Fungal Diversity
https://doi.org/10.1007/s13225-020-00444-8
Taxonomic and phylogenetic contributions to fungi associated
with the invasive weed Chromolaena odorata (Siam weed)
Ausana Mapook1,2,3,4 · Kevin D. Hyde1,3,5 · Eric H. C. McKenzie6 · E. B. Gareth Jones7 · D. Jayarama Bhat8 ·
Rajesh Jeewon9 · Marc Stadler4 · Milan C. Samarakoon3 · Maitree Malaithong2 · Benjawan Tanunchai2 ·
François Buscot2,11 · Tesfaye Wubet2,10,11 · Witoon Purahong2
Received: 1 February 2020 / Accepted: 19 March 2020
© MUSHROOM RESEARCH FOUNDATION 2020
Abstract
This article provides morphological descriptions and illustrations of microfungi associated with the invasive weed, Chromolaena odorata, which were mainly collected in northern Thailand. Seventy-seven taxa distributed in ten orders, 23
families (of which Neomassarinaceae is new), 12 new genera (Chromolaenicola, Chromolaenomyces, Longiappendispora,
Pseudocapulatispora, Murichromolaenicola, Neoophiobolus, Paraleptospora, Pseudoroussoella, Pseudostaurosphaeria,
Pseudothyridariella, Setoarthopyrenia, Xenoroussoella), 47 new species (Aplosporella chromolaenae, Arthrinium chromolaenae, Chromolaenicola chiangraiensis, C. lampangensis, C. nanensis, C. thailandensis, Chromolaenomyces appendiculatus,
Diaporthe chromolaenae, Didymella chromolaenae, Dyfrolomyces chromolaenae, Leptospora chromolaenae, L. phraeana,
Longiappendispora chromolaenae, Memnoniella chromolaenae, Montagnula chiangraiensis, M. chromolaenae, M. chromolaenicola, M. thailandica, Murichromolaenicola chiangraiensis, M. chromolaenae, Muyocopron chromolaenae, M. chromolaenicola, Neomassarina chromolaenae, Neoophiobolus chromolaenae, Neopyrenochaeta chiangraiensis, N. chromolaenae,
N. thailandica, N. triseptatispora, Nigrograna chromolaenae, Nothophoma chromolaenae, Paraleptospora chromolaenae,
P. chromolaenicola, Patellaria chromolaenae, Pseudocapulatispora longiappendiculata, Pseudoroussoella chromolaenae,
Pseudostaurosphaeria chromolaenae, P. chromolaenicola, Pseudothyridariella chromolaenae, Pyrenochaetopsis chromolaenae, Rhytidhysteron chromolaenae, Setoarthopyrenia chromolaenae, Sphaeropsis chromolaenicola, Tremateia chiangraiensis,
T. chromolaenae, T. thailandensis, Xenoroussoella triseptata, Yunnanensis chromolaenae), 12 new host records, three new
taxonomic combinations (Chromolaenicola siamensis, Pseudoroussoella elaeicola, Pseudothyridariella mahakashae), and
two reference specimens (Torula chromolaenae, T. fici) are described and illustrated. Unlike some other hosts, e.g. bamboo
(Poaceae) and Pandanaceae, the dominant group of fungi on Siam weed were Dothideomycetes. Only 15 species previously
recorded from northern Thailand were found in this study. Most of the taxa are likely to have jumped hosts from surrounding
plants and are unlikely to be a specialist to Siam weed. Most fungal families found on Siam weed had divergence estimates
with stem ages in the Cretaceous, which coincided with the expected origin of the host family (Asteraceae). This further
indicates that the species have jumped hosts, as it is unlikely that the taxa on the alien Siam weed came from the Americas
with its host. They may, however, have jumped from other Asteraceae hosts. In a preliminary screening 40 (65%) of the
62 species tested showed antimicrobial activity and thus, the fungi associated with C. odorata may be promising sources
of novel bioactive compound discovery. We provide a checklist of fungi associated with C. odorata based on the USDA
Systematic Mycology and Microbiology Laboratory (SMML) database, relevant literature and our study. In total, 130 taxa
(116 identified and 14 unidentified species) are distributed in 20 orders, 48 families and 85 genera. Pseudocercospora is the
most commonly encountered genus on Siam weed.
Keywords 60 new taxa · Antimicrobial activity · Ascomycota · Checklist · Dothideomycetes · Evolutionary divergence
times · Multi-gene phylogenetics · Sordariomycetes
* Kevin D. Hyde
kdhyde3@gmail.com
Extended author information available on the last page of the article
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Fungal Diversity
Introduction
Invasive plants potentially affect ecosystem processes
including decomposition and nutrient cycling as they may
be more readily decomposed than native species and have
a higher nitrogen concentration and lower carbon to nitrogen ratio (Liao et al. 2008; Bassett et al. 2010). They may
also influence micro-environmental conditions and stimulate the activity of microbial decomposers (Březina et al.
2006; Heneghan et al. 2006; Tůma et al. 2009; Esperschütz
et al. 2011, 2013). Chromolaena odorata (L.) King & Robinson is a serious invasive weed in the old-world tropics and
subtropics (Holm et al. 1977; Gautier 1992; den Breeÿen
et al. 2006). It is known by several common names including “Siam weed” and belongs to the flowering plant family
Asteraceae (Jeffrey 2007).
Siam weed is native to the Americas, where it occurs
from southern Florida and Texas in the USA to northern
Argentina (Gautier 1992). This important invasive weed
has spread to tropical regions of Africa, Asia, and Oceania,
impacting economically, ecologically and environmentally
on forests (Holm et al. 1977; Moore 2004; Muniappan et al.
2005; Zachariades et al. 2013; Catarino et al. 2019). Two
biotypes of Siam weed have been recognized, the South
African biotype (SA), and the Asian/West African biotype
(AWA). Based on comparison of both haplotype and multilocus genotypes of native and invasive populations, Trinidad and Tobago was proposed as the source location of the
Siam weed Asian biotype (Yu et al. 2014), and the invasive
species shows higher competitive ability under nutrient-rich
conditions than in its native regions (Qin et al. 2013; Shao
et al. 2018). The invasive weed also affects conservation and
ecotourism, as it has spread to large areas of agricultural
land, causing reduced cropland and crop yields, and reduced
plant native species richness (Macdonald 1983; Cronk and
Fuller 1995; Goodall and Erasmus 1996; Rose 1997; Matthews and Brand 2004). Siam weed has, however, also been
considered as a nutrient sink with potential benefit to crops,
as a source of organic matter and nutrients (Koutika and
Rainey 2010). The weed can adapt to acidic soils better than
some leguminous plants (Koutika and Rainey 2010) and
has been identified as a hyper-accumulator of heavy metals, especially cadmium and zinc. These two harmful heavy
metals play an important role in ecosystems, affecting the
activity of soil microbial communities (Giller et al. 1998;
Yao et al. 2000; Tanhan et al. 2007; Okoronkwo et al. 2014;
Ruangdech et al. 2017).
Siam weed also has numerous biological activities,
such as antiviral (Pisutthanan et al. 2005; Vital and Rivera
2009; Atindehou et al. 2013; Stanley et al. 2014; Hanphakphoom and Krajangsang 2016), antibacterial (Suksamrarn
et al. 2004; Johari et al. 2012; Kigigha and Zige 2013),
13
antifungal (Naidoo et al. 2011; Ngono et al. 2006; Vijayaraghavan et al. 2013), anthelmintic and antiprotozoal (Vital
and Rivera 2009), antibiofilm (Yahya et al. 2014), antihepatotoxic (Asomugha et al. 2014), antioxidant (Akinmoladun
et al. 2007; Boudjeko et al. 2015), anticancer (Kouamé et al.
2013; Adedapo et al. 2016), anticonvulsant or antiepileptic
(Amazu et al. 2013; Kanase and Shaikh 2018), antidiabetic
(Onkaramurthy et al. 2013; Aba et al. 2015) anti-diarrheal
(Aba et al. 2015), anti-inflammatory (Pandith et al. 2013b;
Hanh et al. 2011) and antiparasitic (Ezenyi et al. 2014).The
weed is widely used as a folk medicine for wound healing
treatments (Vaisakh and Pandey 2012; Pandith et al. 2013a;
Sirinthipaporn and Jiraungkoorskul 2017) in India (Ayyanar and Ignacimuthu 2009; Thomas et al. 2014), Pakistan
(Abbasi et al. 2010), Bangladesh (Rani and Mohammed
2012), and Ghana (Barku et al. 2014). Microbial endophytes
and other microorganisms associated with Siam weed have
also been reported for their biological properties. Buzugbe
et al. (2018) found that an unidentified endophytic fungus,
isolated from leaves of Siam weed, produced secondary
metabolites with antimicrobial and antioxidant properties.
Interestingly, microorganisms associated with weeds and
medicinal plants are beneficial resources for increasing
resistance of the host plants from unfavorable conditions
and enhancing the growth of the host plant through their
interactions (Köberl et al. 2013; Jia et al. 2016; Trognitz
et al. 2016; Huang et al. 2018). For example, Trichoderma
species, which are commonly soil inhabitants and can also
be saprotrophs, mycoparasites, endophytes, as well as human
pathogens (Walsh et al. 2004; Samuels 2006; Jaklitsch 2009;
Kantarcioğlu et al. 2009; Chaverri and Samuels 2013), and
have been reported to have antimicrobial and antiproliferative activities, as well as plant growth regulator activity (Chaverri and Samuels 2013; Leylaie and Zafari 2018;
Zhang et al. 2018). Numerous Trichoderma species can also
potentially protect their host plant from disease infection
and have been used as biological control agents for plant
protection (Harman and Kubicek 1998; Harman et al. 2004;
Hyde et al. 2019b).
Plant-associated microbes, especially microfungi associated with some selected groups of plants in Thailand and
adjacent areas in China, have been studied to provide a hostfungus database for future studies and increase knowledge
of fungal diversity, as well as new fungal discovery. For
example, Dai et al. (2017) studied microfungi from bamboo, resulting in the discovery of 27 new species. Doilom
et al. (2017) reported 28 fungal species on Tectona grandis,
and Tibpromma et al. (2018) described 65 new species of
microfungi on Pandanaceae. Although 68 fungal species
have been recorded from Siam weed according to Farr and
Rossman (2020; from https ://nt.ars-grin.gov/funga ldata
bases /), few taxa have been reported from Thailand. For
example, Tibpromma et al. (2017) described a new species,
Fungal Diversity
Hermatomyces chromolaenae and Li et al. (2017) described
a new species, Torula chromolaenae with a new host record
for T. fici from Siam weed in northern Thailand. Crous et al.
(2018a) introduced a new genus, Neocochlearomyces, which
was collected from leaves of Siam weed in Thailand. However, the diversity of fungi associated with Siam weed is still
relatively unstudied and is likely to be much higher, as have
been shown for other groups of fungi in northern Thailand
(Hyde et al. 2018a, b).
In this study, we provide morphological descriptions and
illustrations of saprobic fungi associated with Chromolaena
odorata, which were mainly collected in northern Thailand.
Both multigene analyses and morphological comparison
were used to confirm the current taxonomic placements of
these fungi. The difference between fungi on Siam weed and
other hosts will be explored for evidence of host-jumping.
Results of preliminary screening for antimicrobial activity
of fungi on Siam weed is provided. Furthermore, a checklist
of fungi associated with C. odorata is provided based on the
USDA Systematic Mycology and Microbiology Laboratory
(SMML) database (Farr and Rossman 2020), together with
relevant literature, and the author’s results from this study.
Table of contents
The numbers of taxa in this study are organized following
the outline of Ascomycota (Wijayawardene et al. 2018) and
updated from recent relevant literature.
Phylum Ascomycota Caval.-Sm.
Class Dothideomycetes sensu O.E. Erikss. & Winka
Subclass Pleosporomycetidae C.L. Schoch et al.
Hysteriales Lindau
Hysteriaceae Chevall.
1. Rhytidhysteron bruguierae Dayarathne, in Dayarathne et al., Mycosphere 11(1): 20 (2020), new
host record.
2. Rhytidhysteron chromolaenae Mapook & K.D.
Hyde, sp. nov.
Pleosporales Luttrell ex M.E. Barr
Acrocalymmaceae Crous & Trakun.
3. Acrocalymma medicaginis Alcorn & J.A.G. Irwin,
Trans. Br. mycol. Soc. 88(2): 163 (1987), new host
record.
Didymellaceae Gruyter et al.
4. Didymella chromolaenae Mapook & K.D. Hyde,
sp. nov.
5. Nothophoma chromolaenae Mapook & K.D.
Hyde, sp. nov.
Didymosphaeriaceae Munk
6. Chromolaenicola Mapook & K.D. Hyde, gen. nov.
7. Chromolaenicola chiangraiensis Mapook & K.D.
Hyde, sp. nov.
8. Chromolaenicola lampangensis Mapook & K.D.
Hyde, sp. nov.
9. Chromolaenicola nanensis Mapook & K.D. Hyde, sp.
nov.
10. Chromolaenicola siamensis (Jayasiri, E.B.G. Jones
& K.D. Hyde) Mapook & K.D. Hyde, comb. nov.
11. Chromolaenicola thailandensis Mapook & K.D.
Hyde, sp. nov.
12. Montagnula chiangraiensis Mapook & K.D. Hyde,
sp. nov.
13. Montagnula chromolaenae Mapook & K.D. Hyde,
sp. nov.
14. Montagnula chromolaenicola Mapook & K.D.
Hyde, sp. nov.
15. Montagnula thailandica Mapook & K.D. Hyde, sp.
nov.
16. Pseudopithomyces palmicola J.F. Li, Ariyaw. &
K.D. Hyde, in Ariyawansa et al., Fungal Divers. 75:
41 (2015), new host record.
17. Tremateia chiangraiensis Mapook & K.D. Hyde,
sp. nov.
18. Tremateia chromolaenae Mapook & K.D. Hyde,
sp. nov.
19. Tremateia thailandensis Mapook & K.D. Hyde, sp.
nov.
Lophiostomataceae Sacc.
20. Flabellascoma minimum A. Hashim., K. Hiray. &
Kaz. Tanaka, in Hashimoto et al., Stud Mycol. 90:
169 (2018), new host record.
21. Pseudocapulatispora Mapook & K.D. Hyde, gen.
nov.
22. Pseudocapulatispora longiappendiculata Mapook
& K.D. Hyde, sp. nov.
Nigrogranaceae Jaklitsch & Voglmayr
23. Nigrograna chromolaenae Mapook & K.D. Hyde,
sp. nov.
Neomassarinaceae Mapook & K.D. Hyde, fam. nov.
24. Neomassarinaceae Mapook & K.D. Hyde, fam.
nov.
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Fungal Diversity
25. Neomassarina chromolaenae Mapook & K.D.
Hyde, sp. nov.
26. Neomassarina thailandica Phook., Jayasiri & K.D.
Hyde, in Hyde et al., Fungal Divers. 80: 138 (2016),
new host record.
Pyrenochaetopsidaceae Valenzuela-Lopez et al.
47. Pyrenochaetopsis chromolaenae Mapook & K.D.
Hyde, sp. nov.
Roussoellaceae J.K. Liu et al.
Neopyrenochaetaceae Valenzuela-Lopez et al.
27. Neopyrenochaeta chiangraiensis Mapook & K.D.
Hyde, sp. nov.
28. Neopyrenochaeta chromolaenae Mapook & K.D.
Hyde, sp. nov.
29. Neopyrenochaeta thailandica Mapook & K.D.
Hyde, sp. nov.
30. Neopyrenochaeta triseptatispora Mapook & K.D.
Hyde, sp. nov.
Phaeosphaeriaceae M.E. Barr
31. Leptospora chromolaenae Mapook & K.D. Hyde,
sp. nov.
32. Leptospora phraeana Mapook & K.D. Hyde, sp.
nov.
33. Leptospora thailandica Phukhams. & K.D. Hyde,
in Hyde et al., Fungal Divers. 80: 100 (2016), new
host record.
34. Murichromolaenicola Mapook & K.D. Hyde, gen.
nov.
35. Murichromolaenicola chiangraiensis Mapook &
K.D. Hyde, sp. nov.
36. Murichromolaenicola chromolaenae Mapook &
K.D. Hyde, sp. nov.
37. Neoophiobolus Mapook & K.D. Hyde, gen. nov.
38. Neoophiobolus chromolaenae Mapook & K.D.
Hyde, sp. nov.
39. Paraleptospora Mapook & K.D. Hyde, gen. nov.
40. Paraleptospora chromolaenae Mapook & K.D.
Hyde, sp. nov.
41. Paraleptospora chromolaenicola Mapook & K.D.
Hyde, sp. nov.
42. Pseudoophiosphaerella huishuiensis J.F. Zhang,
J.K. Liu & Z.Y. Liu, in Zhang et al., Mycosphere
8(1): 207 (2019), new host record.
43. Pseudostaurosphaeria Mapook & K.D. Hyde, gen.
nov.
44. Pseudostaurosphaeria chromolaenae Mapook &
K.D. Hyde, sp. nov.
45. Pseudostaurosphaeria chromolaenicola Mapook
& K.D. Hyde, sp. nov.
46. Yunnanensis chromolaenae Mapook & K.D. Hyde,
sp. nov.
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48. Pseudoroussoella Mapook & K.D. Hyde, gen. nov.
49. Pseudoroussoella chromolaenae Mapook & K.D.
Hyde, sp. nov.
50. Pseudoroussoella elaeicola (Konta & K.D. Hyde)
Mapook & K.D. Hyde, comb. nov.
51. Setoarthopyrenia Mapook & K.D. Hyde, gen. nov.
52. Setoarthopyrenia chromolaenae Mapook & K.D.
Hyde, sp. nov.
53. Xenoroussoella Mapook & K.D. Hyde, gen. nov.
54. Xenoroussoella triseptata Mapook & K.D. Hyde,
sp. nov.
Thyridariaceae Q. Tian & K.D. Hyde
55. Chromolaenomyces Mapook & K.D. Hyde, gen.
nov.
56. Chromolaenomyces appendiculatus Mapook &
K.D. Hyde, sp. nov.
57. Pseudothyridariella Mapook & K.D. Hyde, gen.
nov.
58. Pseudothyridariella chromolaenae Mapook & K.D.
Hyde, sp. nov.
59. Pseudothyridariella mahakashae (Devadatha, V.V.
Sarma, D.N. Wanas., K.D. Hyde & E.B.G. Jones)
Mapook & K.D. Hyde, comb. nov.
Torulaceae Corda
60. Torula chromolaenae Li, Phook., Mapook & K.D.
Hyde, Mycol. Progr. 16(4): 454 (2017), reference
specimen.
61. Torula fici Crous, IMA Fungus 6 (1): 192 (2015),
reference specimen.
62. Torula polyseptata C.G. Lin & K.D. Hyde, in
Hyde et al., Fungal Divers. 96: 71 (2019), new host
record.
Dothideomycetes orders incertae sedis
Botryosphaeriales C.L. Schoch et al.
Aplosporellaceae Slippers et al.
63. Aplosporella chromolaenae Mapook & K.D. Hyde,
sp. nov.
Fungal Diversity
64. Aplosporella hesperidica Speg., Anal. Soc. cient.
argent. 13(1): 18 (1882), new host record.
Botryosphaeriaceae Theiss. & P. Syd., Ann. Mycol. 16
(1–2): 16 (1918)
65. Dothiorella oblonga F.J.J. Van der Walt, Slippers
& G.J. Marais, in Slippers et al., Persoonia 33: 163
(2014), new host record.
66. Sphaeropsis chromolaenicola Mapook & K.D.
Hyde, sp. nov.
Dyfrolomycetales K.L. Pang et al.
Pleurotremataceae Walt. Watson (= Dyfrolomycetaceae
K.D. Hyde et al.)*
67. Dyfrolomyces chromolaenae Mapook & K.D.
Hyde, sp. nov.
Muyocopronales Mapook et al.
Muyocopronaceae K.D. Hyde
Subclass Xylariomycetidae O.E. Erikss & Winka
Amphisphaeriales D. Hawksw. & O.E. Erikss.*
Apiosporaceae K.D. Hyde et al.
75. Arthrinium chromolaenae Mapook & K.D. Hyde,
sp. nov.
Xylariales Nannf.
Cainiaceae J.C. Krug
76. Longiappendispora Mapook & K.D. Hyde, gen.
nov.
77. Longiappendispora chromolaenae Mapook & K.D.
Hyde, sp. nov.
Materials and methods
Collection, examination, and isolation of fungi
68. Muyocopron chromolaenae Mapook & K.D. Hyde,
sp. nov.
69. Muyocopron chromolaenicola Mapook & K.D.
Hyde, sp. nov.
70. Muyocopron lithocarpi Mapook, Boonmee & K.D.
Hyde, Phytotaxa 265(3): 235 (2016), new host
record
Patellariales D. Hawksw. & O.E. Erikss.
Patellariaceae Corda
71. Patellaria chromolaenae Mapook & K.D. Hyde, sp.
nov.
Class Sordariomycetes O.E. Erikss. & Winka
Subclass Diaporthomycetidae Senan. et al.
Diaporthales Nannf.*
Diaporthaceae Höhn. ex Wehm.
72. Diaporthe chromolaenae Mapook & K.D. Hyde,
sp. nov.
Subclass Hypocreomycetidae O.E. Erikss. & Winka
Hypocreales Lindau
Hypocreaceae De Not.
73. Trichoderma guizhouense Q.R. Li, McKenzie &
Yong Wang bis, in Li et al., Mycol. Progr. 12(2):
170 (2012) [2013], new host record.
Stachybotryaceae Lombard & Crous
74. Memnoniella chromolaenae Mapook & K.D. Hyde,
sp. nov.
Fresh materials were collected from sampling sites in Thailand (Chiang Mai, Chiang Rai, Lampang, Mae Hong Son,
Nan, Phetchaburi and Phrae Provinces) during 2015–2017.
Dead aerial stems of Chromolaena odorata (Siam weed)
were randomly collected from roadsides, abandoned fields
or disturbed forests and examined using a Motic SMZ 168
Series microscope. A Carl Zeiss GmbH stereo microscope
fitted with an AxioCam ERC 5S camera was used for image
capture. Sections of ascomata were made by free hand. Fungal material was mounted in water, lactoglycerol, 5% KOH
and/or Indian ink and photographed with a Nikon ECLIPSE
80i compound microscope fitted with a Canon EOS 550D
digital camera, or a Nikon ECLIPSE Ni compound microscope fitted with a Canon EOS 600D and 750D digital
camera. Measurements of microscopic characters were calculated using Tarosoft Image Frame Work program (IFW)
version 0.97 and all photoplates were made using Adobe
Photoshop CS6 version 13.1.2.
Single spore isolations were obtained following the
methods of Chomnunti et al. (2014). Spores were germinated on malt extract agar (MEA: 33.6 g/l sterile distilled
water, Difco malt extract) and observed within 24 h using a
Motic SMZ 168 Series microscope; germinated spores were
transferred to new MEA media. Pure cultures were used for
molecular studies. All specimens were kept in separate ziplock bags and envelopes with collection details and deposited in the herbaria of Mae Fah Luang University (Herb.
MFLU) and Herbarium of Cryptogams, Kunming Institute
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Fungal Diversity
of Botany Academia Sinica (HKAS), China. Living cultures
are also deposited in collection of Mae Fah Luang University
(MFLUCC). Faces of fungi and Index Fungorum numbers
were obtained as in Jayasiri et al. (2015) and Index Fungorum (2020).
Preliminary screening of antimicrobial activity
Preliminary screening of antimicrobial activity against different test organisms such as Bacillus subtilis (Gram-positive
bacteria, DSM10), Escherichia coli (Gram-negative bacteria, DSM498), and Mucor plumbeus (filamentous fungus,
MUCL 49355) were determined by the zone of inhibition
using agar plug diffusion method (Balouiri et al. 2016). Penicillin and ciproloxacin were used as positive control for antibacterial screening, and nystatin was used as positive control
for antifungal screening (El-Desoukey 2018). Sterile Mueller–Hinton agar media and yeast malt agar media (pH 6.3)
were used for testing antibacterial activity and antifungal
activity, respectively. Microbial suspensions were prepared
by cell counting (6.7 × 105 cells/ml for bacteria and 5 × 104
cells/ml for filamentous fungi) and added to the sterile agar
media prior to its solidification. Fungal mycelium plugs from
our isolates were transferred to a solid media plate and incubated for 24 to 48 h at room temperature. Inhibition zones
for antimicrobial activity was measured and compared with
positive control.
DNA extraction, PCR amplification and sequencing
Genomic DNA was extracted using the ZR Soil Microbe
DNA MiniPrep kit (Zymo Research, Irvine, CA, USA) following the manufacturer’s instructions. DNA quality and
quantity were measured by spectrophotometric quantification with a NanoDrop ND-8000 V1.1.1 spectrophotometer (Thermo Fisher Scientific, Dreieich, Germany). DNA
amplifications were performed by polymerase chain reaction (PCR) to amplify partial gene regions using primers
shown in Table 1. PCR amplification and sequencing were
carried out following Mapook et al. (2016a). The PCR products were sequenced with primers mentioned in Table 1 at
SeqLab GmbH (Microsynth AG), Göttingen, Germany.
GenBank based on recent publications. Forward and reverse
sequences were assembled using ContigExpress (a component of Vector NTI Suite 6.0). Single gene sequences were
aligned using MAFFT v.7 online program (https ://mafft
.cbrc.jp/alignment/server/) (Katoh et al. 2019). The alignments were checked and uninformative gaps minimized
manually where necessary in MEGA6 (Tamura et al. 2013).
The align sequence datasets were combined using MEGA5
and MEGA6 (Tamura et al. 2011, 2013). The combined
alignments in FASTA fomat were transform to PHYLIP
formats by ALTER online program (http://www.sing-group
.org/ALTER/) (Glez-Peña et al. 2010).
Phylogenetic analysis of the combined aligned dataset
was analyze based on maximum likelihood (ML) and Bayesian inference (BI). Maximum likelihood analysis was performed in RAxML (Stamatakis et al. 2008) implemented in
raxmlGUI v.0.9b2 (Silvestro and Michalak 2010) with 1000
rapid bootstrap replicates using the GTR + GAMMA model
of nucleotide substitution. Maximum likelihood bootstrap
values (MLBP) equal or greater than 60% are given above
each node. Posterior probabilities (PP) (Rannala and Yang
1996; Zhaxybayeva and Gogarten 2002) were determined by
Markov chain Monte Carlo sampling (MCMC) in MrBayes
v.3.0b4 (Huelsenbeck and Ronquist 2001). MrModeltest 2.2
(Nylander 2004) was used to perform the model of nucleotide substitution for each gene. Six simultaneous Markov
chains were run for 5,000,000 generations or depending on
individual settings for the fungal group, and trees were sampled every 100th generation. Tracer v1.6 (Rambaut et al.
2014) was used to examine the log-likelihood scores to
decide extra runs and determine the stationary phase. The
first 20% of trees were discarded as the burn-in phase. The
remaining trees were used for calculating PP in the majority
rule consensus tree. The run was stopped when the standard deviation of split frequencies was reached below 0.01
(Ariyawansa et al. 2013; Maharachchikumbura et al. 2015;
Udayanga et al. 2015). Bayesian posterior probabilities (PP)
equal to or greater than 0.95 are given above the nodes. Phylogenetic trees were drawn using FigTree 1.4.2 (Rambaut
2009) and edited by Microsoft Office PowerPoint 2013 and
Adobe Illustrator CS6 (Adobe Systems, USA).
Taxonomy
Phylogenetic analysis
The most closely related taxa for analyses were selected
following nucleotide BLAST searches in GenBank (http://
www.ncbi.nlm.nih.gov/) as well as sequence data from
13
Phylum Ascomycota Caval.-Sm.
Class Dothideomycetes sensu O.E. Erikss. & Winka
Subclass Pleosporomycetidae C.L. Schoch et al.
Fungal Diversity
Table 1 Details of genes/loci
with PCR primers and protocols
Family or genus
Class Dothideomycetes
Hysteriaceae
Didymosphaeriaceae
Nigrogranaceae
Phaeosphaeriaceae
Acrocalymmaceae
Lophiostomataceae
Neomassarinaceae
Roussoellaceae, Thyridariaceae
Torulaceae, Muyocopronaceae,
Patellariaceae
Didymellaceae
Neopyrenochaetaceae
Pyrenochaetopsidaceae
Aplosporellaceae
Botryosphaeriaceae
Pleurotremataceae
Class Sordariomycetes
Diaporthaceae
Hypocreaceae
Stachybotryaceae
Apiosporaceae
Cainiaceae
Hysteriales Lindau
Hysteriales was introduced by Lindau (1897b) with a
monotypic family, Hysteriaceae. In this study, we follow
the latest treatment and updated accounts of Hysteriales in
Jayasiri et al. (2018) and Wijayawardene et al. (2018), with
updated accounts of the genus Rhytidhysteron from recent
relevant literature (Jayasiri et al. 2018; Dayarathne et al.
2020; Kumar et al. 2019).
Gene/loci PCR primers (forward/reverse) References for primer
LSU
SSU
ITS
TEF1
LSU
SSU
ITS
TEF1
RPB2
LROR/LR5
NS1/NS4
ITS5/ITS4
983F/2218R
LROR/LR5
NS1/NS4
ITS5/ITS4
983F/2218R
fRPB2-5f/fRPB2–7cR
Vilgalys and Hester (1990)
White et al. (1990)
White et al. (1990)
Rehner (2001)
Vilgalys and Hester (1990)
White et al. (1990)
White et al. (1990)
Rehner (2001)
Liu et al. (1999)
LSU
ITS
TEF1
RPB2
LSU
ITS
RPB2
LSU
ITS
TEF1
ITS
TEF1
LSU
SSU
TEF1
LROR/LR5
ITS5/ITS4
983F/2218R
fRPB2-5f/fRPB2–7cR
LROR/LR5
ITS5/ITS4
fRPB2-5f/fRPB2–7cR
LROR/LR5
ITS5/ITS4
983F/2218R
ITS5/ITS4
983F/2218R
LROR/LR5
NS1/NS4
983F/2218R
Vilgalys and Hester (1990)
White et al. (1990)
Rehner (2001)
Liu et al. (1999)
Vilgalys and Hester (1990)
White et al. (1990)
Liu et al. (1999)
Vilgalys and Hester (1990)
White et al. (1990)
Rehner (2001)
White et al. (1990)
Rehner (2001)
Vilgalys and Hester (1990)
White et al. (1990)
Rehner (2001)
ITS
TEF1
RPB2
TEF1
LSU
ITS
TEF1
RPB2
LSU
ITS
TEF1
LSU
ITS
ITS5/ITS4
983F/2218R
fRPB2-5f/fRPB2–7cR
983F/2218R
LROR/LR5
ITS5/ITS4
983F/2218R
fRPB2-5f/fRPB2–7cR
LROR/LR5
ITS5/ITS4
983F/2218R
LROR/LR5
ITS5/ITS4
White et al. (1990)
Rehner (2001)
Liu et al. (1999)
Rehner (2001)
Vilgalys and Hester (1990)
White et al. (1990)
Rehner (2001)
Liu et al. (1999)
Vilgalys and Hester (1990)
White et al. (1990)
Rehner (2001)
Vilgalys and Hester (1990)
White et al. (1990)
Hysteriaceae Chevall.
Hysteriaceae was introduced by Chevallier (1826) with
Hysterium as the type genus. The family has been classified in several orders such as Pseudosphaeriales (Nannfeldt
1932; Gäumann 1949), Dothiorales (Müller and von Arx
1950; von Arx and Müller 1954), Dothideales (von Arx and
Müller 1975) and presently, Hysteriales, which is closely
related to the Pleosporales (Luttrell 1955; Kirk et al. 2008;
Thambugala et al. 2016; Jayasiri et al. 2018; Wijayawardene
13
Fungal Diversity
et al. 2018). Jayasiri et al. (2018) accepted 13 genera, which
includes nine genera based on both morphology and phylogenetic analyses (Gloniopsis, Graphyllium, Hysterium,
Hysterobrevium, Hysterodifractum, Oedohysterium, Ostreichnion, Psiloglonium, Rhytidhysteron), and four genera
(Actidiographium, Gloniella, Hysterocarina, Hysteropycnis)
based on morphology only. The divergence time estimates
for this family are crown age of 149 Mya (90–213) in the
late Jurassic and stem age of 219 Mya (161–282) in the late
Triassic (Liu et al. 2017).
Rhytidhysteron Speg.
Rhytidhysteron was introduced by Spegazzini (1881a)
to accommodate two species, R. brasiliense and R. viride.
Subsequently, R. brasiliense was designated as the type species (Clements and Shear 1931). There are 21 epithets listed
in Index Fungorum (2020). The genus has been classified
within the family Patellariaceae (Kutorga and Hawksworth
1997; Eriksson 2006; Lumbsch and Huhndorf 2010a, b).
However, recent studies based on multi-gene analyses have
shown that the genus should be placed in Hysteriaceae
Fig. 1 Phylogram generated
from maximum likelihood
analysis based on combined
dataset of LSU, SSU, ITS and
TEF1 sequence data. Thirty
strains are included in the
combined sequence analysis,
which comprise 3568 characters with gaps. Tree topology
of the ML analysis was similar
to the BYPP. The best scoring
RAxML tree with a final likelihood value of − 9583.200312
is presented. The matrix had
646 distinct alignment patterns, with 33.57% of undetermined characters or gaps.
Estimated base frequencies
were as follows: A = 0.245845,
C = 0.235270, G = 0.278609,
T = 0.240276; substitution rates:
AC = 1.066264, AG = 2.632538,
AT = 1.024795, CG = 0.715396,
CT = 6.041010, GT = 1.000000;
gamma distribution shape
parameter α = 0.079339. Bootstrap support values for ML
equal to or greater than 60% and
BYPP equal to or greater than
0.90 are given above the nodes.
Newly generated sequences
are in dark red bold and type
species are in bold. Mytilinidion
mytilinellum (EB 0386, CBS
303.34) are used as outgroup
taxa
13
(Boehm et al. 2009a, b; de Almeida et al. 2014; Wijayawardene et al. 2014a; Jayasiri et al. 2018; Wijayawardene
et al. 2018; Kumar et al. 2019) with seven species having
available sequences in GenBank (R. hysterinum, R. mangrovei, R. neorufulum, R. opuntiae, R. rufulum, R. thailandicum, R. tectonae). Most species were found in Thailand
(Thambugala et al. 2016; Doilom et al. 2017; Kumar et al.
2019) and isolated from diverse habitats, such as saprobic on
decaying woody branches, stems or twigs in terrestrial habitats, as well as marine habitats from mangrove wood (Kumar
et al. 2019). The genus has not been previously reported on
C. odorata nor on the host family Asteraceae. Dayarathne
et al. (2020) introduced a new species, R. bruguierae from
submerged branches of Bruguiera sp. in Thailand. In this
study, we introduce a new species, R. chromolaenae with a
new host record for R. bruguierae, based on morphology and
molecular data, together with descriptions and illustrations
(Figs. 2, 3, 4). A phylogenetic tree based on combined LSU,
SSU, ITS and TEF1 sequence data is presented in Fig. 1.
Fungal Diversity
Fig. 2 Rhytidhysteron bruguierae (new host record).
a, b Appearance of ascomata on substrate. c Section
through ascoma. d Exciple. e
Pseudoparaphyses. f–i Asci.
j–o Ascospores. Scale bars:
a = 500 µm, b, c = 200 µm, d,
f–i = 50 µm, e, j–o = 10 µm
Rhytidhysteron bruguierae Dayarathne, Mycosphere 11(1):
20 (2020)
Facesoffungi number: FoF 06154; Fig. 2
Saprobic on dead stems of Chromolaena odorata. Sexual
morph: Ascomata 250–500 µm high, 500–1000 µm diam.
( x̄ = 360 × 720 µm, n = 10), hysterothecial, boat-shaped,
superficial, with a carbonaceous rim, scattered, closed at
first and opening at maturity, elongate, elliptic, dark brown
to black with dark orange at the center, when dry folded
at the margin. Exciple 45–60(–110) µm wide, comprising
hyaline or pale brown to brown cells arranged in textura
globulosa to textura angularis, continuous to the base (hypothecium). Hamathecium comprising 1.5–2.5 µm wide,
cylindrical to filiform, septate, branching pseudoparaphyses,
13
Fungal Diversity
Fig. 3 Rhytidhysteron chromolaenae (holotype) a, b Appearance of ascomata on substrate. c Section through ascoma. d Exciple. e–g Pseudoparaphyses. h–k Asci. l–q Ascospores. Scale bars: a = 500 µm, b = 200 µm, c, h–k = 100 µm, d = 20 µm, e–g, l–q = 10 µm
13
Fungal Diversity
Fig. 4 Culture characteristics on MEA: a Rhytidhysteron bruguierae (MFLUCC 17-1515). b Rhytidhysteron chromolaenae
(MFLUCC 17-1516)
slightly swollen and rounded at the apex, forming pale red
to pinkish brown epithecium above the asci when mounted
in lactoglycerol and slightly orange to yellowish orange
epithecium above the asci when mounted in water. Asci
(120–)130–140 × 10–15 µm ( x̄ = 130 × 11 µm, n = 10),
8-spored, bitunicate, fissitunicate, cylindrical, straight
or slightly curved, with a short pedicel, apically rounded
with an ocular chamber. Ascospores 18–22 × 7–9 µm
( x̄ = 20 × 8 µm, n = 30), overlapping, uniseriate, pale brown
to brown, broadly fusiform, with upper part or second cell
slightly wider, 1-septate when immature, becoming 3-euseptate when mature, slightly constricted at the central septum, straight or slightly curved, guttulate, without terminal
appendages. Asexual morph: Undetermined.
Culture characteristics: Ascospores germinating on
MEA within 24 h. at room temperature and germ tubes produced from second cell from apex. Colonies on MEA filamentous, mycelium velvety, fluffy, undulate to filamentous,
white aerial hyphae at the surface and in reverse (Fig. 4a).
Pre-screening for antimicrobial activity: Rhytidhysteron bruguierae (MFLUCC 17-1515) showed antimicrobial
activity against Mucor plumbeus with an 18.5 mm inhibition
zone, observable as partial inhibition, when compared with
the positive control (17 mm), but no inhibition of Bacillus
subtilis and Escherichia coli.
Known hosts and distribution: On submerged branches
of Bruguiera sp. (Rhizophoraceae) in Thailand (Dayarathne
et al. 2020).
Material examined: THAILAND, Chiang Rai Province,
Doi Pui, on dead stems of Chromolaena odorata, 7 April
2017, A. Mapook (DP106, MFLU 20-0363); living culture
MFLUCC 17-1515 (new host record); Phrae Province,
Doi Pha Klong, on dead stems of C. odorata, 22 September
2016, A. Mapook (DPKP5, MFLU 20-0364); living culture
MFLUCC 17-1511; Chiang Mai Province, Fah Hom Pok, on
dead stems of C. odorata, 27 September 2016, A. Mapook
(FHP4, MFLU 20-0365); living culture MFLUCC 17-1502;
Mae Hong Son Province, Mae Yen, Pai, on dead stems of C.
odorata, 25 June 2016, A. Mapook (MY8, MFLU 20-0366);
living culture MFLUCC 17-1509.
GenBank numbers: LSU: MN632455, MN632454,
MN632453, MN632452, ITS: MN632460, MN632459,
MN632458, MN632457, SSU: MN632466, MN632465,
MN632464, MN632463, TEF1: MN635662, MN635661
Notes: In a BLASTn search of NCBI GenBank, the
closest match of the ITS sequence of our four strains with
100% similarity was Rhytidhysteron rufulum (strain 510A,
EU020063), while the closest match with the LSU, SSU and
TEF1 sequences with 99.34%, 99.78% and 96.67% similarity, respectively, was R. thailandicum (strain MFLUCC
14-0503). In the present phylogenetic analysis, MFLUCC
17-1515, MFLUCC 17-1511, MFLUCC 17-1502 and
MFLUCC 17-1509 cluster with R. bruguierae (MFLUCC
18-0398) with high bootstrap support (100% ML and 1.00
BYPP, Fig. 1). We therefore, identify our four strains as R.
bruguierae based on phylogenetic analyses with morphological comparison (Table 2) and the four isolates are introduced
here as a new host record from Chromolaena odorata collected in Thailand. Rhytidhysteron bruguierae is also closely
related to R. thailandicum (Fig. 1). However, R. bruguierae
differs from R. thailandicum in having slightly smaller asci
[(120–)130–140 × 10–15 µm vs. 135–160 × 10.5–15 µm] and
pale brown to brown, smaller ascospores (18–22 × 7–9 µm
vs. 20–28(–31) × 7.5–12 µm) (Table 2).
Rhytidhysteron chromolaenae Mapook & K.D. Hyde, sp.
nov.
Index Fungorum number: IF557360, Facesoffungi number: FoF 07780; Fig. 3
Etymology: Name reflects the host genus Chromolaena,
from which this species was isolated.
Holotype: MFLU 20-0367
Saprobic on dead stems of Chromolaena odorata. Sexual morph: Ascomata 275–290(–380) µm
high × (685–)750–885 µm diam. ( x̄ = 305 × 780 µm, n = 5),
hysterothecial, boat-shaped, superficial, with a carbonaceous
rim, not perpendicularly striate, scattered, closed at first and
opening at maturity, elongate, elliptic, dark brown to black
with yellowish green on the margin, orange or dark brown to
black at the center. Exciple (50–)60–85 µm wide, comprising
pale brown or brown to dark brown cells, arranged in textura
globulosa to textura angularis, continuous to the base (hypothecium). Hamathecium comprising (1.5–)2–3 µm wide,
cylindrical to filiform, septate, branching pseudoparaphyses, slightly swollen and rounded at the apex, forming
pale orange to yellowish orange epithecium above the asci
when mounted in water, orange to red epithecium above
the asci when mounted in lactoglycerol and becoming
slightly purple epithecium above the asci when mounted in
5% KOH. Asci 130–155 × (8–)11–14 µm ( x̄ = 145 × 12 µm,
n = 10), 7–8-spored, bitunicate, fissitunicate, cylindrical,
13
Fungal Diversity
straight or slightly curved, apically rounded. Ascospores
23–28 × 8–11 µm ( x̄ = 26.5 × 9 µm, n = 25), overlapping,
uniseriate, broadly fusiform, widest at the center, hyaline
to yellowish brown, 1-septate when immature, becoming
brown to dark brown, 3-septate when mature, slightly constricted at the central septum, straight or slightly curved,
guttulate, without terminal appendages. Asexual morph:
Undetermined.
