
62<br />
100<br />
CPC 11683<br />
EXF-718<br />
EXF-720<br />
EXF-727<br />
72<br />
63<br />
79<br />
99<br />
EXF-977<br />
EXF-1072<br />
dH 12991<br />
dH 12862<br />
dH 13911<br />
EXF-564 C. halotolerans<br />
EXF-380<br />
EXF-703<br />
EXF-646<br />
EXF-228<br />
EXF-572<br />
<strong>CBS</strong> 280.49 “Mycosphaerella” hyperici<br />
<strong>CBS</strong> 177.71 C. bruhnei<br />
<strong>CBS</strong> 812.71<br />
EXF-595 <strong>Cladosporium</strong> sp.<br />
EXF-454 C. ramotenellum<br />
ATCC 66670 Davidiella tassiana<br />
EXF-333<br />
EXF-382<br />
EXF-334<br />
EXF-343 C. subinflatum<br />
C. spinulosum<br />
C. cladosporioides<br />
C. oxysporum<br />
C. tenuissimum<br />
C. sphaerospermum<br />
C. dominicanum<br />
C. psychrotolerans<br />
C. langeronii<br />
C. fusiforme<br />
C. velox<br />
Fig. 2. One of 600 equally most parsimonious <strong>and</strong> equally looking phylogenetic trees based on a heuristic tree search using aligned sequences of the internal transcribed<br />
spacer regions 1 <strong>and</strong> 2 <strong>and</strong> the 5.8S rDNA. <strong>The</strong> tree was r<strong>and</strong>omly selected. Support based on 10 000 replicates of a fast step-wise addition bootstrap analysis is indicated near<br />
the branches. Trees were rooted with the strains of <strong>Cladosporium</strong> salinae. Most monophyletic species clades received high, but some deeper branches moderate, bootstrap<br />
support (CI = 0.804, RI = 0.975, PIC = 68).<br />
RESULTS<br />
Descriptive statistical parameters of phylogenetic analyses <strong>and</strong><br />
calculated tree scores for each analysed sequence locus are<br />
summarised in Table 2. Mainly reference material such as extype<br />
or ex-neotype strains was analysed on the level of SSU<br />
rDNA sequences. Downloaded <strong>and</strong> newly generated SSU rDNA<br />
sequences of members of <strong>Cladosporium</strong> s. str. were compared<br />
with related taxa of the Mycosphaerellaceae, Dothioraceae <strong>and</strong><br />
Dothideaceae. <strong>The</strong> somewhat more distantly related Fusicladium<br />
effusum (Venturiaceae) (Braun et al. 2003: Fig. 2) was selected<br />
as outgroup. Anungitopsis amoena R.F. Castañeda & Dugan (now<br />
placed in Fusicladium Bonord., see Crous et al. 2007b), also a<br />
member of the Venturiaceae, was included in the analyses. All taxa<br />
included in the SSU rDNA analysis belong to the Dothideomycetes<br />
www.studiesinmycology.org<br />
(Schoch et al. 2006), within which the ingroup is represented by<br />
the orders Capnodiales (Davidiellaceae, Mycosphaerellaceae,<br />
Teratosphaeriaceae) <strong>and</strong> Dothideales (Dothioraceae, Dothideaceae)<br />
(see also Schoch et al. 2006). <strong>The</strong> <strong>genus</strong> <strong>Cladosporium</strong>, of which<br />
some species are linked to Davidiella Crous & U. Braun teleomorphs<br />
(Braun et al. 2003), forms a statistically strongly supported<br />
monophyletic group (Davidiellaceae). It also accommodates species<br />
newly described in this paper, namely, C. halotolerans Zalar, de<br />
Hoog & Gunde-Cimerman, C. fusiforme Zalar, de Hoog & Gunde-<br />
Cimerman, C. dominicanum Zalar, de Hoog & Gunde-Cimerman, C.<br />
salinae, C. psychrotolerans Zalar, de Hoog & Gunde-Cimerman, C.<br />
velox Zalar, de Hoog & Gunde-Cimerman <strong>and</strong> C. spinulosum Zalar,<br />
de Hoog & Gunde-Cimerman (Fig. 1). A sister group relationship<br />
of <strong>Cladosporium</strong> s. str. with a clade of taxa characterised,<br />
among others, by Mycosphaerella Johanson teleomorphs,<br />
containing various anamorphic genera such as Septoria Sacc.,<br />
163
Zalar et al.<br />
97 EXF-322<br />
<strong>CBS</strong> 119413<br />
EXF-604<br />
10 steps<br />
89<br />
C. salinae<br />
96<br />
61<br />
87<br />
93<br />
91<br />
98<br />
98 <strong>CBS</strong> 601.84<br />
<strong>CBS</strong> 101880<br />
dH 12459<br />
<strong>CBS</strong> 189.54<br />
<strong>CBS</strong> 109868<br />
100<br />
dH 13833<br />
dH 11736<br />
EXF-332<br />
EXF-326<br />
EXF-391<br />
EXF-714<br />
100 <strong>CBS</strong> 177.71 C. bruhnei<br />
ATCC 66670 Davidiella tassiana<br />
<strong>CBS</strong> 170.54 C. cladosporioides<br />
EXF-343 C. subinflatum<br />
EXF-382<br />
C. spinulosum<br />
EXF-334<br />
ATCC 66669<br />
<strong>CBS</strong> 125.80<br />
<strong>CBS</strong> 119417 C. velox<br />
C. oxysporum<br />
EXF-572<br />
EXF-564<br />
EXF-1072<br />
EXF-380<br />
C. halotolerans<br />
EXF-703<br />
EXF-228<br />
EXF-646<br />
EXF-977<br />
<strong>CBS</strong> 280.49 “Mycosphaerella” hyperici<br />
98<br />
dH 12862<br />
100<br />
63<br />
84<br />
100<br />
95 93 100<br />
100<br />
dH 12941<br />
dH 13911<br />
dH 12991<br />
EXF-727<br />
90 8862<br />
100<br />
99<br />
EXF-458<br />
98<br />
EXF-455<br />
<strong>CBS</strong> 102045<br />
EXF-1061<br />
<strong>CBS</strong> 193.54<br />
EXF-738<br />
<strong>CBS</strong> 109.14<br />
EXF-739<br />
EXF-962<br />
EXF-397<br />
EXF-449<br />
<strong>CBS</strong> 452.71<br />
78<br />
99<br />
EXF-720<br />
EXF-732<br />
EXF-718<br />
CPC 11683<br />
EXF-696<br />
C. halotolerans<br />
C. sphaerospermum<br />
C. fusiforme<br />
C. langeronii<br />
C. psychrotolerans<br />
C. dominicanum<br />
Fig. 3. One of 90 equally most parsimonious <strong>and</strong> equally looking phylogenetic trees based on a heuristic tree search using aligned exons <strong>and</strong> introns of a part of the β-tubulin<br />
gene. <strong>The</strong> tree was r<strong>and</strong>omly selected. Support based on 10 000 replicates of a fast step-wise addition bootstrap analysis is indicated near the branches. Trees were rooted with<br />
the strains of <strong>Cladosporium</strong> salinae. Most monophyletic species clades received high, but deeper branches weak or no, bootstrap support (CI = 0.538, RI = 0.883, PIC = 220).<br />
Ramularia Unger, Cercospora Fresen., Pseudocercospora Speg.,<br />
“Trimmatostroma” Corda (now Catenulostroma Crous & U. Braun)<br />
(see Crous et al. 2004, 2007a – this volume) <strong>and</strong> the somewhat<br />
cladosporium-like <strong>genus</strong> Devriesia Seifert & N.L. Nick. (Seifert et al.<br />
2004), was statistically only moderately supported (Fig. 1), whereas<br />
in an analogous analysis by Braun et al. (2003: Fig. 2) it was highly<br />
supported. <strong>The</strong>se data also support the conclusion by Braun et al.<br />
(2003) <strong>and</strong> Crous et al. (2006) that <strong>Cladosporium</strong> is not a member<br />
of the distantly related Herpotrichiellaceae (Chaetothyriomycetes),<br />
which is also rich in cladosporium-like taxa (Crous et al. 2006).<br />
None of the fungi isolated from hypersaline environments belonged<br />
to the Herpotrichiellaceae. <strong>The</strong> SSU rDNA sequences do not<br />
resolve a phylogenetic structure within <strong>Cladosporium</strong> s. str. Only<br />
a moderately supported clade comprising C. halotolerans, C.<br />
dominicanum, C. velox, C. sphaerospermum <strong>and</strong> C. fusiforme is<br />
somewhat distinguished from a statistically unsupported clade with<br />
C. herbarum (Pers. : Fr.) Link, C. cladosporioides (Fresen.) G.A.<br />
de Vries, C. oxysporum Berk. & Broome, C. spinulosum, <strong>and</strong> C.<br />
psychrotolerans, etc. Because C. salinae appeared most distinct<br />
within the <strong>genus</strong> <strong>Cladosporium</strong> in analyses of the SSU rDNA (Fig.<br />
1), it was used as outgroup in analyses of the ITS rDNA <strong>and</strong> the<br />
β-tubulin <strong>and</strong> actin genes.<br />
Analyses of the more variable ITS rDNA <strong>and</strong> partial β-tubulin<br />
<strong>and</strong> actin gene introns <strong>and</strong> exons supported the species clades of<br />
C. halotolerans, C. dominicanum, C. sphaerospermum, C. fusiforme<br />
<strong>and</strong> C. velox (Figs 2–4), of which C. velox was distinguished in<br />
the β-tubulin tree by a particular long terminal branch of the only<br />
sequenced strain (Fig. 3). <strong>Cladosporium</strong> salinae also clustered as a<br />
well-supported species clade in preliminary analyses using various<br />
Mycosphaerella species as outgroup (not shown). All strains of C.<br />
langeronii (Fonseca, Leão & Nogueira) Vuill. are particularly well<br />
distinguishable from other <strong>Cladosporium</strong> species by strikingly slow-<br />
164
<strong>Cladosporium</strong> sphaerospermum species complex<br />
EXF-322<br />
<strong>CBS</strong> 119413<br />
EXF-604<br />
60<br />
100<br />
C. salinae<br />
<strong>CBS</strong> 189.54<br />
71 <strong>CBS</strong> 601.84<br />
<strong>CBS</strong> 101880<br />
dH 12459<br />
dH 13833<br />
96 <strong>CBS</strong> 109868<br />
dH 11736<br />
62 EXF-332<br />
EXF-391<br />
77 EXF-714<br />
<strong>CBS</strong> 177.71 C. bruhnei<br />
66 89 ATCC 66670 Davidiella tassiana<br />
67 EXF-343 C. subinflatum<br />
85<br />
C. psychrotolerans<br />
87 EXF-696<br />
EXF-718<br />
100<br />
EXF-732<br />
C. dominicanum<br />
CPC 11683<br />
EXF-598<br />
100<br />
<strong>CBS</strong> 102045<br />
EXF-458<br />
EXF-1069<br />
EXF-455<br />
72<br />
EXF-1061<br />
77EXF-739<br />
76<br />
EXF-962<br />
C. sphaerospermum<br />
81 EXF-738<br />
76 <strong>CBS</strong> 109.14<br />
<strong>CBS</strong> 193.54<br />
<strong>CBS</strong> 300.96 <strong>Cladosporium</strong> sp.<br />
EXF-449<br />
100 90<br />
EXF-397 C. fusiforme<br />
<strong>CBS</strong> 452.71<br />
100<br />
EXF-466<br />
EXF-471<br />
<strong>CBS</strong> 119417<br />
C. velox<br />
84 98 EXF-334<br />
EXF-382<br />
C. spinulosum<br />
<strong>CBS</strong> 170.54 C. cladosporioides<br />
89<br />
<strong>CBS</strong> 125.80<br />
ATCC 66669<br />
C. oxysporum<br />
dH 12862<br />
dH 13911<br />
EXF-646<br />
EXF-703<br />
EXF-228<br />
EXF-1072 C. halotolerans<br />
EXF-572<br />
EXF-977<br />
EXF-564<br />
EXF-380<br />
<strong>CBS</strong> 280.49 “Mycosphaerella” hyperici<br />
C. langeronii<br />
10 steps<br />
Fig. 4. One of 32 equally most parsimonious <strong>and</strong> equally looking phylogenetic trees based on a heuristic tree search using aligned exons <strong>and</strong> introns of the partial actin gene.<br />
<strong>The</strong> tree was r<strong>and</strong>omly selected. Support based on 10 000 replicates of a fast step-wise addition bootstrap analysis is indicated near the branches. Trees were rooted with<br />
the strains of <strong>Cladosporium</strong> salinae. Most monophyletic species clades received high, but deeper branches weak or no, bootstrap support (CI = 0.586, RI = 0.885, PIC = 103).<br />
growing colonies at all tested temperatures <strong>and</strong> relatively large,<br />
oblong conidia. However, phylogenetic analyses of the β-tubulin <strong>and</strong><br />
actin gene indicate that C. langeronii presents two cryptic species<br />
(Figs 3–4). <strong>The</strong> species clade of C. psychrotolerans is moderately<br />
supported in analyses of the actin gene but highly by means of<br />
the β-tubulin gene. It is evident from all three analyses (Figs 2–<br />
4) that C. langeronii <strong>and</strong> C. psychrotolerans are closely related<br />
species. <strong>The</strong> species node of <strong>Cladosporium</strong> spinulosum, which is<br />
morphologically clearly distinguished from all other species by its<br />
conspicuous ornamentation consisting of digitate projections (Fig.<br />
5), is supported by β-tubulin (Fig. 3) <strong>and</strong> actin (Fig. 4) sequence<br />
data but not by those of the ITS rDNA (Fig. 2). Analyses of all loci,<br />
however, indicate that it is a member of the C. herbarum complex.<br />
<strong>The</strong> analyses of sequences of the ITS <strong>and</strong> the β-tubulin<br />
<strong>and</strong> actin gene introns <strong>and</strong> exons (Figs 2–4) do not allow the<br />
full elucidation of phylogenetic relationships among these<br />
<strong>Cladosporium</strong> species. Statistical support of the interior tree<br />
branches resulting from analyses of the β-tubulin <strong>and</strong> actin genes<br />
is low (bootstrap values mostly < 50 %). While the sister group<br />
relationship of C. sphaerospermum <strong>and</strong> C. fusiforme is highly<br />
supported in the analysis based on the β-tubulin gene, analysis<br />
of the ITS rDNA indicate that these two species are unrelated, <strong>and</strong><br />
that C. sphaerospermum is closely related to C. dominicanum. It is<br />
clear from the data that the species morphologically resembling C.<br />
sphaerospermum are not phylogenetically closely related <strong>and</strong> that<br />
the data we present here do not allow their classification in natural<br />
subgroups of the <strong>genus</strong> <strong>Cladosporium</strong>. Only C. spinulosum was<br />
placed in all analyses among species of the C. herbarum complex<br />
www.studiesinmycology.org<br />
165
Zalar et al.<br />
Fig. 5. Conidial scars <strong>and</strong> surface ornamentation of ramoconidia <strong>and</strong> conidia (SEM). A. C. dominicanum (strain EXF-732). B. C. fusiforme (strain EXF-449). C. C. halotolerans<br />
(strain EXF-572). D. C. langeronii (strain <strong>CBS</strong> 189.54). E. C. psychrotolerans (strain EXF-391). F. C. salinae (strain EXF-335 = <strong>CBS</strong> 119413). G. C. sphaerospermum (strain<br />
<strong>CBS</strong> 193.54). H. C. spinulosum (strain EXF-334). I. C. velox (strain <strong>CBS</strong> 119417). Scale bars = 5 µm. (Photos: K. Drašlar).<br />
<strong>and</strong> all analyses supported close relatedness of C. langeronii <strong>and</strong><br />
C. psychrotolerans.<br />
<strong>The</strong> majority of species described here have slightly<br />
ornamented conidia ranging from minutely verruculose (C.<br />
fusiforme, C. langeronii, C. psychrotolerans, C. sphaerospermum,<br />
C. velox) to verrucose (C. halotolerans) (Fig. 5). <strong>The</strong> verrucose<br />
conidia of C. halotolerans can be recognised also under the<br />
light microscope <strong>and</strong> used as a distinguishing character. Almost<br />
smooth to minutely verruculose conidia are encountered in C.<br />
dominicanum <strong>and</strong> C. salinae (Fig. 5). <strong>Cladosporium</strong> spinulosum,<br />
a member of the C. herbarum species complex, has conidia with<br />
a digitate ornamentation that can appear spinulose under the light<br />
microscope; however, when using the SEM it became clear that its<br />
projections have parallel sides <strong>and</strong> a blunt end (Fig. 5).<br />
DISCUSSION<br />
<strong>The</strong> <strong>genus</strong> <strong>Cladosporium</strong> was established by Link (1816) who<br />
originally included four species, of which C. herbarum is the<br />
type species of the <strong>genus</strong> (Clements & Shear 1931). In 1950,<br />
von Arx reported a teleomorph connection for this species with<br />
Mycosphaerella tassiana (De Not.) Johanson. Based on SSU<br />
rDNA data the majority of Mycosphaerella species, including<br />
the type species of the <strong>genus</strong>, M. punctiformis (Pers.) Starbäck,<br />
clustered within the Mycosphaerellaceae, a family separated<br />
from M. tassiana (Braun et al. 2003). <strong>The</strong>refore, Mycosphaerella<br />
tassiana was reclassified as Davidiella tassiana (De Not.) Crous<br />
& U. Braun, the type of the new <strong>genus</strong> Davidiella. All anamorphs<br />
with a cladosporium- <strong>and</strong> heterosporium-like appearance <strong>and</strong> with<br />
a supposed Dothideomycetes relationship were maintained under<br />
the anamorph name <strong>Cladosporium</strong>, morphologically characterised<br />
by scars with a protuberant hilum consisting of a central dome<br />
surrounded by a raised rim (David 1997).<br />
<strong>The</strong> concept of distinguishing ramoconidia from secondary<br />
ramoconidia has been adopted from Schubert et al. (2007). In<br />
the species described here, ramoconidia have been observed<br />
often in C. sphaerospermum, sometimes in C. psychrotolerans,<br />
C. langeronii <strong>and</strong> C. spinulosum, <strong>and</strong> only sporadically in all other<br />
species. <strong>The</strong>refore, ramoconidia can be seen as important for<br />
distinguishing species although sometimes, they can be observed<br />
only with difficulty. When using ramoconidia as a diagnostic<br />
criterion, colonies only from SNA <strong>and</strong> not older than 7 d should be<br />
taken into account.<br />
<strong>Cladosporium</strong> sphaerospermum was described by Penzig<br />
(1882) from decaying Citrus leaves <strong>and</strong> branches in Italy. He<br />
described C. sphaerospermum as a species with (i) branched,<br />
septate <strong>and</strong> dark conidiophores having a length of 150–300 µm <strong>and</strong><br />
a width of the main conidiophore stipe of 3.5–4 µm, (ii) spherical to<br />
ellipsoid, acrogenously formed conidia of 3.4–4 µm diam, <strong>and</strong> (iii)<br />
ramoconidia of 6–14 × 3.5–4 µm. Penzig’s original material is not<br />
known to be preserved. Later, a culture derived from <strong>CBS</strong> 193.54,<br />
originating from a human nail, was accepted as typical of C.<br />
sphaerospermum. However, de Vries (1952), incorrectly cited it as<br />
“lectotype”, <strong>and</strong> thus the same specimen is designated as neotype<br />
in this study (see below), with the derived culture (<strong>CBS</strong> 193.54)<br />
166
<strong>Cladosporium</strong> sphaerospermum species complex<br />
used as ex-neotype strain. Numerous strains with identical or very<br />
<strong>similar</strong> ITS rDNA sequences as <strong>CBS</strong> 193.54 were isolated from<br />
hypersaline water or organic substrata including plants or walls of<br />
bathrooms. It is not clear yet whether surfaces in bathrooms <strong>and</strong> of<br />
plants, colonised by C. sphaerospermum, can have a <strong>similar</strong> low<br />
water activity as salterns. In our experiments, the strains of this<br />
species, however, grew under in vitro conditions at a water activity<br />
of up to 0.860, while Hocking et al. (1994) <strong>and</strong> Aihara et al. (2002)<br />
reported that it can grow even at 0.815. <strong>The</strong>refore, we consider<br />
C. sphaerospermum as halo- or osmotolerant. Hardly any reports<br />
are available unambiguously proving that C. sphaerospermum is<br />
a human pathogen. It is therefore possible that <strong>CBS</strong> 193.54 was<br />
not involved in any disease process but rather occurred as a<br />
contaminant on dry nail material. <strong>Cladosporium</strong> sphaerospermum<br />
is a phylogenetically well-delineated species (Figs 2–4).<br />
Strains of C. halotolerans were isolated sporadically from<br />
substrata such as peanut cell suspension, tissue culture, bathroom<br />
walls <strong>and</strong> as culture contaminants. This surprising heterogeneity<br />
of substrata suggests that C. halotolerans is distributed by air <strong>and</strong><br />
that it can colonise whatever substrata available, although it may<br />
have its natural niche elsewhere. We have recurrently isolated it<br />
from hypersaline water of salterns <strong>and</strong> other saline environments<br />
<strong>and</strong> it was also detected with molecular methods (but not isolated)<br />
from skin of a salt water dolphin. <strong>The</strong>re are only few reports of<br />
this species from plants (Table 1). It is therefore possible that C.<br />
halotolerans is a species closely linked to salty or hypersaline<br />
environments although additional sampling is necessary to prove<br />
that. <strong>Cladosporium</strong> halotolerans is morphologically recognisable<br />
by relatively oblong to spherical, coarsely rough-walled conidia.<br />
<strong>The</strong> ITS rDNA sequence of a fungus in the skin of a bottlenose<br />
dolphin, suffering from lobomycosis, is identical to the sequences<br />
of C. halotolerans. This sequence was deposited as Lacazia<br />
loboi Taborda, V.A. Taborda & McGinnis (GenBank AF035674) by<br />
Haubold et al. (1998), who apparently concluded wrongly that a<br />
fungus with a cladosporium-like ITS rDNA sequence <strong>similar</strong> to that<br />
of C. halotolerans can be the agent of lobomycosis. Later, Herr<br />
et al. (2001) showed that Lacazia loboi phylogenetically belongs<br />
to the Onygenales on the basis of amplified SSU rDNA <strong>and</strong> chitin<br />
synthase-2 gene sequences generated from tissue lesions. By<br />
this, they confirmed an earlier supposition by Lacaz (1996) who<br />
reclassified the organism as Paracoccidioides loboi O.M. Fonseca<br />
& Silva Lacaz (Onygenales). It is therefore possible that C.<br />
halotolerans was not the main etiologic agent for the lobomycosis<br />
<strong>and</strong> it was colonising the affected dolphin skin secondarily while<br />
inhabiting other seawater habitats.<br />
<strong>Cladosporium</strong> langeronii <strong>and</strong> C. psychrotolerans are closely<br />
related but C. langeronii is particularly well distinguishable from<br />
all other <strong>Cladosporium</strong> species by its slow growing colonies<br />
(1–7 mm diam / 14 d) <strong>and</strong> relatively large conidia (4–5.5 × 3–4<br />
μm). <strong>Cladosporium</strong> psychrotolerans has smaller conidia (3–4 ×<br />
2.5–3 μm) but a <strong>similar</strong> length : width ratio <strong>and</strong> faster exp<strong>and</strong>ing<br />
colonies (8–18 mm diam / 14 d). <strong>Cladosporium</strong> langeronii is most<br />
likely a complex of at least two species. Strains isolated from the<br />
Arctic <strong>and</strong> the Antarctic may need to be distinguished from C.<br />
langeronii s. str. on species level. This inference is particularly<br />
supported by analyses of the β-tubulin <strong>and</strong> actin genes (Figs 3–4).<br />
<strong>Cladosporium</strong> langeronii s. str., represented by an authentic strain<br />
of Hormodendrum langeronii Fonseca, Leão & Nogueira, <strong>CBS</strong><br />
189.54 (Trejos 1954), has been isolated from a variety of substrata<br />
but is tolerating only up to 10 % NaCl. It was originally described<br />
by da Fonseca et al. (1927a, b) <strong>and</strong> subsequently reclassified<br />
as <strong>Cladosporium</strong> langeronii by Vuillemin (1931). <strong>The</strong> authentic<br />
www.studiesinmycology.org<br />
strain derived from an ulcerating nodular lesion on the arm of a<br />
human patient. Because other strains of this species are ubiquitous<br />
saprobes originating from various substrata, we suspect that C.<br />
langeronii is not an important human pathogen. <strong>Cladosporium</strong><br />
psychrotolerans has been isolated from hypersaline environments<br />
only, <strong>and</strong> tolerates up to 20 % NaCl in culture media.<br />
In general, the human- or animal-pathogenic role of the C.<br />
sphaerospermum-like species described here seems to be limited.<br />
It is possible that pathogenic species of Cladophialophora Sacc.<br />
have been misidentified as C. sphaerospermum or as other<br />
species of <strong>Cladosporium</strong> (de Hoog et al. 2000). Alternatively, true<br />
<strong>Cladosporium</strong> species isolated as clinical strains could have been<br />
secondary colonisers since they are able to dwell on surfaces poor<br />
in nutrients, possibly in an inconspicuous dormant phase <strong>and</strong> may<br />
then be practically invisible. More likely, they could be air-borne<br />
contaminations of lesions, affected nails etc. (Summerbell et al.<br />
2005) or are perhaps disseminated by insufficiently sterilised medical<br />
devices, as melanised fungi can be quite resistant to disinfectants<br />
(Phillips et al. 1992). <strong>The</strong>y can easily be isolated <strong>and</strong> rapidly<br />
become preponderant at isolation <strong>and</strong> thus difficult to exclude as<br />
etiologic agents of a disease. For example, in 2002, a case report<br />
on an intrabronchial lesion by C. sphaerospermum in a healthy,<br />
non-asthmatic woman was described (Yano et al. 2002), but we<br />
judge the identification of the causal agent to remain uncertain, as<br />
it was based on morphology alone <strong>and</strong> no culture is available. <strong>The</strong><br />
present authors have the opinion that all clinical cases ascribed to<br />
<strong>Cladosporium</strong> species need careful re-examination.<br />
General characteristics <strong>and</strong> description of <strong>Cladosporium</strong><br />
sphaerospermum-like species<br />
<strong>The</strong> present paper focuses on <strong>Cladosporium</strong> strains isolated from<br />
hypersaline environments. Comparison of data from deliberate<br />
sampling <strong>and</strong> analysis of reference strains from culture collections<br />
inevitably leads to statistical bias, <strong>and</strong> therefore a balanced<br />
interpretation of ecological preferences of the species presented is<br />
impossible. Nevertheless, some species appeared to be consistent<br />
in their choice of habitat, <strong>and</strong> for this reason we summarise<br />
isolation data for all species described. Strains belonging to a single<br />
molecular clade proved to have <strong>similar</strong> cultural characteristics <strong>and</strong><br />
microscopic morphology. Although within most of the species there<br />
was some molecular variation noted (particularly when intron-rich<br />
genes were analysed), some consistent phenetic trends could be<br />
observed.<br />
Conidiophores of all C. sphaerospermum-like species lack<br />
nodose inflations (McKemy & Morgan-Jones 1991). <strong>The</strong>y are<br />
usually ascending <strong>and</strong> can sometimes be poorly differentiated from<br />
their supporting hyphae. Though the initiation point of conidiophore<br />
stipes could sometimes be determined only approximately,<br />
their lengths were in some cases useful for distinguishing<br />
morphologically <strong>similar</strong> species when observed in slide cultures.<br />
Generally, the branched part of a conidiophore forms a complex<br />
tree-like structure. <strong>The</strong> number <strong>and</strong> orientation of early formed<br />
secondary ramoconidia, however, determines whether it is<br />
rotationally symmetric or unilateral.<br />
<strong>The</strong> variability in ITS rDNA sequences observed in all C.<br />
sphaerospermum-like species (about 10 %) spans the variation<br />
observed in all members of the <strong>genus</strong> <strong>Cladosporium</strong> sequenced<br />
to date. Thus, the C. sphaerospermum-like species described here<br />
may not present a single monophyletic group but may belong to<br />
various species complexes within <strong>Cladosporium</strong>. Verifying existing<br />
literature with sequence data of these species (Wirsel et al. 2002,<br />
Park et al. 2004), we noticed that names of the common saprobes<br />
seem to be distributed nearly at r<strong>and</strong>om over phylogenetic trees.<br />
167
Zalar et al.<br />
For most commonly used names, no type material is available for<br />
sequencing. Also verification of published reports is difficult without<br />
available voucher strains.<br />
<strong>Cladosporium</strong> cladosporioides was incorrectly lectotypified<br />
based on <strong>CBS</strong> 170.54 (de Vries 1952), which Bisby considered a<br />
st<strong>and</strong>ard culture of C. herbarum. <strong>The</strong> C. cladosporioides species<br />
complex requires revision, <strong>and</strong> will form the basis of a future study.<br />
<strong>Cladosporium</strong> herbarum is maintained as a dried specimen in the<br />
Leiden herbarium; Prasil & de Hoog (1988) selected <strong>CBS</strong> 177.71<br />
as a representative living strain. Strains, earlier accepted as living<br />
representatives of C. herbarum, <strong>CBS</strong> 177.71 <strong>and</strong> <strong>CBS</strong> 812.71<br />
(Prasil & de Hoog 1988, Wirsel et al. 2002) <strong>and</strong> ATCC 66670<br />
(Braun et al. 2003, as Davidiella tassiana) have been re-identified<br />
as C. bruhnei Linder by Schubert et al. (2007 – this volume). Ho<br />
et al. (1999) used strain ATCC 38027 as a representative of C.<br />
tenuissimum Cooke <strong>and</strong> this strain has identical ITS sequences as<br />
the non-deposited C. tenuissimum material used by Moricca et al.<br />
(1999). We tentatively accept this concept although we could not<br />
include ATCC 38027 in our analyses. <strong>The</strong> ITS sequence of strain<br />
<strong>CBS</strong> 125.80, identified by Wirsel et al. (2002) as C. oxysporum,<br />
is identical to the sequence of ATCC 38027. Strain ATCC 76499,<br />
published by Ho et al. (1999) as C. oxysporum, appears to be<br />
identical to a number of currently unidentified <strong>Cladosporium</strong> strains<br />
from Slovenian salterns that compose a cluster separate from all<br />
remaining species. Strains of this cluster, represented in Fig. 2 by<br />
strain ATCC 76499, morphologically resemble C. oxysporum.<br />
Strain <strong>CBS</strong> 300.96 has not been identified to species level<br />
in the present study. It clusters outside the species clade of<br />
C. sphaerospermum, with the latter being its nearest relative.<br />
<strong>CBS</strong> 300.96 differs from C. sphaerospermum by having smaller<br />
structures: conidiophore stipes [(5–)20–80(–150) × (2–)2.5–3(–4)<br />
μm], 0–1 septate ramoconidia [(13–)19–27(–32) × 2–2.5 μm],<br />
conidia [(2.5–)3–3.5(–4) × (2–)2–2.5(–3) μm] <strong>and</strong> secondary<br />
ramoconidia [(5–)9–18(–30) × (2–)2.5–2.5(–3) μm]. However,<br />
based on a single isolate, we currently refrain from describing it as<br />
a new species.<br />
Key to species treated in this study<br />
Macro-morphological characters used in the key are from colonies grown on PDA <strong>and</strong> MEA 14 d at 25 °C, if not stated otherwise; microscopical<br />
characters are from SNA slide cultures grown for 7 d at 25 °C.<br />
1. Conidial ornamentation conspicuously echinulate / digitate because of up to 1.3 µm long projections that have more or less parallel sides<br />
............................................................................................................................................................................................. C. spinulosum<br />
1. Conidial ornamentation verruculose to verrucose or smooth, not conspicuously echinulate or digitate .................................................... 2<br />
2. Conidiophores micronematous, poorly differentiated, once or several times geniculate-sinuous, short, up to 60 µm long; terminal conidia<br />
obovoid ....................................................................................................................................................................................... C. salinae<br />
2. Conidiophores micro- or macronematous, not geniculate or only slightly so, usually up to 100 µm or 220 µm long or even longer; terminal<br />
conidia globose, subglobose to ovoid or fusiform ...................................................................................................................................... 3<br />
3. Secondary ramoconidia 0–3(–4)-septate; septa of conidiophores <strong>and</strong> conidia darkened <strong>and</strong> thickened .................................................. 4<br />
3. Secondary ramoconidia 0–1(–2)-septate; septa neither darkened nor thickened ..................................................................................... 5<br />
4. Conidiophores (5–)10–50(–300) × (2–)2.5–3(–5.5) μm; terminal conidia (2–)3–4(–6) × (2–)2.5–3(–5) μm; secondary ramoconidia (5–)7–<br />
12(–37.5) × (2–)2.5–3(–6.5) μm; ramoconidia sporadically formed ................................................................................... C. halotolerans<br />
4. Conidiophores mostly longer <strong>and</strong> somewhat wider, (10–)45–130(–300) × (2.5–)3–4(–6) μm; terminal conidia mostly wider, (2.5–)3–4(–7)<br />
× (2–)3–3.5(–4.5) μm; secondary ramoconidia (4–)8.5–16(–37.5) × (2–)3–3.5(–5) μm; ramoconidia often formed, up to 40 µm long, with<br />
up to 5 septa ............................................................................................................................................................. C. sphaerospermum<br />
5. Terminal conidia usually fusiform ............................................................................................................................................ C. fusiforme<br />
5. Terminal conidia globose, subglobose or ovoid ......................................................................................................................................... 6<br />
6. Conidia <strong>and</strong> secondary ramoconidia irregularly verruculose to sometimes loosely verrucose; radial growth on PDA at 25 °C after 14 d<br />
typically less than 5 mm ......................................................................................................................................................... C. langeronii<br />
6. Conidia <strong>and</strong> secondary ramoconidia smooth to minutely verruculose; radial growth on PDA at 25 °C after 14 d typically more than 10 mm<br />
.................................................................................................................................................................................................................... 7<br />
7. Conidiophores (3–)3.5–4(–7.5) μm wide, thick-walled; conidiogenous loci <strong>and</strong> conidial hila 0.5–2 μm diam; ramoconidia sometimes<br />
formed with a broadly truncate, up to 2 µm wide non-cladosporioid base; no growth observed after 14 d at 30 °C on MEA<br />
..................................................................................................................................................................................... C. psychrotolerans<br />
7. Conidiophores mostly narrower, 2–4 μm wide, only with slightly thickened walls; conidiogenous loci <strong>and</strong> conidial hila narrower, 0.5–1.5<br />
μm diam; ramoconidia rarely formed; colony showing at least weak growth after 14 d at 30 °C on MEA ................................................. 8<br />
8. Secondary ramoconidia (4–)6.5–13(–24.5) μm long; no visible colony growth after 14 d at 10 °C on MEA.................... C. dominicanum<br />
8. Secondary ramoconidia mostly longer, (3.5–)5.5–19(–42) μm; radial growth of colonies after 14 d at 10 °C on MEA more than 5 mm<br />
........................................................................................................................................................................................................ C. velox<br />
168
<strong>Cladosporium</strong> sphaerospermum species complex<br />
Description of <strong>Cladosporium</strong> species<br />
<strong>Cladosporium</strong> dominicanum Zalar, de Hoog & Gunde-Cimerman,<br />
sp. nov. MycoBank MB510995. Fig. 6.<br />
Etymology: Refers to the the Dominican Republic, where most<br />
strains were encountered.<br />
Conidiophora lateralia vel terminalia ex hyphis rectis oriunda; stipes longitudine<br />
variabili, (5–)10–100(–200) × (1.5–)2–2.5(–3.5) μm, olivaceo-brunneus, levis vel<br />
leniter verruculosus, tenuitunicatus, plerumque unicellularis, simplex vel ramosus.<br />
Conidiorum catenae undique divergentes, ad 8 conidia in parte continua continentes.<br />
Cellulae conidiogenae indistinctae. Conidia levia vel leniter verruculosa, dilute<br />
brunnea, unicellularia, plerumque breviter ovoidea, utrinque angustata, (2.5–)<br />
3–3.5(–5.5) × (2–)2–2.5(–2.5) μm, long.: lat. 1.4–1.6; ramoconidia secundaria<br />
cylindrica vel quasi globosa, 0–1-septata, (4–)6.5–13(–24.5) × (2–)2.5–3(–4.5) μm,<br />
ad 4 cicatrices terminales ferentia; cicatrices inspissatae, protuberantes, 0.5–1.2 μm<br />
diam. Hyphae vagina polysaccharidica carentes.<br />
Mycelium without extracellular polysaccharide-like material.<br />
Conidiophores arising laterally <strong>and</strong> terminally on erect hyphae,<br />
micronematous <strong>and</strong> semimacronematous, stipes of variable length,<br />
(5–)10–100(–200) × (1.5–)2–2.5(–3.5) μm, olivaceous-brown,<br />
smooth to minutely verruculose, thin-walled, almost non-septate,<br />
unbranched or branched. Conidial chains branching in all directions,<br />
up to eight conidia in the unbranched parts. Conidiogenous cells<br />
undifferentiated. Ramoconidia rarely formed. Conidia smooth<br />
to minutely verruculose, subhyaline to light brown, non-septate,<br />
usually short-ovoid, narrower at both ends, length : width ratio =<br />
1.4–1.6; (2.5–)3–3.5(–5.5) × (2–)2–2.5(–2.5) μm [av. (± SD) 3.4<br />
(± 0.6) × 2.2 (± 0.2)]; secondary ramoconidia cylindrical to almost<br />
spherical, 0–1-septate, (4–)6.5–13(–24.5) × (2–)2.5–3(–4.5) μm<br />
[av. (± SD) 10.3 (± 5.2) × 2.7 (± 0.6)], with up to four distal scars.<br />
Conidiogenous scars thickened <strong>and</strong> conspicuous, protuberant,<br />
0.5–1.2 μm diam.<br />
Cultural characteristics: Colonies on PDA reaching 18–36 mm<br />
diam, olive-yellow (2D6), hairy granular, flat or slightly furrowed,<br />
with flat margin. Droplets of light reseda-green (2E6) exudate<br />
sometimes present. Reverse dark green to black. Colonies on OA<br />
reaching 19–34 mm diam, olive (2F5), loosely powdery with raised<br />
central part due to fasciculate bundles of conidiophores. Reverse<br />
dark green. Colonies on MEA reaching 30–32 mm diam, reseda<br />
green (2E6), velvety, furrowed, with undulate margin. Reverse dark<br />
green-brown. Colonies on MEA + 5 % NaCl reaching 37–41 mm<br />
diam, reseda-green (2E6), radially furrowed, velvety, sporulating in<br />
the central part or all over the colony, margin white <strong>and</strong> regular.<br />
Reverse brownish green.<br />
Maximum tolerated salt concentration: 75 % of tested strains<br />
develop colonies at 20 % NaCl after 7 d, while after 14 d all strains<br />
grow <strong>and</strong> sporulate.<br />
Cardinal temperatures: No growth at 4 <strong>and</strong> 10 °C, optimum 25 °C<br />
(30–32 mm diam), maximum 30 °C (2–15 mm diam), no growth at<br />
37 °C.<br />
Specimen examined: Dominican Republic, from hypersaline water of salt lake<br />
Enriquillo, coll. Nina Gunde-Cimerman, Jan. 2001, isol. P. Zalar 25 Feb. 2001, <strong>CBS</strong><br />
H-19733, holotype, culture ex-type EXF-732 = <strong>CBS</strong> 119415.<br />
Habitats <strong>and</strong> distribution: Fruit surfaces; hypersaline waters in<br />
(sub)tropical climates.<br />
Differential parameters: No growth at 10 °C, ovoid conidia, large<br />
amounts of sterile mycelium.<br />
Strains examined: CPC 11683, EXF-696, EXF-718, EXF-720, EXF-<br />
727, EXF-732 (= <strong>CBS</strong> 119415; ex-type strain).<br />
www.studiesinmycology.org<br />
Note: Cultures of C. dominicanum sporulate less abundantly than<br />
C. sphaerospermum <strong>and</strong> C. halotolerans <strong>and</strong> tend to lose their<br />
ability to sporulate with subculturing.<br />
<strong>Cladosporium</strong> fusiforme Zalar, de Hoog & Gunde-Cimerman, sp.<br />
nov. MycoBank MB510997. Fig. 7.<br />
Etymology: Refers to its usually fusiform conidia.<br />
Conidiophora erecta, lateralia vel terminalia ex hyphis rectis oriunda; stipes<br />
longitudine variabili, (10–)25–50(–100) × (2–)2–3.5(–4) μm, olivaceo-brunneus,<br />
levis, crassitunicatus, compluries septatus (cellulis 9–23 μm longis), plerumque<br />
simplex. Conidiorum catenae undique divergentes, in parte continua ad 5 conidia<br />
continentes. Cellulae conidiogenae indistinctae. Conidia leniter verruculosus, dilute<br />
brunnea, unicellularia, plerumque fusiformia, utrinque angustata, (2.5–)3.5–5(–<br />
6.5) × (2–)2–2.5(–3) μm, long. : lat. 1.8–2.0; ramoconidia secundaria cylindrica,<br />
0(–1)-septata, (5–)6–11(–22) × (2.5–)2.5–3(–3) μm, ad 4 cicatrices terminales<br />
ferentia; cicatrices inspissatae, conspicuae, 0.7–1.0 μm diam. Hyphae vagina<br />
polysaccharidica carentes.<br />
Mycelium without extracellular polysaccharide-like material.<br />
Conidiophores erect, arising laterally <strong>and</strong> terminally from straight<br />
hyphae, stipes of variable length, (10–)25–50(–100) × (2–)2–3.5(–<br />
4) μm, olivaceous-brown, smooth- <strong>and</strong> thick-walled, regularlyseptate<br />
(cell length 9–23 μm), mostly unbranched. Conidial chains<br />
branching in all directions, up to 5 conidia in the unbranched parts.<br />
Conidiogenous cells undifferentiated. Ramoconidia rarely formed.<br />
Conidia minutely verruculose, light brown, aseptate, usually fusiform<br />
<strong>and</strong> narrower at both ends, length : width ratio = 1.8–2.0; (2.5–)3.5–<br />
5(–6.5) × (2–)2–2.5(–3) μm [av. (± SD) 4.4 (± 0.8) × 2.2 (± 0.2)];<br />
secondary ramoconidia cylindrical, 0(–1)-septate, (5–)6–11(–22) ×<br />
(2.5–)2.5–3(–3) μm [av. (± SD) 9.0 (± 4.7) × 2.6 (± 0.3)], with up<br />
to 4 distal scars. Conidiogenous scars thickened <strong>and</strong> conspicuous,<br />
protuberant, 0.7–1.0 μm diam.<br />
Cultural characteristics: Colonies on PDA reaching 20–26 mm diam,<br />
dull green (30E3), granular due to profuse sporulation, flat, with flat<br />
margin. Sterile mycelium absent. Reverse blackish green. Colonies<br />
on OA reaching 24–28 mm diam, olive (3F3), granular in concentric<br />
circles, consisting of two kinds of conidiophores (low <strong>and</strong> high), flat,<br />
with flat margin. Reverse black. Colonies on MEA reaching 23–28<br />
mm diam, olive (3E5), deeply furrowed, velvety (sporulating all over)<br />
with undulate, white margin. Reverse brownish green. Colonies on<br />
MEA + 5 % NaCl reaching 28–43 mm diam, olive (3E6), granular<br />
due to profuse sporulation, slightly furrowed with flat, olive-grey<br />
(3F2) margin. Reverse dark green.<br />
Maximum tolerated salt concentration: Only one of three strains<br />
tested (<strong>CBS</strong> 452.71) developed colonies at 17 % NaCl after 14 d,<br />
the other two strains grew until 10 % NaCl.<br />
Cardinal temperatures: For one of three strains (<strong>CBS</strong> 452.71) the<br />
minimum temperature of growth was 4 °C (6 mm diam), for the<br />
other two 10 °C (8–9 mm diam); optimum 25 °C (23–28 mm diam),<br />
maximum 30 °C (only strain <strong>CBS</strong> 452.71 grew 5 mm diam), no<br />
growth at 37 °C.<br />
Specimen examined: Slovenia, from hypersaline water of Sečovlje salterns, coll.<br />
<strong>and</strong> isol. L. Butinar, Dec. 1999, <strong>CBS</strong> H-19732, holotype, culture ex-type EXF-449<br />
= <strong>CBS</strong> 119414.<br />
Habitats <strong>and</strong> distribution: Osmotic environments worldwide.<br />
Differential parameters: Oblong conidia, relatively low degree of<br />
halotolerance.<br />
Strains examined: <strong>CBS</strong> 452.71, EXF-397, EXF-449 (= <strong>CBS</strong> 119414;<br />
ex-type strain).<br />
169
Zalar et al.<br />
Fig. 6. <strong>Cladosporium</strong> dominicanum. Macro- <strong>and</strong> micromorphological characters. A–D. Colony surface grown on PDA (A), OA (B), MEA (C) <strong>and</strong> MEA plus 5 % NaCl (D) of strains<br />
incubated for 14 d at 25 ºC in darkness. E–F. Habit of conidiophores. G. Conidiophore. H–I. Secondary ramoconidia <strong>and</strong> conidia. E–I. All from 7-d-old SNA slide cultures. A, D,<br />
F–H, from EXF-2519; B, C, E from EXF-727; I, EXF-732 (ex-type strain). Scale bars A–D = 10 mm, E = 100 µm, F = 30 µm, G–I = 10 µm.<br />
170
<strong>Cladosporium</strong> sphaerospermum species complex<br />
Fig. 7. <strong>Cladosporium</strong> fusiforme. Macro- <strong>and</strong> micromorphological characters. A–D. Colony surface grown on PDA (A), OA (B), MEA (C) <strong>and</strong> MEA plus 5 % NaCl (D) of strains<br />
incubated for 14 d at 25 ºC in darkness. E–G. Habit of conidiophores. H–I. Ramoconidia <strong>and</strong> conidia. E–I. All from 7-d-old SNA slide cultures. A–H, from EXF-449 (ex-type strain);<br />
I, from <strong>CBS</strong> 452.71. Scale bars A–D = 10 mm, E = 100 µm, F–G = 30 µm, H–I = 10 µm.<br />
www.studiesinmycology.org<br />
171
Zalar et al.<br />
Fig. 8. <strong>Cladosporium</strong> halotolerans. Macro- <strong>and</strong> micromorphological characters. A–D. Colony surface grown on PDA (A), OA (B), MEA (C) <strong>and</strong> MEA plus 5 % NaCl (D) of strains<br />
incubated for 14 d at 25 ºC in darkness. E–H. Habit of conidiophores. I. Conidiophore. J. Succession of secondary ramoconidia. K. Conidia. E–K. All from 7-d-old SNA slide<br />
cultures. A–B, from EXF-572 (ex-type strain); C–D, from EXF-977; E, G, from EXF-972; F, from EXF-564; H, I, K, from EXF-1072; J, from dH 12862. Scale bars A–D = 10 mm,<br />
E = 100 µm, F–G = 50 µm, H = 30 µm, I–K = 10 µm.<br />
<strong>Cladosporium</strong> halotolerans Zalar, de Hoog & Gunde-Cimerman<br />
sp. nov. MycoBank MB492439. Fig. 8.<br />
Etymology: Refers to its halotolerant habit.<br />
Conidiophora erecta, lateralia vel terminalia ex hyphis rectis oriunda; stipes<br />
longitudine variabili, (5–)10–50(–300) × (2–)2.5–3(–5.5) μm, pallide olivaceobrunneus,<br />
levis vel leniter verruculosus, tenuitunicatus, 0–3-septatus, interdum<br />
pluriseptatus, simplex, denticulatus. Conidiorum catenae undique divergentes,<br />
terminales ad 9 conidia continentes. Cellulae conidiogenae indistinctae. Conidia<br />
verrucosa, brunnea vel fusca, unicellularia, plerumque subglobosa vel globosa,<br />
raro breviter ovoidea, utrinque angustata, (2–)3–4(–6) × (2–)2.5–3(–5) μm, long.<br />
: lat. 1.2–1.5; ramoconidia secundaria cylindrica vel quasi globosa, 0(–1)-septata,<br />
(5–)7–12(–37.5) × (2–)2.5–3(–6.5) μm , ad 4 cicatrices terminales ferentia; cicatrices<br />
inspissatae, conspicuae, protuberantes, 0.7–1.0(–1.5) μm diam. Hyphae vagina<br />
polysaccharidica carentes.<br />
172
<strong>Cladosporium</strong> sphaerospermum species complex<br />
Mycelium partly submerged, partly superficial; hyphae without<br />
extracellular polysaccharide-like material. Conidiophores erect,<br />
arising laterally <strong>and</strong> terminally from straight hyphae, stipes of<br />
variable length, (5–)10–50(–300) × (2–)2.5–3(–5.5) μm, pale<br />
olivaceous-brown, smooth to minutely verruculose, thin-walled, 0–<br />
3-septate, unbranched, with pronounced denticles. Conidial chains<br />
branching in all directions, terminal chains with up to 9 conidia.<br />
Conidiogenous cells undifferentiated. Ramoconidia rarely formed.<br />
Conidia verrucose, brown to dark brown, non-septate, usually<br />
subglobose to globose, less often short-ovoid, narrower at both<br />
ends, length : width ratio = 1.2–1.5; (2–)3–4(–6) × (2–)2.5–3(–5)<br />
μm [av. (± SD) 3.5 (± 0.7) × 2.7 (± 0.5)]; secondary ramoconidia<br />
cylindrical to almost spherical, 0–1-septate, (5–)7–12(–37.5) ×<br />
(2–)2.5–3(–6.5) μm [av. (± SD) 10.3 (± 4.8) × 2.9 (± 0.6)], with up<br />
to 4 distal scars. Conidiogenous scars thickened <strong>and</strong> conspicuous,<br />
protuberant, 0.7–1.0(–1.5) μm diam.<br />
Cultural characteristics: Colonies on PDA reaching 27–43 mm diam,<br />
olive (2F5), slightly furrowed, often covered with grey secondary<br />
mycelium, except at the marginal area where only sporulating<br />
structures can be observed. Margin white <strong>and</strong> regular, with<br />
submerged hyphae. Reverse pale green to black. Colonies on OA<br />
reaching 29–40 mm diam, olive (2F6), flat, uniform, granular due<br />
to profuse sporulation <strong>and</strong> fasciculate bundles of conidiophores,<br />
without sterile mycelium. Reverse dark green to black. Colonies on<br />
MEA reaching 18–44 mm diam, highly variable in colour, but mainly<br />
olive (2E5), <strong>and</strong> from flat with regular margin to deeply furrowed<br />
with undulate margin. Colony centre wrinkled with crater-shaped<br />
appearance. Reverse pale to dark green. Colonies on MEA + 5 %<br />
NaCl reaching 24–48 mm diam, olive (3E8), furrowed, velvety, with<br />
more pale, undulate margins. Reverse dark green to black.<br />
Maximum tolerated salt concentration: Only 15 % of tested strains<br />
develop colonies at 20 % NaCl after 7 d, whereas after 14 d all<br />
cultures grow <strong>and</strong> sporulate.<br />
Cardinal temperatures: No growth at 4 °C, optimum 25 °C (18–44<br />
mm diam), maximum 30 °C (6–23 mm diam). No growth at 37 °C.<br />
Specimen examined: Namibia, from hypersaline water of salterns, coll. Nina Gunde-<br />
Cimerman, 1 Sep. 2000, isol. P. Zalar, 1 Oct. 2000, <strong>CBS</strong> H-19734, holotype, culture<br />
ex-type EXF-572 = <strong>CBS</strong> 119416.<br />
Habitats <strong>and</strong> distribution: Hypersaline water in subtropical climates;<br />
indoor environments; Arctic ice; contaminant in lesions of humans<br />
<strong>and</strong> animals; plant phyllosphere; rock.<br />
Literature: Haubold et al. (1998), Meklin et al. (2004).<br />
Differential parameters: Verrucose conidia, short unbranched <strong>and</strong><br />
non-septate conidiophores which arise laterally alongside erect<br />
hyphae.<br />
Strains examined: <strong>CBS</strong> 191.54, <strong>CBS</strong> 573.78, <strong>CBS</strong> 626.82, dH<br />
12862, dH 12991, dH 13911, EXF-228, EXF-380, EXF-565, EXF-<br />
567, EXF-571, EXF-572 (= <strong>CBS</strong> 119416; ex-type strain), EXF-646,<br />
EXF-698, EXF-703, EXF-944, EXF-972, EXF-977, EXF-1072,<br />
EXF-2372.<br />