Culture characteristics: Ascospores germinating on
MEA within 24 h. at room temperature and germ tubes
produced from second cell from apex. Colonies on MEA
irregular with slightly raised, undulate, white aerial hyphae
spreading from the center of the colony, become smoke-gray
to olivaceous brown at the surface and olivaceous brown
with creamy white at the margin in reverse (Fig. 4b).
Pre-screening for antimicrobial activity: Rhytidhysteron chromolaenae (MFLUCC 17-1516) showed antimicrobial activity against M. plumbeus with a 20 mm inhibition
zone, observable as partial inhibition, when compared with
the positive control (17 mm), but no inhibition of B. subtilis
and E. coli.
Material examined: THAILAND, Chiang Rai Province,
Doi Pui, on dead stems of Chromolaena odorata, 7 April
2017, A. Mapook (DP107, MFLU 20-0367, holotype); extype culture MFLUCC 17-1516.
GenBank numbers: LSU: MN632456, ITS: MN632461,
SSU: MN632467, TEF1: MN635663
Notes: Rhytidhysteron chromolaenae is similar to R.
columbiense (CUVC 62421) in having ascomata with
yellowish green margins and (1–)3-septate ascospores,
but differs in having smaller ascomata (274–290(–380
) × (685–)750–885 µm vs. 600–700 × 1200–1800 µm);
smooth, not perpendicularly striate at margin, smaller asci
(130–155 × (8–)11–14 µm vs. 175–190 × 14–18 µm) and
smaller ascospores (23–28 × 8–11 µm vs. 38–52 × 13–18 µm)
(Table 2). Rhytidhysteron columbiense was described from
Colombia without molecular data (Soto-Medina and Lücking 2017). In the present phylogenetic analysis, R. chromolaenae forms a well-separated branch from other Rhytidhysteron species and is closely related to R. mangrovei,
R. thailandicum and R. bruguierae (Fig. 1). In a BLASTn
search of NCBI GenBank, the closest match of the ITS
sequence for MFLUCC 17-1516 is R. rufulum with 99.81%
similarity to the strain B1a081-2-P30 (JQ388942), while the
closest match with the LSU sequence was with R. neorufulum (strain MFLUCC 13-0216, NG_059649) with 98.90%
similarity and closest matches with the TEF1 sequence were
R. thailandicum (strain MFLUCC 14-0503, KU497490) with
95.90% similarity. Therefore, R. chromolaenae is described
13
as a new species based on phylogeny and morphological
characters.
Pleosporales Luttrell ex M.E. Barr
Pleosporales was introduced by Luttrell (1955). The
order is highly diverse with more than 75 families (Wijayawardene et al. 2018). We follow the latest treatment and
updated accounts of Pleosporales in Liu et al. (2017) and
Wijayawardene et al. (2018), together with recent relevant
literature for updated accounts of each family.
Acrocalymmaceae Crous & Trakun.
Acrocalymmaceae was introduced by Trakunyingcharoen
et al. (2014b) to accommodate the genus Acrocalymma. The
divergence time estimates for this family are crown age of 23
Mya (8–44) in the Neogene period and stem age of 114 Mya
(71–156) in the Cretaceous (Liu et al. 2017).
Acrocalymma Alcorn & J.A.G. Irwin, Trans. Br. mycol. Soc.
88(2): 163 (1987)
Acrocalymma is an ecologically diverse genus, containing
plant pathogens, endophytes, and saprobes from terrestrial
or freshwater as well as human superficial tissue (Alcorn
and Irwin 1987; Zhang et al. 2012a, b; Trakunyingcharoen
et al. 2014b; Valenzuela-Lopez et al. 2017; Jin et al. 2018;
Jayasiri et al. 2019). The genus was introduced by Alcorn
and Irwin (1987) with A. medicaginis as the type species.
Jayasiri et al. (2019) introduced a new species, A. pterocarpi
from a fallen pod of Pterocarpus indicus in Thailand. Seven
epithets are listed in Index Fungorum (2020). In this study,
the first record of Acrocalymma on Chromolaena odorata
is introduced, based on morphology and molecular data
with a description and illustrations (Fig. 6). A phylogenetic
tree based on combined LSU, ITS, SSU, TEF1 and RPB2
sequence data is presented in Fig. 5.
Acrocalymma medicaginis Alcorn & J.A.G. Irwin, Trans.
Br. mycol. Soc. 88(2): 163 (1987)
Facesoffungi number: FoF 07099; Fig. 6
Saprobic on dead stems of Chromolaena odorata. Sexual
morph: Ascomata 150–250 µm high × (115–)140–155 µm
diam. ( x̄ = 195 × 140 µm, n = 5), immersed, appearing as
black spots, coriaceous, solitary or scattered, subglobose to
obpyriform, brown to dark brown. Ostiole long neck, papillate. Peridium 10–20 µm wide, 4–5 layers, comprising pale
brown to brown cells of textura angularis. Hamathecium
comprising 0.5–1.5 µm wide, cylindrical to filiform, septate, branching pseudoparaphyses. Asci 70–85 × 8–11 µm
( x̄ = 78 × 9 µm, n = 10), 8-spored, bitunicate, fissitunicate,
Fungal Diversity
Fig. 5 Phylogram generated
from maximum likelihood
analysis based on combined
dataset of LSU, ITS, SSU,
TEF1 and RPB2 sequence data.
Sixteen strains are included in
the combined sequence analysis,
which comprise 4699 characters with gaps. Tree topology
of the ML analysis was similar
to the BYPP. The best scoring
RAxML tree with a final likelihood value of − 11536.335405
is presented. The matrix had
503 distinct alignment patterns, with 54.55% of undetermined characters or gaps.
Estimated base frequencies
were as follows: A = 0.242971,
C = 0.232109, G = 0.270331,
T = 0.254589; substitution rates:
AC = 1.527101, AG = 2.317230,
AT = 1.662072, CG = 1.025372,
CT = 5.286531, GT = 1.000000;
gamma distribution shape
parameter α = 0.208825. Bootstrap support values for ML
equal to or greater than 60% and
BYPP equal to or greater than
0.90 are given above the nodes.
Newly generated sequences
are in dark red bold and type
species are in bold. Massarina
eburnea (CBS 473.64) is used
as outgroup taxon
Table 2 Synopsis of Rhytidhysteron species with similar morphological features discussed in this study
Species
Asocomata (µm)
Exciple (µm)
R. columbiense
600–700 high × 1200– 60–90, textura angu(CUVC 62421)
(1590)–1800 diam.
laris
R. bruguierae
410–520 high × 548– 148–162, textura
(MFLUCC 18-0398)
570 diam.
angularis
R. bruguierae
250–500 high × 500– 45–60(–110), textura
(MFLUCC 17-1515)
1000 diam.
globulosa to textura
angularis
(50–)60–85, textura
R. chromolaenae
274–290(–380)
globulosa
(MFLUCC 17-1516)
high × (685–)750–
885 diam.
360–640 high × 53072–130, textura
R. thailandicum
750 diam.
angularis
(MFLUCC 14-0503)
135–160 × 10.5–15
cylindric-clavate, straight or slightly curved, with a
short pedicel, apically rounded, with an ocular chamber.
Ascospores 15–21 × 3–5 µm (x̄ = 18 × 4 µm, n = 20), overlapping, 1–2-seriate, hyaline, oblong to broadly fusiform with
Asci (µm)
Ascospores (µm)
References
175–190 × 14–18
38–(43.5)–52 × 13–
(14.7)–18
14–26 × 6.2–9
(120–)130–
140 × 10–15
18–22 × 7–9
Soto-Medina and
Lücking (2017)
Dayarathne et al.
(2020)
This study
130–155 × (8–)11–14
23–28 × 8–11
This study
20–28(–31) × 7.5–12
Thambugala et al.
(2016)
128–148 × 10–14
obtuse to slightly obtuse ends, uniseptate, straight or slightly
curved, smooth, constricted at the septum, surrounded by
hyaline gelatinous sheath observed clearly when mounted
in Indian ink. Asexual morph: Undetermined.
13
Fungal Diversity
Fig. 6 Acrocalymma medicaginis (new host record) a,
b Appearance of ascomata on
substrate. c Section through
ascoma. d Peridium. e Pseudoparaphyses. f–i Asci. j–n
Ascospores. o Ascospores surrounded by hyaline gelatinous
sheath in Indian ink. Scale bars:
a, b = 500 µm, c = 50 µm, d–i,
o = 10 µm, j–n = 5 µm
13
Fungal Diversity
Culture characteristics: Ascospores germinating on
MEA within 24 h at room temperature, germ tubes produced from both cells. Colonies on MEA irregular, mycelium slightly raised, moderately fluffy, filiform, white aerial
hyphae at the surface, spreading from the center and dark
brown to olivaceous-brown in reverse from the center with
creamy white at rim (Fig. 7).
Pre-screening for antimicrobial activity: Acrocalymma
medicaginis (MFLUCC 17-1439) showed antimicrobial
activity against B. subtilis with a 12 mm inhibition zone and
against M. plumbeus with a 13 mm inhibition zone, observable as partial inhibition, when compared with the positive
control (25 mm and 17 mm, respectively), but no inhibition
of E. coli.
Known hosts and distribution: Root and crown rotting
of Medicago sativa (Fabaceae) in Australia (Alcorn and
Irwin 1987; Trakunyingcharoen et al. 2014a, b)
Material examined: THAILAND, Chiang Rai Province,
Doi Pui, on dead stems of Chromolaena odorata, 8 July
2015, A. Mapook (DP5, MFLU 20-0296); living culture
MFLUCC 17-1423 (new host record); 5 August 2015, A.
Mapook (DP23, MFLU 20-0297); living culture MFLUCC
17-1439.
GenBank numbers: LSU: MT214432, MT214433, ITS:
MT214338, MT214339, SSU: MT214387, MT214388,
TEF1: MT235797, MT235798
Notes: Multigene phylogenetic analyses (Fig. 5) show that
two strains MFLUCC 17-1423, MFLUCC 17-1439 grouped
within the Acrocalymma medicaginis clade. A BLASTn
search of the LSU sequence data showed that the two
strains are identical to A. medicaginis (strain CPC 24345,
KP170718) with 100% similarity while the closest match
with the ITS sequence of our two strains are A. medicaginis
with 99.59% similarity to the strain CPC 24342 (KP170622).
Fig. 7 Culture characteristic on MEA: Acrocalymma medicaginis
(MFLUCC 17-1423)
Therefore, we identify our isolates as A. medicaginis based
on phylogenetic analyses. Only two species, A. walkeri and
A. pterocarpi have been reported with a sexual morph, the
other species being coelomycetous. In this study, we isolated
the sexual morph of A. medicaginis from Chromolaena odorata collected in Thailand for the first time, and the isolates
are also introduced here as a new host record. We did not
obtain the asexual morph of our isolates in culture.
Didymellaceae Gruyter et al.
Didymellaceae contains numerous plant pathogenic, saprobic and endophytic species associated with a wide range
of hosts, as well as some species of clinical or environmental origin. The family was introduced by De Gruyter et al.
(2009) to accommodate Phoma and phoma-like genera, with
Didymella as the type genus. Hyde et al. (2013) accepted
13 genera in the family based on both morphology and
phylogenetic analyses (Ascochyta, Boeremia, Chaetasbolisia, Dactuliochaeta, Didymella, Epicoccum, Leptosphaerulina, Macroventuria, Microsphaeropsis, Monascostroma,
Phoma, Piggotia, Pithomyces). Chen et al. (2015) introduced nine additional genera to the family (Allophoma, Calophoma, Heterophoma, Neoascochyta, Neodidymelliopsis,
Nothophoma, Paraboeremia, Phomatodes, Xenodidymella)
based on morphological observations with multi-locus phylogenetic analyses of ITS, LSU, RPB2 and TUB2 sequence
data. In addition, Didymellocamarosporium, Didysimulans,
Endocoryneum, Mixtura, Peyronellaea, Phaeomycocentrospora, Platychora, Pseudohendersonia and Stagonosporopsis were accepted in Outline of Ascomycota 2017
(Wijayawardene et al. 2018). Recently, Valenzuela-Lopez
et al. (2018) introduced six new genera (Cumuliphoma,
Ectophoma, Juxtiphoma, Remotididymella, Similiphoma,
Vacuiphoma) of clinical or environmental origin and synonymized Peyronellaea under the genus Didymella. Divergence time estimates for this family are crown age of 63 Mya
(35–97) in the Paleogene period and stem age of 115 Mya
(84–149) in the Cretaceous (Liu et al. 2017). Soleimani et al.
(2018) reported divergence time estimates with a focus on
Didymellaceae and suggested that the family diverged from
Aigialaceae at 86.7 Mya (53.9–155.4) in the Cretaceous and
initial divergence of the family happened in the late Eocene.
Didymella Sacc. ex D. Sacc.
Didymella was introduced by Saccardo (1880) with D.
exigua as the type species and contains some species which
are important serious plant pathogens, also endophytes and
saprobes on a wide range of substrates, as well as species
of clinical or environmental origin (Aveskamp et al. 2010;
Chen et al. 2015, 2017; Ahmadpour et al. 2017; Jayasiri
et al. 2017; Valenzuela-Lopez et al. 2018). The genus was
presented as monophyletic by Chen et al. (2015) and comprises 35 known species, with two unidentified species,
13
Fungal Diversity
Fig. 8 Phylogram generated
from maximum likelihood
analysis based on combined
dataset of LSU, ITS, TUB2 and
RPB2 sequence data. Sixtytwo strains are included in the
combined sequence analysis,
which comprise 3028 characters with gaps. Tree topology
of the ML analysis was similar
to the BYPP. The best scoring
RAxML tree with a final likelihood value of − 14613.624205
is presented. The matrix had
624 distinct alignment patterns, with 13.98% of undetermined characters or gaps.
Estimated base frequencies
were as follows: A = 0.240327,
C = 0.243303, G = 0.275771,
T = 0.240599; substitution rates:
AC = 0.969862, AG = 4.713232,
AT = 1.514683, CG = 0.711109,
CT = 10.328441,
GT = 1.000000; gamma
distribution shape parameter
α = 0.102562. Bootstrap support values for ML equal to or
greater than 60% and BYPP
equal to or greater than 0.90 are
given above the nodes. Newly
generated sequences are in dark
red bold and type species are
in bold. Epicoccum poae (LC
8160) and Epicoccum nigrum
(CBS 173.73) are used as outgroup taxa
based on morphological observations and multi-locus phylogenetic analyses of ITS, LSU, RPB2 and TUB2 sequence
data. Subsequently, Chen et al. (2017) introduced eleven
new Didymella species while studying the distribution and
biodiversity of Didymellaceae. Valenzuela-Lopez et al.
(2018) described two new species D. brunneospora and D.
keratinophila from flower-stalk and human superficial tissue,
respectively. Jayasiri et al. (2019) introduced a new species,
D. magnolia (MFLUCC 18-1560) collected from Magnolia
grandiflora cone in China. In this study, D. chromolaenae
13
is introduced as a new species based on morphology and
molecular data (Fig. 9). A phylogenetic tree based on combined LSU, ITS, TUB2 and RPB2 sequence data is presented
in Fig. 8.
Didymella chromolaenae Mapook & K.D. Hyde, sp. nov.
Index Fungorum number: IF557295, Facesoffungi number: FoF 07781; Fig. 9
Etymology: Name reflects the host genus Chromolaena,
from which this species was isolated.
Holotype: MFLU 20-0309
Fungal Diversity
Fig. 9 Didymella chromolaenae (holotype) a Appearance
of ascomata on substrate. b
Section through ascoma. c
Peridium. d–f Immature and
mature asci. g–k Ascospores.
Scale bars: a = 500 µm, b,
d–f = 50 µm, c = 20 µm,
g–k = 10 µm
Saprobic on dead stems of Chromolaena odorata. Sexual morph: Ascomata 130–145 µm high × 145–160 µm
diam. ( x̄ = 140 × 152 µm, n = 5), semi-immersed to superficial, appearing as small black spots, coriaceous, solitary
or scattered, globose to irregular shape, brown to dark
brown. Ostiole central. Peridium 20–30 µm wide, 2–5 layers of thin-walled, pale brown to hyaline cells of textura
angularis. Hamathecium without pseudoparaphyses. Asci
80–85 × 25–35 µm ( x̄ = 83 × 29 µm, n = 5), 8-spored, bitunicate, cylindric-clavate to clavate, with a short pedicel,
slightly curved, apically rounded with an ocular chamber.
Ascospores 15–22 × 9–13 µm ( x̄ = 19 × 11 µm, n = 20), overlapping, irregularly arranged, yellow, oval to obovoid, with
obtuse ends, widest at the upper cell and tapering towards
the narrow-rounded ends, uniseptate, with small guttules,
constricted at the septum, surrounded with mucilaginous
sheath. Asexual morph: Undetermined.
Culture characteristics: Ascospores germinating on
MEA within 24 h. at room temperature and germ tubes produced from the apex. Colonies on MEA circular, mycelium
flat, filiform, white at first, becoming gray to light grayish
brown hyphae spreading at the surface, yellow white with
grayish brown in reverse (Fig. 10).
Pre-screening for antimicrobial activity: Didymella
chromolaenae (MFLUCC 17-1459) showed no inhibition
against E. coli, B. subtilis and M. plumbeus.
Material examined: THAILAND, Lampang Province,
Chaehom, on dead stems of Chromolaena odorata, 24 September 2016, A. Mapook (JH5, MFLU 20-0309, holotype);
ex-type culture MFLUCC 17-1459.
GenBank numbers: LSU: MT214457, ITS: MT214363,
SSU: MT214409, TEF1: MT235799
Notes: In a BLASTn search of NCBI GenBank, the closest match of the ITS sequence of Didymella chromolaenae
(MFLUCC 17-1459, ex-holotype) is Didymella glomerata
with 98.92% similarity to the strain FK2 (KC802087), while
the closest match of the LSU sequence was identical with
99.78% to D. omnivirens (strain CBS 341.86, MH873653).
Based on the present phylogenetic analysis (Fig. 8), D. chromolaenae groups in the clade comprising D. viburnicola
(CBS 523.73), D. pteridis (CBS 379.96), D. ellipsoidea
(CGMCC 3.18350) and D. macrostoma (CBS 223.96) but
13
Fungal Diversity
Fig. 10 Culture characteristic on MEA: Didymella chromolaenae
(MFLUCC 17-1459)
low bootstrap support. However, based on morphological
comparison, D. chromolaenae (MFLUCC 17-1459) resembles species of Didymella based on its immersed to semiimmersed ascomata with one-celled ascospores and distinct from D. macrostoma in having oval to obovoid, yellow
ascospores surrounded with a mucilaginous sheath, while D.
macrostoma has ellipsoid to obovoid, hyaline ascospores. A
comparison of the ITS (+ 5.8S) gene region of D. chromolaenae with D. ellipsoidea and D. macrostoma reveals 9 base
pair differences (1.85%) across 486 nucleotides. Therefore,
D. chromolaenae is described as a new species.
Nothophoma Q. Chen & L. Cai
Nothophoma was introduced by Chen et al. (2015) with
N. infossa as the type species, and four new combinations
(N. anigozanthi, N. arachidis-hypogaeae, N. gossypiicola, N.
quercina) based on morphology and phylogeny. Crous et al.
(2016) introduced a new species, N. macrospora isolated
from human respiratory tract. Abdel-Wahab (2017) introduced a new endophytic species, N. multilocularis isolated
from the medicinal plant Rhazya stricta, and reported on
its antimicrobial activity. Crous et al. (2017) introduced N.
raii, isolated from soil and Valenzuela-Lopez et al. (2018)
introduced N. variabilis which was also isolated from human
respiratory tract. There are nine epithets listed in Index Fungorum (2020). Nothophoma chromolaenae is introduced as
a new species based on morphology and molecular data,
together with description and illustrations (Fig. 12). A phylogenetic tree based on combined LSU, ITS, TUB2 and
RPB2 sequence data is presented in Fig. 11.
13
Nothophoma chromolaenae Mapook & K.D. Hyde, sp. nov.
Index Fungorum number: IF557346, Facesoffungi number: FoF 07782; Fig. 12
Etymology: Name reflects the host genus Chromolaena,
from which this species was isolated.
Holotype: MFLU 20-0342
Saprobic on dead stems of Chromolaena odorata. Sexual morph: Ascomata 55–85 µm high × 85–100(–130) µm
diam. (x̄ = 73 × 98 µm, n = 10), immersed to semi-immersed,
appearing as small black spots, coriaceous, solitary or scattered, globose, dark brown to reddish brown. Ostiole central,
papillate. Peridium (5–)10–20(–30) µm wide, 2–3 layers of
pale brown to brown cells of textura angularis. Hamathecium without pseudoparaphyses. Asci 45–70 × 13–16 µm
( x̄ = 55 × 14.5 µm, n = 15), 8-spored, bitunicate, cylindricclavate, straight or slightly curved, with a short pedicel, apically round with an ocular chamber. Ascospores
7–17 × 4–6.5 µm ( x̄ = 10 × 5 µm, n = 15), overlapping,
irregularly arranged, hyaline, broadly fusiform, aseptate,
widest at the center and tapering towards narrow rounded
ends, straight or slightly curved, rough surface with terminal
appendages. Asexual morph: Undetermined.
Culture characteristics: Ascospores germinating on
MEA within 24 h at room temperature and germ tubes produced from both ends. Colonies on MEA circular, mycelium
slightly raised, filiform, dark brown aerial hyphae at the surface and dark brown in reverse (Fig. 13).
Pre-screening for antimicrobial activity: Nothophoma
chromolaenae (MFLUCC 17-1443) showed no inhibition of
E. coli, B. subtilis and M. plumbeus.
Material examined: THAILAND, Phetchaburi Province,
Cha-am, Khao Nang Panthurat, on dead stems of Chromolaena odorata, 28 July 2015, A. Mapook (NPR6, MFLU
20-0342, holotype); ex-type culture MFLUCC 17-1443.
GenBank numbers: LSU: MT214458, ITS: MT214364,
SSU: MT214410
Notes: In a BLASTn search of NCBI GenBank, the closest match of the ITS sequence of Nothophoma chromolaenae (MFLUCC 17-1443, ex-holotype) is N. gossypiicola
with 99.79% similarity to the strain UTHSC:DI16-294
(LT592943), while the closest match of the LSU sequence
were identical with 98.72% to Macrophoma lageniformis
(strain CBS 364.65, MH870253) and Nothophoma macrospora (strain UTHSC:DI16-294, LN907437). In the
present phylogenetic analysis, N. chromolaenae groups in
the clade comprising, N. gossypiicola, N. macrospora, N.
multilocularis and N. raii (Fig. 11). Those species have
been recorded as asexual morphs. We compared those
asexual morphs (Table 3) and found that they are morphologically distinct. Nothophoma gossypiicola differs
Fungal Diversity
Fig. 11 Phylogram generated
from maximum likelihood
analysis based on combined
dataset of LSU, ITS, TEF1 and
RPB2 sequence data. Eleven
strains are included in the
combined sequence analysis,
which comprise 2757 characters with gaps. Tree topology
of the ML analysis was similar
to the BYPP. The best scoring
RAxML tree with a final likelihood value of − 5700.668844
is presented. The matrix had
226 distinct alignment patterns, with 17.08% of undetermined characters or gaps.
Estimated base frequencies
were as follows: A = 0.243238,
C = 0.237511, G = 0.277502,
T = 0.241749; substitution rates:
AC = 0.970562, AG = 4.188061,
AT = 1.216550, CG = 0.588503,
CT = 14.649953,
GT = 1.000000; gamma
distribution shape parameter
α = 0.020000. Bootstrap support values for ML equal to or
greater than 60% and BYPP
equal to or greater than 0.90 are
given above the nodes. Newly
generated sequences are in dark
red bold and type species are in
bold. Phoma herbarum (CBS
615.75) is used as outgroup
taxon
from N. multilocularis in having smaller conidiomata
(100–250 µm vs. 175–1500 µm), smaller conidiogenous
cells (5–8 × 5–8 µm vs. 11–17 × 9–18 µm) and slightly
smaller conidia (10–12.5 × 2.5–3.5 µm vs. 9–20 × 3–4(–5)
µm), while N. multilocularis, N. gossypiicola and N. macrospora differs from N. raii in having wider conidia. However,
our strain, N. chromolaenae was found as the sexual morph
in nature and we could not obtain its asexual morph in culture. A comparison of the ITS (+5.8S) gene region of N.
chromolaenae and N. raii reveals seven base pair differences
(1.6%) across 434 nucleotides. Therefore, N. chromolaenae
is described here as a new species based on morphology and
phylogeny (Table 3).
Didymosphaeriaceae Munk, Dansk bot. Ark. 15(no. 2): 128
(1953)
Didymosphaeriaceae was introduced by Munk (1953)
with Didymosphaeria as type genus. Ariyawansa et al.
(2014b) provided an updated account of the family and
accepted sixteen genera (Alloconiothyrium, Barria, Bimuria,
Deniquelata, Didymocrea, Didymosphaeria, Julella,
Kalmusia, Karstenula, Letendraea, Montagnula, Neokalmusia, Paraconiothyrium, Paraphaeosphaeria, Phaeodothis, Tremateia). They clarified the taxonomic relationships
within the families Didymosphaeriaceae and Montagnulaceae, and synonymized Montagnulaceae under Didymosphaeriaceae based on the oldest name. Wijayawardene et al.
(2014a, b) introduced two new asexual genera Paracamarosporium and Pseudocamarosporium. Subsequently, Crous
et al. (2015b, c) introduced two new genera Verrucoconiothyrium and Xenocamarosporium. Ariyawansa et al. (2015)
referred Austropleospora and Pseudopithomyces to the family. Wanasinghe et al. (2016) introduced Laburnicola and
Paramassariosphaeria within the family and synonymized
two Munkovalsaria species under the genus Montagnula.
Thambugala et al. (2017b) introduced a new genus Kalmusibambusa which was collected from living culms of bamboo
in Thailand. Jayasiri et al. (2019) introduced a new genus
Cylindroaseptospora and Phookamsak et al. (2019) also
added a new genus Vicosamyces in the family. Divergence
13
Fungal Diversity
Fig. 12 Nothophoma chromolaenae (holotype) a Appearance
of ascomata on substrate. b Section through ascoma. c Peridium. d–f Immature and mature
asci. g–l Ascospores. Scale
bars: a = 400 µm, b = 50 µm, c,
d–f = 20 µm, g–l = 10 µm
time estimates for this family are crown age of 72 Mya
(48–101) and stem age of 109 Mya (83–139) during the
Cretaceous (Liu et al. 2017).
Chromolaenicola Mapook & K.D. Hyde, gen. nov.
Index Fungorum number: IF557279, Facesoffungi number: FoF 07783
Etymology: Named after the host genus, Chromolaena.
Saprobic on dead stems. Sexual morph: Ascomata
immersed to semi-immersed, solitary or scattered, appearing as small dark spots, coriaceous, globose to subglobose,
brown to dark brown. Ostiolar neck protruding. Peridium
3–4 layers, comprising dark brown cells of textura angularis. Hamathecium composed of cylindrical to filiform,
septate, branching pseudoparaphyses. Asci 6–8-spored,
bitunicate, cylindrical, straight or slightly curved, apically
rounded, pedicellate with an ocular chamber. Ascospores
slightly overlapping, uni-seriate, initially hyaline, 1-septate
when immature, becoming reddish brown to brown at maturity, ellipsoid to broadly fusiform, muriform, 3 transverse
septate, and 1 vertical septum, constricted at the central septum, straight or slightly curved, without gelatinous sheath.
Asexual morph: Conidiomata pycnidial, solitary, immersed
13
Fig. 13 Culture characteristic on MEA: Nothophoma chromolaenae (MFLUCC 17-1443)
to semi-immersed, uni-loculate, globose to obpyriform, yellowish brown to brown, sometimes appearing as colonies
on the host surface, superficial, scattered, gregarious, dark
brown to black, not easy to remove from the host surface.
Fungal Diversity
Table 3 Morphological features of Nothophoma species discussed in this study
Species
Conidiomata (µm)
Conidiogenous cells
(µm)
Conidia (µm)
Hosts
References
N. gossypiicola (CBS.
377.67)
N. macrospora
(UTHSC: DI16-276)
N. multilocularis
(AUMC 12003)
N. raii (MCC 1082)
100–250
5–8 × 5–8
10–12.5 × 2.5–3.5
De Gruyter (2002)
100–300
5–10 diam.
175–1500
11–17 × 9–18
(9–)10–15 × 2.5–3(–
3.5)
9–20 × 3–4(–5)
194.3–315.5 × 195.6–
411.3
–
11–14.5 × 1.5–2.5
Parasitic on Gossypium spp.
Human clinical specimen
Endophyte of Rhazya
stricta.
Soil from industrial
area
Crous et al. (2016)
Abdel-Wahab (2017)
Crous et al. (2017)
Table 4 Synopsis of asexual morph of Chromolaenicola species with similar morphological features discussed in this study
Species
Conidiomata (μm)
Conidiogenous cells
(μm)
Conidia (μm)
C. chiangraiensis
(MFLUCC 17-1493)
–
3.5–6.5 × 1–2
9–14 × 6–9
C. lampangensis
(MFLUCC 17-1462)
150–230 high × (155–
–
)170–270(–345) diam.
C. siamensis (= Cylindroaseptospora
siamensis, MFLUCC
17–2527)
110–165 high × 140–
190 diam.
6.5–7.4 × 3.2–4.7
Ostiole central, papillate. Pycnidial wall comprising 2–4
layers, pale brown to light brown cells of textura angularis. Conidiophores reduced to conidiogenous cells. Conidiogenous cells hyaline and unbranched, smooth, elongated,
broadly filiform to ampulliform. Conidia oblong or oval
to ellipsoid, globose to subglobose, hyaline to pale brown,
aseptate when immature, becoming reddish brown to brown,
1-septate when mature, not constricted at the septum, thickwalled finely verruculose.
Type species: Chromolaenicola nanensis Mapook &
K.D. Hyde
Notes: Multigene phylogenetic analyses based on combined LSU, ITS, SSU and TEF1 sequence data show that
Chromolaenicola species form a monophyletic clade in
Didymospheriaceae (0.92 BYPP) and basal to Cylindroaseptospora leucaenae (MFLUCC 17-2424) with low bootstrap support (Fig. 14). Chromolaenicola species differ
from Cy. leucaenae in having oblong or oval to ellipsoid,
globose to subglobose conidia, hyaline to pale brown and
aseptate when immature, becoming dark and 1-septate at
maturity, thick-walled, verruculose, not constricted at the
septum, whereas, Cy. leucaenae has cylindrical conidia, hyaline, aseptate with smooth thin walls (Jayasiri et al. 2019).
Hosts
References
This study
Thailand, Chiang Rai
Province on dead
stems of Chromolaena
odorata
This study
12–15 × 4–6.5
Thailand, Lampang
Province on dead
stems of Chromolaena
odorata
Jayasiri et al. (2019)
7.2–9.4 × 5.4–6.5 Thailand, Lampang
Province on decaying
pod of Leucaena sp.
(Fabaceae)
Therefore, we introduce Chromolaenicola as a new genus
to accommodate four new species with one new taxonomic
combination, based on morphological comparison and phylogenetic analyses.
Chromolaenicola chiangraiensis Mapook & K.D. Hyde,
sp. nov.
Index Fungorum number: IF557280, Facesoffungi number: FoF 07784; Fig. 15
Etymology: Referring to the location where the specimen
was collected, Chiang Rai Province, Thailand.
Holotype: MFLU 20-0301
Saprobic on dead stems of Chromolaena odorata. Sexual
morph: Undetermined. Asexual morph: Colonies superficial, covering the host, scattered, gregarious, dark brown
to black, not easy to remove from the host surface. Conidiophores reduced to conidiogenous cells. Conidiogenous
cells 3.5–6.5 × 1–2 μm ( x̄ = 5 × 1.7 µm, n = 10), holoblastic, hyaline, branched, smooth, elongated, broadly filiform
to ampulliform. Conidia 9–14 × 6–9 μm ( x̄ = 11 × 7.5 µm,
n = 45), oval to ellipsoid, 1-septate, thick-walled, not constricted at the septum, reddish brown, verruculose.
Culture characteristics: Conidia germinating on MEA
within 24 h at room temperature and germ tubes produced
13
Fungal Diversity
Fig. 14 Phylogram generated
from maximum likelihood
analysis based on combined
dataset of LSU, ITS, SSU and
TEF1 sequence data. Ninetyfive strains are included in the
combined sequence analysis,
which comprise 3280 characters with gaps. Tree topology
of the ML analysis was similar
to the BYPP. The best scoring
RAxML tree with a final likelihood value of − 17325.094501
is presented. The matrix had
1124 distinct alignment patterns, with 34.85% of undetermined characters or gaps.
Estimated base frequencies
were as follows: A = 0.241071,
C = 0.243154, G = 0.273420,
T = 0.242354; substitution rates:
AC = 1.437040, AG = 2.295778,
AT = 1.367419, CG = 0.976650,
CT = 7.785933, GT = 1.000000;
gamma distribution shape
parameter α = 0.200569. Bootstrap support values for ML
equal to or greater than 60% and
BYPP equal to or greater than
0.90 are given above the nodes.
Newly generated sequences
are in dark red bold and type
species are in bold. Pleospora
tarda (CBS 714.68), Pleospora
herbarum (CBS 191.86) and
Pleospora herbarum (IT 956)
are used as outgroup taxa
from the apex. Colonies on MEA circular or irregular, mycelium slightly flattened, filamentous, cultures white to creamy
white on surface and pale brown to brown from the centre
of the colony, olivaceous to olivaceous brown appearing
as concentric rings pattern with white margin in reverse
(Fig. 19a).
13
Pre-screening for antimicrobial activity: Chromolaenicola chiangraiensis (MFLUCC 17-1493) showed no inhibition against E. coli, B. subtilis and M. plumbeus.
Material examined: THAILAND, Chiang Rai Province,
Doi Pui, on dead stems of Chromolaena odorata, 2 February 2017, A. Mapook (DP92, MFLU 20-0301, holotype);
ex-type culture MFLUCC 17-1493.
Fungal Diversity
Fig. 15 Chromolaenicola
chiangraiensis (holotype) a,
b Appearance of colonies on
substrate. c–i Conidia with
conidiogenous cells. Scale
bars: a, b = 500 µm, c = 10 µm,
d–i = 5 µm
GenBank numbers: LSU: MN325005, ITS: MN325017
SSU: MN325011, TEF1: MN335650, RPB2: MN335655
Notes: In a BLASTn search of NCBI GenBank, the closest match of the ITS, LSU and TEF1 sequences of Chromolaenicola chiangraiensis (MFLUCC 17-1493, ex-holotype) is Cylindroaseptospora siamensis (strain MFLUCC
17-2527) with 100% (MK347760), 99.88% (MK347976)
and 99.32% (MK360048) similarity, respectively, while
the closest match with the SSU sequence with 99.89%
similarity was Didymosphaeria variabile (strain STE-U
6311, NG_064914). In the present phylogenetic analysis,
Chromolaenicola chiangraiensis (MFLUCC 17-1493) is
closely related to C. siamensis MFLUCC 17-2527 with
high bootstrap support (100% ML and 1.00 BYPP, Fig. 14).
Chromolaenicola chiangraiensis is similar to C. siamensis
(= Cylindroaseptospora siamensis, MFLUCC 17–2527) in
having 1-septate, verruculose conidia but differs in having smaller conidiogenous cells (3.5–6.5 × 1–2 μm vs.
6.5–7.4 × 3.2–4.7 μm) and larger conidia (9–14 × 6–9 μm
vs. 7.2–9.4 × 5.4–6.5 μm) (Table 4). Therefore, we introduced a new species based on phylogeny and morphological
comparison.
Chromolaenicola lampangensis Mapook & K.D. Hyde, sp.
nov.
Index Fungorum number: IF557281, Facesoffungi number: FoF 07785; Fig. 16
Etymology: Referring to the location where the specimen
was collected, Lampang Province, Thailand.
Holotype: MFLU 20-0302
Saprobic on dead stems of Chromolaena odorata.
Sexual morph: Undetermined. Asexual morph: Conidiomata 150–230 µm high × (155–)170–270(–345) µm diam.
( x̄ = 190 × 215 µm, n = 15), pycnidial, solitary, immersed
to semi-immersed, uni-loculate, globose to obpyriform,
yellowish brown to brown. Ostiole central, papillate. Pycnidial wall (5–)10–20 µm wide, comprising 2–4 layers of
pale brown to light brown cells of textura angularis. Conidiophores reduced to conidiogenous cells. Conidiogenous
cells holoblastic, ampulliform, hyaline, unbranched. Conidia
12–15 × 4–6.5 μm ( x̄ = 13.5 × 5 µm, n = 40), oblong to oval,
hyaline to pale brown, aseptate when immature, becoming
reddish brown to brown and 1-septate when mature, not constricted at septum, thick-walled, finely verruculose.
Culture characteristics: Conidia germinating on MEA
within 48 h. at room temperature and germ tubes produced
from both ends. Colonies on MEA circular or irregular,
mycelium slightly flattened, filamentous, cultures creamy
white to yellow from the centre of the colony with white
to pale green on surface and pale brown to brown from the
13
Fungal Diversity
Table 5 Synopsis of sexual morph of Chromolaenicola species with similar morphological features discussed in this study
Species
Asocomata (µm)
Peridium (µm)
Asci (µm)
Ascospores (µm)
References
C. nanensis
(MFLUCC
17-1473)
C. thailandensis
(MFLUCC
17-1510)
210–230 × 200–220
(10–)15–20(–25)
(90–)110–145 × 10–12.5
16–20 × 7.5–9
This study
145–225(–250) × (150–
)175–240(–285)
10–20(–35)
90–160 × 10–14
16–24 × 9–11
This study
Fig. 16 Chromolaenicola
lampangensis (holotype) a,
b Appearance of conidiomata
on substrate. c Section through
conidioma. d Ostiole. e Peridium. f–h Conidiogenous cells
and developing conidia. i–m
Conidia. Scale bars: a = 500 µm,
b = 200 µm, c, d = 50 µm,
e = 20 µm, h = 10 µm, f, g,
i–m = 5 µm
centre of the colony, olivaceous to olivaceous brown appearing as concentric rings pattern with white margin in reverse
(Fig. 19b).
Pre-screening for antimicrobial activity: Chromolaenicola lampangensis (MFLUCC 17-1462) showed antimicrobial activity against M. plumbeus with a 14 mm inhibition
zone, observable as partial inhibition, when compared to
13
the positive control (16 mm), but no inhibition of B. subtilis
and E. coli.
Material examined: THAILAND, Lampang Province,
Chaehom, on dead stems of Chromolaena odorata, 24 September 2016, A. Mapook (JH8, MFLU 20-0302, holotype);
ex-type culture MFLUCC 17-1462.
GenBank numbers: LSU: MN325004, ITS: MN325016,
SSU: MN325010, TEF1: MN335649, RPB2: MN335654
Fungal Diversity
Notes: In a BLASTn search of NCBI GenBank, the closest match of the ITS sequence of Chromolaenicola lampangensis (MFLUCC 17-1462, ex-holotype) was Paraconiothyrium sp. with 98.75% similarity to the strain PNB15_1B1
(MH268018). The closest match with the LSU sequence
with 99.02% similarity was Coniothyrium nitidae (strain
CBS 119209, EU552112). The closest match with the TEF1
sequences with 98.97% similarity was Cylindroaseptospora
siamensis (strain MFLUCC 17-2527, MK360048), while
the closest match with the SSU sequence with 99.89%
similarity was Paraconiothyrium thysanolaenae (strain
MFLUCC 10-0550, NG_063570). In the present phylogenetic analysis, C. lampangensis (MFLUCC 17-1462)
is closely related to C. nanensis with high bootstrap support (90% ML and 1.00 BYPP, Fig. 14). Chromolaenicola
nanensis is introduced as a new species in this study and
found as the sexual morph in nature; however, we failed
to obtain the sexual morph of C. lampangensis. Our strain,
C. lampangensis is similar to C. siamensis (= Cylindroaseptospora siamensis, MFLUCC 17–2527) and C. chiangraiensis (MFLUCC 17-1493) in having 1-septate with
verruculose conidia but differs from C. siamensis in having larger conidiomata (150–230 × (155–)170–270(–345)
μm vs. 110–165 × 140–190 μm) with larger conidia
(12–15 × 4–6.5 μm vs. 7.2–9.4 × 5.4–6.5 μm), and differs from C. chiangraiensis in having narrower conidia
(12–15 × 4–6.5 μm vs. 9–14 × 6–9 μm) (Table 4). A comparison of the RPB2 gene region of C. lampangensis and
C. nanensis reveals 20 base pair differences (1.87%) across
1068 nucleotides. Therefore, we introduce a new species
based on phylogeny and morphological comparison.
Chromolaenicola nanensis Mapook & K.D. Hyde, sp. nov.
Index Fungorum number: IF557282, Facesoffungi number: FoF 07786; Fig. 17
Etymology: Referring to the location where the specimen
was collected, Nan Province, Thailand.
Holotype: MFLU 20-0304
Saprobic on dead stems of Chromolaena odorata. Sexual
morph: Ascomata 210–230 µm high × 200–220 µm diam.
( x̄ = 216 × 211 µm, n = 5), immersed to semi-immersed,
solitary or scattered, appearing as small dark spots, coriaceous, globose to subglobose, brown to dark brown. Ostiolar
neck protruding. Peridium (10–)15–20(–25) µm wide, 3–4
layers, comprising dark brown cells of textura angularis.
Hamathecium comprising 1.5–2.5(–3) µm wide, cylindrical to filiform, septate, branching pseudoparaphyses. Asci
(90–)110–145 × 10–12.5 µm ( x̄ = 120 × 11 µm, n = 20),
6–8-spored, bitunicate, cylindrical, straight or slightly
curved, apically rounded, pedicellate with an ocular chamber. Ascospores 16–20 × 7.5–9 µm ( x̄ = 18.5 × 8 µm, n = 45),
slightly overlapping, uni-seriate, initially hyaline, 1-septate when immature, becoming reddish brown to brown
at maturity, ellipsoid to broadly fusiform, muriform, and
3-transversely septate, with 1-vertical septum when mature,
constricted at the central septum, straight or slightly curved,
without gelatinous sheath. Asexual morph: Undetermined.
Culture characteristics: Ascospores germinating on
MEA within 24 h at room temperature and germ tubes produced from both ends. Colonies on MEA circular, mycelium
slightly flattened, filamentous, cultures white to pale green
on surface, creamy white to yellow in reverse from the centre
of the colony, light green to olivaceous appearing as concentric rings with white margin (Fig. 19c).
Pre-screening for antimicrobial activity: Chromolaenicola nanensis (MFLUCC 17-1473) showed antimicrobial
activity against M. plumbeus with a 12 mm inhibition zone,
observable as partial inhibition, when compared to the positive control (17 mm), but no inhibition of B. subtilis and E.
coli.
Material examined: THAILAND, Nan Province, Doi
Phu Kha, on dead stems of Chromolaena odorata, 23 September 2016, A. Mapook (DPK5, MFLU 20-0304, holotype); ex-type culture MFLUCC 17-1473; (DPK11, MFLU
20-0303); living culture MFLUCC 17-1477.
GenBank numbers: LSU: MN325003, MN325002, ITS:
MN325015, MN325014, SSU: MN325009, MN325008,
TEF1: MN335648, MN335647, RPB2: MN335653
Notes: In a BLASTn search of NCBI GenBank, the closest match of the ITS sequence of Chromolaenicola nanensis (MFLUCC 17-1473, ex-holotype) was Paraconiothyrium sp. with 98.75% similarity to the strain PNB15_1B1
(MH268018). The closest match with the LSU sequence
with 99.01% similarity was Coniothyrium nitidae (strain
CBS 119209, EU552112). The closest match with the TEF1
sequence with 98.68% similarity was Cylindroaseptospora
siamensis (strain MFLUCC 17-2527, MK360048), while the
closest match with the SSU sequence with 99.79% similarity
was Paraconiothyrium thysanolaenae (MFLUCC 10-0550,
NG_063570). Chromolaenicola nanensis (MFLUCC
17-1473) is found as sexual morph in nature and we could
not obtain its asexual morph in culture. The strain is similar
to C. thailandensis (MFLUCC 17-1510) but differs in having
slightly smaller ascomata (210–230 × 200–220 µm vs. 145–2
25(–250) × (150–)175–240(–285) µm), slightly smaller asci
[(90–)110–145 × 10–12.5 µm vs. 90–160 × 10–14 µm] and
smaller ascospores (16–20 × 7.5–9 µm vs. 16–24 × 9–11 µm)
(Table 5). In the present phylogenetic analysis, C. nanensis
groups with C. lampangensis (MFLUCC 17-1462) which is
found as an asexual morph in nature. However, they differ in
culture characteristics on MEA (Fig. 19). A comparison of
the RPB2 gene region of C. nanensis and C. lampangensis
reveals 20 base pair differences (1.87%) across 1068 nucleotides. Therefore, C. nanensis is described as a new species
based on phylogeny and morphological comparison.
13
Fungal Diversity
Fig. 17 Chromolaenicola
nanensis (holotype) a, b
Appearance of immersed ascomata on substrate. c Section
through ascoma. d Ostiole. e
Peridium. f Pseudoparaphyses.
g–j Immature and mature asci.
k–q Ascospores. Scale bars: a,
b = 200 µm, c, g–j = 50 µm, d,
e = 20 µm, f, k–q = 10 µm
Chromolaenicola siamensis (Jayasiri, E.B.G. Jones & K.D.
Hyde) Mapook & K.D. Hyde, comb. nov.
Index Fungorum number: IF557283, Facesoffungi number: FoF 07787
13
≡ Cylindroaseptospora siamensis Jayasiri, E.B.G. Jones
& K.D. Hyde, in Jayasiri et al., Mycosphere 10(1): 68 (2019)
Holotype: THAILAND, Lampang Province on decaying pod of Leucaena sp. (Fabaceae), 18 August 2017, S.C.
Fungal Diversity
Fig. 18 Chromolaenicola
thailandensis (holotype)
a, b Appearance of ascomata on substrate. c Section
through ascoma. d Ostiole. e
Peridium. f Pseudoparaphyses.
g–j Immature and mature asci.
k–p Ascospores. Scale bars: a,
b = 500 µm, c, g–j = 50 µm, d,
e = 20 µm, f, k–p = 10 µm
Jayasiri, C 329 (MFLU 18–2147, holotype; KUN-HKAS
102427, isotype); ex-type culture MFLUCC 17–2527,
KUMCC 18–0227.
Morphological description: See Jayasiri et al. (2019)
Notes: In our multigene phylogenetic study, Cylindroaseptospora siamensis (MFLUCC 17-2527) was in a clade
separate from the type species, Cylindroaseptospora leucaenae (MFLUCC 17-2424) and clustered with Chromolaenicola chiangraiensis (MFLUCC 17-1493) with high bootstrap
support (100% ML and 1.00 BYPP, Fig. 14). The species
also shares similar morphological characters with other
Chromolaenicola spp., such as ampulliform conidiogenous
cells and globose or subglobose to oval or ellipsoid conidia,
hyaline to pale brown, aseptate when immature, becoming
dark and 1-septate at maturity, thick-walled, not constricted
at the septum, and verruculose, while Cy. leucaenae has
cylindrical conidia that are hyaline, aseptate, smooth and
thin-walled. Therefore, we transfer Cylindroaseptospora
siamensis as Chromolaenicola siamensis.
Chromolaenicola thailandensis Mapook & K.D. Hyde, sp.
nov.
13
Fungal Diversity
Index Fungorum number: IF557284, Facesoffungi number: FoF 07788; Fig. 18
Etymology: Named after Thailand, where the fungus was
discovered.
Holotype: MFLU 20-0306
Saprobic on dead stems of Chromolaena odorata. Sexual morph: Ascomata 145–225(–250) µm
high × (150–)175–240(–285) µm diam. ( x̄ = 195 × 215 µm,
n = 5), immersed, solitary or scattered, appearing as dark
spots, coriaceous, globose, light brown to brown. Ostiole central. Peridium 10–20(–35) µm wide, 3–4 layers at
side, thin-walled, inner layers comprising hyaline to pale
brown cells of textura angularis, outer layers comprising brown cells of textura intricata. Hamathecium comprising (1.5–)2–2.5(–3) µm wide, cylindrical, septate,
branching pseudoparaphyses. Asci 90–160 × 10–14 µm
( x̄ = 124 × 11.5 µm, n = 20), 8-spored, bitunicate, cylindrical, straight or slightly curved, apically rounded, pedicellate. Ascospores 16–24 × 9–11 µm ( x̄ = 21 × 10 µm, n = 35),
uni-seriate, initially hyaline to pale brown, 1-septate when
immature, becoming reddish brown to brown at maturity,
ellipsoid to broadly fusiform, muriform, 3-transversely septate, with none or 1-vertical septum, constricted at the central septum, straight or slightly curved, without gelatinous
sheath. Asexual morph: Undetermined.
Culture characteristics: Ascospores germinating on
MEA within 24 h. at room temperature and germ tubes produced from both ends. Colonies on MEA circular, mycelium
flattened, curled, creamy white to pale yellow with white
margin on surface, pale orange from the centre of the colony
with white margin in reverse (Fig. 19d).
Pre-screening for antimicrobial activity: Chromolaenicola thailandensis (MFLUCC 17-1510) showed no inhibition of E. coli, B. subtilis and M. plumbeus.
Material examined: THAILAND, Mae Hong Son Province, Mae Yen, Pai, on dead stems of Chromolaena odorata,
25 June 2016, A. Mapook (MY11, MFLU 20-0306, holotype); ex-type culture MFLUCC 17-1510; Nan Province,
Doi Phu Kha, on dead stems of C. odorata, 23 September
2016, A. Mapook (DPK7, MFLU 20-0305); living culture
MFLUCC 17-1475.
GenBank numbers: LSU: MN325006, MN325007, ITS:
MN325018, MN325019, SSU: MN325012, MN325013,
TEF1: MN335651, MN335652, RPB2: MN335656
Notes: In a BLASTn search of NCBI GenBank, the closest match of the ITS and TEF sequences of Chromolaenicola
thailandensis (MFLUCC 17-1510, ex-holotype) with 100%
(MK347760) and 98.98% (MK360048) similarity, respectively, was Cylindroaseptospora siamensis strain MFLUCC
17-2527. The closest match with the LSU sequence with
99.19% similarity was Alloconiothyrium aptrootii (strain
CBS 981.95, JX496235), while the closest match of the SSU
sequence with 99.89% similarity was Paraconiothyrium thysanolaenae (strain MFLUCC 10-0550, NG_063570). In the
present phylogenetic analysis, C. thailandensis (MFLUCC
17-1510) forms a sister taxon with other Chromolaenicola
species (Fig. 14). The strain is found as a sexual morph in
nature and we could not obtain its asexual morph in culture.
Chromolaenicola thailandensis is similar to C. nanensis in
having uni-seriate ascospores that are 1-septate when immature, becoming reddish brown to brown at maturity, ellipsoid
to broadly fusiform, muriform, and 3-transversely septate,
with 1-vertical septum when mature, constricted at the central septum, without gelatinous sheath, but differs in having slightly larger ascomata (145–225(–250) × (150–)175–
240(–285) µm vs. 210–230 × 200–220 µm), slightly larger
asci (90–160 × 10–14 µm vs. (90–)110–145 × 10–12.5 µm)
and slightly larger ascospores (16–24 × 9–11 µm vs.
16–20 × 7.5–9 µm) (Table 5). A comparison of the TEF1
gene region of C. thailandensis and C. nanensis reveals 12
base pair differences (1.5%) across 798 nucleotides. Therefore, C. thailandensis is described as a new species based on
phylogeny and morphological comparison.
Montagnula Berl., Icon. fung. (Abellini) 2: 68. 1896.
Montagnula was introduced by Berlese (1896) to accommodate M. infernalis as the type species together with M.
gigantean based on morphology. The genus comprises saprobes growing on dead wood, branches, stems, bark and
Fig. 19 Culture characteristics on MEA: a Chromolaenicola chiangraiensis (MFLUCC 17-1493). b Chromolaenicola lampangensis
(MFLUCC 17-1462). c Chromolaenicola nanensis (MFLUCC 17-1473). d Chromolaenicola thailandensis (MFLUCC 17-1510)
13
Fungal Diversity
leaves, which were placed in the family Didymosphaeriaceae
by Ariyawansa et al. (2014b). Liu et al. (2015) introduced
a new species, M. graminicola and Hongsanan et al. (2015)
introduced two new species M. bellevaliae and M. scabiosae which were collected in Italy. Wanasinghe et al. (2016)
introduced a new species, M. saikhuensis from Thailand and
synonymized two Munkovalsaria species (M. donacina and
M. appendiculata) under the genus Montagnula. Hyde et al.
(2016b) introduced M. cirsii from Italy, and M. jonesii was
introduced as a new species by Tennakoon et al. (2016).
Niranjan and Sarma (2018) introduced M. vakrabeejae as a
new species from India, based on morphological comparison. Tibpromma et al. (2018) introduced M. krabiensis from
Pandanaceae in Thailand. Presently 40 epithets are listed in
Index Fungorum (2020). In this study, four new Montagnula
species are introduced, based on morphology and molecular
data, together with descriptions and illustrations (Figs. 21,
22, 23, 24). A phylogenetic tree based on combined LSU,
ITS, SSU and TEF1 sequence data is presented in Fig. 20.
Montagnula chiangraiensis Mapook & K.D. Hyde, sp. nov.
Index Fungorum number: IF557296, Facesoffungi number: FoF 07789; Fig. 21
Etymology: Referring to the location where the specimen
was collected, Chiang Rai Province, Thailand.
Holotype: MFLU 20-0322
Saprobic on dead stems of Chromolaena odorata. Sexual morph: Ascomata (130–)150–220 µm
high × (170–)200–230 µm diam. ( x̄ = 172 × 207 µm, n = 10),
immersed to erumpent, solitary, scattered, globose, coriaceous, reddish brown to brown. Ostiole papillate, protruding from substratum. Peridium (7–)10–20 µm wide, comprising 2–3-layers of thin-walled, reddish brown to light
brown cells of textura angularis. Hamathecium comprising
1.5–3.5 µm wide, cylindrical to filiform, septate, branching
Fig. 20 Phylogram generated
from maximum likelihood
analysis based on combined
dataset of LSU, ITS, SSU and
TEF1 sequence data. Twenty
strains are included in the
combined sequence analysis,
which comprise 4029 characters with gaps. Tree topology
of the ML analysis was similar
to the BYPP. The best scoring
RAxML tree with a final likelihood value of − 12550.757272
is presented. The matrix had
911 distinct alignment patterns, with 34.97% of undetermined characters or gaps.
Estimated base frequencies
were as follows: A = 0.244653,
C = 0.247314, G = 0.272188,
T = 0.235846; substitution rates:
AC = 1.302293, AG = 2.261934,
AT = 1.509666, CG = 1.082059,
CT = 5.323888, GT = 1.000000;
gamma distribution shape
parameter α = 0.176588. Bootstrap support values for ML
equal to or greater than 60% and
BYPP equal to or greater than
0.90 are given above the nodes.
Newly generated sequences
are in dark red bold and type
species are in bold. Pleospora
herbarum (IT 956) is used as
outgroup taxon
13
Fungal Diversity
Fig. 21 Montagnula chiangraiensis (holotype) a,
b Appearance of immersed
ascomata on substrate. c Section
through ascoma. d Ostiole. e
Peridium. f Pseudoparaphyses.
g–j Immature and mature asci.
k–o Ascospores (immature and
mature) with terminal appendages. p Ascospores in Indian
ink. Scale bars: a = 500 µm,
b = 200 µm, c = 50 µm, d,
g–j = 20 µm, e, k–p = 10 µm,
f = 5 µm
pseudoparaphyses. Asci (45–)60–75 × (6–)8–11 µm ( x̄
= 65 × 9.5 µm, n = 20), 8-spored, bitunicate, fissitunicate, cylindric-clavate, slightly curved, with a short, bulbous long pedicel, with small ocular chamber. Ascospores
11–15 × 4–6 µm ( x̄ = 12 × 5 µm, n = 25), overlapping
2–3-seriate, pale brown when immature and dark reddish
brown to dark brown when mature, broadly fusiform to
13
ellipsoid, 1-septate, constricted at the septum, slightly widest at the upper cell and tapering towards ends, straight to
slightly curved, with small guttules; sheath drawn out to
form polar appendages 3–6 × 1–2.5 µm ( x̄ = 5 × 2 µm, n = 10),
from both ends of the ascospores, straight or slightly curved.
Asexual morph: Undetermined.
Fungal Diversity
Fig. 22 Montagnula chromolaenae (holotype) a, b Appearance of immersed ascomata on
substrate. c Section through
ascoma. d Ostiole. e Peridium.
f Pseudoparaphyses. g–j Asci.
k–p Ascospores with terminal
appendages. Scale bars: a,
b = 200 µm, c = 50 µm, d, e,
g–j = 20 µm, f, k–p = 10 µm
Culture characteristics: Ascospores germinating on
MEA within 24 h. at room temperature and germ tubes produced from both ends. Colonies on MEA irregular, mycelium slightly flattened, undulate, cultures white to creamy
white on surface, brown to dark brown in reverse from the
centre of the colony, creamy white at margin with white
mycelium appearing as small spots spreading around the
colony (Fig. 25a).
Pre-screening for antimicrobial activity: Montagnula
chiangraiensis (MFLUCC 17-1420) showed antimicrobial
activity against B. subtilis with an 8 mm inhibition zone and
against M. plumbeus with an 11 mm inhibition zone, observable as partial inhibition, when compared to the positive
control (26 mm and 17 mm, respectively), but no inhibition
of E. coli.
13
Fungal Diversity
Fig. 23 Montagnula chromolaenicola (holotype) a, b
Appearance of ascomata on
substrate. c Section through
ascoma. d Ostiole. e Peridium. f Pseudoparaphyses.
g–j Immature and mature asci.
k–p Ascospores. Scale bars:
a, b = 500 µm, c = 100 µm,
g–j = 50 µm, d, e = 20 µm, f,
k–p = 5 µm
Material examined: THAILAND, Chiang Rai Province,
Doi Pui, on dead stems of Chromolaena odorata, 8 July
2015, A. Mapook, (DP1, MFLU 20-0322, holotype); extype culture MFLUCC 17-1420.
GenBank numbers: LSU: MT214443, ITS: MT214349,
SSU: MT214397
Notes: In a BLASTn search of NCBI GenBank, the closest match of the ITS sequence of Montagnula chiangraiensis
(MFLUCC 17-1420, ex-holotype) with 98.91% similarity
13
was M. appendiculata (strain M-1, DQ435529). The closest
match with the LSU sequence with 98.34% similarity was
M. aloes (strain CBS 132531, NG_042676), while the closest match with the SSU sequence with 97.76% similarity
was Montagnula sp. (strain MFLUCC 11-0576, KJ188099).
In the present phylogenetic analysis, M. chiangraiensis is
closely related to M. appendiculata with high bootstrap
support (70% ML and 0.98 BYPP, Fig. 20). Montagnula
chiangraiensis differs from M. appendiculata in its culture
Fungal Diversity
Fig. 24 Montagnula thailandica (holotype) a Appearance
of ascomata on substrate. b
Section through ascoma with
ostiole. c Peridium. d Pseudoparaphyses. e–h Immature and
mature asci. i–n Ascospores
(immature and mature). Scale
bars: a = 500 µm, b = 100 µm,
e–h = 50 µm, c = 20 µm,
i–n = 10 µm, d = 5 µm
Fig. 25 Culture characteristics on MEA: a Montagnula chiangraiensis (MFLUCC 17-1420). b Montagnula chromolaenae (MFLUCC
17-1435). c Montagnula chromolaenicola (MFLUCC 17-1469). d Montagnula thailandica (MFLUCC 17-1508)
characteristics on MEA. Montagnula chiangraiensis has
white mycelium appearing as small spots spreading around
the colony, which are not reported for M. appendiculata
(Aptroot 2004). Moreover, M. chiangraiensis has dark reddish brown to dark brown ascospores, while M. appendiculata has yellowish brown to brown ascospores. However,
we could not compare size of asci as this feature was not
given for the holotype of M. appendiculata (Aptroot 2004)
(Table 6). A comparison of the ITS (+5.8S) gene region
of M. chiangraiensis and M. appendiculata reveals 14 base
pair differences (1.53%) across 914 nucleotides. Therefore,
13
Fungal Diversity
Table 6 Synopsis of Montagnula species with similar morphological features discussed in this study
Species
Asocomata (µm)
Peridium (µm)
Asci (µm)
Ascospores (µm)
Polar appendages
(µm)
References
M. appendiculata
(CBS 109027)
M. chiangraiensis (MFLUCC
17-1420, DP1)
M. chromolaenae (MFLUCC
17-1435, DP18)
M. chromolaenicola (MFLUCC
17-1469, MY1)
M. donacina
(HVVV01)
100–200 diam.
–
Not reported
12–15 × 4–5
4–7 × 1.5–2.5
Aptroot (2004)
(130–)150–220
high × (170–
)200–230 diam.
170–175(–230)
high × (140–
)170–190 diam.
300–320
high × 215–310
diam.
–
(7–)10–20
(45–)60–
75 × (6–)8–11
11–15 × 4–6
3–6 × 1–2.5
This study
10–25
85–105 × 9–15
15–16.5 × 5–6
6–11 × (1.8–)2–3
This study
10–25
80–100 × 10–13
15–17 × 5–6.5
1-septate, without
polar appendages
This study
–
Not reported
M. donacina
500 diam.
–
90–100 × 12–13
M. graminicola
(MFLUCC
13-0352)
37–117.22 diam.
14.9–16
(45–)50–132(–
137) × (6–)8–
13(–15)
10–20
70–100 × 10–12
(13.6–)14.8–15.2(– 1-septate, without Pitt et al. (2014)
mucilaginous
17.3) × (6.6–
sheath
)7.5–7.7(–8.3)
12–15 × 4
1-septate, without Saccardo (1882)
mucilaginous
sheath
Liu et al. (2015)
1-septate, sur(7.8–)9.8–13(–
rounded by a
15) × (2.8–)3.8–
mucilaginous
5.5(–6.5)
sheath
12–16 × 4–6
1-septate, without Wanasinghe et al.
(2016)
mucilaginous
sheath
(12–)14–17 × 4.5– 1-septate, without This study
7.5
polar appendages
400–450
M. saikhuensis
high × 400–500
(MFLUCC
diam.
16-0315)
(315–)405–415
M. thailandica
high × 330–350
(MFLUCC
diam.
17-1508, FHP16)
(10–)15–20(–30) 80–100 × 9–15
M. chiangraiensis is described as a new species based on
phylogeny and morphological comparison.
Montagnula chromolaenae Mapook & K.D. Hyde, sp. nov.
Index Fungorum number: IF557297, Facesoffungi number: FoF 07790; Fig. 22
Etymology: Name reflects the host genus Chromolaena,
from which this species was isolated.
Holotype: MFLU 20-0323
Saprobic on dead stems of Chromolaena odorata. Sexual morph: Ascomata 170–175(–230) µm
high × (140–)170–190 µm diam. ( x̄ = 185 × 172 µm, n = 5),
immersed, solitary or scattered, globose to subglobose,
coriaceous, reddish brown to brown. Ostiole central,
papillate. Peridium 10–25 µm wide, comprising of 1–2
layers of pale brown to yellowish brown cells of textura
angularis. Hamathecium comprising (2.5–)3–4(–5) µm
wide, cylindrical, septate, branching pseudoparaphyses.
Asci 85–105 × 9–15 ( x̄ = 93 × 12 µm, n = 15), bitunicate,
(4–)6–8-spored, cylindric-clavate, straight or slightly curved,
long pedicel. Ascospores 15–16.5 × 5–6 ( x̄ = 16 × 5.5 µm,
n = 15), overlapping bi-seriate, hyaline to pale brown
when immature and yellowish brown to light brown when
mature, broadly fusiform to ellipsoid, 1-septate, constricted
13
at the septum, slightly widest at the upper cell and tapering
towards ends, straight to slightly curved; sheath drawn out to
form polar appendages 6–11 µm long × (1.8–)2–3 µm wide
( x̄ = 8 × 2.5 µm, n = 20), from both ends of the ascospores,
straight or slightly curved. Asexual morph: Undetermined.
Culture characteristics: Ascospores germinating on
MEA within 24 h. at room temperature and germ tubes produced from both ends. Colonies on MEA irregular, mycelium slightly flattened, undulate, cultures white to creamy
white on surface, brown to dark brown in reverse (Fig. 25b).
Pre-screening for antimicrobial activity: Montagnula
chromolaenae (MFLUCC 17-1435) showed antimicrobial
activity against M. plumbeus with an 18 mm inhibition zone,
observable as partial inhibition, when compared to the positive control (17 mm), but no inhibition of B. subtilis and E.
coli.
Material examined: THAILAND, Chiang Rai Province,
Doi Pui, on dead stems of Chromolaena odorata, 5 August
2015, A. Mapook (DP18, MFLU 20-0323, holotype); extype culture MFLUCC 17-1435.
GenBank numbers: LSU: MT214444, ITS: MT214350,
SSU: MT214398
Notes: In a BLASTn search of NCBI GenBank, the closest match of the ITS sequence of Montagnula chromolaenae
Fungal Diversity
(MFLUCC 17-1435, ex-holotype) with 97.60% similarity
was M. appendiculata (strain M-1, DQ435529). The closest match with the LSU sequence with 99.09% similarity
was Montagnula sp. (strain MFLUCC 11-0576, KJ188098),
while the closest match of the SSU sequence with 99.16%
similarity was Montagnula sp. (strain MFLUCC 11-0576,
KJ188099). In the present phylogenetic analysis, M. chromolaenae is closely related to M. appendiculata and M. chiangraiensis (Fig. 20). However, M. chromolaenae differs
from M. appendiculata in having larger ascomata (170–1
75(–230) × (140–)170–190 µm vs. 100–200 µm diam.) and
larger ascospores (15–16.5 × 5–6 µm vs. 12–15 × 4–5 µm)
with longer polar appendages (6–11 × (1.8–)2–3 µm vs.
4–7 × 1.5–2.5 µm) (Table 6). A comparison of the ITS
(+5.8S) gene region of M. chromolaenae and M. appendiculata reveals 25 base pair differences (2.7%) across 914 nucleotides. Therefore, M. chromolaenae is described as a new
species based on phylogeny and morphological comparison.
Montagnula chromolaenicola Mapook & K.D. Hyde, sp.
nov.
Index Fungorum number: IF557298, Facesoffungi number: FoF 07791; Fig. 23
Etymology: Name reflects the host genus Chromolaena,
on which this species was growing.
Holotype: MFLU 20-0324
Saprobic on dead stems of Chromolaena odorata. Sexual
morph: Ascomata 300–320 µm high × 215–310 µm diam.
(x̅ = 310 × 275 µm, n = 5), semi-immersed to erumpent,
solitary or scattered, globose to obpyriform, coriaceous,
brown to dark brown. Ostiole papillate, protruding from
substratum. Peridium 10–25 µm wide, comprising several
layers of thin-walled, pale brown to brown cells of textura
angularis. Hamathecium comprising 1–2 µm wide, cylindrical to filiform, septate, branching pseudoparaphyses.
Asci 80–100 × 10–13 µm ( x̄ = 90 × 12 µm, n = 10), 8-spored,
bitunicate, elongate-clavate, slightly curved, long pedicel.
Ascospores 15–17 × 5–6.5 µm ( x̄ = 15.5 × 6 µm, n = 15), overlapping 1–2-seriate, pale brown to yellowish brown when
immature, becoming brown to dark brown when mature,
broadly fusiform to ellipsoid, 1-septate, constricted at the
septum, slightly wider upper cell and tapering towards ends,
straight to slightly curved, without terminal appendages.
Asexual morph: Undetermined.
Culture characteristics: Ascospores germinating on
MEA within 24 h. at room temperature and germ tubes
produced from the apex. Colonies on MEA circular, mycelium slightly flattened, entire, cultures grayish green to
light brown on surface and dark brown to black in reverse
(Fig. 25c).
Pre-screening for antimicrobial activity: Montagnula
chromolaenicola (MFLUCC 17-1469) showed no inhibition
of E. coli, B. subtilis and M. plumbeus.
Material examined: THAILAND, Mae Hong Son Province, Mae Yen, Pai, on dead stems of Chromolaena odorata,
25 June 2016, A. Mapook (MY1, MFLU 20-0324, holotype); ex-type culture MFLUCC 17-1469.
GenBank numbers: LSU: MT214445, ITS: MT214351,
SSU: MT214399, TEF1: MT235773, RPB2: MT235809
Notes: In a BLASTn search of NCBI GenBank, the closest match of the ITS sequence of Montagnula chromolaenicola (MFLUCC 17-1469, ex-holotype) with 99.82% similarity was Aporospora terricola (strain 2711, EU272515).
The closest match with the LSU sequence with 99.22%
similarity was Munkovalsaria donacina (strain HVVV01,
KJ628377). The closest match with the SSU sequence with
99.00% similarity was Didymocrea sadasivanii (strain CBS
438, DQ384066), while the closest match with the TEF1
sequence with 97.05% similarity was Montagnula sp. (strain
UTHSC: DI16-251, LT797091). In the present phylogenetic
analysis, M. chromolaenicola forms a separate branch and
clusters with M. donacina (Fig. 20). However, M. chromolaenicola differs from M. donacina in having smaller ascomata (300–320 × 215–310 µm vs. 500 µm diam.) and slightly
smaller asci (80–100 × 10–13 µm vs. 90–100 × 12–13 µm)
with longer ascospores (15–17 µm × 5–6.5 vs. 12–15 × 4 µm)
(Table 6). Therefore, M. chromolaenicola is described as
a new species based on phylogeny and morphological
comparison.
Montagnula thailandica Mapook & K.D. Hyde, sp. nov.
Index Fungorum number: IF557299, Facesoffungi number: FoF 07792; Fig. 24
Etymology: The name reflects the country, where the
specimen was collected, Thailand.
Holotype: MFLU 20-0325
Saprobic on dead stems of Chromolaena odorata. Sexual
morph: Ascomata (315–)405–415 µm high × 330–350 µm
diam. ( x̄ = 380 × 340 µm, n = 5), immersed to erumpent,
solitary or scattered, globose to obpyriform, coriaceous,
brown to dark brown. Ostiole papillate, protruding from
substratum. Peridium (10–)15–20(–30) µm wide, comprising several layers of thin-walled, pale brown to brown cells
of textura angularis. Hamathecium comprising 1–2.5 µm
wide, cylindrical to filiform, septate, branching pseudoparaphyses. Asci 80–100 × 9–15 µm ( x̄ = 90 × 11 µm, n = 15),
8-spored, bitunicate, fissitunicate, elongate-clavate, slightly
curved, long pedicel. Ascospores (12–)14–17 × 4.5–7.5 µm
( x̄ = 15 × 5.5 µm, n = 25), overlapping 1–2-seriate, hyaline
or pale brown to yellowish brown when immature and
becoming brown to reddish brown when mature, broadly
fusiform to ellipsoid, 1-septate, constricted at the septum,
slightly wider upper cell and tapering towards ends, straight
to slightly curved, without terminal appendages. Asexual
morph: Undetermined.
13
Fungal Diversity
Culture characteristics: Ascospores germinating on
MEA within 48 h. at room temperature and germ tubes produced from the apex. Colonies on MEA circular, mycelium
raised, velvety with moderately fluffy, filamentous at margin,
cultures grey with white from the centre of the colony on
surface and creamy white in reverse with pale greyish brown
margin (Fig. 25d).
Pre-screening for antimicrobial activity: Montagnula
thailandica (MFLUCC 17-1508) showed no inhibition of E.
coli, B. subtilis and M. plumbeus.
Material examined: THAILAND, Chiang Mai Province,
Fah Hom Pok, on dead stems of Chromolaena odorata, 27
September 2016, A. Mapook (FHP16, MFLU 20-0325,
holotype); ex-type culture MFLUCC 17-1508.
GenBank numbers: LSU: MT214446, ITS: MT214352,
SSU: MT214400, TEF1: MT235774, RPB2: MT235810
Notes: In a BLASTn search of NCBI GenBank, the closest match of the ITS sequence of Montagnula thailandica
(MFLUCC 17-1508, ex-holotype) with 99.29% similarity was Aporospora terricola (strain 2711, EU272515).
The closest match with SSU sequences with 98.91%
similarity was Didymocrea sadasivanii (strain CBS 438,
DQ384066). The closest match with the LSU sequence
with 99.22% similarity was Munkovalsaria donacina (strain
HVVV01, KJ628377), while the closest match with the
TEF1 sequence with 96.75% similarity was Paraconiothyrium cyclothyrioides (UTHSC: DI16-327, LT797124). In
the present phylogenetic analysis, M. thailandica forms a
separate branch and groups with Montagnula chromolaenicola, M. donacina, M. saikhuensis and M. graminicola
(Fig. 20). Our strain is similar to those Montagnula species in having long pedicellate asci and broadly fusiform
to ellipsoid, 1-septate ascospores, without terminal appendages. However, our strain differs from M. graminicola in
having larger ascomata [(315–)405–415 × 330–350 µm
vs. 37–117.22 µm diam.] and larger ascospores
[(12–)14–17 × 4.5–7.5 µm vs. (7.8–)9.8–13(–15) × (2.8–
)3.8–5.5(–6.5) µm], without gelatinous sheath, while M.
graminicola has ascospores surrounded by a mucilaginous
sheath and our strain also differs from M. saikhuensis in
having smaller ascomata [(315–)405–415 × 330–350 µm
vs. 400–450 × 400–500 µm) with slightly larger asci
(80–100 × 9–15 µm vs. 70–100 × 10–12 µm) and
slightly larger ascospores [(12–)14–17 × 4.5–7.5 µm
vs. 12–16 × 4–6 µm] (Table 6). Montagnula thailandica also differs from M. chromolaenicola in having larger ascomata [(315–)405–415 × 330–350 µm
vs. 300–320 × 215–310 µm], slightly wider asci
(80–100 × 9–15 µm vs. 80–100 × 10–13 µm), with brown to
reddish brown ascospores, while M. chromolaenicola has
brown to dark brown ascospores. A comparison of the ITS
(+5.8S) gene region of M. thailandica and M. chromolaenicola reveals 13 base pair differences (1.51%) across 858
13
nucleotides. Therefore, M. thailandica is described as a new
species based on phylogeny and morphological comparison.
Pseudopithomyces Ariyaw. & K.D. Hyde
Pseudopithomyces was introduced by Ariyawansa et al.
(2015) with P. chartarum as the type species, along with a
new species, P. palmicola. Crous et al. (2016) introduced a
new species, P. diversisporus from human toe nail with two
new combinations (P. atro-olivaceus and P. karoo). Hyde
et al. (2017) introduced a new species, P. kunmingensis from
China. Wanasinghe et al. (2018) introduced P. rosae from
Italy and Tibpromma et al. (2018) introduced P. pandanicola from Thailand, while Crous et al. (2018a) introduced P.
angolensis from leaf spot of unknown host plant in Angola.
Jayasiri et al. (2019) introduced a new species, P. entadae
from pod of Entada phaseoloides in Thailand. We present a
new host record for P. palmicola isolated from C. odorata,
together with a description and illustrations (Figs. 27, 28). A
phylogenetic tree based on combined ITS, LSU, RPB2 and
GAPDH sequence data is presented in Fig. 26.
Pseudopithomyces palmicola J.F. Li, Ariyaw. & K.D. Hyde,
in Ariyawansa et al., Fungal Divers. 75: 41 (2015)
Facesoffungi number: FoF 00939; Fig. 27
Saprobic on dead stems of Chromolaena odorata. Sexual
morph: Undetermined. Asexual morph: Hyphomycetous.
Colonies superficial or partly immersed on the host surface,
scattered, gregarious, dark brown to black. Mycelium consisting of septate, branched, smooth, thin-walled, hyaline
hyphae. Conidiophores reduced to conidiogenous cells.
Conidiogenous cells (4–)6.5–11(–15) × (1.9–)2.5–4 µm
( x̄ = 9 × 3 µm, n = 15), holoblastic, monoblastic, terminal,
hyaline, cylindrical. Conidia (6.5–)17–30 × (4.5–)8–25 µm
( x̄ = 23.5 × 16 µm, n = 50), globose or ellipsoid to amygdaloid, muriform, 2–3-transversely septate, with 1–2-vertical septa, slightly verruculose to echinulate, brown to dark
brown, sometimes slightly constricted at the septa with dark
bands at the septa.
Culture characteristics: Conidia germinating on MEA
within 24 h at room temperature and germ tubes produced
from both ends. Colonies on MEA circular, mycelium
slightly raised, velvety with moderately fluffy, entire, cultures pale brown to creamy brown on surface from the centre
of the colony with white margin, and brown to dark brown
in reverse from the centre of the colony with creamy white
margin (Fig. 28).
Pre-screening for antimicrobial activity: Pseudopithomyces palmicola (MFLUCC 17-1506) showed antimicrobial
activity against M. plumbeus with a 10 mm inhibition zone,
observable as partial inhibition, when compared to the positive control (18 mm), but no inhibition of B. subtilis and E.
coli.
Fungal Diversity
Fig. 26 Phylogram generated
from maximum likelihood
analysis based on combined
dataset of ITS, LSU, RPB2 and
GAPDH sequence data. Fortytwo strains are included in the
combined sequence analysis,
which comprise 3066 characters with gaps. Tree topology
of the ML analysis was similar
to the BYPP. The best scoring
RAxML tree with a final likelihood value of − 10725.898987
is presented. The matrix had
826 distinct alignment patterns, with 28.11% of undetermined characters or gaps.
Estimated base frequencies
were as follows: A = 0.237796,
C = 0.263846, G = 0.271148,
T = 0.227210; substitution rates:
AC = 1.281129, AG = 2.845304,
AT = 1.125930, CG = 1.002622,
CT = 6.844818, GT = 1.000000;
gamma distribution shape
parameter α = 0.149330. Bootstrap support values for ML
equal to or greater than 60% and
BYPP equal to or greater than
0.90 are given above the nodes.
Newly generated sequences
are in dark red bold and type
species are in bold. Laburnicola hawksworthii (MFLUCC
13-0602) and L. muriformis
(MFLUCC 16-0290) are used as
outgroup taxa
Known hosts and distribution: On leaves lesions of
Vitis vinifera (Vitaceae), Phaseolus vulgaris (Fabaceae),
Poa annua (Poaceae) and Fragaria sp. (Rosaceae) in Italy
(Liu et al. 2018); on dead leaves of Pandanus amaryllifolius
(Pandanaceae) in Thailand (Tibpromma et al. 2018); on dead
leaves of unidentified grass in China (Hyde et al. 2017b); on
leaves of Acoelorrhaphe wrightii (Arecaceae) in Thailand
(Ariyawansa et al. 2015).
Material examined: THAILAND, Chiang Mai Province, Fah Hom Pok, on dead stems of C. odorata, 27 September 2016, A. Mapook (FHP10, MFLU 20-0355); living
culture MFLUCC 17-1506 (new host record); Chiang Rai
Province, Doi Pui, on dead stems of C. odorata, 2 February
2017, A. Mapook (DP95, MFLU 20-0354); living culture
MFLUCC 17-1496.
GenBank numbers: LSU: MT214447, MT214448, ITS:
MT214353, MT214354, RPB2: MT235811, MT235812
13
Fungal Diversity
Fig. 27 Pseudopithomyces palmicola (new host record) a, b Appearance of colonies on substrate. c–f Conidia and conidiophores. g–l Conidia.
Scale bars: a = 200 µm, b = 100 µm, d–l = 10 µm, c = 5 µm
Notes: A phylogenetic analyses showed that two strains
MFLUCC 17-1506 and MFLUCC 17-1496 grouped within
the Pseudopithomyces palmicola clade (Fig. 26). In a
BLASTn search of NCBI GenBank, the closest match of the
ITS sequence of our strains with 100% similarity was Pseudopithomyces palmicola (strain DTO 391-A6, MN788110).
The closest match with the LSU sequence with 100% similarity was Pseudopithomyces sp. (strain C449, MK348017),
while the closest match with the RPB2 sequence with 100%
similarity was Pseudopithomyces palmicola (strain UC15,
MH249026). We therefore, identify our isolates as P. palmicola based on phylogenetic analyses. Morphological characters also indicated that our strains belong to the genus
Pseudopithomyces. In this study, we isolated P. palmicola
from Chromolaena odorata collected in Thailand, and the
isolates are introduced here as a new host record. We also
treat P. pandanicola (MFLUCC 18-0116) and P. kunmingensis (MFLUCC 17-0314) under P. palmicola based on
phylogeny with morphological comparison (Table 7).
13
Tremateia Kohlm., Volkm.-Kohlm. & O.E. Erikss., Bot.
Mar. 38(2): 165 (1995)
Tremateia was introduced by Kohlmeyer et al. (1995)
with T. halophila as the type species. Hyde et al. (2016b)
introduced two new species T. arundicola and T. guiyangensis which were collected from dead herbaceous stems,
based on morphology and phylogeny support. Feng et al.
(2019) introduced a new species, T. murispora from China.
We introduce two new Tremateia species from C. odorata,
together with descriptions and illustrations (Figs. 30, 31,
32). A phylogenetic tree based on combined SSU, LSU, ITS,
TEF1 and RPB2 sequence data is presented in Fig. 29.
Tremateia chiangraiensis Mapook & K.D. Hyde, sp. nov.
Index Fungorum number: IF557364, Facesoffungi numberx: FoF 07793; Fig. 30
Etymology: Referring to the location where the specimen
was collected, Chiang Rai Province, Thailand.
Holotype: MFLU 20-0374
Fungal Diversity
Fig. 28 Culture characteristic on MEA: Pseudopithomyces palmicola (MFLUCC 17-1506)
Saprobic on dead stems of Chromolaena odorata. Sexual
morph: Ascomata 220–250 µm high × 215–260(–280) µm
diam. ( x̄ = 230 × 240 µm, n = 10), immersed, solitary or scattered, appearing as small dark spots, coriaceous, globose,
reddish brown to brown. Ostiolar neck protruding. Peridium
10–20(–25) µm wide, 4–5 layers, inner layers comprising
hyaline to pale brown cells of textura epidermoidea, outer
layers comprising brown to dark brown cells of textura
angularis. Hamathecium comprising (1.8–)2–3 µm wide,
cylindrical to filiform, septate, branching pseudoparaphyses. Asci 90–120(–140) × 14–20 µm ( x̄ = 106.5 × 17.5 µm,
n = 15), 8-spored, bitunicate, fissitunicate, cylindric-clavate,
straight or slightly curved, apically rounded, pedicellate.