Notes: <strong>Cladosporium</strong> halotolerans strongly resembles C.<br />
sphaerospermum. Several strains of this species such as dH<br />
12862, dH 12941, <strong>CBS</strong> 191.54 <strong>and</strong> UAMH 7686 have been<br />
isolated sporadically from various indoor habitats in Europe, Brazil<br />
<strong>and</strong> the U.S.A. <strong>and</strong> repeatedly from bathrooms in Slovenia (Table<br />
1). Probably sometimes as uncertain culture contaminations, it<br />
has been isolated from plants (GenBank accession no. L25433),<br />
www.studiesinmycology.org<br />
inner organs of a diseased frog (AY361982) <strong>and</strong> human brain<br />
(Kantarcioglu et al. 2002). <strong>The</strong> presence of C. halotolerans<br />
species in gypsum sediments entrapped in Arctic ice, the fact that<br />
it was repeatedly isolated from hypersaline water <strong>and</strong> possibly its<br />
presence in dolphin skin (see Discussion) suggest that it has a<br />
clear preference for (hyper)osmotic habitats. This is supported by<br />
its ability to grow at 20 % NaCl.<br />
<strong>The</strong> teleomorph of C. halotolerans is predicted to be a<br />
Davidiella species. Strain <strong>CBS</strong> 280.49 was isolated by J.A. von Arx<br />
from teleomorphic material of a fungus labelled as Mycosphaerella<br />
hyperici (Auersw.) Starbäck on Hypericum perforatum in<br />
Switzerl<strong>and</strong>. According to Aptroot (2006) this species may belong<br />
in Davidiella <strong>and</strong> produces a Septoria anamorph. In the original<br />
herbarium specimen, <strong>CBS</strong> H-4867, a Mycosphaerella teleomorph<br />
was present, but no sign of a <strong>Cladosporium</strong> anamorph. We assume<br />
that <strong>CBS</strong> 280.49 was a culture contaminant.<br />
<strong>Cladosporium</strong> langeronii (Fonseca, Leão & Nogueira) Vuill.,<br />
Champ. Paras.: 78. 1931. Fig. 9.<br />
Basionym: Hormodendrum langeronii Fonseca, Leão & Nogueira,<br />
Sci. Med. 5: 563. 1927.<br />
≡ <strong>Cladosporium</strong> langeronii (Fonseca, Leão & Nogueira) Cif., Manuale di<br />
Micologia Medica, ed. 2: 488 (1960), comb. superfl.<br />
Mycelium partly submerged, partly superficial; hyphae sometimes<br />
enveloped in polysaccharide-like material. Conidiophores erect or<br />
ascending, micronematous <strong>and</strong> macronematous, stipes of variable<br />
length, (20–)50–130(–200) × (3–)3.5–4.5(–6.5) μm, dark brown,<br />
rough- <strong>and</strong> thick-walled, regularly septate (cell length 9–22 μm),<br />
arising laterally <strong>and</strong> terminally from submerged or aerial hyphae,<br />
branched. Conidial chains dichotomously branched, up to 6 conidia<br />
in the unbranched parts. Conidiogenous cells undifferentiated,<br />
sometimes seceding <strong>and</strong> forming ramoconidia. Ramoconidia<br />
cylindrical, 0–1 septate, (10–)11–22(–42) × (3–)3.5–4.5(–5) µm,<br />
base broadly truncate, 2–3.5 µm wide, slightly thickened <strong>and</strong><br />
somewhat darkened. Conidia irregularly verruculose to sometimes<br />
loosely verrucose, dark brown, non-septate, usually ovoid, length<br />
: width ratio = 1.3–1.5; conidial size (3–)4–5.5(–8) × (2–)3–4(–5)<br />
μm [av. (± SD) 4.8 (± 1.0) × 3.5 (± 0.6)]; secondary ramoconidia<br />
cylindrical to almost spherical, mostly 0–1(–2)-septate, (5.5–)7.5–<br />
12.5(–35.5) × (2.5–)3–4.5(–5.5) μm [av. (± SD) 10.7 (± 4.7) × 3.6 (±<br />
0.8)], with 2, rarely 3 distal scars. Conidiogenous scars thickened<br />
<strong>and</strong> conspicuous, protuberant, 0.9–1.5(–2.3) μm diam.<br />
Cultural characteristics: Colonies on PDA, OA <strong>and</strong> MEA with<br />
restricted growth, attaining 2.5–4.5, 1.5–7.0 <strong>and</strong> 1.0–5.5 mm<br />
diam, respectively. Colonies flat or heaped (up to 3 mm), dark<br />
green (30F4), with black reverse <strong>and</strong> slightly undulate margin with<br />
immersed mycelium. Sporulating on all media. On MEA + 5 % NaCl<br />
growth is faster, colonies attaining 8.5–12.0 mm diam, sporulating<br />
<strong>and</strong> growing deeply into the agar.<br />
Maximum tolerated salt concentration: All strains develop colonies<br />
at 17 % NaCl after 14 d.<br />
Cardinal temperatures: No growth at 4 °C, optimum / maximum 25<br />
°C (1.0–5.5 mm diam), no growth at 30 °C.<br />
Specimen examined: Brazil, from man ulcero-nodular mycosis of h<strong>and</strong> <strong>and</strong> arm,<br />
1927, coll. <strong>and</strong> isol. da Fonseca, <strong>CBS</strong> H-19737, holotype, culture ex-type <strong>CBS</strong><br />
189.54.<br />
Habitats <strong>and</strong> distribution: Polar ice <strong>and</strong> biomats; conifer wood <strong>and</strong><br />
window frame in Europe; humans; strains originating from nasal<br />
mucus (Buzina et al. 2003) have 100 % sequence homology with<br />
173
Zalar et al.<br />
Fig. 9. <strong>Cladosporium</strong> langeronii. Macro- <strong>and</strong> micromorphological characters. A–D. Colony surface grown on PDA (A), OA (B), MEA (C) <strong>and</strong> MEA plus 5 % NaCl (D) of strains<br />
incubated for 14 d at 25 ºC in darkness. E–F. Habit of conidiophores. G–I. Ramoconidia <strong>and</strong> conidia. E–I. All from 7-d-old SNA slide cultures. A–D, from <strong>CBS</strong> 189.54 (ex-type<br />
strain); E, from <strong>CBS</strong> 109868; F–I, from EXF-999. Scale bars A, C–D = 10 mm, B = 5 mm, E = 100 µm, F = 30 µm, G–I = 10 µm.<br />
the strains studied, as well as with a clone from mycorrhizal roots<br />
(Menkis et al. 2005). <strong>The</strong> species is distributed worldwide, without<br />
any apparent predilection for a particular habitat. <strong>The</strong> strains from<br />
clinical cases probably were culture contaminants.<br />
Literature: da Fonseca et al. (1927a, b).<br />
Differential parameters: Restricted growth; lowest salt halotolerance<br />
taxon of all C. sphaerospermum-like species.<br />
Strains examined: <strong>CBS</strong> 189.54 (ex-type strain), <strong>CBS</strong> 601.84, <strong>CBS</strong><br />
101880, <strong>CBS</strong> 109868, dH 11736, dH 12459 = EXF-999, dH 13833<br />
= EXF-1933.<br />
Notes: De Vries (1952) synonymised the isolate identified as<br />
Hormodendrum langeronii with C. sphaerospermum. Strains of this<br />
species have often been identified as C. cladosporioides (Buzina<br />
et al. 2003, Menkis et al. 2005) although it has slightly longer<br />
conidia.<br />
174
<strong>Cladosporium</strong> sphaerospermum species complex<br />
<strong>Cladosporium</strong> psychrotolerans Zalar, de Hoog & Gunde-<br />
Cimerman, sp. nov. MycoBank MB492428. Fig. 10.<br />
Etymology: Refers to its ability to grow at low temperatures.<br />
Mycelium partim submersum; hyphae vagina polysaccharidica carentes.<br />
Conidiophora erecta vel adscendentia; stipes (10–)50–100(–150) × (3–)3.5–4(–7.5)<br />
μm, olivaceo-brunneus, levis, crassitunicatus, compluries regulariter septatus<br />
(cellulis 10–40 μm longis), identidem dichotome ramosus. Conidiorum catenae<br />
undique divergentes, terminales partes simplices ad 4 conidia continentes. Cellulae<br />
conidiogenae indistinctae. Ramoconidia primaria cylindrica, (18–)19–22(–43) ×<br />
(2.5)3–3.5(–4.5) µm, 0(–1)-septata. Conidia leves vel leniter verruculosa, dilute<br />
brunnea, unicellularia, globosa vel ovoidea, (2.5–)3–4(–4.5) × (2–)2.5–3(–3) μm,<br />
long.: lat. 1.3–1.4; ramoconidia secundaria cylindrica, 0–1(–2)-septata, (5–)8–16(–<br />
36) × (2–)2.5–3(–5) μm, ad 4 cicatrices terminales ferentia; cicatrices inspissatae,<br />
conspicuae, 0.5–2 μm diam.<br />
Mycelium partly superficial partly submerged; hyphae without<br />
extracellular polysaccharide-like material. Conidiophores erect or<br />
ascending, macronematous, stipes (10–)50–100(–150) × (3–)3.5–<br />
4(–7.5) μm, olivaceous-brown, smooth or almost so, thick-walled,<br />
regularly septate (cell length 10–40 μm), arising laterally from<br />
aerial hyphae, repeatedly dichotomously branched. Conidial chains<br />
branching in all directions, up to 4 conidia in the unbranched parts.<br />
Ramoconidia sometimes formed, cylindrical, (18–)19–22(–43) ×<br />
(2.5)3–3.5(–4.5) µm, aseptate, rarely 1-septate, with a broadly<br />
truncate base, up to 2 µm wide, unthickened or slightly thickened,<br />
somewhat darkened-refractive. Conidia smooth to minutely<br />
verruculose, light brown, non-septate, spherical to ovoid, length :<br />
width ratio = 1.3–1.4; conidial size (2.5–)3–4(–4.5) × (2–)2.5–3(–3)<br />
μm [av. (± SD) 3.4 (± 0.5) × 2.5 (± 0.2)]; secondary ramoconidia<br />
cylindrical, 0–1(–2)-septate, (5–)8–16(–36) × (2–)2.5–3(–5) μm<br />
[av. (± SD) 12.7 (± 6.5) × 3.0 (± 0.5)], with up to 4 distal scars.<br />
Conidiogenous scars thickened <strong>and</strong> conspicuous, protuberant,<br />
0.5–2 μm diam.<br />
Cultural characteristics: Colonies on PDA reaching 13–18 mm<br />
diam, velvety, olive (3F4) due to profuse sporulation, flat with<br />
straight margin. Reverse dark green. Colonies on OA reaching 13–<br />
15 mm diam, olive (2F8), of granular appearance due to profuse<br />
sporulation; aerial mycelium sparse. Margin regular. Reverse black.<br />
Colonies on MEA reaching 8–15 mm diam, olive (2F4), velvety,<br />
radially furrowed with undulate white margin. Colonies on MEA<br />
with 5 % NaCl growing faster than on other media, reaching 25–27<br />
mm diam, olive (3E6) <strong>and</strong> granular due to profuse sporulation,<br />
either slightly furrowed or heavily wrinkled with regular or undulate<br />
margin. Reverse dark green.<br />
Maximum tolerated salt concentration: 17 % NaCl after 14 d.<br />
Cardinal temperatures: Minimum at 4 °C (5 mm diam), optimum<br />
<strong>and</strong> maximum at 25 °C (8–15 mm diam).<br />
Specimen examined: Slovenia, from hypersaline water of Sečovlje salterns, coll.<br />
<strong>and</strong> isol. S. Sonjak, May 1999, <strong>CBS</strong> H-19730, holotype, culture ex-type EXF-391<br />
= <strong>CBS</strong> 119412.<br />
Habitats <strong>and</strong> distribution: Hypersaline water in the Mediterranean<br />
basin.<br />
Differential parameters: Growth at 4 °C; maximal NaCl<br />
concentration 17 % NaCl, which differentiates it from other species<br />
with <strong>similar</strong> conidia, like C. sphaerospermum, C. halotolerans <strong>and</strong><br />
C. dominicanum.<br />
Strains examined: EXF-326, EXF-332, EXF-391 (= <strong>CBS</strong> 119412;<br />
ex-type strain), EXF-714.<br />
<strong>Cladosporium</strong> salinae Zalar, de Hoog & Gunde-Cimerman, sp.<br />
nov. MycoBank MB492438. Fig. 11.<br />
Etymology: Refers to salterns (= Latin salinae) as the habitat of<br />
this species.<br />
Mycelium partim submersum; hyphae multa rostra lateralia ferentes, hyphae vagina<br />
polysaccharidica involutae. Conidiophora vix distincta, lateralia vel terminalia ex<br />
hyphis aeriis oriunda; stipes longitudine variabili, (5–)25–50(–60) × (2–)2.5–3(–4)<br />
μm, olivaceo-brunneus, levis vel leniter verruculosus, crassitunicatus, irregulariter<br />
dense septatus (cellulis 6–29 μm longis), simplex, interdum ramosus. Conidiorum<br />
catenae undique divergentes, terminales ad 6 conidia continentes. Cellulae<br />
conidiogenae nonnumquam integratae, in summo sequentiam sympodialem<br />
denticulorum formantes. Conidia levia, interdum leniter verruculosa, dilute brunnea,<br />
unicellularia, plerumque fusiformia, (4.5–)5.5–7.5(–10) × (2–)2.5–3(–3.5) μm,<br />
long. : lat. 1.9–2.4; ramoconidia secundaria cylindrica, 0–1(–2)-septata, (7.5–)9.5–<br />
13.5(–19) × (2.5–)2.5–3.5(–4.5) μm, ad 5 cicatrices terminales ferentia; cicatrices<br />
inspissatae, conspicuae, protuberantes, 0.7–1.8 μm diam.<br />
Mycelium partly superficial partly submerged, with numerous<br />
lateral pegs, consistently enveloped in polysaccharide-like<br />
material. Conidiophores poorly differentiated, micronematous,<br />
stipes (5–)25–50(–60) × (2–)2.5–3(–4) μm, olivaceous-brown,<br />
smooth to often minutely verruculose or irregularly rough-walled,<br />
thick-walled, irregularly densely septate (length of cells 6–29 μm),<br />
arising laterally <strong>and</strong> terminally from aerial hyphae, unbranched,<br />
occasionally branched. Conidial chains branching in all directions,<br />
terminal chains with up to 6 conidia. Conidiogenous cells sometimes<br />
integrated, producing sympodial clusters of pronounced denticles<br />
at their distal ends. Conidia usually smooth, occasionally minutely<br />
verruculose, light brown, aseptate, usually oblong ellipsoidal to<br />
fusiform, length : width ratio = 1.9–2.4; (4.5–)5.5–7.5(–10) × (2–)<br />
2.5–3(–3.5) μm [av. (± SD) 6.7 (± 1.3) × 2.9 (± 0.4)]; secondary<br />
ramoconidia cylindrical, 0–1(–2)-septate, (7.5–)9.5–13.5(–19)<br />
× (2.5–)2.5–3.5(–4.5) μm [av. (± SD) 12.1 (± 3.3) × 3.2 (± 0.6)],<br />
with up to 5 distal scars. Conidiogenous scars thickened <strong>and</strong><br />
conspicuous, protuberant, 0.7–1.8 μm diam.<br />
Cultural characteristics: Colonies on PDA reaching 10–27 mm<br />
diam, granular, olive (2E4) due to profuse sporulation, with white<br />
undulate margin. Aerial mycelium absent. Colonies either heaped<br />
or radially furrowed, in the marginal area growing deeply into the<br />
agar. Reverse dark brown to dark green. Colonies on OA reaching<br />
7–20 mm diam, olive (3E6), of granular appearance due to profuse<br />
sporulation, aerial mycelium present. Margin either undulate or<br />
arachnoid, deeply furrowed. Reverse pale brown to dark green.<br />
Colonies on MEA reaching 8–19 mm diam, velvety, reseda-green<br />
(2E6), heaped. Margin furrowed, growing deeply into the agar.<br />
Colonies on MEA with 5 % NaCl growing much faster than on<br />
other media, reaching 25–38 mm diam, of different colours, mostly<br />
reseda-green (2E6) <strong>and</strong> granulate due to profuse sporulation,<br />
margin olive-yellow (2D6). Reverse yellow to dark green.<br />
Maximum tolerated salt concentration: MEA + 17 % NaCl after 14 d.<br />
Cardinal temperatures: No growth at 4 °C, optimum <strong>and</strong> maximum<br />
temperature at 25 °C (8–19 mm diam), no growth at 30 °C.<br />
Specimen examined: Slovenia, from hypersaline water of Sečovlje salterns, coll.<br />
<strong>and</strong> isol. S. Sonjak, Feb. 1999, <strong>CBS</strong> H-19731, holotype, culture ex-type EXF-335<br />
= <strong>CBS</strong> 119413.<br />
Habitats <strong>and</strong> distribution: Hypersaline water in the Mediterranean<br />
basin.<br />
Differential parameters: Sympodial conidiogenous cells with<br />
pronounced denticles, narrow temperature amplitude.<br />
www.studiesinmycology.org<br />
175
Zalar et al.<br />
Fig. 10. <strong>Cladosporium</strong> psychrotolerans. Macro- <strong>and</strong> micromorphological characters. A–D. Colony surface grown on PDA (A), OA (B), MEA (C) <strong>and</strong> MEA plus 5 % NaCl (D) of<br />
strains incubated for 14 d at 25 ºC in darkness. E–F. Conidiophores. G. Apical part of a conidiophore. H–I. Secondary ramoconidia <strong>and</strong> conidia. E–I. All from 7-d-old SNA slide<br />
cultures. All but C, from EXF-391 (ex-type strain); C, from EXF-714. Scale bars A–D = 10 mm, E = 100 µm, F = 50 µm, G–I = 10 µm.<br />
Strains examined: EXF-322, EXF-335 (= <strong>CBS</strong> 119413; ex-type<br />
strain), EXF-604.<br />
Notes: <strong>Cladosporium</strong> salinae morphologically resembles species<br />
of the <strong>genus</strong> Fusicladium because its conidia are oblong ellipsoidal<br />
to fusiform <strong>and</strong> conidiogenous loci of ramoconidia are placed<br />
closely together. As any other <strong>Cladosporium</strong> species, its conidia<br />
show typical cladosporioid scar structures, however. <strong>Cladosporium</strong><br />
salinae seems to have a separate position within the <strong>genus</strong><br />
<strong>Cladosporium</strong> since it seems to be distantly related to any other<br />
described <strong>Cladosporium</strong> species or currently known species<br />
complex within <strong>Cladosporium</strong>.<br />
176
<strong>Cladosporium</strong> sphaerospermum species complex<br />
Fig. 11. <strong>Cladosporium</strong> salinae. Macro- <strong>and</strong> micromorphological characters. A–D. Colony surface grown on PDA (A), OA (B), MEA (C) <strong>and</strong> MEA plus 5 % NaCl (D) of strains<br />
incubated for 14 d at 25 ºC in darkness. E–F. Habit of conidiophores. G. Conidiophore. H. Detail of apical part of conidiophore. I. Conidia. J. Secondary ramoconidia <strong>and</strong> conidia.<br />
E–J. All from 7-d-old SNA slide cultures. A–D, from EXF-604; E–J, from EXF-335 (ex-type strain). Scale bars A–C = 5 mm, D = 10 mm, E = 100 µm, F = 50 µm, G = 30 µm,<br />
H–J = 10 µm.<br />
<strong>Cladosporium</strong> sphaerospermum Penzig, Michelia 2(8): 473.<br />
1882. Fig. 12.<br />
Mycelium partly submerged, partly superficial; hyphae thick, darkly<br />
pigmented <strong>and</strong> densely septate in submerged mycelium, not<br />
enveloped in polysaccharide-like material. Conidiophores erect or<br />
ascending, micronematous <strong>and</strong> macronematous, stipes of variable<br />
length, (10–)45–130(–300) × (2.5–)3–4(–6) μm, olivaceous-brown,<br />
smooth to minutely verruculose, thick-walled, with relatively<br />
dense septation (cells mostly 4.5–23 long), septa darkened <strong>and</strong><br />
www.studiesinmycology.org<br />
somewhat thickened, arising laterally <strong>and</strong> terminally from immersed<br />
or aerial hyphae, either unbranched or branched. Conidial chains<br />
branching in all directions, up to 6 conidia in the unbranched parts.<br />
Conidiogenous cells not differentiated. Ramoconidia often formed,<br />
cylindrical, (11.5–)20.5–40(–48) × (2.5–)3(–3.5) µm, with up to 5<br />
septa, base broadly truncate, 2 µm wide, slightly thickened <strong>and</strong><br />
somewhat darkened-refractive. Conidia verruculose, brown to<br />
dark brown, non-septate, usually subspherical to spherical, less<br />
often short-ovoid, narrower at both ends, with length : width ratio<br />
= 1.1–1.5; conidial size (2.5–)3–4(–7) × (2–)3–3.5(–4.5) μm [av. (±<br />
177
Zalar et al.<br />
Fig. 12. <strong>Cladosporium</strong> sphaerospermum. Macro- <strong>and</strong> micromorphological characters. A–D. Colony surface grown on PDA (A), OA (B), MEA (C) <strong>and</strong> MEA plus 5 % NaCl (D) of<br />
strains incubated for 14 d at 25 ºC in darkness. E–F. Habit of conidiophores. G–I. Ramoconidia <strong>and</strong> conidia. E–I. All from 7-d-old SNA slide cultures. A, C–D, F–H, from <strong>CBS</strong><br />
193.54 (ex-neotype strain); B, from EXF-738; E, EXF-455; I, EXF-458. Scale bars A–D = 10 mm; E = 100 µm; F = 50 µm; G–I = 10 µm.<br />
SD) 3.8 (± 0.8) × 3.1 (± 0.4)]; secondary ramoconidia cylindrical to<br />
almost spherical, 0–3(–4) septate, (4–)8.5–16(–37.5) × (2–)3–3.5(–<br />
5) μm [av. (± SD) 13.1 (± 6.3) × 3.2 (± 0.5)], with up to 4, rarely up<br />
to 6 distal scars. Conidiogenous scars thickened <strong>and</strong> conspicuous,<br />
protuberant, 0.9–1.1(–1.4) μm diam.<br />
Cultural characteristics: Colonies on PDA reaching 21–44 mm diam,<br />
velvety, olive (2F5) due to profuse sporulation, either with white<br />
<strong>and</strong> regular, or exceptionally undulate margin. Aerial mycelium<br />
sparse. Colonies flat or rarely radially furrowed with elevated<br />
colony centre. Exudates not prominent, some strains release<br />
green soluble pigments into the agar. Reverse blackish blue to pale<br />
green. Growth deep into the agar. Colonies on OA reaching 21–<br />
38 mm diam, olive (2F8), of granular appearance due to profuse<br />
<strong>and</strong> uniform sporulation, almost no aerial mycelium. Margin either<br />
regular or arachnoid, deeply radially furrowed. Reverse black.<br />
Colonies on MEA reaching 15–35 mm diam, velvety, linden-green<br />
(2C5), radially furrowed. Colony centre wrinkled, forming a craterlike<br />
structure; margin furrowed, lighter in colour, consisting of<br />
178
<strong>Cladosporium</strong> sphaerospermum species complex<br />
Fig. 13. <strong>Cladosporium</strong> spinulosum. Macro- <strong>and</strong> micromorphological characters. A–D. Colony surface grown on PDA (A), OA (B), MEA (C) <strong>and</strong> MEA plus 5 % NaCl (D) of strains<br />
incubated for 14 d at 25 ºC in darkness. E. Habit of conidiophores. F–J. Conidiophores. K–L. Conidia (also visible in I–J). E–L. All from 7-d-old SNA slide cultures. A–L, from<br />
EXF-334 (ex-type strain). Scale bars A–D = 10 mm, E = 100 µm, F = 30 µm, G–L = 10 µm.<br />
submerged mycelium. Reverse pale to dark brown. Colonies on<br />
MEA with 5 % NaCl growing faster than on other media, reaching<br />
31–60 mm diam, mainly olive (2D4), either being almost flat or<br />
radially furrowed, with margin of superficial mycelium; sporulation<br />
dense. Reverse ochraceous or dark green.<br />
Maximum tolerated salt concentration: On MEA + 20 % NaCl 89 %<br />
of all strains tested develops colonies after 7 d, 96 % after 14 d.<br />
Cardinal temperatures: No growth at 4 °C, optimum 25 °C (15–35<br />
mm diam), maximum 30 °C (2–15 mm diam). No growth at 37 °C.<br />
Specimen examined: Netherl<strong>and</strong>s, from nail of man, 1949, coll. <strong>and</strong> isol. R.W.<br />
Zappey, <strong>CBS</strong> H-19738, neotype designated here, incorrectly selected by de Vries<br />
(1952) as “lectotype”, culture ex-neotype <strong>CBS</strong> 193.54 = ATCC 11289 = IMI 049637.<br />
Habitats <strong>and</strong> distribution: Hypersaline water in mediterranean<br />
<strong>and</strong> tropics; soil <strong>and</strong> plants in temperate climates; indoor wet<br />
cells; humans. <strong>The</strong> species does not seem to have any particular<br />
preference. Human isolates were probably culture contaminants.<br />
Literature: Penzig (1882), de Vries (1952), Ellis (1971), de Hoog et<br />
al. (2000), Samson et al. (2002).<br />
Diagnostic parameters: Thick-walled, melanised, densely septate<br />
mycelium, almost spherical, verruculose to verrucose terminal<br />
conidia, growth on 20 % NaCl after 7 d.<br />
Strains examined: <strong>CBS</strong> 109.14, <strong>CBS</strong> 122.63, <strong>CBS</strong> 190.54, <strong>CBS</strong><br />
192.54, <strong>CBS</strong> 193.54 (ex-neotype strain), <strong>CBS</strong> 102045, CPC 10944,<br />
EXF-131, EXF-328, EXF-385, EXF-446, EXF-455, EXF-458, EXF-<br />
461, EXF-464, EXF-465, EXF-598, EXF-644, EXF-645, EXF-649,<br />
EXF-715, EXF-738, EXF-739, EXF-781, EXF-962, EXF-965, EXF-<br />
1061, EXF-1726, EXF-1732.<br />
www.studiesinmycology.org<br />
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Zalar et al.<br />
Fig. 14. <strong>Cladosporium</strong> velox. Macro- <strong>and</strong> micromorphological characters. A–D. Colony surface grown on PDA (A), OA (B), MEA (C) <strong>and</strong> MEA plus 5 % NaCl (D) of strains<br />
incubated for 14 d at 25 ºC in darkness. E–F. Habit of conidiophores. G. Conidiophore. H–I. Secondary ramoconidia <strong>and</strong> conidia. E–I. All from 7-d-old SNA slide cultures. A–D,<br />
G, from <strong>CBS</strong> 119417 (ex-type strain); E–F, H–I, from EXF-466. Scale bars A–D = 10 mm, E = 100 µm, F = 50 µm, G = 30 µm, H–I = 10 µm.<br />
<strong>Cladosporium</strong> spinulosum Zalar, de Hoog & Gunde-Cimerman,<br />
sp. nov. MycoBank MB501099. Fig. 13.<br />
Etymology: Refers to its conspicuously digitate conidia.<br />
Conidiophora erecta vel adscendentia; stipites longitudine variabili, (15–)25–50(–<br />
155) × (2.5–)3–4(–5) μm, olivaceo-brunneus, levis, crassitunicatus, 0–6(–9)-septatus<br />
(cellulis 6–20 μm longis), ex hyphis submersis vel aeriis lateraliter vel terminaliter<br />
oriundus, simplex vel ramosus. Conidiorum catenae undique ramosae, ad 4 conidiis<br />
in partibus linearibus continuis cohaerentibus. Cellulae conidiogenae integratae vel<br />
discretae, acervos distales denticulorum conspicuorum sympodialium proferentes.<br />
Conidia echinulata vel digitata, brunnea vel fusca, continua, vulgo subglobosa vel<br />
globosa, (4.5–)5.5–7(–8) × (3–)4–4.5(–5) μm, long.: lat. = 1.1–1.6, digiti 0.6–1.3<br />
μm longi; ramoconidia secundaria etiam digitata, cylindrica vel subglobosa, 0(–1)-<br />
septata, (6–)6.5–8(–18) × (4–)4.5–5(–5.5) μm, 1–3 cicatrices distales ferentia.<br />
Cicatrices inspissatae, conspicuae, protuberantes, 0.8–1.2 μm diam. Hyphae<br />
nonnumquam polysaccharido circumdatae.<br />
Hyphae sometimes enveloped in polysaccharide-like material.<br />
Conidiophores erect or ascending, stipes of variable length, (15–)<br />
180
<strong>Cladosporium</strong> sphaerospermum species complex<br />
25–50(–155) × (2.5–)3–4(–5) μm, olivaceous-brown, smooth,<br />
sometimes irregularly rough-walled to verrucose near the base,<br />
thick-walled, 0–6(–9)-septate (cells mostly 6–20 μm long), arising<br />
laterally <strong>and</strong> terminally from immersed or aerial hyphae, either<br />
unbranched or branched, somewhat tapering towards the apex.<br />
Conidial chains branching in all directions, up to 4 conidia in the<br />
unbranched parts. Conidiogenous cells sometimes integrated,<br />
producing sympodial clusters of pronounced denticles at their distal<br />
ends. Ramoconidia rarely formed. Conidial wall ornamentation<br />
conspicuously digitate, with up to 1.3 μm long projections having<br />
parallel sides <strong>and</strong> blunt ends. Conidia brown to dark brown,<br />
aseptate, usually subspherical to spherical, length : width ratio =<br />
1.1–1.6; conidial size (4.5–)5.5–7(–8) × (3–)4–4.5(–5) μm [av. (±<br />
SD) 6.2 (± 1.0) × 4.2 (± 0.5)]; secondary ramoconidia ornamented<br />
as conidia, cylindrical to almost spherical, 0(–1)-septate, (6–)6.5–<br />
8(–18) × (4–)4.5–5(–5.5) μm [av. (± SD) 8.6 (± 4.0) × 4.8 (± 0.4)],<br />
with up to 3 distal scars. Conidiogenous scars thickened <strong>and</strong><br />
conspicuous, protuberant, 0.8–1.2 μm diam.<br />
Cultural characteristics: Colonies on PDA reaching 20–30 mm<br />
diam, velvety, dull green (29E4) to dark green (29F6) due to profuse<br />
sporulation, either with white <strong>and</strong> regular, or undulate margin. Aerial<br />
mycelium sparse. Colonies flat or radially furrowed with elevated<br />
colony centre. Growth deep into the agar. Exudates not prominent.<br />
Colonies on OA reaching 20–25 mm diam, dull green (29E4) to dark<br />
green (29F6), sometimes olive (3D4), of granular appearance due<br />
to profuse <strong>and</strong> uniform sporulation; almost without aerial mycelium.<br />
Margin arachnoid. Reverse pale brown to black. Colonies on MEA<br />
reaching 17–28 mm diam, velvety, dull green (29E4) to dark green<br />
(29F6), either flat or radially furrowed. Colony centre wrinkled,<br />
forming a crater-like structure; margin furrowed, paler in colour,<br />
consisting of submerged mycelium only. Reverse pale to dark<br />
green. Colonies on MEA with 5 % NaCl reaching 12–18 mm diam,<br />
of different colours, greenish grey (29D2), greyish green (29D5)<br />
to dark green (29F6); colony appearance variable, mostly either<br />
being almost flat with immersed colony centre or radially furrowed,<br />
with white to dark green margin consisting of superficial mycelium;<br />
sporulation dense. Reverse pale to dark green.<br />
Maximum tolerated salt concentration: On MEA + 17 % NaCl, two<br />
of three strains tested developed colonies after 14 d.<br />
Cardinal temperatures: Growth at 4 °C, optimum <strong>and</strong> maximum at<br />
25 °C (17–28 mm). No growth at 30 °C.<br />
Specimen examined: Slovenia, from hypersaline water of Sečovlje salterns, coll.<br />
<strong>and</strong> isol. S. Sonjak, Feb. 1999, <strong>CBS</strong> H-19796, holotype, culture ex-type EXF-334<br />
= <strong>CBS</strong> 119907.<br />
Habitats <strong>and</strong> distribution: Hypersaline water in temperate climate.<br />
Diagnostic parameters: Conidia <strong>and</strong> ramoconidia with a digitate<br />
ornamentation.<br />
Strains examined: EXF-334 (= <strong>CBS</strong> 119907; ex-type strain), EXF-<br />
382.<br />
Notes: <strong>Cladosporium</strong> spinulosum is a member of the C. herbarum<br />
species complex (Figs 2–4) although its globoid conidia are<br />
reminiscent of C. sphaerospermum. Within <strong>Cladosporium</strong>, the<br />
species is unique in having conspicuously digitate conidia <strong>and</strong><br />
ramoconidia. <strong>The</strong> two strains are differing in the size of conidia.<br />
<strong>The</strong> average size of conidia in EXF-334 is 6.2 (± 0.9) × 4.2 (± 0.5)<br />
μm, <strong>and</strong> in EXF-382 it is 3.9 (± 0.6) × 3.3 (± 0.4) μm.<br />
<strong>Cladosporium</strong> velox Zalar, de Hoog & Gunde-Cimerman, sp. nov.<br />
MycoBank MB492435. Fig. 14.<br />
Etymology: Refers to the quick growth of strains of this species.<br />
Mycelium partim submersum; hyphae vagina polysaccharidica carentes.<br />
Conidiophora erecta, lateralia vel terminalia ex hyphis aeriis oriunda; stipes (10–)<br />
25–150(–250) × (2.5–)3–4(–4.5) μm, olivaceo-brunneus, levis, crassitunicatus, ad<br />
7–septatus (cellulis 10–60 μm longis), identidem dichotome ramosus. Conidiorum<br />
catenae undique divergentes, terminales partes simplices ad 5 conidia continentes.<br />
Cellulae conidiogenae indistinctae. Conidia levia vel leniter verruculosa, dilute<br />
brunnea, unicellularia, ovoidea, (2–)3–4(–5.5) × (1.5–)2–2.5(–3) μm, long. : lat.<br />
1.4–1.7; ramoconidia secundaria cylindrica, 0–1-septata, (3.5–)5.5–19(–42) ×<br />
(2–)2.5–3(–4.5) μm, ad 4(–5) cicatrices terminales ferentia; cicatrices inspissatae,<br />
protuberantes, conspicuae, 0.5–1.5 μm diam.<br />
Mycelium partly superficial partly submerged; hyphae without<br />
extracellular polysaccharide-like material. Conidiophores erect,<br />
stipes (10–)25–150(–250) × (2.5–)3–4(–4.5) μm, slightly attenuated<br />
towards the apex, olivaceous-brown, smooth- <strong>and</strong> thick-walled,<br />
arising terminally <strong>and</strong> laterally from aerial hyphae, dichotomously<br />
branched [up to 5(–7)-septate, cell length 10–60 μm]. Ramoconidia<br />
rarely formed. Conidial chains branching in all directions, terminal<br />
chains with up to 5 conidia. Conidia smooth to very finely<br />
verruculose, pale brown, non-septate, ovoid, length : width ratio<br />
= 1.4–1.7; (2–)3–4(–5.5) × (1.5–)2–2.5(–3) μm [av. (± SD) 3.6 (±<br />
0.6) × 2.3 (± 0.2)]; secondary ramoconidia cylindrical, 0–1-septate,<br />
(3.5–)5.5–19(–42) × (2–)2.5–3(–4.5) μm [av. (± SD) 13.4 (± 10.2)<br />
× 2.8 (± 0.5)], with up to 4(–5) distal scars. Conidiogenous scars<br />
thickened <strong>and</strong> conspicuous, protuberant, 0.5–1.5 μm diam.<br />
Cultural characteristics: Colonies on PDA reaching 35–45 mm<br />
diam, velvety, dark green due to profuse sporulation, on some parts<br />
covered with white sterile mycelium, flat with straight white margin.<br />
Reverse dark green to black. Colonies on OA reaching 30–43 mm<br />
diam, dark green, mycelium submerged, aerial mycelium sparse.<br />
Margin regular. Reverse black. Colonies on MEA reaching 30–42<br />
mm diam, pale green, radially furrowed, with raised, crater-shaped<br />
central part, with white, undulate, submerged margin. Sporulation<br />
poor. Colonies on MEA with 5 % NaCl reaching 35–45 mm diam,<br />
pale green, velvety, flat with regular margin. Reverse pale green.<br />
Sporulation poor.<br />
Maximum tolerated salt concentration: 20 % NaCl after 14 d.<br />
Cardinal temperatures: Minimum at 10 °C (9 mm diam), optimum at<br />
25 °C (30–42 mm diam) <strong>and</strong> maximum at 30 °C (5–18 mm diam).<br />
Specimen examined: India, Charidij, isolated from Bambusa sp., W. Gams, <strong>CBS</strong><br />
H-19735, holotype, culture ex-type <strong>CBS</strong> 119417.<br />
Habitats <strong>and</strong> distribution: Hypersaline water in Slovenia; bamboo,<br />
India.<br />
Strains examined: <strong>CBS</strong> 119417 (ex-type strain), EXF-466, EXF-<br />
471.<br />
ACKNOWLEDGEMENTS<br />
<strong>The</strong> authors are grateful to Walter Gams for his comments on the manuscript<br />
<strong>and</strong> for providing Latin diagnoses, <strong>and</strong> to Konstanze Schubert for contributions to<br />
species descriptions. We thank Kazimir Drašlar <strong>and</strong> Marko Lutar for preparing SEM<br />
illustrations, <strong>and</strong> Kasper Luijsterburg, Špela Štrekelj <strong>and</strong> Barbara Kastelic-Bokal for<br />
their technical assistance. <strong>The</strong> research was partially supported by the European<br />
Union-funded Integrated Infrastructure Initiative grant SYNTHESYS Project NL-<br />
TAF-1070.<br />
www.studiesinmycology.org<br />
181
Zalar et al.<br />
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183
available online at www.studiesinmycology.org<br />
doi:10.3114/sim.2007.58.07<br />
Studies in Mycology 58: 185–217. 2007.<br />
Opportunistic, human-pathogenic species in the Herpotrichiellaceae are<br />
phenotypically <strong>similar</strong> to saprobic or phytopathogenic species in the<br />
Venturiaceae<br />
P.W. Crous 1* , K. Schubert 2 , U. Braun 3 , G.S. de Hoog 1 , A.D. Hocking 4 , H.-D. Shin 5 <strong>and</strong> J.Z. Groenewald 1<br />
1<br />
<strong>CBS</strong> Fungal Biodiversity Centre, P.O. Box 85167, 3508 AD, Utrecht, <strong>The</strong> Netherl<strong>and</strong>s; 2 Botanische Staatssammlung München, Menzinger Strasse 67, D-80638 München,<br />
Germany; 3 Martin-Luther-Universität, Institut für Biologie, Geobotanik und Botanischer Garten, Herbarium, Neuwerk 21, D-06099 Halle, Germany; 4 CSIRO Food Science<br />
Australia, 11 Julius Avenue, North Ryde, NSW 2113, Australia; 5 Division of Environmental Science & Ecological Engineering, Korea University, Seoul 136-701, Korea<br />
*Correspondence: Pedro W. Crous, p.crous@cbs.knaw.nl<br />
Abstract: Although morphologically <strong>similar</strong>, species of Cladophialophora (Herpotrichiellaceae) were shown to be phylogenetically distinct from Pseudocladosporium<br />
(Venturiaceae), which was revealed to be synonymous with the older <strong>genus</strong>, Fusicladium. Other than being associated with human disorders, species of Cladophialophora<br />
were found to also be phytopathogenic, or to occur as saprobes on organic material, or in water, fruit juices, or sports drinks, along with species of Exophiala. Caproventuria<br />
<strong>and</strong> Metacoleroa were confirmed to be synonyms of Venturia, which has Fusicladium (= Pseudocladosporium) anamorphs. Apiosporina, based on A. collinsii, clustered<br />
basal to the Venturia clade, <strong>and</strong> appears to represent a further synonym. Several species with a pseudocladosporium-like morphology in vitro represent a sister clade to the<br />
Venturia clade, <strong>and</strong> are unrelated to Polyscytalum. <strong>The</strong>se taxa are newly described in Fusicladium, which is morphologically close to Anungitea, a heterogeneous <strong>genus</strong> with<br />
unknown phylogenetic affinity. In contrast to the Herpotrichiellaceae, which were shown to produce numerous synanamorphs in culture, species of the Venturiaceae were<br />
morphologically <strong>and</strong> phylogenetically more uniform. Several new species <strong>and</strong> new combinations were introduced in Cladophialophora, Cyphellophora (Herpotrichiellaceae),<br />
Exophiala, Fusicladium, Venturia (Venturiaceae), <strong>and</strong> Cylindrosympodium (incertae sedis).<br />
Taxonomic novelties: Cladophialophora australiensis Crous & A.D. Hocking, sp. nov., Cladophialophora chaetospira (Grove) Crous & Arzanlou, comb. nov., Cladophialophora<br />
hostae Crous, U. Braun & H.D. Shin, sp. nov., Cladophialophora humicola Crous & U. Braun, sp. nov., Cladophialophora potulentorum Crous & A.D. Hocking, sp. nov.,<br />
Cladophialophora scillae (Deighton) Crous, U. Braun & K. Schub., comb. nov., Cladophialophora sylvestris Crous & de Hoog, sp. nov., Cylindrosympodium lauri Crous &<br />
R.F. Castañeda, sp. nov., Cyphellophora hylomeconis Crous, de Hoog & H.D. Shin, sp. nov., Exophiala eucalyptorum Crous, sp. nov., Fusicladium africanum Crous, sp. nov.,<br />
Fusicladium amoenum (R.F. Castañeda & Dugan) Crous, K. Schub. & U. Braun, comb. nov., Fusicladium brevicatenatum (U. Braun & Feiler) Crous, U. Braun & K. Schub.,<br />
comb. nov., Fusicladium fagi Crous & de Hoog, sp. nov., Fusicladium intermedium (Crous & W.B. Kendr.) Crous, comb. nov., Fusicladium matsushimae (U. Braun & C.F. Hill)<br />
Crous, U. Braun & K. Schub., comb. nov., Fusicladium pini Crous & de Hoog, sp. nov., Fusicladium ramoconidii Crous & de Hoog, sp. nov., Fusicladium rhodense Crous & M.J.<br />
Wingf., sp. nov., Venturia hystrioides (Dugan, R.G. Roberts & Hanlin) Crous & U. Braun, comb. nov.<br />
Key words: Anungitea, Anungitopsis, Cladophialophora, Exophiala, Fusicladium, phylogeny, Pseudocladosporium, systematics, Venturia.<br />
Introduction<br />
Species of Cladophialophora Borelli are relatively simple<br />
hyphomycetes with brown hyphae that give rise to branched chains<br />
of pale brown conidia. Phylogenetically they are defined to belong to<br />
the Chaetothyriales (Haase et al. 1999, Untereiner 2000), an order<br />
containing numerous opportunists (de Hoog et al. 2000); teleomorph<br />
relationships are with Capronia Sacc. in the Herpotrichiellaceae.<br />
In several cases cladophialophora-like synanamorphs are found<br />
accompanying black yeasts of the <strong>genus</strong> Exophiala J.W. Carmich.<br />
(de Hoog et al. 1995). Braun & Feiler (1995) placed several saprobic<br />
hyphomycetes in Cladophialophora, <strong>and</strong> described Capronia<br />
hanliniana U. Braun & Feiler as teleomorph of Cladophialophora<br />
brevicatenata U. Braun & Feiler. This work was continued by Dugan<br />
et al. (1995), who described an additional teleomorph, Capronia<br />
hystrioides Dugan, R.G. Roberts & Hanlin for Cladophialophora<br />
hachijoensis (Matsush.) U. Braun & Feiler. Untereiner (1997)<br />
reduced Capronia hystrioides to synonymy with C. hanliniana,<br />
<strong>and</strong> placed them in Venturia Sacc. (Venturiaceae, Pleosporales).<br />
<strong>The</strong> concept of Cladophialophora hachijoensis, which is based<br />
on Phaeoramularia hachijoensis Matsush. (Matsushima 1975) is<br />
confused, however, <strong>and</strong> phylogenetic studies have revealed that<br />
isolates attributed to this name in recent studies, were in fact<br />
representatives of three different species in phylogenetically distinct<br />
genera (Braun et al. 2003). <strong>The</strong> separation of Cladophialophora<br />
with Capronia teleomorphs (Herpotrichiellaceae, Chaetothyriales;<br />
commonly isolated as human pathogens), from predominantly<br />
saprobic or phytopathogenic isolates in the Dothideomycetes<br />
was recognised by Braun (1998). Recently the cactus endophyte<br />
Cladophialophora yegresii de Hoog was reported to be the nearest<br />
neighbour of C. carrionii (Trejos) de Hoog et al., a major agent<br />
of human chromoblastomycosis (de Hoog et al. 2007), so that<br />
the main distinction between the two anamorph genera remains<br />
in their phylogenetic positions. Capronia hanliniana <strong>and</strong> C.<br />
hystrioides were again recognised as distinct species, <strong>and</strong> placed<br />
in a new <strong>genus</strong>, Caproventuria U. Braun (Venturiaceae), while their<br />
anamorphs were accommodated in Pseudocladosporium U. Braun.<br />
Caproventuria was primarily distinguished from Venturia based on<br />
its distinct Pseudocladosporium anamorphs. Recently, Crous et al.<br />
(2007b) introduced a third <strong>genus</strong>, namely Sympoventuria Crous &<br />
Seifert, which produces a sympodiella-like anamorph in culture. To<br />
complicate matters further, Beck et al. (2005) concluded, based on<br />
an ITS DNA phylogeny, that the morphology attributed to the form<br />
genera Spilocaea Fr., Pollaccia E. Bald. & Cif., <strong>and</strong> Fusicladium<br />
Bonord. has evolved several times within Venturia, <strong>and</strong> that a single<br />
anamorph <strong>genus</strong> should be used for Venturia, namely Fusicladium<br />
(see Schubert et al. 2003 for additional generic synonyms).<br />
In their treatment of Venturia anamorphs, Schubert et al.<br />
(2003) excluded Pseudocladosporium, <strong>and</strong> stated that its status<br />
needs to be confirmed along with that of other genera such as<br />
Anungitea B. Sutton, Fusicladium <strong>and</strong> Polyscytalum Riess. In the<br />
study by Beck et al. (2005) an isolate of Caproventuria hystrioides<br />
(Pseudocladosporium sp.) was included to confirm the link to<br />
the Venturiaceae, though this was not well resolved, nor was the<br />
status of the older generic names mentioned above addressed.<br />
185
Crous et al.<br />
<strong>The</strong> aim of the present study, therefore, was to use DNA sequence<br />
comparisons in conjunction with morphology in an attempt to clarify<br />
these generic issues, as well as to determine which morphological<br />
characters could be used to distinguish Pseudocladosporium from<br />
Cladophialophora.<br />
Materials <strong>and</strong> methods<br />
Isolates<br />
Cultures were obtained from the Centraalbureau voor<br />
Schimmelcultures (<strong>CBS</strong>) in Utrecht, the Netherl<strong>and</strong>s, or isolated<br />
from plant material incubated in moist chambers to promote<br />
sporulation. Isolates were cultured on 2 % malt extract plates<br />
(MEA; Gams et al. 2007), by obtaining single conidial colonies as<br />
explained in Crous (2002). Colonies were subcultured onto fresh<br />
MEA, oatmeal agar (OA), potato-dextrose agar (PDA) <strong>and</strong> synthetic<br />
nutrient-poor agar (SNA) (Gams et al. 2007), <strong>and</strong> incubated at 25<br />
°C under continuous near-ultraviolet light to promote sporulation.<br />
DNA extraction, amplification <strong>and</strong> phylogeny<br />
Fungal colonies were established on agar plates, <strong>and</strong> genomic<br />
DNA was isolated following the CTAB-based protocol described<br />
in Gams et al. (2007). <strong>The</strong> primers V9G (de Hoog & Gerrits van<br />
den Ende 1998) <strong>and</strong> LR5 (Vilgalys & Hester 1990) were used to<br />
amplify part (ITS) of the nuclear rDNA operon spanning the 3’ end<br />
of the 18S rRNA gene (SSU), the first internal transcribed spacer<br />
(ITS1), the 5.8S rRNA gene, the second ITS region <strong>and</strong> the 5’<br />
end of the 28S rRNA gene (LSU). Four internal primers, namely<br />
ITS4 (White et al. 1990), LR0R (Rehner & Samuels 1994), LR3R<br />
(www.biology.duke.edu/fungi/mycolab/primers.htm), <strong>and</strong> LR16<br />
(Moncalvo et al. 1993), were used for sequencing to ensure good<br />
quality overlapping sequences were obtained. <strong>The</strong> PCR conditions,<br />
sequence alignment <strong>and</strong> subsequent phylogenetic analysis followed<br />
the methods of Crous et al. (2006a). <strong>The</strong> ITS1, ITS2 <strong>and</strong> 5.8S rRNA<br />
gene were only sequenced for isolates of which these data were<br />
not available. <strong>The</strong> ITS data were not included in the analyses but<br />
deposited in GenBank where applicable. Gaps longer than 10<br />
bases were coded as single events for the phylogenetic analyses;<br />
the remaining gaps were treated as missing data. Sequence data<br />
were deposited in GenBank (Table 1) <strong>and</strong> alignments in TreeBASE<br />
(www.treebase.org).<br />
Taxonomy<br />
Structures were mounted in lactic acid, <strong>and</strong> 30 measurements<br />
(× 1 000 magnification) determined wherever possible, with the<br />
extremes of spore measurements given in parentheses. Colony<br />
colours (surface <strong>and</strong> reverse) were assessed after 2–4 wk on OA<br />
<strong>and</strong> PDA at 25 °C in the dark, using the colour charts of Rayner<br />
(1970). All cultures obtained in this study are maintained in the <strong>CBS</strong><br />
collection (Table 1). Nomenclatural novelties <strong>and</strong> descriptions were<br />
deposited in MycoBank (www.MycoBank.org).<br />
Results<br />
DNA phylogeny<br />
Amplicons of approximately 1 700 bases were obtained for the<br />
isolates listed in Table 1. <strong>The</strong>se sequences were used to obtain<br />
additional sequences from GenBank which were added to the<br />
alignment. <strong>The</strong> manually adjusted LSU alignment contained 116<br />
sequences (including the two outgroup sequences) <strong>and</strong> 1 157<br />
characters including alignment gaps (available in TreeBASE).<br />
Of the 830 characters used in the phylogenetic analysis, 326<br />
were parsimony-informative, 79 were variable <strong>and</strong> parsimonyuninformative,<br />
<strong>and</strong> 425 were constant. Neighbour-joining analyses<br />
using three substitution models on the sequence data yielded trees<br />
with identical topologies to one another. <strong>The</strong> neighbour-joining trees<br />
support the same clades as obtained from the parsimony analysis,<br />
but with a different arrangement at the deep nodes, for example,<br />
the clade containing Protoventuria alpina (Sacc.) M.E. Barr (<strong>CBS</strong><br />
140.83) is placed as sister to the Venturiaceae using parsimony but<br />
basal to the Herpotrichiellaceae using neighbour-joining. Because<br />
of the large number of different strain associations in the Venturia<br />
clade (see the small number of strict consensus branches for this<br />
clade in Fig. 1), only the first 5 000 equally most parsimonious trees<br />
(TL = 1 752 steps; CI = 0.392; RI = 0.849; RC = 0.333) were saved,<br />
one of which is shown in Fig. 1.<br />
Bayesian analysis was conducted on the same aligned LSU<br />
data set using a general time-reversible (GTR) substitution model<br />
with inverse gamma rates <strong>and</strong> dirichlet base frequencies. <strong>The</strong><br />
Markov Chain Monte Carlo (MCMC) analysis of 4 chains started<br />
from a r<strong>and</strong>om tree topology <strong>and</strong> lasted 2 000 000 generations.<br />
Trees were saved each 1 000 generations, resulting in 2 000 saved<br />
trees. Burn-in was set at 500 000 generations after which the<br />
likelihood values were stationary, leaving 1 500 trees from which<br />
the consensus tree (Fig. 2) <strong>and</strong> posterior probabilities (PP’s) were<br />
calculated. <strong>The</strong> average st<strong>and</strong>ard deviation of split frequencies<br />
was 0.06683 at the end of the run. <strong>The</strong> same overall topology as<br />
that observed using parsimony was obtained, with the exception<br />
of the position of Anungitopsis speciosa R.F. Castañeda & W.B.<br />
Kendr., which is placed between the Leotiomycetes <strong>and</strong> the<br />
Sordariomycetes based on the Bayesian analysis. Also, <strong>similar</strong> to<br />
the results obtained using neighbour-joining, the clade containing<br />
Protoventuria alpina (<strong>CBS</strong> 140.83) is placed as sister to the<br />
Herpotrichiellaceae <strong>and</strong> not to the Venturiaceae. <strong>The</strong> phylogenetic<br />
affinity of specific genera or species are discussed below.<br />
Taxonomy<br />
Several collections represented novel members of the<br />
Herpotrichiellaceae <strong>and</strong> Venturiaceae, <strong>and</strong> these are described<br />
below. Taxa that were cladophialophora- or pseudocladosporiumlike,<br />
but that clustered elsewhere, are treated under excluded<br />
species.<br />
Members of Chaetothyriales, Herpotrichiellaceae<br />
Cladophialophora australiensis Crous & A.D. Hocking, sp. nov.<br />
MycoBank MB504525. Fig. 3.<br />
Etymology: Named after its country of origin, Australia.<br />
Cladophialophorae carrionii similis, sed conidiis secundis majoribus, (7–)8–12(–15)<br />
× 3–4 µm.<br />
In vitro: Mycelium consisting of branched, septate, smooth, pale<br />
brown, guttulate, 2–3 µm wide hyphae; hyphal coils not seen.<br />
Conidiophores dimorphic; macroconidiophores mononematous,<br />
subcylindrical, multi-septate, straight to curved, up to 150 µm long<br />
(including conidiogenous cells), <strong>and</strong> 4 µm wide, pale to medium<br />
brown, smooth, guttulate; microconidiophores integrated with<br />
hyphae, which terminate in subcylindrical conidiogenous cells that<br />
give rise to branched chains of conidia; conidiophores (including<br />
186
Herpotrichiellaceae <strong>and</strong> Venturiaceae<br />
conidiogenous cells) up to 5-septate, 50 µm long, with terminal <strong>and</strong><br />
lateral conidiogenous cells. Conidiogenous cells pale to medium<br />
brown, smooth, guttulate, terminal <strong>and</strong> lateral, subcylindrical,<br />
20–35 × 2–3.5 µm, or reduced to indistinct subtruncate to truncate<br />
loci, scars up to 2 µm wide, mono- to polyblastic, proliferating<br />
sympodially, scars neither darkened, thickened, nor refractive.<br />
Conidia pale to medium brown, guttulate, smooth; ramoconidia<br />
subcylindrical, 0–1-septate, 20–35 × 2–3 µm, hila subtruncate,<br />
inconspicuous, up to 2 µm wide, giving rise to branched chains<br />
of conidia; conidia ellipsoid, pale brown, but becoming dark brown<br />
<strong>and</strong> thick-walled in older cultures, guttulate, tapering towards<br />
subtruncate terminal loci, 0–1-septate, occurring in chains of up to<br />
20 conidia, (7–)8–12(–15) × 3–4 µm (older, dark brown conidia are<br />
ellipsoid, up to 5 µm wide).<br />
Cultural characteristics: Colonies erumpent, somewhat spreading,<br />
margins crenate, feathery, aerial mycelium sparse; colonies on<br />
PDA olivaceous-grey to iron-grey (surface); reverse iron-grey; on<br />
OA <strong>and</strong> SNA olivaceous-grey. Colonies reaching 5 mm diam after 2<br />
wk at 25 °C in the dark; colonies fertile. Not able to grow at 37 °C.<br />
Specimen examined: Australia, isolated from apple juice, Dec. 1986, A.D. Hocking,<br />
holotype <strong>CBS</strong> H-19899, culture ex-type <strong>CBS</strong> 112793 = CPC 1377.<br />
Notes: Cladophialophora australiensis is one of two novel species<br />
of Cladophialophora originally isolated from sports drinks in<br />
Australia. Cladophialophora spp. are commonly associated with<br />
human disorders (Honbo et al. 1984, de Hoog et al. 2000, Levin<br />
et al. 2004), <strong>and</strong> thus their occurrence in sports drinks is cause for<br />
concern. However, none of the new species described here had<br />
the ability to grow at 37 °C, <strong>and</strong> therefore it is not expected that<br />
they could pose a danger to humans. Comparing ITS diversity, the<br />
species shows more than 12 % difference to established pathogens<br />
such as C. carrionii <strong>and</strong> C. bantiana (Sacc.) de Hoog et al.<br />
Cladophialophora chaetospira (Grove) Crous & Arzanlou, comb.<br />
nov. MycoBank MB504526. Fig. 4.<br />
Basionym: Septocylindrium chaetospira Grove, J. Bot. Lond. 24:<br />
199. 1886.<br />
≡ Septonema chaetospira (Grove) S. Hughes, Naturalist, London 840: 9.<br />
1952.<br />
≡ Heteroconium chaetospira (Grove) M.B. Ellis, in Ellis, More Dematiaceous<br />
Hyphomycetes: 64. 1976.<br />
In vitro: Mycelium consisting of branched, septate, smooth,<br />
medium brown hyphae, 2–3.5 µm wide. Conidiophores reduced<br />
to conidiogenous cells, or a single supporting cell, 20–40 × 3–4<br />
µm. Conidiogenous cells subcylindrical, erect, straight to irregularly<br />
curved, medium brown, smooth, 15–30 × 3–4 µm. Conidia in<br />
branched, acropetal chains with up to 30 conidia; subcylindrical to<br />
fusiform, medium brown, smooth, tapering slightly at subtruncate<br />
ends, 1(–3)-septate, thin-walled, becoming slightly constricted at<br />
septa of older conidia, (20–)25–30(–45) × 3–4(–5) µm; conidia<br />
remaining attached in long chains; hila neither thickened, nor<br />
darkened-refractive.<br />
Cultural characteristics: Colonies erumpent, convex, spreading,<br />
with sparse to dense aerial mycelium; margins smooth, undulate;<br />
on PDA iron-grey (surface), margins olivaceous-black; reverse<br />
olivaceous-black; on OA olivaceous-grey in the middle due to fluffy<br />
aerial mycelium, iron-grey in wide outer margin; on SNA olivaceousgrey.<br />
Colonies reaching 12 mm diam after 2 wk on PDA at 25 °C in<br />
the dark. Not able to grow at 37 °C.<br />
Specimens examined: China, Yunnan, Yiliang, isolated from Phyllostachys<br />
bambusoides (Gramineae), decaying bamboo, freshwater, 6 Jul. 2003, L. Cai, <strong>CBS</strong><br />
114747; China, Yunnan, stream in Kunming, isolated from bamboo wood, 15 Jun.<br />
2003, C. Lei, <strong>CBS</strong> 115468. Denmark, isolated from roots of Picea abies (Pinaceae),<br />
isol. by D.S. Malla, <strong>CBS</strong> 491.70. Germany, Schleswig-Holstein, Kiel-Kitzeberg,<br />
isolated from wheat field soil, isol. by W. Gams, <strong>CBS</strong> 514.63 = ATCC 16274 = MUCL<br />
8310.<br />
Notes: Two cultures of Heteroconium chaetospira were originally<br />
deposited as Spadicoides minuta L. Cai, McKenzie & K.D. Hyde (Cai<br />
et al. 2004), but later found to represent Heteroconium chaetospira,<br />
a species commonly found on rotting wood in Europe (Ellis 1976).<br />
<strong>The</strong> <strong>genus</strong> Heteroconium Petr. has in recent years been used<br />
to name leaf spotting fungi with chains of brown, disarticulating<br />
conidia (Crous et al. 2006b), which have phylogenetic affinities<br />
to several orders, obviously being polyphyletic. <strong>The</strong> type species<br />
of Heteroconium, H. citharexyli Petr., is a plant pathogen on<br />
Cytharexylum (Petrak 1949) with hitherto unknown phylogenetic<br />
position. <strong>The</strong> fact that H. chaetospira is linked to the Chaetothyriales,<br />
was rather unexpected. <strong>The</strong> species appears to be <strong>similar</strong> to others<br />
placed in Cladophialophora by having short, lateral conidiogenous<br />
cells, <strong>and</strong> long chains of branched subcylindrical conidia that largely<br />
remain attached. It is, however, quite distinct from other members of<br />
Cladophialophora in having medium brown conidia, <strong>and</strong> in lacking<br />
the ellipsoid conidia observed in several species.<br />
Cladophialophora hostae Crous, U. Braun & H.D. Shin, sp. nov.<br />
MycoBank MB504527. Figs 5–6.<br />
Etymology: Epithet derived from the host <strong>genus</strong>, Hosta.<br />
Cladophialophorae scillae similis, sed conidiophoris in vitro brevioribus et leniter<br />
angustioribus, 10–15 × 1.5–2 µm, conidiis brevioribus, (7–)10–15(–20) µm.<br />
In vivo: Leaf spots amphigenous, subcircular to somewhat angularirregular,<br />
1–5 mm wide, scattered to aggregated, sometimes<br />
confluent, pale to medium brown or with a reddish brown tinge,<br />
later greyish brown, margin indefinite or on the upper leaf surface<br />
with a narrow slightly raised marginal line or very narrow lighter<br />
halo, yellowish, ochraceous to brownish. Caespituli epiphyllous,<br />
punctiform to confluent, dingy greyish brown. Mycelium immersed,<br />
forming fusicladium-like hyphal str<strong>and</strong>s or plates; hyphae septate,<br />
sometimes with constrictions at the septa, thin-walled, pale<br />
olivaceous, 1.5–7 µm wide. Stromata immersed, small, 10–40 µm<br />
diam, composed of swollen hyphal cells, subcircular to somewhat<br />
angular-irregular in outline, 2–8 µm diam, wall somewhat<br />
thickened, brown. Conidiophores in small to moderately large<br />
fascicles, loose, divergent to moderately dense, rarely solitary,<br />
arising from stromatic hyphal aggregations, erumpent, erect,<br />
usually unbranched, rarely branched, straight, subcylindrical to<br />
distinctly geniculate-sinuous, 5–40 × 2–5 µm, 0–6-septate, pale<br />
to medium olivaceous to olivaceous-brown, thin-walled, up to 0.5<br />
µm, smooth. Conidiogenous cells integrated, terminal, 5–15(–20)<br />
µm long, sympodial, conidiogenous loci rather inconspicuous<br />
to subdenticulate, flat-tipped, 1–1.5 µm diam, unthickened or<br />
almost so, not to slightly darkened-refractive. Conidia in simple or<br />
branched chains, narrowly ellipsoid-subcylindrical, 10–15 × 1.5–<br />
3.5 µm, 0–1-septate, subhyaline to pale olivaceous, thin-walled,<br />
smooth, ends truncate or with two denticle-like hila in ramoconidia,<br />
(0.75–)1–1.5(–2) µm diam, unthickened or almost so, at most<br />
slightly darkened-refractive.<br />
In vitro: Mycelium composed of branched, smooth, pale olivaceous<br />
to medium brown hyphae, frequently forming hyphal coils, guttulate,<br />
septa inconspicuous, not constricted, hyphae somewhat irregular<br />
in width, 1–2 µm wide. Conidiophores reduced to conidiogenous<br />
cells, integrated in hyphae, terminal, subcylindrical, pale olivaceous<br />
to pale brown, smooth, 0–1-septate, proliferating sympodially at<br />
www.studiesinmycology.org<br />
187
Crous et al.<br />
10 changes<br />
Athelia epiphylla AY586633<br />
Paullicorticium ansatum AY586693<br />
100<br />
Incertae sedis<br />
Fasciatispora petrakii AY083828<br />
Pidoplitchkoviella terricola AF096197<br />
Anungitopsis speciosa <strong>CBS</strong> 181.95<br />
100<br />
94 100<br />
98<br />
81 97<br />
90<br />
90<br />
87<br />
53<br />
100<br />
94<br />
100<br />
Polyscytalum fecundissimum <strong>CBS</strong> 100506<br />
Phlogicylindrium eucalypti DQ923534<br />
Cyphellophora laciniata <strong>CBS</strong> 190.61<br />
Cladophialophora proteae <strong>CBS</strong> 111667<br />
100<br />
84<br />
82<br />
67<br />
100<br />
Phaeococcomyces catenatus AF050277<br />
Exophiala sp. 3 CPC 12171<br />
Exophiala sp. 3 CPC 12173<br />
Exophiala sp. 3 CPC 12172<br />
Glyphium elatum AF346420<br />
67 100<br />
“<strong>Cladosporium</strong>” adianticola DQ008144<br />
“<strong>Cladosporium</strong>” adianticola DQ008143<br />
Metulocladosporiella musae DQ008162<br />
Metulocladosporiella musicola DQ008159<br />
Thysanorea papuana EU041871<br />
Veronaea botryosa EU041874<br />
Ceramothyrium carniolicum AY004339<br />
Cyphellophora hylomeconis <strong>CBS</strong> 113311<br />
Exophiala eucalyptorum CPC 11261<br />
98<br />
100<br />
100<br />
100<br />
Cladophialophora humicola AF050263<br />
Cladophialophora sylvestris <strong>CBS</strong> 350.83<br />
Cladophialophora hostae CPC 10737<br />
Cladophialophora scillae <strong>CBS</strong> 116461<br />
Exophiala jeanselmei AF050271<br />
Exophiala oligosperma AF050289<br />
Exophiala dermatitidis AF050270<br />
Capronia mansonii AY004338<br />
Capronia munkii AF050250<br />
Ramichloridium mackenziei AF050288<br />
Capronia pilosella AF050254<br />
Exophiala sp. 1 <strong>CBS</strong> 115142<br />
Exophiala sp. 2 <strong>CBS</strong> 115143<br />
53<br />
68<br />
90<br />
Exophiala pisciphila DQ823101<br />
Fonsecaea pedrosoi AF050276<br />
Ramichloridium anceps AF050285<br />
Capronia acutiseta AF050241<br />
Fonsecaea pedrosoi AF050275<br />
Cladophialophora australiensis <strong>CBS</strong> 112793<br />
Cladophialophora potulentorum <strong>CBS</strong> 115144<br />
Cladophialophora potulentorum <strong>CBS</strong> 114772<br />
85 89<br />
98<br />
100<br />
Cladophialophora potulentorum <strong>CBS</strong> 112222<br />
Cladophialophora carrionii AF050262<br />
Phialophora americana AF050283<br />
Phialophora americana AF050280<br />
Cladophialophora chaetospira <strong>CBS</strong> 114747<br />
Cladophialophora chaetospira <strong>CBS</strong> 514.63<br />
Cladophialophora chaetospira <strong>CBS</strong> 491.70<br />
Cladophialophora chaetospira <strong>CBS</strong> 115468<br />
Sordariomycetes,<br />
Xylariomycetidae,<br />
Xylariales<br />
Chaetothyriomycetes, Chaetothyriales, Herpotrichiellaceae<br />
Fig. 1. (Page 188–189). One of 5 000 equally most parsimonious trees obtained from a heuristic search with 100 r<strong>and</strong>om taxon additions of the LSU sequence alignment using<br />
PAUP v. 4.0b10. <strong>The</strong> scale bar shows 10 changes, <strong>and</strong> bootstrap support values from 1 000 replicates are shown at the nodes. Thickened lines indicate the strict consensus<br />
branches <strong>and</strong> ex-type sequences are printed in bold face. <strong>The</strong> tree was rooted to two sequences obtained from GenBank (Athelia epiphylla AY586633 <strong>and</strong> Paullicorticium<br />
ansatum AY586693).<br />
apex via 1–2(–3) flat-tipped, minute, denticle-like loci, 1–1.5 µm<br />
wide, 10–15 × 1.5–2 µm; scars minutely darkened <strong>and</strong> thickened,<br />
but not refractive. Conidia in extremely long chains (–60), simple<br />
or branched, subcylindrical, or narrowly ellipsoid, smooth, pale<br />
olivaceous, 0–1-septate, (7–)10–15(–20) × (1.5–)2(–2.5) µm,<br />
hila truncate, 1–1.5 µm wide, minutely thickened <strong>and</strong> darkenedrefractive.<br />
Cultural characteristics: Colonies on PDA erumpent, spreading,<br />
with smooth, undulate margins <strong>and</strong> dense aerial mycelium; surface<br />
hazel (middle), outer zone isabelline; reverse fuscous-black in<br />
middle, isabelline in outer zone. Colonies reaching 25 mm diam<br />
on SNA, <strong>and</strong> 40 mm diam on PDA after 1 mo at 25 °C in the dark;<br />
colonies fertile.<br />
Specimen examined: Korea, Pyongchang, Hosta plantaginea (Hostaceae), 20 Sep.<br />
2003, H.D. Shin, HAL 2030 F, holotype, culture ex-type SMK 19664, CPC 10737 =<br />
<strong>CBS</strong> 121637, CPC 10738–10739.<br />
Notes: Although this species is morphologically <strong>similar</strong> to<br />
Cladophialophora scillae (Deighton) Crous, U. Braun & K.<br />
Schub. described below in this paper, C. hostae is treated as a<br />
separate taxon due to the differences in the length <strong>and</strong> width of its<br />
conidiophores <strong>and</strong> conidia in vitro, as well as 17 bp differences in<br />
the ITS DNA sequence data <strong>and</strong> a distinct ecology causing leafspots<br />
on a different, unrelated host. Based on disease symptoms<br />
caused on the living host leaves, C. hostae is a very unusual,<br />
unexpected member of the <strong>genus</strong> Cladophialophora. In vivo, the<br />
mycelium forms obvious hyphal str<strong>and</strong>s <strong>and</strong> plates which are<br />
characteristic for Fusicladium species. <strong>The</strong> conidiophores <strong>and</strong><br />
conidia are also fusicladium-like. Nevertheless, this species clusters<br />
within the Herpotrichiellaceae, i.e., it has to be placed in the <strong>genus</strong><br />
Cladophialophora. Biotrophic species like C. hostae <strong>and</strong> C. scillae<br />
without phialidic synanamorphs render the differentiation between<br />
Cladophialophora <strong>and</strong> Fusicladium (incl. Pseudocladosporium)<br />
almost impossible without sequence data. Furthermore, the<br />
188
Herpotrichiellaceae <strong>and</strong> Venturiaceae<br />
10 changes<br />
Satchmopsis brasiliensis DQ195797<br />
Neofabraea malicorticis AY544662<br />
Protoventuria alpina <strong>CBS</strong> 140.83<br />
Clathrosporium intricatum AY616235<br />
Zeloasperisporium hyphopodioides <strong>CBS</strong> 218.95<br />
96 72<br />
94<br />
99<br />
86<br />
71<br />
65<br />
88<br />
59<br />
71<br />
73<br />
91<br />
66<br />
100<br />
99<br />
81<br />
95<br />
100<br />
100<br />
80<br />
Veronaeopsis simplex EU041877<br />
Fusicladium africanum CPC 12828<br />
Fusicladium africanum CPC 12829<br />
Sympoventuria capensis DQ885906<br />
Sympoventuria capensis DQ885904<br />
Sympoventuria capensis DQ885905<br />
100<br />
Fusicladium amoenum <strong>CBS</strong> 254.95<br />
Fusicladium intermedium <strong>CBS</strong> 110746<br />
Fusicladium rhodense CPC 13156<br />
Fusicladium pini <strong>CBS</strong> 463.82<br />
Fusicladium ramoconidii <strong>CBS</strong> 462.82<br />
90<br />
100<br />
Cylindrosympodium lauri <strong>CBS</strong> 240.95<br />
Venturia fraxini <strong>CBS</strong> 374.55<br />
Venturia macularis <strong>CBS</strong> 477.61<br />
Venturia maculiformis <strong>CBS</strong> 377.53<br />
Metacoleroa dickiei DQ384100<br />
Venturia alpina <strong>CBS</strong> 373.53<br />
Fusicladium fagi <strong>CBS</strong> 621.84<br />
Venturia sp. <strong>CBS</strong> 681.74<br />
Caproventuria hanliniana AF050290<br />
Venturia hystrioides <strong>CBS</strong> 117727<br />
Venturia lonicerae <strong>CBS</strong> 445.54<br />
Venturia chlorospora <strong>CBS</strong> 466.61<br />
Fusicladium catenosporum <strong>CBS</strong> 447.91<br />
Venturia helvetica <strong>CBS</strong> 474.61<br />
Venturia minuta <strong>CBS</strong> 478.61<br />
Venturia polygoni-vivipari <strong>CBS</strong> 114207<br />
Venturia atriseda <strong>CBS</strong> 378.49<br />
Venturia viennotii <strong>CBS</strong> 690.85<br />
Venturia anemones <strong>CBS</strong> 370.55<br />
Venturia aceris <strong>CBS</strong> 372.53<br />
Venturia cephalariae <strong>CBS</strong> 372.55<br />
Venturia tremulae var. tremulae <strong>CBS</strong> 257.38<br />
Venturia ditricha <strong>CBS</strong> 118894<br />
Venturia populina <strong>CBS</strong> 256.38<br />
Venturia atriseda <strong>CBS</strong> 371.55<br />
Venturia inaequalis <strong>CBS</strong> 535.76<br />
Fusicladium pomi <strong>CBS</strong> 309.31<br />
Venturia chlorospora <strong>CBS</strong> 470.61<br />
Fusicladium m<strong>and</strong>shuricum <strong>CBS</strong> 112235<br />
Venturia tremulae var. gr<strong>and</strong>identatae <strong>CBS</strong> 695.85<br />
Venturia tremulae var. populi-albae <strong>CBS</strong> 694.85<br />
Fusicladium radiosum <strong>CBS</strong> 112625<br />
Venturia saliciperda <strong>CBS</strong> 214.27<br />
Venturia saliciperda <strong>CBS</strong> 480.61<br />
Fusicladium convolvularum <strong>CBS</strong> 112706<br />
Fusicladium oleagineum <strong>CBS</strong> 113427<br />
Fusicladium phillyreae <strong>CBS</strong> 113539<br />
Venturia nashicola <strong>CBS</strong> 793.84<br />
Venturia pyrina <strong>CBS</strong> 120825<br />
Venturia pyrina <strong>CBS</strong> 331.65<br />
Venturia aucupariae <strong>CBS</strong> 365.35<br />
Venturia crataegi <strong>CBS</strong> 368.35<br />
Apiosporina collinsii CPC 12229<br />
Fusicladium effusum CPC 4524<br />
Fusicladium effusum CPC 4525<br />
60<br />
91<br />
Venturia carpophila AY849967<br />
Fusicladium carpophilum <strong>CBS</strong> 497.62<br />
Venturia cerasi <strong>CBS</strong> 444.54<br />
Leotiomycetes, Helotiales<br />
Dothideomycetes, Pleosporales, Venturiaceae<br />
Fig 1. (Continued).<br />
morphology of C. hostae in vivo <strong>and</strong> in vitro shows remarkable<br />
differences in conidiophore morphology, i.e., the growth in vivo is<br />
characteristically fusicladium-like (conidiophores macronematous,<br />
long, septate), whereas habit in vitro is rather pseudocladosporiumlike<br />
(conidiophores less developed, usually reduced to conidiogenous<br />
cells, short). However, several Fusicladium species have also been<br />
observed to exibit a Pseudocladosporium growth habit in culture,<br />
suggesting this growth plasticity to be rather common, <strong>and</strong> strongly<br />
influenced by growth conditions.<br />
Cladophialophora humicola Crous & U. Braun, sp. nov.<br />
MycoBank MB504528. Figs 7–8.<br />
www.studiesinmycology.org<br />
Etymology: Named after its ecology, namely occurring in soil.<br />
Cladophialophorae bantianae similis, sed conidiis majoribus, (8–)11–14(–17) ×<br />
(1.5–)2(–2.5) µm, locis conidiogenis et hilis angustioribus, 1–1.5 µm latis.<br />
In vitro: Mycelium composed of branched, smooth, pale<br />
olivaceous to pale brown hyphae, frequently forming hyphal coils,<br />
prominently guttulate, not to slightly constricted at the septa, 1–2<br />
µm wide, cells somewhat uneven in width. Conidiophores solitary,<br />
mostly inconspicuous <strong>and</strong> integrated in hyphae, varying from<br />
inconspicuously truncate lateral loci on hyphal cells, 1–1.5 µm wide,<br />
to occasionally terminal conidiophores, 0–3-septate, subcylindrical,<br />
proliferating sympodially, 10–30 × 1.5–3 µm, pale brown, smooth.<br />
189
Crous et al.<br />
Table 1. Isolates used for sequence analysis.<br />
Anamorph Teleomorph Accession number 1 Host Country Collector GenBank numbers 2<br />
Anungitopsis speciosa <strong>CBS</strong> 181.95*; INIFAT C94/135<br />
(ITS, LSU)<br />
Leaf litter of Buchenavia<br />
capitata Cuba R.F. Castañeda EU035401, EU035401<br />
Cladophialophora australiensis <strong>CBS</strong> 112793*; CPC 1377 Sports drink Australia – EU035402, EU035402<br />
Cladophialophora chaetospira <strong>CBS</strong> 114747 Phyllostachys bambusoides China L. Cai EU035403, EU035403<br />
<strong>CBS</strong> 115468; HKUCC 10147 Bamboo China – EU035404, EU035404<br />
<strong>CBS</strong> 491.70 Roots of Picea abies Denmark – EU035405, EU035405<br />
<strong>CBS</strong> 514.63; ATCC 16274; MUCL 8310 Soil, wheat field Germany – EU035406, EU035406<br />
Cladophialophora hostae CPC 10737* Hosta plantaginea Korea H.D. Shin EU035407, EU035407<br />
Cladophialophora humicola <strong>CBS</strong> 117536*; BBA 65570 Soil, arable Germany Z. Zaspel & H. Nirenberg EU035408, AF050263<br />
Cladophialophora potulentorum <strong>CBS</strong> 112222; CPC 1376; FRR 4946 Sports drink Australia N.J. Charley EU035409, EU035409<br />
<strong>CBS</strong> 114772; CPC 1375; FRR 4947 Sports drink Australia N.J. Charley EU035410, EU035410<br />
<strong>CBS</strong> 115144*; CPC 11048; FRR 3318 Apple juice – – DQ008141, DQ008141<br />
Cladophialophora proteae <strong>CBS</strong> 111667*; CPC 1514 Protea cynaroides South Africa L. Viljoen EU035411, EU035411<br />
Cladophialophora scillae <strong>CBS</strong> 116461* Scilla peruviana New Zeal<strong>and</strong> C.F. Hill EU035412, EU035412<br />
Cladophialophora sylvestris <strong>CBS</strong> 350.83 Pinus sylvestris Netherl<strong>and</strong>s – EU035413, EU035413<br />
“<strong>Cladosporium</strong>” adianticola <strong>CBS</strong> 735.87*; ATCC 200931; INIFAT C87/40 Adiantum sp. Cuba R.F. Castañeda & G. Arnold DQ008125, DQ008144<br />
Cylindrosympodium lauri <strong>CBS</strong> 240.95*; INIFAT C95/3-2 Laurus sp.<br />
Spain, Canary<br />
Isl<strong>and</strong>s R.F. Castañeda EU035414, EU035414<br />
Cyphellophora hylomeconis <strong>CBS</strong> 113311* Helomeco velane Korea H.D. Shin EU035415, EU035415<br />
Cyphellophora laciniata <strong>CBS</strong> 190.61*; ATCC 14166; MUCL 9569 Man, skin Switzerl<strong>and</strong> K.M. Wissel EU035416, EU035416<br />
Exophiala eucalyptorum CPC 11261* Eucalyptus sp. New Zeal<strong>and</strong> J. Stalpers EU035417, EU035417<br />
Exophiala sp. 1 <strong>CBS</strong> 115142; CPC 11044; FRR 5582 Fruit-based drink – – DQ008139, EU035418<br />
Exophiala sp. 2 <strong>CBS</strong> 115143*; CPC 11047; FRR 5599 Bottled spring water – – DQ008140, EU035419<br />
Exophiala sp. 3 CPC 12171 Prunus sp. Canada K.A. Seifert EU035420, EU035420<br />
CPC 12172 Prunus sp. Canada K.A. Seifert EU035421, EU035421<br />
CPC 12173 Prunus sp. Canada K.A. Seifert EU035422, EU035422<br />
Fusicladium africanum CPC 12828* Eucalyptus sp. South Africa P.W. Crous EU035423, EU035423<br />
CPC 12829 Eucalyptus sp. South Africa P.W. Crous EU035424, EU035424<br />
Fusicladium amoenum <strong>CBS</strong> 254.95*; ATCC 200947; CPC 3681; IMI 367525; Leaf litter of Eucalyptus Cuba R.F. Castañeda<br />
INIFAT C94/155; MUCL 39143<br />
gr<strong>and</strong>is<br />
EU035425, EU035425<br />
Fusicladium carpophilum Venturia carpophila <strong>CBS</strong> 497.62; ETH 4568 Prunus sp. Switzerl<strong>and</strong> – EU035426, EU035426<br />
190
Herpotrichiellaceae <strong>and</strong> Venturiaceae<br />
Fusicladium catenosporum <strong>CBS</strong> 447.91* Salix tri<strong>and</strong>ra Germany H. Butin EU035427, EU035427<br />
Fusicladium convolvularum <strong>CBS</strong> 112706*; CPC 3884; IMI 383037 Convolvulus arvensis New Zeal<strong>and</strong> C.F. Hill AY251082, EU035428<br />
Fusicladium effusum CPC 4524 Carya illinoinensis U.S.A. K. Stevenson AY251084, EU035429<br />
CPC 4525 Carya illinoinensis U.S.A. K. Stevenson AY251085, EU035430<br />
Fusicladium fagi <strong>CBS</strong> 621.84*; ATCC 200937 Fagus sylvatica Netherl<strong>and</strong>s G.S. de Hoog EU035431, EU035431<br />
Fusicladium intermedium <strong>CBS</strong> 110746*; CPC 778; IMI 362702 Eucalyptus sp. Madagascar P.W. Crous EU035432, EU035432<br />
Fusicladium m<strong>and</strong>shuricum Venturia m<strong>and</strong>shurica <strong>CBS</strong> 112235*; CPC 3639 Populus simonii China – EU035433, EU035433<br />
Fusicladium oleagineum <strong>CBS</strong> 113427 Olea europaea New Zeal<strong>and</strong> C.F. Hill EU035434, EU035434<br />
Fusicladium phillyreae <strong>CBS</strong> 113539; UPSC 1329 – Portugal B. d’Oliveira EU035435, EU035435<br />
Fusicladium pini <strong>CBS</strong> 463.82* Pinus sylvestris Netherl<strong>and</strong>s G.S. de Hoog EU035436, EU035436<br />
Fusicladium pomi Venturia inaequalis <strong>CBS</strong> 309.31 – – – EU035437, EU035437<br />
<strong>CBS</strong> 535.76 Sorbus aria Switzerl<strong>and</strong> – EU035460, EU035460<br />
Fusicladium radiosum Venturia tremulae <strong>CBS</strong> 112625; CPC 3638 Populus tremula France – EU035438, EU035438<br />
Fusicladium ramoconidii <strong>CBS</strong> 462.82*; CPC 3679 Pinus sp. Netherl<strong>and</strong>s G.S. de Hoog AY251086, EU035439<br />
Fusicladium rhodense CPC 13156* Ceratonia siliqua Greece P.W. Crous EU035440, EU035440<br />
Polyscytalum fecundissimum <strong>CBS</strong> 100506 Fagus sylvatica Netherl<strong>and</strong>s W. Gams EU035441, EU035441<br />
Zeloasperisporium hyphopodioides <strong>CBS</strong> 218.95*; IMI 367520; INIFAT C94/114; MUCL 39155 Air Cuba R. F. Castañeda EU035442, EU035442<br />
Apiosporina collinsii CPC 12229 Amelanchier alnifolia Canada L.J. Hutchinson EU035443, EU035443<br />
Protoventuria alpina <strong>CBS</strong> 140.83 Arctostaphylos uva-ursi Switzerl<strong>and</strong> – EU035444, EU035444<br />
Sympoventuria capensis <strong>CBS</strong> 120136; CPC 12838 Eucalyptus sp. South Africa P.W. Crous DQ885906, DQ885906<br />
CPC 12839 Eucalyptus sp. South Africa P.W. Crous DQ885905, DQ885905<br />
CPC 12840 Eucalyptus sp. South Africa P.W. Crous DQ885904, DQ885904<br />
Venturia aceris <strong>CBS</strong> 372.53 Acer pseudoplatanus Switzerl<strong>and</strong> – EU035445, EU035445<br />
Venturia alpina <strong>CBS</strong> 373.53 Arctostaphylos alpina Switzerl<strong>and</strong> – EU035446, EU035446<br />
Venturia anemones <strong>CBS</strong> 370.55; IMI 163998 Anemone alpina France – EU035447, EU035447<br />
Venturia atriseda <strong>CBS</strong> 371.55 Gentiana punctata Switzerl<strong>and</strong> – EU035448, EU035448<br />
<strong>CBS</strong> 378.49 Gentiana lutea Switzerl<strong>and</strong> J.A. von Arx EU035449, EU035449<br />
Venturia aucupariae <strong>CBS</strong> 365.35; IMI 163987 Sorbus aucuparia Germany – EU035450, EU035450<br />
Venturia cephalariae <strong>CBS</strong> 372.55 Cephalaria alpina Switzerl<strong>and</strong> – EU035451, EU035451<br />
Venturia cerasi <strong>CBS</strong> 444.54; ATCC 12119; IMI 163988 Prunus cerasus Germany – EU035452, EU035452<br />
Venturia chlorospora <strong>CBS</strong> 466.61; ETH 543 Salix caesia Switzerl<strong>and</strong> J. Nüesch EU035453, EU035453<br />
<strong>CBS</strong> 470.61 Salix daphnoides France J. Nüesch EU035454, EU035454<br />
Venturia crataegi <strong>CBS</strong> 368.35 Crataegus sp. Germany – EU035455, EU035455<br />
www.studiesinmycology.org<br />
191
Crous et al.<br />
Table 1. (Continued).<br />
Anamorph Teleomorph Accession number 1 Host Country Collector GenBank numbers 2<br />
Venturia ditricha <strong>CBS</strong> 118894 Betula pubescens var.<br />
tortuosa<br />
(ITS, LSU)<br />
Finl<strong>and</strong> – EU035456, EU035456<br />
Venturia fraxini <strong>CBS</strong> 374.55 Fraxinus excelsior Switzerl<strong>and</strong> – EU035457, EU035457<br />
Venturia helvetica <strong>CBS</strong> 474.61; ETH 2571; IMI 163990 Salix helvetica Switzerl<strong>and</strong> J. Nüesch EU035458, EU035458<br />
Venturia hystrioides <strong>CBS</strong> 117727*; ATCC 96019; CPC 5391 Prunus avium cv. Bing U.S.A. R.G. Roberts EU035459, EU035459<br />
Venturia lonicerae <strong>CBS</strong> 445.54; IMI 163997 Lonicera coerulea Switzerl<strong>and</strong> – EU035461, EU035461<br />
Venturia macularis <strong>CBS</strong> 477.61; ETH 2831 Populus tremula France – EU035462, EU035462<br />
Venturia maculiformis <strong>CBS</strong> 377.53 Epilobium montanum France – EU035463, EU035463<br />
Venturia minuta <strong>CBS</strong> 478.61; ETH 523; IMI 163991 Salix nigricans Switzerl<strong>and</strong> J. Nüesch EU035464, EU035464<br />
Venturia nashicola <strong>CBS</strong> 793.84 Pyrus serotina Japan – EU035465, EU035465<br />
Venturia polygoni-vivipari <strong>CBS</strong> 114207; UPSC 2754 Polygonum viviparum Norway K. & L. Holm EU035466, EU035466<br />
Venturia populina <strong>CBS</strong> 256.38; IMI 163996 Populus canadensis Italy – EU035467, EU035467<br />
Venturia pyrina <strong>CBS</strong> 120825 Pyrus communis Brazil – EU035468, EU035468<br />
<strong>CBS</strong> 331.65 Pyrus sp. – – EU035469, EU035469<br />
Venturia saliciperda <strong>CBS</strong> 214.27; IMI 163993 – – – EU035470, EU035470<br />
<strong>CBS</strong> 480.61; ETH 2836 Salix cordata Switzerl<strong>and</strong> – EU035471, EU035471<br />
Venturia sp. <strong>CBS</strong> 681.74 Cedrus atlantica France W. Gams EU035472, EU035472<br />
Venturia tremulae var. gr<strong>and</strong>identatae <strong>CBS</strong> 695.85 Populus tremuloides Canada – EU035473, EU035473<br />
Venturia tremulae var. populi-albae <strong>CBS</strong> 694.85 Populus alba France – EU035474, EU035474<br />
Venturia tremulae var. tremulae <strong>CBS</strong> 257.38 Populus tremula Italy – EU035475, EU035475<br />
Venturia viennotii <strong>CBS</strong> 690.85 Populus tremula France – EU035476, EU035476<br />
1 ATCC: American Type Culture Collection, Virginia, U.S.A.; BBA: Biologische Bundesanstalt für L<strong>and</strong>- und Forstwirtschaft, Berlin-Dahlem, Germany; <strong>CBS</strong>: Centraalbureau voor Schimmelcultures, Utrecht, <strong>The</strong> Netherl<strong>and</strong>s; CPC: Culture collection of Pedro<br />
Crous, housed at <strong>CBS</strong>; ETH: Eidgenössische Technische Hochschule, Institute for Special Botany, Zürich, Switzerl<strong>and</strong>; FRR: Division of Food Research, CSIRO, North Ryde, N.S.W., Australia; HKUCC: <strong>The</strong> University of Hong Kong Culture Collection, Dept.<br />
of Ecology <strong>and</strong> Biodiversity, University of Hong Kong, Pokfulam Road, China; IMI: International Mycological Institute, CABI-Bioscience, Egham, Bakeham Lane, U.K.; INIFAT: Alex<strong>and</strong>er Humboldt Institute for Basic Research in Tropical Agriculture, Ciudad<br />
de La Habana, Cuba; MUCL: Mycotheque de l’ Université Catholique de Louvain, Louvain-la-Neuve, Belgium; UPSC: Uppsala University Culture Collection of Fungi, Museum of Evolution, Botany Section, Evolutionary Biology Centre, Uppsala, Sweden.<br />
2 ITS: internal transcribed spacer regions, LSU: partial 28S rDNA sequence.<br />
*Ex-type cultures.<br />
192
Herpotrichiellaceae <strong>and</strong> Venturiaceae<br />
0.1 expected changes per site<br />
Athelia epiphylla AY586633<br />
Paullicorticium ansatum AY586693<br />
0.92 0.98 Neofabraea malicorticis AY544662<br />
Protoventuria alpina <strong>CBS</strong> 140.83<br />
0.94 Clathrosporium intricatum AY616235<br />
Leotiomycetes, Helotiales<br />
Satchmopsis brasiliensis DQ195797<br />
1.00<br />
Anungitopsis speciosa <strong>CBS</strong> 181.95 Incertae sedis<br />
0.98 1.00 Fasciatispora petrakii AY083828<br />
Sordariomycetes,<br />
Pidoplitchkoviella terricola AF096197<br />
0.98 Polyscytalum fecundissimum <strong>CBS</strong> 100506 Xylariomycetidae,<br />
1.00 Phlogicylindrium eucalypti DQ923534 Xylariales<br />
1.00<br />
0.94<br />
Cladophialophora humicola AF050263<br />
0.84 Cladophialophora sylvestris <strong>CBS</strong> 350.83<br />
1.00 Cladophialophora hostae CPC 10737<br />
Cladophialophora scillae <strong>CBS</strong> 116461<br />
0.50 Cladophialophora proteae <strong>CBS</strong> 111667<br />
Phaeococcomyces catenatus AF050277<br />
1.00 Glyphium elatum AF346420<br />
1.00 Exophiala sp. 3 CPC 12171<br />
1.00 Exophiala sp. 3 CPC 12173<br />
1.00 Exophiala sp. 3 CPC 12172<br />
“<strong>Cladosporium</strong>” adianticola DQ008144<br />
1.00<br />
1.00<br />
1.00<br />
0.85<br />
0.56<br />
1.00<br />
1.00<br />
1.00<br />
“<strong>Cladosporium</strong>” adianticola DQ008143<br />
Metulocladosporiella musae DQ008162<br />
Metulocladosporiella musicola DQ008159<br />
Thysanorea papuana EU041871<br />
Veronaea botryosa EU041874<br />
Exophiala jeanselmei AF050271<br />
Exophiala oligosperma AF050289<br />
Ramichloridium mackenziei AF050288<br />
Ramichloridium anceps AF050285<br />
Capronia pilosella AF050254<br />
Capronia acutiseta AF050241<br />
Exophiala dermatitidis AF050270<br />
Capronia mansonii AY004338<br />
Capronia munkii AF050250<br />
0.71<br />
1.00<br />
0.81 Exophiala pisciphila DQ823101<br />
0.56 Fonsecaea pedrosoi AF050276<br />
0.98 Exophiala sp. 1 <strong>CBS</strong> 115142<br />
0.60 Exophiala sp. 2 <strong>CBS</strong> 115143<br />
0.80 Cyphellophora laciniata <strong>CBS</strong> 190.61<br />
Ceramothyrium carniolicum AY004339<br />
1.00 Cyphellophora hylomeconis <strong>CBS</strong> 113311<br />
1.00 Exophiala eucalyptorum CPC 11261<br />
Fonsecaea pedrosoi AF050275<br />
0.77 0.68<br />
Cladophialophora australiensis <strong>CBS</strong> 112793<br />
Cladophialophora potulentorum <strong>CBS</strong> 115144<br />
1.00 Cladophialophora potulentorum <strong>CBS</strong> 114772<br />
1.00 Cladophialophora potulentorum <strong>CBS</strong> 112222<br />
1.00<br />
0.95<br />
1.00<br />
1.00<br />
1.00<br />
Cladophialophora carrionii AF050262<br />
Phialophora americana AF050283<br />
Phialophora americana AF050280<br />
Cladophialophora chaetospira <strong>CBS</strong> 114747<br />
Cladophialophora chaetospira <strong>CBS</strong> 514.63<br />
Cladophialophora chaetospira <strong>CBS</strong> 491.70<br />
Cladophialophora chaetospira <strong>CBS</strong> 115468<br />
Fig. 2. (Page 193–194). Consensus phylogram (50 % majority rule) of 1 500 trees resulting from a Bayesian analysis of the LSU sequence alignment using MrBayes v. 3.1.2.<br />
Bayesian posterior probabilities are indicated at the nodes. Ex-type sequences are printed in bold face. <strong>The</strong> tree was rooted to two sequences obtained from GenBank (Athelia<br />
epiphylla AY586633 <strong>and</strong> Paullicorticium ansatum AY586693).<br />
Chaetothyriomycetes, Chaetothyriales, Herpotrichiellaceae<br />
Conidiogenous cells integrated, inconspicuous, truncate, lateral<br />
loci 1–1.5 µm wide, or conidiogenous cells subcylindrical with 1–3<br />
sympodial loci (which appear as minute lateral denticles), 7–17 ×<br />
1.5–2 µm; scars inconspicuous, neither darkened, refractive nor<br />
thickened. Conidia in short chains of up to 10, simple or branched,<br />
subcylindrical to narrowly ellipsoid, 0–1-septate, (8–)11–14(–17)<br />
× (1.5–)2(–2.5) µm, pale olivaceous to olivaceous-brown or pale<br />
brown, smooth, hila truncate, 1–1.5 µm wide, unthickened, neither<br />
darkened, nor refractive.<br />
Cultural characteristics: Colonies erumpent, spreading, with<br />
uneven, feathery margins <strong>and</strong> dense aerial mycelium on PDA;<br />
pale olivaceous-grey in the middle, becoming olivaceous-grey<br />
in the outer zone (surface); reverse olivaceous-black, with greyolivaceous<br />
margins. Colonies reaching 7 mm diam after 2 wk at 25<br />
°C in the dark; colonies fertile.<br />
Specimen examined: Germany, Br<strong>and</strong>enburg, Müncheberg, from soil, Zaspel,<br />
Zalf & H. Nirenberg, holotype <strong>CBS</strong> H-19902, culture ex-type BBA 65570 = <strong>CBS</strong><br />
117536.<br />
Notes: Phylogenetically Cladophialophora humicola is closely<br />
related to C. sylvestris Crous & de Hoog (see below). Morphologically<br />
the two species can be distinguished in that C. humicola lacks<br />
ramoconidia, <strong>and</strong> has 1-septate conidia, while those of C. sylvestris<br />
are 0–3-septate.<br />
Cladophialophora potulentorum Crous & A.D. Hocking, sp. nov.<br />
MycoBank MB504529. Figs 9–10.<br />
Etymology: Refers to its presence in fruit juices <strong>and</strong> sports drinks.<br />
Cladophialophorae carrionii similis, sed conidiis secundis majoribus, (6–)8–10(–13)<br />
× 2–3 µm.<br />
In vitro: Mycelium consisting of branched, septate, smooth,<br />
pale brown, guttulate, 1.5–2.5 µm wide hyphae. Conidiophores<br />
solitary, macronematous, well distinguishable under the dissecting<br />
microscope from aerial mycelium, pale to medium brown,<br />
subcylindrical, straight to somewhat curved, erect, with apical<br />
apparatus appearing as a tuft due to extremely long conidial<br />
www.studiesinmycology.org<br />
193
Crous et al.<br />
0.1 expected changes per site<br />
Cylindrosympodium lauri <strong>CBS</strong> 240.95<br />
1.00 Fusicladium amoenum <strong>CBS</strong> 254.95<br />
1.00 Fusicladium intermedium <strong>CBS</strong> 110746<br />
1.00<br />
Fusicladium rhodense CPC 13156<br />
Fusicladium pini <strong>CBS</strong> 463.82<br />
1.00 1.00 1.00 Fusicladium ramoconidii <strong>CBS</strong> 462.82<br />
1.00<br />
Veronaeopsis simplex EU041877<br />
Fusicladium africanum CPC 12828<br />
Fusicladium africanum CPC 12829<br />
0.85<br />
0.95<br />
0.78<br />
0.56<br />
0.71<br />
1.00<br />
0.69<br />
0.79<br />
0.94<br />
0.56<br />
0.97<br />
0.91<br />
1.00<br />
0.97<br />
1.00<br />
1.00<br />
0.68<br />
0.55<br />
0.65<br />
0.61<br />
0.92<br />
0.94<br />
0.72<br />
0.95<br />
Sympoventuria capensis DQ885906<br />
Sympoventuria capensis DQ885904<br />
Sympoventuria capensis DQ885905<br />
Zeloasperisporium hyphopodioides <strong>CBS</strong> 218.95<br />
Metacoleroa dickiei DQ384100<br />
Venturia alpina <strong>CBS</strong> 373.53<br />
Venturia fraxini <strong>CBS</strong> 374.55<br />
Venturia macularis <strong>CBS</strong> 477.61<br />
Venturia maculiformis <strong>CBS</strong> 377.53<br />
Apiosporina collinsii CPC 12229<br />
Fusicladium fagi <strong>CBS</strong> 621.84<br />
Venturia sp. <strong>CBS</strong> 681.74<br />
Caproventuria hanliniana AF050290<br />
Venturia hystrioides <strong>CBS</strong> 117727<br />
Venturia viennotii <strong>CBS</strong> 690.85<br />
Venturia inaequalis <strong>CBS</strong> 535.76<br />
Venturia saliciperda <strong>CBS</strong> 214.27<br />
Fusicladium radiosum <strong>CBS</strong> 112625<br />
Venturia saliciperda <strong>CBS</strong> 480.61<br />
Venturia tremulae var. populi-albae <strong>CBS</strong> 694.85<br />
Venturiatremulae var. gr<strong>and</strong>identatae <strong>CBS</strong> 695.85<br />
Fusicladium m<strong>and</strong>shuricum <strong>CBS</strong> 112235<br />
Fusicladium pomi <strong>CBS</strong> 309.31<br />
Venturia atriseda <strong>CBS</strong> 371.55<br />
Venturia populina <strong>CBS</strong> 256.38<br />
Venturia chlorospora <strong>CBS</strong> 470.61<br />
Venturia ditricha <strong>CBS</strong> 118894<br />
Venturia tremulae var. tremulae <strong>CBS</strong> 257.38<br />
Fusicladium catenosporum <strong>CBS</strong> 447.91<br />
Venturia helvetica <strong>CBS</strong> 474.61<br />
Venturia minuta <strong>CBS</strong> 478.61<br />
Venturia chlorospora <strong>CBS</strong> 466.61<br />
Venturia polygoni-vivipari <strong>CBS</strong> 114207<br />
Venturia lonicerae <strong>CBS</strong> 445.54<br />
Venturia carpophila AY849967<br />
Fusicladium carpophilum <strong>CBS</strong> 497.62<br />
Venturia cerasi <strong>CBS</strong> 444.54<br />
Venturia atriseda <strong>CBS</strong> 378.49<br />
Venturia anemones <strong>CBS</strong> 370.55<br />
Venturia aceris <strong>CBS</strong> 372.53<br />
Venturia cephalariae <strong>CBS</strong> 372.55<br />
Fusicladium convolvularum <strong>CBS</strong> 112706<br />
Venturia aucupariae <strong>CBS</strong> 365.35<br />
Venturia crataegi <strong>CBS</strong> 368.35<br />
Venturia nashicola <strong>CBS</strong> 793.84<br />
Venturia pyrina <strong>CBS</strong> 120825<br />
Venturia pyrina <strong>CBS</strong> 331.65<br />
Fusicladium effusum CPC 4524<br />
Fusicladium effusum CPC 4525<br />
Fusicladium oleagineum <strong>CBS</strong>113427<br />
Fusicladium phillyreae <strong>CBS</strong> 113539<br />
Dothideomycetes, Pleosporales, Venturiaceae<br />
Fig. 2. (Continued).<br />
Fig. 3. Cladophialophora australiensis (<strong>CBS</strong> 112793). A. Conidiophore. B–C. Subcylindrical ramoconidia, <strong>and</strong> ellipsoid conidia. Scale bar = 10 µm.<br />
194
Herpotrichiellaceae <strong>and</strong> Venturiaceae<br />
Fig. 4. Cladophialophora chaetospira (<strong>CBS</strong> 114747). A–C. Hyphae giving rise to conidiophores with catenulate conidia. D–F. Conidia become up to 3-septate, frequently<br />
remaining attached in chains. Scale bars = 10 µm.<br />
Fig. 5. Cladophialophora hostae (CPC 10737). A–B. Conidiogenous loci (arrows). C. Hyphal coil. D–F. Branched conidial chains. G–H. Conidia. Scale bar = 10 µm.<br />
www.studiesinmycology.org<br />
195
Crous et al.<br />
Fig. 6. Cladophialophora hostae (CPC 10737). Branched conidial chains with<br />
ramoconidia <strong>and</strong> conidia. Scale bar = 10 µm.<br />
Fig. 7. Cladophialophora humicola (<strong>CBS</strong> 117536). Conidiophore with branched<br />
conidial chains. Scale bar = 10 µm.<br />
Fig. 8. Cladophialophora humicola (<strong>CBS</strong> 117536). A. Hyphal coil. B. Conidiophore. C–F. Conidial chains with ramoconidia <strong>and</strong> conidia. Scale bar = 10 µm.<br />
196
Herpotrichiellaceae <strong>and</strong> Venturiaceae<br />
Fig. 9. Cladophialophora potulentorum (<strong>CBS</strong> 115144). A. Colony on PDA. B. Conidiophore. C–D. Conidial chains. E–F. Ramoconidia <strong>and</strong> conidia. Scale bar = 10 µm.<br />
chains; conidiophores up to 5-septate, <strong>and</strong> 100 µm tall (excluding<br />
conidiogenous cells). Conidiogenous cells pale brown, smooth,<br />
terminal <strong>and</strong> lateral, subcylindrical, tapering towards subtruncate<br />
to truncate loci, 1 µm wide, somewhat darkened, thickened, but not<br />
refractive, loci appearing subdenticulate on lateral conidiogenous<br />
cells, mono- to polyblastic, proliferating sympodially, 10–35 × 1.5–2<br />
µm. Conidia pale brown, smooth, guttulate, occurring in branched<br />
chains of up to 60; hila somewhat darkened <strong>and</strong> thickened, but not<br />
refractive, 0.5 µm wide; ramoconidia subcylindrical, 0–1-septate,<br />
15–17(–20) × 2.5–3 µm; conidia ellipsoid, (6–)8–10(–13) × 2–3<br />
µm.<br />
Cultural characteristics: Colonies erumpent, spreading, with smooth<br />
margins <strong>and</strong> dense aerial mycelium on PDA, olivaceous-grey<br />
(surface), with a thin, olivaceous-black margin; reverse olivaceousblack;<br />
on OA olivaceous-grey (surface) with a wide olivaceousblack<br />
margin. Colonies reaching 25–30 mm diam after 1 mo at 25<br />
°C in the dark; colonies fertile, also sporulating in the agar. Not able<br />
to grow at 37 °C.<br />
Specimens examined: Australia, isolated from apple juice drink, Dec. 1986, A.D.<br />
Hocking, holotype <strong>CBS</strong> H-19901, culture ex-type <strong>CBS</strong> 115144 = CPC 11048;<br />
Australia, isolated from sports drink, Feb. 1996, A.D. Hocking, <strong>CBS</strong> 114772 = CPC<br />
1375 = FRR 4947; Australia, isolated from sports drink, Feb. 1996, A.D. Hocking,<br />
<strong>CBS</strong> 112222 = FRR 4946.<br />
Fig. 10. Cladophialophora potulentorum (<strong>CBS</strong> 115144). Conidiophores with chains<br />
of ramoconidia <strong>and</strong> conidia. Scale bar = 10 µm.<br />
www.studiesinmycology.org<br />
Notes: Originally this taxon, isolated from fruit <strong>and</strong> sports drinks,<br />
was thought to be an undescribed species of Pseudocladosporium<br />
(= Fusicladium, see below). However, upon closer examination, this<br />
197
Crous et al.<br />
Fig. 11. Cladophialophora proteae (<strong>CBS</strong> 111667). A. Colony on OA. B–C. Conidiophores. D–H. Catenulate conidia. Scale bars = 10 µm.<br />
proved not to be the case. Conidiophores appear as distinct tufts<br />
under the dissecting microscope, <strong>and</strong> are readily distinguishable<br />
from the superficial mycelium, as is normally observed in species<br />
of Fusicladium, but the conidial chains are extremely long, <strong>and</strong> the<br />
conidia tend to be more ellipsoid than the predominantly fusiform or<br />
subcylindrical conidia observed in species of Fusicladium. Hyphal<br />
coils were also not observed in cultures of C. potulentorum, but<br />
are rather common in species of Fusicladium. <strong>The</strong> phylogenetic<br />
position of this taxon within the Herpotrichiellaceae clade also<br />
supports inclusion in the <strong>genus</strong> Cladophialophora.<br />
Cladophialophora proteae Viljoen & Crous, S. African J. Bot. 64:<br />
137. 1998. Fig. 11.<br />
≡ Pseudocladosporium proteae (Viljoen & Crous) Crous, in Crous et al.,<br />
Cultivation <strong>and</strong> Diseases of Proteaceae: Leucadendron, Leucospermum<br />
<strong>and</strong> Protea: 101. 2004.<br />
In vitro: Mycelium consisting of branched, septate hyphae, often<br />
forming str<strong>and</strong>s, anastomosing, smooth to finely verruculose,<br />
frequently constricted at septa, olivaceous, 3–4 µm wide; hyphal<br />
cells in older cultures becoming swollen, up to 6 µm wide.<br />
Conidiophores reduced to conidiogenous cells. Conidiogenous<br />
cells holoblastic, integrated, forming short, truncate protuberances,<br />
2–3 × 1.5–2 µm, concolorous with mycelium, subcylindrical.<br />
Conidia in vitro arranged in long acropetal chains (up to 20), simple<br />
or branched, subcylindrical to oblong-doliiform, (9–)13–17(–22) ×<br />
2.5–3(–4) µm in vitro on MEA, (9–)16–22(–25) × (2.5–)3–4(–6)<br />
µm on SNA; 0–1(–4)-septate, pale brown to pale olivaceous,<br />
smooth, hila subtruncate to truncate, not thickened, but somewhat<br />
refractive.<br />
Cultural characteristics: Colonies erumpent, with sparse aerial<br />
mycelium on PDA; margins irregular, feathery; greyish rose, with<br />
patches of pale olivaceous-grey (surface); reverse olivaceous-grey.<br />
Colonies reaching 10 mm diam after 2 wk at 25 °C in the dark;<br />
colonies fertile.<br />
Specimen examined: South Africa, Western Cape Province, Stellenbosch, J.S.<br />
Marais Nature Reserve, leaves of Protea cynaroides (Proteaceae), 26 Aug. 1996, L.<br />
Viljoen, holotype PREM 55345, culture ex-type <strong>CBS</strong> 111667.<br />
Notes: Cladophialophora proteae differs from species of Fusicladium<br />
(= Pseudocladosporium) based on its colony colour, the slimy<br />
nature of colonies, as well as its conidia that have inconspicuous,<br />
unthickened hila (Fig. 11) (Crous et al. 2004), unlike those observed<br />
in species of Fusicladium. Sequence data show that this species is<br />
not allied to the Venturiaceae, but to the Herpotrichiellaceae.<br />
Cladophialophora scillae (Deighton) Crous, U. Braun & K. Schub.,<br />
comb. nov. MycoBank MB504530. Fig. 12.<br />
Basionym: <strong>Cladosporium</strong> scillae Deighton, N. Zeal<strong>and</strong> J. Bot. 8:<br />
55. 1970.<br />
≡ Fusicladium scillae (Deighton) U. Braun & K. Schub., IMI Descriptions of<br />
Fungi <strong>and</strong> Bacteria 152: 1518. 2002.<br />
198
Herpotrichiellaceae <strong>and</strong> Venturiaceae<br />
Fig. 12. Cladophialophora scillae (<strong>CBS</strong> 116461). A–C. Conidiophores. D–F. Catenulate conidia. Scale bar = 10 µm.<br />
In vivo: see Schubert & Braun (2002a) <strong>and</strong> Schubert et al. (2003).<br />
In vitro: Mycelium consisting of branched, septate, smooth, greenbrown<br />
to medium brown, guttulate hyphae, variable in width, 1.5–3<br />
µm diam. Conidiophores lateral or terminal on hyphae, erect,<br />
straight to slightly flexuous, solitary, in some cases aggregated,<br />
subcylindrical, curved to geniculate-sinuous, unbranched, up to 55<br />
µm long, 2–3 µm wide, 0–7-septate, septa in short succession,<br />
pale to medium brown, somewhat paler towards apices, smooth.<br />
Conidiogenous cells integrated, terminal or lateral as individual loci<br />
on hyphal cells, straight to curved, subcylindrical, up to 14(–18) µm<br />
long <strong>and</strong> 2 µm wide, pale to medium brown, smooth, with a single or<br />
few subdenticulate to denticulate loci at the apex due to sympodial<br />
proliferation, or reduced to individual loci, 0.8–1.5(–2) µm wide;<br />
scars minutely thickened <strong>and</strong> darkened, but not refractive. Conidia<br />
occurring in long, unbranched or loosely branched chains (–30),<br />
straight to slightly curved, ellipsoid to mostly narrowly subcylindrical,<br />
obclavate in some larger, septate conidia, (5–)10–20(–35) × 1.5–3<br />
µm, 0–1(–3)-septate, sometimes slightly constricted at the septa,<br />
subhyaline to pale brown, smooth, guttulate, tapering at ends<br />
to subtruncate hila, 0.8–1.5 µm wide, minutely thickened <strong>and</strong><br />
darkened, but not refractive; microcyclic conidiogenesis occurring.<br />
Cultural characteristics: Colonies erumpent, spreading, with<br />
smooth, even margins <strong>and</strong> dense, abundant aerial mycelium on<br />
PDA; grey-olivaceous (surface); reverse dark olivaceous. Colonies<br />
on OA olivaceous-grey, smoke-grey due to profuse sporulation,<br />
www.studiesinmycology.org<br />
reverse olivaceous-grey to iron-grey, velvety, aerial mycelium<br />
sparse, diffuse. Colonies reaching 20 mm diam on SNA, <strong>and</strong> 40<br />
mm on PDA after 1 mo at 25 °C in the dark; colonies fertile.<br />
Specimens examined: New Zeal<strong>and</strong>, Levin, on Scilla peruviana (Hyacinthaceae),<br />
21 Dec. 1965, G.F. Laudon, IMI 116997 holotype; Auckl<strong>and</strong>, Manurewa, Auckl<strong>and</strong><br />
Botanic Gardens, on leaf spots of Scilla peruviana, 25 Apr. 2004, C.F. Hill, 1044,<br />
<strong>CBS</strong> H-19903, epitype designated here, culture ex-type <strong>CBS</strong> 116461.<br />
Notes: In culture Cladophialophora scillae forms a<br />
pseudocladosporium-like state, though the scars are somewhat<br />
darkened <strong>and</strong> thickened, but not refractive. Conidiophores are<br />
reduced to conidiogenous cells that are integrated in the mycelium,<br />
terminal or lateral, frequently also as an inconspicuous lateral<br />
denticle, with a flat-tipped scar. Conidia occur in long, branched<br />
chains, which are subcylindrical to narrowly ellipsoid, <strong>and</strong> are<br />
up to 35 µm long, 1.5–3 µm wide, thus longer <strong>and</strong> thinner than<br />
reported on the host, which were 0–3-septate, subcylindrical to<br />
ellipsoid-ovoid, 7–22 × 2.5–4 µm. Due to the fusicladioid habit of<br />
this species in vivo, Schubert & Braun (2002a) reallocated it to<br />
Fusicladium. Based on ITS sequence data, morphology <strong>and</strong> cultural<br />
characteristics, Cladophialophora scillae was almost identical to<br />