Ascospores 23–27.5 × 5–7 µm ( x̄ = 25 × 9 µm, n = 35),
overlapping, 1–2 seriate, initially hyaline, 1-septate when
immature, becoming golden-brown to brown at maturity,
ellipsoid to broadly fusiform, muriform, 5–7-transversely
septate, with 1 vertical septum, slightly constricted at the
central septum, straight or slightly curved, surrounded by
hyaline gelatinous sheath observed clearly when mounted
in Indian ink. Asexual morph: Undetermined.
Culture characteristics: Ascospores germinating on
MEA within 24 h. at room temperature and germ tubes produced from several cells. Colonies on MEA circular, mycelium slightly raised, filamentous, cultures white on surface,
white to creamy white in reverse (Fig. 33a).
Pre-screening for antimicrobial activity: Tremateia chiangraiensis (MFLUCC 17-1428) showed no inhibition of E.
coli, B. subtilis and M. plumbeus.
Material examined: THAILAND, Chiang Rai Province,
Huai Kang Pla waterfall, on dead stems of Chromolaena
odorata, 23 June 2015, A. Mapook (HKP4, MFLU 20-0374,
holotype); ex-type culture MFLUCC 17-1428, (HKP5,
MFLU 20-0375); living culture MFLUCC 17-1429.
GenBank numbers: LSU: MT214449, MT214450, ITS:
MT214355, MT214356, SSU: MT214401, MT214402,
TEF1: MT235775, MT235776, RPB2: MT235813,
MT235814
Notes: In a BLASTn search of NCBI GenBank, the
ITS sequence of Tremateia chiangraiensis (MFLUCC
17-1428, ex-holotype) with 97.66% similarity was T. guiyangensis (strain GZAAS01, KX274240). The LSU and
SSU sequences with 97.17% (KX274248) and 97.51%
(KX274254) similarity, respectively, were T. arundicola
strain MFLUCC 16-1275, while the closest match with the
TEF1 and RPB2 sequences with 97.90% (LT797094) and
90.02% (LT797014) similarity, respectively, were Kalmusia
Table 7 Morphology of Pseudopithomyces species with similar morphological features discussed in this study
Species
Conidiogenous cells (µm)
Conidia (µm)
Conidia septation
P. chartarum (MUCL 15905)
P. chartarum (UTHSC
05-2460, UTHSC 03-2472)
P. palmicola (MFLU
15-1474, holotype)
P. palmicola (CBS 143933,
CBS 143932, CBS 143935
and CBS 143934)
P. palmicola (MFLUCC
17-1496 and MFLUCC
17-1496)
P. palmicola (= P. pandanicola MFLUCC 18-0116)
P. palmicola (= P. kunmingensis MFLUCC 17-0314)
–
–
18–29 × 10–17
12–33 × 9–19
3–4-transverse 1–2 vertical Ariyawansa et al. (2015)
3 transverse, 1–2 vertical
da Cunha et al. (2014)
3.5–5.5 × 2.5–3.5
21.5–30.5 × 10–16.5
2–3 transverse, 1–3 vertical Ariyawansa et al. (2015)
–
3 transverse, 2 vertical
(11.5–)21.1(–
27.5) × (7.5–)11.7(–
16.5)
(6.5–)17–30 × (4.5–)8–25 2–3 transverse 1–2 vertical
(4–)6.5–11(–15) × (1.9–
)2.5–4
References
Liu et al. (2018)
This study
–
10–25 × 7–15
2–3 transverse, 1–2 vertical Tibpromma et al. (2018)
3–6 × 5–6
19–25 × 10–15
2–3 transverse, 1–2 vertical Hyde et al. (2017b)
13
Fungal Diversity
Fig. 29 Phylogram generated
from maximum likelihood
analysis based on combined
dataset of ITS, LSU, SSU,
TEF1 and RPB2 sequence data.
Twelve strains are included in
the combined sequence analysis,
which comprise 4634 characters with gaps. Tree topology
of the ML analysis was similar
to the BYPP. The best scoring
RAxML tree with a final likelihood value of − 10566.353641
is presented. The matrix had
559 distinct alignment patterns, with 22.71% of undetermined characters or gaps.
Estimated base frequencies
were as follows: A = 0.235911,
C = 0.270697, G = 0.272162,
T = 0.221230; substitution rates:
AC = 1.376817, AG = 3.496002,
AT = 1.093429, CG = 1.659156,
CT = 6.548636, GT = 1.000000;
gamma distribution shape
parameter α = 0.147928. Bootstrap support values for ML
equal to or greater than 60% and
BYPP equal to or greater than
0.90 are given above the nodes.
Newly generated sequences
are in dark red bold and type
species are in bold. Bimuria
novae-zelandiae (CBS 107.79)
is used as outgroup taxon
sp. strain UTHSC: DI16-256. In the present phylogenetic
analysis, our two strains of T. chiangraiensis form a sister taxon with T. thailandensis (MFLUCC 17-1430) with
high bootstrap support (100% ML and 1.00 BYPP, Fig. 29).
However, T. chiangraiensis differs from T. thailandensis in having larger ascomata (220–250 × 215–260(–280)
µm vs. 200–230 × 150–175 µm) and slightly smaller asci
(90–120(–140) × 14–20 µm vs. (80–)90–125 × 15–25 µm)
and slightly smaller ascospores (23–27.5 × 5–7 µm vs.
20–26 × 5.5–9 µm) (Table 8). A comparison of the ITS
(+5.8S) gene region of T. chiangraiensis and T. thailandensis reveals 17 base pair differences (3%) across 565 nucleotides. Therefore, T. chiangraiensis is described as a new
species based on phylogeny and morphological comparison.
Tremateia chromolaenae Mapook & K.D. Hyde, sp. nov.
Index Fungorum number: IF557365, Facesoffungi number: FoF 07794; Fig. 31
Etymology: Name reflects the host genus Chromolaena,
from which this species was isolated.
13
Holotype: MFLU 20-0378
Saprobic on dead stems of Chromolaena odorata. Sexual
morph: Ascomata 150–220 µm high × 170–200(–220) µm
diam. ( x̄ = 183 × 191.5 µm, n = 10), immersed, solitary or
scattered, appearing as small dark spots, coriaceous, globose, dark brown to brown. Ostiolar neck protruding. Peridium 10–15 µm wide, comprising 2–3 layers, brown to dark
brown cells of textura angularis. Hamathecium comprising
2–3(–3.5) µm wide, broadly cylindrical, septate, branching pseudoparaphyses. Asci (65–)75–105 × (10–)15–20 µm
( x̄ = 89 × 15.5 µm, n = 15), 8-spored, bitunicate, fissitunicate,
clavate to cylindric-clavate, straight or slightly curved, pedicellate, apically rounded. Ascospores 16.5–20 × 6.5–10 µm
( x̄ = 18 × 8 µm, n = 30), overlapping, 1–2-seriate, initially
hyaline, 1-septate when immature, becoming golden-brown
at maturity, oval to ellipsoidal, muriform, 3–4-transversely
septate, with 1–2 vertical septa, upper part wider, slightly
constricted at the central septum, straight or slightly curved,
guttulate, surrounded by hyaline gelatinous sheath observed
Fungal Diversity
Fig. 30 Tremateia chiangraiensis (holotype) a, b Appearance of immersed ascomata on
substrate. c Section through
ascoma. d Ostiole. e Peridium.
f Pseudoparaphyses. g–j Asci.
k–p Ascospores q Ascospore
with gelatinous sheath in Indian
ink. Scale bars: a = 500 µm,
b = 200 µm, c, d, g–j = 50 µm,
e = 20 µm, k–q = 10 µm,
f = 5 µm
clearly when mounted in Indian ink. Asexual morph:
Undetermined.
Culture characteristics: Ascospores germinating on
MEA within 24 h at room temperature and germ tubes produced from several cells. Colonies on MEA circular, mycelium slightly raised, initially white, filamentous, becoming pale pinkish white on surface, brown to pale brown in
reverse from the centre of the colony, white to creamy white
at margin (Fig. 33b).
Pre-screening for antimicrobial activity: All isolates of
Tremateia chromolaenae showed no inhibition of E. coli, B.
subtilis and M. plumbeus.
Material examined: THAILAND, Chiang Rai Province,
Doi Pui, on dead stems of Chromolaena odorata, 5 August
2015, A. Mapook (DP26, MFLU 20-0376); living culture
MFLUCC 17-1442; Chiang Rai Province, Doi Pui, on dead
stems of C. odorata, 8 July 2015, A. Mapook (DP8, MFLU
20-0377); living culture MFLUCC 17-1424; (DP9, MFLU
20-0378, holotype); ex-type culture MFLUCC 17-1425;
Lampang Province, Chaehom, on dead stems of C. odorata,
13
Fungal Diversity
Fig. 31 Tremateia chromolaenae (holotype) a, b Appearance of immersed ascomata on
substrate. c Section through
ascoma. d Ostiole. e Peridium.
f Pseudoparaphyses. g–j Asci.
k–p Ascospores q Ascospore
with gelatinous sheath in Indian
ink. Scale bars: a, b = 500 µm,
c, g–j = 50 µm, d = 20 µm, e,
k–q = 10 µm, f = 5 µm
24 September 2016, A. Mapook (JH4, MFLU 20-0379); living culture MFLUCC 17-1458.
GenBank numbers: LSU: MT214451, MT214452,
MT214453, MT214454, ITS: MT214357, MT214358,
MT214359, MT214360, SSU: MT214403, MT214404,
MT214405, MT214406, TEF1: MT235777, MT235778,
MT235779, MT235780, RPB2: MT235815, MT235816,
MT235817, MT235818
Notes: In a BLASTn search of NCBI GenBank, the closest
match with the ITS and SSU sequences of Tremateia chromolaenae (MFLUCC 17-1442, ex-holotype) with 99.34%
(KY038612) and 99.90% (KX274253) similarity, respectively, was Tremateia guiyangensis strain GZAAS01. The
closest match with the LSU sequence with 99.88% similarity
13
was T. arundicola (strain MFLUCC 16-1275, KX274248),
while the closest match with the TEF1 and RPB2 sequences
with 97.58% (LT797094) and 92.31% (LT797014) similarity, respectively, was Kalmusia sp. strain UTHSC: DI16256. In the present phylogenetic analysis, four strains of
T. chromolaenae form a separate clade and cluster with T.
guiyangensis (GZAAS01) and T. arundicola (MFLUCC
16-1275), with high bootstrap support (Fig. 29). However,
T. chromolaenae differs from T. guiyangensis and T. arundicola in having smaller ascomata (150–220 × 170–200(–220)
µm vs. 130–280 × 190–400 µm and 200–300 × 250–350 µm),
smaller asci [(65–)75–105 × (10–)15–20 µm vs.
152–160 × 21–27 µm and 170–200 × 15–20 µm] and smaller
ascospores (16.5–20 × 6.5–10 µm vs. 20–28 × 9–12 µm
Fungal Diversity
Fig. 32 Tremateia thailandensis (holotype) a, b Appearance of immersed ascomata on
substrate. c Section through
ascoma. d Ostiole. e Peridium.
f Pseudoparaphyses. g–j Asci.
k–o Ascospores p Ascospore
with gelatinous sheath in Indian
ink. Scale bars: a = 500 µm,
b = 200 µm, c, g–j = 50 µm,
d = 20 µm, e, f, k–p = 10 µm
Fig. 33 Culture characteristic
on MEA: a Tremateia chiangraiensis (MFLUCC 17-1428).
b Tremateia chromolaenae
(MFLUCC 17-1424). c Tremateia thailandensis (MFLUCC
17-1430)
13
Fungal Diversity
Table 8 Synopsis of Tremateia species with similar morphological features discussed in this study
Species
Ascomata (µm)
Peridium (µm) Asci (µm)
Ascospores (µm) Transverse
septa
References
T. arundicola
(MFLUCC 16-1275)
T. chiangraiensis
(MFLUCC 17-1428)
T. chromolaenae
(MFLUCC 17-1442)
T. guiyangensis
(GZAAS01)
T. thailandensis
(MFLUCC 17-1430)
200–300 high × 250–
350 diam.
220–250 high × 215–
260(–280) diam.
150–220 high × 170–
200(–220) diam.
130–280 high × 190–
400 diam.
200–230 high × 150–
175 diam.
10–20
170–200 × 15–20
20–30 × 6–9
3–6
Hyde et al. (2016b)
10–20(–25)
90–120(–140) × 14–20
23–27.5 × 5–7
5–7
This study
10–15
16.5–20 × 6.5–10 3–4
This study
Up to 9–16
(65–)75–105 × (10–
)15–20
152–160 × 21–27
20–28 × 9–12
3–5
Hyde et al. (2016b)
10–25
(80–)90–125 × 15–25
20–26 × 5.5–9
4–5
This study
and 20–30 × 6–9 µm) (Table 8). A comparison of the ITS
(+5.8S) gene region of T. chromolaenae and T. guiyangensis
reveals 126 base pair differences (22.3%) across 565 nucleotides, and a comparison of the ITS (+5.8S) gene region of
T. chromolaenae and T. arundicola reveals 136 base pair
differences (22.9%) across 593 nucleotides. Therefore, T.
chromolaenae is described as a new species based on phylogeny and morphological comparison.
Tremateia thailandensis Mapook & K.D. Hyde, sp. nov.
Index Fungorum number: IF557366, Facesoffungi number: FoF 07795; Fig. 32
Etymology: Named after Thailand, where the fungus was
first discovered.
Holotype: MFLU 20-0380
Saprobic on dead stems of Chromolaena odorata. Sexual morph: Ascomata 200–230 µm high × 150–175 µm
diam. ( x̄ = 220 × 166 µm, n = 5), immersed, solitary or scattered, coriaceous, subglobose to obpyriform, dark brown
to brown. Ostiolar neck protruding. Peridium 10–25 µm
wide, comprising 2–4 layers, brown to dark brown cells
of textura angularis. Hamathecium comprising 2–3.5 µm
wide, broadly cylindrical, septate, branching pseudoparaphyses. Asci (80–)90–125 × 15–25 µm ( x̄ = 103.5 × 18 µm,
n = 30), 8-spored, bitunicate, fissitunicate, clavate to cylindric-clavate, straight or slightly curved, apically rounded,
pedicellate. Ascospores 20–26 × 5.5–9 µm ( x̄ = 23 × 8 µm,
n = 35), overlapping, 2 seriate, initially hyaline, 1-septate
when immature, becoming golden-brown to brown at maturity, ellipsoid to broadly fusiform, muriform, 4–5-transversely septate, with 1 vertical septum, upper part wider and
shorter, slightly constricted at the central septum, straight
or slightly curved, guttulate, surrounded by hyaline gelatinous sheath observed clearly when mounted in Indian ink.
Asexual morph: Undetermined.
Culture characteristics: Ascospores germinating on
MEA within 24 h. at room temperature and germ tubes
13
produced from several cells. Colonies on MEA circular,
mycelium slightly raised, filamentous, cultures white on
surface, white to creamy white in reverse (Fig. 33c).
Pre-screening for antimicrobial activity: Tremateia
thailandensis (MFLUCC 17-1430) showed no inhibition of
E. coli, B. subtilis and M. plumbeus.
Material examined: THAILAND, Chiang Rai Province,
Huai Kang Pla waterfall, on dead stems of Chromolaena
odorata, 23 June 2015, A. Mapook (HKP6, MFLU 20-0380,
holotype); ex-type culture MFLUCC 17-1430.
GenBank numbers: LSU: MT214455, ITS: MT214361,
SSU: MT214407, TEF1: MT235781, RPB2: MT235819
Notes: In a BLASTn search of NCBI GenBank, the
ITS sequence of Tremateia thailandensis (MFLUCC
17-1430, ex-holotype) with 95.48% similarity was T. guiyangensis (strain GZAAS01, KX274240). The LSU and
SSU sequences with 97.29% (KX274248) and 96.66%
(KX274254) similarity, respectively, was T. arundicola
strain MFLUCC 16-1275, while the closest match with the
TEF1 and RPB2 sequences with 97.77% (LT797094) and
90.00% (LT797014) similarity, respectively, was Kalmusia
sp. strain UTHSC: DI16-256. In the present phylogenetic
analysis, T. thailandensis clusters with T. chiangraiensis with
high bootstrap support (100% ML and 1.00 BYPP, Fig. 29).
However, T. thailandensis differs from T. chiangraiensis
in having smaller ascomata (200–230 × 150–175 µm vs.
220–250 × 215–260(–280) µm) and slightly larger asci
[(80–)90–125 × 15–25 µm vs. 90–120(–140) × 14–20 µm]
and ascospores (20–26 × 5.5–9 µm vs. 23–27.5 × 5–7 µm)
(Table 8). A comparison of the ITS (+5.8S) gene region of
T. thailandensis and T. chiangraiensis reveals 17 base pair
differences (3%) across 565 nucleotides. Therefore, T. thailandensis is described as a new species based on phylogeny
and morphological comparison.
Lophiostomataceae Sacc.
Lophiostomataceae was introduced by Saccardo (1883)
with Lophiostoma as the type genus. The family contains
Fungal Diversity
saprobes found on woody plants from terrestrial, freshwater,
and marine habitats, as well as on herbaceous twigs and pods
(Thambugala et al. 2015; Devadatha et al. 2017; Hashimoto
et al. 2018; Tennakoon et al. 2018; Jayasiri et al. 2019).
Thambugala et al. (2015) accepted 16 genera in the family, which included eleven new genera (Alpestrisphaeria,
Biappendiculispora, Capulatispora, Coelodictyosporium,
Dimorphiopsis, Guttulispora, Lophiohelichrysum, Lophiopoacea, Lophiostoma, Neotrematosphaeria, Paucispora,
Platystomum, Pseudolophiostoma, Pseudoplatystomum,
Sigarispora, Vaginatispora). Wanasinghe et al. (2018) introduced a new genus Neopaucispora based on morphology
and phylogeny, while Hashimoto et al. (2018) introduced
seven new genera (Crassiclypeus, Flabellascoma, Lentistoma, Leptoparies, Neovaginatispora, Parapaucispora,
Pseudopaucispora) based on morphological observations
and phylogenetic analyses. Divergence time estimates for the
family was stem age at 130 Mya (87–178) in the Cretaceous
period (Liu et al. 2017).
Fig. 34 Phylogram generated
from maximum likelihood analysis based on combined dataset
of LSU, ITS, SSU, RPB2 and
TEF1 sequence data. Seventytwo strains are included in the
combined sequence analysis,
which comprise 5256 characters with gaps. Tree topology
of the ML analysis was similar
to the BYPP. The best scoring
RAxML tree with a final likelihood value of − 30152.374563
is presented. The matrix had
1782 distinct alignment patterns, with 29.14% of undetermined characters or gaps.
Estimated base frequencies
were as follows: A = 0.251190,
C = 0.244578, G = 0.267209,
T = 0.237023; substitution rates:
AC = 1.507173, AG = 4.140389,
AT = 1.272744, CG = 1.390194,
CT = 8.158020, GT = 1.000000;
gamma distribution shape
parameter α = 0.167204. Bootstrap support values for ML
equal to or greater than 60% and
BYPP equal to or greater than
0.90 are given above the nodes.
Newly generated sequences
are in dark red bold and type
species are in bold. Teichospora
rubriostiolata (TR 7) and T.
trabicola (C 134) are used as
outgroup taxa
13
Fungal Diversity
Fig. 35 Flabellascoma
minimum (new host record)
a Appearance of ascomata on
substrate. b Section through
ascoma. c Ostiole. d Peridium.
e Pseudoparaphyses. f–g Asci.
h–k Ascospores. Scale bars:
a = 200 µm, b = 50 µm, c, d,
f, g = 20 µm, h–k = 10 µm,
e = 5 µm
Flabellascoma A. Hashim., K. Hiray. & Kaz. Tanaka (2018)
Flabellascoma was introduced by Hashimoto et al. (2018)
to accommodate two new species, F. cycadicola and F. minimum the type species. Jayasiri et al. (2019) introduced a new
host record for F. minimum from fallen pods of Leucaena
leucocephala in Thailand. In this study, we record F. minimum on Siam weed (Fig. 35). A phylogenetic tree based on
combined LSU, ITS, SSU, RPB2 and TEF1 sequence data
is presented in Fig. 34.
13
Flabellascoma minimum A. Hashim., K. Hiray. & Kaz.
Tanaka, in Hashimoto et al., Studies in Mycology 90: 169
(2018)
Facesoffungi number: FoF 05263; Fig. 35
Saprobic on dead stems of Chromolaena odorata. Sexual
morph: Ascomata 210–220(–235) µm high × 165–190(–260)
µm diam. ( x̄ = 221 × 193 µm, n = 5), immersed, appearing
as black spots, coriaceous, solitary or scattered, globose
to obpyriform, brown. ostiole long neck, carbonaceous,
Fungal Diversity
papillate, crest-like, elongated and laterally compressed,
with hyaline periphyses. Peridium 15–20(–25) µm wide,
comprising several layers, outer layers comprising brown
to dark brown cells of textura prismatica, inner layers comprising light brown to hyaline cells of textura angularis.
Hamathecium comprising 1–2(–2.5) µm wide, cylindrical to filiform, septate, branching pseudoparaphyses. Asci
(50–)60–75(–85) × 7–10 µm ( x̄ = 66.5 × 8.5 µm, n = 15),
8-spored, bitunicate, fissitunicate, cylindric-clavate, straight
or slightly curved, with a short pedicel, apically rounded,
with an ocular chamber. Ascospores 4–16 × 4.5–6 µm
( x̄ = 15 × 5 µm, n = 20), overlapping, 1–2-seriate, hyaline,
broadly fusiform with obtuse ends, uniseptate, straight
or slightly curved, guttulate, constricted at the septum;
sheath drawn out to form polar appendages, (4–)6–9 µm
long × 1.5–2.3 µm wide ( x̄ = 7 × 2 µm, n = 30), from apex
of ascospores, with a lateral pad-like structure. Asexual
morph: Undetermined.
Culture characteristics: Ascospores germinating on
MEA within 48 h. at room temperature, at first becoming
4 celled and then germ tubes produced from both ends of
the ascospores. Colonies on MEA filamentous, mycelium
slightly raised, flattened, filiform, white aerial hyphae,
spreading from the center, becoming grayish to light brown
at the surface and dark olivaceous-brown to black in reverse
from the center with creamy white at rim (Fig. 36).
Pre-screening for antimicrobial activity: Flabellascoma minimum (MFLUCC 17-1474) showed antimicrobial
activity against B. subtilis with a 15 mm inhibition zone and
against M. plumbeus with a 17 mm inhibition zone, observable as partial inhibition, when compared to the positive
control (26 mm and 17 mm, respectively), but no inhibition
of E. coli.
Known hosts and distribution: On petioles of Arenga
engleri (Arecaceae), on pods of Bauhinia purpurea
(Fabaceae) in Taiwan (Hashimoto et al. 2018); on pods of
Fig. 36 Culture characteristic on MEA: Flabellascoma minimum
(MFLUCC 17-1474)
Leucaena leucocephala (Fabaceae) in Thailand (Jayasiri
et al. 2019).
Material examined: THAILAND, Nan Province, Doi
Phu Kha, on dead stems of Chromolaena odorata, 23 September 2016, A. Mapook (DPK6, MFLU 20-0312); living
culture MFLUCC 17-1474 (new host record).
GenBank numbers: LSU: MT214461, ITS: MT214367,
SSU: MT214414, TEF1: MT235782, RPB2: MT235820
Notes: A phylogenetic analyses show the strain MFLUCC
17-1474 grouped with Flabellascoma minimum (Fig. 34).
In a BLASTn search of NCBI GenBank, the closest match
of the ITS and RPB2 sequences of MFLUCC 17-1474 with
100% (LC312503) and 99.61% (LC312590) similarity was
F. minimum strain KT 2013, while the closest match with the
LSU, SSU and TEF1 sequences with 99.65% (MK347975),
99.72% (MK347865) and 98.78% (MK360054) similarity
was F. minimum strain MFLUCC 18-0233. Therefore, we
identify our isolate as F. minimum based on the phylogenetic
analysis. Morphological characters also indicated that our
strain is this species. We isolated F. minimum from Chromolaena odorata collected in Thailand, and the isolate is
introduced here as a new host record.
Pseudocapulatispora Mapook & K.D. Hyde, gen. nov.
Index Fungorum number: IF557285, Facesoffungi number: FoF 07796
Etymology: Referring to its similarity with Capulatispora.
Saprobic on dead stems. Sexual morph: Ascomata
immersed, coriaceous, solitary or scattered, ovoid, light
brown to brown. Ostiole long neck, carbonaceous, papillate,
crest-like, elongated and laterally compressed, with hyaline
periphyses. Peridium comprising several layers, outer layers comprising light brown to brown cells of textura prismatica, inner layers comprising pale brown to hyaline cells
of textura angularis. Hamathecium comprising cylindrical to filiform, septate, branching pseudoparaphyses. Asci
8-spored, bitunicate, fissitunicate, cylindric-clavate, with
a short pedicel, apically rounded, with an ocular chamber.
Ascospores overlapping, 1–2-seriate, hyaline, broadly fusiform, uniseptate, with a narrow sheath; sheath drawn out to
form polar appendages from apex of ascospores. Asexual
morph: Undetermined.
Type species: Pseudocapulatispora longiappendiculata
Mapook & K.D. Hyde
Notes: A phylogenetic analyses based on combined dataset of LSU, ITS, SSU, RPB2 and TEF1 sequence data show
that two strains of Pseudocapulatispora longiappendiculata
form a monophyletic clade within the family Lophiostomataceae with high bootstrap support (83% ML and 0.99 BYPP,
Fig. 34). Pseudocapulatispora species are also morphologically distinct in having a long sheath drawn out to form polar
appendages from apex of ascospores with tips of the sheath
13
Fungal Diversity
Fig. 37 Pseudocapulatispora
longiappendiculata (holotype)
a, b Appearance of ascomata
on substrate. c Section through
ascoma. d Ostiole. e Peridium.
f Pseudoparaphyses. g–j Asci.
k–p Ascospores. Scale bars:
a = 500 µm, b = 200 µm, c,
g–j, l–p = 50 µm, d, k = 20 µm,
e = 10 µm, f = 5 µm
capped. Therefore, we introduce Pseudocapulatispora as a
new genus with a new species P. longiappendiculata, based
on morphology and phylogeny.
Pseudocapulatispora longiappendiculata Mapook & K.D.
Hyde, sp. nov.
13
Index Fungorum number: IF557286, Facesoffungi number: FoF 07797; Fig. 37
Etymology: Referring to ascospores with long polar
appendages.
Holotype: MFLU 20-0347
Fungal Diversity
Saprobic on dead stems of Chromolaena odorata. Sexual
morph: Ascomata (195–)250–265 µm high × 105–150 µm
diam. ( x̄ = 240 × 135 µm, n = 5), immersed, appearing as
black spots, coriaceous, solitary or scattered, ovoid, light
brown to brown. Ostiole long neck, carbonaceous, papillate,
crest-like, elongated and laterally compressed, with hyaline
periphyses. Peridium 15–25 µm wide, comprising several
layers, outer layers comprising light brown to brown cells
of textura prismatica, inner layers comprising pale brown
to hyaline cells of textura angularis. Hamathecium comprising (1–)2–3 µm wide, cylindrical to filiform, septate,
branching pseudoparaphyses. Asci 75–120 × 14–20 µm
( x̄ = 100 × 15.5 µm, n = 18), 8-spored, bitunicate, fissitunicate, cylindric-clavate, straight or slightly curved, with
a short pedicel, apically rounded, with an ocular chamber.
Ascospores 24–29 × 6–9 µm ( x̄ = 26 × 7.5 µm, n = 20), overlapping, 1–2-seriate, hyaline, broadly fusiform, uniseptate,
straight or slightly curved, guttulate, constricted at the
septum, with a narrow sheath; sheath drawn out to form
polar appendages, (5.5–)10–40 µm long × (2.5–)3–5(–9) µm
wide ( x̄ = 25 × 4.5 µm, n = 30), from apex of the ascospores
terminating in a glose drop at their tips. Asexual morph:
Undetermined.
Culture characteristics: Ascospores germinating on
MEA within 24 h. at room temperature, at first becoming 4
celled and then germ tubes produced around the ascospores.
Colonies on MEA, initial irregular, becoming filamentous
in old culture, mycelium slightly raised, flattened, filiform,
white aerial hyphae, spreading from the center, becoming
olivaceous-brown at the surface and olivaceous-brown to
black in reverse, MEA change to yellow (Fig. 38).
Pre-screening for antimicrobial activity: Pseudocapulatispora longiappendiculata (MFLUCC 17-1452) showed
antimicrobial activity against B. subtilis with a 7 mm inhibition zone and against M. plumbeus with a 23 mm inhibition
zone, observable as partial inhibition, when compared to the
Fig. 38 Culture characteristic on MEA: Pseudocapulatispora longiappendiculata (MFLUCC 17-1452)
positive control (26 mm and 20 mm, respectively), but no
inhibition of E. coli.
Material examined: THAILAND, Chiang Rai Province,
Doi Pui, on dead stems of Chromolaena odorata, 5 August
2015, A. Mapook (DP45, MFLU 20-0347, holotype); extype culture MFLUCC 17-1452; Lampang Province, Chaehom, on dead stems of Chromolaena odorata, 24 September 2016, A. Mapook (JH3, MFLU 20-0348); living culture
MFLUCC 17-1457.
GenBank numbers: LSU: MT214462, MT214463, ITS:
MT214368, MT214369, SSU: MT214415, MT214416,
TEF1: MT235783, MT235784, RPB2: MT235821
Notes: In a BLASTn search of NCBI GenBank, the closest match of the ITS sequence of Pseudocapulatispora
longiappendiculata (MFLUCC 17-1452, ex-holotype)
with 88.99% similarity was Lophiostoma sp. (strain MA
4558, AJ972793). The closest match with the LSU and
SSU sequences with 98.02% (AY016369) and 99.37%
(AY016354) similarity, respectively, were Lophiostoma heterospora strain CBS 644.86, while the closest match with
the TEF1 sequences with 96.04% similarity was Trematosphaeria sp. (LT797069).
Nigrogranaceae Jaklitsch & Voglmayr
Nigrogranaceae was introduced by Jaklitsch and
Voglmayr (2016) to accommodate the monotypic genus
Nigrograna, which is reported as ecologically diverse
(Kolařík et al. 2017; Kolařík 2018) and contains 11 epithets
(Index Fungorum 2019). The divergence time estimates for
this family are crown age of 72 Mya (44–124) and stem age
of 131 Mya (86–180) during Cretaceous period (Lui et al.
2017).
Nigrograna Gruyter et al.
Nigrograna is an ecologically diverse genus, comprising
plant endophytes, saprobes, marine or estuarine and human
mycetoma (Jaklitsch and Voglmayr 2016; Ahmed et al.
2018; Hyde et al. 2017b; Kolařík et al. 2017; Tibpromma
et al. 2017; Kolařík 2018). The genus was described by de
Gruyter et al. (2013) with the type species, N. mackinnonii. It has been considered as a synonym of Biatriospora
by Ahmed et al. (2014), based on the sequence related to
B. marina. Kolařík et al. (2017) described four new endophytic species of Biatriospora (B. antibiotica, B. carollii, B.
peruviensis, B. yasuniana) from woody plants in temperate
forests and in tropical regions based on morphology and
multigene analyses. These species were synonymized under
Nigrograna by Kolařík et al. (2018). Subsequently, Zhao
et al. (2018) introduced a new species, N. locuta-pollinis
from hive-stored pollen in China. In this study, we introduce a new species, Nigrograna chromolaenae, based on
morphology and molecular data, and provide a description
and illustrations (Fig. 40). A phylogenetic tree based on
13
Fungal Diversity
Fig. 39 Phylogram generated
from maximum likelihood
analysis based on combined
dataset of LSU, ITS, SSU and
TEF1 sequence data. Thirtytwo strains are included in the
combined sequence analysis,
which comprise 3190 characters with gaps. Tree topology
of the ML analysis was similar
to the BYPP. The best scoring
RAxML tree with a final likelihood value of − 7547.223829
is presented. The matrix had
431 distinct alignment patterns, with 29.75% of undetermined characters or gaps.
Estimated base frequencies
were as follows: A = 0.245036,
C = 0.242903, G = 0.269102,
T = 0.242959; substitution rates:
AC = 1.661081, AG = 2.575440,
AT = 1.432192, CG = 0.774269,
CT = 13.455559,
GT = 1.000000; gamma
distribution shape parameter
α = 0.020000. Bootstrap support values for ML equal to or
greater than 60% and BYPP
equal to or greater than 0.90 are
given above the nodes. Newly
generated sequences are in dark
red bold and type species are
in bold. Seriascoma didymospora (MFLUCC 11-0194) and
S. didymospora (MFLUCC
11-0179) are used as outgroup
taxa
combined LSU, ITS, SSU and TEF1 sequence data is presented in Fig. 39.
Nigrograna chromolaenae Mapook & K.D. Hyde, sp. nov.
Index Fungorum number: IF557345, Facesoffungi number: FoF 07297; Fig. 40
Etymology: Name reflects the host genus Chromolaena,
from which this species was isolated.
Holotype: MFLU 20-0341
Saprobic on dead stems of Chromolaena odorata. Sexual
morph: Ascomata 160–280 µm high × 115–130(–150) µm
diam. ( x̄ = 205 × 128 µm, n = 5), immersed to erumpent,
appearing as black spots, coriaceous, solitary or scattered,
13
subglobose to obpyriform, brown to dark brown. Ostiole
long papillate, with hyaline periphyses. Peridium 15–25 µm
wide, comprising several layers, thick-walled cells, outer layers comprising brown to dark brown cells of textura angularis, inner layers comprising light brown to hyaline cells
of textura angularis. Hamathecium comprising 1–2(–2.5)
µm wide, cylindrical to filiform, septate, branching pseudoparaphyses. Asci 40–55(–65) × 7–10 µm ( x̄ = 50 × 8.5 µm,
n = 15), 8-spored, bitunicate, fissitunicate, cylindricclavate, straight or slightly curved, with a short, bulbous
pedicel, apically rounded. Ascospores 10.5–15 × 3.5–5 µm
( x̄ = 12 × 4 µm, n = 30), overlapping, 1–2-seriate, greyish
brown to dark brown, broadly fusiform to inequilateral,
Fungal Diversity
Fig. 40 Nigrograna chromolaenae (holotype) a, b Appearance
of ascomata on substrate. c Section through ascoma. d Ostiole.
e Peridium. f Pseudoparaphyses.
g–j Asci. k–p Ascospores. Scale
bars: a = 500 µm, b = 200 µm,
c = 50 µm, d, g–j = 20 µm,
e = 10 µm, f, k–p = 5 µm
with slightly obtuse ends, upper part or second cell slightly
wider, 1-septate when immature, becoming 3-euseptate
when mature, slightly constricted at the primary median septum, straight or slightly curved, guttulate, without terminal
appendages. Asexual morph: Undetermined.
Culture characteristics: Ascospores germinating on
MEA within 24 h at room temperature and germ tubes
produced from several cells. Colonies on MEA irregular,
mycelium slightly raised, entire, white aerial hyphae at
first, spreading from the center at the surface, becoming
olivaceous-grey to brown and dark olivaceous-grey to black
in reverse (Fig. 41).
Pre-screening for antimicrobial activity: Nigrograna
chromolaenae (MFLUCC 17-1437) showed antimicrobial
activity against M. plumbeus with an 11 mm inhibition zone,
observable as partial inhibition, when compared to the positive control (17 mm), but no inhibition of B. subtilis and E.
coli.
Material examined: THAILAND, Chiang Rai Province,
Doi Pui, on dead stems of Chromolaena odorata, 5 August
13
Fungal Diversity
Fig. 41 Culture characteristic on MEA: Nigrograna chromolaenae
(MFLUCC 17-1437)
2015, A. Mapook (DP21, MFLU 20-0341, holotype); extype culture MFLUCC 17-1437.
GenBank numbers: LSU: MT214473, ITS: MT214379,
TEF1: MT235801
Notes: In a BLASTn search of NCBI GenBank, the closest match with the ITS sequence of Nigrograna chromolaenae (MFLUCC 17-1437, ex-holotype) with 99.32% similarity was Nigrograna mackinnonii (strain L3396, KP263091),
while the closest match with the LSU and TEF1 sequences
with 100% (LN907384) and 100% (LT797087) similarity,
respectively, was N. mackinnonii strain UTHSC: DI16241. In the present phylogenetic analysis, N. chromolaenae
clusters with N. marina strain CY 1228 with low bootstrap
support (Fig. 39). However, N. chromolaenae differs from
N. marina in morphology, N. chromolaenae having broadly
fusiform to inequilateral ascospores, with slightly obtuse
ends, upper part or second cell slightly wider, 1-septate when
immature, becoming 3-euseptate when mature, slightly constricted at the primary median septum, guttulate without terminal appendages, while N. marina has unusual ascospores
that are dark brown, fusiform with hyaline, rounded swollen
tips at both ends and 1–4 septa situated near the ends (Hyde
et al. 2013). A comparison of the ITS (+5.8S) gene region
of N. chromolaenae and N. marina reveals 15 base pair differences (1.77%) across 847 nucleotides. Therefore, N. chromolaenae is described as a new species based on phylogeny
and morphological comparison.
Neomassarinaceae Mapook & K.D. Hyde, fam. nov.
Index Fungorum number: IF557341, Facesoffungi number: FoF 07798
Etymology: Referring to the type genus, Neomassarina
Saprobic on dead leaf or stems of herbaceous plant.
Sexual morph: Ascomata immersed or semi-immersed
to erumpent, globose to subglobose or obpyriform, light
brown to brown, coriaceous, solitary or scattered. Ostiole
long neck, carbonaceous, papillate, crest-like, elongated and
13
laterally compressed, with or without hyaline periphyses.
Peridium comprising 2–5 layers, pale brown to hyaline cells
or dark brown to black cells of textura angularis. Hamathecium composed of cylindrical to filiform, septate, branching
pseudoparaphyses. Asci 8-spored, bitunicate, fissitunicate,
cylindrical to cylindric-clavate, with a short pedicel, apically
rounded, with an ocular chamber. Ascospores overlapping,
1–2-seriate, hyaline to pale brown at maturity, fusiform,
uniseptate, with a narrow sheath or surrounded by hyaline
gelatinous sheath; sheath drawn out to form polar appendages from both ends of the ascospores, straight or slightly
curved. Asexual morph: Undetermined.
Type genus: Neomassarina Phook., Jayasiri & K.D.
Hyde, in Hyde et al., Fungal Divers. 80: 136 (2016)
Notes: Based on a phylogenetic tree of combined LSU,
ITS, SSU, TEF1 and RPB2 sequence data, Neomassarinaceae forms a distinct family in the clade comprising
Sporormiaceae and Amorosiaceae (Fig. 42). Neomassarinaceae differs from Sporormiaceae in morphology. The
new family has similarity with Amorosiaceae in its crestlike ostiole, cylindrical to cylindric-clavate asci with an
ocular chamber and surrounded by a mucilaginous sheath
(Thambugala et al. 2015; Tibpromma et al. 2017). However, they differ in crest-like, elongated and laterally compressed, carbonaceous ostiolar necks, with or without hyaline periphyses.
Neomassarina Phook., Jayasiri & K.D. Hyde, in Hyde et al.,
Fungal Divers. 80: 136 (2016)
Neomassarina was introduced by Hyde et al. (2016b)
with Neomassarina thailandica as the type species collected from Agave angustifolia (Asparagaceae) in Thailand
and placed in Pleosporales genera incetae sedis based on
phylogenetic analyses. Hyde et al. (2018a, b) included the
genus in Sporormiaceae and introduced a new species, N.
pandanicola from Pandanus sp. (Pandanaceae) in Thailand,
based on both morphology and phylogenetic support. In this
study, a new species, Neomassarina chromolaenae is introduced together with a new host record for N. thailandica,
together with descriptions and illustrations (Figs. 43, 44). A
phylogenetic tree based on combined LSU, ITS, SSU, TEF1
and RPB2 sequence data is presented in Fig. 42.
Neomassarina chromolaenae Mapook & K.D. Hyde, sp.
nov.
Index Fungorum number: IF557342, Facesoffungi number: FoF 07799; Fig. 43
Etymology: Name reflects the host genus Chromolaena,
from which this species was isolated.
Holotype: MFLU 20-0333
Saprobic on dead stems of Chromolaena odorata. Sexual
morph: Ascomata 235–295 µm high × 125–155 µm diam.
( x̄ = 270 × 145 µm, n = 5), immersed, coriaceous, solitary
Fungal Diversity
Fig. 42 Phylogram generated
from maximum likelihood analysis based on combined dataset
of LSU, ITS, SSU, TEF1 and
RPB2 sequence data. Ninetyeight strains are included in the
combined sequence analysis,
which comprise 5181 characters with gaps. Tree topology
of the ML analysis was similar
to the BYPP. The best scoring
RAxML tree with a final likelihood value of − 56728.891914
is presented. The matrix had
2700 distinct alignment patterns, with 44.08% of undetermined characters or gaps.
Estimated base frequencies
were as follows: A = 0.247521,
C = 0.249316, G = 0.271783,
T = 0.231379; substitution rates:
AC = 1.589623, AG = 3.631327,
AT = 1.592103, CG = 1.373577,
CT = 7.761405, GT = 1.000000;
gamma distribution shape
parameter α = 0.271790. Bootstrap support values for ML
equal to or greater than 60% and
BYPP equal to or greater than
0.90 are given above the nodes.