an isolate obtained from leaf spots of Hosta plantaginea in Korea.<br />
<strong>The</strong>se isolates appeared to resemble species of Fusicladium, but<br />
phylogenetically they clustered in the Herpotrichiellaceae. <strong>The</strong>refore,<br />
“Fusicladium” scillae was placed in the <strong>genus</strong> Cladophialophora.<br />
As far as we are aware, this species <strong>and</strong> C. hostae are first reports<br />
of phytopathogenic species within the <strong>genus</strong> Cladophialophora.<br />
199
Crous et al.<br />
Fig. 13. Cladophialophora sylvestris (<strong>CBS</strong> 350.83). A–B. Conidiophores. C. Catenulate conidia. D. Conidial mass. Scale bar = 10 µm.<br />
Fig. 14. Cyphellophora hylomeconis (<strong>CBS</strong> 113311). A. Colony on PDA. B–C. Hyphae with truncate conidiogenous loci. D–F. Conidia. Scale bars = 10 µm.<br />
Cladophialophora sylvestris Crous & de Hoog, sp. nov.<br />
MycoBank MB504531. Fig. 13.<br />
Etymology: Refers to its host, Pinus sylvestris.<br />
Cladophialophorae humicolae similis, sed conidiis 0–3-septatis, (7–)10–16(–20) ×<br />
1.5–2 µm.<br />
Mycelium composed of branched, smooth, pale olivaceous to<br />
pale brown hyphae, frequently forming hyphal coils, not to slightly<br />
constricted at the septa, 1–2 µm wide. Conidiophores medium<br />
brown, subcylindrical, flexuous, mononematous, multiseptate, up<br />
to 50 µm long, <strong>and</strong> 2–3 µm wide. Conidiogenous cells apical,<br />
sympodial, pale brown, 5–12 × 2–3 µm; scars somewhat darkened<br />
<strong>and</strong> thickened, not refractive. Conidia occurring in branched chains;<br />
ramoconidia up to 2 µm wide, giving rise apically to disarticulating<br />
chains of conidia; smooth, 0–3-septate, pale olivaceous,<br />
subcylindrical, (7–)10–16(–20) × 1.5–2 µm, with truncate ends; hila<br />
somewhat darkened <strong>and</strong> thickened, not refractive.<br />
Cultural characteristics: Colonies erumpent on PDA, with smooth,<br />
catenulate margins; iron-grey (surface); reverse greenish black.<br />
Colonies reaching 15 mm diam after 1 mo at 25 °C in the dark;<br />
colonies fertile.<br />
Specimen examined: Netherl<strong>and</strong>s, Kootwijk, needle litter of Pinus sylvestris<br />
(Pinaceae), 8 Nov. 1982, G.S. de Hoog, holotype <strong>CBS</strong> H-19917, culture ex-type<br />
<strong>CBS</strong> 350.83.<br />
Notes: Morphologically <strong>CBS</strong> 350.83 was originally identified as<br />
Polyscytalum griseum Sacc., but the latter is reported to have<br />
conidia that are 5–5.5 × 1 µm (Saccardo 1877), which is much<br />
smaller than that observed for the present isolate. Furthermore,<br />
the type species of Polyscytalum, P. fecundissimum Riess (<strong>CBS</strong><br />
100506), does not cluster within the Herpotrichiellaceae, thus<br />
suggesting that <strong>CBS</strong> 350.83 is best treated as a new species of<br />
Cladophialophora.<br />
Cyphellophora hylomeconis Crous, de Hoog & H.D. Shin, sp.<br />
nov. MycoBank MB504532. Fig. 14.<br />
Etymology: Named after its host <strong>genus</strong>, Hylomecon.<br />
Cyphellophorae lacinatae similis, sed conidiis longioribus et leniter angustioribus,<br />
(15–)25–35(–55) × (2.5–)3(–4) µm.<br />
Mycelium consisting of branched, greenish brown, septate,<br />
branched, smooth, 3–5 µm wide hyphae, constricted at septa.<br />
Conidiogenous cells phialidic, intercalary, appearing denticulate,<br />
1 µm tall, 1.5–2 µm wide, with minute collarettes (at times<br />
200
Herpotrichiellaceae <strong>and</strong> Venturiaceae<br />
proliferating percurrently). Conidia sickle-shaped, smooth, medium<br />
brown, guttulate, (1–)3(–5)-septate, constricted at septa, widest in<br />
middle, or lower third of the conidium; apex subacutely rounded,<br />
base subtruncate, or having a slight constriction, giving rise to a<br />
foot cell, 1 µm long, 0.5–1 µm wide, subacutely rounded, (15–)25–<br />
35(–55) × (2.5–)3(–4) µm; a marginal frill is visible above the foot<br />
cell, suggesting this foot cell may be the onset of basal germination;<br />
conidia also anastomose <strong>and</strong> undergo microcyclic conidiation in<br />
culture.<br />
Cultural characteristics: Colonies slow-growing, slimy, aerial<br />
mycelium absent, margins smooth, catenate; surface crumpled,<br />
olivaceous-black to iron-grey. Colonies reaching 20 mm diam<br />
after 1 mo at 25 °C in the dark on PDA, 12 mm on SNA; colonies<br />
fertile.<br />
Specimen examined: Korea, Yangpyeong, on leaves of Hylomecon verlance<br />
(Papaveraceae), 4 Jun. 2003, H.D. Shin, holotype <strong>CBS</strong> H-19907, isotype SMK<br />
19550, culture ex-type <strong>CBS</strong> 113311.<br />
Notes: Cyphellophora hylomeconis is related to the type species<br />
of the <strong>genus</strong>, Cyphellophora laciniata G.A. de Vries, which also<br />
resides in the Herpotrichiellaceae. <strong>The</strong> <strong>genus</strong> Cyphellophora G.A.<br />
de Vries is phenetically distinguished from Pseudomicrodochium<br />
B.C. Sutton, typified by P. aciculare B.C. Sutton (1975) by melanized<br />
versus hyaline thalli. Phylogenetic confirmation is pending due to<br />
unavailability of sequence data. Decock et al. (2003) synonymised<br />
the hyaline <strong>genus</strong> Kumbhamaya M. Jacob & D.J. Bhat (Jacob &<br />
Bhat 2000) with Cyphellophora, but as no cultures of this fungus are<br />
available this decision seems premature. Nearly all Cyphellophora<br />
species accepted by Decock et al. (2003) have been found to be<br />
involved in cutaneous infections in humans. This also holds true<br />
for the species originally described as being environmental, C.<br />
vermispora Walz & de Hoog, which is closely related to C. suttonii<br />
(Ajello et al.) Decock <strong>and</strong> C. fusarioides (C.K. Campbell & B.C.<br />
Sutton) Decock known from proven human <strong>and</strong> animal infections.<br />
Decock et al. (2003) added the melanized species C. guyanensis<br />
Decock & Delgado, isolated as a saprobe from tropical leaf litter.<br />
Cyphellophora hylomeconis is the first species of the <strong>genus</strong><br />
infecting a living plant host. ITS sequences are remote from those of<br />
the remaining Cyphellophora species, the nearest neighbour being<br />
C. pluriseptata G.A. de Vries, Elders & Luykx at 19.1 % distance<br />
(data not shown). Cyphellophora hylomeconis can be distinguished<br />
based on its conidial dimensions <strong>and</strong> septation. Conidia are larger<br />
than those of C. fusarioides (11–20 × 2–2.5 µm, 1–2-septate), <strong>and</strong><br />
those of C. laciniata (11–25 × 2–5 µm, 1–3-septate) (for a key to<br />
the species see Decock et al. 2003).<br />
Exophiala sp. 1. Fig. 15.<br />
Mycelium consisting of smooth, branched, septate, medium brown,<br />
2–3 µm wide hyphae, regular in width, forming hyphal str<strong>and</strong>s<br />
<strong>and</strong> hyphal coils, with free yeast-like cells present in culture;<br />
chlamydospores terminal on hyphae, frequently forming clusters<br />
or chains, medium brown, ellipsoid, 0–1-septate, up to 10 µm<br />
long <strong>and</strong> 5 µm wide. Conidiophores reduced to conidiogenous<br />
cells, or consisting of one supporting cell, giving rise to a single<br />
conidiogenous cell, subcylindrical to ellipsoid, medium brown,<br />
smooth, 5–12 × 3.5–4 µm, with 1(–3) phialidic loci, somewhat<br />
protruding, appearing subdenticulate at first glance under the<br />
light microscope. Conidiogenous cells integrated as lateral loci on<br />
hyphal cells, inconspicuous, 1–1.5 µm wide, with a slightly flaring<br />
collarette, (1–)1.5(–2) µm long. Conidia ellipsoid, smooth, guttulate,<br />
becoming brown, swollen <strong>and</strong> elongated, <strong>and</strong> at times 1-septate,<br />
4–5(–7) × (2.5–)3(–4) µm (description based on <strong>CBS</strong> 115142).<br />
www.studiesinmycology.org<br />
Cultural characteristics: Colonies erumpent, spreading, with sparse<br />
to dense aerial mycelium on PDA, olivaceous-grey (surface), with a<br />
thin to wide, smooth, olivaceous-black margins; reverse olivaceousblack;<br />
on OA olivaceous-grey (surface) with wide, olivaceous-black<br />
margins. Colonies reaching 40–50 mm diam after 1 mo at 25 °C in<br />
the dark; colonies fertile, but sporulation sparse. Not able to grow<br />
at 37 °C.<br />
Specimen examined: Australia, from a fruit drink, May 2002, N.J. Charley, <strong>CBS</strong><br />
115142 = CPC 11044 = FRR 5582.<br />
Notes: Species of Exophiala are frequently observed as agents<br />
of human mycoses in immunocompromised patients (de Hoog<br />
et al. 2000). <strong>The</strong>y are found in the environment as slow-growing,<br />
oligotrophic colonisers of moist substrates. For example the<br />
thermotolerant species E. dermatitidis (Kano) de Hoog <strong>and</strong> E.<br />
phaeomuriformis (Matsumoto et al.) Matos et al. are common<br />
in public steam baths (Matos et al. 2003), while E. mesophila<br />
Listemann & Freiesleben can be found in showers <strong>and</strong> swimming<br />
pools (unpubl. data). Both species are able to cause infections<br />
in humans (Zeng et al. 2007). Several other species have been<br />
associated primarily with infections in fish <strong>and</strong> cold-blooded animals<br />
(Richards et al. 1978) <strong>and</strong> are occasionally found on humans<br />
(Madan et al. 2006). <strong>The</strong> occurrence of the present species in fruit<br />
drinks, therefore, is cause of concern, although it was unable to<br />
grow at 37 °C. This species forms part of a larger study, <strong>and</strong> will be<br />
treated elsewhere.<br />
Exophiala sp. 2. Fig. 16.<br />
Mycelium consisting of smooth, branched, septate, pale brown,<br />
1.5–3 µm wide hyphae, forming hyphal str<strong>and</strong>s <strong>and</strong> hyphal coils;<br />
hyphae at times terminating in chains of ellipsoid chlamydospores<br />
that are medium brown, smooth, up to 10 µm long <strong>and</strong> 5 µm wide.<br />
Conidiophores subcylindrical, medium brown, smooth, consisting<br />
of a supporting cell <strong>and</strong> a single conidiogenous cell, or reduced<br />
to a conidiogenous cell, straight to curved, up to 30 µm long<br />
<strong>and</strong> 2–3 µm wide. Conidiogenous cells pale to medium brown,<br />
subcylindrical to narrowly ellipsoid or subclavate, with 1–3 apical,<br />
phialidic loci, 1 µm wide, 1–2 µm tall, collarette somewhat flaring,<br />
but mostly cylindrical, 7–20 × 2–2.5 µm; at times proliferating<br />
percurrently. Conidia ellipsoid, smooth, guttulate, hyaline, becoming<br />
pale olivaceous, apex obtuse, base subtruncate, (4–)5–7(–10) ×<br />
2–2.5(–3) µm.<br />
Cultural characteristics: Colonies spreading with smooth,<br />
submerged margins, moderate aerial mycelium on PDA, sparse on<br />
OA, on PDA <strong>and</strong> OA olivaceous-grey (surface), with a wide, irongrey<br />
margin; reverse iron-grey. Colonies reaching 40–50 mm diam<br />
after 1 mo at 25 °C in the dark; colonies fertile. Not able to grow<br />
at 37 °C.<br />
Specimen examined: Australia, from bottled spring water, May 2003, N.J. Charley,<br />
<strong>CBS</strong> 115143 = CPC 11047 = FRR 5599.<br />
Notes: This strain represents another taxon occurring in bottled<br />
drinks destined for human consumption. As it is unable to grow at<br />
37 °C, it does not appear to pose any serious threat to human<br />
health. This species forms part of a larger study, <strong>and</strong> will be treated<br />
elsewhere.<br />
201
Crous et al.<br />
Fig. 15. Exophiala sp. 1 (<strong>CBS</strong> 115142). A. Colony on PDA. B. Hyphal coil. C. Hyphal str<strong>and</strong>. D–H. Conidiogenous cells <strong>and</strong> loci. I–O. Conidiogenous cells <strong>and</strong> conidia. Scale<br />
bars = 10 µm.<br />
Fig. 16. Exophiala sp. 2 (<strong>CBS</strong> 115143). A. Conidiogenous cells. B. Conidiophore with hyphal coil. C. Conidiogenous cell with hyphal str<strong>and</strong>. D. Conidia. Scale bar = 10 µm.<br />
202
Herpotrichiellaceae <strong>and</strong> Venturiaceae<br />
Fig. 17. Exophiala eucalyptorum (CPC 11261). A. Colony on PDA. B–H. Hyphae, conidiogenous cells <strong>and</strong> conidia. Scale bars = 10 µm.<br />
Exophiala eucalyptorum Crous, sp. nov. MycoBank MB504533.<br />
Fig. 17.<br />
Etymology: Named after its occurrence on Eucalyptus leaves.<br />
Exophialae spiniferae similis, sed conidiis fusoidibus-ellipsoideis, (5–)6–8(–10)<br />
× (3–)4–5(–7) µm, et cellulis conidiogenis saepe catenatis, in catenis brevibus,<br />
dividentibus.<br />
Mycelium consisting of smooth to finely verruculose, branched,<br />
septate, 2–4 µm wide hyphae, at times giving rise to chains of<br />
dark brown, fusoid-ellipsoid chlamydospores, which can still have<br />
phialides, suggesting they were conidiogenous cells; hyphae<br />
becoming constricted at septa when fertile. Conidiophores reduced<br />
to conidiogenous cells. Conidiogenous cells numerous, terminal<br />
<strong>and</strong> lateral, mono- to polyphialidic, 5–15 × 3–5 µm; loci 1–1.5 µm<br />
wide <strong>and</strong> tall, with inconspicuous collarettes, at time proliferating<br />
percurrently; conidiogenous cells fusoid-ellipsoid, <strong>and</strong> frequently<br />
breaking off, appearing as short chains of conidia, but distinct in<br />
having conidiogenous loci. Conidia fusoid-ellipsoid, apex acutely<br />
rounded, base subtruncate, (5–)6–8(–10) × (3–)4–5(–7) µm;<br />
frequently becoming fertile, septate <strong>and</strong> brown with age.<br />
Cultural characteristics: Colonies erumpent, convex, smooth, slimy,<br />
margins feathery to crenate <strong>and</strong> smooth; aerial mycelium absent,<br />
growth yeast-like. Colonies on PDA, OA <strong>and</strong> SNA chestnut on<br />
surface <strong>and</strong> reverse. Colonies reaching 4 mm diam after 2 wk on<br />
PDA at 25 °C in the dark.<br />
Specimen examined: New Zeal<strong>and</strong>, Wellington Botanical Garden, on leaf litter of<br />
Eucalyptus sp. (Myrtaceae), Mar. 2004, J.A. Stalpers, holotype <strong>CBS</strong> H-19905,<br />
culture ex-type <strong>CBS</strong> 121638 = CPC 11261.<br />
www.studiesinmycology.org<br />
Notes: Exophiala eucalyptorum is rather characteristic in that,<br />
in culture, chains of conidiogenous cells frequently detach from<br />
hyphae, appearing as short, intact chains of fertile conidia.<br />
Its phylogenetic position is somewhat outside the core of the<br />
Herpotrichiellaceae containing most Capronia teleomorphs <strong>and</strong><br />
the remaining opportunistic Exophiala species, but still within the<br />
Chaetothyriales (Figs 1–2).<br />
Members of Venturiaceae<br />
Anungitea B. Sutton <strong>and</strong> Anungitopsis R.F. Castañeda &<br />
W.B. Kendr.<br />
Sutton (1973) erected the <strong>genus</strong> Anungitea to accommodate<br />
species with brown, mononematous conidiophores bearing<br />
apically aggregated, flat-tipped, subdenticulate conidiogenous loci<br />
that give rise to chains of pale brown subcylindrical conidia with<br />
thickened, darkened hila. He compared the type species, A. fragilis<br />
B. Sutton with anamorph genera of the Mycosphaerellaceae,<br />
but did not compare it to Fusicladium, to which it is remarkably<br />
<strong>similar</strong>. Castañeda & Kendrick (1990b) introduced the <strong>genus</strong><br />
Anungitopsis based on A. speciosa R.F. Castañeda & W.B. Kendr.<br />
This <strong>genus</strong> was distinguished from Anungitea by its formation of<br />
subdenticulate conidiogenous loci distributed along the apical<br />
region of the conidiophore, <strong>and</strong> by the relatively poorly defined<br />
appearance of these loci. No cultures are available of the extype<br />
species of Anungitea, but we studied strains of Anungitopsis<br />
203
Crous et al.<br />
Fig. 18. Cylindrosympodium lauri (<strong>CBS</strong> 240.95). A–C. Conidiophores with conidiogenous loci. D. Conidia. Scale bar = 10 µm.<br />
amoena R.F. Castañeda & Dugan (<strong>CBS</strong> 254.95, ex-type), <strong>and</strong><br />
Anungitopsis intermedia Crous & W.B. Kendr. (<strong>CBS</strong> 110746,<br />
ex-epitype), <strong>and</strong> found them to cluster adjacent to Fusicladium<br />
(Venturiaceae). However, the ex-type strain of Anungitopsis<br />
speciosa (<strong>CBS</strong> 181.95), type species of Anungitopsis, clustered<br />
distantly from all other species, confirming that the <strong>genus</strong> name<br />
Anungitopsis is not available for any of the taxa treated here. In<br />
any case, A. speciosa has unusual subdenticulate conidiogenous<br />
loci with indistinct marginal frills, <strong>and</strong> these are obviously different<br />
from those of anungitea- <strong>and</strong> fusicladium-like anamorphs, including<br />
A. amoena <strong>and</strong> A. intermedia. <strong>The</strong> latter two species previously<br />
referred to as Anungitopsis belong to a sister clade of the Venturia<br />
(Fusicladium, incl. Pseudocladosporium) clade. Sympoventuria<br />
(Crous et al. 2007b), which produces a sympodiella-like anamorph<br />
in culture, is the only teleomorph of this clade hitherto known. <strong>The</strong><br />
venturia-like habit of Sympoventuria, connected with fusicladium-<br />
/ pseudocladosporium-like anamorphs distributed in both clades,<br />
indicates a close relation between these clades, suggesting a<br />
placement in the Venturiaceae. Schubert et al. (2003) referred to<br />
the difficulty to distinguish between Anungitea <strong>and</strong> Fusicladium.<br />
Anungitea is undoubtedly heterogeneous. Anungitea rhabdospora<br />
P.M. Kirk (Kirk 1983) is, for instance, intermediate between<br />
Anungitea (conidiophores with a terminal denticulate conidiogenous<br />
cell, but conidia disarticulating in an arthroconidium-like manner)<br />
<strong>and</strong> Sympodiella B. Kendr. (conidiophores distinctly sympodial,<br />
forming arthroconidia). Other species assigned to Anungitea<br />
possess a distinctly swollen, lobed conidiophore base, e.g. A.<br />
heterospora P.M. Kirk (Kirk 1983), which is comparable with other<br />
morphologically <strong>similar</strong> genera, e.g., Parapleurotheciopsis P.M.<br />
Kirk (Kirk 1982), Rhizocladosporium Crous & U. Braun (see Crous<br />
et al. 2007a – this volume), <strong>and</strong> Subramaniomyces Varghese &<br />
V.G. Rao (Varghese & Rao 1979, Kirk 1982). <strong>The</strong> application of<br />
Anungitea depends, however, on the affinity of A. fragilis, the type<br />
species, of which sequence data are not yet available. <strong>The</strong> best<br />
solution for this problem is the widened application of Fusicladium<br />
(incl. Pseudocladosporium) to both sister clades, i.e., to the whole<br />
Venturiaceae. Morphologically a distinction between fusicladioid<br />
anamorphs of both clades is impossible. <strong>The</strong> more “fusicladium-like”<br />
growth is mainly characteristic for the fruiting in vivo, above all in<br />
biotrophic taxa, whereas the more “pseudocladosporium-like” habit<br />
is typical for the growth in vitro <strong>and</strong> in saprobic taxa, a phenomenon<br />
which is also evident in species of the morphologically <strong>similar</strong><br />
<strong>genus</strong> Cladophialophora (see C. hostae <strong>and</strong> C. scillae). A potential<br />
placement of Anungitea fragilis within the Venturiaceae, which has<br />
still to be proven, would render the <strong>genus</strong> Anungitea a synonym of<br />
Fusicladium, but in the case of a quite distinct phylogenetic position<br />
a new circumscription of this <strong>genus</strong>, excluding the Venturiaceae<br />
anamorphs, would be necessary. Thus, a final conclusion about<br />
Anungitea has to be postponed, awaiting cultures <strong>and</strong> sequence<br />
analyses of its type species.<br />
<strong>The</strong> taxonomic placement of a fungus from the Canary<br />
Isl<strong>and</strong>s, isolated from leaf litter of Laurus sp. (<strong>CBS</strong> 240.95), is<br />
somewhat problematic. It clusters within the Venturiaceae, but<br />
not within Venturia s. str. itself, <strong>and</strong> it does not fit into the current<br />
morphological concept of Fusicladium (incl. Pseudocladosporium).<br />
Based on its solitary, cylindrical, hyaline conidia <strong>and</strong> pale brown<br />
conidiogenous structures, it resembles species accommodated in<br />
Cylindrosympodium W.B. Kendr. & R.F. Castañeda (Castañeda &<br />
Kendrick 1990a, Marvanová & Laichmanová 2007).<br />
Cylindrosympodium lauri Crous & R.F. Castañeda, sp. nov.<br />
MycoBank MB504534. Fig. 18.<br />
Etymology: Named after the host <strong>genus</strong> it was collected from,<br />
Laurus.<br />
Cylindrosympodii variabilis similis, sed conidiophoris longioribus, ad 70 µm, conidiis<br />
subhyalinis vel dilute olivaceis.<br />
Mycelium consisting of brown, smooth, septate, branched hyphae,<br />
1.5–2.5 µm wide. Conidiophores macronematous, mononematous,<br />
solitary, erect, subcylindrical, straight to geniculate-sinuous,<br />
medium brown, smooth, 35–70 × 2.5–4 µm, 1–5-septate.<br />
Conidiogenous cells terminal, integrated, pale to medium brown,<br />
smooth, 10–35 × 2–3 µm, proliferating sympodially, with one to<br />
several flat-tipped loci, 1.5–2 µm wide; scars somewhat darkened,<br />
minutely thickened, but not refractive. Conidia solitary, subacicular<br />
to narrowly subcylindrical, apex subobtuse, base truncate, or<br />
somewhat swollen, straight or curved, smooth, subhyaline to very<br />
pale olivaceous, guttulate, (45–)60–70(–80) × 2.5–3(–3.5) µm,<br />
(4–)6–8-septate; scars are somewhat darkened, minutely thickened,<br />
but not refractive, 2.5–3 µm wide.<br />
Cultural characteristics: Colonies erumpent, convex, with smooth,<br />
lobed margins, <strong>and</strong> moderate, dense aerial mycelium on PDA;<br />
mouse-grey in the central part, <strong>and</strong> dark mouse-grey in the outer<br />
zone (surface); reverse dark mouse-grey. Colonies reaching 5 mm<br />
diam after 2 wk at 25 °C in the dark; colonies fertile.<br />
204
Herpotrichiellaceae <strong>and</strong> Venturiaceae<br />
Specimen examined: Spain, Canary Isl<strong>and</strong>s, leaf litter of Laurus sp. (Lauraceae), 4<br />
Jan. 1995, R.F. Castañeda, holotype <strong>CBS</strong> H-19909, culture ex-type <strong>CBS</strong> 240.95.<br />
Note: <strong>The</strong> present fungus differs from Cylindrosympodium variabile<br />
(de Hoog) W.B. Kendr. & R.F. Castañeda (de Hoog 1985) in that the<br />
conidiophores are much longer, the conidia are subhyaline to very<br />
pale olivaceous, <strong>and</strong> the scars <strong>and</strong> hila are thin, slightly darkened,<br />
but not refractive.<br />
Venturia Sacc. <strong>and</strong> its anamorph Fusicladium<br />
Venturia Sacc., Syll. fung. (Abellini) 1: 586. 1882.<br />
= Apiosporina Höhn., Sitzungsber. Kaiserl. Akad. Wiss., Math.-Naturw. Cl., Abt.<br />
1, 119: 439. 1910, syn. nov.<br />
= Metacoleroa Petr., Ann. Mycol. 25: 332. 1927, syn. nov.<br />
= Caproventuria U. Braun, A Monograph of Cercosporella, Ramularia <strong>and</strong> Allied<br />
Genera (Phytopathogenic Hyphomycetes) 2: 396. 1998, syn. nov.<br />
For additional synonyms see Sivanesan, <strong>The</strong> bitunicate<br />
Ascomycetes <strong>and</strong> their anamorphs: 604. 1984.<br />
Anamorph: Fusicladium Bonord., H<strong>and</strong>b. Mykol.: 80. 1851.<br />
= Pseudocladosporium U. Braun, A Monograph of Cercosporella, Ramularia<br />
<strong>and</strong> Allied Genera (Phytopathogenic Hyphomycetes) 2: 392. 1998, syn. nov.<br />
For additional synonyms, see Schubert et al. (2003).<br />
Notes: <strong>The</strong> <strong>genus</strong> Caproventuria, based on C. hanliniana (U. Braun<br />
& Feiler) U. Braun, was erected to accommodate saprobic, soil-borne<br />
venturia-like ascomycetes with numerous ascomatal setae, <strong>and</strong> an<br />
anamorph quite distinct from Fusicladium (Braun 1998). <strong>The</strong> <strong>genus</strong><br />
Metacoleroa is based on M. dickiei (Berk. & Broome) Petr., which<br />
clusters in the Venturiaceae, adjacent to Caproventuria, which has<br />
Pseudocladosporium anamorphs. Metacoleroa was retained by Barr<br />
(1987) as separate from Venturia based on its superficial ascomata<br />
with a thin, stromatic layer beneath the ascomata. Whether these<br />
criteria still justify the separation of Caproventuria <strong>and</strong> Metacoleroa<br />
from Venturia is debatable, <strong>and</strong> the names Venturia dickiei (Berk. &<br />
Broome) Ces. & de Not. <strong>and</strong> Venturia hanliniana (U. Braun & Feiler)<br />
Unter. are available for these organisms. <strong>The</strong> <strong>genus</strong> Apiosporina,<br />
which is based on Apiosporina collinsii (Schwein.) Höhn., clusters in<br />
the Venturiaceae, as was to be expected based on its Fusicladium<br />
anamorph (Schubert et al. 2003). It was distinguished from Venturia<br />
species by having ascospores strictly septate near the lower end<br />
(Sivanesan 1984).<br />
<strong>The</strong> anamorph <strong>genus</strong> Fusicladium has been monographed by<br />
Schubert et al. (2003). Morphological as well as molecular studies<br />
(Beck et al. 2005) demonstrated that the <strong>genus</strong> Venturia with its<br />
Fusicladium anamorphs is monophyletic. A separation of Venturia<br />
into various uniform subclades based on the previous anamorph<br />
genera Fusicladium, Pollaccia <strong>and</strong> Spilocaea was not evident <strong>and</strong><br />
could be rejected. As in cercosporoid anamorphs of Mycosphaerella,<br />
features such as the arrangement of the conidiophores (solitary,<br />
fasciculate, sporodochial), the proliferation of conidiogenous cells<br />
(sympodial, percurrent) <strong>and</strong> shape, size as well as formation of<br />
conidia (solitary, catenate) proved to be of little taxonomic value at<br />
generic level. Hence, Schubert et al. (2003) proposed to maintain<br />
Fusicladium emend. as sole anamorph <strong>genus</strong> for Venturia.<br />
<strong>The</strong> <strong>genus</strong> Fusicladosporium Partridge & Morgan-Jones (type<br />
species: <strong>Cladosporium</strong> carpophilum Thüm.) (Partridge & Morgan-<br />
Jones 2003), recently erected to accommodate fusicladium-like<br />
species with catenate conidia, represents a further synonym of<br />
Fusicladium.<br />
Similar to their occurrence in vivo the conidiophores in<br />
vitro of species previously referred to the genera Spilocaea<br />
<strong>and</strong> Pollaccia are usually micronematous, conidia often appear<br />
to be directly formed on the mycelium, unilocal, determinate,<br />
mostly reduced to conidiogenous cells, sometimes forming a few<br />
percurrent proliferations, whereas the conidiophores of species<br />
of Fusicladium s. str. are mostly macronematous, but sometimes<br />
also micronematous. <strong>The</strong>y are often initiated as short lateral,<br />
peg-like outgrowths of hyphae which proliferate sympodially,<br />
becoming slightly geniculate, forming a single, several or numerous<br />
subdenticulate to denticulate, truncate, unthickened or only slightly<br />
thickened, somewhat darkened-refractive conidiogenous loci.<br />
<strong>The</strong> <strong>genus</strong> Pseudocladosporium was described to be quite<br />
distinct from Fusicladium by being saprobic <strong>and</strong> connected with a<br />
different teleomorph, viz. Caproventuria (Braun 1998). However,<br />
since the type species of Caproventuria, C. hanliniana, with its<br />
anamorph Pseudocladosporium brevicatenatum (U. Braun &<br />
Feiler) U. Braun clusters together with numerous Venturia species,<br />
the <strong>genus</strong> Pseudocladosporium should be reduced to synonymy<br />
with Fusicladium. Morphologically there is no clear delimitation<br />
between Fusicladium <strong>and</strong> Pseudocladosporium. <strong>The</strong> typically<br />
pseudocladosporium-like habit, characterised by forming solitary<br />
conidiophores, often reduced to conidiogenous cells or even<br />
micronematous, <strong>and</strong> conidia formed in long chains, is mainly found<br />
in culture, above all in saprobic taxa. <strong>The</strong> fusicladium-like growth<br />
with well-developed macronematous conidiophores is usually more<br />
evident in vivo, above all in biotrophic taxa. <strong>The</strong>re are, however, all<br />
kinds of transitions between these two genera.<br />
Fusicladium africanum Crous, sp. nov. MycoBank MB504535.<br />
Fig. 19.<br />
Etymology: Named after the continent from which it was collected,<br />
Africa.<br />
Fusicladio brevicatenato similis, sed conidiophoris brevioribus, 5–10 µm longis,<br />
conidiis minoribus, ad 20 × 3.5 µm, 0(–1)-septatis, locis conidiogenis et hilis<br />
angustioribus, 1–1.5 µm latis.<br />
Mycelium composed of smooth, medium brown, branched,<br />
septate, 1.5–2 µm wide hyphae, frequently forming hyphal coils.<br />
Conidiophores reduced to conidiogenous cells, solitary, pale to<br />
medium brown, smooth, inconspicuous, integrated in hyphae,<br />
varying from small, truncate lateral loci on hyphal cells, 1–1.5 µm<br />
wide, to micronematous conidiogenous cells, 5–10 × 2–3 µm; monoto<br />
polyblastic, sympodial, scars inconspicuous, 1 µm wide. Conidia<br />
in long, branched chains of up to 40, subcylindrical, 0(–1)-septate,<br />
pale brown, smooth; hila truncate, 1 µm wide, unthickened, neither<br />
darkened nor refractive; ramoconidia (11–)15–17(–20) × 2–3(–3.5)<br />
µm; conidia (8–)11–17 × 2–2.5 µm.<br />
Cultural characteristics: Colonies somewhat erumpent, with<br />
moderate aerial mycelium <strong>and</strong> smooth, lobate margins on PDA,<br />
ochreous to umber (surface); reverse dark umber; on OA umber;<br />
on SNA ochreous. Colonies reaching 9 mm diam on PDA after 2 wk<br />
at 25 °C in the dark; colonies fertile.<br />
Specimen examined: South Africa, Western Cape Province, Malmesbury,<br />
Eucalyptus leaf litter, Jan. 2006, P.W. Crous, holotype <strong>CBS</strong> H-19904, cultures extype<br />
CPC 12828 = <strong>CBS</strong> 121639, CPC 12829 = <strong>CBS</strong> 121640.<br />
Notes: Fusicladium africanum is a somewhat atypical member of the<br />
<strong>genus</strong>, as its conidial hila are quite unthickened <strong>and</strong> inconspicuous.<br />
Among biotrophic, leaf-spotting Fusicladium species a wider<br />
morphological variation was found pertaining to the structure of the<br />
conidiogenous loci <strong>and</strong> conidial hila, ranging from being indistinct,<br />
unthickened <strong>and</strong> not darkened-refractive to unthickened or almost<br />
so, but somewhat darkened-refractive (Schubert et al. 2003).<br />
Fusicladium africanum was found occurring with Sympoventuria<br />
capensis Crous & Seifert on Eucalyptus leaf litter in South Africa<br />
(Crous et al. 2007b).<br />
www.studiesinmycology.org<br />
205
Crous et al.<br />
Fig. 19. Fusicladium africanum (CPC 12828). A. Colony on MEA. B. Hyphal coil. C. Branched conidial chain. D–F. Conidiophores with catenulate conidia. Scale bar = 10 µm.<br />
Fig. 20. Fusicladium amoenum (<strong>CBS</strong> 254.95). A–E. Conidiophores with conidiogenous loci. F. Conidia. Scale bar = 10 µm.<br />
206
Herpotrichiellaceae <strong>and</strong> Venturiaceae<br />
Fig. 21. Fusicladium convolvularum (<strong>CBS</strong> 112706). A–B, D–I. Conidiophores with conidiogenous loci. C, J–K. Ramoconidia <strong>and</strong> conidia. Scale bars = 10 µm.<br />
Fusicladium amoenum (R.F. Castañeda & Dugan) Crous, K.<br />
Schub. & U. Braun, comb. nov. MycoBank MB504536. Fig. 20.<br />
Basionym: Anungitopsis amoena R.F. Castañeda & Dugan,<br />
Mycotaxon 72: 118. 1999.<br />
≡ <strong>Cladosporium</strong> amoenum R.F. Castañeda, in Untereiner et al., 1998,<br />
nom. nud.<br />
Specimen examined: Cuba, Santiago de Cuba, La Gran Piedra, fallen leaves of<br />
Eucalyptus sp. (Myrtaceae), 2 Nov. 1994, R.F. Castañeda, (Ho et al. 1999: 117,<br />
figs 2–3) iconotype, culture ex-type <strong>CBS</strong> 254.95 = ATCC 200947 = IMI 367525 =<br />
INIFAT C94/155 = MUCL 39143.<br />
Note: In culture F. amoenum has a typical pseudocladosporium-like<br />
morphology, though the scars are neither prominently thickened,<br />
nor refractive.<br />
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207
Crous et al.<br />
Fig. 22. Fusicladium fagi (<strong>CBS</strong> 621.84). A. Conidiophore with truncate conidiogenous loci. B. Hypha with conidiogenous loci. C–G. Conidial chains. Scale bars = 10 µm.<br />
Fusicladium caruanianum Sacc., Ann. Mycol. 11: 20. 1913.<br />
≡ Pseudocladosporium caruanianum (Sacc.) U. Braun, Schlechtendalia<br />
9: 114. 2003.<br />
Fusicladium convolvularum Ondřej, Česká Mycol. 25: 171. 1971.<br />
Fig. 21.<br />
In vivo: Schubert et al. (2003: 37).<br />
In vitro on SNA: Mycelium unbranched or only sparingly branched,<br />
2–3 µm wide, septate, not constricted at septa, subhyaline to pale<br />
brown, smooth, walls unthickened or almost so. Conidiophores<br />
laterally arising from hyphae, erect, straight to somewhat flexuous,<br />
sometimes geniculate, unbranched, (6–)12–75 × (2.5–)3–4.5<br />
µm, aseptate or septate, pale brown or pale medium brown,<br />
smooth, walls somewhat thickened, sometimes only as short<br />
lateral conical prolongations of hyphae, occasionally irregular in<br />
shape. Conidiogenous cells integrated, terminal or conidiophores<br />
reduced to conidiogenous cells, sometimes geniculate, 6–29 µm<br />
long, proliferation sympodial, with several denticle-like loci, broadly<br />
truncate, 1.5–2(–2.5) µm wide, unthickened, somewhat refractive<br />
or darkened. Ramoconidia occurring, 20–28 × 5 µm, 0–1-septate,<br />
somewhat darker, pale medium brown, with a broadly truncate<br />
base, 3–4 µm wide, usually with several denticle-like apical loci.<br />
Conidia catenate, formed in unbranched or loosely branched<br />
chains, straight to sometimes curved, cells sometimes irregularly<br />
swollen, fusiform, subcylindrical, sometimes obpyriform, 13–35 ×<br />
3.5–5.5(–6) µm, 0–3-septate, occasionally slightly constricted at<br />
the median septum, few very large conidia with up to five septa,<br />
up to 75 µm long, 4.5–6 µm wide, subhyaline to pale brown,<br />
smooth, walls slightly thickened, slightly attenuated towards apex<br />
<strong>and</strong> base, hila broadly truncate, 1–2 µm wide, unthickened or<br />
only slightly thickened, somewhat darkened-refractive; microcyclic<br />
conidiogenesis occurring, conidia often germinating.<br />
Cultural characteristics: Colonies on PDA spreading, somewhat<br />
erumpent, with moderate aerial mycelium <strong>and</strong> regular, but feathery<br />
margins; surface fuscous black, <strong>and</strong> reverse dark fuscous black.<br />
Colonies reaching 15 mm diam after 1 mo on PDA at 25 °C in the<br />
dark.<br />
Specimens examined: Czech Republic, Libina, okraj pole pod nadrazim (okr.<br />
Sumperk), on Convolvulus arvensis (Convolvulaceae), 7 Sep. 1970, Ondřej,<br />
holotype BRA. New Zeal<strong>and</strong>, on leaves of Convolvulus arvensis, 7 Nov. 2000,<br />
C.F. Hill, epitype designated here <strong>CBS</strong> H-19911, culture ex-epitype <strong>CBS</strong> 112706<br />
= CPC 3884 = IMI 383037.<br />
Note: Conidiophores are somewhat longer <strong>and</strong> narrower in vitro<br />
than in vivo, <strong>and</strong> ramoconidia occur (Schubert & Braun 2002b,<br />
Schubert et al. 2003).<br />
208
Herpotrichiellaceae <strong>and</strong> Venturiaceae<br />
Fig. 23. Fusicladium intermedium (<strong>CBS</strong> 110746). A–G. Conidiophores with sympodial conidiogenous loci. H. Conidia. Scale bar = 10 µm.<br />
Fusicladium fagi Crous & de Hoog, sp. nov. MycoBank MB504537.<br />
Fig. 22.<br />
Etymology: Named after its host, Fagus sylvatica.<br />
Fusicladio brevicatenato similis, sed conidiis secundis minoribus, (8–)11–17(–20) ×<br />
3–3.5 µm, locis conidiogenis et hilis angustioribus, 1–1.5 µm latis.<br />
Mycelium consisting of pale to medium brown, smooth to finely<br />
verruculose, branched, 2–3 µm wide hyphae. Conidiophores<br />
integrated, terminal on hyphae, 0–1-septate, mostly reduced to<br />
conidiogenous cells, also lateral, visible as small, protruding,<br />
denticle-like loci, 10–15 × 2–3.5 µm. Conidiogenous cells<br />
subcylindrical, 5–15 × 2–3.5 µm, pale to medium brown, smooth<br />
to finely verruculose, tapering to 1–3 apical loci, 1–1.5 µm wide;<br />
scars inconspicuous. Conidia pale brown, smooth, guttulate,<br />
subcylindrical to narrowly ellipsoid, occurring in simple or branched<br />
chains, 0–1(–2)-septate, tapering towards subtruncate ends, 1.5–<br />
2.5 µm wide, aseptate conidia (8–)11–17(–20) × 3–3.5 µm, septate<br />
conidia up to 40 µm long <strong>and</strong> 4 µm wide; hila inconspicuous, i.e.<br />
neither thickened nor darkened-refractive; microcyclic conidiation<br />
common in older cultures.<br />
Cultural characteristics: Colonies erumpent, spreading, with<br />
abundant aerial mycelium on PDA, <strong>and</strong> feathery to smooth margins;<br />
isabelline to patches of fuscous-black due to the absence of aerial<br />
mycelium, which collapses with age (surface); reverse fuscousblack.<br />
Colonies reaching 50 mm diam after 1 mo at 25 °C in the<br />
dark; colonies fertile.<br />
Specimen examined: Netherl<strong>and</strong>s, Baarn, Maarschalksbosch, decaying leaves of<br />
Fagus sylvatica (Fagaceae), 1 Oct. 1984, G.S. de Hoog, holotype <strong>CBS</strong> H-10366,<br />
culture ex-type <strong>CBS</strong> 621.84 = ATCC 200937.<br />
Notes: Isolate <strong>CBS</strong> 621.84 was until recently preserved at the<br />
<strong>CBS</strong> as representative of <strong>Cladosporium</strong> nigrellum Ellis & Everh., a<br />
species known from bark of Robinia sp. in the U.S.A. Morphologically<br />
it is, however, quite distinct in having somewhat larger, <strong>and</strong> more<br />
subcylindrical to ellipsoid conidia. Conidia of C. nigrellum are<br />
fusiform to limoniform, 0–3-septate, 5–15 × 4–7 µm (Ellis 1976),<br />
possessing the typical cladosporioid scars with a central convex<br />
dome <strong>and</strong> a periclinal rim which characterise it as a true member<br />
of the <strong>genus</strong> <strong>Cladosporium</strong> Link, which has been confirmed by a<br />
re-examination of type material of C. nigrellum (on inner bark of<br />
railroad ties, U.S.A., West Virginia, Fayette Co., Nuttallburg, 20<br />
Oct. 1893, L.A. Nuttall, Flora of Fayette County No. 172, NY; also<br />
Ellis & Everh., N. Amer. Fungi 3086 <strong>and</strong> Fungi Columb. 382, BPI,<br />
NY, PH).<br />
Fusicladium intermedium (Crous & W.B. Kendr.) Crous, comb.<br />
nov. Mycobank MB504538. Fig. 23.<br />
Basionym: Anungitopsis intermedia Crous & W.B. Kendr. S. Afr. J.<br />
Bot. 63: 286. 1997.<br />
Specimens examined: South Africa, Mpumalanga, from leaf litter of Eucalyptus<br />
sp. (Myrtaceae), Oct. 1992, M.J. Wingfield, PREM 51438 holotype. Madagascar,<br />
Tamatave, Eucalyptus leaf litter, Apr. 1994, P.W. Crous, <strong>CBS</strong> H-19918, epitype<br />
designated here, culture ex-epitype CPC 778 = IMI 362702 = <strong>CBS</strong> 110746.<br />
Note: Conidiophores are dimorphic in culture, being macronematous,<br />
anungitopsis-like, <strong>and</strong> micronematous, more pseudocladosporiumlike.<br />
Fusicladium matsushimae (U. Braun & C.F. Hill) Crous, U. Braun<br />
& K. Schub., comb. nov. Mycobank MB504539.<br />
Basionym: Pseudocladosporium matsushimae U. Braun & C.F. Hill,<br />
Australas. Pl. Pathol. 33: 492. 2004.<br />
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209
Crous et al.<br />
Fig. 24. Fusicladium pini (<strong>CBS</strong> 463.82). A–F. Conidiogenous cells with conidiogenous loci. G. Conidia. Scale bars = 10 µm.<br />
Fusicladium m<strong>and</strong>shuricum (M. Morelet) Ritschel & U. Braun,<br />
Schlechtendalia 9: 62. 2003.<br />
Basionym: Pollaccia m<strong>and</strong>shurica M. Morelet, Ann. Soc. Sci. Nat.<br />
Archéol. Toulon Var 45(3): 218. 1993.<br />
= Pollaccia sinensis W.P. Wu & B. Sutton, in herb. (IMI).<br />
Teleomorph: Venturia m<strong>and</strong>shurica M. Morelet, Ann. Soc. Sci.<br />
Nat. Archéol. Toulon Var 45(3): 219. 1993.<br />
In vivo: Schubert et al. (2003: 62).<br />
In vitro on OA: Mycelium loosely branched, filiform to narrowly<br />
cylindrical-oblong, 1–4 µm wide, later somewhat wider, up to<br />
7 µm, septate, sometimes slightly constricted at the septa,<br />
sometimes irregular in outline due to small swellings, subhyaline<br />
to pale brown, smooth, walls unthickened, sometimes aggregating,<br />
forming compact conglomerations of slightly swollen hyphal cells.<br />
Conidiophores usually reduced to conidiogenous cells, arising<br />
terminally or laterally from hyphae, subcylindrical to cylindrical,<br />
unbranched, 9–20 × (2.5–)4–5(–6) µm, aseptate, very rarely 1-<br />
septate, very pale brown, smooth, walls unthickened, monoblastic,<br />
unilocal, determinate, later occasionally becoming percurrent,<br />
enteroblastically proliferating, forming a few (up to five) annellations,<br />
loci broadly truncate, (2–)3–5 µm wide, unthickened, not darkened.<br />
Conidia solitary, straight to curved, fusiform to obclavate, distinctly<br />
apiculate, 24–45(–57) × (6–)7–9(–10.5) µm, (1–)2–4(–5)-septate,<br />
more or less constricted at septa, sometimes up to 85 µm long<br />
with up to 7 septa, septa often somewhat darkened, second cell<br />
often bulging, pale medium to medium olivaceous-brown or brown,<br />
smooth, walls somewhat thickened, somewhat attenuated towards<br />
the base, hilum broadly truncate, (2–)3–5 µm wide, unthickened,<br />
not darkened; microcyclic conidiogenesis not observed.<br />
Cultural characteristics: Colonies on OA iron-grey to olivaceousgrey<br />
due to aerial mycelium <strong>and</strong> sporulation (surface); reverse<br />
iron-grey to black, somewhat velvety; margin glabrous, olivaceous;<br />
aerial mycelium sparsely formed, loose, diffuse; sporulating.<br />
Specimens examined: China, Liaoning, on Populus simonii × P. nigra, 17 Jun. 1992,<br />
M. Morelet, holotype PC (PFN 1466); P. simonii, 20 Apr. 1993, epitype designated<br />
here <strong>CBS</strong> H-19912, culture ex-epitype <strong>CBS</strong> 112235 = CPC 3639 = MPFN 307.<br />
Note: Conidiophores are densely fasciculate in vivo, forming<br />
sporodochial conidiomata, cylindrical to ampulliform, 5–7 × 6–7.5<br />
µm (Schubert et al. 2003).<br />
Fusicladium pini Crous & de Hoog, sp. nov. MycoBank MB504540.<br />
Fig. 24.<br />
Etymology: Named after its host, Pinus.<br />
Fusicladio africano similis, sed conidiis minoribus, (6–)10–12(–17) × 1.5–2(–2.5)<br />
µm, locis conidiogenis et hilis angustioribus, 0.5–1 µm latis.<br />
Mycelium consisting of smooth, medium brown, branched,<br />
1.5–2 µm wide hyphae, giving rise to solitary, micronematous<br />
conidiophores. Conidiophores reduced to conidiogenous cells,<br />
medium to dark brown, erect, thick-walled, smooth, subcylindrical,<br />
widest at the base, tapering to a subtruncate apex, 5–15 × 2–3<br />
µm; scars flat-tipped, somewhat darkened <strong>and</strong> thickened, one to<br />
several in the apical region, somewhat protruding, 0.5–1 µm wide.<br />
Conidia in branched or unbranched chains of up to 15, medium<br />
brown, smooth, subcylindrical, 0–1-septate, widest in the middle,<br />
tapering to subtruncate ends, straight to slightly curved, (6–)10–<br />
12(–17) × 1.5–2(–2.5) µm; hila somewhat darkened <strong>and</strong> thickened,<br />
not refractive, 0.5–1 µm wide.<br />
Cultural characteristics: Colonies erumpent, with sparse aerial<br />
mycelium <strong>and</strong> smooth margins on PDA, greyish sepia (surface);<br />
reverse fuscous-black; on OA patches of greyish sepia <strong>and</strong> fuscousblack<br />
(surface); on SNA umber (surface). Colonies reaching 15 mm<br />
diam on PDA after 1 mo at 25 °C in the dark; colonies fertile.<br />
210
Herpotrichiellaceae <strong>and</strong> Venturiaceae<br />
Fig. 25. Fusicladium ramoconidii (<strong>CBS</strong> 462.82). A. Colony on OA. B. Hyphal coil. C–F. Conidiophores reduced to conidiogenous cells. G–H. Conidiophores. I. Conidia. Scale<br />
bars = 10 µm.<br />
Specimen examined: Netherl<strong>and</strong>s, Baarn, De Vuursche, needle of Pinus sylvestris<br />
(Pinaceae), 12 Apr. 1982, G.S. de Hoog, holotype <strong>CBS</strong> H-1610, culture ex-type<br />
<strong>CBS</strong> 463.82.<br />
Notes: This fungus was originally maintained in the <strong>CBS</strong> collection<br />
as Anungitea uniseptata Matsush. In culture, however, only a<br />
pseudocladosporium-like state was observed. Conidiophores are<br />
reduced to conidiogenous cells, <strong>and</strong> have several apical loci as<br />
in Fusicladium, but are not subdenticulate; scars are somewhat<br />
darkened <strong>and</strong> thickened, not refractive. Conidia of F. africanum are<br />
(8–)11–17(–20) × 2–3(–3.5) µm, thus <strong>similar</strong>, but somewhat larger<br />
than the mean conidial size range (10–12 × 1.5–2 µm) observed<br />
in F. pini. <strong>The</strong> conidiogenous loci <strong>and</strong> conidial hila of F. africanum<br />
are also somewhat larger. Although the LSU sequence of F. pini is<br />
identical to that of F. ramoconidii, the ITS sequence <strong>similar</strong>ity is 97<br />
% (572/585 nucleotides).<br />
Fusicladium ramoconidii Crous & de Hoog, sp. nov. MycoBank<br />
MB504541. Figs 25–26.<br />
Etymology: Named after the presence of its characteristic<br />
ramoconidia.<br />
Fusicladio brevicatenato similis, sed ramoconidiis minoribus, (12–)15–17(–20) ×<br />
2(–3) µm, locis conidiogenis et hilis minoribus, 0.5–1 µm diam.<br />
Mycelium consisting of branched, septate, 1.5–2 µm wide hyphae,<br />
pale brown, smooth, frequently with hyphal coils. Conidiophores<br />
integrated into hyphae, <strong>and</strong> reduced to small, lateral protruding<br />
conidiogenous cells, concolorous with hyphae, or macronematous,<br />
dark brown, erect, thick-walled, 10–40 × 3–4 µm, 0–3-septate.<br />
Conidiogenous cells terminal, integrated, subcylindrical, tapering to<br />
a rounded apex, concolorous with hyphae (as hyphal pegs), or dark<br />
www.studiesinmycology.org<br />
211
Crous et al.<br />
Fusicladium rhodense Crous & M.J. Wingf., sp. nov. MycoBank<br />
MB504542. Fig. 27.<br />
Etymology: Named after the Greek Isl<strong>and</strong>, Rhodos, where it was<br />
collected.<br />
Fusicladio africano similis, sed locis conidiogenis angustioribus, 1.5–2 µm latis, et<br />
differt a F. pini ramoconidiis formantibus.<br />
Mycelium consisting of smooth to finely roughened, medium brown,<br />
branched, septate, 1.5–3 µm wide hyphae, frequently forming<br />
hyphal coils, giving rise to solitary, micronematous conidiophores.<br />
Conidiophores reduced to conidiogenous cells that are terminal<br />
or lateral on hyphae, medium brown, smooth, subcylindrical,<br />
subdenticulate, erect, or more distinct, up to 15 µm tall, 1.5–2<br />
µm wide, mono- to polyblastic; scars flat-tipped, somewhat<br />
darkened <strong>and</strong> thickened, but not refractive. Conidia in branched<br />
or unbranched chains of up to 15, pale brown in younger conidia,<br />
becoming medium brown, smooth, subcylindrical, 0–3-septate,<br />
tapering slightly towards the subtruncate ends, straight, but at times<br />
slightly curved, (8–)12–16(–20) × (2–)2.5–3(–4) µm; ramoconidia<br />
(0–)1(–3)-septate, 12–20 × 3–4 µm; conidia (0–)1-septate, 8–17<br />
× 2–3 µm; hila somewhat darkened <strong>and</strong> thickened, not refractive,<br />
1–1.5 µm wide.<br />
Fig. 26. Fusicladium ramoconidii (<strong>CBS</strong> 462.82). Conidiogenous cells with<br />
ramoconidia <strong>and</strong> conidia. Scale bar = 10 µm.<br />
brown on mononematous conidiophores, smooth, 3–15 × 2–3(–4)<br />
µm; proliferating sympodially, loci slightly thickened, darkened <strong>and</strong><br />
refractive, 0.5–1 µm wide. Conidia occurring in branched chains,<br />
narrowly ellipsoid to subcylindrical, pale olivaceous, guttulate;<br />
ramoconidia (0–)1(–3)-septate, (12–)15–17(–20) × 2(–3) µm;<br />
conidia occurring in short chains (–15), 0–1-septate, (8–)10–12(–<br />
16) × 2(–3) µm; hila slightly thickened <strong>and</strong> darkened, not refractive,<br />
0.5–1 µm wide.<br />
Cultural characteristics: Colonies erumpent, with sparse aerial<br />
mycelium <strong>and</strong> smooth margins on PDA, hazel to fawn (surface),<br />
with a thin, submerged margin; reverse brown-vinaceous; on OA<br />
hazel to fawn (surface) with a wide, fawn, submerged margin.<br />
Colonies reaching 25 mm diam on PDA after 1 mo at 25 °C in the<br />
dark; colonies fertile.<br />
Specimen examined: Netherl<strong>and</strong>s, Baarn, De Vuursche, needle of Pinus sp.<br />
(Pinaceae), 12 Apr. 1982, G.S. de Hoog, holotype <strong>CBS</strong> H-19908, culture ex-type<br />
<strong>CBS</strong> 462.82.<br />
Notes: This strain has been deposited in the <strong>CBS</strong> collection as<br />
Pseudocladosporium hachijoense (Matsush.) U. Braun. However,<br />
its ramoconidia <strong>and</strong> conidia are smaller than those cited by<br />
Matsushima (1975) (ramoconidia up to 30 µm long, conidia 10–21<br />
× 2–4 µm). Although it clusters with F. pini in the LSU phylogeny,<br />
there are 13 bp differences in their ITS sequence data. Furthermore,<br />