Newly generated sequences
are in dark red bold and type
species are in bold. Melanomma
pulvis-pyrius (CBS 124080) is
used as outgroup taxon
or scattered, globose or subglobose to obpyriform, light
brown to brown. Ostiole long neck, carbonaceous, papillate, crest-like, elongated and laterally compressed, with
hyaline periphyses. Peridium 15–20 µm wide, comprising
2–3-layers of pale brown to hyaline cells of textura angularis. Hamathecium comprising 1–2 µm wide, cylindrical
to filiform, septate, branching pseudoparaphyses. Asci
70–100 × 12–13 µm ( x̄ = 88.5 × 12.5 µm, n = 10), 8-spored,
bitunicate, fissitunicate, cylindric-clavate, straight or slightly
curved, with a short pedicel, apically rounded, with an ocular chamber. Ascospores 20–30 × 4–6.5 µm ( x̄ = 24 × 5 µm,
n = 25), overlapping, 1–2-seriate, hyaline, fusiform,
13
Fungal Diversity
Fig. 43 Neomassarina
chromolaenae (holotype) a,
b Appearance of ascomata on
substrate. c Section through
ascoma. d Ostiole. e Peridium.
f Pseudoparaphyses. g–h
Asci. i–n Ascospores with a
gelatinous sheath forming an
appendage apicaly. Scale bars:
a, b = 500 µm, c = 100 µm,
d = 20 µm, g, h = 50 µm, e, f,
i–n = 10 µm
uniseptate, straight or slightly curved, constricted at the
septum, smooth, with a narrow sheath; sheath drawn out
to form polar appendages 4.5–9.5 µm long × 2–4 µm wide
( x̄ = 6.5 × 3 µm, n = 30), from both ends of the ascospores,
straight or slightly curved. Asexual morph: Undetermined.
Culture characteristics: Ascospores germinating on
MEA within 48 h. at room temperature, germ tubes produced from the apex. Colonies on MEA circular, mycelium
slightly crateriform, flattened, filiform, white aerial hyphae
at the surface and creamy brown to yellowish-brown at
the center in reverse, white to yellow-white at the margin
(Fig. 45a).
13
Pre-screening for antimicrobial activity: Neomassarina
chromolaenae (MFLUCC 17-1480) showed antimicrobial
activity against E. coli with an 11 mm inhibition zone when
compared to the positive control (9 mm), but no inhibition
of B. subtilis and M. plumbeus.
Material examined: THAILAND, Lampang Province,
Ngao, on dead stems of Chromolaena odorata, 21 September 2016, A. Mapook (LP1, MFLU 20-0333, holotype); extype culture MFLUCC 17-1480.
GenBank numbers: LSU: MT214466, ITS: MT214372,
SSU: MT214419, TEF1: MT235785, RPB2: MT235822
Notes: In a BLASTn search of NCBI GenBank,
the closest match of the ITS and TEF1 sequences for
Fungal Diversity
Fig. 44 Neomassarina thailandica (new host record) a,
b Appearance of ascomata on
substrate. c Section through
ascoma. d Ostiole. e Peridium. f
Pseudoparaphyses. g–j Asci. k–
n Ascospores. o–p Ascospores
with gelatinous sheath in Indian
ink. Scale bars: a = 200 µm,
b = 100 µm, c = 50 µm, d,
g–j = 20 µm, e, k–p = 10 µm,
f = 5 µm
Neomassarina chromolaenae (MFLUCC 17-1480, exholotype) is N. thailandica with 94.72% (NR_154244)
and 96.30% (KX672163), respectively, similarity to the
strain MFLU 11-0144, while the closest match with the
LSU and SSU sequences were with N. pandanicola strain
MFLUCC 16-0270 with 99.67% (MG298946) and 98.34%
(MG298947) similarity. In the present phylogenetic analysis, N. chromolaenae is closely related to N. thailandica
with high bootstrap support (100% ML and 1.00 BYPP,
Fig. 42). However, N. chromolaenae differs from N. thailandica in having wider asci (70–100 × 12–13 µm vs.
(70–)75–90(–93) × 7–8(–8.5) µm) and larger ascospores
(20–30 × 4–6.5 µm vs. (17.5–)18–20 × 3–4(–5) µm) with
a narrow sheath drawn out to form polar appendages
(4.5–9.5 × 2–4 µm vs. 4.5–8 × 1.5–3.5 µm) (Table 9). A
comparison of the ITS (+5.8S) gene region of N. chromolaenae and N. thailandica reveals 30 base pair differences
(6%) across 495 nucleotides. Therefore, N. chromolaenae is
described as a new species based on phylogeny with morphological comparison.
Neomassarina thailandica Phook., Jayasiri & K.D. Hyde,
in Hyde et al., Fungal Divers. 80: 138 (2016)
Facesoffungi number: FoF 02260; Fig. 44
13
Fungal Diversity
Fig. 45 Culture characteristics on MEA: a Neomassarina chromolaenae (MFLUCC 17-1480). b Neomassarina thailandica (MFLUCC
17-1432)
Saprobic on dead stems of Chromolaena odorata. Sexual morph: Ascomata (90–)135–160 µm high × 95–130 µm
diam. ( x̄ = 125 × 110 µm, n = 5), immersed, coriaceous,
solitary or scattered, obpyriform, flattened base, light
brown to brown. Ostiole long neck, carbonaceous, papillate, crest-like, elongated and laterally compressed, with
hyaline periphyses. Peridium 5–15 µm wide, comprising 2
layers of pale brown to hyaline cells of textura angularis.
Hamathecium comprising (1–)1.5–2.3 µm wide, cylindrical to filiform, septate, branching pseudoparaphyses. Asci
55–70 × 7–9 µm ( x̄ = 60 × 8 µm, n = 15), 8-spored, bitunicate,
fissitunicate, cylindric-clavate, straight or slightly curved,
with a short pedicel, apically rounded, with an ocular chamber. Ascospores 17.5–20 × 2.5–4.5 µm ( x̄ = 18.5 × 3.5 µm,
n = 25), overlapping, 1–2-seriate, hyaline, fusiform, uniseptate, straight or slightly curved, guttulate, constricted
at the septum, surrounded by hyaline gelatinous sheath;
sheath drawn out to form polar appendages 4.5–8 µm
long × 1.5–3.5 µm wide ( x̄ = 6 × 2.5 µm, n = 10), observed
clearly when mounted in Indian ink. Asexual morph:
Undetermined.
Culture characteristics: Ascospores germinating on
MEA within 24 h at room temperature, germ tubes produced from both ends. Colonies on MEA circular, mycelium
slightly crateriform, flattened, filiform, initial white aerial
hyphae, becoming pale brown from the center at the surface
and initial white to yellow-white at the margin, brown to
dark brown at the center in reverse, becoming light brown
to brown in old culture (Fig. 45b).
Pre-screening for antimicrobial activity: Neomassarina thailandica (MFLUCC 17-1432) showed antimicrobial activity against M. plumbeus with a 30 mm inhibition
zone, observable as partial inhibition, when compared to
the positive control (17 mm), but no inhibition of B. subtilis
and E. coli.
Known hosts and distribution: On dead bract-like
leaves from flower stalk of Agave angustifolia (Asparagaceae) in Thailand (Hyde et al. 2016)
Material examined: THAILAND, Chiang Rai Province,
Doi Pui, on dead stems of Chromolaena odorata, 5 August
2015, A. Mapook (DP15, MFLU 20-0334); living culture
MFLUCC 17-1432 (new host record).
GenBank numbers: LSU: MT214467, ITS: MT214373,
SSU: MT214420, TEF1: MT235786, RPB2: MT235823
Notes: In the present phylogenetic analysis, MFLUCC
17-1432 is closely related to N. thailandica with high bootstrap support (100% ML and 1.00 BYPP, Fig. 42). In a
BLASTn search of NCBI GenBank, the closest match of
ITS and TEF1 sequences showed that the strains is identical to N. thailandica with 100% (NR_154244) and 99.31%
(KX672163), respectively, while the closest match with the
LSU and SSU sequences were with N. pandanicola strain
MFLUCC 16-0270 with 100% (MG298946) and 98.06%
(MG298947) similarity. Therefore, we identify our isolates
as N. thailandica based on phylogenetic analyses with morphological comparison (Table 9) and the isolates are introduced here as a new host record from Chromolaena odorata
collected in Thailand.
Neopyrenochaetaceae Valenzuela-Lopez et al.
Neopyrenochaetaceae was introduced by ValenzuelaLopez et al. (2018) to accommodate the monotypic genus
Neopyrenochaeta based on phylogeny.
Neopyrenochaeta Valenzuela-Lopez et al.
Neopyrenochaeta was introduced by Valenzuela-Lopez
et al. (2018) for a new species N. fragariae. The genus
Table 9 Synopsis of Neomassarina species with similar morphological features discussed in this study
Species
Ascomata (µm)
Asci (µm)
N. chromolaenae
(MFLUCC 17-1480)
N. pandanicola (MFLUCC
16-0270)
N. thailandica (MFLUCC
17-1432)
N. thailandica (MFLUCC
10-0552)
235–295 × 125–155
70–100 × 12–13 20–30 × 4–6.5
4.5–9.5 × 2–4
This study
140–160 × 130–180
50–80 × 6.5–10 11–20 × 3–5
3.5–6 long
Hyde et al. (2018a, b)
4.5–8 × 1.5–3.5
This study
13
Ascospores (µm)
Polar appendages (µm) References
(90–)135–160 × 95–130 55–70 × 7–9
17.5–20 × 2.5–4.5
130–180 × 100–200
(17.5–)18–20 × 3–4(–5) Surrounded by a distinct mucilaginous
sheath
(70–)75–90(–
93) × 7–8(–
8.5)
Hyde et al. (2016b)
Fungal Diversity
Fig. 46 Phylogram generated
from maximum likelihood
analysis based on combined
dataset of LSU, ITS and RPB2
sequence data. Fourty-nine
strains are included in the
combined sequence analysis,
which comprise 2556 characters with gaps. Tree topology
of the ML analysis was similar
to the BYPP. The best scoring
RAxML tree with a final likelihood value of − 18819.011208
is presented. The matrix had
977 distinct alignment patterns, with 12.78% of undetermined characters or gaps.
Estimated base frequencies
were as follows: A = 0.247993,
C = 0.237502, G = 0.272601,
T = 0.241905; substitution rates:
AC = 1.771498, AG = 5.599945,
AT = 1.899441, CG = 1.222461,
CT = 10.018389,
GT = 1.000000; gamma
distribution shape parameter
α = 0.168302. Bootstrap support values for ML equal to or
greater than 60% and BYPP
equal to or greater than 0.90 are
given above or below the nodes.
Newly generated sequences
are in dark red bold and type
species are in bold. Pleospora
herbarum (CBS 191.86) and P.
herbarum (IT956) are used as
outgroup taxa
belonging in Cucurbitariaceae (Aveskamp et al. 2010; de
Gruyter et al. 2010, 2013; Doilom et al. 2013) based on
phylogenetic analysis and available sequences from GenBank. The genus is mostly found as the asexual morph, and
has been reported from diverse habitats. N. acicula was
recorded from a waterpipe, N. fragariae from fruit (Fragaria
ananassa), N. inflorescentiae from plants (Protea neriifolia), N. telephoni from a mobile phone (Crous et al. 2015d;
Valenzuela-Lopez et al. 2018), and N. cercidis on pods of
Cercis chinensis in China based on morphology and phylogeny (Jayasiri et al. 2019). Five epithets are listed in Index
Fungorum (2020) with N. acicula as the type species. We
introduce the sexual morphs of four new Neopyrenochaeta
species, which were collected from dead aerial stems of
Chromolaena odorata, based on morphology and molecular
data, together with descriptions and illustrations (Figs. 47,
48, 49, 50). A phylogenetic tree based on combined LSU,
ITS and RPB2 sequence data is presented in Fig. 46.
Neopyrenochaeta chiangraiensis Mapook & K.D. Hyde,
sp. nov.
Index Fungorum number: IF557311, Facesoffungi number FoF 07482; Fig. 47
13
Fungal Diversity
Fig. 47 Neopyrenochaeta
chiangraiensis (holotype) a,
b Appearance of superficial
ascomata on substrate. c Section through ascoma. d Dark
brown setae. e Peridium. f
Pseudoparaphyses. g–j Asci.
k–p Ascospores. Scale bars:
a = 500 µm, b = 200 µm, c,
g–j = 20 µm, d, e, k–p = 10 µm,
f = 5 µm
Etymology: Referring to the location where the specimen
was collected, Chiang Rai Province, Thailand.
Holotype: MFLU 20-0337
Saprobic on dead stems of Chromolaena odorata. Sexual
morph: Ascomata 100–110 µm high × 95–105 µm diam.
( x̄ = 105 × 100 µm, n = 5), superficial, appearing as small
dark spots, coriaceous, solitary or scattered, globose to obpyriform, brown to dark brown. Ostiole short papillate, with
numerous external brown setae. Peridium (3.5–)5–12 µm
wide, comprising 1–2 layers of thin-walled, pale brown to
13
brown cells of textura angularis. Hamathecium comprising
1–2 µm wide, cylindrical, septate, branching pseudoparaphyses. Asci (45–)50–65 × 8–12 µm ( x̄ = 55 × 9.5 µm, n = 20),
8-spored, bitunicate, fissitunicate, cylindric-clavate, straight
or slightly curved, with a short bulbous pedicel, apically
rounded. Ascospores 14–18 × 3.5–5 µm ( x̄ = 16 × 4.5 µm,
n = 15), overlapping, bi-seriate, light olivaceous-brown to
pale yellowish brown, cylindric-fusiform to inequilateral,
with slightly obtuse ends, widest at the second cell from
the apex and tapering towards ends, 3-septate, straight or
Fungal Diversity
Fig. 48 Neopyrenochaeta
chromolaenae (holotype) a,
b Appearance of superficial
ascomata on substrate. c Section through ascoma. d Dark
brown setae. e Peridium. f
Pseudoparaphyses. g–j Asci.
k–p Ascospores. Scale bars:
a = 500 µm, b = 100 µm, c,
g–j = 20 µm, d, e, k–p = 10 µm,
f = 5 µm
slightly curved, small guttule, slightly constricted at the
septa, without terminal appendages. Asexual morph:
Undetermined.
Culture characteristics: Ascospores germinating on
MEA within 24 h. at room temperature and germ tubes produced from both ends. Colonies on MEA circular, mycelium
flat, filamentous, white to light grey at first, become greyishbrown at the surface and greyish-brown to dark brown in
reverse with age (Fig. 51a).
Pre-screening for antimicrobial activity: Not tested.
Material examined: THAILAND, Chiang Rai Province,
Doi Pui, on dead stems of Chromolaena odorata, 5 August
2015, A. Mapook (DP38, MFLU 20-0337, holotype); extype culture MFLUCC 17-1445.
GenBank numbers: LSU: MT214468, ITS: MT214374
Notes: In a BLASTn search of NCBI GenBank, the
closest match with the ITS sequence of Neopyrenochaeta
chiangraiensis (MFLUCC 17-1445, ex-holotype) with
96.07% similarity was Setophaeosphaeria badalingensis
(strain S139, MK304112), while the closest match with
the LSU sequence with 96.04% similarity was Neopyrenochaeta cercidis (strain C136, MK347932). In the present phylogenetic analysis, N. chiangraiensis clusters
with N. chromolaenae (MFLUCC 17-1446) with high
bootstrap support (99% ML and 1.00 BYPP, Fig. 46).
13
Fungal Diversity
Fig. 49 Neopyrenochaeta thailandica (holotype) a, b Appearance of superficial ascomata on
substrate. c Ascoma d Section
through ascoma. e Brown setae.
f Peridium. g Pseudoparaphyses. h–k Asci. l–q Ascospores.
Scale bars: a, b = 200 µm, c, d,
h–k = 50 µm, e–f = 20 µm, g,
l–q = 10 µm
However, N. chiangraiensis differs from N. chromolaenae in having smaller asci [(45–)50–65 × 8–12 µm vs.
(55.5–)60–80 × (14.5–)15–20 µm] and smaller ascospores
(14–18 × 3.5–5 µm vs. 23–26 × 6–8 µm) that are light olivaceous-brown to pale yellowish brown with 3 septa, while
N. chromolaenae has pale olivaceous-brown to smoke-grey,
5-septate ascospores (Table 10). A comparison of the ITS
(+5.8S) gene region of N. chiangraiensis and N. chromolaenae reveals 23 base pair differences (4.4%) across 517 nucleotides. Therefore, N. chiangraiensis is described as a new
species based on phylogeny and morphological comparison.
Neopyrenochaeta chromolaenae Mapook & K.D. Hyde,
sp. nov.
13
Index Fungorum number: IF557312, Facesoffungi number: FoF 07800; Fig. 48
Etymology: Name reflects the host genus Chromolaena,
from which this species was isolated.
Holotype: MFLU 20-0338
Saprobic on dead stems of Chromolaena odorata. Sexual morph: Ascomata 95–110 µm high × 70–75(–145) µm
diam. ( x̄ = 100 × 108 µm, n = 5), superficial, appearing as
small dark spots, coriaceous, solitary or scattered, globose
to obpyriform, brown to dark brown. Ostiole papillate,
with numerous external brown setae. Peridium 5–15 µm
wide, comprising 1–2 layers of thin-walled, pale brown
to brown cells of textura angularis. Hamathecium comprising 1–2.5 µm wide, cylindrical, septate, branching,
Fungal Diversity
Fig. 50 Neopyrenochaeta
triseptatispora (holotype) a,
b Appearance of superficial
ascomata on substrate. c Section
through ascoma. d Ostiole with
brown setae. e Brown seta. f
Peridium. g Pseudoparaphyses.
h–k Immature and mature asci.
l–q Ascospores. Scale bars:
a = 200 µm, b = 100 µm, c,
h–k = 20 µm, d, f = 10 µm, e, g,
l–q = 5 µm
pseudoparaphyses. Asci (55.5–)60–80 × (14.5–)15–20 µm
( x̄ = 70 × 16.5 µm, n = 20), 8-spored, bitunicate, fissitunicate, cylindric-clavate to broadly clavate, straight or slightly
curved, with a short bulbous pedicel, apically rounded.
Ascospores 23–26 × 6–8 µm ( x̄ = 24.5 × 7 µm, n = 25), overlapping 3–5-seriate, pale olivaceous-brown to smoke-grey,
cylindric-fusiform, with slightly obtuse ends, widest at
the center and tapering towards the rounded ends, 5-septate, straight or slightly curved, guttulate, constricted at
the septa, without terminal appendages. Asexual morph:
Undetermined.
Culture characteristics: Ascospores germinating on
MEA within 24 h. at room temperature and germ tubes
produced from both ends. Colonies on MEA circular, mycelium flat, filamentous, white at first with brown from the
center of the colony, become greyish to dark olivaceous at
the surface and dark olivaceous in reverse (Fig. 51b).
Pre-screening for antimicrobial activity: Neopyrenochaeta chromolaenae (MFLUCC 17-1446) showed antimicrobial activity against B. subtilis and E. coli (13 mm and
10 mm inhibition zone, respectively), when compared to the
positive control (26 mm and 17 mm, respectively), but no
inhibition of M. plumbeus.
Material examined: THAILAND, Chiang Rai Province,
Doi Pui, on dead stems of Chromolaena odorata, 5 August
13
Fungal Diversity
Fig. 51 Culture characteristics on MEA: a Neopyrenochaeta chiangraiensis (MFLUCC 17-1445). b Neopyrenochaeta chromolaenae
(MFLUCC 17-1446). c Neopyrenochaeta thailandica (MFLUCC 17-1461). d Neopyrenochaeta triseptatispora (MFLUCC 17-1436)
2015, A. Mapook (DP39, MFLU 20-0338, holotype); extype culture MFLUCC 17-1446.
GenBank numbers: LSU: MT214469, ITS: MT214375,
SSU: MT214421, TEF1: MT235787, RPB2: MT235824
Notes: In a BLASTn search of NCBI GenBank, the closest match with the ITS sequence of Neopyrenochaeta chromolaenae (MFLUCC 17-1446, ex-holotype) with 95.06%
similarity was Setophaeosphaeria badalingensis (strain 41,
MK311292). The closest match with the LSU sequence
with 99.42% similarity was Neopyrenochaeta cercidis
(strain C136, MK347932), while the closest match with
the RPB2 sequence with 94.25% similarity was Setophaeosphaeria citricola (strain CBS 143179, MH108008). In
the present phylogenetic analysis, N. chromolaenae clusters with N. chiangraiensis (MFLUCC 17-1445) with
high bootstrap support (99% ML and 1.00 BYPP, Fig. 46).
However, N. chromolaenae differs from N. chiangraiensis
in having larger asci [(55.5–)60–80 × (14.5–)15–20 µm vs.
(45–)50–65 × 8–12 µm] and ascospores (23–26 × 6–8 µm vs.
14–18 × 3.5–5 µm) that are pale olivaceous-brown to smokegrey with 5 septa, while N. chiangraiensis has light olivaceous-brown to pale yellowish brown, 3-septate ascospores
(Table 10). A comparison of the ITS (+5.8S) gene region
of N. chromolaenae and N. chiangraiensis reveals 23 base
pair differences (4.4%) across 517 nucleotides. Therefore,
N. chromolaenae is described as a new species based on
phylogeny and morphological comparison.
Neopyrenochaeta thailandica Mapook & K.D. Hyde, sp.
nov.
Index Fungorum number: IF557313, Facesoffungi number: FoF 07801; Fig. 49
Etymology: The name reflects the country, where the
specimen was collected, Thailand.
Holotype: MFLU 20-0339
Saprobic on dead stems of Chromolaena odorata. Sexual morph: Ascomata 115–160 µm high × 110–150 µm
diam. ( x̄ = 133 × 125 µm, n = 5), superficial, appearing as
small dark spots, coriaceous, solitary or scattered, subglobose to obpyriform, reddish brown to brown. Ostiole
short papillate, with numerous external reddish-brown
setae. Peridium 5–12 µm wide, comprising 2–3-layers of
thin-walled, pale brown to brown cells of textura angularis. Hamathecium comprising (1.5–)2–2.5 µm wide, cylindrical to broadly filiform, septate, branching pseudoparaphyses. Asci (55–)60–80 × 13–20 µm ( x̄ = 67.5 × 16.5 µm,
Table 10 Morphological features of Neopyrenochaeta species discussed in this study
Species
Asocomata (µm)
Peridium (µm) Asci (µm)
Ascospores (µm)
References
N. chiangraiensis
(MFLUCC 17-1445)
100–110 high × 95–105
diam.
(3.5–)5–12
(45–)50–65 × 8–12
This study
N. chromolaenae
(MFLUCC 17-1446)
95–110 high × 70–75
(–145) diam.
5–15
(55.5–)60–80 ×
(14.5–)15–20
N. thailandica (MFLUCC
17-1461)
115–160 high × 110–150
diam.
5–12
(55–)60–80 × 13–20
N. triseptatispora
(MFLUCC 17-1436)
75–110 high × 70–90 diam. 5–10(–15)
14–18 × 3.5–5; light
olivaceous-brown to
pale yellowish brown,
3-septate
23–26 × 6–8; pale
olivaceous-brown to
smoke-grey, 5-septate
(21.5–)23–32 × 5–9; pale
olivaceous-grey to
slightly yellowish brown,
3–5-septate
15–20 × 4–7; hyaline,
3-septate
13
45–65 × 9.5–13
This study
This study
This study
Fungal Diversity
n = 15), 8-spored, bitunicate, fissitunicate, cylindricclavate to broadly clavate, straight or slightly curved, apically rounded, with a short, bulbous pedicel. Ascospores
(21.5–)23–32 × 5–9 µm ( x̄ = 26 × 6.5 µm, n = 40), overlapping 1–3-seriate, pale olivaceous-grey to slightly yellowish brown, cylindrical to broadly fusiform, with slightly
obtuse ends, widest at the center and tapering towards the
rounded ends, 3–5-septate, straight or slightly curved, third
cell from top slightly swollen, guttulate, slightly constricted
at the septa, without terminal appendages. Asexual morph:
Undetermined.
Culture characteristics: Ascospores germinating on
MEA within 48 h. at room temperature and germ tubes produced from the apex. Colonies on MEA circular, mycelium
slightly raised, entire, greyish to dark olivaceous at the surface and dark olivaceous in reverse (Fig. 51c).
Pre-screening for antimicrobial activity: Neopyrenochaeta thailandica (MFLUCC 17-1461) showed no inhibition of E. coli, B. subtilis and M. plumbeus.
Material examined: THAILAND, Lampang Province,
Chaehom, on dead stems of Chromolaena odorata, 24 September 2016, A. Mapook (JH7, MFLU 20-0339, holotype);
ex-type culture MFLUCC 17-1461.
GenBank numbers: LSU: MT214470, ITS: MT214376,
SSU: MT214422, TEF1: MT235788, RPB2: MT235825
Notes: In a BLASTn search of NCBI GenBank, the closest match with the ITS sequence of Neopyrenochaeta thailandica (MFLUCC 17-1461, ex-holotype) with 98.96% similarity was Setophaeosphaeria hemerocallidis (strain A582,
KX463035). The closest match with the LSU sequence with
99.12% similarity was N. cercidis (strain C136, MK347932),
while the closest match with the RPB2 sequence with
88.88% similarity was Setophaeosphaeria citricola (strain
CBS 143179, MH108008). In the present phylogenetic analysis, N. thailandica forms a sister taxon with N. chromolaenae (MFLUCC 17-1446), N. chiangraiensis (MFLUCC
17-1445) and N. triseptatispora (MFLUCC 17-1436) with
bootstrap support (1.00 BYPP, Fig. 46). However, N. thailandica differs from N. chiangraiensis and N. triseptatispora in having larger asci [(55–)60–80 × 13–20 µm vs.
(45–)50–65 × 8–12 µm and 45–65 × 9.5–13 µm] and larger
ascospores [(21.5–)23–32 × 5–9 µm vs. 14–18 × 3.5–5 µm
and 15–20 × 4–7 µm] that are 3–5-septate, while N. chiangraiensis and N. triseptatispora have 3-septate ascospores
(Table 10). A comparison of the ITS (+5.8S) gene region
of N. thailandica and N. triseptatispora reveals 66 base pair
differences (12.15%) across 543 nucleotides. Therefore, N.
thailandica is described as a new species based on phylogeny and morphological comparison.
Neopyrenochaeta triseptatispora Mapook & K.D. Hyde,
sp. nov.
Index Fungorum number: IF557314, Facesoffungi number: FoF 07802; Fig. 50
Etymology: The epithet “triseptatispora” refers to the
3-septate ascospores.
Holotype: MFLU 20-0340
Saprobic on dead stems of Chromolaena odorata. Sexual morph: Ascomata 75–110 µm high × 70–90 µm diam.
( x̄ = 95 × 80 µm, n = 5), superficial, appearing as, small dark
spots, coriaceous, solitary or scattered, globose to obpyriform, reddish brown to brown. Ostiole short papillate, with
numerous external light brown to brown setae. Peridium
5–10(–15) µm wide, comprising 2–3 layers of thin-walled,
hyaline or pale brown to brown cells of textura angularis.
Hamathecium comprising 1–3 µm wide, cylindrical, septate, branching pseudoparaphyses. Asci 45–65 × 9.5–13 µm
( x̄ = 56 × 11 µm, n = 20), 8-spored, bitunicate, fissitunicate,
cylindric-clavate, straight or slightly curved, with a short bulbous pedicel, apically rounded. Ascospores 15–20 × 4–7 µm
( x̄ = 18 × 6 µm, n = 40), overlapping, 2–3-seriate, hyaline,
cylindric-fusiform to inequilateral, with obtuse ends, slightly
widest at the second cell from the apex and tapering towards
the rounded ends, 3-septate, straight or slightly curved, small
guttule, constricted at the septa, without terminal appendages. Asexual morph: Undetermined.
Culture characteristics: Ascospores germinating on
MEA within 24 h. at room temperature and germ tubes produced from several cells. Colonies on MEA circular, mycelium flat, filamentous, white at first, become greyish to dark
olivaceous at the surface and dark olivaceous in reverse with
age (Fig. 51d).
Pre-screening for antimicrobial activity: Neopyrenochaeta triseptatispora (MFLUCC 17-1436) showed antimicrobial activity against B. subtilis with a 16 mm inhibition
zone and against M. plumbeus with a 24 mm inhibition zone,
observable as partial inhibition, when compared to the positive control (25 mm and 17 mm, respectively), but no inhibition of E. coli.
Material examined: THAILAND, Chiang Rai Province,
Doi Pui, on dead stems of Chromolaena odorata, 5 August
2015, A. Mapook (DP20, MFLU 20-0340, holotype); extype culture MFLUCC 17-1436.
GenBank numbers: LSU: MT214471, ITS: MT214377,
SSU: MT214423, TEF1: MT235789, RPB2: MT235826
Notes: In a BLASTn search of NCBI GenBank, the
closest match with the ITS sequence of Neopyrenochaeta
triseptatispora (MFLUCC 17-1436, ex-holotype) with
99.62% similarity was Setophaeosphaeria citricola (strain
CBS 143179, MH107916). The closest match with the
LSU sequence with 98.97% similarity was N. cercidis
(strain C136, MK347932), while the closest match with
the RPB2 sequence with 93.75% similarity was Setophaeosphaeria citricola (strain CBS 143179, MH108008).
In the present phylogenetic analysis, N. triseptatispora
13
Fungal Diversity
is forms a sister taxon with N. chromolaenae (MFLUCC
17-1446) and N. chiangraiensis (MFLUCC 17-1445) with
high bootstrap support (98% ML and 1.00 BYPP, Fig. 46).
However, N. triseptatispora differs from N. chromolaenae in having smaller ascomata (75–110 × 70–90 µm vs.
95–110 × 70–75(–145) µm), smaller asci (45–65 × 9.5–13 µm
vs. (55.5–)60–80 × (14.5–)15–20 µm) and smaller ascospores
(15–20 × 4–7 µm vs. 23–26 × 6–8 µm) that are 3-septate,
while N. chromolaenae has 5-septate ascospores (Table 10).
N. triseptatispora is also similar to N. chiangraiensis in
having 3-septate ascospores but differs in slightly larger
ascomata (75–110 × 70–90 µm vs. 100–110 × 95–105 µm)
and ascospores (15–20 × 4–7 µm vs. 14–18 × 3.5–5 µm). A
comparison of the ITS (+5.8S) gene region of N. triseptatispora and N. chiangraiensis reveals 47 base pair differences
(9.1%) across 515 nucleotides. Therefore, N. triseptatispora
is described as a new species based on phylogeny and morphological comparison.
Phaeosphaeriaceae M.E. Barr
Fig. 52 Phylogram generated
from maximum likelihood
analysis based on combined
dataset of LSU, ITS, SSU,
TEF1 and RPB2 sequence data.
Thirteen strains are included in
the combined sequence analysis,
which comprise 4416 characters with gaps. Tree topology
of the ML analysis was similar
to the BYPP. The best scoring
RAxML tree with a final likelihood value of − 8333.195072
is presented. The matrix had
327 distinct alignment patterns, with 24.82% of undetermined characters or gaps.
Estimated base frequencies
were as follows: A = 0.252190,
C = 0.233838, G = 0.264447,
T = 0.249525; substitution rates:
AC = 1.127465, AG = 2.895211,
AT = 2.022869, CG = 0.534488,
CT = 8.448227, GT = 1.000000;
gamma distribution shape
parameter α = 0.020000. Bootstrap support values for ML
equal to or greater than 60% and
BYPP equal to or greater than
0.90 are given above the nodes.
Newly generated sequences are
in dark red bold and type species are in bold. Phaeosphaeria
oryzae (CBS 110110) is used as
outgroup taxon
13
Phaeosphaeriaceae was introduced by Barr (1979) with
Phaeosphaeria as type genus and monographed most
recently by Phookamsak et al. (2014). Fifty-two genera were
accepted in Wijayawardene et al. (2018). Subsequently, Yang
et al. (2019) introduced a new genus, Neostagonosporella
with Neostagonosporella sichuanensis as a new species.
Zhang et al. (2019) introduced two new genera Hydeopsis and Pseudoophiosphaerella from Karst landforms in
Guizhou Province, China, based on morphology and phylogeny. Recently, Phookamsak et al. (2019) introduced a
new genus, Brunneomurispora from China. Divergence time
estimates for this family are crown age of 75 Mya (46–102)
and stem age of 99 Mya (73–129) in the Cretaceous (Liu
et al. 2017).
Leptospora Rabenh.
Leptospora was introduced by Rabenhorst (1857) with
L. rubella as the type species. Hyde et al. (2016b) reported
a reference specimen of L. rubella from UK with two new
species L. galii from Italy and L. thailandica from Thailand.
Fungal Diversity
Zhang et al. (2019) introduced a new species, L. hydei from
China. We introduce two new Leptospora species with a
new host record of L. thailandica from C. odorata, together
with descriptions and illustrations (Figs. 53, 54, 55, 56). A
phylogenetic tree based on combined LSU, ITS, SSU, TEF1
and RPB2 sequence data is presented in Fig. 52.
Leptospora chromolaenae Mapook & K.D. Hyde, sp. nov.
Index Fungorum number: IF557289, Facesoffungi number: FoF 07803; Fig. 53
Etymology: Name reflects the host genus Chromolaena,
from which this species was isolated.
Holotype: MFLU 20-0313
Saprobic on dead stems of Chromolaena odorata. Sexual
morph: Ascomata 200–260 µm high × 200–250 µm diam.
( x̄ = 230 × 220 µm, n = 10), semi-immersed to superficial,
solitary or scattered, appearing as dark spots, coriaceous,
globose to obpyriform, reddish brown or dark brown to
black. Ostiolar neck protruding, with hyaline periphyseslike. Peridium (5–)10–20 µm wide, 2–3 layers, inner layers comprising of hyaline to pale brown cells of textura
angularis, outer layers comprising of reddish brown to yellowish brown cells of textura epidermoidea. Hamathecium
comprising 2–3.5 µm wide, cylindrical, septate, branching pseudoparaphyses, anastomosing above the asci. Asci
80–115 × 9.5–15 µm ( x̄ = 93 × 11 µm, n = 20), overlapping,
8-spored, bitunicate, fissitunicate, cylindric-clavate, straight
or slightly curved, apically rounded, with a short, bulbous
pedicel, ocular chamber visible when immature. Ascospores
60–95 × 2.5–3.5 µm ( x̄ = 77 × 3.2 µm, n = 25), fasciculate,
in parallel, scolecosporous, hyaline to pale brown, filiform,
(22–)27–28-septate, minute guttule in each cell, straight or
slightly curved, with globose polar appendages, observed
clearly when mounted in Indian ink. Asexual morph:
Undetermined.
Culture characteristics: Ascospores germinating on
MEA within 24 h. at room temperature and germ tubes produced from both ends. Colonies on MEA circular, mycelium slightly raised, moderately fluffy, entire to filamentous,
cultures white to greyish-brown at the surface and initially
creamy-white in reverse, becoming dark brown from the centre of the colony with creamy-white at the margin (Fig. 56a).
Pre-screening for antimicrobial activity: Leptospora
chromolaenae (MFLUCC 17-1421) showed antimicrobial
activity against B. subtilis with a 13 mm inhibition zone and
against M. plumbeus with a 20 mm inhibition zone, observable as partial inhibition, when compared to the positive
control (26 mm and 17 mm, respectively), but no inhibition
of E. coli.
Material examined: THAILAND, Chiang Rai Province,
Doi Pui, on dead stems of Chromolaena odorata, 8 July
2015, A. Mapook (DP2, MFLU 20-0313, holotype); extype culture MFLUCC 17-1421; 5 August 2015, A. Mapook
(DP41, MFLU 20-0314); living culture MFLUCC 17-1448;
(DP44, MFLU 20-0315); living culture MFLUCC 17-1451;
Lampang Province, Chaehom, on dead stems of C. odorata,
24 September 2016, A. Mapook (JH6, MFLU 20-0317);
living culture MFLUCC 17-1460; (JH19, MFLU 20-0316);
living culture MFLUCC 17-1501.
GenBank numbers: LSU: MN994552, MN994553,
MN994554, MN994555, MN994556, ITS: MN994575,
MN994576, MN994577, MN994578, MN994579, SSU:
MN994598, MN994599, MN994600, MN994601,
MN994602, TEF1: MN998156, MN998157, MN998158,
MN998159, MN998160
Notes: In a BLASTn search of NCBI GenBank, the
closest match with the ITS sequence of Leptospora chromolaenae (MFLUCC 17-1421, ex-holotype) with 95.93%
similarity was L. rubella (strain CAP17F, JQ936327). The
closest match with the LSU sequence with 99.88% similarity
was L. rubella (strain CPC 11006, DQ195792). The closest
match with the SSU sequence with 99.91% similarity was
Phaeosphaeria sp. (strain sn23-1, EU189215). The closest
match with the TEF1 sequence with 96.04% similarity was
L. hydei (strain GZCC 19-0004, MK523387), while the closest match with the RPB2 sequence with 77.11% similarity
was Parastagonospora fusiformis (strain MFLUCC 13-0215,
KX863711). In the present phylogenetic analysis, L. chromolaenae forms a monophyletic clade and clusters with the
clade comprising, L. hydei, L. phraeana and L. thailandica
with hight bootstrap support (89% ML and 1.00 BYPP,
Fig. 52). However, L. chromolaenae differs from these species in having (22–)27–28-septate ascospores (Table 11).
Therefore, L. chromolaenae is described as a new species
based on phylogeny and morphological comparison.
Leptospora phraeana Mapook & K.D. Hyde, sp. nov.
Index Fungorum number: IF557291, Facesoffungi number: FoF 07804; Fig. 54
Etymology: Referring to the location where the specimen
was collected, Phrae Province, Thailand.
Holotype: MFLU 20-0318
Saprobic on dead stems of Chromolaena odorata. Sexual
morph: Ascomata (130–)180–235 µm high × 125–185 µm
diam. ( x̄ = 200.5 × 155 µm, n = 10), semi-immersed, solitary
or scattered, appearing as dark spots, coriaceous, globose
to obpyriform, reddish brown to yellowish brown or brown.
Ostiolar neck protruding, with hyaline periphyses-like cells.
Peridium (10–)15–20 µm wide, 2–3 layers, comprising light
brown to brown cells of textura angularis. Hamathecium
comprising 2–3.5 µm wide, cylindrical, septate, branching pseudoparaphyses, anastomosing above the asci. Asci
(60–)70–90 × 8–13 µm ( x̄ = 80 × 11 µm, n = 10), overlapping, 8-spored, bitunicate, fissitunicate, cylindric-clavate,
straight or slightly curved, apically rounded, with a short
pedicel, ocular chamber observed clearly when immature.
13
Fungal Diversity
Fig. 53 Leptospora chromolaenae (holotype) a, b Appearance of superficial ascomata on
substrate. c Section through an
ascoma. d Ostiole. e Peridium. f
Pseudoparaphyses. g, h Asci. i–l
Ascospores with globose polar
appendages. m Ascospores
with polar appendages in Indian
ink. Scale bars: a, b = 500 µm,
c, g–m = 50 µm, d = 20 µm, e,
f = 10 µm
Ascospores 50–70 × 2–3 µm ( x̄ = 63 × 2.6 µm, n = 15), fasciculate, in parallel, scolecosporous, pale brown to yellowish brown, cylindrical to broadly filiform, 13-septate with
minute guttule in each cell, straight or slightly curved, with
polar appendages. Asexual morph: Undetermined.
Culture characteristics: Ascospores germinating on
MEA within 48 h. at room temperature and germ tubes produced from both ends. Colonies on MEA circular, mycelium
slightly raised, moderately fluffy, entire, cultures white at the
surface and initially creamy-white to pale brown in reverse,
becoming dark brown from the centre of the colony with
creamy-white at the margin (Fig. 56b).
13
Pre-screening for antimicrobial activity: Leptospora
phraeana (MFLUCC 17-1478) showed antimicrobial activity against M. plumbeus with a 22 mm inhibition zone,
observable as partial inhibition, when compared to the positive control (17 mm), but no inhibition of B. subtilis and E.
coli.
Material examined: THAILAND, Phrae Province, Doi
Pha Klong, on dead stems of Chromolaena odorata, 22 September 2016, A. Mapook (DPKP2, MFLU 20-0318, holotype); ex-type culture MFLUCC 17-1478.
GenBank numbers: LSU: MN994557, ITS: MN994580,
SSU: MN994603, TEF1: MN998161
Fungal Diversity
Fig. 54 Leptospora phraeana
(holotype) a, b Appearance
of superficial ascomata on
substrate. c Section through
ascoma. d Ostiole. e Peridium.
f Pseudoparaphyses. g, h
Asci. i–m Ascospores with
polar appendages. Scale bars:
a = 500 µm, b = 200 µm, c, g,
h = 50 µm, d, e, i–m = 20 µm,
f = 10 µm
Notes: In a BLASTn search of NCBI GenBank, the closest match with the ITS sequence of Leptospora phraeana
(MFLUCC 17-1478, ex-holotype) with 96.26% similarity
was L. rubella (strain CAP17F, JQ936327). The closest
match with the LSU sequence with 99.65% similarity was L.
rubella (strain CPC 11006, DQ195792). The closest match
with the SSU sequence with 100% similarity was Phaeosphaeria sp. (strain sn23-1, EU189215). The closest match
with the TEF1 sequence with 95.70% similarity was L. hydei
(strain GZCC 19-0004, MK523387), while the closest match
with the RPB2 sequence with 78.72% similarity was Neophaeosphaeria filamentosa (strain CBS 102202, GU371773).
In the present phylogenetic analysis, L. phraeana forms
a separate branch and clusters with L. thailandica with
hight bootstrap support (85% ML and 1.00 BYPP, Fig. 52).
However, L. phraeana differs from L. thailandica in having slightly smaller asci [(60–)70–90 × 8–13 µm vs.
68–114 × 7–13 µm] and slightly smaller ascospores
(50–70 × 2–3 µm vs. 63–89 × 1.8–3.8 µm) that are 13-septate, while L. thailandica has (14–)20–22-septate ascospores
(Table 11). A comparison of the ITS (+5.8S) gene region of
L. phraeana and L. thailandica reveals 17 base pair differences (3.3%) across 506 nucleotides. Therefore, L. phraeana
is described as a new species based on phylogeny and morphological comparison.
13
Fungal Diversity
Fig. 55 Leptospora thailandica (new host record) a, b
Appearance of superficial
ascomata on substrate. c Section
through ascoma. d Ostiole. e
Peridium. f Pseudoparaphyses.
g, h Asci. i–m Ascospores with
polar appendages. Scale bars:
a = 200 µm, b = 100 µm, c, g,
h = 50 µm, d, e, i–m = 20 µm,
f = 5 µm
Fig. 56 Culture characteristics
on MEA: a Leptospora chromolaenae (MFLUCC 17-1421).
b Leptospora phraeana
(MFLUCC 17-1478). c Leptospora thailandica (MFLUCC
17-1468)
Leptospora thailandica Phukhams. & K.D. Hyde, in
Hyde et al., Fungal Divers. 80: 100 (2016)
Facesoffungi number: FoF 02381; Fig. 55
13
Saprobic on dead stems of Chromolaena odorata. Sexual
morph: Ascomata 150–170 µm high × 120–160 µm diam.
( x̄ = 162 × 136 µm, n = 5), immersed to semi-immersed,
This study
63–89 × 1.8–3.8
(60–)75–90(–100) × 2.5–
3.5
68–114 × 7–13
(65–)70–115 × 9.5–15
13–15-septate, with polar
appendages
Hyde et al. (2016b)
This study
13-septate, with polar
appendages
(14–)20–22-septate
50–70 × 2–3
(130–)180–235
high × 125–185 diam.
188–207 high × 112–170
diam.
150–170 high × 120–160
diam.
L. phraeana (MFLUCC
17-1478)
L. thailandica (MFLUCC
16-0385)
L. thailandica (MFLUCC
17-1468)
(60–)70–90 × 8–13
Zhang et al. (2019)
(104–)124–138(–
146) × 4–4.8
196–267 high × 168–187
diam.
L. hydei (GZCC 19-0004)
147–165(–180) × 13–17(–
19.5)
11-septate
This study
(22–)27–28-septate, with
polar appendages
60–95 × 2.5–3.5
80–115 × 9.5–15
(5–)10–20, 2–3 layers
of textura angularis to
textura epidermoidea
11–24, several layers of
textura angularis to
prismatica
(10–)15–20, 2–3 layers of
textura angularis
(5–)10–24(–27), 5–7 layers
of textura angularis
10–25, 2–4 layers of
textura angularis
200–260 high × 200–250
diam.
L. chromolaenae
(MFLUCC 17-1448)
Ascospores (µm)
Asci (µm)
Peridium (µm)
Asocomata (µm)
Species
Table 11 Synopsis of Leptospora species with similar morphological features discussed in this study
Septation of ascospores
References
Fungal Diversity
solitary or scattered, appearing as dark spots, coriaceous,
globose to obpyriform, reddish brown to brown. Ostiolar
neck protruding. Peridium 10–25 µm wide, 2–4 layers,
inner layers comprising hyaline cells of textura angularis, outer layers comprising light brown to brown cells of
textura angularis. Hamathecium comprising 2–2.5 µm wide,
oblong to cylindrical, septate, branching pseudoparaphyses,
anastomosing above the asci. Asci (65–)70–115 × 9.5–15 µm
( x̄ = 89 × 13 µm, n = 15), overlapping, 8-spored, bitunicate, fissitunicate, cylindric-clavate, straight or slightly
curved, apically rounded, with a short, bulbous pedicel, ocular chamber visible when immature. Ascospores
(60–)75–90(–100) × 2.5–3.5 µm ( x̄ = 83 × 2.9 µm, n = 15),
fasciculate, in parallel, scolecosporous, hyaline to pale
brown, cylindrical to broadly filiform, 1-septate at the
center of the ascospores when immature, 13–15-septate when mature, minute guttule in each cell, straight or
slightly curved, with polar appendages. Asexual morph:
Undetermined.
Culture characteristics: Ascospores germinating on
MEA within 24 h at room temperature and germ tubes produced from both ends. Colonies on MEA circular, mycelium slightly raised, entire, cultures white at the surface and
initially creamy-white to pale brown in reverse, becoming
creamy-brown to brown from the centre of the colony with
creamy-white at the margin (Fig. 56c).
Pre-screening for antimicrobial activity: Leptospora
thailandica (MFLUCC 17-1468) showed antimicrobial
activity against B. subtilis, E. coli and M. plumbeus (13 mm,
8 mm and 24 mm inhibition zone, respectively), observable
as partial inhibition, when compared to the positive control
(26 mm, 9 mm, and 17 mm, respectively).
Known hosts and distribution: On dead branches of
Duranta sp. (Verbenaceae) in Thailand (Hyde et al. 2016a,
b)
Material examined: THAILAND, Chiang Rai Province,
Doi Mae Salong, on dead stems of Chromolaena odorata,
8 April 2017, A. Mapook (DMS5, MFLU 20-0319); living
culture MFLUCC 17-1468 (new host record).
GenBank numbers: LSU: MN994558, ITS: MN994581,
SSU: MN994604, TEF1: MN998162
Notes: In a BLASTn search of NCBI GenBank, the closest match with the ITS sequence of MFLUCC 17-1468 with
99.65% similarity was L. thailandica (strain MFLUCC
16-0385, NR_154133). The closest match with the LSU
sequence with 99.40% similarity was L. hydei (strain
GZCC 19-0004, MK522497). The closest match with the
SSU sequence with 99.90% similarity was Phaeosphaeria
sp. (strain sn23-1, EU189215). The closest match with the
TEF1 sequence with 96.02% similarity was Populocrescentia ammophilae (strain MFLUCC 17-0665, MG829231),
while the closest match with the RPB2 sequence with
80.18% similarity was Paraphoma fimeti (strain UTHSC:
13
Fungal Diversity
Fig. 57 Phylogram generated
from maximum likelihood
analysis based on combined
dataset of LSU, ITS, SSU
and TEF1 sequence data. One
hundred sixty-seven strains
are included in the combined
sequence analysis, which
comprise 3312 characters with
gaps. The best scoring RAxML
tree with a final likelihood
value of − 33364.909121 is
presented. The matrix had
1254 distinct alignment patterns, with 21.74% of undetermined characters or gaps.
Estimated base frequencies
were as follows: A = 0.241300,
C = 0.264151, G = 0.235962,
T = 0.258586; substitution rates:
AC = 0.989272, AG = 3.620674,
AT = 1.917765, CG = 0.739088,
CT = 4.661336, GT = 1.000000;
gamma distribution shape
parameter α = 0.156765. Bootstrap support values for ML
equal to or greater than 60%
and BYPP equal to or greater
than 0.90 are given at the nodes.
Newly generated sequences
are in dark red bold and type
species are in bold. Staurosphaeria rhamnicola (MFLUCC
17-0814) and S. rhamnicola
(MFLUCC 17-0813) are used as
outgroup taxa
13
Fungal Diversity
Fig. 57 (continued)
DI16-296, LT797032). In the present phylogenetic analysis, MFLUCC 17-1468 clusters with L. thailandica strain
MFLUCC 16-0385 with hight bootstrap support (100% ML
and 1.00 BYPP, Fig. 52). Therefore, we identify MFLUCC
17-1468 as Leptospora thailandica based on phylogenetic
analyses with morphological comparison (Table 11) and the
isolates are introduced here as a new host record from Chromolaena odorata collected in Thailand.
Murichromolaenicola Mapook & K.D. Hyde, gen. nov.
Index Fungorum number: IF557338, Facesoffungi number: FoF 07805
Etymology: The generic epithet reflects the muriform ascospore, which was growing on the host genus
Chromolaena.
Saprobic on dead stems. Sexual morph: Ascomata semiimmersed to superficial, solitary or scattered, appearing as
dark spots, coriaceous, globose to obpyriform, light brown to
brown. Ostiolar neck protruding, with hyaline periphyses-like
cells. Peridium comprising several layers of brown to dark
13
Fungal Diversity
brown cells of textura angularis. Hamathecium composed
of cylindrical, septate, branching pseudoparaphyses. Asci
8-spored, bitunicate, fissitunicate, cylindric-clavate, straight
or slightly curved, apically rounded, pedicellate. Ascospores
overlapping, 1–2-seriate, initially hyaline to golden-yellow, 1-septate when immature, becoming golden-brown to
brown at maturity, ellipsoid to broadly fusiform, muriform,
5–7-transversely septate, with 1–2-vertical septum, guttulate,
slightly constricted at the central septum, straight or slightly
curved, surrounded by hyaline gelatinous sheath observed
clearly when mounted in Indian ink. Asexual morph: Conidiomata pycnidial, solitary, immersed to semi-immersed in the
host surface, uni-loculate, globose, yellowish brown to brown.
Pycnidial wall comprising 4–5 layers of thick-walled, dense,
yellowish brown to brown cells of textura angularis. Conidiophores reduced to conidiogenous cells. Conidiogenous cells
phialidic, ampulliform to cylindrical, hyaline, unbranched.
Conidia ellipsoid to broadly fusiform, muriform, 5–7-transversely septate, with 1–2-vertical septa, small guttule, not constricted at the septa, yellowish brown to brown, with polar
appendages, observed clearly when mounted in Indian ink.
Type species: Murichromolaenicola chromolaenae
Mapook & K.D. Hyde
Notes: A phylogenetic analyses based on combined dataset of LSU, ITS, SSU and TEF1 sequence data show that
two new Murichromolaenicola species form a distinct lineage with Wojnowicia and Wojnowiciella with low bootstrap support (Fig. 57). Murichromolaenicola species are
distinct morphologially in having ellipsoid to broadly fusiform, muriform ascospores, 5–7-transversely septate with
1–2-vertical septa, while Wojnowiciella has subcylindrical
to ellipsoid conidia and Wojnowicia has fusiform to cylindrical conidia. Therefore, we introduce Murichromolaenicola
as a new genus with two new species M. chiangraiensis and
M. chromolaenae, based on morphology and phylogeny,
together with a comparison of the TEF1 gene region.
Murichromolaenicola chiangraiensis Mapook & K.D.
Hyde, sp. nov.
Index Fungorum number: IF557339, Facesoffungi number: FoF 07806; Fig. 58
Etymology: Referring to the location where the specimen
was collected, Chiang Rai Province, Thailand.
Holotype: MFLU 20-0326
Saprobic on dead stems of Chromolaena odorata. Sexual
morph: Ascomata 460–555 µm high × (350–)500–530 µm
diam. ( x̄ = 515 × 475 µm, n = 5), semi-immersed to superficial, solitary or scattered, appearing as dark spots, coriaceous,
globose to obpyriform, light brown to brown. Ostiolar neck
protruding, with hyaline periphyses-like. Peridium 30–50 µm
wide, comprising several layers of brown to dark brown
cells of textura angularis. Hamathecium comprising 3–4 µm
wide, cylindrical, septate, branching pseudoparaphyses. Asci
13
(80–)110–135 × (15–)17–20 µm ( x̄ = 117 × 18 µm, n = 10),
8-spored, bitunicate, fissitunicate, cylindric-clavate, straight
or slightly curved, apically rounded, pedicellate. Ascospores
25–35 × 10–13 µm ( x̄ = 29 × 11 µm, n = 20), overlapping,
1–2-seriate, initially hyaline to golden-yellow, 1-septate when
immature, becoming golden-brown to brown at maturity, ellipsoid to broadly fusiform, muriform, 5–7-transversely septate,
with 1–2-vertical septum, guttulate, slightly constricted at
the central septum, straight or slightly curved, surrounded by
hyaline gelatinous sheath observed clearly when mounted in
Indian ink. Asexual morph: Undetermined.
Culture characteristics: Ascospores germinating on
MEA within 24 h. at room temperature and germ tubes produced from several cells. Colonies on MEA circular, mycelium crateriform, entire, cultures white with greyish from the
centre of the colony at the surface, pale olivaceous-brown
from the centre of the colony in reverse with creamy-white
at the margin (Fig. 60a).
Pre-screening for antimicrobial activity: Not tested.
Material examined: THAILAND, Chiang Rai Province,
Doi Pui, on dead stems of Chromolaena odorata, 2 April
2017, A. Mapook (DP86, MFLU 20-0326, holotype); extype culture MFLUCC 17-1488.
GenBank numbers: LSU: MN994559, ITS: MN994582,
SSU: MN994605, TEF1: MN998163
Notes: In a BLASTn search of NCBI GenBank, the closest
match with the ITS sequence of Murichromolaenicola chiangraiensis (MFLUCC 17-1488, ex-holotype) with 94.02% similarity was Wojnowicia dactylidicola (strain MFLUCC 13-0738,
NR_154501). The closest match with the LSU and TEF1
sequences with 91.68% (KX430001) and 83.80% (KX430003)
similarity, respectively, were W. italica strain MFLU 14-0732,
while the closest match with the SSU sequence with 93.15%
similarity was Pyrenochaeta corni (strain CBS 102828,
GQ387548). In the present phylogenetic analysis, M. chiangraiensis clusters with M. chromolaenae (MFLUCC 17-1489)
with high bootstrap support (100% ML and 1.00 BYPP,
Fig. 52). Although we could not compare the morphological
characteristics of those species; M. chiangraiensis is found as
sexual morph in nature and we could not obtain its asexual
morph in culture, while M. chromolaenae is found as asexual
morph in nature, they differ in culture characteristics on MEA
(Fig. 60). A comparison of the TEF1 gene region of M. chiangraiensis and M. chromolaenae reveals 41 base pair differences
(4.7%) across 872 nucleotides. Therefore, M. chiangraiensis is
described here as a new species based on phylogeny.
Murichromolaenicola chromolaenae Mapook & K.D.
Hyde, sp. nov.
Index Fungorum number: IF557340, Facesoffungi number: FoF 07807; Fig. 59
Etymology: Name reflects the host genus Chromolaena,
from which this species was isolated.
Fungal Diversity
Fig. 58 Murichromolaenicola
chiangraiensis (holotype)
a, b Appearance of ascomata on substrate. c Section
through ascoma. d Peridium.
e Pseudoparaphyses. f–i Asci.
j–n Ascospores. o Ascospores
with gelatinous sheath in Indian
ink. Scale bars: a = 500 µm,
b = 200 µm, c = 100 µm,
f–i = 50 µm, d, j–o = 20 µm,
e = 5 µm
Holotype: MFLU 20-0327
Saprobic on dead stems of Chromolaena odorata.
Sexual morph: Undetermined. Asexual morph: Conidiomata 200–235 µm high × 195–230(–260) µm diam.
( x̄ = 228 × 226 µm, n = 5), pycnidial, solitary, immersed to
semi-immersed in the host surface, uni-loculate, globose,
yellowish brown to brown. Pycnidial wall 10–20 µm wide,
comprising 4–5 layers of thick-walled, dense, yellowish
brown to brown cells of textura angularis. Conidiophores
reduced to conidiogenous cells. Conidiogenous cells phialidic, ampulliform to cylindrical, hyaline, unbranched.
Conidia 14–25 × 6.5–11 μm ( x̄ = 20.5 × 8.5 µm, n = 50),
ellipsoid to broadly fusiform, muriform, 5–7-transversely
septate, with 1–2-vertical septa, small guttule, not constricted at the septa, yellowish brown to brown, with polar
appendages, observed clearly when mounted in Indian ink.
Culture characteristics: Conidia germinating on MEA
within 24 h at room temperature and germ tubes produced
13
Fungal Diversity
Fig. 59 Murichromolaenicola
chromolaenae (holotype) a,
b Appearance of conidiomata
on substrate. c Section through
conidioma. d Peridium. e–g
Conidiogenous cells and developing conidia. h–j Conidia.
k Conidia with polar appendages in Indian ink. Scale bars:
a, b = 500 µm, c = 100 µm,
d–k = 10 µm
from both ends. Colonies on MEA circular, mycelium crateriform, undulate, cultures white with greyish from the centre
of the colony at the surface, olivaceous from the centre of the
colony in reverse with creamy-white at the margin (Fig. 60b).
Pre-screening for antimicrobial activity: Murichromolaenicola chromolaenae (MFLUCC 17-1489) showed antimicrobial activity against E. coli with a 17 mm inhibition
zone, when compared to the positive control (9 mm), but no
inhibition of B. subtilis and M. plumbeus.
Material examined: THAILAND, Chiang Rai Province,
Doi Pui, on dead stems of Chromolaena odorata, 2 April
2017, A. Mapook (DP88, MFLU 20-0327, holotype); extype culture MFLUCC 17-1489.
GenBank numbers: LSU: MN994560, ITS: MN994583,
SSU: MN994606, TEF1: MN998164
Notes: In a BLASTn search of NCBI GenBank, the closest
match with the ITS sequence of Murichromolaenicola chromolaenae (MFLUCC 17-1489, ex-holotype) with 90.65% similarity was Phaeosphaeria herpotrichoides (strain CBS 306.71,
MH860137). The closest match with the LSU sequence with
98.90% similarity was Allophaeosphaeria muriformia (strain
MFLUCC 13-0277, KX910089). The closest match with the
SSU sequence with 99.82% similarity was Phaeosphaeria avenaria f. sp. triticae (strain ATCC 26370, EU189210), while the
closest match with the TEF1 sequence with 99.82% similarity
was Wojnowicia italica (strain MFLU 14-0732, KX430003).
In the present phylogenetic analysis, M. chromolaenae clusters
13
Fig. 60 Culture characteristics on MEA: a Murichromolaenicola
chiangraiensis (MFLUCC 17-1488). b Murichromolaenicola chromolaenae (MFLUCC 17-1489)
with M. chiangraiensis (MFLUCC 17-1488) with high bootstrap support (100% ML and 1.00 BYPP, Fig. 52). Although
we could not compare the morphological characteristics of
these species; M. chromolaenae is found as asexual morph in
nature, while M. chiangraiensis is found as sexual morph in
nature and we could not obtain its asexual morph in culture,
but they differ in culture characteristics on MEA (Fig. 60).
A comparison of the TEF1 gene region of M. chromolaenae and M. chiangraiensis reveals 41 base pair differences
(4.7%) across 872 nucleotides. Therefore, M. chromolaenae
is described as a new species based on phylogeny.
Neoophiobolus Mapook & K.D. Hyde, gen. nov.
Fungal Diversity
Index Fungorum number: IF557343, Facesoffungi number: FoF 07808
Etymology: Etymology: Referring to its similarity with
Ophiobolus.
Saprobic on dead stems. Sexual morph: Ascomata
immersed to semi-immersed, solitary, scattered, globose to subglobose, coriaceous, dark brown. Ostiole short papillate, with
hyaline periphyses-like. Peridium comprising several layers of
hyaline or pale brown to dark brown, pseudoparenchymatous
cells, arranged in a textura angularis. Hamathecium composed
of cylindrical to broadly filiform, septate, branching pseudoparaphyses, anastomosing above the asci. Asci overlapping,
8-spored, bitunicate, cylindrical to cylindric-clavate, straight
or slightly curved, apically rounded, with ocular chamber visible when immature. Ascospores fasciculate, in parallel, scolecosporous, hyaline to pale yellow brown, filiform, 10–20-septate with minute guttule in each cell, slightly curved, constricted
at the central septum where the spore separates into two parts,
without polar appendages. Asexual morph: Undetermined.
Type species: Neoophiobolus chromolaenae Mapook &
K.D. Hyde
Notes: Neoophiobolus is similar to the genus Ophiobolus
in having scolecosporous ascospores with a swollen cell.
However, phylogenetic analyses based on combined dataset
of LSU, ITS, SSU and TEF1 sequence data show that N. chromolaenae is phylogenetically distant from Ophiobolus and
forms a distinct lineage within the family Phaeosphaeriaceae
(Fig. 52). Therefore, we introduce Neoophiobolus as a new
genus to accommodate a new species N. chromolaenae.
Neoophiobolus chromolaenae Mapook & K.D. Hyde, sp.
nov.
Index Fungorum number: IF557344, Facesoffungi number: FoF 07809; Fig. 61
Etymology: Name reflects the host genus Chromolaena,
from which this species was isolated.
Holotype: MFLU 20-0335
Saprobic on dead stems of Chromolaena odorata. Sexual
morph: Ascomata 230–400 µm high × 200–350 µm diam.
( x̄ = 323.5 × 296 µm, n = 5), immersed to semi-immersed,
solitary, scattered, globose to subglobose, coriaceous, dark
brown. Ostiole short papillate, with hyaline periphyses-like.
Peridium 15–30 µm wide, comprising several layers of hyaline or pale brown to dark brown, pseudoparenchymatous
cells, arranged in a textura angularis. Hamathecium comprising 1.5–2.5 µm wide, cylindrical to broadly filiform, septate,
branching pseudoparaphyses, anastomosing above the asci.
Asci (90–)100–135 × 8–13 µm ( x̄ = 115 × 10 µm, n = 20), overlapping, 8-spored, bitunicate, cylindrical to cylindric-clavate,
straight or slightly curved, apically rounded, with ocular chamber visible when immature. Ascospores 85–110 × 2.5–3.5 µm
( x̄ = 97 × 3 µm, n = 20), fasciculate, in parallel, scolecosporous, hyaline to pale yellow brown, filiform, 10–20-septate with
minute guttule in each cell, slightly curved, constricted at the
central septum where the spore separates into two parts, without polar appendages. Asexual morph: Undetermined.
Culture characteristics: Ascospores germinating on
MEA within 24 h. at room temperature and germ tubes produced from both ends. Colonies on MEA circular, mycelium slightly raised, moderately fluffy, entire to filamentous,
cultures white at the surface, creamy-white in reverse with
brown to dark brown from the centre of the colony (Fig. 62).
Pre-screening for antimicrobial activity: Neoophiobolus chromolaenae (MFLUCC 17-1467) showed no inhibition
of E. coli, B. subtilis and M. plumbeus.
Material examined: THAILAND, Chiang Rai Province, Doi
Mae Salong, on dead stems of Chromolaena odorata, 8 April
2017, A. Mapook (DMS4, MFLU 20-0335, holotype); ex-type
culture MFLUCC 17-1467; Chiang Rai Province, Doi Pui, on
dead stems of Chromolaena odorata, 5 August 2015, A. Mapook
(DP42, MFLU 20-0336); living culture MFLUCC 17-1449.
GenBank numbers: LSU: MN994561, MN994562, ITS:
MN994584, MN994585, SSU: MN994607, MN994608,
TEF1: MN998165, MN998166
Notes: In a BLASTn search of NCBI GenBank, the closest match with the ITS and TEF1 sequences of Neoophiobolus chromolaenae (MFLUCC 17-1467, ex-holotype) with
99.31% (LT796836) and 99.16% (LT797076) similarity, respectively, was Trematophoma sp. strain UTHSC:
DI16-210. The closest match with the LSU sequence
with 99.12% similarity was Paraphoma fimeti (strain CBS
127796, MH876144), while the closest match with the SSU
sequences with 96.89% similarity was Ophiosphaerella narmari (strain ATCC 64688, KC848510).
Paraleptospora Mapook & K.D. Hyde, gen. nov.
Index Fungorum number: IF557347, Facesoffungi number: FoF 07810
Etymology: Referring to its similarity with Leptospora.
Saprobic on dead stems. Sexual morph: Ascomata
immersed to semi-immersed, solitary or scattered, gregarious, coriaceous, globose or subglobose to ampulliform, brown
to dark brown, appearing as dark spot with red area on host
surface. Ostiole short papillate. Peridium several layers, comprising dense, thick-walled, reddish brown to dark brown,
pseudoparenchymatous cells, arranged in textura angularis.
Hamathecium composed of filiform or broadly filiform to cylindrical, septate, branching, pseudoparaphyses, anastomosing
above the asci. Asci 8-spored, bitunicate, fissitunicate, cylindrical to cylindric-subclavate, straight or slightly curved, pedicellate, apically rounded, with an ocular chamber. Ascospores
overlapping, 1–3-seriate, hyaline or pale yellow to yellowish
brown, cylindric-fusiform, tapering towards narrow the rounded
ends, (5–)6–7(–8)-septate, broader at the center and slightly
constricted at septa, straight to slightly curved, guttulate, with
or without polar appendages. Asexual morph: Undetermined.
13
Fungal Diversity
Fig. 61 Neoophiobolus
chromolaenae (holotype) a,
b Appearance of superficial
ascomata on substrate. c Section
through ascoma. d Ostiole. e
Peridium. f Pseudoparaphyses.
g, h Asci. i–m Ascospores.
Scale bars: a = 500 µm,
b = 200 µm, c = 100 µm, d, g,
h = 50 µm, e, i–m = 20 µm,
f = 5 µm
Type species: Paraleptospora chromolaenae Mapook &
K.D. Hyde
Notes: Paraleptospora is similar to the genus Leptospora
in staining the host surface red. However, phylogenetic analyses based on combined dataset of LSU, ITS, SSU and TEF1
sequence data show that Paraleptospora species are phylogenetically distant from Leptospora and form a sister lineage with Acericola, Jeremyomyces and Phaeosphaeriopsis (Fig. 52). Therefore,
we introduce Paraleptospora as a new genus to accommodate
two new species P. chromolaenae and P. chromolaenicola.
Paraleptospora chromolaenae Mapook & K.D. Hyde, sp. nov.
Index Fungorum number: IF557348, Facesoffungi number: FoF 07811; Fig. 63
13
Etymology: Name reflects the host genus Chromolaena,
from which this species was isolated.
Holotype: MFLU 20-0343
Saprobic on dead stems of Chromolaena odorata. Sexual
morph: Ascomata 220–305 µm high × 210–340 µm diam.
( x̄ = 265 × 260 µm, n = 5), immersed to semi-immersed, solitary or scattered, gregarious, globose to subglobose, coriaceous, dark brown, appearing as dark spot with red area on host
surface. Ostiole short papillate. Peridium 15–20 µm wide, several layers, comprising dense, thick-walled, reddish brown to
dark brown, pseudoparenchymatous cells, arranged in textura
angularis. Hamathecium comprising 1–2 µm wide, broadly
filiform, septate, branching, pseudoparaphyses, anastomosing
above the asci. Asci 80–120 × 7.5–11 µm ( x̄ = 100 × 9 µm,
Fungal Diversity
present phylogenetic analysis, P. chromolaenae clusters with
P. chromolaenicola with high bootstrap support (100% ML
and 1.00 BYPP, Fig. 52). However, P. chromolaenae differs
from P. chromolaenicola in having slightly larger ascomata
(220–305 × 210–340 µm vs. 140–310 × 120–300 µm) and
slightly larger ascospores (30–40 × 3–4 µm vs. 28–38 × 3–4 µm)
with 6–7-septa, and without polar appendages, while P. chromolaenicola has (5–)7–8-septate ascospores, and very few
ascospores with polar appendages (Table 12). A comparison
of the ITS (+5.8S) gene region of P. chromolaenae and P. chromolaenicola reveals 23 base pair differences (4.6%) across 503
nucleotides. Therefore, P. chromolaenae is described as a new
species based on phylogeny and morphological comparison.
Fig. 62 Culture characteristic on MEA: Neoophiobolus chromolaenae (MFLUCC 17-1467)
n = 30), 8-spored, bitunicate, fissitunicate, cylindrical to
cylindric-subclavate, slightly curved, pedicellate, apically
rounded, with an ocular chamber. Ascospores 30–40 × 3–4 µm
( x̄ = 35 × 3.5 µm, n = 20), overlapping, 1–2-seriate, hyaline to
pale yellow, cylindric-fusiform, tapering towards narrow the
rounded ends, 6–7-septate, broader at the center and slightly
constricted at septa, straight to slightly curved, guttulate, without polar appendages. Asexual morph: Undetermined.
Pre-screening for antimicrobial activity: Paraleptospora chromolaenae (MFLUCC 17-1481) showed antimicrobial activity against E. coli with an 8 mm inhibition zone,
when compared to the positive control (9 mm), but no inhibition of B. subtilis and M. plumbeus.
Culture characteristics: Ascospores germinating on
MEA within 48 h. at room temperature and germ tubes produced from both ends. Colonies on MEA circular, mycelium
umbonate, cultures white at the surface, creamy-white in
reverse (Fig. 65a).
Material examined: THAILAND, Lampang Province,
Ngao, on dead stems of Chromolaena odorata, 21 September 2016, A. Mapook (LP3, MFLU 20-0343, holotype); extype culture MFLUCC 17-1481.
GenBank numbers: LSU: MN994563, ITS: MN994586,
SSU: MN994609, TEF1: MN998167
Notes: In a BLASTn search of NCBI GenBank, the closest
match with the ITS sequence of Paraleptospora chromolaenae
(MFLUCC 17-1481, ex-holotype) with 91.36% similarity was
Phaeosphaeriaceae sp. (strain MUT 4404, KC339239). The
closest match with the LSU sequence with 99.34% similarity
was Neostagonospora arrhenatheri (strain MFLUCC 15–0464,
KX910091). The closest match with the SSU sequence with
99.71% similarity was Parastagonospora nodorum (strain
LSNZN10, MH269310), while the closest match with the
TEF1 sequence with 94.91% similarity was Yunnanensis
phragmitis (strain MFLUCC 17-0365, MF683625). In the
Paraleptospora chromolaenicola Mapook, Samarakoon &
K.D. Hyde, sp. nov.
Index Fungorum number: IF557349, Facesoffungi number: FoF 07812; Fig. 64
Etymology: Name reflects the host genus Chromolaena,
on which this species was growing.
Holotype: MFLU 18-0836
Saprobic on dead stems of Chromolaena odorata. Sexual
morph: Ascomata 140–310 µm high × 120–300 µm diam.,
semi-immersed, solitary or scattered, gregarious, subglobose
to ampulliform, coriaceous, brown to dark brown, appearing
as dark spot with red area on host surface. Ostiole short papillate, flat top, red stained hyphae, with periphyses. Peridium
14–27.5 μm wide, several layers, comprising dense, thick-walled,
brown to dark brown, pseudoparenchymatous cells, arranged
in textura angularis. Hamathecium comprising 1–2 µm wide,
filiform, septate, branching, pseudoparaphyses, anastomosing
above the asci. Asci 100–120 × 8–11 µm ( x̄ = 107.5 × 9.4 μm,
n = 30), 8-spored, bitunicate, fissitunicate, cylindrical to cylindric-clavate, slightly curved, pedicellate, apically rounded, with
an ocular chamber. Ascospores 28–38 × 3–4 µm (x̄ = 33 × 3.5 μm,
n = 30), overlapping, 2–3 seriate, hyaline or pale yellow to yellowish brown, cylindric-fusiform, tapering towards narrow the
rounded ends, (5–)7–8-septate, broader at the center and slightly
constricted at septa, straight to slightly curved, guttulate, very
few ascospores with polar appendages, observed clearly when
mounted in Indian ink. Asexual morph: Undetermined.
Culture characteristics: Ascospores germinating on
MEA within 24 h. at room temperature and germ tubes produced from both ends. Colonies on MEA circular, mycelium
slightly raised, moderately fluffy, entire to filamentous, cultures white at the surface, brown to dark brown from the
centre of the colony with creamy-white at the margin in
reverse (Fig. 65b).
Pre-screening for antimicrobial activity: Paraleptospora chromolaenicola (MFLUCC 17-1450) showed antimicrobial activity against E. coli with a 12 mm inhibition
zone, when compared to the positive control (9 mm), but no
inhibition of B. subtilis and M. plumbeus.
13
Fungal Diversity
Fig. 63 Paraleptospora
chromolaenae (holotype) a,
b Appearance of ascomata on
substrate. c Section through
ascoma. d Ostiole. e Peridium.
f Pseudoparaphyses. g–j Asci.
k–q Ascospores. Scale bars:
a, b = 500 µm, c = 100 µm,
g–j = 50 µm, d, e, k–q = 20 µm,
f = 5 µm
Material examined: THAILAND, Chiang Mai Province,
Mae Taeng, on dead stems of Chromolaena odorata, 1 September 2017, Milan C. Samarakoon, (SAMC016, MFLU
18-0836, holotype; HKAS 102326, isotype) ex-type culture
MFLUCC 17-2670; Chiang Rai Province, Doi Pui, on dead
13
stems of Chromolaena odorata, 5 August 2015, A. Mapook
(DP43, MFLU 20-0344); living culture MFLUCC 17-1450.
GenBank numbers: LSU: MN994564, MN994565, ITS:
MN994587, MN994588, SSU: MN994610, MN994611,
TEF1: MN998168, MN998169
Fungal Diversity
Table 12 Synopsis of Paraleptospora species with similar morphological features discussed in this study
Species
Asocomata (µm)
Peridium (µm)
Asci (µm)
Ascospores (µm)
Septation of ascospores
References
P. chromolaenae
(MFLUCC
17-1481)
P. chromolaenicola
(MFLUCC
17-1450)
220–305 high × 210–340 diam.
15–20
80–120 × 7.5–11
30–40 × 3–4
6–7-septate
This study
140–310 high × 120–300 diam.
14–27.5
100–120 × 8–11
28–38 × 3–4
(5–)7–8-septate
This study
Notes: In a BLASTn search of NCBI GenBank, the closest match with the ITS sequence of Paraleptospora chromolaenicola (MFLUCC 17-2670, ex-holotype) with 88.97%
similarity was Phaeosphaeriaceae sp. (strain MUT 4404,
KC339239). The closest match with the LSU sequence
with 99.02% similarity was Loratospora aestuarii (strain
CBS 117592, MH874575). The closest match with the
SSU sequence with 100% similarity was Neosetophoma
rosarum (strain MFLU 17-0308, NG_065146), while the
closest match with the TEF1 sequence with 95.40% similarity was Neosetophoma sp. (strain UTHSC: DI16-337,
LT797130). In the present phylogenetic analysis, P. chromolaenicola clusters with P. chromolaenae with high bootstrap support (100% ML and 1.00 BYPP, Fig. 52). However, P. chromolaenicola has (5–)7–8-septate ascospores,
and very few ascospores with polar appendages, while
P. chromolaenae differs from P. chromolaenicola in having slightly larger ascomata (220–305 × 210–340 µm vs.
140–310 × 120–300 µm) and slightly larger ascospores
(30–40 × 3–4 µm vs. 28–38 × 3–4 µm), 6–7-septate, without polar appendages (Table 12). A comparison of the ITS
(+5.8S) gene region of P. chromolaenicola and P. chromolaenae reveals 23 base pair differences (4.6%) across 503
nucleotides. Therefore, P. chromolaenicola is described
as a new species based on phylogeny and morphological
comparison.
Pseudoophiosphaerella J.F. Zhang, J.K. Liu & Z.Y. Liu
2019
Pseudoophiosphaerella was introduced by Zhang et al.
(2019) with P. huishuiensis as the type species from China,
based on morphology and phylogeny. We introduce a new
host record of Pseudoophiosphaerella huishuiensis based
on morphology and molecular data, together with a description and illustrations (Fig. 66). A phylogenetic tree based on
combined LSU, ITS, SSU, TEF1 and RPB2 sequence data
is presented in Fig. 52.
Pseudoophiosphaerella huishuiensis J.F. Zhang, J.K. Liu &
Z.Y. Liu, in Zhang, Liu, Jeewon, Wanasinghe & Liu, Mycosphere 8(1): 207 (2019)
Facesoffungi number: FoF 05836; Fig. 66
Saprobic on dead stems of Chromolaena odorata. Sexual
morph: Ascomata 275–350 µm high × 160–190 µm diam.
(x̅ = 300 × 170 µm, n = 5), immersed to semi-immersed,
solitary or scattered, appearing as dark spots, coriaceous,
subglobose to obpyriform, dark brown to black with hyaline
at side to basal. Ostiolar neck protruding, papillate, with
hyaline periphyses-like. Peridium (15–)25–50 µm wide,
comprising several layers, of hyaline to dark brown, pseudoparenchymatous cells, arranged in a textura angularis.
Hamathecium comprising 1.5–3 µm wide, cylindrical, septate, branching pseudoparaphyses, anastomosing above the
asci. Asci 140–175 × 7.5–10 µm ( x̄ = 150 × 8.5 µm, n = 20),
overlapping, 8-spored, bitunicate, cylindrical to cylindricclavate, straight or slightly curved, apically rounded, with
a small ocular chamber. Ascospores 130–170 × 2–2.5 µm
( x̄ = 147 × 2.3 µm, n = 25), fasciculate, arranged spirally in
the centre, scolecosporous, hyaline when immature, becoming pale brown to yellowish brown when mature, filiform,
15–16-septate with minute guttule in each cell, slightly
curved, not constricted at the septa, without polar appendages. Asexual morph: Undetermined.
Culture characteristics: Ascospores germinating on
MEA within 24 h. at room temperature and germ tubes
produced from several cells. Colonies on MEA irregular,
initially mycelium white with creamy-white from the centre
of the colony at the surface, slightly raised, velvety with
moderately fluffy, undulate to lobate, yellowish brown from
the centre of the colony in reverse with creamy-white at the
margin, becoming grayish-brown on surface in old cultures,
dark brown to black in reverse (Fig. 67a, b).
Pre-screening for antimicrobial activity: Pseudoophiosphaerella huishuiensis (MFLUCC 17-1453) showed antimicrobial activity against B. subtilis, E. coli and M. plumbeus
(10 mm, 8 mm and 10 mm inhibition zone, respectively),
observable as partial inhibition, when compared to the positive control (26 mm, 9 mm, and 17 mm, respectively).
Known hosts and distribution: On dead culms of unidentified herbaceous plant in China (Zhang et al. 2019).
Material examined: THAILAND, Chiang Rai Province,
Doi Pui, on dead stems of Chromolaena odorata, 5 August
13
Fungal Diversity
Fig. 64 Paraleptospora
chromolaenicola (holotype)
a, b Appearance of ascomata
on substrate. c Section through
ascoma. d Ostiole. e Peridium.
f Pseudoparaphyses. g–i Asci.
j–m Ascospores. Scale bars:
a = 1000 µm, c, d = 50 µm, e,
g–m = 20 µm, f = 10 µm
2015, A. Mapook (DP46, MFLU 20-0350); living culture
MFLUCC 17-1453 (new host record); (DP40, MFLU
20-0349); living culture MFLUCC 17-1447; Chiang Mai
Province, Fah Hom Pok, on dead stems of C. odorata, 27
September 2016, A. Mapook (FHP1, MFLU 20-0351); living culture MFLUCC 17-1463; Mae Hong Son Province,
Mae Yen, Pai, on dead stems of C. odorata, 25 June 2016, A.
Mapook (MY5, MFLU 20-0352); living culture MFLUCC
17-1471.
GenBank numbers: LSU: MN994566, MN994567,
MN994568, MN994569, ITS: MN994589, MN994590,
MN994591, MN994592, SSU: MN994612, MN994613,
MN994614, MN994615, TEF1: MN998170, MN998171,
MN998172, MN998173
Notes: In a BLASTn search of NCBI GenBank, the
closest match with the ITS and TEF1 sequences of
13
MFLUCC 17-1453 with 99.47% (MK522509) and 99.03%
(MK523389) similarity, respectively, was Pseudoophiosphaerella huishuiensis strain HS-13. The closest match with
the LSU sequence with 99.56% similarity was Dematiopleospora alliariae (strain MFLUCC 13-0070, KX494877),
while the closest match with the SSU sequence with 99.46%
similarity was Phaeosphaeriaceae sp. (strain SYPF 7948,
MF588884). In the present phylogenetic analysis, MFLUCC
17-1453 clusters with Pseudoophiosphaerella huishuiensis strain HS-13 with hight bootstrap support (100% ML
and 1.00 BYPP, Fig. 52). We therefore, identify MFLUCC
17-1453 as Pseudoophiosphaerella huishuiensis based on
phylogenetic analyses with morphological comparison
(Table 13), and the isolates are introduced here as a new host
record from Chromolaena odorata collected in Thailand.
Fungal Diversity
Pseudostaurosphaeria Mapook & K.D. Hyde, gen. nov.
Index Fungorum number: IF557354, Facesoffungi number: FoF 07813
Etymology: Refer r ing to its similar ity with
Staurosphaeria.
Saprobic on dead stems. Sexual morph: Undetermined.
Asexual morph: Conidiomata pycnidial, solitary, immersed
or semi-immersed to superficial, uni-loculate, globose to
subglobose, pale brown to brown or dark brown. Ostiole
central, short papillate or apapillate. Pycnidial wall comprising 3–4 layers of thick-walled, hyaline or pale brown to light
brown cells of textura angularis. Conidiophores reduced
to conidiogenous cells. Conidiogenous cells holoblastic,
ampulliform to cylindrical or oblong, hyaline, unbranched.
Conidia globose or oblong to obovoid, 1–2-transverse septa,
later developing 1–2-vertical septa, muriform, smoothwalled, slightly constricted at the septa, yellowish brown to
brown or reddish brown, with single polar appendage from
apex, observed clearly when mounted in Indian ink.
Type species: Pseudostaurosphaeria chromolaenicola
Mapook & K.D. Hyde
Notes: Pseudostaurosphaeria is similar to the asexual
morph of Staurosphaeria, which belongs to the family
Coniothyriaceae (Wanasinghe et al. 2017), in having reddish
brown conidia with a transverse septum, and later developing vertical septa dividing the conidium into four compartments. A phylogenetic analyses based on combined dataset
of LSU, ITS, SSU and TEF1 sequence data show that Pseudostaurosphaeria species are phylogenetically distant from
Staurosphaeria and form a sister lineage with Yunnanensis
instead (Fig. 52). However, the asexual morph of Yunnanensis has been reported as having muriform conidia, ellipsoidal to obovoid with 3-transversely septate, and 1 vertical
septum. Therefore, we introduce Pseudostaurosphaeria as
a new genus to accommodate two new species Pseu. chromolaenae and Pseu. chromolaenicola.