F. ramoconidii has ramoconidia which are absent in F. pini, <strong>and</strong><br />
has a faster growth rate, <strong>and</strong> hazel to fawn colonies, compared<br />
to the greyish sepia colonies of F. pini. <strong>The</strong> well-developed,<br />
septate conidiophores <strong>and</strong> ramoconidia are reminiscent of F.<br />
brevicatenatum, which differs, however, by its longer <strong>and</strong> wider<br />
ramoconidia, up to 30 × 6(–7) µm, as well as larger conidiogenous<br />
loci <strong>and</strong> conidial hila, 1.5–3 µm diam.<br />
Cultural characteristics: Colonies spreading, somewhat erumpent,<br />
with moderate aerial mycelium <strong>and</strong> crenate margins on PDA,<br />
uneven, greyish sepia (surface), margins fuscous-black; reverse<br />
fuscous-black; on OA smooth, spreading, with sparse aerial<br />
mycelium <strong>and</strong> even, regular margins, greyish sepia; on SNA<br />
spreading, smooth, even margins, sparse aerial mycelium, greyish<br />
sepia (surface). Colonies reaching 9 mm diam on PDA after 2 wk at<br />
25 °C in the dark; colonies fertile.<br />
Specimen examined: Greece, Rhodos, on branches of Ceratonia siliqua (Fabaceae),<br />
1 Jun. 2006, P.W. Crous & M.J. Wingfield, holotype <strong>CBS</strong> H-19910, culture ex-type<br />
<strong>CBS</strong> 121641 = CPC 13156.<br />
Note: Fusicladium rhodense has a typical pseudocladosporiumlike<br />
morphology in culture, with conidial scars that are somewhat<br />
darkened <strong>and</strong> thickened.<br />
Venturia hanliniana (U. Braun & Feiler) Unter., Mycologia 89: 129.<br />
1997.<br />
Basionym: Capronia hanliniana U. Braun & Feiler, Microbiol. Res.<br />
150: 90. 1995.<br />
≡ Caproventuria hanliniana (U. Braun & Feiler) U. Braun, in Braun,<br />
A Monograph of Cercosporella, Ramularia <strong>and</strong> Allied Genera<br />
(Phytopathogenic Hyphomycetes) 2: 396. 1998.<br />
Anamorph: Fusicladium brevicatenatum (U. Braun & Feiler)<br />
Crous, U. Braun & K. Schub., comb. nov. MycoBank MB504543.<br />
Basionym: Cladophialophora brevicatenata U. Braun & Feiler,<br />
Microbiol. Res. 150: 84. 1995.<br />
≡ Pseudocladosporium brevicatenatum (U. Braun & Feiler) U. Braun,<br />
in Braun, A Monograph of Cercosporella, Ramularia <strong>and</strong> Allied Genera<br />
(Phytopathogenic Hyphomycetes) 2: 393. 1998.<br />
Venturia hystrioides (Dugan, R.G. Roberts & Hanlin) Crous & U.<br />
Braun, comb. nov. MycoBank MB504544. Fig. 28.<br />
Basionym: Capronia hystrioides Dugan, R.G. Roberts & Hanlin,<br />
Mycologia 87: 713. 1995.<br />
≡ Caproventuria hystrioides (Dugan, R.G. Roberts & Hanlin) U. Braun,<br />
in Braun, A monograph of Cercosporella, Ramularia <strong>and</strong> allied genera<br />
(Phytopathogenic Hyphomycetes). Vol. 2: 396. 1998.<br />
Anamorph: Fusicladium sp.<br />
Only the anamorph was observed on OA, PDA <strong>and</strong> SNA in culture.<br />
212
Herpotrichiellaceae <strong>and</strong> Venturiaceae<br />
Fig. 27. Fusicladium rhodense (CPC 13156). A. Colony on OA. B. Conidial chains <strong>and</strong> hyphal coil. C–F. Chains of ramoconidia <strong>and</strong> conidia. Scale bar = 10 µm.<br />
Fig. 28. Venturia hystrioides (<strong>CBS</strong> 117727). A. Conidiophores giving rise to catenulate conidia. B. Ramoconidium giving rise to conidia. C–D. Conidial chains. E. Conidia <strong>and</strong><br />
conidiogenous cell with conidiogenous loci. F. Ramoconidium. G. Conidia. Scale bars = 10 µm.<br />
www.studiesinmycology.org<br />
213
Crous et al.<br />
Excluded taxa<br />
Polyscytalum fecundissimum Riess, Bot. Zeitung (Berlin) 11:<br />
138. 1853. Fig. 29.<br />
Cultural characteristics: Colonies erumpent, spreading, aerial<br />
mycelium sparse, margins smooth; colonies sienna to umber on<br />
PDA, with patches of greyish sepia; reverse chestnut-brown; on<br />
OA whitish due to moderate aerial mycelium, with diffuse umber<br />
pigment in the agar; whitish on SNA. Colonies reaching 15 mm<br />
diam on PDA after 3 wk at 25 °C in the dark.<br />
Specimen examined: Netherl<strong>and</strong>s, Schovenhorst, leaf litter of Fagus<br />
sylvatica (Fagaceae), 8 Nov. 1997, W. Gams, <strong>CBS</strong> H-6049, culture <strong>CBS</strong><br />
100506.<br />
Fig. 29. Polyscytalum fecundissimum (<strong>CBS</strong> 100506). Conidiophores giving rise to<br />
catenulate conidia. Scale bar = 10 µm.<br />
Mycelium consisting of branched, septate, smooth, guttulate,<br />
1.5–2.5 µm wide hyphae, pale brown, forming hyphal str<strong>and</strong>s.<br />
Conidiophores mostly reduced to conidiogenous cells, or if<br />
present, micronematous, consisting of a supporting cell, <strong>and</strong> single<br />
conidiogenous cell. Conidiogenous cells integrated in hyphae as<br />
lateral loci, or terminal, frequently disarticulating, subcylindrical,<br />
pale to medium brown, smooth, mono- to polyblastic, loci 1–1.5<br />
µm wide, 2.5 µm tall; conidiogenous cells subcylindrical, up to<br />
40 µm tall, <strong>and</strong> 2–2.5 µm wide. Conidia in long chains of up to<br />
60, branched or not, subcylindrical to narrowly ellipsoid, pale<br />
olivaceous to pale brown, smooth; ramoconidia 0–1(–3)-septate,<br />
15–20(–30) × 2–3(–3.5) µm; conidia 0(–1)-septate, 6–8(–12) ×<br />
2–3(–3.5) µm; hila 1–1.5 µm wide, inconspicuous to somewhat<br />
darkened, subtruncate.<br />
Cultural characteristics: Colonies erumpent, with sparse aerial<br />
mycelium on PDA, <strong>and</strong> smooth, even margins; olivaceous-grey<br />
to iron-grey (surface); reverse greenish black; on OA dark mousegrey<br />
(surface), with even, smooth margins. Colonies reaching 40<br />
mm diam after 2 wk at 25 °C in the dark; colonies fertile.<br />
Specimen examined: U.S.A., Washington, Wenatchee, on bing cherry fruit, Prunus<br />
avium cv. Bing (Rosaceae), R.G. Roberts, culture ex-type, ATCC 96019 = <strong>CBS</strong><br />
117727.<br />
Note: Dugan et al. (1995) commented that although <strong>similar</strong> to<br />
“Phaeoramularia” hachijoensis, the conidia of this species were<br />
predominantly aseptate <strong>and</strong> somewhat shorter than those described<br />
by Matsushima (1975).<br />
Notes: Polyscytalum fecundissimum is the type species of the <strong>genus</strong><br />
Polyscytalum. Several isolates of this species were investigated<br />
here to determine if Polyscytalum would be available for taxa that<br />
have a pseudocladosporium-like morphology. <strong>The</strong> clustering of<br />
<strong>CBS</strong> 681.74 within the Venturiaceae was surprising. However, this<br />
culture proved to be sterile, <strong>and</strong> therefore its identity could not be<br />
confirmed.<br />
Isolate <strong>CBS</strong> 109882 sporulated profusely. Colonies were greyolivaceous<br />
with olivaceous margins on PDA; conidiophores pale,<br />
<strong>and</strong> not dark brown as depicted for Polyscytalum in Ellis (1971);<br />
conidial chains were greenish yellow in mass, <strong>and</strong> pale olivaceousgreen<br />
under the dissecting microscope, somewhat roughened,<br />
polyblastic; on ITS sequence this isolate is identical to U57492,<br />
Cistella acuum (Alb. & Schwein.) Svrček (Helotiales), but the latter<br />
species should have a phialidic anamorph, so it is possible that<br />
this GenBank sequence is incorrect. <strong>The</strong> identity of <strong>CBS</strong> 109882<br />
therefore remains unresolved.<br />
Although isolate <strong>CBS</strong> 100506 is poorly sporulating, illustrations<br />
made in vitro when it was collected show this isolate to be<br />
authentic for the species <strong>and</strong> the <strong>genus</strong> Polyscytalum. Based on<br />
its LSU sequence, it is allied to Phlogicylindrium eucalypti Crous,<br />
Summerb. & Summerell (<strong>CBS</strong> 120080; Summerell et al. 2006), <strong>and</strong><br />
is therefore unrelated to the Venturiaceae.<br />
Zeloasperisporium R.F. Castañeda, Mycotaxon 60: 285. 1996,<br />
emend.<br />
Hyphomycetes. Mycelium mostly superficial, hyphae septate, brown<br />
to olivaceous. Hyphopodia absent. Conidiophores differentiated,<br />
mononematous, erect, aseptate or septate, brown to olivaceous.<br />
Conidiogenous cells integrated, terminal, proliferation sympodial,<br />
polyblastic, with subdenticulate, somewhat thickened <strong>and</strong> darkened<br />
scars. Conidia solitary, fusiform to obclavate or cylindrical, septate,<br />
asperulate to verrucose, olivaceous to brown, tips always hyaline,<br />
thinner-walled <strong>and</strong> smooth, forming mucoid app<strong>and</strong>ages, often only<br />
visible as a thickened frill. Synanamorph present, micronematous.<br />
Conidiogenous cells short cylindrical, antenna or hyphopodiumlike,<br />
phialidic, colarette sometimes present, aseptate, subhyaline.<br />
Conidia solitary, obovoid, ellipsoid, aseptate, brown to olivaceous,<br />
verruculose.<br />
Zeloasperisporium hyphopodioides R.F. Castañeda, Mycotaxon<br />
60: 285. 1996. Fig. 30.<br />
In vitro on OA: Mycelium internal to superficial, unbranched to<br />
sparingly branched, 1.5–3 µm wide, loosely septate, septa almost<br />
invisible, pale brown, smooth to asperulate, minutely verruculose,<br />
walls unthickened, sometimes inflated at the base of conidiophores.<br />
214
Herpotrichiellaceae <strong>and</strong> Venturiaceae<br />
Fig. 30. Zeloasperisporium hyphopodioides (<strong>CBS</strong> 218.95). A–B. Conidiogenous cells. C. Conidia with apical mucoid caps. D. Conidiogenous cell with sympodial proliferation.<br />
E–G. Conidiogenous cells of micronematous synanamorph. H. Conidia, <strong>and</strong> microconidia of synanamorph. Scale bars = 10 µm.<br />
Conidiophores macronematous, arising usually laterally from<br />
plagiotropous hyphae, erect, straight, subcylindrical or conical,<br />
not geniculate, usually unbranched, rarely branched, 13–45 ×<br />
3–4(–5) µm, slightly to distinctly attenuated towards the apex,<br />
tapered, aseptate, rarely with a single septum, pale brown to pale<br />
medium brown, smooth or minutely verruculose, walls unthickened,<br />
often somewhat constricted near the base. Conidiogenous cells<br />
integrated or conidiophores usually reduced to conidiogenous<br />
cells, subcylindrical to conical, proliferation sympodial, with a single<br />
or several subdenticulate to denticulate conidiogenous loci mostly<br />
crowded at or towards the apex, protuberant, truncate, 0.8–1.2 µm<br />
wide, thickened <strong>and</strong> darkened-refractive. Conidia solitary, straight<br />
to curved, ellipsoid, fusiform to obclavate, distinctly tapered towards<br />
the apex, apiculate, (12–)15–32 × 3.5–5.5 µm, (0–)1–2(–3)-septate,<br />
mainly 1-septate, usually constricted at the septa, pale brown to<br />
pale medium brown, asperulate to verruculose, walls unthickened or<br />
almost so, tips always hyaline, thinner-walled <strong>and</strong> smooth, forming<br />
mucoid appendages, often only visible as a thickened frill, base<br />
somewhat rounded or slightly bulbous, hila often situated on short<br />
peg-like prolongations, truncate, 0.8–1(–1.2) µm wide, thickened,<br />
darkened-refractive; microcyclic conidiogenesis occurring, conidia<br />
forming secondary conidiophores.<br />
Synanamorph micronematous. Conidiophores reduced<br />
to conidiogenous cells, numerous, occurring as short lateral<br />
prolongations of hyphae, antenna or telescope-like, cylindrical,<br />
www.studiesinmycology.org<br />
unbranched, conidiogenesis unclear, at times appearing<br />
phialidic, or having one to two apical scars; up to 5 µm long,<br />
1–1.5 µm wide, aseptate, subhyaline, smooth. Conidia of the<br />
micronematous anamorph quite different from the conidia formed<br />
by the macronematous conidiophores, solitary, obovoid, ellipsoid<br />
to somewhat fusiform, 5–9 × 2.5–3 µm, aseptate, pale to pale<br />
medium brown, verruculose, somewhat attenuated towards the<br />
base, hila flat, unthickened to somewhat thickened, appearing to<br />
have the ability to form a slime appendage at the apex.<br />
Cultural characteristics: Colonies on OA iron-grey to olivaceous due<br />
to abundant sporulation (surface); reverse black, velvety; margin<br />
regular to undulate, feathery; aerial mycelium absent or sparse,<br />
sporulation profuse.<br />
Specimen examined: Cuba, isolated from air, 2 Oct. 1994, R.F. Castañeda, INIFAT<br />
C94/114, holotype, <strong>CBS</strong>-H 5624, H-5639, isotypes, culture ex-type <strong>CBS</strong> 218.95 =<br />
INIFAT C94/114 = MUCL 39155 = IMI 367520.<br />
Notes: Within the course of the recent phylogenetic studies<br />
in Herpotrichiellaceae <strong>and</strong> Venturiaceae the type culture of<br />
Zeloasperisporium hyphopodioides has been included since it was<br />
deposited at the <strong>CBS</strong> as “Fusicladium hyphopodioides”. When<br />
the culture was re-examined, the described short appressoriumlike,<br />
inflated hyphopodia with slightly warted to lobed apices<br />
(Castañeda et al. 1996) could be recognised as conidiogenous<br />
cells of a synanamorph forming a second conidial type. In addition,<br />
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Crous et al.<br />
the conidial tips are hyaline, unthickened <strong>and</strong> smooth, <strong>and</strong> have the<br />
ability to form mucoid appendages that are often only visible as a<br />
thickened frill. <strong>The</strong>se two features, viz., the synanamorph <strong>and</strong> the<br />
conidia with mucoid appendages, easily distinguish this <strong>genus</strong> from<br />
morphologically <strong>similar</strong> genera such as Fusicladium, Asperisporium<br />
Maubl., <strong>and</strong> Passalora Fr. Phylogenetically Zeloasperisporium<br />
clusters basal to the Venturiaceae.<br />
Discussion<br />
<strong>The</strong> present paper was initiated to clarify the status of<br />
Cladophialophora <strong>and</strong> Pseudocladosporium spp., which appear<br />
morphologically <strong>similar</strong>. Confusion occurs when strains with this<br />
morphology are identified based solely on microscopic <strong>and</strong> cultural<br />
characteristics. <strong>The</strong> results clarify that Cladophialophora is allied to<br />
the Herpotrichiellaceae <strong>and</strong> Pseudocladosporium (= Fusicladium)<br />
to the Pleosporales (Dothideomycetes). <strong>The</strong> plant-pathogenic<br />
Cladophialophora species compose a separate clade within the<br />
order (Fig. 1). Another, somewhat remote chaetothyrialean clade<br />
contains extremotolerant, rock-inhabiting species around the<br />
<strong>genus</strong> Coniosporium Link (Cluster 5 of Haase et al. 1999). Both<br />
clades are significantly distinct from the prevalently hyperparasitic<br />
or oligotrophic, frequently opportunistic species of the remainder of<br />
the order (Fig. 1). This remainder includes all Capronia teleomorphs<br />
sequenced to date, <strong>and</strong> is thus likely to represent the family<br />
Herpotrichiellaceae. <strong>The</strong> ecological trends in each of the main<br />
clades of Chaetothyriales are thus quite different (Braun 1998).<br />
Several novelties are introduced within the preponderantly plantassociated<br />
clade of Chaetothyriales, including two new species<br />
associated with leaf spots. Cladophialophora is distinguished from<br />
Polyscytalum, which clusters outside the Herpotrichiellaceae, <strong>and</strong><br />
appears allied to Phlogicylindrium Crous, Summerb. & Summerell,<br />
a recently introduced <strong>genus</strong> for species occurring on Eucalyptus<br />
leaves (Summerell et al. 2006). Surprisingly Heteroconium<br />
chaetospira clusters in the Herpotrichiellaceae, <strong>and</strong> is placed<br />
in Cladophialophora as a distinctively pigmented member of the<br />
<strong>genus</strong>. Some species of Cladophialophora <strong>and</strong> Exophiala are<br />
newly described from a range of substrates such as fruit juices,<br />
drinking water <strong>and</strong> leaf litter, revealing the potential of these<br />
materials as ecological sources of inoculum for taxa associated<br />
with opportunistic human <strong>and</strong> animal infections.<br />
Furthermore, Pseudocladosporium belongs to the Venturiaceae,<br />
<strong>and</strong> is best treated as a synonym of Fusicladium, along with other<br />
genera as proposed by Schubert et al. (2003) <strong>and</strong> Beck et al. (2005).<br />
Although numerous isolates of the Venturiaceae were included for<br />
study, it was surprising to find relatively little variation within the<br />
family, suggesting that previously proposed teleomorph genera<br />
such as Apiosporina, Metacoleroa <strong>and</strong> Caproventuria should be<br />
best treated as synonyms of Venturia. <strong>The</strong> Venturiaceae is further<br />
extended with the inclusion of a novel sister clade of hyphomycetes<br />
with a pseudocladosporium-like morphology, which are also<br />
referred to as Fusicladium, thus widening the generic concept of<br />
the latter to encompass all pseudocladosporium-like anamorphs<br />
within the family. Some species assigned to Anungitopsis proved<br />
to cluster within the Venturiaceae, but the type species of the<br />
latter <strong>genus</strong>, A. speciosa, clustered elsewhere <strong>and</strong> possesses<br />
distinct conidiogenous loci, i.e., Anungitopsis cannot be reduced<br />
to synonymy with Fusicladium. <strong>The</strong> anamorphs of this sister clade<br />
of the main Venturia clade are morphologically rather close to<br />
taxa assigned to Anungitea. However, species of Anungitea <strong>and</strong><br />
Fusicladium are morphologically barely distinguishable (Schubert<br />
et al. 2003), but the true affinity of Anungitea depends on its type<br />
species of which cultures <strong>and</strong> sequence data are not yet available.<br />
Several anamorph genera with divergent morphologies were<br />
found to cluster together, suggesting that these are either different<br />
synanamorphs of the same teleomorph <strong>genus</strong>, or that they may<br />
represent cryptic clades that will diverge further once additional<br />
species are added in future studies. Although the Herpotrichiellaceae<br />
appeared to represent quite a diverse assembledge of morphotypes,<br />
the Venturiaceae were again surprisingly uniform.<br />
Acknowledgements<br />
Several colleagues from different countries provided material without which this work<br />
would not have been possible. We thank M. Vermaas for preparing the photographic<br />
plates, H.-J. Schroers for some gDNA <strong>and</strong> sequences used in this work, <strong>and</strong> A.<br />
van Iperen for maintaining the cultures. K. Schubert was financially supported by<br />
a Synthesys grant (No. 2559) which is gratefully acknowledged. R.C. Summerbell<br />
is thanked for his comments on an earlier version of the script. L. Hutchison is<br />
acknowledged for collecting Apiosporina collinsii for us.<br />
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Studies in Mycology 58: 219–234. 2007.<br />
Molecular analysis <strong>and</strong> pathogenicity of the Cladophialophora carrionii complex,<br />
with the description of a novel species<br />
G.S. de Hoog 1,2* , A.S. Nishikaku 3 , G. Fern<strong>and</strong>ez-Zeppenfeldt 4 , C. Padín-González 4 , E. Burger 3 , H. Badali 1 , N. Richard-Yegres 4 <strong>and</strong> A.H.G.<br />
Gerrits van den Ende 1<br />
1<br />
<strong>CBS</strong> Fungal Biodiversity Centre, Utrecht, <strong>The</strong> Netherl<strong>and</strong>s; 2 Institute for Biodiversity <strong>and</strong> Ecosystem Dynamics, University of Amsterdam, Amsterdam, <strong>The</strong> Netherl<strong>and</strong>s;<br />
3<br />
Laboratory of Immunopathology of Mycosis, Department of Immunology, São Paulo University, São Paulo, Brazil; 4 National Experimental University “Francisco de Mir<strong>and</strong>a”<br />
(UNEFM), Coro, Venezuela<br />
*Correspondence: Sybren de Hoog, s.hoog@cbs.knaw.nl<br />
Abstract: Cladophialophora carrionii is one of the four major etiologic agents of human chromoblastomycosis in semi-arid climates. This species was studied using sequence<br />
data of the internal transcribed spacer region of rDNA, the partial β-tubulin gene <strong>and</strong> an intron in the translation elongation factor 1-alpha gene, in addition to morphology.<br />
With all genes a clear bipartition was observed, which corresponded with minute differences in conidiophore morphology. A new species, C. yegresii, was introduced, which<br />
appeared to be, in contrast to C. carrionii, associated with living cactus plants. All strains from humans, <strong>and</strong> a few isolates from dead cactus debris, belonged to C. carrionii, for<br />
which a lectotype was designated. Artificial inoculation of cactus plants grown from seeds in the greenhouse showed that both fungi are able to persist in cactus tissue. When<br />
reaching the spines they produce cells that morphologically resemble the muriform cells known as the “invasive form” in chromoblastomycosis. <strong>The</strong> tested clinical strain of C.<br />
carrionii proved to be more virulent in cactus than the environmental strain of C. yegresii that originated from the same species of cactus, Stenocereus griseus. <strong>The</strong> muriform<br />
cell expressed in cactus spines can be regarded as the extremotolerant survival phase, <strong>and</strong> is likely to play an essential role in the natural life cycle of these organisms.<br />
Taxonomic novelty: Cladophialophora yegresii de Hoog, sp. nov.<br />
Key words: Cactus, chromoblastomycosis, Cladophialophora, endophyte, extremotolerance, phylogeny, taxonomy.<br />
INTRODUCTION<br />
Cladophialophora carrionii (Trejos) de Hoog, Kwon-Chung &<br />
McGinnis is one of the most frequent etiologic agents of human<br />
chromoblastomycosis, a chronic cutaneous disease characterised<br />
by verrucose skin lesions eventually leading to emerging, cauliflowerlike<br />
eruptions. <strong>The</strong> species is particularly observed in arid <strong>and</strong> semiarid<br />
climates of e.g. South <strong>and</strong> Central America (Lavelle 1980) <strong>and</strong><br />
Australia (Trejos 1954, Riddley 1957). <strong>The</strong> current hypothesis is that<br />
patients suffering from chromoblastomycosis are rural workers who<br />
acquire the infection after being pricked by cactus thorns or splinters<br />
(Rubin et al. 1991, Fernández-Zeppenfeldt et al. 1994). A classical<br />
case reported by O’Daly (1943) concerns traumatic inoculation with<br />
thorns of “guazábara” (Opuntia caribaea), a common xerophilic<br />
plant in semi-arid Venezuela. This hypothesised traumatic route of<br />
infection was later supported by Richard-Yegres & Yegres (Richard-<br />
Yegres & Yegres 1987; strain SR3 = <strong>CBS</strong> 863.96) <strong>and</strong> Fernández-<br />
Zeppenfeldt et al. (1994), who isolated strains from Prosopis<br />
juliflora litter. Cladophialophora Borelli has also been detected in<br />
association with spines of the common xerophyte Aloe vera <strong>and</strong> of<br />
the Cactaceae: Opuntia caribaeae, O. caracasana, Stenocereus<br />
griseus <strong>and</strong> Cereus lanuginosus (Borelli 1972, Yegres et al. 1996).<br />
Thorny American cacti are important components of the xerophyte<br />
flora of the arid climate of our study area in Falcon State, Venezuela<br />
(Richard-Yegres et al. 1992). Muriform cells are produced in<br />
human skin <strong>and</strong> represent the supposed pathogenic invasive form<br />
of fungi causing chromoblastomycosis (Mendoza et al. 1993).<br />
Early experiments involving the inoculation of several species of<br />
cold-blooded animals have shown the abundant production of the<br />
characteristic muriform cells in vivo (Trejos 1953).<br />
A <strong>similar</strong> plant origin of chromoblastomycosis has been<br />
supposed for a related agent of chromoblastomycosis, Fonsecaea<br />
pedrosoi (Brumpt) Negroni. Marques et al. (2006) isolated this<br />
species from the shells of Babassu coconuts (Orbignya phalerata).<br />
<strong>The</strong> habit of local people to sit on these shells might explain the<br />
frequent occurrence of lesions on the buttocks (Silva et al. 1995).<br />
Salgado et al. (2004) found the species on the thorns of a Mimosa<br />
pudica plant which a patient could identify as the source of traumatic<br />
onset of his chromoblastomycosis.<br />
Recently, with the development of molecular tools for species<br />
identification, doubt has arisen about the correctness of this<br />
supposed route of infection. <strong>The</strong> question whether environmental<br />
<strong>and</strong> clinical strains represent exactly the same species needs to be<br />
re-determined. In order to establish this for C. carrionii-associated<br />
chromoblastomycosis, reference strains from the <strong>CBS</strong> culture<br />
collection, supplemented with a large set of strains from semiarid<br />
Venezuela, have been verified using molecular tools that are<br />
currently routinely employed to answer taxonomic questions in<br />
black yeasts <strong>and</strong> their filamentous relatives (de Hoog et al. 2003),<br />
particularly the internal transcribed spacer (ITS) region of rDNA,<br />
the partial β-tubulin gene (BT2), <strong>and</strong> an intron in the translation<br />
elongation factor 1-alpha (EF1). In addition, a series of three<br />
experiments has been conducted concerning inoculation into<br />
<strong>and</strong> superficial application onto germlings of Stenocereus griseus<br />
obtained by cultivation in vitro, mature plants of S. griseus from the<br />
wild, <strong>and</strong> in spines of S. griseus collected in the semi-arid area of<br />
study. Our aim is to reveal the role of the cactus S. griseus in the<br />
life cycle of its associated Cladophialophora spp., <strong>and</strong> to determine<br />
whether a link could be made to C. carrionii for obtaining a better<br />
underst<strong>and</strong>ing of human chromoblastomycosis.<br />
MATERIALS AND METHODS<br />
Fungal strains <strong>and</strong> morphology<br />
Strains studied are listed in Table 1. This list comprises strains which<br />
have morphologically been identified as C. carrionii. Reference<br />
strains from the <strong>CBS</strong> culture collection, as well as fresh isolates<br />
from patients <strong>and</strong> the environment have been included. Strains<br />
were lyophilised <strong>and</strong> stored in liquid nitrogen soon after deposit at<br />
<strong>CBS</strong>. Stock cultures for transient working collections were grown<br />
on slants of 2 % malt extract agar (MEA) <strong>and</strong> oatmeal agar (OA) at<br />
24 °C. For morphological observation, slide cultures were made of<br />
strains grown on potato-dextrose agar (PDA) (de Hoog et al. 2000)<br />
<strong>and</strong> mounted in lactophenol cotton blue.<br />
219
De Hoog et al.<br />
Table 1. Isolation data of Cladophialophora strains examined.<br />
Name <strong>CBS</strong> nr. Other reference(s) 1 GenBank mtDNA*<br />
(Kawasaki<br />
ITS, BT2, EF1 et al. 1993)<br />
Source [human: duration, localization, sex, age<br />
(Pérez-Blanco et al. 2003)].<br />
Geography<br />
A / I / 1: C. carrionii<br />
117904 UNEFM 0004-02 = dH 14480 EU137281, –, – Chromoblastomycosis; 14 y; hip, thigh, leg; male 38 Falcon State, Venezuela<br />
117891 UNEFM 0002-00 = dH 14475 EU137278, –, EU137222 Chromoblastomycosis; 1 y; male 62 Falcon State, Venezuela<br />
117906 UNEFM 0014-96 = dH 14504 EU137288, EU137171, EU137231 Chromoblastomycosis; 0.5 y; h<strong>and</strong>; male 45 Falcon State, Venezuela<br />
117897 UNEFM 0011-03 = dH 14497 EU137314, –, EU137254 Chromoblastomycosis; 0.5 y; h<strong>and</strong>; male 42 Falcon State, Venezuela<br />
859,96 UNEFM 9617 = dH 10703 EU137295, EU137178, EU137237 Dry plant debris, arid zone Falcon State, Venezuela<br />
117898 UNEFM 0010-98 = dH 14496 EU137308, –, EU137246 Chromoblastomycosis; 20 y; h<strong>and</strong>; female 59 Falcon State, Venezuela<br />
117889 UNEFM 0003-04 = dH 14478 –, EU137190, – Chromoblastomycosis; 20 y; thigh, leg; female 78 Falcon State, Venezuela<br />
114392 UNEFM 82267 = dH 13261 EU137267, EU137150, EU137211 Chromoblastomycosis; leg; female Falcon State, Venezuela<br />
114394 UNEFM 9803 = dH 13263 EU137307, –, EU137245 Chromoblastomycosis; h<strong>and</strong>; male 22 Falcon State, Venezuela<br />
114396 UNEFM 2001/1 = dH 13265 EU137269, EU137152, EU137213 Chromoblastomycosis; arm; male 35 Falcon State, Venezuela<br />
114399 UNEFM 2003/2 = dH 13268 EU137272, EU137155, EU137216 Chromoblastomycosis; arm; female 64 Falcon State, Venezuela<br />
114401 UNEFM 9901 = dH 13270 EU137274, EU137157, EU137218 Chromoblastomycosis; arm; female 40 Falcon State, Venezuela<br />
114402** UNEFM 9902 = dH 13271 EU137275, EU137158, EU137219 Chromoblastomycosis; arm; female 40 Falcon State, Venezuela<br />
114403 UNEFM 95195 = dH 13272 EU137276, EU137159, EU137220 Chromoblastomycosis; arm; male Falcon State, Venezuela<br />
117899 UNEFM 0010-04 = dH 14495 EU137301, EU137183, EU137241 Chromoblastomycosis; 2 y; h<strong>and</strong>; male 57 Falcon State, Venezuela<br />
117901 UNEFM 0009-03 = dH 14492 EU137312, EU137197, EU137252 Chromoblastomycosis; 8 y; arm; female 41 Falcon State, Venezuela<br />
114393 UNEFM 9801 = dH 13262 EU137268, EU137151, EU137212 Chromoblastomycosis; h<strong>and</strong>; male 72 Falcon State, Venezuela<br />
108.97** UNEFM 9501 = dH 10704 EU137306, EU137188, EU137265 Chromoblastomycosis; skin Falcon State, Venezuela<br />
114397 UNEFM 84020 = dH 13266 EU137270, EU137153, EU137214 Chromoblastomycosis; h<strong>and</strong>, arm; male 54 Falcon State, Venezuela<br />
114404 UNEFM 95656 = dH 13273 EU137311, EU137196, EU137251 Chromoblastomycosis; arm; male Falcon State, Venezuela<br />
117902 UNEFM 0008-03 = dH 14489 EU137283, EU137166, EU137226 Chromoblastomycosis; 3 y; arm; male 42 Falcon State, Venezuela<br />
117893 UNEFM 0001-00 = dH 14470 EU137316, EU137200, – Chromoblastomycosis; 2 y; knee; male 19 Falcon State, Venezuela<br />
117892 UNEFM 0001-02 = dH 14471 EU137277, EU137160, EU137221 Chromoblastomycosis; 8 y; knee; male 52 Falcon State, Venezuela<br />
117908 UNEFM 0013-04 = dH 14502 –, EU137191, – Chromoblastomycosis; 6 y; back; male 13 Falcon State, Venezuela<br />
109.97** UNEFM 9503 = dH 10706 –, –, – Chromoblastomycosis; skin Falcon State, Venezuela<br />
857.96 UNEFM 9408 = dH 10707 EU137294, EU137177, EU137236 Chromoblastomycosis; skin Falcon State, Venezuela<br />
114398 UNEFM 2003/1 = dH 13267 EU137271, EU137154, EU137215 Chromoblastomycosis; arm; female 67 Falcon State, Venezuela<br />
114400 UNEFM 2003/3 = dH 13269 EU137273, EU137156, EU137217 Chromoblastomycosis; arm; male 50 Falcon State, Venezuela<br />
117909 UNEFM 0013-00 = dH 14501 EU137287, EU137170, EU137230 Chromoblastomycosis; arm; male Falcon State, Venezuela<br />
114395 UNEFM 9802 = dH 13264 EU137299, EU137182, EU137240 Chromoblastomycosis; leg; female 22 Falcon State, Venezuela<br />
166.54 MUCL 10088 EU137290, EU137173, – Skin lesion in human Falcon State, Venezuela<br />
862.96 UNEFM 9603 = dH 10700 EU137315, EU137199, EU137255 Dry plant debris, semi-arid zone Falcon State, Venezuela<br />
863.96** IFM 41444 = UNEFM SR3 = dH 10699 AB109169 / EU137296, EU137179, EU137238 Dry spine (Opuntia caribaea) on soil, semi-arid zone Falcon State, Venezuela<br />
861.96 UNEFM 9607 = dH 10701 EU137309, EU137194, EU137249 Dry plant debris, semi-arid zone Falcon State, Venezuela<br />
117896 dH 14498 EU137285, –, EU137228 H<strong>and</strong> lesion Falcon State, Venezuela<br />
114397 UNEFM 84020 = dH 13266 EU137270, EU137153, EU137214 Chromoblastomycosis, h<strong>and</strong> <strong>and</strong> arm Falcon State, Venezuela<br />
220
Cladophialophora carrionii complex<br />
117905 dH 14505 EU137300, –, – Chromoblastomycosis, h<strong>and</strong>, male Falcon State, Venezuela<br />
117900 dH 14493 EU137284, –, EU137227 Chromoblastomycosis, h<strong>and</strong>, male Falcon State, Venezuela<br />
114392 UNEFM 82267 = dH 13261 EU137267, EU137150, EU137211 Chromoblastomycosis, leg, female Falcon State, Venezuela<br />
– FMC 248 AF397181, –, – Chromoblastomycosis Venezuela<br />
– IFM 41807 AB109175, –, – Group mt-I – Venezuela<br />
– IFM 4812 AB109168, –, – Group mt-I – Venezuela<br />
– IMTSP 690 AF397180, –, – Chromoblastomycosis Brazil<br />
410.96 UAMH 4392 = NCMH 1010 = DUKE 2403 EU137310, EU137195, EU137250 Chromoblastomycosis –<br />
163.54 EU137304, EU137186, EU137243 Chromoblastomycosis Australia<br />
117903 dH 14482 EU137282, –, EU137225 Chromoblastomycosis, forearm, male –<br />
362.70 M.J. Campos 4555 = dH 15806 EU137302, EU137184, EU137242 Human Mozambique<br />
260.83 CDC B-1352 = FMC 282 = ATCC 44535 (ex-T of C. ajelloi) EU137292, EU137175, EU137234 Group mt-I Skin lesion in human Ug<strong>and</strong>a<br />
986.96 UAMH 5717 EU137297, EU137180, – Clinical material –<br />
– IFM 4805 AB087204, –, – – –<br />
– IFM 4811 AB109178, –, – – –<br />
– IFM 41814 AB109176, –, – – –<br />
B / II / 2: C. carrionii<br />
160.54 ATCC 16264 = CDC A-835 = MUCL 40053 = IFM 4808 (ex-LT of C. carrionii) AB109177 / EU137266, EU137201, EU137210 Group mt-II Chromoblastomycosis, human Australia<br />
– Todd Pryce 200867 = dH 13218 Human Australia<br />
406.96 MRL 1114 = UAMH 4366 = dH 15847 EU137317, EU137202, EU137256 Human Queensl<strong>and</strong>, Australia<br />
100434 ATCC 32279 = dH 10745 = IP 518 = RV 16499 EU137289, EU137172, EU137232 Human Madagascar<br />
– IFM 4810 AB109170, –, – – –<br />
– IFM 41446 = DCU 606 AB109171, –, – – –<br />
C: C. carrionii<br />
– IFM 41651 AB109174, –, – Group mt-I – China<br />
– IFM 41650 AB109173, –, – Group mt-I – China<br />
– IFM 41641 AB109172, –, – Group mt-I – China<br />
– IFM 4985 AB109179, –, – – –<br />
– IFM 4986 AB109180, –, – – –<br />
D / III / 3: C. yegresii<br />
114406 UNEFM SgSR1 = dH 13275 EU137323, EU137208, EU137263 Stenocereus griseus asymptomatic plant Falcon State, Venezuela<br />
114407 UNEFM SgSR2 = dH 13276 EU137324, –, EU137264 Stenocereus griseus asymptomatic plant Falcon State, Venezuela<br />
114405** UNEFM SgSR3 = dH 13274 (ex-T of C. yegresii) EU137322, EU137209, EU137262 Stenocereus griseus asymptomatic plant Falcon State, Venezuela<br />
1 Abbreviations: ATCC = American Type Culture Collection, Manassas, U.S.A.; <strong>CBS</strong> = Centraalbureau voor Schimmelcultures, Utrecht, <strong>The</strong> Netherl<strong>and</strong>s; CDC = Centers for Disease Control <strong>and</strong> Prevention, Atlanta, U.S.A.; DCU = Department of<br />
Dermatology, School of Medicine, Chiba, Japan; dH = G.S. de Hoog working collection; FMC = Faculdade de Medicina, Caracas, Venezuela; ITMSP = Instituto de Medicina Tropical de São Paulo, São Paulo, Brazil; IFM = Research Center for<br />
Pathogenic Fungi <strong>and</strong> Microbial Toxicoses, Chiba University, Chiba, Japan; IP = Institut Pasteur, Paris, France; MUCL = Mycotheque de l’Université de Louvain, Louvain-la-Neuve, Belgium; RV = Prince Leopold Institute of Tropical Medicine, Antwerp,<br />
Belgium; UAMH = <strong>The</strong> University of Alberta Microfungus Collection <strong>and</strong> Herbarium, Edmonton, Canada; UNEFM = Universidade Nacional Experimental Francisco de Mir<strong>and</strong>a, Coro, Falcon, Venezuela.<br />
Ex-T = Type strain; ex-LT = Lectotype strain.<br />
*Type I <strong>and</strong> II groups based on mitochondrial DNA restriction fragment length polymorphism: H-1 <strong>and</strong> H-2 = restriction patterns 1 <strong>and</strong> 2, respectively, with HaeIII enzyme; M-1 <strong>and</strong> M-2 = restriction patterns 1 <strong>and</strong> 2, respectively, with MspI enzyme; S-1<br />
<strong>and</strong> S-2 = restriction patterns 1 <strong>and</strong> 2, respectively, with Sau3AI enzyme.<br />
** Used in plant <strong>and</strong> mouse inoculation experiments.<br />
www.studiesinmycology.org<br />
221
De Hoog et al.<br />
Table 2. Results from MrAic using corrected Akaike Information Criterion (AICc).<br />
Fragment/Gene Model df* lnL* AICc* wAICc*<br />
rRNA ITS TrNG 89 -21.840.556 4575.9992” 0.3080<br />
EF-1α HKYG 88 -19.392.355 41.147.171 0.5242<br />
ß-Tubulin SYMIG 90 -28.547.745 59.261.933 0.1913<br />
*df = degrees of freedom; lnL = log likelihood; AICc = corrected AIC; wAICc = weighted corrected AIC.<br />
DNA extraction<br />
Approximately 1 cm 2 mycelium of 30-d-old cultures was transferred<br />
to a 2 mL Eppendorf tube containing 300 µL TES-buffer (Tris 1.2 %<br />
w/v, Na-EDTA 0.38% w/v, SDS 2 % w/v, pH 8.0) <strong>and</strong> about 80 mg<br />
of a silica mixture (Silica gel H, Merck 7736, Darmstadt, Germany<br />
/ Kieselguhr Celite 545, Machery, Düren, Germany, 2 : 1, w/w).<br />
Cells were disrupted mechanically in a tight-fitting sterile pestle for<br />
approximately 1 min. Subsequently 200 µL TES-buffer was added,<br />
the mixture was vortexed, 10 µL proteinase K was added <strong>and</strong><br />
incubated for 10 min at 65 °C. After addition of 140 µL of 5 M NaCl<br />
<strong>and</strong> 1/10 vol CTAB 10 % (cetyltrimethylammoniumbromide) buffer,<br />
the material was incubated for 30 min at 65 °C. Subsequently<br />
700 µL SEVAG (24 : 1, chloroform : isoamylalcohol) was mixed to<br />
solution, incubated during 30 min on ice water <strong>and</strong> centrifuged for<br />
10 min at 14 000 rpm. <strong>The</strong> supernatant was transferred to a new<br />
tube with 225 µL 5 M NH 4<br />
-acetate, incubated on ice water <strong>and</strong><br />
centrifuged again for 10 min at 14 000 rpm. <strong>The</strong> supernatant was<br />
transferred to another Eppendorf tube with 0.55 vol isopropanol <strong>and</strong><br />
spun for 5 min at 14 000 rpm. Subsequently, the pellet was washed<br />
with ice cold 70 % ethanol. After drying at room temperature it was<br />
re-suspended in 48.5 µL TE buffer (Tris 0.12 % w/v, Na-EDTA 0.04<br />
% w/v) plus 1.5 µL RNAse 20 U/mL <strong>and</strong> incubated for 15–30 min<br />
at 37 °C.<br />
Sequencing <strong>and</strong> phylogenetic reconstruction<br />
Three loci, namely the internal transcribed spacers (ITS), ß-<br />
tubulin (BT2) <strong>and</strong> translation elongation factor 1-α (EF1), were<br />
sequenced. For ITS sequencing, amplification was performed<br />
with V9G (5’-TTACGTCCCTGCCCTTTGTA-3’) <strong>and</strong> LS266 (5’-<br />
GCATTCCCAAACAACTCGACTC-3’). Sequencing reactions<br />
were conducted with ITS1 <strong>and</strong> ITS4 primers (White et al.<br />
1990). For BT2 amplification <strong>and</strong> sequencing, primers Bt2a<br />
(5’-GGTAACCAAATCGGTGCTGCTTTC-3’) <strong>and</strong> Bt2b (5’-<br />
ACCCTCAGTGTAGTGACCCTTGGC-3’) were used (Glass &<br />
Donaldson 1995) <strong>and</strong> for EF1 amplification <strong>and</strong> sequencing, primers<br />
EF1-728F (5’CATCGAGAAGTTCGAGAAGG-3’) <strong>and</strong> EF1-986R<br />
(5’-TACTTGAAGGAACCCTTACC-3’) (Carbone & Kohn, 1999).<br />
Sequences were aligned in BioNumerics v. 4.5 (Applied Maths,<br />
Kortrijk, Belgium), exported <strong>and</strong> converted into Phylip interleaved<br />
format (Felsenstein 1993).<br />
Calculation of ILD (incongruence length difference) was<br />
performed in PAUP v. 4.0b10 (Swofford 2003). A combined data set<br />
of ITS, EF1 <strong>and</strong> BT2 sequences was created. Optimality criterion<br />
was set to parsimony. <strong>The</strong> total number of characters was 1 263<br />
with equal weight, while 677 characters were constant, <strong>and</strong> 396<br />
parsimony-informative. Gaps were treated as missing, <strong>and</strong> treebisection-reconnection<br />
(TBR) was used as branch-swapping<br />
algorithm. Maximum number of trees was set to 100 <strong>and</strong> left<br />
unchanged.<br />
Substitution model testing<br />
<strong>The</strong> program MrAic (www.abc.se/~nyl<strong>and</strong>er/; Nyl<strong>and</strong>er 2004)<br />
was used to select a substitution model. MrAic is a Perl script for<br />
calculating the Akaike Information Criterion (AIC), corrected Akaike<br />
Information Criterion (AICc), Bayesian Information Criterion (BIC),<br />
<strong>and</strong> Akaike weights for nucleotide substitution models <strong>and</strong> model<br />
uncertainty. Using an ML algorithm, likelihood scores under different<br />
models were estimated using Phyml (http://atgc.lirmm.fr/phyml/). All<br />
56 models implemented in Modeltest (Posada & Cr<strong>and</strong>all 1998)<br />
were evaluated. <strong>The</strong>se models were also combined with proportion<br />
of invariable sites (I) <strong>and</strong>/or gamma distribution shape parameter<br />
(G). A difference between Modeltest <strong>and</strong> MrAic is that the latter<br />
does not evaluate all models on the same, approximate topology<br />
as in PAUP (Swofford 1981). Instead, Phyml was used to try to find<br />
the maximum of the likelihood function under all models. This is<br />
necessary for finding AIC, AICc, or BIC for the models. <strong>The</strong> AICc<br />
calculation (Table 2) was used to select the right model for the ratio<br />
of parameters to characters (Nchar/Nparameters < 40; Burnham &<br />
Anderson 2002) for all loci. <strong>The</strong> substitution matrix of the models<br />
is printed next to the trees. Another advantage of using MrAic in<br />
combination with Phyml was the obtained accuracy of tree topology<br />
<strong>and</strong> the greater calculation speed (Guindon & Gascuel 2003).<br />
Population genetic analyses<br />
In order to confirm the intraspecific diversity shown in the MP trees,<br />
the number of populations in the C. carrionii complex was inferred<br />
with Structure v. 2.2 (Pritchard et al. 2000) using genotype data<br />
of the ITS regions of rRNA gene <strong>and</strong> of the partial EF1 <strong>and</strong> BT2<br />
genes. Genotypes of these three loci of 43 isolates were sorted<br />
on the basis of sequence <strong>similar</strong>ity. Structure is a model-based<br />
clustering method for using multilocus genotype data to infer<br />
population structure <strong>and</strong> assign individuals to populations. <strong>The</strong><br />
parameters were as follows: the length of burn-in period was<br />
set to 10 6 , number of MCMC repeats after burn-in 30 000; the<br />
ancestry model: admixture (individuals have mixed ancestry <strong>and</strong><br />
is recommended as starting point for most analyses). Uniform prior<br />
for ALPHA was set to 1.0 (default) <strong>and</strong> all allele frequencies were<br />
taken as independent among populations with λ set to 1.0 (default).<br />
Probability of the data (for estimating K) was also computed<br />
(Falush et al. 2003). <strong>The</strong> burn-in period length <strong>and</strong> number of<br />
MCMC repetitions after burn-in were set as 10 000 <strong>and</strong> 100 000,<br />
<strong>and</strong> admixture model <strong>and</strong> allele frequencies correlated model were<br />
chosen for analysis. <strong>The</strong> number of populations (K) was assumed<br />
from two to four.<br />
Association of multilocus genotypes was screened with the<br />
multilocus option in BioNumerics. To test for reproductive mode in<br />
each population, index of association (I A<br />
, a measure of multilocus<br />
linkage disequilibrium) was calculated with Multilocus v. 1.2.2 (www.<br />
bio.ic.ac.uk/evolve/software/multilocus). <strong>The</strong> null hypothesis for this<br />
analysis was complete panmixia. <strong>The</strong> values of I A<br />
were compared<br />
between observed <strong>and</strong> r<strong>and</strong>omised data sets. <strong>The</strong> hypothesis would<br />
be rejected when p < 0.05. Population differentiation (index: theta,<br />
θ) was also detected using the same software <strong>and</strong> a null hypothesis<br />
for this analysis is no population differentiation. When observed θ is<br />
statistically significantly different from those of r<strong>and</strong>om datasets (p<br />
< 0.05), population differentiation should be considered.<br />
222
Cladophialophora carrionii complex<br />
A reticulogram was reconstructed using T-rex (Makarenkov<br />
2001, Makarenkov & Legendre 2004) (www.labunix.uqam.ca/<br />
~makarenv/trex.html) on C. carrionii / Cladophialophora sp. <strong>The</strong><br />
program first computed a classical additive tree using one of the<br />
five available tree reconstruction algorithms. Subsequently, at each<br />
step of the procedure, a reticulation (a new edge) was chosen that<br />
minimised the least-squares or the weighted least-squares loss<br />
function; it was added to the growing reticulogram. Two statistical<br />
criteria (Q1 <strong>and</strong> Q2) were proposed to measure the gain in fit when<br />
reticulations were added. <strong>The</strong> minimum of each of these criteria<br />
may suggest a stopping rule for addition of reticulations. With<br />
HGT (horizontal gene transfer) reticulogram reconstruction option<br />
(Makarenkov 2001) the program mapped the gene tree into the<br />
species tree using the least-squares method. Horizontal transfers<br />
of the considered gene were then shown in the species tree. <strong>The</strong><br />
reticulate network was created in the ITS tree, which served as<br />
a species tree <strong>and</strong> compared with a gene tree, EF1. Degrees of<br />
recombination or horizontal gene transfer were also visualised<br />
using SplitsTree v. 4.8 software (Huson & Bryant 2006). Split<br />
decomposition (B<strong>and</strong>elt & Dress 1992) was applied on three loci of<br />
the entire C. carrionii complex. Calculation was done with default<br />
settings of characters transformation using uncorrected P-values,<br />
equal angles <strong>and</strong> optimise box iterations set to 1. Star- or brushlike<br />
trees indicate clonal development, while reticulation indicates<br />
genetic exchange.<br />
Isolation of fungi for inoculation experiments<br />
Nine plants of Stenocereus griseus, located within 50 m radius<br />
of the house of a patient with chromoblastomycosis due to strain<br />
UNEFM 9902 = <strong>CBS</strong> 114402 (C. carrionii) in Sabaneta (Mir<strong>and</strong>a,<br />
Falcón State, Venezuela), were analysed. Four fragments of<br />
approx. 2 × 3 × 1 cm were excised from each plant at brownish<br />
superficial lesions in upper branches. Sampled fragments were<br />
soaked in mineral oil for 15 min at 23 °C under agitation at 150<br />
rpm (Fernández-Zeppenfeldt et al. 1994). Subsequently four<br />
cultivations were made per sample on agar slants. Strains with<br />
cultural characteristics <strong>and</strong> morphology <strong>similar</strong> to C. carrionii (de<br />
Hoog et al. 2000) were selected. Final identification was made by<br />
sequencing of the ITS region, by determining the ability of strains<br />
to grow at 35, 37, 38 <strong>and</strong> 40 °C, <strong>and</strong> whether they could break<br />
down 20 % gelatin (Richard-Yegres & Yegres 1987, Fernández-<br />
Zeppenfeldt et al. 1994). Environmental strain UNEFM-SgSR3 =<br />
<strong>CBS</strong> 114405 (Cladophialophora sp.) <strong>and</strong> clinical strain UNEFM<br />
9902 = <strong>CBS</strong> 114402 (C. carrionii) were selected for the inoculation<br />
experiments.<br />
Inoculum preparation<br />
Approximately 1 cm 2 of a culture on Sabouraud’s glucose agar<br />
(SGA) was transferred to 50 mL YPG medium (yeast extract 0.5<br />
%, peptone 0.5 %, glucose 2 %) (de Hoog et al. 2000), shaken at<br />
150 rpm <strong>and</strong> incubated for 3 d at 23 °C (Yegres et al. 1991). Five<br />
mL aliquots of the starter culture were transferred serially every 4<br />
d to 500 mL flasks containing 100 mL synthetic medium (d-glucose<br />
2 %, KH 2<br />
PO 4<br />
0.2 %, NH 4<br />
SO 4<br />
0.1 %, urea 0.03 %, MgSO 4<br />
0.03<br />
%, CaCl 2<br />
0.003 %; pH 6.2) shaken at 150 rpm at 23 °C. After 4 d<br />
the suspensions, which were predominantly conidia, were filtered<br />
through sterile gauze, ground in 50 mL 0.85 % saline, centrifuged<br />
at 2 000 rpm, <strong>and</strong> repeatedly washed with saline until a clear<br />
supernatant was obtained. <strong>The</strong> suspensions were adjusted to 5 ×<br />
10 6 cells/mL (Yegres et al. 1991, Cermeño & Torres 1998). Inocula<br />
of 2 mL were checked for viability in lactritmel medium (de Hoog et<br />
al. 2000).<br />
www.studiesinmycology.org<br />
Experimental cactus germlings<br />
Young cactus plants (Stenocereus griseus) were obtained in the<br />
laboratory (Clausnitzer 1978) by cultivation from seeds of a single<br />
cardon fruit collected near the house of the patient infected with <strong>CBS</strong><br />
114402 in the endemic area for chromoblastomycosis in Falcon<br />
State, Venezuela. <strong>The</strong> seeds were rinsed with sterile distilled water,<br />
the contents washed by agitation for 10 min at 120 rpm in 250 mL<br />
sodium hypochlorite 4 % (v/v), <strong>and</strong> subsequently with sterile distilled<br />
water at 120 rpm for 5 min. <strong>The</strong> supernatant was decanted, 250 mL<br />
HCl 20 % was added, the seeds were incubated for 3 h, decanted<br />
<strong>and</strong> washed repeatedly with sterile distilled water. Seeds were<br />
then dried for 24 h on filter paper at 37 °C. Onset of germination<br />
was obtained by incubation of the seeds in a moist chamber on<br />
filter paper for 15 d under alternately 8 h of continuous white light<br />
(26 W) <strong>and</strong> 16 h of darkness; bud emergence was observed daily.<br />
Germlings of 1 cm in length, with green colour <strong>and</strong> having two<br />
leaves were transplanted to 128-container germinators until roots<br />
developed. <strong>The</strong> sterile substrate contained 5 parts Sogemix® <strong>and</strong><br />
1 part river soil from the region where the fruit was collected. <strong>The</strong><br />
daily light regime was as above; plants were watered every 10 d<br />
with 5 mL sterile tap water for 1 yr.<br />
Inoculation of S. griseus germlings<br />
Fungal suspensions (0.1 mL) were either injected using a syringe<br />
(13 × 0.4 mm) at a depth of approximately 5 mm into cortical<br />
tissue (Fig. 1A), or superficially applied onto (Fig. 1B) 96 r<strong>and</strong>omly<br />
selected 1-yr-old plants: 50 % using clinical strain <strong>CBS</strong> 114402<br />
(C. carrionii) <strong>and</strong> 50 % using environmental strain <strong>CBS</strong> 114405<br />
(Cladophialophora sp.). <strong>The</strong> controls were 64 plants which were<br />
treated <strong>similar</strong>ly, but using sterile saline (0.85 %). <strong>The</strong> growth<br />
chambers with inoculated plants stayed in the laboratory under the<br />
conditions specified above. From day 15 post inoculation onwards<br />
every 15 th day, six plants of each treatment were sectioned<br />
longitudinally from the apex <strong>and</strong> transversely by means of a h<strong>and</strong>held<br />
microtome, examined directly in glycerin water (25 %), <strong>and</strong><br />
cultured in lactritmel medium (Fernández-Zeppenfeldt et al. 1994).<br />
Experimental cactus plants<br />
A total of 150 whole S. griseus plants ≤ 15 cm tall <strong>and</strong> without<br />
macroscopically visible lesions, were dug from an area within a<br />
50 m radius of the house of the patient with chromoblastomycosis<br />
as specified above. Plants were transported to the laboratory <strong>and</strong><br />
transplanted individually into polyethylene bags with a capacity of<br />
1 kg, using as substrate river soil from the same area. Plants were<br />
maintained outside, directly adjacent to the laboratory to adjust<br />
at average temperatures of 32 °C <strong>and</strong> with natural daylight. <strong>The</strong>y<br />
were watered with tap water every 15 th d for a period of 6 mo.<br />
Scar formation in mature S. griseus plants<br />
For inoculation purposes, 150 sharp, wooden toothpicks 4 × 0.3 ×<br />
0.2 cm were washed <strong>and</strong> boiled for 3 min in tap water to eliminate<br />
resins (Yegres & Richard-Yegres 2002). This procedure was<br />
repeated three times. Three batches of 50 toothpicks each were<br />
kept separate in Petri dishes. Plates were incubated for 15 d at<br />
23 °C after inoculating each batch with 1 mL fungal suspension (5<br />
× 10 6 cells/mL) of either strain <strong>CBS</strong> 114405 or <strong>CBS</strong> 114402, with<br />
sterile water as control. A total of 50 r<strong>and</strong>omly selected plants were<br />
inoculated (Fig. 1C) halfway up the shaft with a toothpick colonised<br />
with <strong>CBS</strong> 114402 (C. carrionii), <strong>CBS</strong> 114405 (Cladophialophora<br />
sp.) or the control (Yegres & Richard-Yegres 2002).<br />
Starting from 2 wk post inoculation, 10 plants were r<strong>and</strong>omly<br />
chosen every 15 d, <strong>and</strong> tissue samples taken at the point of<br />
223
De Hoog et al.<br />
+ + + - -<br />
-<br />
+<br />
+<br />
A B C D<br />
Clinical C. carrionii<br />
> Environmental C. yegresii<br />
Fig. 1. Diagram of inoculation experiments with results. A. Inoculation of young cactus; B. Superficial application of young cactus; C. Traumatic application of mature cactus,<br />
with brown resulting scar; D. Superficial application of mature spines. Indications +/- refer to positive resp. negative results of re-isolated strains. Lower line: circles represent<br />
production of muriform cells, filaments represent hyphal growth.<br />
Fig. 2. Split decomposition of the C. carrionii complex using SplitsTree with uncorrected (P-value) distances. Nodes are shown only with different genotypes; hence EF1 shows<br />
the largest number of nodes. Extensive reticulation is noted in all loci.<br />
224
Cladophialophora carrionii complex<br />
0.1<br />
99<br />
<strong>CBS</strong>102227<br />
<strong>CBS</strong>83496<br />
92<br />
94<br />
85<br />
<strong>CBS</strong>25983<br />
<strong>CBS</strong>45482<br />
100<br />
<strong>CBS</strong>117901<br />
<strong>CBS</strong>117891<br />
<strong>CBS</strong>117906<br />
<strong>CBS</strong>117892<br />
<strong>CBS</strong>114403<br />
<strong>CBS</strong>117899<br />
<strong>CBS</strong>114392<br />
<strong>CBS</strong>114393<br />
<strong>CBS</strong>41096<br />
<strong>CBS</strong>117903<br />
<strong>CBS</strong>114395<br />
<strong>CBS</strong>114398<br />
<strong>CBS</strong>114404<br />
<strong>CBS</strong>117900<br />
<strong>CBS</strong>117909<br />
dH16363<br />
<strong>CBS</strong>114399<br />
<strong>CBS</strong>117905<br />
<strong>CBS</strong>16654<br />
<strong>CBS</strong>114396<br />
<strong>CBS</strong>117904<br />
<strong>CBS</strong>114402<br />
<strong>CBS</strong>114397<br />
<strong>CBS</strong>85796<br />
<strong>CBS</strong>114394<br />
<strong>CBS</strong>86396<br />
<strong>CBS</strong>86196<br />
<strong>CBS</strong>16354<br />
<strong>CBS</strong>117896<br />
<strong>CBS</strong>85896<br />
<strong>CBS</strong>36270<br />
<strong>CBS</strong>26083<br />
<strong>CBS</strong>40696<br />
<strong>CBS</strong>100434<br />
<strong>CBS</strong>16054<br />
<strong>CBS</strong>114407<br />
<strong>CBS</strong>114405<br />
<strong>CBS</strong>114406<br />
dH14614<br />
K=5 K=4<br />
Venezuela<br />
Venezuela<br />
Venezuela<br />
Venezuela<br />
Venezuela<br />
Venezuela<br />
Venezuela<br />
Venezuela<br />
unknown<br />
Venezuela<br />
Venezuela<br />
Venezuela<br />
Venezuela<br />
Venezuela<br />
Venezuela<br />
unknown<br />
Venezuela<br />
Venezuela<br />
Venezuela<br />
Venezuela<br />
Venezuela<br />
Venezuela<br />
Venezuela<br />
Venezuela<br />
Venezuela<br />
Venezuela<br />
Venezuela<br />
Australia<br />
Venezuela<br />
Venezuela<br />
Mozambique<br />
Ug<strong>and</strong>a<br />
Australia<br />
Madagascar<br />
Australia<br />
Venezuela<br />
Venezuela<br />
Venezuela<br />
I<br />
II<br />
III<br />
Fig. 3. Phylogenetic tree (Neighbour-joining) of the C. carrionii complex based on EF1 (with grouping I–III) using the same strains of Fig. 1, generated using the HKY+G model.<br />
<strong>The</strong> model was calculated using ML in MrAic software. Bootstrap cut-off = 80 %. <strong>CBS</strong> 834.96 was taken as outgroup. Columns were generated with Structure software<br />
hypothesising K = 4 <strong>and</strong> K = 5, <strong>and</strong> alleles independent. Geographical origins in black refer to isolates from humans (chromoblastomycosis); origins in green refer to isolates<br />
from plant material.<br />
inoculation, <strong>and</strong> from the thorns directly adjacent to this area.<br />
Samples were rinsed with 4 % sodium hypochlorite for 3 min, <strong>and</strong><br />
subsequently washed in sterile distilled water for re-isolations, <strong>and</strong><br />
for histological study by means of light microscopy (Fernández-<br />
Zeppenfeldt et al. 1994).<br />
Experiments with spines of S. griseus<br />
Ninety cactus spines of 2.5 cm av. length were collected from<br />
a single S. griseus plant located near the home of the patient<br />
infected with <strong>CBS</strong> 114402, at approx. 2.5 m height, superficially<br />
sterilised, <strong>and</strong> divided into three groups, of which 30 spines were<br />
inoculated with <strong>CBS</strong> 114402 (C. carrionii), 30 with <strong>CBS</strong> 114405<br />
(Cladophialophora sp.) <strong>and</strong> 30 to be used as control, inoculated<br />
with a saline solution (Fig. 1D). A <strong>similar</strong> series composed of 90<br />
spines of 1.5 cm average length was collected at approx. 1 m<br />
height. All spines were incubated in sterile Petri dishes with filter<br />
paper (Whatman #1) with 2 mL saline solution; subsequently 0.1<br />
mL fungal suspension was applied. Twenty spines were analysed<br />
weekly by means of longitudinal sectioning with a h<strong>and</strong>-held<br />
microtome, cultured as above <strong>and</strong> observed microscopically until<br />
day 75 post incubation.<br />
www.studiesinmycology.org<br />
225
De Hoog et al.<br />
ROOT<br />
<strong>CBS</strong>114398<br />
<strong>CBS</strong>117892<br />
<strong>CBS</strong>16654<br />
<strong>CBS</strong>86196<br />
<strong>CBS</strong>41096<br />
<strong>CBS</strong>114404<br />
<strong>CBS</strong>114395<br />
<strong>CBS</strong>114392<br />
<strong>CBS</strong>85796<br />
<strong>CBS</strong>117903<br />
<strong>CBS</strong>117899<br />
<strong>CBS</strong>117909<br />
<strong>CBS</strong>114403<br />
<strong>CBS</strong>114399<br />
5<br />
<strong>CBS</strong>114397<br />
<strong>CBS</strong>85896<br />
<strong>CBS</strong>114402<br />
<strong>CBS</strong>114396<br />
<strong>CBS</strong>117896<br />
<strong>CBS</strong>117904<br />
dH16363<br />
<strong>CBS</strong>16354<br />
<strong>CBS</strong>86396<br />
<strong>CBS</strong>117906<br />
<strong>CBS</strong>117900<br />
<strong>CBS</strong>117891<br />
<strong>CBS</strong>117905<br />
<strong>CBS</strong>114394<br />
<strong>CBS</strong>117901<br />
<strong>CBS</strong>114393<br />
<strong>CBS</strong>40696<br />
<strong>CBS</strong>16054<br />
<strong>CBS</strong>100434<br />
<strong>CBS</strong>36270<br />
1 7<br />
4 6<br />
3 2 9<br />
10<br />
8<br />
<strong>CBS</strong>26083<br />
A<br />
B<br />
<strong>CBS</strong>114406<br />
<strong>CBS</strong>114405<br />
<strong>CBS</strong>114407<br />
A<br />
Unknown<br />
Australia<br />
Australia<br />
Australia<br />
Madagascar<br />
Mozambique<br />
D<br />
Ug<strong>and</strong>a<br />
C. carrionii<br />
C. yegresii<br />
Fig. 4. Reticulogram of South American strains of Cladophialophora species <strong>and</strong> strains from other continents (mentioned) constructed with T-rex software. ITS rDNA (with<br />
grouping A, B, D) was used as species tree <strong>and</strong> compared with the ß-tubulin gene tree. First 10 reticulations are shown with numbers.<br />
Statistics<br />
Survival of the cactus seedlings <strong>and</strong> collected plants following<br />
inoculation were evaluated using the X 2 -test (P = 0.05 was<br />
considered significant). Stem lesions resulting from inoculations<br />
were analysed with Student’s T-test (P = 0.01 was considered<br />
significant).<br />
RESULTS<br />
<strong>The</strong> rDNA ITS region was sequenced for 43 strains identified as<br />
C. carrionii based on morphology. Sequences of 16 additional<br />
strains were downloaded from GenBank. Five distantly related,<br />
unidentified cladophialophora-like species were added, with <strong>CBS</strong><br />
834.96 as outgroup. In C. carrionii, 203 positions were compared<br />
in ITS1, 158 in the 5.8S rRNA gene <strong>and</strong> 182 in ITS2 (Table 3).<br />
<strong>The</strong> sequences could be aligned with confidence over their entire<br />
lengths. Over the data set, 35 positions were polymorphic, of which<br />
33 were phylogenetically informative (Table 3), the two remaining<br />
being variable T-repeats near the ends of ITS1 <strong>and</strong> 2.<br />
For ITS sequences the AICc selected the TrN+G model (TrNG;<br />
Tamura & Nei 1993). <strong>The</strong> base frequency of ITS: T = 0.2467, C<br />
= 0.2897, A = 0.2247, G = 0.2390, TC = 0.5364, AG = 0.4636.<br />
<strong>The</strong> EF1 tree was built with substitution model HKY+G; the base<br />
frequency of EF1: T = 0.2990, C = 0.2665, A = 0.2123, G = 0.2221,<br />
TC = 0.5655, AG = 0.4345. <strong>The</strong> best model for BT2 sequences<br />
was the SYM+I+G (symmetrical model). <strong>The</strong> base frequency for<br />
BT2: T = 0.2255, C = 0.2953, A = 0.2463, G = 0.2328, TC = 0.5208,<br />
AG = 0.4792. Bootstrap values of the EF1 tree were calculated<br />
with PAUP using parsimony <strong>and</strong> with maxtrees set to 500 <strong>and</strong> 500<br />
replicates (data not shown). Total number of characters was 191 of<br />
which 101 were parsimony-informative. Tree length was 365 <strong>and</strong><br />
had the following indices: Consistency Index = 0.685, Retention<br />
Index = 0.542 <strong>and</strong> Homoplasy Index = 0.315.<br />
<strong>The</strong> original tree length, L o<br />
was 1 055, the tree length of the<br />
combined data, L c<br />
was 1 062. <strong>The</strong> resulting incongruence length<br />
difference L = (L c<br />
–L o<br />
) was 7 (P = 0.24). <strong>The</strong> observed ILD was not<br />
significantly greater than expected by chance <strong>and</strong> it was concluded<br />
that the sequences were congruent <strong>and</strong> could be used together in<br />
a combined analyses.<br />
Split decomposition based on the same alignment generated<br />
extensive recombination. <strong>The</strong> structure found with three loci was<br />
robust, with the exception of separation of <strong>CBS</strong> 834.96 <strong>and</strong> <strong>CBS</strong><br />
102227 with EF1 (Fig. 2).<br />
<strong>The</strong> core of the network, comprising the strains listed in Table 1,<br />
was analysed in more detail. With ITS, four groups were recognised<br />
(A–D; Table 1). (A) was the main group with 36 strains / sequences;<br />
FMC 248 differed only by a small T-repeat <strong>and</strong> was regarded as a<br />
member of (A). <strong>The</strong> remaining groups were smaller, differing from<br />
group (A) maximally by two consistent positions (Table 3). Group<br />
(C) mainly comprised sequences from GenBank <strong>and</strong> all originated<br />
from Abliz et al. (2004). One of the strains of group (C), IFM 4808,<br />
concerned a subculture of <strong>CBS</strong> 160.54, which is an original isolate<br />
of Trejos (1954) representing C. carrionii. Re-sequencing indicated<br />
that it was a member of group (B). Analysis of our electropherograms<br />
of this isolate was not suggestive of heterothallism. None of the<br />
positions characterising groups (A)–(C) were also found to differ in<br />
group (D), which deviated in 16 mutations in ITS1 <strong>and</strong> 8 in ITS2;<br />
17 of the mutations were transitions, 7 were transversions <strong>and</strong> 7<br />
indels. Group (D) was clearly distinct from the complex of (A)–(C),<br />
226
Cladophialophora carrionii complex<br />
Table 3. Nucleotide variability of ITS1-2 ribosomal DNA regions of<br />
Cladophialophora carrionii (A – C) <strong>and</strong> C. yegresii (D).<br />
rDNA domains (length), with variable nucleotide positions.<br />
ITS1 (201-203) A B C D<br />
16 C C C T<br />
17 T C T T<br />
19 T T T C<br />
51 A A A G<br />
57 A A A T<br />
90-92 TG- TG- TG- CGT<br />
101 T T T C<br />
103 C C C T<br />
104 G A G G<br />
106 A A A G<br />
114 T T T C<br />
122 T T T C<br />
132 C C C T<br />
137 A A A C<br />
141 C C C T<br />
145 - - - A<br />
163-170 6-10T 6-10T 6-10T TTGTATCT<br />
180 - - - A<br />
183 G G G A<br />
190 T/A A A A<br />
5.8S (158) Monomorphic<br />
ITS2 (178-182) A B C D<br />
36 C T T T<br />
48 T T G T<br />
49 T T T C<br />
51 - - - C<br />
114 C C C G<br />
140 A A A G<br />
155 - - - T<br />
178-179 -- -- -- CT<br />
with a total of 27 mutations.<br />
For multilocus analysis with ITS, EF1 <strong>and</strong> BT2 a smaller set<br />
of strains was compared. Sequences of the 205 bp long element<br />
of EF1 contained 32 phylogenetically informative mutations. Three<br />
entities were distinguished (I–III; Fig. 3). With BT2, three groups<br />
with the same composition were recognised. Strains of ITS group<br />
(C) were not available for study.<br />
On the basis of multilocus screening in BioNumerics, concordant<br />
groups (A)–(D) were tested with the Structure programme. When<br />
K was set at 4 or 5, consistent groupings were noted, indicated as<br />
I, II <strong>and</strong> III (Fig. 3), corresponding with ITS groups (A), (B) <strong>and</strong> (D),<br />
respectively in Table 1.<br />
<strong>The</strong> possibility that group (D) / (III) included a member of another,<br />
morphologically <strong>similar</strong> but phylogenetically unrelated group of fungi<br />
was excluded by SSU sequencing. Genera morphologically <strong>similar</strong><br />
to Cladophialophora, such as <strong>Cladosporium</strong> Link, Devriesia Seifert<br />
www.studiesinmycology.org<br />
& N.L. Nick., Phaeoramularia Munt.-Cvetk., Pseudocladosporium<br />
U. Braun <strong>and</strong> Stenella Syd. proved to be remote (data not shown).<br />
With T-rex, interaction between groups (B) <strong>and</strong> (D) was noted,<br />
rather than between groups (B) <strong>and</strong> (A), despite the high sequence<br />
<strong>similar</strong>ity of (A) <strong>and</strong> (B) (Fig. 4).<br />
Morphological observation revealed that representatives of<br />
ITS groups (A)–(C) generally had conidiophores that arise at right<br />
angles from creeping hyphae (Fig. 5), while those of (D) tend to<br />
be ascending, hyphae gradually becoming conidiophore-like. Since<br />
slight correspondence was found in independent markers <strong>and</strong><br />
phenetic criteria, we considered group (D) to represent a separate<br />
species, which is described as follows.<br />
Cladophialophora yegresii de Hoog, sp. nov. MycoBank<br />
MB500208. Figs 6, 7D–F.<br />
Etymology: Named after Francisco Yegres, Venezuelean<br />
mycologist.<br />
Coloniae in agaro PDA dicto 22 °C planae, olivaceo-virides, pulverulentae vel<br />
velutinae, margine integra; reversum olivaceo-atrum. Hyphae fertiles dilute olivaceovirides,<br />
ascendentes, paulatim in catenas conidiorum concolorium vertentes.<br />
Conidiorum catenae ramosae, conidia dilute olivaceo-viridia, levia et tenuitunicata,<br />
4.5–6 × 2.5 µm, faciliter liberata, cicatricibus modice pigmentatis. Chlamydosporae<br />
et cellulae zymosae absentes. Synanamorphe phialidica non visa. Teleomorphe<br />
ignota.<br />
Holotypus cultura sicca <strong>CBS</strong> H-18464 in herbarium <strong>CBS</strong> praeservatur.<br />
Colonies on PDA at 22 °C evenly olivaceous green, powdery to<br />
velvety, with entire margin; reverse olivaceous black. Fertile hyphae<br />
pale olivaceous green, ascending, gradually changing over into<br />
concolorous chains of conidia. Conidial system profusely branched.<br />
Conidia pale olivaceous green, smooth- <strong>and</strong> thin-walled, 4.5–6<br />
× 2.5 µm, detached rather easily, with slightly pigmented scars.<br />
Chlamydospores <strong>and</strong> yeast cells absent. Phialidic synanamorph<br />
not observed. Teleomorph unknown.<br />
Specimen examined: Venezuela, Falcon state, from asymptomatic Stenocereus<br />
griseus cactus, G. Fernández-Zeppenfeldt, <strong>CBS</strong> H-18464 holotype, culture ex-type<br />
<strong>CBS</strong> 114405 = UNEFM SgSR3.<br />
Notes: Of the 48 <strong>dematiaceous</strong> isolates obtained from 36 fragments<br />
of the cactus Stenocereus griseus, four strains originating from<br />
four different plants of S. griseus presented morphological <strong>and</strong><br />
physiological key characteristics of Cladophialophora carrionii or<br />
C. yegresii (de Hoog et al. 2000, 2006). Gelatin liquefaction was<br />
negative in all strains <strong>and</strong> the maximum growth temperature was<br />
37 °C. After identification to species level using sequence data (de<br />
Hoog et al. 2006), both C. carrionii <strong>and</strong> C. yegresii appeared to be<br />
among the strains isolated.<br />
A total of 256 plants obtained at the end of 1 yr from germlings,<br />
had ribs, spines, <strong>and</strong> an average height of 15 cm. <strong>The</strong> 96<br />
germlings inoculated with fungal suspensions of the test strains<br />
<strong>CBS</strong> 114402 (C. carrionii, clinical) <strong>and</strong> <strong>CBS</strong> 114405 (C. yegresii,<br />
environmental) remained without visible external lesions during the<br />
year of experimentation. Histological sections of the 96 inoculated<br />
plants consistently revealed internal growth of the fungi in their<br />
filamentous form. Muriform cells were not observed, neither on<br />
the epidermis, nor in the internal tissue, spines or roots. <strong>The</strong> reisolated<br />
cultures demonstrated the viability of the fungi during the<br />
entire experimental process: <strong>CBS</strong> 114402 (C. carrionii) was grown<br />
from 26 (54.16 %) of the plants <strong>and</strong> <strong>CBS</strong> 114405 (C. yegresii) in<br />
23 (47.90 %) of the plants. <strong>The</strong> X 2 test did not reveal significant<br />
differences between the isolates (X 2 c = 0.0729 < X 2 t = 3.84). <strong>The</strong><br />
32 control plants remained without external lesions, <strong>and</strong> in the<br />
227
De Hoog et al.<br />
Fig. 5. Microscopic morphology of C. carrionii, strain <strong>CBS</strong> 160.54. Conidiophore erect, i.e. mostly arising at 90° from creeping hypha (sketch). Scale bar = 10 µm.<br />
histological sections no internal or external fungal elements were<br />
observed. None of the fungi isolated from the control plants proved<br />
to be a species of Cladophialophora.<br />
With 96 plants with superficial application of spore suspension<br />
(48 plants for each isolate, either clinical or environmental) neither<br />
internal nor external lesions were observed. Histological sectioning<br />
did not reveal fungal elements in or on plant tissue. Short hyphal<br />
elements <strong>and</strong> meristematic cells were occasionally seen around<br />
<strong>and</strong> inside the outer layers of the spines. <strong>The</strong> re-isolated strains<br />
proved that the fungi survived during the entire experimental<br />
procedure: <strong>CBS</strong> 114402 (C. carrionii) was isolated from 32 (66.67<br />
%) plants <strong>and</strong> <strong>CBS</strong> 114405 (C. yegresii) from 33 (68.75 %). <strong>The</strong> X 2<br />
test did not detect significant differences in survival rates among<br />
the isolates (X 2 c = 0.4375 < X 2 t = 3.84).<br />
Mature plants inoculated using colonised toothpicks showed<br />
average scarring of 1.88 cm diam with C. carrionii <strong>and</strong> 1.33 cm<br />
diam with C. yegresii, around the point of inoculation. In histological<br />
sections of 100 plants, dark, septate hyphae with inflated elements<br />
were observed at the points of inoculation. Muriform cells were not<br />
observed. Re-isolated strains were evidence of isolate viability:<br />
228
Cladophialophora carrionii complex<br />
Fig. 6. Microscopic morphology of C. yegresii, strain <strong>CBS</strong> 114405. Conidiophore ascending, i.e. mostly emerging from hyphal end that is gradually growing upwards to become<br />
a conidiophore (sketch). Scale bar = 10 µm.<br />
<strong>CBS</strong> 114402 (C. carrionii) was grown from 36 (72 %) plants <strong>and</strong> <strong>CBS</strong><br />
114405 (C. yegresii) from 30 (60 %). <strong>The</strong> fungi could not be isolated<br />
from spines. <strong>The</strong> 50 plants used as controls showed scarring of<br />
1.06 cm diam on average around the point of inoculation. No fungal<br />
elements were seen in direct examinations <strong>and</strong> histological sections<br />
of these plants. <strong>The</strong> retro-cultures were negative. <strong>The</strong> scarring<br />
responses of the plants to the clinical strain, environmental strain<br />
<strong>and</strong> control proved to be highly significant:<br />
Clinical <strong>CBS</strong> 114402 vs. environmental <strong>CBS</strong> 114405: þ = 0.000832,<br />
P = 0.01;<br />
Clinical <strong>CBS</strong> 114402 vs. control: þ = 0.00003128, P = 0.01;<br />
Environmental <strong>CBS</strong> 114405 vs. control: þ = 0.005343, P = 0.01.<br />
Spines 2.5 cm av. in length, seeded with suspensions of <strong>CBS</strong><br />
114402 (C. carrionii) <strong>and</strong> <strong>CBS</strong> 114405 (C. yegresii), developed<br />
toruloid hyphal elements with some dark, swollen cells <strong>similar</strong><br />
to muriform cells known in human tissue. <strong>The</strong> re-isolated strains<br />
proved the species to survive during the experimental procedure<br />
(< 75 d). Similar results were obtained with the spines 1.5 cm av.<br />
in length. No cladophialophora-like fungi were isolated from the<br />
controls.<br />
www.studiesinmycology.org<br />
229
De Hoog et al.<br />
Fig. 7. Conidial morphology in selected branches of (upper row: A–C) C. carrionii, strain <strong>CBS</strong> 260.83; (lower row: D–F) C. yegresii, strain <strong>CBS</strong> 114405. In this respect the two<br />
species are identical. Scale bar = 10 µm.<br />
DISCUSSION<br />
Taxonomy of Cladophialophora<br />
Judging from SSU rDNA phylogeny data, all Cladophialophora<br />
species that are consistently associated with pathology to humans<br />
belong to the Herpotrichiellaceae in the order Chaetothyriales<br />
(Haase et al. 1999). Within this order, the <strong>genus</strong> Cladophialophora<br />
is polyphyletic. Conidia of all species are produced in branched<br />
chains on poorly differentiated hyphae. This very simply structured<br />
conidial system may lead to confusion with morphologically<br />
<strong>similar</strong> but unrelated fungi that are encountered as contaminants<br />
in the hospital environment. <strong>The</strong> <strong>genus</strong> <strong>Cladosporium</strong> comprises<br />
ubiquitous airborne fungi which mostly have erect, more or less<br />
differentiated conidiophores, <strong>and</strong> dark conidial scars. <strong>The</strong>y are<br />
associated with Davidiella Crous & U. Braun teleomorphs <strong>and</strong><br />
belong to the Dothideomycetes, family Davidiellaceae (Braun et al.<br />
2003, Schoch et al. 2006). Pseudocladosporium was introduced by<br />
Braun (1998) with three species differing from Cladophialophora<br />
mainly by intercalary hyphal cells with lateral extensions that bear<br />
conidial chains, having Caproventuria U. Braun teleomorphs (see<br />
Crous et al. 2007 – this volume). <strong>The</strong> group is classified in the<br />
Venturiaceae <strong>and</strong> Mycosphaerellaceae in the Dothideales (Braun et<br />
al. 2003). <strong>The</strong> anamorph <strong>genus</strong> Devriesia comprises thermophilic<br />
saprobes with a cladophialophora-like appearance <strong>and</strong> producing<br />
230
Cladophialophora carrionii complex<br />
dark, multi-celled chlamydospores alongside the hyphae.<br />
Phylogenetically this <strong>genus</strong> is related to the Mycosphaerellaceae,<br />
in the Dothideomycetes (Seifert et al. 2004).<br />
Cladophialophora carrionii was originally introduced by Trejos<br />
(1954) on the basis of 46 strains from Venezuela, Australia <strong>and</strong><br />
South Africa. He did not indicate a holotype. For this reason isolate<br />
Trejos 27 = Emmons 8619 = <strong>CBS</strong> 160.54, the first strain mentioned<br />
by Trejos (1954), is selected here as representative for C. carrionii.<br />
A dried specimen of this strain has been deposited as lectotype<br />
in the Herbarium of the Centraalbureau voor Schimmelcultures as<br />
<strong>CBS</strong> H-18465.<br />
<strong>The</strong> ex-type strain of Cladophialophora ajelloi Borelli, <strong>CBS</strong><br />
260.83, proved to be indistinguishable from C. carrionii, which was<br />
also known to be able to produce phialides in addition to catenate<br />
conidia (Honbo et al. 1984). Remarkably, a strain identified as C.<br />
ajelloi from Samoa (<strong>CBS</strong> 259.83; Goh et al. 1982) proved to be<br />
related to but consistently different from all strains of the C. carrionii<br />
complex. <strong>The</strong> 43-year-old male patient in otherwise good health<br />
carrying this fungus had a 5 × 3 cm erythematous, scaling lesion<br />
on his arm. Muriform cells were present in superficial dermis <strong>and</strong><br />
stratum corneum. This clearly represents yet another agent of<br />
human chromoblastomycosis. <strong>The</strong> name C. ajelloi is not available<br />
for this taxon, as this is a synonym of C. carrionii. <strong>The</strong> taxon will be<br />
formally described in a forthcoming paper.<br />
Members of ITS groups (A)–(D) were shown to be close to<br />
each other in SSU phylogeny (data not shown) underlining that all<br />
analysed species were correctly assigned to Cladophialophora.<br />
This <strong>genus</strong> was defined by melanised acropetal chains of conidia,<br />
near absence of conidiophores, <strong>and</strong> phylogenetic affinity to the<br />
order Chaetothyriales. Strains (A)–(D) clustered in a clade which<br />
contained a mixture of species of Cladophialophora, Fonsecaea<br />
Negroni <strong>and</strong> Phialophora Medlar. From a point of view of human<br />
disease, the species of the clade were known as agents of brain<br />
infection [C. bantiana (Sacc.) de Hoog et al., F. monophora (M. Moore<br />
& F.P. Almeida) de Hoog et al.], disseminated disease [C. devriesii<br />
(A.A. Padhye & Ajello) de Hoog et al.], cutaneous disease [C. boppii<br />
(Borelli) de Hoog et al.] <strong>and</strong> particularly chromoblastomycosis (C.<br />
carrionii, Fonsecaea, Phialophora).<br />
Diversity of Cladophialophora carrionii / C. yegresii<br />
Infraspecific variability was observed within C. carrionii. <strong>The</strong> groups<br />
(A)–(C) were separated on the basis of five mutations in the ITS<br />
region, which were supported by mutations in EF1 <strong>and</strong> BT2, as<br />
confirmed by analysis in Structure, where the same separation<br />
(K = 5) of entities was observed. Furthermore, K = 4 unites groups<br />
(B/II) <strong>and</strong> (D/III), despite the fact that the sequence of (B) is more<br />
close to those of (A). With T-rex software a <strong>similar</strong> relationship<br />
between [(B), C. carrionii] <strong>and</strong> [(D), C. yegresii] was noted,<br />
suggesting horizontal gene flow between these entities. This is<br />
remarkable, since (C) strains predominantly inhabit remote deserts<br />
in Madagascar <strong>and</strong> Australia, while (D) is found in equally remote<br />
localities in Venezuela. Extensive reticulation was observed in all<br />
genes with SplitsTree. With ITS <strong>and</strong> BT2, <strong>CBS</strong> 834.96 <strong>and</strong> <strong>CBS</strong><br />
102227 cluster closely together, while in the more variable EF1<br />
data these are all widely apart, suggesting that in Cladophialophora<br />
other mechanisms than recombination may occur.<br />
Group (C) contained ITS sequences taken from the public<br />
domain, originating from a single study (Abliz et al. 2004).<br />
Remarkably, strain IFM 4808 found in group (C) on the basis of data<br />
from Abliz et al. (2004), was the same isolate as <strong>CBS</strong> 160.54, which<br />
was found repeatedly in group (B) in our data set (Table 1). A <strong>similar</strong><br />
www.studiesinmycology.org<br />
phenomenon was observed with strain IFM 41444 = <strong>CBS</strong> 863.96,<br />
of which GenBank deposition AB109169 consistently deviated from<br />
our data in a frequently observed mutation. A possible explanation<br />
of these consistent sequence conflicts is heterozygosity. Although<br />
most chaetothyrialean fungi are supposed to be haploid (Szaniszlo<br />
2002; Zeng et al. 2007), some strains have a double DNA content<br />
in yeast cells (Ohkusu et al. 1999). Teleomorphs are not known<br />
in Cladophialophora <strong>and</strong> related black yeasts, but many species<br />
are known to form profuse hyphal anastomoses (de Hoog et al.<br />
2006), allowing parasexual processes <strong>and</strong> mitotic recombination.<br />
However, all electropherograms including those from the study of<br />
Abliz et al. (2004), which were kindly sent by K. Fukushima (Chiba,<br />
Japan), were unambiguous, without double peaks. This matched<br />
with the observation of preponderant clonality despite frequent<br />
anastomoses in Exophiala J.W. Carmich. (Zeng et al. 2007). An<br />
alternative explanation might be the occurrence of paralogous ITS<br />
repeats, as reported earlier in Fusarium Link (O’Donnell & Cigelnik<br />
1997).<br />
<strong>The</strong> remaining diversity within C. carrionii as confirmed by<br />
Structure shows some geographical structuring of populations,<br />
in that group (A) does not occur in Asia, group (B) is limited to<br />
Australia <strong>and</strong> Africa, <strong>and</strong> group (C) has thus far only been reported<br />
from Asia. <strong>The</strong> wide distribution of most genotypes suggests,<br />
however, that worldwide occurrence is likely to become apparent<br />
when more strains have been analysed. All climate zones where<br />
C. carrionii was isolated were semi-arid to arid, desert-like.<br />
Genotypes were not limited to the endemic semi-arid areas, <strong>and</strong><br />
thus a relatively rapid vector of dispersal has to be hypothesised<br />
enabling the fungus to cross climate zones where the saprobic<br />
phase is unable to survive. Kawasaki et al. (1993) analysed three<br />
further loci in mtDNA using RFLP. Only some of their strains were<br />
available for sequencing. <strong>The</strong>se had all identical mtDNA profiles,<br />
with the exception of IFM 4808 = <strong>CBS</strong> 160.54, that differed in two<br />
markers (Table 1). If we assume that there is no real separation of<br />
ITS groups (A) <strong>and</strong> (C) (see above), the conclusion is warranted<br />
that mtDNA allows distinction of polymorphism at the same level of<br />
diversity as detected in this study with ITS, EF1 <strong>and</strong> BT2.<br />
South America harbours group (D) which represents a second<br />
species, C. yegresii. This species thus far has not been found<br />
on humans, <strong>and</strong> seems to be restricted to living Stenocereus<br />
cactus plants. Nishimura et al. (1989) published a strain from<br />
chromoblastomycosis in China which matched the morphology of<br />
strains now classified as C. yegresii, but as far as we are aware this<br />
strain has not been sequenced.<br />
Ecology <strong>and</strong> virulence of Cladophialophora carrionii / C.<br />
yegresii<br />
Cladophialophora carrionii was preponderantly found as an agent<br />
of human infection <strong>and</strong> only occasionally on dead plant debris,<br />
mainly seceded cactus needles. <strong>The</strong> only three strains available<br />
of C. yegresii were isolated from living, asymptomatic Stenocereus<br />
plants surrounding the cabin of a symptomatic patient from whom C.<br />
carrionii, <strong>CBS</strong> 114402 was isolated. Although in some publications<br />
convincing evidence was presented that infections originate from<br />
puncture by plant material (e.g., Salgado et al. 2004), it now<br />
becomes clear that the environmental look-alikes of clinical strains<br />
do not necessarily belong to the same species (Crous et al. 2006,<br />
Mostert et al. 2006), but may be members of other, related taxa with<br />
slightly different ecology; an unambiguous connection between a<br />
clinical <strong>and</strong> an environmental strain still has to be proven.<br />
<strong>The</strong> endemic area of the two species, C. carrionii <strong>and</strong> C.<br />
231
De Hoog et al.<br />
yegresii, has a semi-arid climate, with average yearly temperatures<br />
of 24 ºC, scarce rainfall (up to 600 annual mL) <strong>and</strong> is located at<br />
moderate altitude (up to 500 m) (Borelli 1979, Richard-Yegres &<br />
Yegres 1987). <strong>The</strong> l<strong>and</strong>scape is dominated by large cacti <strong>and</strong> other<br />
xerophytes. Stenocereus griseus is a columnar American cactus<br />
with a very strong, protective external epidermis that allows the<br />
accumulation of water in the shaft <strong>and</strong> enables tolerance of extreme<br />
drought. <strong>The</strong> species produces ovoidal, thorny fruits of about 5 cm<br />
diam, which are commonly eaten by the local population. It has<br />
therefore been suggested that patients with chromoblastomycosis<br />
acquire their infection by traumatic inoculation with cactus spines,<br />
<strong>similar</strong> to the supposed infection process of Madurella mycetomatis<br />
(Laveran) Brumpt in the arid climate of Africa (Ahmed et al. 2002).<br />
<strong>The</strong> frequent occurrence of 16 / 1 000 for chromoblastomycosis in<br />
areas endemic for Cladophialophora in Venezuela (Yegres et al.<br />
1985; Yegűez-Rodriguez et al. 1992) indicates a marked invasive<br />
potential for C. carrionii. Local goat-keepers are particularly at risk:<br />
in 1984, 14 of 18 patients investigated had these occupational<br />
characteristics (Yegres et al. 1985). Nevertheless, virulence of C.<br />
carrionii is low when inoculated into the footpads of mice (Yegres<br />
et al. 1998); also an environmental strain of C. carrionii failed to<br />
produce lesions in mice <strong>and</strong> in a volunteer (Richard-Yegres &<br />
Yegres 1987).<br />
We performed inoculation experiments with C. carrionii <strong>and</strong><br />
C. yegresii using freshly grown, healthy cacti in the greenhouse.<br />
<strong>The</strong> plants were followed over a 1-yr period; during all this time<br />
the control plants remained without lesions. Both Cladophialophora<br />
strains were able to produce infection when syringe-inoculated<br />
deep into young cactus tissue. Histopathology showed septate<br />
hyphae between host cells, <strong>and</strong> the shaft was maintained over<br />
prolonged periods without causing visible damage. This absence<br />
of appreciable destruction would categorise them as endophytes.<br />
Cactus tissue is rich in carbohydrates, vitamins <strong>and</strong> minerals (Vélez<br />
& Chávez 1980) which may promote endophyte growth.<br />
In contrast, suspensions applied superficially lead to growth<br />
on <strong>and</strong> in spines only. <strong>The</strong> absence of infection after superficial<br />
application indicates that the fungi are unable to invade healthy<br />
plant tissue from the surface <strong>and</strong> thus they cannot be characterised<br />
as obligatory phytopathogens.<br />
<strong>The</strong> two species differed in the degree of scarring after traumatic<br />
inoculation into mature plants: the clinical strain C. carrionii was<br />
consistently more virulent than C. yegresii that originated from<br />
the same host plant. Both species showed the same viability in<br />
re-isolated cultures. In nature, the fungi are likely to invade only<br />
when the integrity of the epidermis is broken, as happens e.g.<br />
by goat feeding or transmission by sap-sucking birds or piercing<br />
insects. <strong>The</strong>y also show the same transformation to meristematic<br />
morphology (González et al. 1990) when entering hard spine<br />
tissue. A possible trigger for this conversion is the dominance of<br />
lignin in the spines. Survival on <strong>and</strong> in spines is enhanced by their<br />
capturing of atmospheric water formed after nightly condensation.<br />
<strong>The</strong> fact that superficial application leads to colonisation around <strong>and</strong><br />
inside the spines suggests that the spines play a role in mechanic<br />
dispersion of the fungi.<br />
A possibly coincidental mechanism of dispersal might be<br />
traumatic inoculation into living tissue of humans or animals, where<br />
the same muriform cells are formed, defining the skin disease<br />
chromoblastomycosis. It may be questioned whether animal/<br />
human inoculation plays a role in the evolution of the fungus. ITS<br />
differences between the two species are observed in 23 positions,<br />
with a ratio of transitions : transversions of 2 : 1 (Table 3). Thus<br />
no saturation of mutations has taken place <strong>and</strong> the diversification<br />
can be regarded as an example of recent sympatric speciation.<br />
Cladophialophora carrionii is widely distributed, <strong>and</strong> shows a higher<br />
degree of diversity than C. yegresii. This would be suggestive for<br />
a longer evolutionary time span of existence <strong>and</strong> C. carrionii then<br />
should be regarded as ancestral to C. yegresii, with the latter<br />
showing a founder effect due to the absence of polymorphisms.<br />
However, such an order of event (a host jump from humans to<br />
cactus) is difficult to imagine. It is more likely that C. yegresii is<br />
the original cactus endophyte exhibiting extremotolerance via its<br />
muriform cells. T-rex data suggested a more direct connection of<br />
C. yegresii with African <strong>and</strong> Australian rather than Venezuelean<br />
strains of C. carrionii. We suppose that the low degree of observed<br />
variation in C. yegresii is not a founder effect, but rather a sampling<br />
effect, as living cacti have thus far not been studied outside the<br />
framework of our study on the patient with Cladophialophora<br />
chromoblastomycosis. <strong>The</strong> difference in virulence may be simply<br />
explained by C. carrionii, which lives as a saprobe on dead cactus<br />
debris for part of its life cycle, <strong>and</strong> is less adapted to an endophytic<br />
life style.<br />
Cladophialophora cf. carrionii is known to occur on lignified<br />
materials, such as wood chips of Eucalyptus crebra <strong>and</strong> wooden<br />
remains of Prosopis juliflora <strong>and</strong> Stenocereus griseus (Riddley<br />
1957, Yegres et al. 1985, Fernández-Zeppenfeldt et al. 1994). This<br />
does not exclude a certain degree of pathogenicity to humans, as<br />
also pathogens like Cryptococcus neoformans (Sanfelice) Vuill.<br />
are known to have an essential part of their life cycle in hollows<br />
of Eucalyptus trees. Cryptococcus neoformans produces diphenol<br />
oxidase to degrade lignin, an aromatic polymer in the cell wall of<br />
plants <strong>and</strong> a component of wood (Cabral 1999). Similar degradation<br />
pathways are present in Cladophialophora carrionii (Prenafeta-<br />
Boldú et al. 2006).<br />
<strong>The</strong> natural occurrence of C. carrionii <strong>and</strong> C. yegresii in<br />
association with xerophytes has been proven, but their environmental<br />
route of dispersal is still unknown. As transformation to meristematic<br />
cells takes place when the hyphae reach the spines <strong>and</strong> on dead<br />
spines, the muriform cell apparently is the extremotolerant phase<br />
of the species. <strong>The</strong> conidial anamorph can be found sporulating on<br />
rotten spines directly after rainfall (Richard-Yegres & Yegres 1987),<br />
but as the fungus has thus far never been isolated from outside air,<br />
it is still unclear how a new host plant is reached.<br />
<strong>The</strong> behaviour of C. carrionii on humans, provoking the very<br />
characteristic disease, chromoblastomycosis, of which the agents<br />
are limited to the ascomycete family Herpotrichiellaceae (de Hoog<br />
et al. 2000) is puzzling. In humans, the extremophilic muriform<br />
anamorph is expressed rather than hyphae, <strong>and</strong> thus humans do<br />
not seem a natural reservoir of the fungus. Nevertheless some<br />
acquired cellular immunity seems to be involved. Albornoz et al.<br />
(1982) demonstrated that a significant share of the local population<br />
of goat keepers (Yegres et al. 1985) is asymptomatically infected<br />
with C. carrionii; Iwatsu et al. (1982) detected cutaneous delayed<br />
hypersensitivity in rats experimentally-infected with agents of<br />
chromoblastomycosis. With murine experimental infection of<br />
the related fungus Fonsecaea pedrosoi, Ahrens et al. (1989)<br />
found enlargement <strong>and</strong> metastasis of lesions in athymic but not<br />
in normal mice, or in mice with defective macrophage function.<br />
Several authors (Kurita 1979, Nishimura & Miyaji 1981, Polak<br />
1984) observed a significant role of acquired cellular immunity in F.<br />
pedrosoi, while Cardona-Castro & Agudelo-Flórez (1999) obtained<br />
chronic infection in immunocompetent mice when inoculated<br />
intraperitoneally. Garcia Pires et al. (2002) noted an unbalance<br />
between protective Th1 <strong>and</strong> less efficient Th2 responses. <strong>The</strong><br />
possible host response leads to different clinical types, referred<br />
232
Cladophialophora carrionii complex<br />
to as tuberculoid <strong>and</strong> suppurative granuloma, respectively.<br />
<strong>The</strong> existence of genetic constitutional factors in susceptibility is<br />
underlined by a marked frequency of family relationships among<br />
symptomatic individuals (Yegűez-Rodriguez et al. 1992). <strong>The</strong><br />
disease is not observed in local animals such as goats, possibly<br />
due to their high body temperature (≈ 39 °C). Nevertheless, hyphal<br />
fragments artificially inoculated into goats led to transformation into<br />
muriform cells, but the lesions disappeared within 60 d (Martínez<br />
et al. 2005). Further animal experiments using strains identified<br />
according to new taxonomy will be necessary to answer questions<br />
on the role of the fungus on warm-blooded animals.<br />
ACKNOWLEDGEMENTS<br />
<strong>The</strong> authors are indebted to F. Yegres, N. Richard-Yegres <strong>and</strong> V. Maigualida Pérez-<br />
Blanco for providing a large set of strains for study <strong>and</strong> information on strain ecology,<br />
<strong>and</strong> to R.C. Summerbell for helpful suggestions on the manuscript. P. Abliz <strong>and</strong> K.<br />
Fukushima are acknowledged for making sequence data available for study. K.F.<br />
Luijsterburg is thanked for technical assistance. <strong>The</strong> study was supported in part<br />
by the Fondo Nacional de Investigaciones Cientificas y Tecnológicas (Fonacit),<br />
Venezuela.<br />
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fungal ribosomal RNA genes for phylogenetics. In: PCR Protocols: a guide to<br />
methods <strong>and</strong> applications (Innis MA, Gelf<strong>and</strong> DH, Sninsky JJ, White TJ, eds).<br />
Academic Press, San Diego, California: 315–322.<br />
Yegres F, Niel F, Gantier JC, Richard-Yegres N (1998). Murine humoral immune<br />
response against Cladophialophora carrionii <strong>and</strong> Fonsecaea pedrosoi infection.<br />
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Yegres F, Richard-Yegres N (2002). Cladophialophora carrionii: Aportes al<br />
conocimiento de la endemia en Venezuela durante el siglo XX. Revista de la<br />
Sociedad Venezuelana de Microbiologίa 2: 153–157.<br />
Yegres F, Richard-Yegres N, Medina-Ruiz E, González-Vivas R (1985).<br />
Cromomicosis por <strong>Cladosporium</strong> carrionii en criadores de caprinos del estado<br />
Falcon. Investigaciόn Clínica 26: 235–246.<br />
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pathogenicity of human <strong>and</strong> environmental isolates of <strong>Cladosporium</strong> carrionii in<br />
new born ddY mice. Mycopathologia 114: 71–76.<br />
Yegres F, Richard-Yegres N, Perez-Blanco M (1996). Cromomicosis. In: Temas<br />
de Micologia Médica. (Bastardo de Albornoz M, ed.). Caracas, Venezuela:<br />
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234
available online at www.studiesinmycology.org<br />
doi:10.3114/sim.2007.58.09<br />
Studies in Mycology 58: 235–245. 2007.<br />
Taxonomy, nomenclature <strong>and</strong> phylogeny of three cladosporium-like hyphomycetes,<br />
Sorocybe resinae, Seifertia azaleae <strong>and</strong> the Hormoconis anamorph of Amorphotheca<br />
resinae<br />
K.A. Seifert, S.J. Hughes, H. Boulay <strong>and</strong> G. Louis-Seize<br />
Biodiversity (Mycology & Botany), Eastern Cereal <strong>and</strong> Oilseed Research Centre, Agriculture & Agri-Food Canada, Ottawa, Ontario K1A 0C6 Canada<br />
*Correspondence: Keith A. Seifert, seifertk@agr.gc.ca<br />
Abstract: Using morphological characters, cultural characters, large subunit <strong>and</strong> internal transcribed spacer rDNA (ITS) sequences, <strong>and</strong> provisions of the International Code<br />
of Botanical Nomenclature, this paper attempts to resolve the taxonomic <strong>and</strong> nomenclatural confusion surrounding three species of cladosporium-like hyphomycetes. <strong>The</strong> type<br />
specimen of Hormodendrum resinae, the basis for the use of the epithet resinae for the creosote fungus {either as Hormoconis resinae or <strong>Cladosporium</strong> resinae) represents<br />
the mononematous synanamorph of the synnematous, resinicolous fungus Sorocybe resinae. <strong>The</strong> phylogenetic relationships of the creosote fungus, which is the anamorph of<br />
Amorphotheca resinae, are with the family Myxotrichaceae, whereas S. resinae is related to Capronia (Chaetothyriales, Herpotrichiellaceae). Our data support the segregation<br />
of Pycnostysanus azaleae, the cause of bud blast of rhododendrons, in the recently described anamorph <strong>genus</strong> Seifertia, distinct from Sorocybe; this species is related to the<br />