Pseudostaurosphaeria chromolaenae Mapook & K.D.
Hyde, sp. nov.
Index Fungorum number: IF557355, Facesoffungi number: FoF 07814; Fig. 68
Etymology: Name reflects the host genus Chromolaena,
from which this species was isolated.
Holotype: MFLU 20-0358
Saprobic on dead stems of Chromolaena odorata. Sexual morph: Undetermined. Asexual morph: Conidiomata
110–145 µm high × 95–140 µm diam. ( x̄ = 130 × 118 µm,
n = 10), pycnidial, solitary, semi-immersed to superficial,
uni-loculate, subglobose, brown to dark brown, ostiolate,
apapillate. Pycnidial wall 9–15 µm wide, comprising 3–4
layers of thick-walled, pale brown to light brown cells of
textura angularis. Conidiophores reduced to conidiogenous cells. Conidiogenous cells holoblastic, ampulliform
Fig. 65 Culture characteristics on MEA: a Paraleptospora chromolaenae (MFLUCC 17-1481), b Paraleptospora chromolaenicola
(MFLUCC 17-1450)
to oblong, hyaline, unbranched. Conidia 7–10 × 5–8.5 μm
( x̄ = 8.5 × 7 µm, n = 50), globose or oblong to obovoid,
1–2-transverse septa, later developing 1–2-vertical septum,
muriform, smooth-walled, slightly constricted at the septa,
yellowish brown to reddish brown, with single polar appendage from apex, observed clearly when mounted in Indian
ink.
Culture characteristics: Conidia germinating on MEA
within 24 h. at room temperature and germ tubes produced from the apex. Colonies on MEA circular, mycelium
umbonate, entire to filamentous, cultures white at the surface, yellowish brown to brown from the centre of the colony
in reverse with creamy-white at the margin (Fig. 70a).
Pre-screening for antimicrobial activity: Pseudostaurosphaeria chromolaenae (MFLUCC 17-1490) showed antimicrobial activity against B. subtilis with a 10 mm inhibition zone and against M. plumbeus with a 25 mm inhibition
zone, observable as partial inhibition, when compared to the
positive control (27 mm and 19 mm, respectively), but no
inhibition of E. coli.
Material examined: THAILAND, Chiang Rai Province,
Doi Pui, on dead stems of Chromolaena odorata, 2 April
2017, A. Mapook (DP89, MFLU 20-0358, holotype); extype culture MFLUCC 17-1490.
GenBank numbers: LSU: MN994570, ITS: MN994593,
SSU: MN994616, TEF1: MN998174
Notes: In a BLASTn search of NCBI GenBank, the closest
match with the ITS sequence of Pseudostaurosphaeria chromolaenae (MFLUCC 17-1490, ex-holotype) with 89.07%
similarity was Neosetophoma italica (strain FC-3846,
LC206635). The closest match with the LSU sequence with
98.90% similarity was Phaeosphaeria sinensis (strain C454,
MK348022). The closest match with the SSU sequence
with 99.45% similarity was Phaeosphaeria avenaria f. sp.
triticae (strain ATCC 26370, EU189210), while the closest
match with the TEF1 sequence with 97.00% similarity was
Paraphoma radicina (strain UTHSC: DI16-209, LT797075).
In the present phylogenetic analysis, Pseu. chromolaenae
clusters with Pseu. chromolaenicola with high bootstrap
13
Fungal Diversity
Fig. 66 Pseudoophiosphaerella
huishuiensis (new host record)
a, b Appearance of superficial
ascomata on substrate. c Section
through ascoma. d Ostiole. e
Peridium. f Pseudoparaphyses.
g, h Asci. i–m Ascospores.
Scale bars: a, b = 500 µm, c,
g–m = 50 µm, d, e = 20 µm,
f = 5 µm
support (100% ML and 1.00 BYPP, Fig. 52). However,
Pseu. chromolaenae differs from Pseu. chromolaenicola
in having larger conidiomata (110–145 × 95–140 μm vs.
105–120 × 85–120 μm) and larger conidia (7–10 × 5–8.5 μm
vs. 6.5–9 × 5–6.5 μm), with globose or oblong to obovoid,
1–2-transverse septa, later developing 1–2-vertical septa,
while Pseu. chromolaenicola has oblong to obovoid conidia
with 1-transverse septum and later developing 1-vertical septum (Table 14). A comparison of the ITS (+5.8S)
gene region of Pseu. chromolaenae and Pseu. chromolaenicola reveals 59 base pair differences (10.9%) across 539
13
nucleotides. Therefore, Pseu. chromolaenae is described
as a new species based on phylogeny and morphological
comparison.
Pseudostaurosphaeria chromolaenicola Mapook & K.D.
Hyde, sp. nov.
Index Fungorum number: IF557356, Facesoffungi number: FoF 07815; Fig. 69
Etymology: Name reflects the host genus Chromolaena,
on which this species was growing.
Holotype: MFLU 20-0360
Fungal Diversity
Fig. 67 Culture characteristics on MEA: a, b Pseudoophiosphaerella huishuiensis (MFLUCC 17-1453)
Saprobic on dead stems of Chromolaena odorata. Sexual morph: Undetermined. Asexual morph: Conidiomata
105–120 µm high × 85–120 µm diam. ( x̄ = 113 × 100 µm,
n = 5), pycnidial, solitary, immersed to semi-immersed, uniloculate, globose to subglobose, pale brown to light brown.
Ostiole central, short papillate. Pycnidial wall 5–15 µm
wide, comprising 3–4 layers of thick-walled, hyaline or
pale brown to light brown cells of textura angularis. Conidiophores reduced to conidiogenous cells. Conidiogenous
cells holoblastic, ampulliform to cylindrical, hyaline and
unbranched. Conidia 6.5–9 × 5–6.5 μm ( x̄ = 7.6 × 6 µm,
n = 45), oblong to obovoid, 1-transverse septum, later developing 1-vertical septum, muriform, smooth-walled, slightly
constricted at the septum, yellowish brown to brown with
single polar appendage from apex, observed clearly when
mounted in Indian ink.
Culture characteristics: Conidia germinating on MEA
within 48 h. at room temperature and germ tubes produced
from both ends. Colonies on MEA circular, mycelium
umbonate, entire, cultures white at the surface, olivaceousbrown to dark brown from the centre of the colony in reverse
with creamy-white at the margin (Fig. 70b).
Pre-screening for antimicrobial activity: Pseudostaurosphaeria chromolaenicola (MFLUCC 17-1497) showed
antimicrobial activity against M. plumbeus with an 18 mm
inhibition zone, observable as partial inhibition, when compared to the positive control (18 mm), but no inhibition of
E. coli and B. subtilis.
Material examined: THAILAND, Chiang Rai Province,
Doi Pui, on dead stems of Chromolaena odorata, 2 April
2017, A. Mapook (DP90, MFLU 20-0359); living culture
MFLUCC 17-1491; (DP96, MFLU 20-0360, holotype); extype culture MFLUCC 17-1497.
GenBank numbers: LSU: MN994571, MN994572, ITS:
MN994594, MN994595, SSU: MN994617, MN994618,
TEF1: MN998175, MN998176
Notes: In a BLASTn search of NCBI GenBank, the closest match with the ITS sequence of Pseudostaurosphaeria
chromolaenicola (MFLUCC 17-1497, ex-holotype) with
93.91% similarity was Neosetophoma sp. (strain CFE-31,
MK614774). The closest match with the LSU sequence with
98.46% similarity was Phaeosphaeria sinensis (strain C454,
MK348022). The closest match with the SSU sequence with
99.82% similarity was Phaeosphaeria avenaria f. sp. triticae
(strain ATCC 26370, EU189210), while the closest match
with the TEF1 sequence with 96.81% similarity was Yunnanensis phragmitis (strain MFLUCC 17-0365, MF683625).
In the present phylogenetic analysis, Pseu. chromolaenicola
clusters with Pseu. chromolaenae (MFLUCC 17-1490)
with high bootstrap support (100% ML and 1.00 BYPP,
Fig. 52). However, Pseu. chromolaenicola differs from
Pseu. chromolaenae in having oblong to obovoid conidia
with 1-transverse septum and later developing 1-vertical
septum, while Pseu. chromolaenae has larger conidiomata
(110–145 × 95–140 μm vs. 105–120 × 85–120 μm) and
larger conidia (7–10 × 5–8.5 μm vs. 6.5–9 × 5–6.5 μm), with
globose or oblong to obovoid, 1–2-transverse septa, later
developing 1–2-vertical septa (Table 14). A comparison of
the ITS (+5.8S) gene region of Pseu. chromolaenicola and
Pseu. chromolaenae reveals 59 base pair differences (10.9%)
across 539 nucleotides. Therefore, Pseu. chromolaenicola is
described as a new species based on phylogeny and morphological comparison.
Yunnanensis Karun., Phook. & K.D. Hyde
Yunnanensis was introduced by Karunarathna et al.
(2017) with Yu. phragmitis as the type species, based on
morphology and phylogeny. The species was collected on
stems of Phragmites australis (Poaceae) from China. We
Table 13 Morphological features of Pseudoophiosphaerella species discussed in this study
Species
Asocomata (µm)
P. huishuiensis
(MFLUCC
17-1463)
P. huishuiensis
(MFLUCC
19-0164)
Peridium (µm)
Asci (µm)
Ascospores (µm)
Septation of ascospores
References
(15–)25–50
275–350
high × 160–190
diam.
140–175 × 7.5–10
130–170 × 2–2.5
15–16-septate
This study
–
143–151.5 × 8–9.5
130.5–151.5 × 2.5–3.5
Multi-septate
Zhang et al. (2019)
25–34
13
Fungal Diversity
Fig. 68 Pseudostaurosphaeria
chromolaenae (holotype) a,
b Appearance of conidiomata
on substrate. c Section through
of conidioma. d Peridium.
e–g Conidiogenous cells and
developing conidia. h–m
Micro- and macro-conidia with
appendages n Conidia with
appendages in Indian ink. Scale
bars: a = 500 µm, b = 200 µm,
c = 50 µm, d, h = 10 µm, e–g,
i–n = 5 µm
introduce a new Yunnanensis species from C. odorata, based
on morphology and molecular data, together with descriptions and illustrations (Fig. 71). A phylogenetic tree based on
combined LSU, ITS, SSU, TEF1 and RPB2 sequence data
is presented in Fig. 52.
Yunnanensis chromolaenae Mapook & K.D. Hyde, sp. nov.
Index Fungorum number: IF557293, Facesoffungi number: FoF 07816; Fig. 71
Etymology: Name reflects the host genus Chromolaena,
from which this species was isolated.
Holotype: MFLU 20-0384
Saprobic on dead stems of Chromolaena odorata. Sexual
morph: Ascomata 130–170(–210) µm high × 140–225 µm
diam. ( x̄ = 167 × 175 µm, n = 10), semi-immersed to superficial, solitary or scattered, gregarious, appearing as dark,
longitudinal spots, coriaceous, globose, brown to dark
brown. Ostiole central. Peridium 10–15.5 µm wide, 2–3 layers, comprising pale brown to brown cells of textura angularis. Hamathecium comprising 1.5–3 µm wide, cylindrical
to broadly filiform, septate, branching pseudoparaphyses.
Asci 95–125(–155) × 19–28 µm ( x̄ = 112 × 23.5 µm, n = 10),
8-spored, bitunicate, fissitunicate, cylindric-clavate to obovoid, straight or slightly curved, apically rounded, pedicellate. Ascospores 20–28 × 10–12.5 µm ( x̄ = 24 × 11 µm,
n = 20), overlapping, 1–2-seriate, initially hyaline to pale
yellow, 1-septate when immature, becoming goldenbrown at maturity, ellipsoid to broadly fusiform, muriform,
4–6-transversely septate, with 1 vertical septum, constricted
at the central septum, straight or slightly curved, surrounded
by hyaline gelatinous sheath observed clearly when mounted
in Indian ink. Asexual morph: Undetermined.
Table 14 Synopsis of Pseudostaurosphaeria species with similar morphological features discussed in this study
Species
Conidiomata (μm)
Pycnidial wall
(μm)
Conidia (μm)
Septate of conidia
P. chromolaenae
(MFLUCC
17-1490)
P. chromolaenicola (MFLUCC
17-1497)
110–145 high × 95–140 diam.
9–15
7–10 × 5–8.5
105–120 high × 85–120 diam.
5–15
6.5–9 × 5–6.5
This study
Globose or oblong to obovoid,
1–2-transverse septa, later developing 1–2-vertical septa
This study
Oblong to obovoid, 1-transverse
septum, later developing 1-vertical septum
13
Reference
Fungal Diversity
Fig. 69 Pseudostaurosphaeria
chromolaenicola (holotype) a,
b Appearance of conidiomata
on substrate. c Section through
of conidioma. d Ostiole. e
Peridium. f-i Conidiogenous
cells and developing conidia.
j-l Conidia with appendages
m Conidia with appendages
in Indian ink. Scale bars:
a = 500 µm, b = 200 µm, c,
d = 20 µm, e-m = 10 µm
Culture characteristics: Ascospores germinating on
MEA within 24 h. at room temperature and germ tubes produced from several cells. Colonies on MEA circular, mycelium crateriform, entire, cultures white at the surface, dark
brown to olivaceous from the centre of the colony in reverse
with creamy-white at the margin (Fig. 72).
Pre-screening for antimicrobial activity: Yunnanensis
chromolaenae (MFLUCC 17-1487) showed antimicrobial
activity against E. coli with an 8 mm inhibition zone and
against M. plumbeus with a 35 mm inhibition zone, observable as partial inhibition, when compared to the positive
control (9 mm and 19 mm, respectively), but no inhibition
of B. subtilis.
Material examined: THAILAND, Chiang Rai Province,
Doi Pui, on dead stems of Chromolaena odorata, 2 April
2017, A. Mapook (DP85, MFLU 20-0384, holotype); extype culture MFLUCC 17-1487; (DP84, MFLU 20-0383);
living culture MFLUCC 17-1486.
GenBank numbers: LSU: MN994573, MN994574, ITS:
MN994596, MN994597, SSU: MN994619, MN994620,
TEF1: MN998177, MN998178
Notes: In a BLASTn search of NCBI GenBank, the closest match with the ITS and TEF1 sequences of Yunnanensis chromolaenae (MFLUCC 17-1487, ex-holotype) with
93.24% (KY768867) and 97.84% (MF683624) similarity,
respectively, was Yunnanensis phragmitis strain MFLUCC
17-0315. The closest match with the LSU sequence with
98.79% similarity was Tintelnotia destructans (strain CBS
127737, NG_058274), while the closest match with the SSU
sequence with 99.82% similarity was Phaeosphaeria sp.
(strain S93-48, EU189216). A phylogenetic analyses based
on combined dataset of LSU, ITS, SSU and TEF1 sequence
data show that Yu. chromolaenae forms a distinct lineage
with Yu. phragmitis with high bootstrap support (99% ML
and 1.00 BYPP, Fig. 52). Yu. chromolaenae is similar to Yu.
phragmitis in having muriform spores, fusiform to ellipsoidal, surrounded by hyaline gelatinous sheath. However, Yu.
13
Fungal Diversity
Fig. 70 Culture characteristics on MEA: a Pseudostaurosphaeria
chromolaenae (MFLUCC 17-1490). b Pseudostaurosphaeria chromolaenicola (MFLUCC 17-1497)
Fig. 71 Yunnanensis chromolaenae (holotype) a, b Appearance of ascomata on substrate.
c Section through ascoma. d
Peridium. e Pseudoparaphyses.
f–i Asci. j–n Ascospores. o
Ascospores with gelatinous
sheath in Indian ink. Scale
bars: a = 200 µm, b = 100 µm,
c, f–i = 50 µm, d = 20 µm,
j–o = 10 µm, e = 5 µm
13
chromolaenae differs from Yu. phragmitis in having larger
semi-immersed to superficial ascomata, appearing as longitudinal spots on host surface and larger ascospores with
4–6-transversely septate, and 1 vertical septum, while Yu.
phragmitis has smaller immersed to erumpent ascomata and
smaller ascospores with 3-transverse septa, and 1 vertical
septum in the second and third cells (Table 15). A comparison of the ITS (+5.8S) gene region of Yu. chromolaenae
and Yu. phragmitis reveals 32 base pair differences (5.97%)
across 536 nucleotides. Therefore, we introduce a new species based on morphology and phylogeny.
Fungal Diversity
Pyrenochaetopsidaceae Valenzuela-Lopez et al.
Pyrenochaetopsidaceae was introduced by ValenzuelaLopez et al. (2018) to accommodate Pyrenochaetopsis as
the type genus, plus two new genera, Neopyrenochaetopsis
and Xenopyrenochaetopsis based on phylogenetic analyses.
The family was described as an asexual morph with seven
new species provided with genomic sequences. The genus
has been isolated from plants, cyst of plant parasitic nematodes, as well as water and soil samples (Boerema et al.
2004; Crous et al. 2014; Papizadeh et al. 2017). ValenzuelaLopez et al. (2018) isolated most of the new species from
human respiratory tract (bronchial wash and sputum sample), human superficial tissue (foot skin, sinusitis tissue, toe
nail, ear lesion and dermatitis sample) and human deep tissue (blood sample).
Pyrenochaetopsis Gruyter, Aveskamp & Verkley, in de
Gruyter, et al., Mycologia 102(5): 1076 (2010)
Pyrenochaetopsis was introduced by de Gruyter et al.
(2010) with P. leptospora as the type species. The genus
is ecologically diverse and had been accepted as a member of Cucurbitariaceae (Gruyter et al. 2010, 2013; Doilom
et al. 2013; Hyde et al. 2013; Papizadeh et al. 2017; Wijayawardene et al. 2012, 2014a, Wijayawardene et al. 2017a, b,
2018). However, Valenzuela-Lopez et al. (2018) showed that
the genus formed a distinct monophyletic clade outside the
Cucurbitariaceae. Presently, 13 epithets are listed in Index
Fungorum (2020). In this study, Pyrenochaetopsis chromolaenae is introduced based on morphology and molecular
data, together with a description and illustrations (Fig. 74).
A phylogenetic tree based on combined LSU, ITS and RPB2
sequence data is presented in Fig. 73.
Pyrenochaetopsis chromolaenae Mapook & K.D. Hyde,
sp. nov.
Index Fungorum number: IF557359, Facesoffungi number: FoF 07817; Fig. 74
Etymology: Name reflects the host genus Chromolaena,
from which this species was isolated.
Holotype: MFLU 20-0362
Saprobic on dead stems of Chromolaena odorata. Sexual morph: Ascomata (60–)100–110 µm high × 75–125 µm
diam. ( x̄ = 95 × 110 µm, n = 5), superficial, appearing as
small dark spots, coriaceous, solitary or scattered, globose, brown to dark brown. Ostiole short papillate, with
numerous external reddish brown setae. Peridium 5–10 µm
wide, comprising 2-layers of thin-walled, pale brown to
brown cells of textura angularis. Hamathecium comprising
1.5–2.5 µm wide, cylindrical to filiform, septate, branching
pseudoparaphyses. Asci 45–90 × 9–12 µm ( x̄ = 65 × 10.5 µm,
n = 15), 8-spored, bitunicate, fissitunicate, cylindric-clavate,
straight or slightly curved, with a short, bulbous pedicel apically rounded. Ascospores 14–24 × 4–6 µm ( x̄ = 19 × 5 µm,
n = 15), overlapping 2–3-seriate, hyaline to pale brown to
slightly yellowish brown, cylindrical to broadly fusiform,
with slightly obtuse ends, widest at the center and tapering towards rounded ends, 3–4-septate, straight or slightly
curved, second cell from apex is slightly inflated, guttulate,
constricted at the septa, without terminal appendages. Asexual morph: Undetermined.
Culture characteristics: Ascospores germinating on
MEA within 24 h. at room temperature and germ tubes produced from both ends. Colonies on MEA circular, mycelium
slightly raised, entire, white aerial hyphae at the surface,
spreading and dark brown to olivaceous in reverse (Fig. 75).
Pre-screening for antimicrobial activity: Pyrenochaetopsis chromolaenae (MFLUCC 17-1440) showed antimicrobial activity against E. coli with a 9 mm inhibition zone
when compared to the positive control (9 mm), but no inhibition of B. subtilis and M. plumbeus.
Material examined: THAILAND, Chiang Rai Province,
Doi Pui, on dead stems of Chromolaena odorata, 5 August
2015, A. Mapook (DP24, MFLU 20-0362, holotype); extype culture MFLUCC 17-1440.
GenBank numbers: LSU: MT214472, ITS: MT214378,
SSU: MT214424, TEF1: MT235790, RPB2: MT235827
Notes: In a BLASTn search of NCBI GenBank, the
closest match with the ITS sequence of Pyrenochaetopsis chromolaenae (MFLUCC 17-1440, ex-holotype) with
99.81% similarity was Py. microspora (strain CBS 102876,
NR_160059). The closest match with the LSU sequence
with 99.88% similarity was Py. leptospora (strain CBS
131.69, MH871006), while the closest match with the RPB2
sequence with 95.35% similarity was Pyrenochaetopsis
sp. (strain UTHSC:DI16-275, LT593072). In the present
phylogenetic analysis, Py. chromolaenae forms a separate
clade and clusters with Py. setosissima (CBS 119739) and
Py. americana (UTHSC:DI16-225), which were found as
asexual morphs from culture, with bootstrap support (67%
ML and 0.98 BYPP, Fig. 73). However, Py. chromolaenae
was found as the sexual morph in nature and we could not
obtain its asexual morph in culture. A comparison of the
RPB2 gene region of Py. chromolaenae and Py. americana
reveals 48 base pair differences (5.4%) across 884 nucleotides. Therefore, Py. chromolaenae is described as a new
species based on phylogeny.
Roussoellaceae Liu et al.
Roussoellaceae was introduced to accommodate three
genera Neoroussoella, Roussoella and Roussoellopsis which
have trabeculate pseudoparaphyses (sensu Liew et al. 2000)
and brown bicelled ascospores (Liu et al. 2014). Previously,
the family has been treated as a synonym of Thyridariaceae
by Jaklitsch and Voglmayr (2016). However, the latest
updated accounts based on multigene phylogenetic analysis
show that Roussoellaceae is well-resolved in Pleosporales
13
Fungal Diversity
Fig. 72 Culture characteristic on MEA: Yunnanensis chromolaenae (MFLUCC 17-1487)
(Dai et al. 2017; Tibpromma et al. 2017; Hyde et al. 2018a,
b; Jayasiri et al. 2019; Jiang et al. 2019; Phookamsak et al.
2019). Wanasinghe et al. (2018) introduced two roussoellalike taxa, Pararoussoella and Pseudoneoconiothyrium in
Thyridariaceae. However, Phookamsak et al. (2019) showed
that these two genera clustered with Roussoella in Roussoellaceae, based on increased taxon sampling in the phylogenetic analysis. This finding was supported by Jayasiri et al.
(2019) and Jiang et al. (2019). Divergence time estimates for
this family are crown age of 62 Mya (34–91) in the Cenozoic
Era (Paleogene period) and stem age of 77 Mya (44–110) in
the late Cretaceous (Liu et al. 2017).
Pseudoroussoella Mapook & K.D. Hyde, gen. nov.
Index Fungorum number: IF557351, Facesoffungi number: FoF 07818
Etymology: Referring to its similarity with Roussoella.
Saprobic on dead stems or petiole. Sexual morph:
Ascomata immersed to erumpent through the host surface,
solitary, appearing as dark spots, coriaceous, globose to
subglobose, dark brown to black, Ostiolar neck protruding. Peridium several layers, inner layers comprising of
hyaline to light brown cells of textura epidermoidea to
textura angularis, outer layers comprising of brown to dark
brown cells of textura intricata. Hamathecium composed of
cylindrical to filiform, septate, trabeculate pseudoparaphyses. Asci 8-spored, bitunicate, fissitunicate, cylindrical to
clavate, straight or slightly curved, apically rounded, short
pedicellate with small ocular chamber. Ascospores uniseriate, initially hyaline to pale brown, septate when immature,
becoming yellowish brown to dark brown at maturity, oval
to ellipsoid, 1-septate, constricted at the septum, straight
or slightly curved, slightly widest at the upper cell and
tapering towards obtuse ends, with a reticulate spore wall
ornamentation, surrounded by hyaline gelatinous sheath
observed clearly when mounted in Indian ink. Asexual
morph: Conidiomata pycnidial, solitary, superficial, uniloculate, globose to obpyriform, yellowish brown to brown.
Ostiole central, papillate. Pycnidial wall comprising of 2–3
layers, hyaline or pale brown to light brown cells of textura
angularis. Conidiophores reduced to conidiogenous cells.
Conidiogenous cells annellidic, ampulliform to oblong, hyaline and unbranched. Conidia oblong to oval, pale brown
to light brown when immature, becoming yellowish brown
to reddish brown when mature, aseptate, thick-walled with
guttulate.
Type species: Pseudoroussoella elaeicola (Konta & K.D.
Hyde) Mapook & K.D. Hyde
Notes: A phylogenetic analysis based on combined dataset of LSU, ITS, TEF1, RPB2 and SSU sequence data show
that two Pseudoroussoella species form a separate clade
and group with Pseudoneoconiothyrium rosae and Roussoella euonymi which were found as asexual morphs, with
bootstrap support (83% ML, Fig. 76). Asexual morph of
Pseudoroussoella species differ from Pseudoneoconiothyrium rosae and R. euonymi in smaller oblong to oval conidia,
while Pseudoneoconiothyrium rosae has globose to irregularly ellipsoid conidia (Table 16). Therefore, we introduce
Pseudoroussoella as a new genus with two new species
based on morphology and phylogeny.
Pseudoroussoella chromolaenae Mapook & K.D. Hyde,
sp. nov.
Index Fungorum number: IF557353, Facesoffungi number: FoF 07332; Fig. 77
Etymology: Name reflects the host genus Chromolaena,
from which this species was isolated.
Table 15 Synopsis of Yunnanensis species with similar morphological features discussed in this study
Species
Asocomata (µm)
Peridium (µm) Asci (µm)
130–170(–210) high × 140–225 10–15.5
Yu. chrodiam.
molaenae
(MFLUCC
17-1487)
100–130 high, 115–170 diam. 7–12
Yu. phragmitis
(MFLUCC
17-0315)
13
Ascospores (µm) Septate of ascospores References
95–125(–155) × 19–28 20–28 × 10–12.5
4–6-transversely sep- This study
tate, with 1-vertical
septum
61–113 × 23–31
3-transversely septate, and 1-vertical
septum
11–13 × 4–7
Karunarathna
et al.
(2017)
Fungal Diversity
Table 16 Synopsis of asexual morph of Pseudoroussoella species with similar morphological features discussed in this study
Species
Conidiomata (µm)
Conidiogenous cells
(µm)
Roussoella euonymi (CBS 143426)
150–300 diam.
5–12 × 5–7
Pseudoneoconiothyrium rosae (MFLUCC 190–240 high × 120–170 diam.
4–7 × 3–7
15-0052)
Pseudoroussoella chromolaenae
130–175(–230) high × 160–230 diam. –
(MFLUCC 17-1492)
Holotype: MFLU 20-0356
Saprobic on dead stems of Chromolaena odorata.
Sexual morph: Undetermined. Asexual morph: Conidiomata 130–175(–230) µm high × 160–230 µm diam.
( x̄ = 165 × 195 µm, n = 5), pycnidial, solitary, superficial,
uni-loculate, globose to obpyriform, yellowish brown to
brown. Ostiole central, papillate. Pycnidial wall 10–20 µm
wide, comprising 2–3 layers of hyaline or pale brown to light
brown cells of textura angularis. Conidiophores reduced
to conidiogenous cells. Conidiogenous cells annellidic,
ampulliform to oblong, hyaline and unbranched. Conidia
5.5–7 × 3.5–5 μm ( x̄ = 6.5 × 4.5 µm, n = 50), oblong to oval,
pale brown to light brown when immature, becoming yellowish brown to reddish brown when mature, aseptate, thickwalled with guttule.
Culture characteristics: Conidia germinating on MEA
within 24 h at room temperature and germ tubes produced
from both ends. Colonies on MEA circular, mycelium
slightly raised, velvety with moderately fluffy, filamentous, cultures white with grayish-brown from the centre
and near the margin of the colony on surface, olivaceous in
reverse from the centre of the colony with white at margin
(Fig. 79b).
Pre-screening for antimicrobial activity: Pseudoroussoella chromolaenae (MFLUCC 17-1492) showed no inhibition of E. coli, B. subtilis and M. plumbeus.
Material examined: THAILAND, Chiang Rai Province,
Doi Pui, on dead stems of Chromolaena odorata, 2 February 2017, A. Mapook (DP91, MFLU 20-0356, holotype);
ex-type culture MFLUCC 17-1492.
GenBank numbers: LSU: MT214439, ITS: MT214345,
SSU: MT214393, TEF1: MT235769
Notes: In a BLASTn search of NCBI GenBank, the closest match with the ITS sequence of Pseudoroussoella chromolaenae (MFLUCC 17-1492, ex-holotype) with 99.10%
similarity was Roussoella sp. (strain MFLUCC 17-2059,
MH744730). The closest match with the LSU sequence
with 98.59% similarity was Roussoella hysterioides (strain
CBS 125434, MH875155). The closest match with the SSU
sequence with 91.77% similarity was Roussoella intermedia
(strain KT 2303, AB524483), while the closest match with
the TEF1 sequence with 96.49% similarity was Arthopyrenia
Conidia (µm)
References
(6–)7(–8) × (4–)5–6 Crous et al. (2018b)
6–8 × 4–7
Wanasinghe et al. (2018)
5.5–7 × 3.5–5
This study
sp. (strain UTHSC: DI16-334, LT797127). In the present
phylogenetic analysis, P. chromolaenae clusters with P.
elaeicola with high bootstrap support (100% ML and 1.00
BYPP, Fig. 76). A comparison of the TEF1 gene region of
P. chromolaenae and P. elaeicola reveals 10 base pair differences (1.1%) across 899 nucleotides and the ITS (+5.8S)
gene region reveals five base pair differences (1.1%) across
443 nucleotides. However, we could not compare the morphological characteristics of those species; P. chromolaenae
is found as asexual morph in nature, while P. elaeicola is
found as sexual morph in nature and we could not obtain
its asexual morph in culture. Therefore, P. chromolaenae
is described here as a new species based on phylogeny and
culture characteristic comparison.
Pseudoroussoella elaeicola (Konta & K.D. Hyde) Mapook
& K.D. Hyde, comb. nov.
Index Fungorum number: IF557352, Facesoffungi number: FoF 07819; Fig. 78
≡ Roussoella elaeicola Konta & K.D. Hyde, in Phookamsak et al., Fungal Divers (2019)
Holotype: THAILAND, Chiang Rai Province, on dead
petiole of Elaeis guineensis (Arecaceae), 25 November
2014, S. Konta, HR02d (MFLU 15-0022), ex-type culture,
MFLUCC 15-0276
Saprobic on dead stems of Chromolaena odorata. Sexual
morph: Ascomata 225–475 µm high × 240–400 µm diam.
( x̄ = 365 × 325 µm, n = 5), immersed, solitary, appearing as
dark spots, coriaceous, subglobose, dark brown to black.
Ostiolar neck protruding. Peridium (20–)30–50 µm wide,
several layers, inner layers comprising hyaline to light brown
cells of textura epidermoidea, outer layers comprising brown
to dark brown cells of textura intricata. Hamathecium comprising 1–2 µm wide, cylindrical to filiform, septate, trabeculate pseudoparaphyses. Asci (70–)95–135 × 6–8.5 µm
( x̄ = 108 × 7 µm, n = 10), 8-spored, bitunicate, cylindrical
to clavate, straight or slightly curved, apically rounded,
short pedicellate with small ocular chamber. Ascospores
10–14 × 4.5–6 µm ( x̄ = 12.5 × 5.5 µm, n = 25), uniseriate, initially hyaline to pale brown, septate when immature, becoming yellowish brown at maturity, oval to ellipsoid, 1-septate,
constricted at the septum, straight or slightly curved, slightly
13
Fungal Diversity
Fig. 73 Phylogram generated
from maximum likelihood
analysis based on combined
dataset of LSU, ITS and RPB2
sequence data. Fourty-nine
strains are included in the
combined sequence analysis,
which comprise 2556 characters with gaps. Tree topology
of the ML analysis was similar
to the BYPP. The best scoring
RAxML tree with a final likelihood value of − 18819.011208
is presented. The matrix had
977 distinct alignment patterns, with 12.78% of undetermined characters or gaps.
Estimated base frequencies
were as follows: A = 0.247993,
C = 0.237502, G = 0.272601,
T = 0.241905; substitution rates:
AC = 1.771498, AG = 5.599945,
AT = 1.899441, CG = 1.222461,
CT = 10.018389,
GT = 1.000000; gamma
distribution shape parameter
α = 0.168302. Bootstrap support values for ML equal to or
greater than 60% and BYPP
equal to or greater than 0.90 are
given above or below the nodes.
Newly generated sequences
are in dark red bold and type
species are in bold. Pleospora
herbarum (CBS 191.86) and P.
herbarum (IT956) are used as
outgroup taxa
widest at the upper cell and tapering towards obtuse ends,
with a reticulate spore wall ornamentation, surrounded by
hyaline gelatinous sheath observed clearly when mounted in
Indian ink. Asexual morph: Undetermined.
Culture characteristics: Ascospores germinating on
MEA within 48 h. at room temperature and germ tubes produced from both ends. Colonies on MEA circular, mycelium
slightly raised, velvety with moderately fluffy, filamentous,
cultures white, grayish-brown from the centre of the colony
with white at margin on surface, brown to pale olivaceousbrown in reverse from the centre of the colony, white to
creamy-white at margin (Fig. 79a).
13
Pre-screening for antimicrobial activity: Pseudoroussoella elaeicola (MFLUCC 17-1483) showed antimicrobial
activity against E. coli with a 10 mm inhibition zone when
compared to the positive control (9 mm), but no inhibition
of B. subtilis and M. plumbeus.
Known hosts and distribution: On dead petiole of
Elaeis guineensis (Arecaceae) in Thailand (Phookamsak
et al. 2019).
Material examined: THAILAND, Lampang Province,
Ngao, on dead stems of Chromolaena odorata, 21 September 2016, A. Mapook (LP5, MFLU 20-0357); living culture
MFLUCC 17-1483 (new host record).
Fungal Diversity
Fig. 74 Pyrenochaetopsis
chromolaenae (holotype)
a, b Appearance of superficial ascomata on substrate.
c Section through ascoma. d
Ostiole with brown setae. e
Peridium. f Pseudoparaphyses. g–j Immature and mature
asci. k–p Ascospores. Scale
bars: a = 500 µm, b = 100 µm,
c = 50 µm, g–j = 20 µm, d, e,
k–p = 10 µm, f = 5 µm
GenBank numbers: LSU: MT214442, ITS: MT214348,
SSU: MT214396, TEF1: MT235772, RPB2: MT235808
Notes: A phylogenetic analyses show that the strain
MFLUCC 17-1483 grouped with Pseudoroussoella elaeicola (= Roussoella elaeicola) (Fig. 76). In a BLASTn search
of NCBI GenBank, the closest match of the ITS sequence of
our strains with 99.80% similarity was Roussoella sp. (strain
MFLUCC 17-2059, MH744730). The closest match with
the LSU sequence with 98.47% similarity was Arthopyrenia
salicis (strain MUT < ITA > :4879, KP671722). The closest match with the SSU sequence with 93.95% similarity
was Roussoella intermedia (strain KT 2303, AB524483).
The closest match with the TEF1 sequence with 96.06%
similarity was Arthopyrenia sp. (strain UTHSC: DI16-334,
LT797127), while the closest match with the RPB2 sequence
with 90.92% similarity was R. euonymi (strain CBS 143426,
MH108007). We therefore, identify our isolates as P. elaeicola based on phylogenetic analyses with morphological
comparison (Table 17). In this study, we isolated P. elaeicola
from Chromolaena odorata collected in Thailand, and the
isolate is introduced here as a new host record.
13
Fungal Diversity
Fig. 75 Culture characteristic on MEA: Pyrenochaetopsis chromolaenae (MFLUCC 17-1440)
Setoarthopyrenia Mapook & K.D. Hyde, gen. nov.
Index Fungorum number: IF557361, Facesoffungi number: FoF 07820
Etymology: Referring to the Ostiolar neck with numerous external setae and its ascospores similarity with
Arthopyrenia.
Saprobic on dead stems. Sexual morph: Ascomata superficial, solitary, appearing as small spots, coriaceous, globose, brown to dark brown. Ostiolar neck protruding, with
numerous external brown to dark brown setae. Peridium
2–3 layers, comprising brown to dark brown cells of textura
angularis. Hamathecium composed of filiform to broadly filiform, septate, branching pseudoparaphyses. Asci 8-spored,
bitunicate, cylindric-clavate, straight or slightly curved, apically rounded, short pedicellate. Ascospores bi-seriate, hyaline, ellipsoid to obovoid, 1-septate, guttulate, constricted
at the septum, straight, without gelatinous sheath. Asexual
morph: Undetermined.
Type species: Setoarthopyrenia chromolaenae Mapook
& K.D. Hyde
Notes: A phylogenetic analysis based on combined dataset of LSU, ITS, TEF1, RPB2 and SSU sequence data show
that Setoarthopyrenia chromolaenae forms a separate clade
with Roussoella clade, with bootstrap support (83% ML and
0.99 BYPP, Fig. 76). Setoarthopyrenia differs from Roussoella in having a protruding ostiole neck, with numerous external brown to dark brown setae, bi-seriate ascospores that are
hyaline, ellipsoid to obovoid, 1-septate without gelatinous
sheath, while Roussoella species have uniseriate ascospores
that are brown, fusiform-ellipsoidal, ornamented and surrounded by a wide mucilaginous sheath (Liu et al. 2014).
Therefore, we introduce Setoarthopyrenia as a new genus
with a new species, S. chromolaenae based on morphology
and phylogeny.
13
Setoarthopyrenia chromolaenae Mapook & K.D. Hyde, sp.
nov.
Index Fungorum number: IF557362, Facesoffungi number: FoF 07821; Fig. 80
Etymology: Name reflects the host genus Chromolaena,
from which this species was isolated.
Holotype: MFLU 20-0368
Saprobic on dead stems of Chromolaena odorata. Sexual morph: Ascomata 65–105 µm high × 80–95 µm diam.
(x̅ = 85 × 88 µm, n = 5), superficial, solitary, appearing as
small spots, coriaceous, globose, brown to dark brown.
Ostiolar neck protruding, with numerous external brown to
dark brown setae. Peridium (4.5–)5–10 µm wide, 2–3 layers, comprising brown to dark brown cells of textura angularis. Hamathecium comprising 0.5–1.5 µm wide, filiform
to broadly filiform, septate, branching pseudoparaphyses.
Asci 40–60 × 12–15 µm (x̅ = 55 × 13.5 µm, n = 10), 8-spored,
bitunicate, cylindric-clavate, straight or slightly curved, apically rounded, short pedicellate. Ascospores 14–17 × 6–8 µm
(x̅ = 15 × 7 µm, n = 25), bi-seriate, hyaline, ellipsoid to obovoid, 1-septate, guttulate, constricted at the septum, straight,
without gelatinous sheath. Asexual morph: Undetermined.
Culture characteristics: Ascospores germinating on
MEA within 24 h. at room temperature and germ tubes produced from both ends. Colonies on MEA circular, mycelium
slightly raised, entire, cultures white, olivaceous-brown at
margin on surface, dark brown to olivaceous-brown from the
centre of the colony in reverse with creamy-white appearing
as concentric ring pattern and dark brown to olivaceousbrown at margin (Fig. 81).
Pre-screening for antimicrobial activity: Setoarthopyrenia chromolaenae (MFLUCC 17-1444) showed antimicrobial activity against B. subtilis with a 16 mm inhibition zone,
when compared to the positive control (26 mm), but no inhibition of M. plumbeus and E. coli.
Material examined: THAILAND, Chiang Rai Province,
Doi Pui, on dead stems of Chromolaena odorata, 5 August
2015, A. Mapook (DP37, MFLU 20-0368, holotype); extype culture MFLUCC 17-1444.
GenBank numbers: LSU: MT214438, ITS: MT214344,
SSU: MT214392, TEF1: MT235768, RPB2: MT235805
Notes: In a BLASTn search of NCBI GenBank, the closest match of the ITS, LSU, TEF1 and RPB2 sequences of
Setoarthopyrenia chromolaenae (MFLUCC 17-1444, exholotype) with 99.83% (LT796905), 100% (LN907505),
99.78% (LT797145) and 96.35% (LT797065) similarity,
respectively, was Arthopyrenia sp. strain UTHSC: DI16-362,
while the closest match of the SSU sequence with 99.81%
similarity was Roussoella intermedia (strain CBS 170.96,
KF443390). In the present phylogenetic analysis, S. chromolaenae (MFLUCC 17-1444) is closely related to Arthopyrenia sp. UTHSC: DI16-362 (Fig. 76). However, we could not
compare the morphological characteristics of both strains as
Fungal Diversity
Fig. 76 Phylogram generated
from maximum likelihood analysis based on combined dataset
of LSU, ITS, TEF1, RPB2 and
SSU sequence data. Eightyfour strains are included in the
combined sequence analysis,
which comprise 4416 characters with gaps. Tree topology
of the ML analysis was similar
to the BYPP. The best scoring
RAxML tree with a final likelihood value of − 35074.673502
is presented. The matrix had
1690 distinct alignment patterns, with 39.45% of undetermined characters or gaps.