Dothideomycetes but its exact phylogenetic placement is uncertain. To formally stabilize the name of the anamorph of the creosote fungus, conservation of Hormodendrum<br />
resinae with a new holotype should be considered. <strong>The</strong> paraphyly of the family Myxotrichaceae with the Amorphothecaceae suggested by ITS sequences should be confirmed<br />
with additional genes.<br />
Key words: Amorphothecaceae, <strong>Cladosporium</strong> resinae, creosote fungus, Hormoconis resinae, jet fuel fungus, kerosene fungus, Myxotrichaceae, Pycnostysanus, resinicolous<br />
fungi.<br />
INTRODUCTION<br />
<strong>The</strong> ascomycete Amorphotheca resinae Parbery (1969) grows in<br />
hydrocarbon-rich substrates such as jet fuel, cosmetics <strong>and</strong> wood<br />
preserved with creosote or coal tar. This fungus is widely known<br />
by the anamorph name Hormoconis resinae (Lindau) Arx & G.A.<br />
de Vries or its obligate synonym <strong>Cladosporium</strong> resinae (Lindau)<br />
G.A. de Vries. It produces lightly pigmented, warty conidiophores,<br />
<strong>and</strong> branched, acropetally developing chains of lightly pigmented<br />
ameroconidia lacking conspicuous scars (Fig. 1B–E). This species<br />
is known colloquially as the “creosote fungus”, the “kerosene<br />
fungus” or the “jet fuel fungus”; to avoid confusion caused by the<br />
many heterotypic names with the epithet “resinae”, in this paper<br />
we generally will use the oldest of these informal names, “creosote<br />
fungus”, when referring to A. resinae or its anamorph. This fungus<br />
grows in jet fuel contaminated with small amounts of water, <strong>and</strong> the<br />
mycelium clogs fuel lines <strong>and</strong> corrodes metal parts. Consequently,<br />
fuel tanks in airports are monitored for this fungus by private<br />
companies using various physiological or biochemical tests.<br />
Sorocybe resinae (Fr.) Fr. produces dark black colonies on conifer<br />
resin, comprising dark synnemata <strong>and</strong> an effuse mononematous<br />
synanamorph, both with cladosporium-like conidiogenous cells<br />
<strong>and</strong> conidia. Unlike the anamorph of the creosote fungus, the<br />
conidia of Sorocybe resinae are dark brown <strong>and</strong> the lateral walls<br />
are conspicuously thicker than the poles (Fig. 2D–G). Colonies<br />
with only the mononematous anamorph sometimes occur, <strong>and</strong><br />
the mononematous anamorph can be sparse on colonies bearing<br />
synnemata. However, the conidia of the mononematous anamorph<br />
have identical pigmentation <strong>and</strong> lateral wall thickening to that of<br />
the synnematous anamorph. <strong>The</strong> mononematous anamorph rarely<br />
has been referred to by its own binomial name although, as we will<br />
show, there is a species epithet available. For the same reasons<br />
given above for Amorphotheca Parbery, generally we will refer to<br />
Sorocybe resinae herein as “the resin fungus”.<br />
Despite the micromorphological differences noted above, there<br />
is disagreement about whether the creosote fungus is conspecific<br />
with the mononematous synanamorph of the resin fungus (Parberry<br />
1969). <strong>The</strong> name for the anamorph of the creosote fungus is based<br />
on Hormodendrum resinae Lindau (1906). Christensen et al.<br />
(1942) presented a study of a cladosporium-like fungus commonly<br />
isolated from wood impregnated with creosote <strong>and</strong> coal tar <strong>and</strong><br />
applied Lindau’s name without examining its type. A later ecological<br />
study by Marsden (1954) employed the same name for the same<br />
fungus. An extra dimension was added to the confusion when de<br />
Vries (1952, using the name <strong>Cladosporium</strong> avellaneum G.A. de<br />
Vries) described four formae for the creosote fungus (differing in<br />
the colours of their conidia, the production of setae, or the total<br />
absence of conidia), each based on single conidium isolates made<br />
from one parent culture. De Vries (1955) <strong>and</strong> Parberry (1969)<br />
examined the holotype of Hormodendrum resinae <strong>and</strong> concluded<br />
that it represented the creosote fungus. Hughes (1958), prior to<br />
the description of Amorphotheca or Hormoconis Arx & G.A. de<br />
Vries, examined the same specimen <strong>and</strong> considered it to be the<br />
mononematous synanamorph of the resin fungus. If Hughes (1958)<br />
is correct, then neither the species Hormodendrum resinae, nor<br />
the <strong>genus</strong> that it typifies, Hormoconis, can represent the creosote<br />
fungus, as intended by Parberry (1969) or von Arx <strong>and</strong> de Vries (in<br />
von Arx 1973).<br />
In this paper, we present micromorphological, cultural <strong>and</strong><br />
molecular evidence that the resin fungus is a different species from<br />
the creosote fungus. Combined with re-examination of the holotype<br />
of Hormodendrum resinae, this information is used to provide a<br />
revised taxonomy <strong>and</strong> nomenclature for these two species. A third<br />
cladosporium-like fungus, Seifertia azaleae, is also considered in<br />
our discussion of generic concepts.<br />
Historical review<br />
<strong>The</strong> history of the fungus now known as Sorocybe resinae began<br />
with Fries (1815), who described Racodium resinae Fr. as follows:<br />
“310. Racodium resinae, expansum molliusculum dense contextum nigrum, filis<br />
inaequalibus.<br />
In resina Pini Abietis in silvis Suecia passim.<br />
Habitu et loco natali distinctum. Fila divaricato-ramosa; alia rigidula apice<br />
capituli sera, sub miscrosc. Coremio Link similia, Demat. villosum Schleich. huic<br />
simile; sed sub microsc. fila maxime differunt.”<br />
235
Seifert et al.<br />
Fig. 1. Amorphotheca resinae, colony characters <strong>and</strong> anamorph micromorphology. A. 10-d-old colony on PDA. B, D–E. Micromorphology of conidiophores, showing acropetal<br />
conidial chains, ramoconidia, <strong>and</strong> conidia. C. Conidia. DAOM 170427; for C, E see scale bar in D.<br />
<strong>The</strong> comparison with Coremium Link indicates the probability of a<br />
synnematous fungus, <strong>and</strong> an authentic specimen of Fries’ fungus,<br />
which as the only known authentic material we interpret as the<br />
holotype, is preserved in Link’s herbarium (see below). It represents<br />
the synnematous form of the resin fungus 1 .<br />
1 Persoon (1822) described a form of R. resinae “β piceum”. Hughes (1968) examined<br />
the holotype of this form, <strong>and</strong> it represents the mycelium of the ascomycete<br />
Strigopodia resinae (Sacc. & Bres.) S.J. Hughes. This taxon is thus not relevant to<br />
the three species that are the focus of this paper.<br />
Fries (1832) later transferred his species to Sporocybe<br />
Fr. (1825), a <strong>genus</strong> then used for relatively conspicuous dark<br />
hyphomycetes with dry spores (Mason & Ellis 1953). <strong>The</strong> 1832<br />
description explicitly stated... “capitulo rotundato inaequali, sporidiis<br />
seriatis, stipite aequali simplici.” <strong>The</strong> use of “capitulo” <strong>and</strong> “stipite”<br />
imply what would now be recognised as a synnematous fungus.<br />
Fries (1832) further characterised the habit of the fungus as “habitu<br />
stipitum Calicii,” a further comparison to a group of black, stipitate<br />
lichenized fungi classified in Calicium Pers., which under a h<strong>and</strong><br />
lens look <strong>similar</strong> to a dark synnematous fungus.<br />
Fries (1849) next described the <strong>genus</strong> Sorocybe Fr. for this<br />
fungus, as follows:<br />
236
Hormoconis resinae <strong>and</strong> morphologically <strong>similar</strong> taxa<br />
Fig. 2. Sorocybe resinae, synnematous form. A. Colony on bark of living, st<strong>and</strong>ing conifer. B. Synnemata. C. Four-month-old colony on DG18. D–G. Acropetally developing<br />
chains of conidia. Note that the lateral walls are conspicuously thickened; compare with Fig. 3. A, C. DAOM 239134. B, D–G. DAOM 11381.<br />
Sorocybe Fr.<br />
Habitus prioris. sed mycelium floccosum densum, stroma corneo-carbonaceum,<br />
sporis moniliformi-concatenatis basi excipulum incompletum praebens.<br />
1. S. resinae. Fr. 1–4. at raro fructif. Klotzsch. exs. C. 2.<br />
Because this description explicitly referred to the Systema, Fries<br />
presumably was segregating the fungus, originally described<br />
as Racodium resinae, into a new monotypic <strong>genus</strong> (McNeill et<br />
al. 2007; Art. 33.3) <strong>and</strong> this interpretation of R. resinae as the<br />
basionym generally has been followed in subsequent treatments<br />
of Sorocybe resinae.<br />
As noted in Table 1, Fries’ Racodium resinae was placed in<br />
several other hyphomycete genera by eighteenth century authors.<br />
<strong>The</strong>se diversions need not be reviewed in detail here because the<br />
modern status of these other genera, <strong>and</strong> their lack of <strong>similar</strong>ity with<br />
Sorocybe, is clear.<br />
Bonorden (1851) described Hormodendrum Bonord., with four<br />
species originally placed in Penicillium Link by Corda (1839); H.<br />
olivaceum (Corda) Bonord. (≡ Penicillium olivaceum Corda 1839)<br />
was designated as lectotype by Clements & Shear (1931). This<br />
<strong>genus</strong> was frequently, but incorrectly, spelled “Hormodendron”.<br />
Bonorden’s descriptions <strong>and</strong> illustrations are of variable quality by<br />
modern st<strong>and</strong>ards, <strong>and</strong> his herbarium is unknown (Stafleu et al.<br />
1995). Consequently the actual identities of the species Bonorden<br />
placed in Hormodendrum are unknown <strong>and</strong> Corda’s <strong>Cladosporium</strong><br />
olivaceum (Corda) Bonord. was dismissed in Penicillium monographs<br />
because the drawing shows branched conidial chains<br />
(Thom 1930), although the specimen has apparently not been<br />
re-examined. <strong>The</strong> generic name was used as a segregate for<br />
<strong>Cladosporium</strong> Link by some authors (e.g. Kendrick 1961), in<br />
particular for species with ameroconidia (de Vries 1952). Although it<br />
sometimes has been considered a synonym of <strong>Cladosporium</strong>, it will<br />
remain a nomen dubium until the type species is properly typified.<br />
Unaware of the resinicolous fungus described by Fries, Lindau<br />
described two species growing on conifer resin, Pycnostysanus<br />
resinae Lindau (1904), the type of this anamorph generic name,<br />
<strong>and</strong> Hormodendrum resinae Lindau (1906). <strong>The</strong> former was<br />
clearly illustrated <strong>and</strong> described as a synnematous species. <strong>The</strong><br />
protologue of the latter concludes with, “Mit Pycnostysanus resinae<br />
hat die Art nichts zu tun.” Clearly, Lindau observed no synnemata<br />
on the specimen of the mononematous fungus <strong>and</strong> he believed<br />
it was a different fungus, rather than what would now be called a<br />
synanamorph of the synnematous fungus that he had described<br />
previously. Lindau (1910) reproduced the 1904 illustration of<br />
Pycnostysanus resinae as Stysanus resinae (Fr.) Sacc. (1906),<br />
thus accepting its identity with the species originally described<br />
as Racodium resinae Fr. Lindau (1910) made no mention of<br />
Hormodendrum resinae, indicating he still made no association<br />
between the synnematous <strong>and</strong> mononematous fungi on resin.<br />
De Vries (1952) described a new species, <strong>Cladosporium</strong><br />
avellaneum G.A. de Vries, isolated from cosmetics. Later, he noted<br />
the <strong>similar</strong>ities between his C. avellaneum <strong>and</strong> the creosote fungus,<br />
<strong>and</strong> suggested that they were the same species (de Vries 1955),<br />
replacing the name of one of his previously described formae,<br />
i.e. viride, with the forma name resinae. He examined Lindau’s<br />
type of Hormodendrum resinae <strong>and</strong> decided that it provided an<br />
earlier epithet for C. avellaneum. He transferred the species into<br />
<strong>Cladosporium</strong> as C. resinae (Lindau) G.A. de Vries, <strong>and</strong> this name<br />
was widely used for the creosote fungus until 1973. This binomial<br />
is still commonly employed in non-taxonomic literature, especially<br />
commercial publications dealing with the creosote fungus.<br />
www.studiesinmycology.org<br />
237
Seifert et al.<br />
Table 1. Nomenclature <strong>and</strong> synonymies for the creosote fungus <strong>and</strong> the resin fungus, showing the use of the same basionym for the two fungi. <strong>The</strong> “false” names <strong>and</strong><br />
synonymies for the anamorph of the resin fungus are indicated by blue text. <strong>The</strong> second nomenclatural solution described in the text would have the effect of switching the<br />
blue text to black for the creosote fungus, <strong>and</strong> to simultaneously switch the equivalent black text to blue for the mononematous synanamorph of the resin fungus. Holotypes<br />
we have examined, <strong>and</strong> the herbarium where they are deposited, are marked with exclamation points, <strong>and</strong> details of these specimens are noted in Materials <strong>and</strong> Methods.<br />
Creosote fungus<br />
Teleomorph: Amorphotheca resinae Parberry, Australian J. Bot. 17: 340. 1969.<br />
Anamorph<br />
Hormodendrum resinae Lindau, in Rabenh. Krypt.-Fl., 2, 1 (Pilze) 8: 699. 1906 (B!).<br />
≡ <strong>Cladosporium</strong> resinae (Lindau) G.A. de Vries, Antonie van Leeuwenhoek 21: 167. 1955.<br />
≡ Hormoconis resinae (Lindau) von Arx & G.A. de Vries, in von Arx, Verh. K. Ned. Akad. Wet., Afd. Natuurk. 61: 62. 1973.<br />
= <strong>Cladosporium</strong> avellaneum G.A. de Vries, Contribution to the knowledge of the <strong>genus</strong> <strong>Cladosporium</strong>, Uitg. Druk. Holl<strong>and</strong>ia, p. 56, 1952.<br />
Resin fungus<br />
Mononematous synanamorph:<br />
Hormodendrum resinae Lindau, in Rabenh. Krypt.-Fl., 2, 1 (Pilze) 8: 699. 1906 (B!)<br />
≡ <strong>Cladosporium</strong> resinae (Lindau) G.A. de Vries, Antonie van Leeuwenhoek 21: 167. 1955.<br />
≡ Hormoconis resinae (Lindau) von Arx & G.A. de Vries, in von Arx, Verh. K. Ned. Akad. Wet., Afd. Natuurk. 61: 62. 1973.<br />
Synnematous anamorph:<br />
Sorocybe resinae (Fr.) Fr., Summa Veg. Scan. 2: 468. 1849.<br />
≡ Racodium resinae Fr., Obs. Mycol. 1: 216. 1815 (basionym) (B!).<br />
≡ Sporocybe resinae (Fr.) Fr., Syst. Mycol. 3: 341. 1832.<br />
≡ Dendryphion resinae (Fr.) Corda, Icon. Fung. 6: 11. 1854.<br />
≡ Stysanopsis resinae (Fr.) Ferr., Flora Ital. Crypt., 1 (Fungi, Hyphales), p. 187. 1910.<br />
? = Dematium nigrum Link, Mag. ges. naturf. Fr. 3: 21. 1809 (B!).<br />
≡ Sporotrichum nigrum (Link) Link, Mag. Ges. naturf. Fr. Berlin 7: 35. 1815.<br />
= Pycnostysanus resinae Lindau, Verh. Bot. Ver. Br<strong>and</strong>enb. 45 : 160. 1904 (B!).<br />
≡ Stysanus resinae (Lindau) Sacc., Syll. Fung. 18: 651. 1906.<br />
In his study of type collections of classical hyphomycetes,<br />
Hughes (1958) included Pycnostysanus resinae Lindau <strong>and</strong><br />
Hormodendrum resinae Lindau as facultative synonyms of<br />
Sorocybe resinae (Fr.) Fr., with Racodium resinae Fr. <strong>and</strong> several<br />
other nomenclatural variants as obligate synonyms (Table 1). <strong>The</strong><br />
synnematous Pycnostysanus resinae was cited as “Pycnostysanus<br />
state [i.e. synanamorph] of Sorocybe resinae”. Hormodendrum<br />
resinae thus remained to represent the mononematous<br />
synanamorph of what was interpreted as a single species.<br />
Parberry (1969) described a cleistothecial ascomycete,<br />
Amorphotheca resinae, for the teleomorph of the creosote fungus.<br />
He also examined the holotype of Hormodendrum resinae <strong>and</strong><br />
agreed with the conclusions of de Vries (1955). He used the<br />
epithet resinae for the teleomorph to correspond with that of the<br />
anamorph. He discounted the possibility that the synnematous<br />
Sorocybe resinae could be the same fungus as Hormodendrum<br />
resinae because synnemata never developed in his cultures of the<br />
creosote fungus.<br />
Von Arx <strong>and</strong> de Vries (in von Arx 1973) described the <strong>genus</strong><br />
Hormoconis, typified by Hormodendrum resinae, with the new<br />
combination Hormoconis resinae (Lindau) Arx & G.A. de Vries.<br />
<strong>The</strong>ir intention was to erect an anamorph <strong>genus</strong> for the anamorph<br />
of the creosote fungus, which they suggested was improperly<br />
classified in <strong>Cladosporium</strong> because it lacked darkened, thickened<br />
secession scars on the conidia.<br />
A third cladosporium-like fungus is relevant to this story.<br />
Seifertia azaleae (Peck) Partridge & Morgan-Jones [until recently<br />
known as Pycnostysanus azaleae (Peck) E.W. Mason] is a<br />
cosmopolitan fungus causing bud blast <strong>and</strong> twig blight of azaleas<br />
<strong>and</strong> rhododendrons. This species is morphologically <strong>similar</strong> to<br />
Sorocybe resinae, but the conidia are paler <strong>and</strong> lack laterally<br />
thickened walls. Sorocybe <strong>and</strong> Pycnostysanus have often been<br />
considered taxonomic synonyms (Ellis 1976, Carmichael et al.<br />
1980); as shown above, both are based on the synnematous form<br />
of the resin fungus. Partridge <strong>and</strong> Morgan-Jones (2002) argued that<br />
Sorocybe resinae <strong>and</strong> “Pycnostysanus azaleae” are not congeneric,<br />
<strong>and</strong> described the new <strong>genus</strong> Seifertia Part. & Morgan-Jones for<br />
the Rhododendron fungus. <strong>The</strong>y observed that the connection<br />
between conidia in Seifertia azaleae is much narrower than in<br />
Sorocybe resinae, <strong>and</strong> that minute denticles are visible on the<br />
conidiogenous cells of the former fungus. <strong>The</strong> broader connections<br />
between conidia of Sorocybe resinae result in broadly protuberant<br />
conidiogenous loci on the conidiogenous cells, <strong>and</strong> more truncate<br />
detached conidia.<br />
MATERIALS AND METHODS<br />
Herbarium material <strong>and</strong> fungal strains<br />
Full details of herbarium material examined are listed below.<br />
Cultures <strong>and</strong> dried herbarium specimens were studied in 90 %<br />
lactic acid without stains; preparations of some exsiccate <strong>and</strong> types<br />
were mounted in glycerin jelly. Cultures were grown on potatodextrose<br />
agar (PDA, Difco), oatmeal agar (OA, Samson et al.<br />
2004), Blakeslee’s malt extract agar (MEA, Samson et al. 2004) <strong>and</strong><br />
dichloran-18 % glycerol agar (DG-18, Samson et al. 2004). Colony<br />
characters were taken from cultures grown at 25 °C in darkness.<br />
Cultures are maintained in the Canadian Collection of Fungal<br />
Cultures (DAOM), Agriculture & Agri-Food Canada, Ottawa.<br />
238
Hormoconis resinae <strong>and</strong> morphologically <strong>similar</strong> taxa<br />
Exsiccati <strong>and</strong> types<br />
Dematium nigrum [scr. Link]. E. Hbr. Link (23) = Sporocybe<br />
resinae III. 341 [scr. ? ] (herb. Link, B).<br />
Hormodendrum resinae Lindau, n. sp. Fl. v. Hamburg 206, auf<br />
Harz an Picea excelsa, Sachsenwald, leg. O. Jaap, 29-4-1906.<br />
[scr. Lindau]. (DAOM 41888, slide prepared from the holotype<br />
preserved in B.)<br />
Pycnostysanus resinae Lindau nov. gen. et nov. spec., Kabát et<br />
Bubák: Fungi imperfecti exsiccati no. 99. Auf erhärteten Fichtenharz<br />
an Brockenweg, am Dreieckigen Pfahl in Harz, Deutschl<strong>and</strong>, leg.<br />
G. Lindau, 13.VIII. 1903 (holotype, B).<br />
Racodium resinae Fries. E. Hbr. Link, Fries legi, Smol. [scr.<br />
Fries]. (DAOM 41890, slide prepared from herb. Link, B). This<br />
is the presumed holotype of R. resinae, the basionym for the<br />
resin fungus, Sorocybe resinae. <strong>The</strong> specimen includes dark,<br />
decapitated synnemata, brown conidia with laterally thickened<br />
walls, <strong>and</strong> acropetal conidial chains, allowing it to be recognised<br />
as the fungus we now know as S. resinae. Fries perhaps sent this<br />
fungus to Link to see if it could be differentiated from Coremium<br />
Link. <strong>The</strong> minimal details, that the fungus was collected by Fries,<br />
presumably in Smål<strong>and</strong> (a province of Sweden), match the details<br />
in the protologue of this species.<br />
Sorocybe resinae. “Fungi Rhenani Fasc. II, 1863, L. Fuckel, no.<br />
129, ad Abietis resinam, raro Hieme, in sylva Hostrichiensi” (as<br />
Myxotrichum resinae Fr., DAOM 55543 ex FH). “Flora Suecica,<br />
2956, Ad resinam piceae, Smål<strong>and</strong>: Femsjö, Prostgaidsshogen,<br />
6 Aug. 1929, leg. J.A. Nannfeldt, s.n.” (as Stysanus resinae (Fr.)<br />
Sacc., DAOM 41891 ex UPS). “Flora Suecica, 4709, Ad resinam<br />
abietinum, Uppl<strong>and</strong>: Bondkysko sin Valsätra, 9 May 1932, leg. J.A.<br />
Nannfeldt” (as Hormodendrum resinae Lindau, DAOM 41889 ex<br />
UPS). “[ on wood scr. Berkeley] J.E. Vize, Hereford 1877” (as Torula<br />
pinophila Fr., DAOM 113425 ex K). “Sydow, Mycotheca germanica,<br />
350. Auf Fichtenharz… am Brockenweg 30.9.1904, leg. P. Sydow”<br />
(DAOM 41893).<br />
Other material examined<br />
Sorocybe resinae. Canada, British Columbia: Burnaby, Central Park, on resin of<br />
Tsuga heterophylla, leg. S. & L. Hughes, 17 Aug. 2000 (DAOM 228572a, 228573a);<br />
Cameron Lake, Cathedral Grove, on Pseudotsuga menziesii, leg. isol. S.J. Hughes,<br />
21 Aug. 1957 (DAOM 56088a). Ladysmith, Ivy Green Park, on resinous exudates,<br />
leg. R.J. B<strong>and</strong>oni no. BC-978, 18 Apr. 1960, det. S.J. Hughes (DAOM 70462).<br />
North Vancouver, Lynn Valley Conservation Area, leg. det. S.J. Hughes, 1 Jul. 1975<br />
(DAOM 139385); North Vancouver, Lynn Valley Conservation Area, on bark of living<br />
conifer (probably Pseudotsuga menziesii), leg. isol. K.A. Seifert no. 1574, 26 May<br />
2002 (single conidium isolate, culture <strong>and</strong> specimen DAOM 239134; ITS GenBank<br />
EU030275, LSU GenBank EU030277); Terrace, near Kalum, on Tsuga heterophylla,<br />
leg. W.G. Ziller no. V-6549, 10 July 1950, det. S.J. Hughes (DAOM 59657); Queen<br />
Charlotte Isl<strong>and</strong>s, east coast of Moresby Isl<strong>and</strong>, north side of Gray Bay, 53°08’ N,<br />
131°47’ W, on Picea sitchensis, leg. I. Brodo, M.J. Schepanek, W.B. Schofield,<br />
28 Sep. 1973, det. S.J. Hughes (DAOM 144757); Queen Charlotte Isl<strong>and</strong>s,<br />
Graham Isl<strong>and</strong>, Tow Hill area, on resin of Picea sitchensis, leg. S.A. Redhead no.<br />
4440, 20 Sep. 1982, det. G.P. White (DAOM 184025); Revelstoke, Wigwam, on<br />
Tsuga heterophylla, leg. W. Ziller V-6567 det. S.J. Hughes, 6 Jun. 1950 (DAOM<br />
59710); Vancouver Isl<strong>and</strong>, Cathedral Grove, Cameron Lake, on Pseudotsuga<br />
menziesii, leg. det. S.J. Hughes, 21 Aug. 1957 (DAOM 56088a); Vancouver Isl<strong>and</strong>,<br />
Caycuse, on resin of Pseudotsuga menziesii, leg. det. S.J. Hughes, 17 Jul. 1972<br />
(DAOM 139355); Vancouver Isl<strong>and</strong>, Lake Cowichan, Honeymoon Bay, on resin<br />
of Pseudotsuga menziesii, leg. J Ginns, det. S.J. Hughes, 29 Oct. 1971 (DAOM<br />
134968); Vancouver Isl<strong>and</strong>, Lake Cowichan, Mesachie Lake Forest Experimental<br />
Station, leg. det. S.J. Hughes, 5 Jul. 1972 (DAOM 139277a, DAOM 139278) <strong>and</strong><br />
6 Jul. 1072 (DAOM 139281). Czechoslovakia, Ještěd near Liberec, leg. det. S.J.<br />
Hughes, on resin of Larix europaea, 10 May 1955 (DAOM 51723). United States,<br />
Oregon: Andrews’ Experimental Forest, Forest Service Rd. no 1553, on resin of<br />
Tsuga heterophylla, leg. det. S.J. Hughes, 10 May 1969 (DAOM 134565); Andrews’<br />
Experimental Forest, Blue River, on resin of conifer, cut wood, leg. det. K.A. Seifert<br />
no. 69, 10 Jul. 1981 (DAOM 228203); Oregon, del Norte Co., J. Smith’s State Park,<br />
on Tsuga heterophylla, leg. det. S.J. Hughes, 11 May 1069 (DAOM 134614); Devil’s<br />
Elbow State Park, Cape Perpetus, on Picea sitchensis, leg. det. S.J. Hughes, 6 May<br />
www.studiesinmycology.org<br />
1969 (DAOM 134615); Linn Co., near Cascadia, on Pseudotsuga menziesii, leg. R.<br />
Fogel, det. S.J. Hughes, 14 May 1969 (DAOM 127885); U.S. Forest Service Rd. no.<br />
126, North fork Cape Creek, on resin of Abies gr<strong>and</strong>is, leg. det. S.J. Hughes, 7 May<br />
1969 (DAOM 134852,134563); Willamette National Forest, McKenzie Bridge Camp<br />
Grounds, leg. det. S.J. Hughes, 10 May 1969 (DAOM 134564). Washington: Kittitas<br />
Co., Wanatchee National Forest, Rocky Run, on Abies nobilis, leg. Field Mycology<br />
Class 1955, 22 Jul. 1955, det. S.J. Hughes, (mononematous synanamorph only,<br />
DAOM 118934 ex WSP 45210, as Helminthosporium sp.); Jefferson Co., Olympic<br />
National Forest, 10 mi Camp, Sec. 17, T26N, R3W, on Pseudotsuga mucronata,<br />
leg. Field Mycology Class, 22 Jul. 1955 (DAOM 113801 ex WSP 45212, as<br />
Helminthosporium); Grays Harbor Co., Twin Harbors Beach State Park, resin of<br />
Picea sitchensis, leg. W.B. & V.G. Cooke, 24 Jul. 1951, det. S.J. Hughes (DAOM<br />
118970 ex WSP 28432).<br />
Amorphotheca resinae. Isolated from jet fuel by P. Emonds (culture, DAOM<br />
170427 = ATCC 22711, ITS GenBank EU030278, LSU GenBank EU030280).<br />
Canada, British Columbia, source unknown, isol. “Mrs. Volkoff”, Jul. 1969 (culture,<br />
DAOM 194228, ITS GenBank EU030279).<br />
Seifertia azaleae. All on flower buds of Rhododendron spp. Canada, British<br />
Columbia: Burnaby, Central Park, leg. S.J. Hughes, 17 Aug. 2000 (DAOM 228571);<br />
Vancouver, Stanley Park, leg. K.A. Seifert no. 1571, 11 May 2002 (culture <strong>and</strong><br />
specimen, DAOM 239136, LSU GenBank EU030276). Irel<strong>and</strong>, Munter, Kerry,<br />
near Glenbeigh (ca. N 52° 03’ W 9° 54’), leg. K.A. Seifert no. 3197, 26 Sep. 2006<br />
(culture <strong>and</strong> specimen, DAOM 239135, ITS GenBank EU030273). Netherl<strong>and</strong>s,<br />
Gelderl<strong>and</strong>, Kröller-Müller Museum, leg. K.A. Seifert no. 1235, 12 May 2000 (DAOM<br />
227136). United Kingdom, Wales, Hafod Estate (ca. N 52° 22’ W 3° 51’), leg. K.A.<br />
Seifert no. 3198, 1 Oct. 2006 (culture <strong>and</strong> specimen, DAOM 239137, ITS GenBank<br />
EU030274).<br />
DNA extraction, amplification <strong>and</strong> sequencing<br />
DNA was isolated using a FastDNA Kit <strong>and</strong> the FastPrep<br />
FP120 (BIO 101 Inc.) or an UltraClean Microbial DNA Isolation<br />
Kit (Mo Bio Laboratories, Inc., Solana Beach, CA, U.S.A.) using<br />
mycelium removed from agar cultures. PCR <strong>and</strong> cycle sequencing<br />
reactions were performed on a Techne Genius thermocycler<br />
(Techne Cambridge Ltd.). PCR reactions were performed using<br />
Ready-To-Go Beads (Amersham Canada Ltd.) in 25 µL volumes,<br />
each containing 20–100 ng of genomic DNA, 2.5 units pure Taq<br />
DNA Polymerase, 10 mM Tris-HCl (pH 9.0), 50 mM KCl, 1.5 mM<br />
MgCl 2<br />
, 200 µM of each dNTP, 0.2 µL of each primer (50 µM), <strong>and</strong><br />
stabilizers including bovine serum albumin. <strong>The</strong> reaction profile<br />
included an initial denaturation for 4 min at 94 °C, followed by 30<br />
cycles of 1.5 min denaturation at 95 °C, 1 min annealing at 56 °C,<br />
2 min extension at 72 °C, with a final extension of 10 min at 72 °C.<br />
Amplicons were purified by ethanol/sodium acetate precipitation<br />
<strong>and</strong> resuspended as recommended for processing on an ABI<br />
PRISM 3100 DNA Analyzer or an ABI 373 Stretch DNA Sequencer<br />
(Applied Biosystems, Foster, CA). Amplification products were<br />
sequenced using the BigDye v. 2.0 Terminator Cycle Sequencing<br />
Ready Reaction Kit (ABI Prism/Applied Biosystems) following the<br />
manufacturer’s directions. An approximately 1 000 bp portion of the<br />
large subunit (LSU) ribosomal DNA was amplified <strong>and</strong> sequenced<br />
using primers LR0R <strong>and</strong> LR6, <strong>and</strong> cycle-sequenced using primers<br />
LR0R, LR3R, LR16 <strong>and</strong> LR6 (Vilgalys & Hester 1990, Rehner &<br />
Samuels 1995; www.biology.duke.edu/fungi/mycolab/primers.<br />
htm). <strong>The</strong> complete ITS <strong>and</strong> 5.8S rRNA genes were amplified<br />
<strong>and</strong> sequenced using the primers ITS5 <strong>and</strong> ITS4, with ITS2 <strong>and</strong><br />
ITS3 primers used for cycle sequencing when necessary (White<br />
et al. 1990). Some sequences were derived from single PCR<br />
amplifications of the ITS5–LR6 region.<br />
Data matrices were subjected to parsimony analysis using<br />
heuristic searches in PAUP* v. 4.0b10 (Swofford 2002) with<br />
simple stepwise addition of taxa, <strong>and</strong> tree bisection-reconnection<br />
(TBR) branch swapping. Uninformative characters were removed<br />
for all analyses. Strict consensus trees were calculated, <strong>and</strong> the<br />
robustness of the phylogenies was tested using full bootstrap<br />
analyses (1 000 replications). For all analyses, GenBank accession<br />
numbers are given on the tree figures, <strong>and</strong> the sequences generated<br />
in this study are indicated in bold.<br />
239
Seifert et al.<br />
Fig. 3. Hormodendrum resinae, A–B. Conidiophores <strong>and</strong> acropetally developing chains of conidia. C. Conidia. Note that the lateral walls are conspicuously thickened compared<br />
to the walls at the poles. From a slide (DAOM 41888) prepared from the holotype (B).<br />
Byssoascus striatisporus U17912<br />
99<br />
96<br />
10 changes<br />
88 88<br />
99 “Mycosphaerella mycopappi” U43480<br />
* Seifertia azaleae DAOM 239136 EU030276<br />
Pleomassaria siparia AY004341<br />
*<br />
Pleospora herbarum AF382386<br />
* Cochliobolus heliconiae AF163978<br />
Setosphaeria monoceras AY016368<br />
Trematosphaeria heterospora AY016369<br />
Iodosphaeria aquatica AF452044<br />
Westerdykella cylindrica AY004343<br />
Letendraea helminthicola AY016362<br />
Lojkania enalia AY016363<br />
Botryosphaeria ribis AY004336<br />
Capnodium citri AY004337<br />
Discosphaerina fagi AY016359<br />
Ceramothyrium carniolicum AY004339<br />
Fonsecaea pedrosoi AF356666<br />
80 Phialophora americana AF050279<br />
99 79 Capronia mansonii AY004338<br />
Glyphium elatum AF346420<br />
97 Terfezia gigantea AF499449<br />
Sorocybe resinae DAOM 239134 EU030277<br />
Ascosphaera apis AY004344<br />
Saccharomyces cerevisiae J01355<br />
Phaeotrichum benjaminii AY004340<br />
Calicium viride AF356670<br />
Pertusaria mammosa AY212831<br />
73 Pseudocyphellaria perpetua AF401954<br />
Xylographa vitiligo AY212849<br />
Neofabraea alba AY064705<br />
Melanelia exasperatula AJ421436<br />
* Lophodermium pinastri AY004334<br />
99 Cudonia lutea AF433138<br />
Spathularia flavida AF433141<br />
Ascocoryne cylichnium AF353580<br />
Bisporella citrina AF335454<br />
Hormoconis resinae DAOM 170427 EU030280<br />
Rhytisma acerinum AF356696<br />
Pleuroascus nicholsonii AF096196<br />
Golovinomyces cichoracearum AB022360<br />
Fig. 4. Parsimony analysis of large subunit sequences, demonstrating the phylogenetic positions of Amorphotheca resinae, Sorocybe resinae <strong>and</strong> Seifertia azaleae (all shown<br />
in bold) in the Ascomycota. One of 12 equally parsimonious trees (1 888 steps, CI = 0.390, RI = 0.554, RC = 0.216, HI = 0.610) with Golovinomyces cichoracearum as the outgroup.<br />
Bootstrap values above 70 % are shown at the relevant nodes, with an asterisk representing 100 % bootstrap support; branches with thick lines occurred in all equally<br />
parsimonious trees.<br />
Leotiomycetes Dothideomycetes<br />
240
Hormoconis resinae <strong>and</strong> morphologically <strong>similar</strong> taxa<br />
<strong>The</strong> large subunit matrix was assembled from the closest<br />
BLAST matches using our sequences for the three fungi of<br />
interest, S. resinae, A. resinae <strong>and</strong> S. azaleae; Golovinomyces<br />
cichoracearum was added as an out-group to root the tree.<br />
Although these sequences were put into a single matrix, there<br />
is no implication that this data set represents the diversity of the<br />
Ascomycota. <strong>The</strong> alignment was calculated using MAFFT (Katoh et<br />
al. 2002) <strong>and</strong> adjusted using Se-Al (Sequence Alignment Program<br />
v. 1.d1; http://evolve.zoo.ox.ac.uk/software/Se-Al/main.html) to<br />
maximise homology.<br />
<strong>The</strong> internal transcribed spacers alignment including Sorocybe<br />
resinae was derived from an alignment of Capronia <strong>and</strong> related<br />
anamorphs used by Davey & Currah (2007), originally produced<br />
using MAFFT. This data set was modified considerably using Se-<br />
Al to maximise homology, but still included several areas where<br />
the homology of aligned sequences was difficult to evaluate. ITS<br />
sequences of Amorphotheca resinae were used to retrieve closely<br />
related sequences using a BLAST search of GenBank, <strong>and</strong> these<br />
relevant sequences were added to an alignment of Oidiodendron<br />
Robak sequences from the study of Hambleton et al. (1998), <strong>and</strong><br />
then adjusted using Se-Al.<br />
We attempted direct PCR from two specimens containing only<br />
the putative mononematous synanamorph of Sorocybe resinae<br />
(DAOM 228772a, 228573a), to allow comparison of sequences<br />
obtained from cultures of the synnematous synanamorph.<br />
<strong>The</strong>se attempts, using the same methods outlined above, were<br />
unsuccessful.<br />
RESULTS<br />
Cultural characters <strong>and</strong> micromorphology<br />
Most micromorphological characters of the resin fungus Sorocybe<br />
resinae (Partridge & Morgan-Jones 2002), the creosote fungus<br />
Amorphotheca resinae (Parbery 1969, de Vries 1952, 1955, Ho<br />
et al. 1999) <strong>and</strong> the rhododendron fungus Seifertia azaleae (Ellis<br />
1976, Partridge & Morgan-Jones 2002, Glawe & Hummel 2006) are<br />
well-described in the literature <strong>and</strong> will not be repeated here.<br />
<strong>The</strong> three species are readily distinguished based on growth<br />
rates <strong>and</strong> overall cultural phenotypes. Agar colonies of Sorocybe<br />
resinae are coal-black, wrinkled, <strong>and</strong> restricted in growth, no matter<br />
what agar medium is employed; even after 3 mo, the colonies are<br />
rarely more than 2 cm diam (Fig. 2C). Synnemata did not form in<br />
our cultures; in vivo, the synnemata produce branched, acropetal<br />
chains of conidia with laterally thickened walls (Figs 2D–G). No<br />
thickened, refractive or darkened secession scars were evident<br />
on individual conidia or ramoconidia. Conidial masses were<br />
removed from the mononematous <strong>and</strong> synnematous parts of a<br />
freshly collected specimen (DAOM 56088a) <strong>and</strong> grown on PDA<br />
<strong>and</strong> sterilised conifer wood. <strong>The</strong>re were no discernable differences<br />
between colonies derived from the two types of conidiophores, in<br />
all cases yielding restricted black colonies, or in their microscopic<br />
characters. <strong>The</strong>refore, we conclude that these two types of<br />
conidiophores represent synanamorphs of one fungus. An identical<br />
conclusion was reached by Partridge & Morgan-Jones (2002). We<br />
documented the occurrence of this fungus in California, Oregon,<br />
<strong>and</strong> Washington State, U.S.A. <strong>and</strong> British Columbia, Canada, on<br />
resinous exudates on Abies nobilis, Picea sitchensis, Pseudotsuga<br />
menziesii <strong>and</strong> Tsuga heterophylla.<br />
Microscopic features from the holotype specimen of<br />
Hormodendrum resinae Lindau are shown in Fig. 3. Dark, thickwalled<br />
conidiophore stipes give rise to branched, acropetally<br />
www.studiesinmycology.org<br />
developing conidial chains. <strong>The</strong> conidia are relatively darkly<br />
pigmented, <strong>and</strong> the lateral walls are more conspicuously thickened<br />
<strong>and</strong> darkened than the polar walls. <strong>The</strong>re are no obvious thickened,<br />
refractive or darkened secession scars on any of the cells. Apart<br />
from the production of synnemata, the characters of the conidia<br />
<strong>and</strong> conidium ontogeny are identical in Lindau’s specimen <strong>and</strong> the<br />
synnematous specimens of Sorocybe resinae examined.<br />
In contrast, both the resin fungus <strong>and</strong> the rhododendron fungus<br />
have spreading rather than restricted agar colonies. Cultures of<br />
the resin fungus are s<strong>and</strong>y brown (Kornerup & Wanscher 1989),<br />
planar <strong>and</strong> powdery, growing 4–4.5 cm diam in 10 d on PDA (Fig.<br />
1A). Cultures of the rhododendron fungus are slower, growing<br />
2.5–3.5 cm diam after 21 d on MEA (not shown). <strong>The</strong>y are planar<br />
<strong>and</strong> greyish brown, with an orange-brown reverse. No synnemata<br />
were observed in our cultures of the rhododendron fungus on MEA,<br />
OA or PDA, but cladosporium-like conidiation occurred in the aerial<br />
mycelium.<br />
Phylogeny<br />
<strong>The</strong> large subunit analysis (LSU) was used to demonstrate the<br />
general phylogenetic relationships of the resin fungus Sorocybe<br />
resinae (DAOM 239134), the creosote fungus Amorphotheca resinae<br />
(DAOM 170427, 194228) <strong>and</strong> the rhododendron fungus Seifertia<br />
azaleae (DAOM 239136), <strong>and</strong> subsequent analyses of the internal<br />
transcribed spacers were used to estimate more precise affinities.<br />
Fig. 4 shows the LSU analysis <strong>and</strong> demonstrates that Sorocybe<br />
resinae appears to be a member of the Herpotrichiellaceae,<br />
Chaetothyriales, A. resinae is related to the inoperculate<br />
discomycetes (Leotiomycetes) <strong>and</strong> Seifertia azaleae is most closely<br />
related to a sequence labelled Mycosphaerella mycopappi A. Funk<br />
& Dorworth, which is unrelated to Mycosphaerella s. str.<br />
For the ITS alignment of Sorocybe resinae, two preliminary<br />
parsimony analyses were conducted, one with informative<br />
characters from the full alignment, the second with a subset with<br />
179 characters excluded from seven ambiguously aligned regions.<br />
<strong>The</strong> consistency indices (full 0.301, partial 0.324), tree topologies,<br />
<strong>and</strong> bootstrap supports for the two analyses were relatively <strong>similar</strong>.<br />
<strong>The</strong>refore, the complete alignment was used for the tree presented<br />
here (Fig. 5). <strong>The</strong> data matrix included 57 taxa, with 352 of 752<br />
characters phylogenetically informative. Sorocybe resinae clearly<br />
is related to Capronia <strong>and</strong> allied anamorph genera, as suggested<br />
by the LSU analysis. In the ITS analysis (Fig. 6) it forms a wellsupported<br />
clade with C. villosa Samuels, that is a well-supported<br />
sister group to species now in three different anamorph genera,<br />
Phaeococcomyces nigricans (M.A. Rich & A.M. Stern) de Hoog,<br />
Ramichloridium cerophilum, <strong>and</strong> an undescribed species of<br />
Heteroconium Petr.<br />
<strong>The</strong> ITS matrix for A. resinae included 42 taxa, with 171<br />
phylogenetically informative characters in the 530 base alignment.<br />
<strong>The</strong> phylogenetic analysis confirmed the relationship of this<br />
species with the Leotiomycetes, <strong>and</strong> provided a more precise<br />
hypothesis of its family-level relationships (Fig. 6). Amorphotheca<br />
resinae DAOM 170427 <strong>and</strong> 194228 had identical ITS sequences<br />
to another strain of the same species reported in GenBank<br />
(AY251067, from Braun et al. 2003), <strong>and</strong> one bp substitution from<br />
a second strain (AF393726 based on the isotype ATCC 200942<br />
= <strong>CBS</strong> 406.68). <strong>The</strong>se four sequences formed a sister group<br />
to two sequences of “<strong>Cladosporium</strong>” breviramosum Morgan-<br />
Jones (AF393683, AF393684). <strong>The</strong> well-supported clade of A.<br />
resinae <strong>and</strong> C. breviramosum, which represent the proposed<br />
family Amorphothecaceae, was previously noted by Braun et<br />
al. (2003). <strong>The</strong> nesting of this clade within two well-supported<br />
241
Seifert et al.<br />
100<br />
76<br />
99<br />
99<br />
100<br />
AY064704 Neofabraea alba<br />
AF281377 Neofabraea alba<br />
AF281397 Neofabraea perennans<br />
AF281388 Neofabraea malicorticis<br />
AF281365 Neofabraea krawtzewii<br />
AY129289 Pseudeurotium ovale var. ovale<br />
AY129290 Pseudeurotium ovale var. milkoi<br />
AY129287 Pseudeurotium bakeri<br />
AY129286 Pseudeurotium zonatum<br />
AY129288 Pseudeurotium desertorum<br />
AF307760 Geomyces pannorum<br />
AJ390390 Geomyces asperulatus<br />
AY789290 Heyderia abietis<br />
Z81441 Piceomphale bulgarioides<br />
AY781230 Leptodontidium elatius<br />
AY348594 Calycina herbarum<br />
AF062802 Oidiodendron periconioides<br />
99<br />
100<br />
79<br />
99<br />
88<br />
100<br />
100<br />
89<br />
88<br />
100<br />
98<br />
100<br />
AF062787 Oidiodendron pilicola<br />
AF062798 Oidiodendron maius<br />
AF062790 Oidiodendron citrinum<br />
AF062789 Oidiodendron chlamydosporicum<br />
AF062797 Oidiodendron griseum<br />
AF307774 Oidiodendron tenuissimum<br />
AF062792 Oidiodendron flavum<br />
AF062810 Myxotrichum arcticum<br />
AF062809 Oidiodendron truncatum<br />
AF062808 Oidiodendron tenuissimum<br />
AF062805 Oidiodendron setiferum<br />
AF062788 Oidiodendron cerealis<br />
AJ635314 Oidiodendron myxotrichoides<br />
AF062811 Myxotrichum cancellatum<br />
AF062817 Byssoascus striatisporus<br />
AF062803 Oidiodendron rhodogenum<br />
AF062814 Myxotrichum deflexum<br />
88<br />
A. resinae DAOM 170427, 194228, EU030278-9<br />
AF393726 Amorphotheca resinae isotype<br />
AY251067 Amorphotheca resinae<br />
AF393684 <strong>Cladosporium</strong> breviramosum<br />
AF393683 <strong>Cladosporium</strong> breviramosum<br />
AF062813 Myxotrichum chartarum<br />
AF062812 Myxotrichum carminoparum<br />
AF062816 Myxotrichum stipitatum<br />
5 changes<br />
Fig. 5. Parsimony analysis of internal transcribed spacers sequences, demonstrating the position of Amorphotheca resinae (shown in bold) in the ascomycete family<br />
Myxotrichaceae. One of 44 equally parsimonious trees (645 steps, CI = 0.460, RI = 0.758, RC = 0.349, HI = 0.540) with mid-point rooting. Bootstrap values above 70 % are<br />
shown at the relevant nodes; branches with thick lines occurred in all equally parsimonious trees.<br />
Myxotrichaceae Pseudeurotiaceae<br />
clades of Myxotrichum spp. <strong>and</strong> the associated anamorph <strong>genus</strong><br />
Oidiodendron, which comprise the family Myxotrichaceae, has not<br />
been documented previously.<br />
<strong>The</strong> ITS sequences of two strains of Seifertia azaleae were<br />
474 bp <strong>and</strong> differed by one bp. BLAST searches with these<br />
sequences revealed significant homologies only with unidentified<br />
fungi, <strong>and</strong> lower probability matches with various members of<br />
the Dothideomycetes. <strong>The</strong>refore, no taxonomically meaningful<br />
phylogenetic analysis can be presented with these ITS sequences.<br />
<strong>The</strong> species does seem to have affinities with the Dothideomycetes,<br />
but the putative relationship with Mycosphaerella, suggested by the<br />
LSU analysis, could not be confirmed with the ITS analysis.<br />
DISCUSSION<br />
Micromorphological comparisons, differences in culture characters,<br />
<strong>and</strong> phylogenetic analysis all support the conclusion that the<br />
mononematous synanamorph of Sorocybe resinae, the resin<br />
fungus, is different from the anamorph of Amorphotheca resinae,<br />
the creosote fungus. Based on ribosomal DNA sequences, the<br />
creosote fungus is related to the family Myxotrichaceae, the <strong>genus</strong><br />
Myxotrichum <strong>and</strong> its Oidiodendron anamorphs (Fig. 5). In this<br />
gene tree, Myxotrichum <strong>and</strong> the Myxotrichaceae are paraphyletic,<br />
with Amorphotheca <strong>and</strong> the Amorphothecaceae nested within<br />
them. Sorocybe appears to be an additional anamorph <strong>genus</strong><br />
phylogenetically associated with Capronia (Herpotrichiellaceae,<br />
Chaetothyriales, Fig. 6). <strong>The</strong> genetic connection between the<br />
synnematous <strong>and</strong> mononematous morphs of S. resinae was<br />
verified by morphological comparison of polyspore isolates derived<br />
from the two synanamorphs. However, the living cultures are no<br />
longer available <strong>and</strong> the connection was not confirmed with single<br />
conidium isolations. <strong>The</strong> type specimen of Hormodendrum resinae<br />
(Fig. 3) is the basis for the application of the most frequently<br />
used anamorph epithet for the creosote fungus. This specimen<br />
represents the mononematous synanamorph of Sorocybe resinae,<br />
not the anamorph of Amorphotheca resinae.<br />
It is difficult to underst<strong>and</strong> how these two fungi were confused<br />
when their micromorphologies are so different. <strong>The</strong> conidia are of<br />
the same general size <strong>and</strong> shape, but in both morphs of Sorocybe<br />
resinae (Figs 2D–G, 3C), the lateral walls are conspicuously<br />
thickened, a condition not present in the creosote fungus (Fig. 1C),<br />
<strong>and</strong> the conidia are much darker. In his monograph of <strong>Cladosporium</strong>,<br />
de Vries (1952) noted that single conidium isolates of C. avellaneum<br />
gave rise to four different colony types. In 1955, he extended these<br />
observations <strong>and</strong> decided that the much darker resin fungus was<br />
242
Fig. 6<br />
Hormoconis resinae <strong>and</strong> morphologically <strong>similar</strong> taxa<br />
87<br />
89<br />
74<br />
100<br />
100<br />
83<br />
93<br />
94<br />
100<br />
93<br />
Capronia epimyces AY156968<br />
Capronia epimyces AF050245<br />
Capronia mansonii AF050247<br />
Capronia munkii AF050250<br />
Exophiala dermatitidis AY857526<br />
Exophiala heteromorpha AY857524<br />
Exophiala heteromorpha AF083207<br />
88<br />
97<br />
94<br />
Rhinocladiella similis AY857529<br />
Melanchlenus oligospermus AY163555<br />
Exophiala oligosperma AY857534<br />
Exophiala nishimurae AY163560<br />
Rhinocladiella basitona AY163561<br />
94<br />
Exophiala spinifera AM176734<br />
Ramichloridium mackenziei AY857540<br />
Rhinocladiella atrovirens AY618683<br />
Melanchlenus eumetabolus AY163554<br />
Capronia dactylotricha AF050243<br />
Ramichloridium anceps DQ826740<br />
Exophiala lecaniicorni AY857528<br />
Exophiala mesophila AF542377<br />
Exophiala attenuata AF549446<br />
Exophiala pisciphila DQ826739<br />
Exophiala crusticola AM048755<br />
Capronia nigerrima AF050251<br />
Capronia parasitica AF050253<br />
Capronia pulcherrima AF050256<br />
Capronia pilosella DQ826737<br />
Capronia pilosella AF050255<br />
Capronia coronata AF050242<br />
Cladophialophora carrionii AY857520<br />
Cladophialophora minourae AY251087<br />
Phialophora verrucosa AF050283<br />
Capronia semiimmersa AF050260<br />
Cladophialophora emmonsii AY857518<br />
Cladophialophora emmonsii AB109184<br />
Cladophialophora bantiana AY857517<br />
Cladophialophora sp. <strong>CBS</strong> 110551 AY857510<br />
Cladophialophora devriesii AB091212<br />
Cladophialophora devriesii AM114417<br />
Cladophialophora sp. <strong>CBS</strong> 102230 AY857508<br />
Cladophialophora sp. <strong>CBS</strong> 114326 AY<br />
Cladophialophora arxii AY857509<br />
Cladophialophora arxii AB109181<br />
Cladophialophora minourae AB091213<br />
Cladophialophora sp. AY781217<br />
Cladophialophora boppii AB109182<br />
Capronia fungicola AF050246<br />
83<br />
90<br />
94<br />
99<br />
98<br />
99<br />
80<br />
100<br />
100<br />
100<br />
Sorocybe resinae DAOM 239134, EU030275<br />
Capronia villosa AF050261<br />
Phaeococcomyces nigricans AY843154<br />
Ramichloridium cerophilum AF050286<br />
Heteroconium sp. AJ748260<br />
Phaeococcomyces catenatus AY843041<br />
Phaeococcomyces chersonesos AJ507323<br />
Metulocladosporiella musae DQ008138<br />
Metulocladosporiella musicola DQ008136<br />
Capronia acutiseta AF050241<br />
10 changes<br />
Fig. 6. Parsimony analysis of internal transcribed spacers sequences, demonstrating the position of Sorocybe resinae (shown in bold) among species of Capronia<br />
(Herpotrichiellaceae, Chaetothyriales) <strong>and</strong> its associated anamorph genera. One of 34 equally parsimonious trees (2 607 steps, CI = 0.301, RI = 0.506, RC = 0.153, HI = 0.699),<br />
with mid-point rooting. Bootstrap values above 70 % are shown at the relevant nodes; branches with thick lines occurred in all equally parsimonious trees.<br />
the same as one of his mutant forms of the creosote fungus,<br />
despite never having isolated such a dark spored form from any of<br />
his cultures. Parbery (1969) implied that the demonstrated ability<br />
of the creosote fungus to grow on a diversity of hydrocarbon-rich<br />
substrates favoured the thought that it would be able to grow on<br />
conifer resin. If cultures of the true Sorocybe resinae had been<br />
available, it is unlikely that this confusion would have persisted for<br />
so long. In vitro, the creosote fungus <strong>and</strong> the resin fungus are so<br />
different (Figs 1A, 2C) that it would difficult to defend the idea that<br />
they were mutants of the same fungus. <strong>The</strong>se differences in the<br />
cultures are reflected by the disparate phylogenetic affinities of<br />
what now are clearly demonstrated to be two different species.<br />
Unfortunately, the name Hormodendrum resinae has been<br />
misapplied to the creosote fungus, a species of economic<br />
importance. Also unfortunately, this species is the type of<br />
www.studiesinmycology.org<br />
Hormoconis, a generic name that the community concerned with<br />
this fungus has been slow to adapt to in the 30 years since its<br />
introduction. <strong>The</strong>re are several possible solutions to this problem.<br />
<strong>The</strong> conventional solution would be to apply names based strictly<br />
on the type specimens <strong>and</strong> accept Hormoconis as a synonym of<br />
Sorocybe, or to use it as a generic name for the mononematous<br />
synanamorph of the resin fungus. A new anamorph <strong>genus</strong> would<br />
then be described for the creosote fungus, making <strong>Cladosporium</strong><br />
avellaneum G.A. de Vries the basionym for its type. However, the<br />
resulting binomial would be unfamiliar to those concerned with the<br />
creosote fungus, <strong>and</strong> the earlier literature citing H. resinae would<br />
be misleading.<br />
A more parsimonious solution is possible. Article 14.9 of the<br />
International Code of Botanical Nomenclature (McNeil et al. 2006)<br />
allows for conservation of a name with a different type from that<br />
243
Seifert et al.<br />
designated by the authors. <strong>The</strong> name Hormodendrum resinae is<br />
not otherwise needed because the mononematous synanamorph<br />
of the resin fungus is rarely referred to by a Latin binomial, <strong>and</strong><br />
because Sorocybe resinae is based on a different type. <strong>The</strong>refore,<br />
a new type specimen could be proposed <strong>and</strong> conserved for<br />
Hormodendrum resinae Lindau, preferably the holotype of A.<br />
resinae (MELU 7130). This would make the anamorph-teleomorph<br />
connection unequivocal, maintain current species epithets <strong>and</strong><br />
taxonomic authorities, <strong>and</strong> ensure that most of the historical<br />
literature can be interpreted easily without the need to consult<br />
complicated nomenclators (Table 1). However, by perpetuating the<br />
use of the epithet “resinae”, this change would also perpetuate the<br />
misunderst<strong>and</strong>ing that resin is a possible substrate for the creosote<br />
fungus. In any case, the use of this epithet for the teleomorph of<br />
the creosote fungus, Amorphotheca resinae, is legitimate <strong>and</strong> valid,<br />
<strong>and</strong> unlikely ever to be changed.<br />
A third option would be an intermediate one. <strong>The</strong> application<br />
of the name <strong>Cladosporium</strong> avellaneum G.A. de Vries has never<br />
been in doubt, <strong>and</strong> it would be possible to conserve this species as<br />
the type of Hormoconis. This has the advantage of maintaining the<br />
familiar generic name Hormoconis, in combination with a species<br />
epithet that has been consistently applied. Furthermore, this solution<br />
would allow the confusion about the application <strong>and</strong> correct author<br />
citation around the epithet “resinae” for the anamorph of creosote<br />
fungus to recede.<br />
<strong>The</strong> second <strong>and</strong> third solutions require formal taxonomic<br />
proposals to be published in Taxon. We will argue the merits of<br />
these possible solutions at more length in that venue.<br />
<strong>The</strong> phylogenetic position of A. resinae raises additional<br />
taxonomic problems. This fungus typifies the monotypic family<br />
Amorphothecaceae, which has been considered incertae<br />
sedis since its description by Parbery (1969). Our phylogenetic<br />
analysis suggests that this family sits within the Myxotrichaceae.<br />
Amorphothecaceae (1969) is the older name, but Myxotrichaceae<br />
(1985) is well-entrenched in the mycological literature. As a<br />
consequence, the Myxotrichaceae are paraphyletic with respect<br />
to the Amorphothecaceae. <strong>The</strong> peridium of A. resinae, the only<br />
species presently placed in this family, lacks the thick-walled<br />
appendages that characterise most species of the Myxotrichaceae.<br />
Furthermore, the acropetal-blastic features of the anamorph of<br />
A. resinae differ from the thallic-arthric conidiogenesis of the<br />
other anamorphs associated with the Myxotrichaceae, principally<br />
Oidiodendron. <strong>The</strong>se morphological differences explain why the<br />
affinity of A. resinae with the Myxotrichaceae was not noted before.<br />
A formal proposal to conserve Myxotrichaceae as the name for this<br />
family might be prudent eventually, but this should await analysis of<br />
additional genes to confirm the phylogenetic relationship.<br />
Whether <strong>Cladosporium</strong> breviramosum, originally isolated from<br />
discoloured wallpaper, is actually a distinct species from A. resinae<br />
requires further study. It is clear that this species, if it is distinct,<br />
would be a member of Hormoconis rather than <strong>Cladosporium</strong>. Apart<br />
from the study of additional specimens, it might be fruitful to attempt<br />
to induce an Amorphotheca-like teleomorph in the two available<br />
cultures of C. breviramosum, <strong>and</strong> to compare the morphology with<br />
that of A. resinae. According to Parbery (1969), A. resinae includes<br />
both homothallic <strong>and</strong> heterothallic strains.<br />
Unfortunately, the phylogenetic affinities of Seifertia azaleae<br />
were not established with certainty in this study. Its closest relative<br />
in the LSU analysis is a sequence identified as Mycosphaerella<br />
mycopappi Funk & Dorworth (U43480, based on the apparent<br />
type culture ATCC 64711), but this sequence does not cluster<br />
with others representing the family Mycosphaerellaceae (data not<br />
shown). Similarly, the ITS sequences of the rhododendron fungus<br />
did not cluster with the many ITS sequences of Mycosphaerella<br />
available. Presently, it seems that Seifertia azaleae fungus is allied<br />
with the Dothideomycetes, but its precise affinities are uncertain. It<br />
is clear that this fungus should not be classified in Pycnostysanus<br />
(a taxonomic synonym of Sorocybe), <strong>and</strong> continued recognition of<br />
the monotypic <strong>genus</strong> Seifertia seems justified.<br />
ACKNOWLEDGEMENTS<br />
We are grateful to Sarah Hambleton <strong>and</strong> Marie Davey for providing ITS alignments<br />
that served as the basis for analyses in this paper. Walter Gams <strong>and</strong> Scott Redhead<br />
provided expert nomenclatural advice <strong>and</strong> helpful reviews of this manuscript.<br />
We thank Uwe Braun for proposing the third option for resolving the name of the<br />
creosote fungus, outlined in the text. <strong>The</strong> Canadian Collection of Fungal Cultures<br />
(DAOM) provided several of the strains for this study. <strong>The</strong> curators of DAOM <strong>and</strong> B<br />
kindly provided access to critical type specimens.<br />
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245
INDEX OF FUNGAL NAMES<br />
Arranged according to <strong>genus</strong> <strong>and</strong> then epithets.<br />
Synonymised names are italicised <strong>and</strong> page numbers of main entries are bolded. A superscript asterisk ( * ) points to pages with illustrations,<br />
a superscript c ( c ) to pages with a cladogram, a superscript t ( t ) to pages with a table <strong>and</strong> a superscript k ( k ) to pages with a key to genera or<br />
species.<br />
Acidomyces, 12<br />
Acidomyces richmondensis, 12<br />
Acladium biophilum, 85<br />
Acladium herbarum, 105<br />
Acrotheca acuta, 84<br />
Acrotheca cerophila, 72<br />
Acrotheca multispora, 84<br />
Acrothecium multisporum, 84<br />
Amorphotheca, 235, 242<br />
Amorphotheca resinae, 235–236 * , 238 t , 239–241, 242 c , 244<br />
Annulatascus triseptatus, 62 c , 64 c<br />
Antennaria cellaris, 75<br />
Anungitea, 54 k , 185, 203–204, 216<br />
Anungitea fragilis, 203–204<br />
Anungitea heterospora, 204<br />
Anungitea longicatenata, 54 k<br />
Anungitea rhabdospora, 204<br />
Anungitea uniseptata, 211<br />
Anungitopsis, 203–204, 216<br />
Anungitopsis amoena, 163, 204, 207<br />
Anungitopsis intermedia, 204, 209<br />
Anungitopsis speciosa, 186, 188 c , 190 t , 193 c , 203–204, 216<br />
Apiosporina, 205, 216<br />
Apiosporina collinsii, 189 c , 191 t , 194 c , 205<br />
Aporothielavia leptoderma, 36 c –37 c<br />
Ascochyta pisi, 36 c –37 c<br />
Ascocoryne cylichnium, 240 c<br />
Ascosphaera apis, 240 c<br />
Ascotaiwania hughesii, 62 c , 64 c<br />
Asperisporium, 31, 216<br />
Athelia epiphylla, 6 c –7 c , 36 c –37 c , 58, 62 c , 64 c , 188 c , 193 c<br />
Aureobasidium caulivorum, 158 t<br />
Batcheloromyces, 4, 12 k<br />
Batcheloromyces eucalypti, 2 t , 6 c –7 c , 12<br />
Batcheloromyces leucadendri, 2 t , 6 c –7 c , 12–13 *<br />
Batcheloromyces proteae, 2 t , 6 c –7 c , 12, 63 c , 65 c , 162 c<br />
Bispora, 52 k<br />
Bisporella citrina, 36 c –37 c , 240 c<br />
Blumeria graminis f. sp. bromi, 36 c –37 c<br />
Botryosphaeria ribis , 97 c , 240 c<br />
Botryosphaeria stevensii , 97 c<br />
Botrytis epichloës, 89<br />
Byssoascus striatisporus, 240 c , 242 c<br />
Cadophora elatum, 46<br />
Calicium, 236<br />
Calicium viride, 240 c<br />
Calycina herbarum, 242 c<br />
Capnobotryella, 11 k –12, 17<br />
Capnobotryella renispora, 2 t , 5, 6 c –7 c , 12<br />
Capnodium citri, 240 c<br />
Capnodium coffeae, 6 c –7 c<br />
Capnodium salicinum, 2 t , 6 c –7 c<br />
Capronia, 55, 92, 185, 203, 216, 241–243<br />
Capronia acutiseta, 188 c , 193 c , 243 c<br />
Capronia coronata, 62 c , 64 c , 243 c<br />
Capronia dactylotricha, 243 c<br />
Capronia epimyces, 243 c<br />
Capronia fungicola, 243 c<br />
Capronia hanliniana, 185, 212<br />
Capronia hystrioides, 185, 212<br />
Capronia mansonii, 6 c –7 c , 62 c , 64 c , 188 c , 193 c , 240 c , 243 c<br />
Capronia munkii, 188 c , 193 c , 243 c<br />
Capronia nigerrima, 243 c<br />
Capronia parasitica, 243 c<br />
Capronia pilosella, 188 c , 193 c , 243 c<br />
Capronia pulcherrima, 62 c , 64 c , 243 c<br />
Capronia semiimmersa, 243 c<br />
Capronia villosa, 241, 243 c<br />
Caproventuria, 185, 216, 205, 230<br />
Caproventuria hanliniana, 189 c , 194 c , 205, 212<br />
Caproventuria hystrioides, 185, 212<br />
Carpoligna, 92<br />
Carpoligna pleurothecii, 62 c , 64 c , 83<br />
Castanedaea, 52 k<br />
Catenulostroma, 12 k , 13–14, 164<br />
Catenulostroma abietis, 2 t , 6 c –7 c , 14 k –15, 17, 63 c , 65 c<br />
Catenulostroma castellanii, 2 t , 6 c –7 c , 36 c –37 c<br />
Catenulostroma chromoblastomycosum, 2 t , 6 c –7 c , 14 k –15 * , 16,<br />
36 c –37 c<br />
Catenulostroma elginense, 2 t , 6 c –7 c , 15 k –16<br />
Catenulostroma excentricum, 10, 14 k , 16<br />
Catenulostroma germanicum, 2 t , 6 c –7 c , 14 k , 16 * –17, 36 c –37 c<br />
Catenulostroma macowanii, 2 t , 6 c –7 c , 15 k , 17, 63 c , 65 c , 162 c<br />
Catenulostroma microsporum, 2 t , 6 c –7 c , 10, 15 k , 17<br />
Catenulostroma protearum, 14–15 k , 17<br />
Catenulostroma sp., 2 t<br />
Ceramothyrium carniolicum, 188 c , 193 c , 240 c<br />
Cercospora, 1, 31, 164<br />
Cercospora apii , 97 c<br />
Cercospora beticola , 97 c , 111 c<br />
Cercospora cruenta, 162 c<br />
Cercospora epicoccoides, 11<br />
Cercospora eucalypti, 26<br />
Cercospora penicillata, 1<br />
Cercospora zebrina, 162 c<br />
Cercosporella centaureicola, 2 t , 6 c –7 c , 63 c , 65 c<br />
Cercostigmina, 30–31<br />
Chaetomium globosum, 36 c –37 c<br />
Chaetomium homopilatum, 36 c –37 c<br />
Chloridium apiculatum, 68<br />
Chloridium indicum, 68, 72–73<br />
Chloridium minus, 76<br />
Chloridium musae, 57, 72<br />
Chloridium schulzerii, 84<br />
Cibiessia, 11 k , 17, 24–25, 30<br />
Cibiessia dimorphospora, 2 t , 6 c –7 c , 17, 63 c , 65 c<br />
Cibiessia minutispora, 2 t , 6 c –7 c<br />
Cibiessia nontingens, 2 t , 6 c –7 c<br />
Cistella acuum, 214<br />
246
Cladophialophora, 52 k , 54 k –55, 167, 185–188, 198–200, 204, 216,<br />
219–220, 226–228, 230–232<br />
Cladophialophora ajelloi, 231<br />
Cladophialophora arxii, 243 c<br />
Cladophialophora australiensis, 186, 187–188 c , 190 t , 193 c –194 *<br />
Cladophialophora bantiana, 187, 231, 243 c<br />
Cladophialophora boppii, 231, 243 c<br />
Cladophialophora brevicatenata, 185, 212<br />
Cladophialophora carrionii, 185, 187–188 c , 193 c , 219, 220 t –221 t ,<br />
222–223, 224 c –226 c , 227–228 * , 229–230 * , 231–232, 243 c<br />
Cladophialophora cf. carrionii, 232<br />
Cladophialophora chaetospira, 54 k , 187–188 c , 190 t , 193 c , 195 *<br />
Cladophialophora devriesii, 231, 243 c<br />
Cladophialophora emmonsii, 243 c<br />
Cladophialophora hachijoensis, 185<br />
Cladophialophora hostae, 187, 188 c –189, 190 t , 193 c , 195 * –196 * ,<br />
199, 204<br />
Cladophialophora humicola, 188 c –189, 190 t , 193 c , 196 *<br />
Cladophialophora minourae, 243 c<br />
Cladophialophora potulentorum, 188 c , 190 t , 193 c , 197 * –198<br />
Cladophialophora proteae, 188 c , 190 t , 193 c , 198 *<br />
Cladophialophora scillae, 188 c , 190 t , 193 c , 198–199 * , 204<br />
Cladophialophora sp., 223, 225, 243 c<br />
Cladophialophora sylvestris, 188 c , 190 t , 193 c , 200 *<br />
Cladophialophora yegresii, 185, 221 t , 224 c , 226 c , 227–228, 229 * –<br />
230 * , 231–232<br />
Cladoriella, 55<br />
Cladoriella eucalypti, 34 t , 36 c –37 c<br />
<strong>Cladosporium</strong>, 8, 21, 30, 33, 35, 38–39, 41, 44, 47, 49, 52 k , 55,<br />
57, 72, 95–96, 103, 105–106 * , 107, 113, 115–116, 137, 142,<br />
153–155, 157, 161–169, 173, 176, 181, 209, 227, 230, 237–<br />
238, 242, 244<br />
“<strong>Cladosporium</strong>” adianticola, 190 t , 193 c<br />
<strong>Cladosporium</strong> alneum, 116–117<br />
<strong>Cladosporium</strong> alopecuri, 142<br />
<strong>Cladosporium</strong> amoenum, 207<br />
<strong>Cladosporium</strong> antarcticum, 108 t , 111 c –112 c , 114 k –115, 116 * –117 * ,<br />
154–155<br />
<strong>Cladosporium</strong> araguatum, 43<br />
<strong>Cladosporium</strong> argillaceum, 50<br />
<strong>Cladosporium</strong> arthoniae, 41, 116<br />
<strong>Cladosporium</strong> arthrinioides, 140<br />
<strong>Cladosporium</strong> arthropodii, 142<br />
<strong>Cladosporium</strong> asterinae, 55, 103<br />
<strong>Cladosporium</strong> avellaneum, 235, 237–238 t , 242–244<br />
<strong>Cladosporium</strong> breviramosum, 241–242 c , 244<br />
<strong>Cladosporium</strong> bruhnei, 2 t , 6 c –7 c , 36 c –37 c , 63 c , 65 c , 97 c , 107–108 t ,<br />
110, 111 c –112 c , 114 k , 118 * –120 * , 145, 154–155, 158 t , 162 c –165 c ,<br />
168<br />
<strong>Cladosporium</strong> brunneum, 133<br />
<strong>Cladosporium</strong> carpophilum, 205<br />
<strong>Cladosporium</strong> castellanii, 43, 44<br />
<strong>Cladosporium</strong> cellare, 75<br />
<strong>Cladosporium</strong> cerophilum, 72<br />
<strong>Cladosporium</strong> chlorocephalum, 95–96, 103<br />
<strong>Cladosporium</strong> cladosporioides, 2 t , 6 c –7 c , 36 c –37 c , 63 c , 65 c , 108 t ,<br />
111 c –112 c , 113 k , 140, 147, 150, 153, 158 t , 162 c –165 c , 168, 174<br />
<strong>Cladosporium</strong> colocasiae, 162 c<br />
<strong>Cladosporium</strong> cubense, 19–20<br />
<strong>Cladosporium</strong> dominicanum, 158 t , 162 c –165 c , 166 * , 168 k –169,<br />
170 * , 175<br />
<strong>Cladosporium</strong> elatum, 46<br />
<strong>Cladosporium</strong> ferrugineum, 20–21<br />
<strong>Cladosporium</strong> fulvum, 55<br />
<strong>Cladosporium</strong> fusiforme, 158 t , 162 c –165 c , 166 * , 168 k –169, 171 *<br />
<strong>Cladosporium</strong> gracile, 133<br />
<strong>Cladosporium</strong> graminum, 133<br />
<strong>Cladosporium</strong> halotolerans, 158 t , 162 c –165 c , 166 * –167, 168 k –169,<br />
172 * –173, 175<br />
<strong>Cladosporium</strong> helicosporum, 18<br />
<strong>Cladosporium</strong> hemileiae, 55<br />
<strong>Cladosporium</strong> herbaroides, 108 t , 111 c –112 c , 114 k , 120, 121 * –123 * ,<br />
154<br />
<strong>Cladosporium</strong> herbarum, 8, 33, 96–97 c , 101, 105, 107–108 t , 110,<br />
111 c –112 c , 113–114 k , 118, 120, 122, 124, 125 * –127 * , 133, 142,<br />
153–155, 158 t , 162 c , 164–166, 168, 181<br />
<strong>Cladosporium</strong> herbarum var. macrocarpum, 105, 129<br />
<strong>Cladosporium</strong> hordei, 118<br />
<strong>Cladosporium</strong> hypophyllum, 140<br />
<strong>Cladosporium</strong> iridis, 36 c –37 c , 97 c , 109 t , 111 c –112 c , 114 k , 125, 127–<br />
128 * , 135, 137, 153<br />
<strong>Cladosporium</strong> langeronii, 159 t , 162 c –165 c , 166 * –167, 168 k , 173–<br />
174 *<br />
<strong>Cladosporium</strong> lichenicola, 116<br />
<strong>Cladosporium</strong> licheniphilum, 116<br />
<strong>Cladosporium</strong> lichenum, 116<br />
<strong>Cladosporium</strong> macrocarpum, 97 c , 105, 107, 109 t –110, 111 c –112 c ,<br />
114 k , 122, 129 * –130 * , 131 * –133, 153–155<br />
<strong>Cladosporium</strong> magnusianum, 133, 135<br />
<strong>Cladosporium</strong> musae, 55<br />
<strong>Cladosporium</strong> nigrellum, 209<br />
<strong>Cladosporium</strong> olivaceum, 237<br />
<strong>Cladosporium</strong> ossifragi, 97 c , 109 t –110, 111 c –112 c , 114 k , 133 * –134 * ,<br />
135, 154<br />
<strong>Cladosporium</strong> oxysporum, 159 t , 162 c –165 c , 168<br />
<strong>Cladosporium</strong> paeoniae, 95–96, 101, 103<br />
<strong>Cladosporium</strong> paeoniae var. paeoniaeanomalae, 95–96<br />
<strong>Cladosporium</strong> pseudiridis, 109 t , 111 c –112 c , 114 k , 135, 136 * –137 *<br />
<strong>Cladosporium</strong> psoraleae, 116–117<br />
<strong>Cladosporium</strong> psychrotolerans, 159 t , 162 c –165 c , 166 * –167, 168 k ,<br />
175–176 *<br />
<strong>Cladosporium</strong> ramotenellum, 107, 109 t , 111 c –112 c , 115 k , 137–138 * ,<br />
139 * –140, 150, 154, 159 t , 163 c<br />
<strong>Cladosporium</strong> resinae, 235, 237–238 t<br />
<strong>Cladosporium</strong> rigidophorum, 21, 22<br />
<strong>Cladosporium</strong> salinae, 159 t , 161, 162 c –165 c , 166 * , 168 k , 175,<br />
176–177 *<br />
<strong>Cladosporium</strong> scillae, 198<br />
<strong>Cladosporium</strong> sinuosum, 109 t , 111 c –112 c , 114 k , 140 * –141 * , 142<br />
<strong>Cladosporium</strong> sp., 109 t , 111 c , 149, 159 t , 163 c , 165 c<br />
<strong>Cladosporium</strong> sphaerospermum, 2 t , 6 c –7 c , 63 c , 65 c , 153, 157, 159 t ,<br />
162 c –165 c , 166 * , 167–168 k , 169, 173–175, 177–178 * , 181<br />
<strong>Cladosporium</strong> spinulosum, 109 t , 111 c –112 c , 114 k , 142 * , 145, 154–<br />
155, 160 t , 162 c –165 c , 166 * , 168 k , 179 * , 180–181<br />
<strong>Cladosporium</strong> staurophorum, 55<br />
<strong>Cladosporium</strong> strumelloideum, 23<br />
<strong>Cladosporium</strong> subinflatum, 109 t , 111 c –112 c , 114 k , 143 * –144 * , 145,<br />
154, 160 t , 163 c –165 c<br />
<strong>Cladosporium</strong> subnodosum, 150<br />
<strong>Cladosporium</strong> subtilissimum, 109 t , 111 c –112 c , 114 k , 145 * –146 * ,<br />
147 * , 149–150, 154–155<br />
<strong>Cladosporium</strong> tenellum, 97 c , 109 t , 111 c –112 c , 115 k , 137, 140, 148 * –<br />
149 * , 150, 153–154<br />
247
<strong>Cladosporium</strong> tenuissimum, 160 t , 162 c –163 c , 168<br />
<strong>Cladosporium</strong> uredinicola, 2 t , 6 c –7 c , 36 c –37 c , 63 c , 65 c , 116, 162 c<br />
<strong>Cladosporium</strong> variabile, 97 c , 110 t , 111 c –112 c , 114 k , 131, 150 * –152 * ,<br />
153–154<br />
<strong>Cladosporium</strong> velox, 160 t , 162 c –165 c , 166 * , 168 k , 180 * –181<br />
Clathrosporium intricatum, 189 c , 193 c<br />
Coccodinium, 28<br />
Coccodinium bartschii, 2 t , 4–5, 6 c –7 c , 28, 29 *<br />
Cochliobolus heliconiae, 240 c<br />
Colletogloeopsis, 24<br />
Colletogloeopsis blakelyi, 26<br />
Colletogloeopsis considenianae, 26<br />
Colletogloeopsis dimorpha, 26<br />
Colletogloeopsis gauchensis, 26<br />
Colletogloeopsis molleriana, 10<br />
Colletogloeopsis nubilosum, 10<br />
Colletogloeopsis stellenboschiana, 26<br />
Colletogloeopsis zuluensis, 26<br />
Colletogloeum nubilosum, 10<br />
Coniella, 25<br />
Conioscypha lignicola, 62 c , 64 c<br />
Conioscyphascus varius, 62 c , 64 c<br />
Coniosporium, 216<br />
Coniothecium macowanii, 17<br />
Coniothecium punctiforme, 17<br />
Coniothyrium palmarum, 34 t , 36 c –37 c<br />
Coniothyrium zuluense, 26<br />
Coremium, 236, 239<br />
Cryptadelphia groenendalensis, 62 c , 64 c<br />
Cryptadelphia polyseptata, 62 c , 64 c<br />
Cryptococcus neoformans, 232<br />
Cudonia lutea, 240 c<br />
Cylindrosympodium, 204<br />
Cylindrosympodium lauri, 189 c –190 t , 194 c , 204 *<br />
Cylindrosympodium variabile, 205<br />
Cyphellophora, 201<br />
Cyphellophora fusarioides, 201<br />
Cyphellophora guyanensis, 201<br />
Cyphellophora hylomeconis, 188 c , 190 t , 193 c , 200 * –201<br />
Cyphellophora laciniata, 188 c , 190 t , 193 c , 201<br />
Cyphellophora pluriseptata, 201<br />
Cyphellophora suttonii, 201<br />
Cyphellophora vermispora, 201<br />
Davidiella, 5 k , 8, 30, 33, 35, 44, 52 k , 92, 96, 106, 115, 153, 155,<br />
163, 166, 173, 230<br />
Davidiella allicina, 2 t , 108 t , 112 c , 115 k , 118, 119 * –120 *<br />
Davidiella macrocarpa, 109 t , 112 c , 115 k , 129–130 * , 132 * –133<br />
Davidiella macrospora, 109 t , 112 c , 125, 128<br />
Davidiella sp., 108 t , 110 t , 111 c –112 c , 155<br />
Davidiella tassiana, 8, 108 t , 112 c , 115 k , 120, 122, 124 * –125, 126 * ,<br />
158 t , 163 c –165 c , 166, 168<br />
Davidiella variabile, 110 t , 112 c , 115 k , 151 * , 152–153<br />
Dematium epiphyllum var. chionanthi, 122, 124<br />
Dematium graminum, 129, 132<br />
Dematium herbarum, 8, 105, 122<br />
Dematium herbarum var. brassicae, 129, 132<br />
Dematium nigrum, 238 t –239<br />
Dematium vulgare var. foliorum, 129, 132<br />
Dematium vulgare var. typharum, 129, 132<br />
Dendryphion resinae, 238 t<br />
Denticularia, 31<br />
Devriesia, 11 k , 17, 43, 54 k –55, 164, 227, 230<br />
Devriesia acadiensis, 34 t , 36 c –37 c<br />
Devriesia americana, 34 t , 36 c –37 c , 42–43 *<br />
Devriesia shelburniensis, 34 t , 36 c –37 c , 43<br />
Devriesia staurophora, 5, 6 c –7 c , 17, 36 c –37 c , 63 c , 65 c , 162 c<br />
Devriesia thermodurans, 34 t , 36 c –37 c<br />
Dichocladosporium, 53 k , 55, 96, 103<br />
Dichocladosporium chlorocephalum, 36 c –37 c , 96, 97 c , 98 t* –99 * ,<br />
100 * –101, 102 *<br />
Didymella bryoniae, 36 c –37 c<br />
Didymella cucurbitacearum, 36 c –37 c<br />
Didymellina macrospora, 126<br />
Digitopodium, 52 k , 55<br />
Diplococcium, 39<br />
Discosphaerina fagi, 162 c , 240 c<br />
Dissoconium, 5 k , 28, 61<br />
Dissoconium aciculare, 2 t , 6 c –7 c , 28, 63 c , 65 c<br />
Dissoconium commune, 2 t , 28<br />
Dissoconium dekkeri, 2 t<br />
Distocercospora, 31<br />
Dothidea insculpta, 162 c<br />
Dothidea ribesia, 162 c<br />
Dothistroma, 4<br />
Exophiala, 75, 92, 185, 201, 203, 216, 231<br />
Exophiala attenuata, 243 c<br />
Exophiala crusticola, 243 c<br />
Exophiala dermatitidis, 6 c –7 c , 62 c , 64 c , 188 c , 193 c , 201, 243 c<br />
Exophiala eucalyptorum, 188 c , 190 t , 193 c , 203 *<br />
Exophiala heteromorpha, 243 c<br />
Exophiala jeanselmei, 6 c –7 c , 62 c , 64 c , 188 c , 193 c<br />
Exophiala lecaniicorni, 243 c<br />
Exophiala mesophila, 201, 243 c<br />
Exophiala nishimurae, 243 c<br />
Exophiala oligosperma, 6 c –7 c , 188 c , 193 c , 243 c<br />
Exophiala phaeomuriformis, 201<br />
Exophiala pisciphila, 188 c , 193 c , 243 c<br />
Exophiala sp. 1, 188 c , 190 t , 193 c , 201–202 *<br />
Exophiala sp. 2, 188 c , 190 t , 193 c , 201–202 *<br />
Exophiala sp. 3, 188 c , 190 t , 193 c<br />
Exophiala spinifera, 243 c<br />
Fasciatispora petrakii, 37 c , 188 c , 193 c<br />
Fonsecaea, 231<br />
Fonsecaea monophora, 231<br />
Fonsecaea pedrosoi, 6 c –7 c , 62 c , 64 c , 188 c , 193 c , 219, 232, 240 c<br />
Friedmanniomyces, 11 k –12, 24<br />
Fumagospora capnodioides, 2 t<br />
Fusarium, 231<br />
Fusicladium, 17, 52 k , 54 k –55, 91, 96, 103, 163, 176, 185, 188–189,<br />
197–199, 203–204, 205, 211, 216<br />
Fusicladium africanum, 189 c –190 t , 194 c , 205–206 * , 211<br />
Fusicladium amoenum, 162 c , 189 c –190 t , 194 c , 206 * –207<br />
Fusicladium brevicatenatum, 212<br />
Fusicladium carpophilum, 189 c –190 t , 194 c<br />
Fusicladium caruanianum, 208<br />
Fusicladium catenosporum, 189 c , 191 t , 194 c<br />
Fusicladium convolvularum, 189 c , 191 t , 194 c , 207 * –208<br />
Fusicladium effusum, 161, 162 c –163, 189 c , 191 t , 194 c<br />
Fusicladium fagi, 189 c , 191 t , 194 c , 208 * –209<br />
“Fusicladium hyphopodioides”, 215<br />
Fusicladium intermedium, 189 c , 191 t , 194 c , 209 *<br />
Fusicladium m<strong>and</strong>shuricum, 189 c , 191 t , 194 c , 210<br />
Fusicladium matsushimae, 209<br />
Fusicladium oleagineum, 189 c , 191 t , 194 c<br />
248
Fusicladium phillyreae, 189 c , 191 t , 194 c<br />
Fusicladium pini, 189 c , 191 t , 194 c , 210 * , 211–212<br />
Fusicladium pomi, 189 c , 191 t , 194 c<br />
Fusicladium radiosum, 189 c , 191 t , 194 c<br />
Fusicladium ramoconidii, 189 c , 191 t , 194 c , 211 * –212 *<br />
Fusicladium rhodense, 189 c , 191 t , 194 c , 212–213 *<br />
Fusicladium scillae, 198–199<br />
Fusicladium sp., 212<br />
Fusicladosporium, 205<br />
Geomyces asperulatus, 242 c<br />
Geomyces pannorum, 242 c<br />
Glyphium elatum, 188 c , 193 c , 240 c<br />
Golovinomyces cichoracearum, 240 c –241<br />
Gomphinaria, 84<br />
Gomphinaria amoena, 84<br />
Graphiopsis chlorocephala, 96<br />
Hansfordia biophila, 85<br />
Hansfordia torvi, 85<br />
Haplographium chlorocephalum, 96<br />
Haplographium chlorocephalum var. ovalisporum, 96<br />
Haplotrichum, 54 k<br />
Helminthosporium, 239<br />
Helminthosporium variabile, 150<br />
Hendersonia gr<strong>and</strong>ispora, 11<br />
Heteroconium, 54 k , 187, 241<br />
Heteroconium chaetospira, 187, 216<br />
Heteroconium citharexyli, 187<br />
Heteroconium sp., 243 c<br />
Heterosporium, 33, 142, 153<br />
Heterosporium iridis, 125<br />
Heterosporium magnusianum, 133, 135<br />
Heterosporium ossifragi, 133, 135<br />
Heterosporium variabile, 150<br />
Heyderia abietis, 242 c<br />
Hormiactis, 51 k<br />
Hormoconis, 54 k , 235, 238, 243–244<br />
Hormoconis resinae, 34 t , 36 c –37 c , 235, 238 t , 240 c , 243<br />
“Hormodendron”, 237<br />
Hormodendrum, 237<br />
“Hormodendrum” elatum, 47<br />
Hormodendrum hordei, 118, 120<br />
Hormodendrum langeronii, 167, 173–174<br />
Hormodendrum olivaceum, 237<br />
Hormodendrum resinae, 54 k , 235, 237–238 t , 239–240 * , 241–244<br />
Hormodendrum viride, 237<br />
Hortaea, 12 k , 17<br />
Hortaea werneckii, 2 t , 6 c –7 c , 17<br />
Hyalodendriella, 46, 52 k<br />
Hyalodendriella betulae, 34 t , 36 c –37 c , 46–47 *<br />
Hyalodendron, 46, 51 k<br />
Iodosphaeria aquatica, 240 c<br />
Kirramyces, 24<br />
Kirramyces destructans, 26<br />
Kirramyces epicoccoides, 11<br />
Kirramyces eucalypti, 26<br />
Kumbhamaya, 201<br />
Lacazia loboi, 167<br />
Lecanosticta, 24<br />
Leptodontidium elatius, 242 c<br />
Leptospora rubella, 36 c –37 c<br />
Letendraea helminthicola, 240 c<br />
Loboa loboi, 159 t<br />
Lojkania enalia, 240 c<br />
Lophodermium pinastri, 240 c<br />
Lylea, 51 k<br />
Madurella mycetomatis, 232<br />
Magnaporthe grisea, 62 c , 64 c<br />
Melanchlenus eumetabolus, 243 c<br />
Melanchlenus oligospermus, 243 c<br />
Melanelia exasperatula, 240 c<br />
Metacoleroa, 205, 216<br />
Metacoleroa dickiei, 62 c , 64 c , 189 c , 194 c , 205<br />
Metulocladosporiella, 18, 51, 53 k , 55, 103<br />
Metulocladosporiella musae, 103, 188 c , 193 c , 243 c<br />
Metulocladosporiella musicola, 188 c , 193 c , 243 c<br />
Mollisia cinerea, 6 c –7 c<br />
Mycosphaerella, 1, 5 k , 8, 28, 30, 31–33, 45, 51 k , 55, 91, 96, 105,<br />
115, 155, 163, 164, 166, 205, 241–242, 244<br />
Mycosphaerella africana, 9, 36 c –37 c<br />
Mycosphaerella alchemillicola, 34 t<br />
Mycosphaerella alistairii, 8–9<br />
Mycosphaerella ambiphylla, 10<br />
Mycosphaerella associata, 8–9<br />
Mycosphaerella aurantia , 97 c<br />
Mycosphaerella bellula, 8, 10<br />
“Mycosphaerella” communis, 2 t , 6 c –7 c , 28, 63 c , 65 c<br />
Mycosphaerella cryptica, 8, 10<br />
Mycosphaerella dendritica, 8, 10<br />
Mycosphaerella endophytica, 63 c , 65 c<br />
Mycosphaerella eucalypti, 8<br />
Mycosphaerella excentrica, 8, 10<br />
Mycosphaerella fibrillosa, 10<br />
Mycosphaerella fimbriata, 8, 10<br />
Mycosphaerella flexuosa, 10<br />
Mycosphaerella gamsii, 10<br />
Mycosphaerella graminicola, 3 t , 6 c –7 c , 63 c , 65 c , 162 c<br />
Mycosphaerella gr<strong>and</strong>is, 10<br />
Mycosphaerella gregaria, 63 c , 65 c<br />
“Mycosphaerella” hyperici, 158 t , 163 c –165 c , 173<br />
Mycosphaerella intermedia, 63 c , 65 c<br />
Mycosphaerella irregulariramosa, 36 c –37 c<br />
Mycosphaerella jonkershoekensis, 8, 10<br />
Mycosphaerella juvenis, 10<br />
Mycosphaerella latebrosa, 162 c<br />
“Mycosphaerella” lateralis, 2 t , 6 c –7 c , 28, 36 c –37 c , 63 c , 65 c , 162 c<br />
Mycosphaerella macrospora, 126<br />
Mycosphaerella madeirae, 63 c , 65 c<br />
Mycosphaerella marksii, 36 c –37 c , 63 c , 65 c<br />
Mycosphaerella maxii, 8, 10<br />
Mycosphaerella mexicana, 8, 10<br />
Mycosphaerella microspora, 10<br />
Mycosphaerella molleriana, 10<br />
“Mycosphaerella mycopappi”, 240 c –241, 244<br />
Mycosphaerella nubilosa, 8, 10<br />
Mycosphaerella ohnowa, 10<br />
Mycosphaerella parkii, 63 c , 65 c<br />
Mycosphaerella parkiiaffinis, 10<br />
Mycosphaerella parva, 10<br />
Mycosphaerella perpendicularis, 10<br />
Mycosphaerella pluritubularis, 10<br />
Mycosphaerella pseudafricana, 11<br />
Mycosphaerella pseudocryptica, 8, 11<br />
Mycosphaerella pseudosuberosa, 8, 11<br />
Mycosphaerella punctiformis, 6 c –7 c , 28, 36 c –37 c , 63 c , 65 c , 97 c ,<br />
166<br />
249
Mycosphaerella quasicercospora, 11<br />
Mycosphaerella readeriellophora, 11<br />
Mycosphaerella secundaria, 11<br />
Mycosphaerella stigminaplatani, 30<br />
Mycosphaerella stramenticola, 11<br />
Mycosphaerella suberosa, 8, 11<br />
Mycosphaerella suttonii, 11<br />
Mycosphaerella tassiana, 122, 166<br />
Mycosphaerella toledana, 11<br />
Mycosphaerella tulasnei, 132<br />
Mycosphaerella vespa, 10<br />
Mycosphaerella walkeri, 63 c , 65 c<br />
Mycovellosiella, 31, 53 k<br />
Myrmecridium, 61 k , 84, 92<br />
Myrmecridium flexuosum, 59 t , 62 c , 64 c , 84–85, 86 *<br />
Myrmecridium schulzeri, 59 t , 62 c , 64 c , 68 * , 84–85 *<br />
Myrmecridium schulzeri var. schulzeri, 84<br />
Myrmecridium schulzeri var. tritici, 84<br />
Myxotrichum, 242<br />
Myxotrichum arcticum, 242 c<br />
Myxotrichum cancellatum, 242 c<br />
Myxotrichum carminoparum, 242 c<br />
Myxotrichum chartarum, 242 c<br />
Myxotrichum deflexum, 36 c –37 c , 242 c<br />
Myxotrichum resinae, 239<br />
Myxotrichum stipitatum, 242 c<br />
Napicladium ossifragi, 133, 135<br />
Neofabraea alba, 36 c –37 c , 240 c , 242 c<br />
Neofabraea krawtzewii, 242 c<br />
Neofabraea malicorticis, 36 c –37 c , 189 c , 193 c , 242 c<br />
Neofabraea perennans, 242 c<br />
Neoovularia, 87<br />
Nothostrasseria, 12 k<br />
Nothostrasseria dendritica, 2 t , 6 c –7 c , 10<br />
Ochrocladosporium, 46, 54 k –55<br />
Ochrocladosporium elatum, 34 t , 36 c –37 c , 46, 48 * –49<br />
Ochrocladosporium frigidarii, 34 t , 36 c –37 c , 47–49 *<br />
Oidiodendron, 241–242, 244<br />
Oidiodendron cerealis, 242 c<br />
Oidiodendron chlamydosporicum, 242 c<br />
Oidiodendron citrinum, 242 c<br />
Oidiodendron flavum, 242 c<br />
Oidiodendron griseum, 242 c<br />
Oidiodendron maius, 242 c<br />
Oidiodendron myxotrichoides, 242 c<br />
Oidiodendron periconioides, 242 c<br />
Oidiodendron pilicola, 242 c<br />
Oidiodendron rhodogenum, 242 c<br />
Oidiodendron setiferum, 242 c<br />
Oidiodendron tenuissimum, 242 c<br />
Oidiodendron truncatum, 242 c<br />
Ophiostoma stenoceras, 62 c , 64 c<br />
Paracercospora, 31<br />
Paracoccidioides loboi, 167<br />
Parahaplotrichum, 54 k<br />
Parapericoniella, 53 k , 55, 103<br />
Parapleurotheciopsis, 51, 53 k –54 k , 204<br />
Parapleurotheciopsis coccolobae, 51, 54 k<br />
Parapleurotheciopsis inaequiseptata, 34 t –35, 37 c , 51<br />
Passalora, 28, 31, 44–45, 53 k , 216<br />
Passalora arachidicola , 97 c<br />
Passalora daleae, 34 t , 36 c –37 c<br />
Passalora dodonaeae, 162 c<br />
Passalora eucalypti, 36 c –37 c<br />
Passalora fulva, 36 c –37 c , 55, 97 c , 162 c<br />
Passalora perplexa, 30<br />
“Passalora” zambiae, 2 t , 6 c –7 c , 28<br />
Paullicorticium ansatum, 6 c –7 c , 36 c –37 c , 58, 62 c , 64 c , 188 c , 193 c<br />
Penicillium, 237<br />
Penicillium olivaceum, 237<br />
Penidiella, 12 k , 17–18, 21–22, 24, 53 k<br />
Penidiella columbiana, 2 t , 6 c –7 c , 17–18 k , 19 * –20 * , 36 c –37 c<br />
Penidiella cubensis, 18 k , 19–21 *<br />
Penidiella nect<strong>and</strong>rae, 3 t , 18 k , 20–21 * , 36 c –37 c<br />
Penidiella rigidophora, 3 t , 18 k , 21, 22 * –23 * , 36 c –37 c<br />
Penidiella strumelloidea, 3 t , 18 k , 23, 23 * –24 * , 36 c –37 c , 53 k<br />
Penidiella venezuelensis, 3 t , 18 k , 22, 24–25 * , 36 c –37 c<br />
Periconia chlorocephala, 95–96, 101, 103<br />
Periconia ellipsospora, 96<br />
Periconia velutina, 64<br />
Periconiella, 17, 53 k , 57–58, 61 k –62, 63, 91<br />
Periconiella arcuata, 59 t , 63 c , 65 c –66 * , 68 *<br />
Periconiella levispora, 59 t , 63 c , 65 c , 67 * –68<br />
Periconiella musae, 57, 72<br />
Periconiella papuana, 60, 80<br />
Periconiella velutina, 17, 57, 59 t –63 c , 64–65 c , 66 *<br />
Pertusaria mammosa, 240 c<br />
Phaeoblastophora, 52 k<br />
Phaeococcomyces catenatus, 188 c , 193 c , 243 c<br />
Phaeococcomyces chersonesos, 243 c<br />
Phaeococcomyces nigricans, 241, 243 c<br />
Phaeoisariopsis, 4, 31<br />
Phaeophleospora, 4, 24<br />
Phaeophleospora destructans, 26<br />
Phaeophleospora epicoccoides, 11<br />
Phaeophleospora eucalypti, 26<br />
Phaeophleospora toledana, 11, 25<br />
Phaeoramularia, 31, 53 k , 227<br />
“Phaeoramularia” hachijoensis, 43, 185, 214<br />
Phaeoseptoria eucalypti, 11<br />
Phaeoseptoria luzonensis, 11<br />
Phaeosphaeria avenaria, 36 c –37 c<br />
Phaeotheca triangularis, 3 t , 6 c –7 c<br />
Phaeothecoidea, 11 k<br />
Phaeothecoidea eucalypti, 3 t , 6 c –7 c<br />
Phaeotrichum benjaminii, 240 c<br />
Phialophora, 231<br />
Phialophora americana, 188 c , 193 c , 240 c<br />
Phialophora verrucosa, 243 c<br />
Phlogicylindrium, 216<br />
Phlogicylindrium eucalypti, 37 c , 188 c , 193 c , 214<br />
Phoma herbarum, 36 c –37 c<br />
Phyllactinia guttata, 132<br />
Phyllactinia sp., 110<br />
Piceomphale bulgarioides, 242 c<br />
Pidoplitchkoviella terricola, 188 c , 193 c<br />
Pilidiella, 25<br />
Plectosphaera eucalypti, 37 c<br />
Pleomassaria siparia, 240 c<br />
Pleospora herbarum, 240 c<br />
Pleuroascus nicholsonii, 240 c<br />
Pleurophoma sp., 3 t<br />
Pleurophragmium acutum, 84<br />
Pleurophragmium tritici, 84<br />
Pleurotheciopsis, 18, 51<br />
250
Ramichloridium epichloës, 60, 89<br />
Pleurothecium, 61 k , 83, 92<br />
243 c Rhinocladiella sp., 60 t , 62 c , 64 c<br />
Pleurothecium obovoideum, 59 t , 62 c , 64 c , 80, 83 * , 92<br />
Pollaccia, 185, 205<br />
Pollaccia m<strong>and</strong>shurica, 210<br />
Pollaccia sinensis, 210<br />
Polyscytalum, 51 k , 55 k , 185, 200, 214, 216<br />
Polyscytalum fecundissimum, 188 c , 191 t , 193 c , 200, 214 *<br />
Polyscytalum griseum, 200<br />
Prathigada, 31<br />
Protoventuria alpina, 186, 189 c , 191 t , 193 c<br />
Pseudeurotium bakeri, 242 c<br />
Pseudeurotium desertorum, 242 c<br />
Pseudeurotium ovale var. milkoi, 242 c<br />
Pseudeurotium ovale var. ovale, 242 c<br />
Pseudeurotium zonatum, 242 c<br />
Pseudocercospora, 4, 12, 30–31, 164<br />
Pseudocercospora angolensis, 162 c<br />
Pseudocercospora epispermogonia, 63 c , 65 c<br />
Pseudocercospora paraguayensis, 36 c –37 c<br />
Pseudocercospora protearum, 162 c<br />
Pseudocercosporidium, 31<br />
Ramichloridium fasciculatum, 60, 75, 77<br />
Ramichloridium indicum, 59 t –61, 63 c , 65 c , 73 *<br />
Ramichloridium mackenziei, 58, 59 t –60 t , 75, 80, 83, 188 c , 193 c ,<br />
243 c<br />
Ramichloridium musae, 57–59 t , 60–61, 63 c , 65 c , 70 * , 72–73, 91<br />
Ramichloridium obovoideum, 60, 83<br />
Ramichloridium pini, 58–59 t , 60–61, 63 c , 65 c , 74<br />
Ramichloridium schulzeri, 58, 84<br />
Ramichloridium schulzeri var. flexuosum, 60, 84<br />
Ramichloridium schulzeri var. schulzeri, 60<br />
Ramichloridium schulzeri var. tritici, 84<br />
Ramichloridium strelitziae, 59 t , 63 c , 65 c , 73 * –74 *<br />
Ramichloridium subulatum, 60, 89<br />
Ramularia, 51 k , 164<br />
Ramularia aplospora, 34 t , 36 c –37 c<br />
Ramularia endophylla, 28<br />
Ramularia miae, 6 c –7 c , 63 c , 65 c<br />
Ramularia pratensis var. pratensis, 3 t , 6 c –7 c , 63 c , 65 c<br />
Ramularia sp., 3 t , 6 c –7 c , 63 c , 65 c<br />
Ramularia torvi, 85<br />
Pseudocladosporium, 17, 43, 52 k , 54 k –55, 185, 186, 188–189, Ramulispora, 4, 31<br />
197–198, 204–205, 216, 227, 230<br />
Pseudocladosporium brevicatenatum, 205, 212<br />
Pseudocladosporium caruanianum, 208<br />
Pseudocladosporium hachijoense, 43, 212<br />
Pseudocladosporium matsushimae, 209<br />
Pseudocladosporium proteae, 198<br />
Pseudocyphellaria perpetua, 240 c<br />
Pseudodidymaria, 87<br />
Pseudomassaria carolinensis, 37 c<br />
Pseudomicrodochium, 201<br />
Pseudomicrodochium aciculare, 201<br />
Pseudotaeniolina, 11 k –12, 24<br />
Pseudotaeniolina convolvuli, 24<br />
Pseudotaeniolina globosa, 3 t , 6 c –7 c , 24<br />
Pseudovirgaria, 61 k , 87, 92<br />
Pseudovirgaria hyperparasitica, 59 t , 63 c , 65 c , 67 * , 86 * –87<br />
Psilobotrys schulzeri, 84<br />
Pycnostysanus, 238, 244<br />
Pycnostysanus azaleae, 238<br />
Pycnostysanus resinae, 237–238 t , 239<br />
Rachicladosporium, 38–39, 52 k , 55<br />
Rachicladosporium luculiae, 34 t , 36 c –37 c , 38 * –39<br />
Racodium, 75<br />
Racodium cellare, 75<br />
Racodium resinae, 235, 237–238 t , 239<br />
Racodium resinae (β) piceum, 236<br />
Racodium rupestre, 75<br />
Radulidium, 61 k , 89, 92<br />
Radulidium epichloës, 59 t , 63 c , 65 c , 87–88 * , 89<br />
Radulidium sp., 59 t , 63 c , 65 c<br />
Radulidium subulatum, 59 t , 63 c , 65 c , 70 * , 87–88 * , 89<br />
Ramichloridium, 17, 57–58, 60–61 k , 68, 75, 84, 89, 91–92<br />
Ramichloridium anceps, 60, 75, 188 c , 193 c , 243 c<br />
Ramichloridium apiculatum, 17, 57, 59 t –61, 63 c , 65 c , 68–69 * , 73<br />
Ramichloridium australiense, 59 t , 63 c , 65 c , 69 * –70 *<br />
Ramichloridium basitonum, 58, 60, 75, 77<br />
Ramichloridium biverticillatum, 59 t , 61, 63 c , 65 c , 71 * –72, 91<br />
Ramichloridium brasilianum, 59 t , 63 c , 65 c , 70 * –71 * , 72<br />
Ramulispora sorghi, 162 c<br />
Rasutoria pseudotsugae, 63 c , 65 c<br />
Rasutoria tsugae, 63 c , 65 c<br />
Readeriella, 11 k –12 k , 17, 24–25, 30<br />
Readeriella blakelyi, 6 c –7 c , 26<br />
Readeriella brunneotingens, 3 t , 6 c –7 c , 26–27 *<br />
Readeriella considenianae, 6 c –7 c , 26, 63 c , 65 c<br />
Readeriella destructans, 3 t , 6 c –7 c , 26<br />
Readeriella dimorpha, 6 c –7 c , 26<br />
Readeriella epicoccoides, 6 c –7 c , 11, 30<br />
Readeriella eucalypti, 3 t , 6 c –7 c , 26<br />
Readeriella gauchensis, 3 t , 6 c –7 c , 26<br />
Readeriella mirabilis, 3 t , 6 c –7 c , 24–25, 27 * , 30, 97 c<br />
Readeriella molleriana, 3 t , 10<br />
Readeriella novaezel<strong>and</strong>iae, 6 c –7 c , 97 c<br />
Readeriella nubilosa, 10<br />
Readeriella ovata, 3 t , 6 c –7 c<br />
Readeriella pulcherrima, 6 c –7 c , 26<br />
Readeriella readeriellophora, 11, 25–27 *<br />
Readeriella stellenboschiana, 3 t , 6 c –7 c , 26<br />
Readeriella toledana, 11<br />
Readeriella zuluensis, 3 t , 6 c –7 c , 26<br />
Repetophragma goidanichii, 62 c , 64 c<br />
Retroconis, 46<br />
Retroconis fusiformis, 34 t , 36 c –37 c , 46<br />
Rhinocladiella, 57–58, 60–61 k , 75–76, 92<br />
Rhinocladiella anceps, 59 t , 62 c , 64 c , 75 * –76<br />
Rhinocladiella apiculata, 68<br />
Rhinocladiella atrovirens, 62 c , 64 c , 75–76, 243 c<br />
Rhinocladiella basitona, 59 t , 62 c , 64 c , 76 * –77, 243 c<br />
Rhinocladiella cellaris, 75<br />
Rhinocladiella elatior, 89<br />
Rhinocladiella fasciculata, 59 t , 62 c , 64 c , 77 *<br />
Rhinocladiella indica, 68, 73<br />
Rhinocladiella mackenziei, 62 c , 64 c , 78 * , 80, 83, 92<br />
Rhinocladiella obovoidea, 83<br />
Rhinocladiella phaeophora, 60 t , 62 c , 64 c , 76<br />
Rhinocladiella schulzeri, 84<br />
Ramichloridium cerophilum, 59 t –61, 63 c , 65 c , 71 * –72, 73, 241, Rhinocladiella similis, 243 c<br />
251
Rhinotrichum multisporum, 84<br />
Rhizocladosporium, 50–51, 54 k –55, 204<br />
Rhizocladosporium argillaceum, 34 t , 36 c –37 c , 50 * –51<br />
Rhodoveronaea, 61 k , 89–90, 92<br />
Rhodoveronaea varioseptata, 60 t , 62 c , 64 c , 70 * , 90–91<br />
Rhytisma acerinum, 240 c<br />
Saccharomyces cerevisiae, 240 c<br />
Sarcoleotia turficola, 36 c –37 c<br />
Satchmopsis brasiliensis, 189 c , 193 c<br />
Schizothyrium, 5 k , 28, 30<br />
Schizothyrium acerinum, 28<br />
Schizothyrium pomi, 6 c –7 c<br />
Scolicotrichum iridis, 125<br />
Scorias spongiosa, 6 c –7 c<br />
Seifertia, 52 k , 238, 244<br />
Seifertia azaleae, 235, 238–239, 240 c , 241–242, 244<br />
Septocylindrium chaetospira, 187<br />
Septonema, 54 k<br />
Septonema chaetospira, 187<br />
Septoria, 164, 173<br />
Septoria pulcherrima, 26<br />
Septoria tritici, 3 t , 6 c –7 c , 63 c , 65 c<br />
Setosphaeria monoceras, 240 c<br />
Sonderhenia, 24<br />
Sorocybe, 52 k , 54 k , 236–238, 242–244<br />
Sorocybe resinae, 235, 237 * –238 t , 239–240 c , 241–243 c , 244<br />
Spadicoides minuta, 187<br />
Spathularia flavida, 240 c<br />
Sphaerella allicina, 118<br />
Sphaerella cryptica, 10<br />
Sphaerella molleriana, 10<br />
Sphaerella nubilosa, 10<br />
Sphaerella tassiana, 8, 122<br />
Sphaeria allicina, 118<br />
Spilocaea, 185, 205<br />
Spilomyces dendriticus, 10<br />
Sporidesmium pachyanthicola, 63 c , 65 c<br />
Sporocybe, 236<br />
Sporocybe resinae, 237–238 t , 239<br />
Sporotrichum anceps, 76<br />
Sporotrichum nigrum, 238 t<br />
Stagonospora pulcherrima, 26<br />
Staninwardia, 12 k , 26<br />
Staninwardia breviuscula, 26<br />
Staninwardia suttonii, 5, 6 c –7 c , 26, 36 c –37 c , 63 c , 65 c<br />
Stenella, 31, 44–45, 53 k , 75, 227<br />
Stenella araguata, 16, 19, 24, 43, 44 * –45 * , 75<br />
“Stenella” cerophilum, 36 c –37 c<br />
Stenellopsis, 31<br />
Stigmidium, 30<br />
Stigmidium schaereri, 30 *<br />
Stigmina, 4, 12, 30–31<br />
Stigmina eucalypti, 12<br />
Stigmina platani, 12<br />
Strigopodia resinae, 236<br />
Stysanopsis resinae, 238 t<br />
Stysanus resinae, 237–238 t , 239<br />
Subramaniomyces, 51, 54 k , 204<br />
Subramaniomyces fusisaprophyticus, 34 t –35, 37 c , 51<br />
Subramaniomyces simplex, 51<br />
Sympodiella, 204<br />
Sympoventuria, 185, 204<br />
Sympoventuria capensis, 189 c , 191 t , 194 c , 205<br />
Taeniolella, 41, 52 k<br />
Taeniolina, 52 k<br />
Taeniolina scripta, 17<br />
Teratosphaeria, 5 k , 8, 28, 30–31, 43, 45, 92, 96<br />
Teratosphaeria africana, 8<br />
Teratosphaeria alistairii, 6 c –7 c , 9, 36 c –37 c , 63 c , 65 c<br />
Teratosphaeria associata, 9<br />
Teratosphaeria bellula, 3 t , 6 c –7 c , 10<br />
Teratosphaeria cryptica, 6 c –7 c , 10, 36 c –37 c<br />
Teratosphaeria dendritica, 2 t , 10<br />
Teratosphaeria excentrica, 10<br />
Teratosphaeria fibrillosa, 3 t –4, 6 c –7 c , 8–9 * , 10, 63 c , 65 c<br />
Teratosphaeria fimbriata, 10<br />
Teratosphaeria flexuosa, 6 c –7 c , 10<br />
Teratosphaeria gamsii, 10<br />
Teratosphaeria jonkershoekensis, 10<br />
Teratosphaeria juvenis, 36 c –37 c<br />
Teratosphaeria maxii, 6 c –7 c , 10<br />
Teratosphaeria mexicana, 3 t , 6 c –7 c , 10<br />
Teratosphaeria microspora, 2 t , 8, 10, 15 k , 97 c<br />
Teratosphaeria molleriana, 3 t , 6 c –7 c , 10, 36 c –37 c , 63 c , 65 c<br />
Teratosphaeria nubilosa, 3 t , 6 c –7 c , 10, 162 c<br />
Teratosphaeria ohnowa, 4 t , 6 c –7 c , 10<br />
Teratosphaeria parkiiaffinis, 10<br />
Teratosphaeria parva, 6 c –7 c , 10, 63 c , 65 c<br />
Teratosphaeria perpendicularis, 10<br />
Teratosphaeria pluritubularis, 10<br />
Teratosphaeria proteaarboreae, 8<br />
Teratosphaeria pseudafricana, 11<br />
Teratosphaeria pseudocryptica, 11<br />
Teratosphaeria pseudosuberosa, 2 t , 6 c –7 c , 11<br />
Teratosphaeria quasicercospora, 11<br />
Teratosphaeria readeriellophora, 6 c –7 c , 11, 63 c , 65 c , 97 c<br />
Teratosphaeria secundaria, 4 t , 6 c –7 c , 11<br />
Teratosphaeria sp., 2 t , 4 t , 6 c –7 c<br />
Teratosphaeria stramenticola, 11<br />
Teratosphaeria suberosa, 6 c –7 c , 11, 63 c , 65 c<br />
Teratosphaeria suttonii, 3 t , 6 c –7 c , 11, 36 c –37 c<br />
Teratosphaeria toledana, 6 c –7 c , 11, 63 c , 65 c<br />
Terfezia gigantea, 240 c<br />
<strong>The</strong>dgonia ligustrina, 34 t , 36 c –37 c<br />
Thielaviopsis basicola, 159 t<br />
Thyridium, 84<br />
Thyridium vestitum, 60, 62 c , 64 c<br />
Thysanorea, 61 k , 63, 80, 92<br />
Thysanorea papuana, 60 t , 62 c , 64 c , 68 * , 78 * –79 * , 80, 188 c , 193 c<br />
Torrendiella eucalypti, 36 c –37 c<br />
Torula pinophila, 239<br />
Toxicocladosporium, 39, 53 k , 55<br />
Toxicocladosporium irritans, 34 t , 36 c –37 c , 39–40 * , 41<br />
Trematosphaeria heterospora, 240 c<br />
Trichosphaeria pilosa, 6 c –7 c<br />
Trimmatostroma, 5, 14, 164<br />
Trimmatostroma abietina, 14–15, 97 c<br />
Trimmatostroma abietis, 14–15, 97 c<br />
Trimmatostroma betulinum, 3 t , 5, 6 c –7 c<br />
Trimmatostroma elginense, 16<br />
Trimmatostroma excentricum, 10<br />
Trimmatostroma macowanii, 17<br />
Trimmatostroma microsporum, 10<br />
Trimmatostroma protearum, 17<br />
252
Trimmatostroma salicis, 3 t , 5, 6 c –7 c , 13 * , 97 c<br />
Uwebraunia, 28<br />
Venturia, 43, 52 k , 55, 185–186, 204–205, 216<br />
Venturia aceris, 189 c , 191 t , 194 c<br />
Venturia alpina, 189 c , 191 t , 194 c<br />
Venturia anemones, 189 c , 191 t , 194 c<br />
Venturia asperata, 62 c , 64 c<br />
Venturia atriseda, 189 c , 191 t , 194 c<br />
Venturia aucupariae, 189 c , 191 t , 194 c<br />
Venturia carpophila, 62 c , 64 c , 189 c –190 t , 194 c<br />
Venturia cephalariae, 189 c , 191 t , 194 c<br />
Venturia cerasi, 189 c , 191 t , 194 c<br />
Venturia chlorospora, 62 c , 64 c , 189 c , 191 t , 194 c<br />
Venturia crataegi, 189 c , 191 t , 194 c<br />
Venturia dickiei, 205<br />
Venturia ditricha, 189 c , 191 t , 194 c<br />
Venturia fraxini, 189 c , 192 t , 194 c<br />
Venturia hanliniana, 62 c , 64 c , 205, 212<br />
Venturia helvetica, 189 c , 192 t , 194 c<br />
Venturia hystrioides, 189 c , 192 t , 194 c , 212–213 *<br />
Venturia inaequalis, 62 c , 64 c , 189 c , 191 t , 194 c<br />
Venturia lonicerae, 189 c , 192 t , 194 c<br />
Venturia macularis, 189 c , 192 t , 194 c<br />
Venturia maculiformis, 189 c , 192 t , 194 c<br />
Venturia m<strong>and</strong>shurica, 191 t , 210<br />
Venturia minuta, 189 c , 192 t , 194 c<br />
Venturia nashicola, 189 c , 192 t , 194 c<br />
Venturia polygonivivipari, 189 c , 192 t , 194 c<br />
Venturia populina, 189 c , 192 t , 194 c<br />
Venturia pyrina, 62 c , 64 c , 189 c , 192 t , 194 c<br />
Venturia saliciperda, 189 c , 192 t , 194 c<br />
Venturia sp., 189 c , 192 t , 194 c<br />
Venturia tremulae, 191 t<br />
Venturia tremulae var. gr<strong>and</strong>identatae, 189 c , 192 t , 194 c<br />
Venturia tremulae var. populialbae, 189 c , 192 t , 194 c<br />
Venturia tremulae var. tremulae, 189 c , 192 t , 194 c<br />
Venturia viennotii, 189 c , 192 t , 194 c<br />
Veronaea, 57–58, 60–61 k , 76, 80, 87, 89, 91, 92<br />
Veronaea apiculata, 68<br />
Veronaea botryosa, 60 t , 62 c , 64 c , 76, 80–81 * , 188 c , 193 c<br />
Veronaea compacta, 60 t , 62 c , 64 c , 81–82 * , 83<br />
Veronaea harunganae, 92<br />
Veronaea indica, 73<br />
Veronaea japonica, 60 t , 62 c , 64 c , 74 * , 82 *<br />
Veronaea musae, 57, 72<br />
Veronaea parvispora, 76<br />
Veronaea simplex, 60, 91<br />
Veronaea verrucosa, 73<br />
Veronaeopsis, 61 k , 91–92<br />
Veronaeopsis simplex, 60 t , 62 c , 64 c , 74 * , 90 * –91, 189 c , 194 c<br />
Verrucisporota, 31, 45<br />
Verrucocladosporium, 41, 53 k , 55<br />
Verrucocladosporium dirinae, 34 t , 36 c –37 c , 41–42 *<br />
Vibrissea flavovirens, 6 c –7 c<br />
Vibrissea truncorum, 6 c –7 c<br />
Virgaria, 87<br />
Websteromyces, 52 k<br />
Wentiomyces, 91<br />
Wentiomyces javanicus, 91<br />
Westerdykella cylindrica, 240 c<br />
Xenomeris juniperi, 63 c , 65 c<br />
Xylographa vitiligo, 240 c<br />
Xylohypha, 52 k<br />
Zasmidium, 45, 61, 74–75<br />
Zasmidium cellare, 60 t –61, 63 c , 65 c , 74 * –75<br />
Zeloasperisporium, 214, 216<br />
Zeloasperisporium hyphopodioides, 189 c , 191 t , 194 c , 214–215 *<br />
Zygosporium, 5 k<br />
253