Estimated base frequencies
were as follows: A = 0.246392,
C = 0.254324, G = 0.268438,
T = 0.230846; substitution rates:
AC = 1.657341, AG = 4.989387,
AT = 2.205689, CG = 1.285998,
CT = 10.615734,
GT = 1.000000; gamma
distribution shape parameter
α = 0.486933. Bootstrap support values for ML equal to or
greater than 60% and BYPP
equal to or greater than 0.90 are
given above or below the nodes.
Newly generated sequences
and new combination are in
dark red bold and type species
are in bold. Occultibambusa
bambusae (MFLUCC 11-0394)
and O. bambusae (MFLUCC
13-0855) are used as outgroup
taxa
the morphology of Arthopyrenia sp. (UTHSC: DI16-362)
has not been reported. Therefore, we introduce a new species
based on morphology and phylogeny.
Index Fungorum number: IF557367, Facesoffungi number: FoF 07822
Etymology: Xeno = ξένος in Greek, distinct;
Roussoella = roussoella-like
Xenoroussoella Mapook & K.D. Hyde, gen. nov.
13
Fungal Diversity
Table 17 Synopsis of sexual morph species with similar morphological features discussed in this study
Species
Ascomata (µm)
Peridium (µm) Asci (µm)
Ascospores (µm)
References
Pseudoroussoella elaeicola
(= Roussoella elaeicola,
MFLUCC 15-0276)
P. elaeicola (MFLUCC
17-1483, LP5)
Xenoroussoella triseptata
(MFLUCC 17-1438, DP22)
315–410 high, 325–350 diam.
25–70
70–140 × 6–9
10–15 × 3–6, 1-septate
225–475 high × 240–400
diam.
(155–)170–220 high × 185–
235 diam.
(20–)30–50
(70–)95–135 × 6–8.5 10–14 × 4.5–6, 1-septate
Phookamsak et al.
(2019)
This study
10–15
55–70 × 10.5–15
13.5–17 × 5–7.5, 3-septate This study
Fig. 77 Pseudoroussoella
chromolaenae (holotype) a,
b Appearance of conidiomata
on substrate. c Section through
of conidioma. d Ostiole e
Peridium. f–g Conidiogenous
cells and developing conidia.
h–i Conidia. Scale bars: a,
b = 200 µm, c = 50 µm, d,
e = 20 µm, h, i = 10 µm, f,
g = 5 µm
Saprobic on dead stems. Sexual morph: Ascomata
immersed, solitary or scattered, appearing as orange spots,
coriaceous, globose to subglobose, brown to dark brown.
Ostiolar neck protruding. Peridium 2–4 layers, comprising
hyaline to brown cells of textura angularis. Hamathecium
composed of oblong to cylindrical, septate, branching pseudoparaphyses. Asci 8-spored, bitunicate, cylindric-clavate to
clavate, straight or slightly curved, apically rounded, short
pedicellate. Ascospores bi-seriate, initially hyaline, 1-septate when immature, becoming pale brown to dark brown
at maturity, ellipsoid to obovoid, 3-septate, with irregular
longitudinal striations, constricted at the central septum,
straight or slightly curved, without gelatinous sheath. Asexual morph: Undetermined.
13
Type species: Xenoroussoella triseptata Mapook & K.D.
Hyde
Notes: A phylogenetic analysis based on combined dataset of LSU, ITS, TEF1, RPB2 and SSU sequence data show
that Xenoroussoella triseptata forms a separate branch
and groups with the clade comprising Pseudoroussoella
species, Pseudoneoconiothyrium rosae and Roussoella
euonymi, with high bootstrap support (99% ML and 1.00
BYPP, Fig. 76). Xenoroussoella differs from Pseudoroussoella in having smaller ascomata and asci, thin peridium,
larger bi-seriate ascospores (Table 16) that are ellipsoid to
obovoid, 3-septate with irregular longitudinal striations,
without gelatinous sheath, while Pseudoroussoella has uniseriate ascospores that are oval to ellipsoid, 1-septate, with
a reticulate spore wall ornamentation, surrounded by hyaline
Fungal Diversity
Fig. 78 Pseudoroussoella
elaeicola (new host record)
a, b Appearance of immersed
ascomata on substrate. c Section
through ascoma. d Ostiole. e
Peridium. f Pseudoparaphyses.
g–j Asci. k–o Ascospores. p
Ascospores with gelatinous
sheath in Indian ink. Scale
bars: a = 500 µm, b = 200 µm,
c = 100 µm, g–j = 50 µm,
d, e = 20 µm, f = 10 µm,
k–p = 5 µm
gelatinous sheath. Therefore, we introduce Xenoroussoella
as a new genus with a new species, Xe. triseptata based on
morphology and phylogeny.
Xenoroussoella triseptata Mapook & K.D. Hyde, sp. nov.
Index Fungorum number: IF557368, Facesoffungi number: FoF 07823; Fig. 82
Etymology: The specific epithet “triseptata” is based on
the 3-septate ascospores
Holotype: MFLU 20-0382
Saprobic on dead stems of Chromolaena odorata. Sexual
morph: Ascomata (155–)170–220 µm high × 185–235 µm
diam. ( x̄ = 185 × 210 µm, n = 5), immersed, solitary or
scattered, appearing as orange spots, coriaceous, globose to subglobose, brown to dark brown. Ostiolar
neck protruding. Peridium 10–15 µm wide, comprising
2–4 layers of hyaline to brown cells of textura angularis. Hamathecium comprising 1.9–3 µm wide, oblong to
cylindrical, septate, branching pseudoparaphyses. Asci
55–70 × 10.5–15 µm ( x̄ = 65 × 12 µm, n = 20), 8-spored,
13
Fungal Diversity
phylogeny and their distinct morphology in having 3-septate
ascospores, with irregular longitudinal striations.
Fig. 79 Culture characteristics on MEA: a Pseudoroussoella
elaeicola (MFLUCC 17-1483). b Pseudoroussoella chromolaenae
(MFLUCC 17-1492)
bitunicate, cylindric-clavate to clavate, straight or slightly
curved, apically rounded, short pedicellate. Ascospores
13.5–17 × 5–7.5 µm ( x̄ = 15 × 6.5 µm, n = 30), bi-seriate,
initially hyaline, 1-septate when immature, becoming pale
brown to dark brown at maturity, ellipsoid to obovoid, 3-septate, with irregular longitudinal striations, constricted at the
central septum, straight or slightly curved, without gelatinous sheath. Asexual morph: Undetermined.
Culture characteristics: Ascospores germinating on
MEA within 24 h. at room temperature and germ tubes produced from both ends. Colonies on MEA circular, mycelium
slightly raised, undulate, cultures white, pale grayish brown
from the centre of the colony with olivaceous to olivaceousbrown at margin on surface, creamy-white in reverse, olivaceous-brown to brown from the centre of the colony with
olivaceous to olivaceous-brown at margin (Fig. 83).
Pre-screening for antimicrobial activity: Xenoroussoella triseptata (MFLUCC 17-1438) showed no inhibition of
E. coli, B. subtilis and M. plumbeus.
Material examined: THAILAND, Chiang Rai Province,
Doi Pui, on dead stems of Chromolaena odorata, 5 August
2015, A. Mapook (DP22, MFLU 20-0382, holotype); extype culture MFLUCC 17-1438.
GenBank numbers: LSU: MT214437, ITS: MT214343,
SSU: MT214391, TEF1: MT235767, RPB2: MT235804
Notes: In a BLASTn search of NCBI GenBank, the closest match of the ITS, LSU, TEF1 and RPB2 sequences of
Xenoroussoella triseptata (MFLUCC 17-1438, ex-holotype)
with 99.65% (LT796887), 99.88% (LN907477), 99.38%
(LT797127) and 97.80% (LT797047) similarity, respectively,
was Arthopyrenia sp. strain UTHSC: DI16-334, while the
closest match of the SSU sequence with 93.39% similarity
was Roussoella intermedia (strain KT 2303, AB524483). In
the present phylogenetic analysis, Xe. triseptata (MFLUCC
17-1438) is closely related to Arthopyrenia sp. UTHSC:
DI16-334 (Fig. 76). However, we could not compare the
morphological characteristics of both strains as the morphology of Arthopyrenia sp. (UTHSC: DI16-334) has not been
reported. Therefore, we introduce a new species based on
13
Thyridariaceae Q. Tian & K.D. Hyde
Family Thyridariaceae was introduced by Hyde et al.
(2013) to accommodate Thyridaria as its type genus. Jaklitsch and Voglmayr (2016) synonymized Roussoellaceae
under Thyridariaceae with introduction of a new genus
Parathyridaria in the family. However, Tibpromma et al.
(2017) reinstated Roussoellaceae based on morphology with
phylogenetic analysis and treated it as a well-resolved family
in Pleosporales. Wanasinghe et al. (2018) introduced three
new genera (Cycasicola, Pseudoneoconiothyrium, Pararoussoella), of which Pseudoneoconiothyrium and Pararoussoella are presently placed in Roussoellaceae. Devadatha et al.
(2018) also introduced a new genus Thyridariella with two
new species in Thyridariaceae. Phookamsak et al. (2019)
introduced a new genus Liua based on morphological distinction and phylogenetic support. Divergence time estimates for this family are crown age of 15 Mya (3–38) in the
Cenozoic period and stem age of 95 Mya (60–138) in the
Cretaceous (Liu et al. 2017).
Chromolaenomyces Mapook & K.D. Hyde, gen. nov.
Index Fungorum number: IF557333, Facesoffungi number: FoF 07824
Etymology: Named after the host genus Chromolaena,
combined with “myces” for fungi.
Saprobic on dead stems. Sexual morph: Ascomata
immersed, coriaceous, solitary or scattered, globose to
subglobose, light brown to brown. Ostiolar neck protruding. Peridium 2–3 layers, pale brown to dark brown cells
of textura angularis. Hamathecium composed of cylindrical to filiform, septate, branching pseudoparaphyses. Asci
8-spored, bitunicate, fissitunicate, cylindrical, straight or
slightly curved, apically rounded, with a short pedicel.
Ascospores uni-seriate, irregular arrangement, hyaline, oval
to broadly fusiform, uni-septate, constricted at the septum,
smooth, with a narrow sheath; sheath drawn out to form
polar appendages from both ends of the ascospores, straight
or slightly curved. Asexual morph: Undetermined.
Type species: Chromolaenomyces appendiculatus
Mapook & K.D. Hyde
Notes: Phylogenetic analysis based on combined dataset
of LSU, ITS, TEF1, RPB2 and SSU sequence data show
that Chromolaenomyces appendiculatus forms a separate
branch and groups with Thyridariella mangrovei, with low
bootstrap support (Fig. 84). However, Chromolaenomyces
differs from Thyridariella in having cylindrical asci with a
short pedicel and ascospores that are uni-seriate, arranged
irregularly, oval to broadly fusiform, uni-septate with a narrow sheath, drawn out to form polar appendages from both
ends of the ascospores, while Thyridariella has clavate asci
Fungal Diversity
Fig. 80 Setoarthopyrenia
chromolaenae (holotype) a,
b Appearance of superficial
ascomata on substrate. c Section
through ascoma. d dark brown
setae. e Peridium. f Pseudoparaphyses. g–j Immature and
mature asci. k–p Ascospores.
Scale bars: a = 500 µm,
b = 100 µm, c, g–j = 20 µm, d,
k–p = 10 µm, e, f = 5 µm
with a moderately long pedicel and ascospores that are uniseriate to bi-seriate, fusiform to ellipsoidal, muriform, and
surrounded by hyaline gelatinous sheath (Devadatha et al.
2018). Therefore, we introduce Chromolaenomyces as a new
genus with a new species, C. appendiculatus based on morphology and phylogeny.
Chromolaenomyces appendiculatus Mapook & K.D. Hyde,
sp. nov.
Index Fungorum number: IF557334, Facesoffungi number: FoF 07825; Fig. 85
Etymology: Referring to ascospores with sheath drawn
out to form polar appendages
Holotype: MFLU 20-0307
Saprobic on dead stems of Chromolaena odorata. Sexual
morph: Ascomata 175–185 µm high × 140–165 µm diam.
( x̄ = 180 × 155 µm, n = 5), immersed, coriaceous, solitary
or scattered, globose to subglobose, light brown to brown.
Ostiolar neck protruding. Peridium 7–12(–15) µm wide, 2–3
layers, pale brown to dark brown cells of textura angularis. Hamathecium comprising 0.5–1.5 µm wide, cylindrical to filiform, septate, branching pseudoparaphyses. Asci
(75–)85–105 × 10–14 µm ( x̄ = 95 × 12 µm, n = 10), 8-spored,
bitunicate, fissitunicate, cylindrical, straight or slightly
curved, apically rounded, with a short pedicel. Ascospores
15–20.5 × 6–7.5 µm ( x̄ = 17.5 × 7 µm, n = 20), uni-seriate,
irregular arrangement, hyaline, oval to broadly fusiform,
uni-septate, constricted at the septum, smooth, with a
13
Fungal Diversity
similarity was Cycasicola leucaenae (strain MFLUCC
17-0914, MK434900). In the present phylogenetic analysis,
C. appendiculatus (MFLUCC 17-1455) is closely related to
Thyridariella mangrovei (Fig. 84). However, morphological
characteristics of both strains are distinct.
Fig. 81 Culture characteristic on MEA: Setoarthopyrenia chromolaenae (MFLUCC 17-1444)
narrow sheath; sheath drawn out to form polar appendages
(4.5–)6.5–12(–16) µm long × 2–3.5 µm wide ( x̄ = 9 × 2.5 µm,
n = 30), from both ends of the ascospores, straight or slightly
curved. Asexual morph: Undetermined.
Culture characteristics: Ascospores germinating on
MEA within 48 h. at room temperature, germ tubes produced from both ends of the ascospores. Colonies on MEA
circular to irregular, mycelium slightly raised, entire, cultures olivaceous-grey at the surface with white margin and
olivaceous-brown at the center in reverse, white to yellowwhite at the margin (Fig. 86a).
Pre-screening for antimicrobial activity: Chromolaenomyces appendiculatus (MFLUCC 17-1455) showed antimicrobial activity against B. subtilis with a 20 mm inhibition
zone and against M. plumbeus with a 13 mm inhibition zone,
observable as partial inhibition, when compared to the positive control (26 mm and 17 mm, respectively), but no inhibition of E. coli.
Material examined: THAILAND, Lampang Province,
Chaehom, on dead stems of Chromolaena odorata, 24 September 2016, A. Mapook (JH1, MFLU 20-0307, holotype);
ex-type culture MFLUCC 17-1455.
GenBank numbers: LSU: MT214440, ITS: MT214346,
SSU: MT214394, TEF1: MT235770, RPB2: MT235806
Notes: In a BLASTn search of NCBI GenBank, the
closest match of the ITS sequences of Chromolaenomyces
appendiculatus (MFLUCC 17-1455, ex-holotype) with
83.59% similarity was Roussoella sp. (strain IFM 64548,
LC317761). The closest match of the LSU sequence with
99.20% similarity was Cycasicola goaensis (strain MFLU
17-0581, NG_059057). The closest match of the SSU
sequence with 92.89% similarity was Parathyridaria percutanea (strain CBS 868.95, NG_062999). The closest match
of the TEF1 sequence with 96.86% similarity was Cycasicola goaensis (strain MFLUCC 17-0754, MG829198),
while the closest match of the RPB2 sequence with 87.21%
13
Pseudothyridariella Mapook & K.D. Hyde, gen. nov.
Index Fungorum number: IF557357, Facesoffungi number: FoF 07826
Etymology: Referring to its similarity with Thyridariella.
Saprobic on dead stems or decaying wood. Sexual
morph: Ascomata immersed to erumpent, solitary or scattered, appearing as dark spots, coriaceous, globose or subglobose to obpyriform, yellowish brown to brown. Ostiolar neck protruding. Peridium several layers, light brown
or brown to yellowish brown cells of textura angularis.
Hamathecium composed of cylindrical to filiform, septate,
branching pseudoparaphyses. Asci 8-spored, bitunicate, fissitunicate, cylindric-clavate, straight or slightly curved, pedicellate at base, with an ocular chamber at apically rounded
apex. Ascospores overlapping, 1–2 seriate, hyaline to greyish
brown, 1-septate when immature, hyaline to brown or olivaceous-brown to dark brown at maturity, ellipsoid to broadly
fusiform, muriform, with (3–)5–8-transverse septa, and 1
vertical septum, slightly constricted at the central septum,
straight or slightly curved, surrounded by hyaline gelatinous
sheath, with central septum observed clearly when mounted
in Indian ink. Asexual morph: Undetermined.
Type species: Pseudothyridariella chromolaenae
Mapook & K.D. Hyde
Notes: A phylogenetic analysis based on combined dataset of LSU, ITS, TEF1, RPB2 and SSU sequence data shows
that Pseudothyridariella chromolaenae forms a sister clade
with a clade comprising Liua, Cycasicola, Thyridariella
and Chromolaenomyces in Thyridariaceae, with bootstrap
support (100% ML and 1.00 BYPP, Fig. 84). Pseudothyridariella is similar to Thyridariella in having ellipsoid
to broadly fusiform muriform ascospores surrounded by a
hyaline gelatinous sheath, but differs in having ascospores
with constriction at the central septum, while ascospores
of Thyridariella are without constriction at the central
septum (Devadatha et al. 2018). Therefore, we introduce
Pseudothyridariella as a new genus with a new species,
Pseudothyridariella chromolaenae based on morphology
and phylogeny.
Pseudothyridariella chromolaenae Mapook & K.D. Hyde,
sp. nov.
Index Fungorum number: IF557358, Facesoffungi number: FoF 07827; Fig. 87
Etymology: Name reflects the host genus Chromolaena,
from which this species was isolated.
Holotype: MFLU 20-0361
Fungal Diversity
Fig. 82 Xenoroussoella triseptata (holotype) a, b Appearance of immersed ascomata on
substrate. c Section through
ascoma. d Ostiole. e Peridium.
f Pseudoparaphyses. g–j
Asci. k–p Ascospores. Scale
bars: a = 200 µm, b = 100 µm,
c = 50 µm, d, g–j = 20 µm,
e = 10 µm, f, k–p = 5 µm
Saprobic on dead stems of Chromolaena odorata.
Sexual morph:Ascomata 170–345 µm high × 95–220 µm
diam. ( x̄ = 225 × 150 µm, n = 5), immersed, solitary or
scattered, appearing as dark spots, coriaceous, obpyriform, yellowish brown to brown. Ostiolar neck protruding. Peridium (10–)15–20 µm wide, several layers, light
brown to yellowish brown cells of textura angularis.
Hamathecium comprising 1.5–2.5 µm wide, cylindrical to filiform, septate, branching pseudoparaphyses. Asci
80–105(–135) × (14–)17–22 µm ( x̄ = 100 × 18.5 µm, n = 10),
8-spored, bitunicate, fissitunicate, cylindric-clavate, straight
or slightly curved, apically rounded, pedicellate. Ascospores
23–28 × 9–12.5 µm ( x̄ = 25 × 10.5 µm, n = 25), overlapping,
1–2 seriate, initially hyaline to greyish brown, 1-septate
when immature, becoming brown or olivaceous-brown to
dark brown at maturity, ellipsoid to broadly fusiform, muriform, with 5–8-transverse septa, and 1 vertical septum,
slightly constricted at the central septum, straight or slightly
curved, surrounded by hyaline gelatinous sheath observed
clearly when mounted in Indian ink. Asexual morph:
Undetermined.
13
Fungal Diversity
closely related to Thyridariella mahakashae NFCCl 4215,
which was treated as Pseudothyridariella mahakashae
in this study (Fig. 84). However, P. chromolaenae differs from P. mahakashae in having smaller ascomata
(170–345 × 95–220 µm vs. 250–550 × 195–500 µm) and asci
(80–105(–135) × (14–)17–22 µm vs. 70–220 × 10–20 µm)
with brown or olivaceous-brown to dark brown ascospores
at maturity with 5–8-transverse septa, while P. mahakashae
has hyaline ascospores with 3–6-transverse septa (Table 18).
A comparison of the ITS (+ 5.8S) gene region of P. chromolaenae and P. mahakashaereveals 82 base pair differences
(17%) across 482 nucleotides. Therefore, P. chromolaenae
is described as a new species based on phylogeny and morphological comparison.
Fig. 83 Culture characteristic on MEA: Xenoroussoella triseptata
(MFLUCC 17-1438)
Culture characteristics: Ascospores germinating on
MEA within 48 h. at room temperature and germ tubes produced from several cells. Colonies on MEA circular, mycelium slightly raised, velvety with moderately fluffy, entire,
cultures white at the surface, creamy-white in reverse with
white at the margin (Fig. 88).
Pre-screening for antimicrobial activity: Pseudothyridariella chromolaenae (MFLUCC 17-1472) showed
antimicrobial activity against M. plumbeus with a 23 mm
inhibition zone, observable as partial inhibition, when compared to the positive control (17 mm), but no inhibition of
B. subtilis and E. coli.
Material examined: THAILAND, Nan Province, Doi
Phu Kha, on dead stems of Chromolaena odorata, 23 September 2016, A. Mapook (DPK1, MFLU 20-0361, holotype); ex-type living culture MFLUCC 17-1472.
GenBank numbers: LSU: MT214441, ITS: MT214347,
SSU: MT214395, TEF1: MT235771, RPB2: MT235807
Notes: In a BLASTn search of NCBI GenBank, the
closest match of the ITS sequences of Pseudothyridariella chromolaenae (MFLUCC 17-1472, ex-holotype) with
93.82% similarity was Thyridariella mahakashae (strain
NFCCl 4215, MG020436). The closest match of the LSU
sequence with 97.59% similarity was Roussoellaceae
sp. (strain MUT 4884, KP671726). The closest match of
SSU sequence with 95.95% similarity was Parathyridaria
percutanea (strain CBS 868.95, NG_062999). The closest match of the TEF1 sequence with 96.58% similarity
was Parathyridaria percutanea (strain UTHSC: DI16300, LT797113), while the closest match of the RPB2
sequence with 87.57% similarity was T. mangrovei (strain
NFCCI 4213, MG020445). In the present phylogenetic
analysis, P. chromolaenae (MFLUCC 17-1472) is found
13
Pseudothyridariella mahakashae (Devadatha, V.V. Sarma,
D.N. Wanas., K.D. Hyde & E.B.G. Jones) Mapook & K.D.
Hyde, comb. nov.
Index Fungorum number: IF557369, Facesoffungi number: FoF 07828
≡ Thyridariella mahakoshae Devadatha, V.V. Sarma,
D.N. Wanas., K.D. Hyde & E.B.G. Jones, in Devadatha,
Sarma, Jeewon, Wanasinghe, Hyde & Jones, Mycol. Progr.:
https://doi.org/10.1007/s11557-018-1387-4, 8 (2018)
Holotype: India, Tamil Nadu, Tiruvarur, Muthupet mangroves (10.4° N 79.5° E), on decaying wood of Avicennia marina (Acanthaceae), 29 March 2017, B. Devadatha
(AMH-9933, holotype), ex-type culture, NFCCI-4215.
Morphological description: See Devadatha et al. (2018)
(Fig. 2).
Notes: In our multigene phylogenetic study, Thyridariella
mahakoshae (NFCCl 4215) was in a clade separate from
the type species, Thyridariella mangrovei (NFCCI-4213)
and clustered with Pseudothyridariella chromolaenae
(MFLUCC 17-1472) with high bootstrap support (100%
ML and 1.00 BYPP, Fig. 84). The species shares similar
morphological characters with P. chromolaenae in having
ellipsoid to broadly fusiform muriform ascospore, surrounded by a hyaline gelatinous sheath with constriction
at the central septum, while T. mangrovei has ascospores
surrounded by a wide gelatinous sheath in circle but not
constricted at the central septum (Devadatha et al. 2018).
Therefore, we transfer Thyridariella mahakoshae as Pseudothyridariella mahakashae based on phylogeny and morphological comparison.
Torulaceae Corda
Torulaceae was introduced by Corda (Sturm 1829) with
Torula as the type genus. Crous et al. (2015a) provided
molecular data for Torula species and also accepted the
genus Dendryphion in the family. Su et al. (2016) introduced
a new genus Neotorula from freshwater habitats and Li et al.
(2016) introduced a new genus Sporidesmioides within the
Fungal Diversity
Fig. 84 Phylogram generated
from maximum likelihood analysis based on combined dataset
of LSU, ITS, TEF1, RPB2 and
SSU sequence data. Eightyfour strains are included in the
combined sequence analysis,
which comprise 4416 characters with gaps. Tree topology
of the ML analysis was similar
to the BYPP. The best scoring
RAxML tree with a final likelihood value of − 35074.673502
is presented. The matrix had
1690 distinct alignment patterns, with 39.45% of undetermined characters or gaps.
Estimated base frequencies
were as follows: A = 0.246392,
C = 0.254324, G = 0.268438,
T = 0.230846; substitution rates:
AC = 1.657341, AG = 4.989387,
AT = 2.205689, CG = 1.285998,
CT = 10.615734,
GT = 1.000000; gamma
distribution shape parameter
α = 0.486933. Bootstrap support values for ML equal to or
greater than 60% and BYPP
equal to or greater than 0.90 are
given above or below the nodes.
Newly generated sequences
and new combination are in
dark red bold and type species
are in bold. Occultibambusa
bambusae (MFLUCC 11-0394)
and O. bambusae (MFLUCC
13-0855) are used as outgroup
taxa
family based on both morphology and phylogenetic analyses. Su et al. (2018) introduced a new genus Rostriconidium
from freshwater habitats, and provided a morphological
comparison and mutigene analyses. Currently, five genera, Torula, Dendryphion, Neotorula, Sporidesmioides and
Rostriconidium are accepted in Torulaceae within the order
Pleosporales (Li et al. 2016a, b, 2017; Su et al. 2016, 2018;
Tibpromma et al. 2018). Divergence time estimates for this
family are crown age of 15 Mya (4–34) in the Cenozoic Era
13
Fungal Diversity
Fig. 85 Chromolaenomyces
appendiculatus (holotype)
a, b Appearance of immersed
ascomata on substrate. c Section
through ascoma. d Ostiole. e
Peridium. f Pseudoparaphyses.
g–j Asci. k–p Ascospores.
Scale bars: a, b = 100 µm, c,
g–j = 50 µm, d, e = 20 µm,
k–p = 10 µm, f = 5 µm
(Neogene period) and stem age of 140 Mya (95–188) in the
early Cretaceous (Liu et al. 2017).
Torula Pers.
Torula was introduced by Persoon (1794) with T. herbarum as the type species. Crous et al. (2015a) introduced
three new species T. fici, T. hollandica and T. masonii based
13
morphology and phylogeny. Subsequently, Li et al. (2017)
introduced four new species (T. chromolaenae, T. mackenziei, T. pluriseptata, T. chiangmaiensis) with two new host
records for T. masonii and T. fici, based on DNA sequence
analyses from nucleotides and protein genes; including two
species T. fici and T. chromolaenae which were collected
from Chromolaena odorata in Thailand. Hyde et al. (2019a)
Fungal Diversity
Fig. 86 Culture characteristic on MEA: Chromolaenomyces
appendiculatus (MFLUCC 17-1455)
introduced two new species T. polyseptata and T. breviconidiophora based on morphology and phylogeny. We present
two reference specimens and a new species from C. odorata, together with descriptions and illustrations (Figs. 90,
91, 92). A phylogenetic tree based on combined LSU, SSU,
TEF1, RPB2 and ITS sequence data is presented in Fig. 89.
Torula chromolaenae Li, Phook., Mapook & K.D. Hyde,
Mycol. Progr. 16(4): 454 (2017)
Facesoffungi number: FoF 02713; Fig. 90
Saprobic on dead stems of Chromolaena odorata. Sexual
morph: Undetermined. Asexual morph: Hyphomycetous.
Colonies effuse on host, black, powdery. Mycelium partly
immersed to superficial on the host surface, comprising
septate, smooth, and pale brown to light brown hyphae.
Conidiophores 4–4.7 × 4–5 µm ( x̄ = 4.5 × 4.6 µm, n = 5),
macronematous, mononematous, solitary, arising from
hypha, erect, pale brown to brown, smooth, thick-walled,
subglobose, comprising 1-cell or reduced to conidiogenous
cell. Conidiogenous cells 3–6 × 3.5–6 µm ( x̄ = 5 × 5 µm,
n = 15), monoblastic, doliiform to subglobose, brown to dark
brown, paler at apex, smooth to minutely verruculose, thickwalled. Conidia 7–18.5 × 4–8 µm ( x̄ = 13.5 × 6 µm, n = 30),
phragmosporous, catenate, acrogenous, branched, oblong,
elongated, 1–3-septate, slightly constricted with dark bands
at the septa, dark brown to black, smooth to minutely verrucose, rounded at both ends and mostly with a dark coronate
conidiogenous cell at the apex.
Culture characteristics: Conidia germinating on MEA
within 24 h. at room temperature and germ tubes produced
from both ends. Colonies on MEA circular, mycelium
slightly raised, entire, cultures greyish brown at the surface
with white at the margin and pale pinkish brown from the
centre of the colony in reverse with creamy-white at the margin (Fig. 93a).
Pre-screening for antimicrobial activity: Torula chromolaenae (MFLUCC 17-1514) showed antimicrobial activity against B. subtilis, E. coli and M. plumbeus (9 mm, 8 mm
and 18 mm inhibition zone, respectively), observable as
partial inhibition, when compared to the positive control
(26 mm, 9 mm, and 17 mm, respectively).
Known hosts and distribution: On dead stems of Chromolaena odorata in Thailand (Li et al. 2017; this study);
on dead leaf of Pandanus tectorius (Pandanaceae) in China
(Tibpromma et al. 2018).
Material examined: THAILAND, Chiang Rai Province,
Doi Pui, on dead stems of Chromolaena odorata, 2 February 2017, A. Mapook (DP105, MFLU 20-0370); living
culture MFLUCC 17-1514 (reference specimen); Chiang
Mai Province, Fah hom pok, on dead stems of Chromolaena
odorata, 27 September 2016, A. Mapook (FHP8, MFLU
20-0371); living culture MFLUCC 17-1504.
GenBank numbers: LSU: MT214477, MT214478, ITS:
MT214383, MT214384, SSU: MT214428, MT214429,
TEF1: MT235792, MT235793, RPB2: MT235831,
MT235832
Notes: A phylogenetic analysis shows that two strains
MFLUCC 17-1514 and MFLUCC 17-1504 are grouped
with Torula chromolaenae (Fig. 89). In a BLASTn search
of NCBI GenBank, the closest match of the ITS, LSU, TEF1
and RPB2 sequences with 99.79% (MH275087), 100%
(KY197860), 99.53% (KY197880) and 99.88% (KY197873)
similarity, respectively, were identical to T. chromolaenae,
while the closest match of the SSU sequence with 99.81%
similarity was T. hollandica (strain CBS 220.69, KF443389).
We therefore, identify our isolates as T. chromolaenae based
on phylogenetic analyses and morphological comparison
(Table 19). Moreover, the type and our strains are collected
from the same host, Chromolaena odorata and from the
same country as that of Li et al. (2017). Thus, we designate
our strain as a reference specimen (sensu Ariyawansa et al.
2014a) for Torula chromolaenae.
Torula fici Crous, IMA Fungus 6 (1): 192 (2015)
Facesoffungi number: FoF 02712; Fig. 91
Saprobic on dead stems of Chromolaena odorata. Sexual
morph: Undetermined. Asexual morph: Hyphomycetous.
Colonies effuse on host, dark brown to black, powdery.
Mycelium partly immersed to superficial on the host surface,
comprising septate, smooth, and pale brown to light brown
hyphae. Conidiophores 2.5–6.5 × 3.9–5.5 µm (x̅ = 5 × 4.5 µm,
n = 15), macronematous, mononematous, solitary, arising from hypha, erect, pale brown to brown, comprising
1–2 cells, smooth, thick-walled, subglobose to oblong.
Conidiogenous cells 4–7 × 5.5–7 µm ( x̄ = 6 × 6.25 µm,
n = 15), mono- to polyblastic, doliiform to subglobose,
dark brown to black, paler at apex, smooth, thick-walled.
Conidia 10–25 × 5.5–8 µm ( x̄ = 18.5 × 7 µm, n = 20),
13
Fungal Diversity
Fig. 87 Pseudothyridariella
chromolaenae (holotype) a,
b Appearance of ascomata on
substrate. c Section through
ascoma. d Ostiole. e Peridium.
f Pseudoparaphyses. g–j Asci.
k–o Ascospores. p Ascospores
with gelatinous sheath in Indian
ink. Scale bars: a–c = 100 µm,
g–j = 50 µm, d, e, p = 20 µm, f,
k–o = 10 µm
phragmosporous, catenate, acrogenous, branched, oblong,
elongated, 1–5-septate, slightly constricted with dark bands
at the septa, brown to dark brown, verrucose, rounded and
mostly dark brown at the apex, straight or slightly curved.
Culture characteristics: Conidia germinating on MEA
within 24 h. at room temperature and germ tubes produced
13
from both ends. Colonies on MEA circular, mycelium
slightly raised, entire, cultures white at the surface and olivaceous-brown from the centre of the colony in reverse with
creamy-white at the margin (Fig. 93b).
Fungal Diversity
Fig. 88 Culture characteristic on MEA: Pseudothyridariella chromolaenae (MFLUCC 17-1472)
Pre-screening for antimicrobial activity: Torula fici
(MFLUCC 17-1494) showed antimicrobial activity against
M. plumbeus with a 13 mm inhibition zone, observable as
partial inhibition, when compared to the positive control
(18 mm), but no inhibition of B. subtilis and E. coli.
Known hosts and distribution: On dead stems of Chromolaena odorata in Thailand (Li et al. 2017; this study);
on decaying cone of Magnolia grandiflora (Magnoliaceae)
in China; on decaying fruit pericarp of Garcinia sp. (Clusiaceae) in Thailand (Jayasiri et al. 2019); on dead leaf of
Pandanus sp. (Pandanaceae) in Thailand (Tibpromma et al.
2018); on submerged decaying wood in China (Su et al.
2018); on Ficus religiosa (Moraceae) in Cuba (Crous et al.
2015).
Material examined: THAILAND, Chiang Rai Province,
Doi Pui, on dead stems of Chromolaena odorata, 2 February
2017, A. Mapook (DP93, MFLU 20-0372); living culture
MFLUCC 17-1494 (reference specimen).
GenBank numbers: LSU: MT214479, ITS: MT214385,
SSU: MT214430, TEF1: MT235794, RPB2: MT235833
Notes: Phylogenetic analyses show that the strain
MFLUCC 17-1494 grouped within Torula fici clade
(Fig. 89). In a BLASTn search of NCBI GenBank, the closest match of the ITS, LSU, SSU, TEF1 and RPB2 sequences
with 99.80% (KF443409), 100% (MK348016), 99.81%
(KF443388), 98.98% (MK360090) and 98.43% (MK434871)
similarity, respectively was Torula fici. We, therefore, identify our isolates as T. fici based on phylogenetic analyses and
morphological comparison (Table 19). Moreover, T. fici has
been collected from the same host, Chromolaena odorata
and from the same country as in study of Li et al. (2017).
Thus, we designate our strain as a reference specimen (sensu
Ariyawansa et al. 2014a) for Torula fici.
Torula polyseptata C.G. Lin & K.D. Hyde, in Hyde et al.,
Fungal Divers. 96: 71 (2019)
Facesoffungi number: FoF 05070; Fig. 92
Saprobic on dead stems of Chromolaena odorata. Sexual
morph: Undetermined. Asexual morph: Hyphomycetous.
Colonies effuse on host, dark brown to black, powdery.
Mycelium partly immersed to superficial on the host surface,
comprising septate, smooth, branched and pale brown to light
brown hyphae. Conidiophores (4.5–)10–27 × 3.5–4.5 µm
(x̅ = 15 × 4 µm, n = 10), micro- to macronematous, mononematous, solitary, arising from hypha, erect, pale brown to
brown, comprising 1–3 cells, long, thick-walled, cylindrical to ampulliform. Conidiogenous cells 5.5–7.7 × 4.5–8 µm
( x̄ = 6 × 6.5 µm, n = 10), mono- to polyblastic, doliiform to
subglobose, brown to dark brown, thick-walled. Conidia
15–55 × 6.5–9 µm ( x̄ = 31 × 8 µm, n = 15), phragmosporous, catenate, acrogenous, unbranched, oblong, elongated,
1–10-septate, constricted with dark bands at the septa, pale
brown to yellowish brown, hyaline to light brown at apex
when immature, becoming dark brown when mature, verrucose, rounded at both ends?, straight or slightly curved.
Culture characteristics: Conidia germinating on MEA
within 24 h. at room temperature and germ tubes produced
from both ends. Colonies on MEA circular, mycelium
slightly flattened, curled, cultures greyish brown at the surface with white at the margin and pinkish brown from the
centre of the colony in reverse with creamy-white at the margin, MEA change to yellow (Fig. 93c).
Pre-screening for antimicrobial activity: Torula polyseptata (MFLUCC 17-1495) showed antimicrobial activity against M. plumbeus with a 12 mm inhibition zone,
Table 18 Synopsis of Pseudothyridariella species with similar morphological features discussed in this study
Species
Ascomata (μm)
Peridium (µm) Asci (µm)
Ascospores (µm)
References
P. chromolaenae
(MFLUCC 17-1472)
170–345 high × 95–220
diam.
(10–)15–20
80–105(–135) × (14–)17–
22
P. mahakashae (= Thyridariella mahakashae
NFCCl 4215)
250–550 high × 195–500
diam.
15–30
70–220 × 10–20
23–28 × 9–12.5, hyaline to This study
greyish brown, 1-septate
when immature, becoming brown or olivaceousbrown to dark brown at
maturity, 5–8-transverse
septa
17–27 × 5–12, hyaline, 3-6 Devadatha
transverse septa
et al.
(2018)
13
Fungal Diversity
Fig. 89 Phylogram generated
from maximum likelihood analysis based on combined dataset
of LSU, SSU, TEF1, RPB2 and
ITS sequence data. Fourtysix strains are included in the
combined sequence analysis,
which comprise 4655 characters with gaps. Tree topology
of the ML analysis was similar
to the BYPP. The best scoring
RAxML tree with a final likelihood value of − 16753.701279
is presented. The matrix had
1163 distinct alignment patterns, with 37.38% of undetermined characters or gaps.
Estimated base frequencies
were as follows: A = 0.245595,
C = 0.259181, G = 0.270709,
T = 0.224516; substitution rates:
AC = 1.706173, AG = 2.694498,
AT = 1.288686, CG = 1.108377,
CT = 7.475019, GT = 1.000000;
gamma distribution shape
parameter α = 0.172063. Bootstrap support values for ML
equal to or greater than 60% and
BYPP equal to or greater than
0.90 are given above the nodes.
Newly generated sequences
are in dark red bold and type
species are in bold. Occultibambusa bambusae (MFLUCC
11-0394) and O. bambusae
(MFLUCC 13-0855) are used as
outgroup taxa
observable as partial inhibition, when compared to the positive control (18 mm), but no inhibition of B. subtilis and E.
coli.
Known hosts and distribution: On submerged decaying
wood in Thailand (Hyde et al. 2019a).
Material examined: THAILAND, Chiang Rai Province,
Doi Pui, on dead stems of Chromolaena odorata, 2 February
2017, A. Mapook (DP94, MFLU 20-0373); living culture
MFLUCC 17-1495 (new host record).
GenBank numbers: LSU: MT214476, ITS: MT214382,
SSU: MT214427, TEF1: MT235791, RPB2: MT235830
13
Notes: Phylogenetic analyses show strain MFLUCC
17-1495 grouped with Torula polyseptata (Fig. 89). In a
BLASTn search of NCBI GenBank, the closest match of
the ITS sequences with 98.16% similarity was T. herbarum
(strain MEF021, KT315411). The closest match of the LSU
sequence with 100% similarity was T. chiangmaiensis (strain
KUMCC 16-0039, KY197856). The closest match of the SSU
and RPB2 sequences with 99.81% (KF443389) and 96.02%
(KF443393) similarity, respectively, was T. hollandica strain
CBS 220.69, while the closest match of the TEF1 sequence
with 98.38% similarity was T. pluriseptata (strain MFLUCC
14-0437, KY197875). We therefore, identify our isolate as T.
Fungal Diversity
Fig. 90 Torula chromolaenae (reference specimen) a,
b Appearance of colonies on
substrate. c, d Conidiophores
and Conidiogenous cells.
e–g Conidia in chains. h–m
Conidia. Scale bars: a = 500 µm,
b = 100 µm, e–m = 10 µm, c,
d = 5 µm
polyseptata based on morphological comparison and phylogenetic analyses. The isolate is introduced here as a new host
record from Chromolaena odorata collected in Thailand.
Dothideomycetes orders incertae sedis
Botryosphaeriales C.L. Schoch et al.
Botryosphaeriales was introduced by Schoch et al.
(2006). We follow the latest treatment and updated accounts
of Botryosphaeriales by Phillips et al. (2019) with recent
relevant literature for updated accounts of each family. Six
families, Aplosporellaceae, Botryosphaeriaceae, Melanopsaceae, Phyllostictaceae, Planistromellaceae, and Saccharataceae were accepted in the order, based on morphology
and molecular data (Phillips et al. 2019).
Aplosporellaceae Slippers et al.
Aplosporellaceae was introduced by Slippers et al. (2013)
to accommodate the genera Bagnisiella and Aplosporella,
with the latter as the type genus and suggested that the
divergence time estimates for the order Botryosphaeriales originated in the Cretaceous period around 103 Mya
(45–188). Sharma et al. (2017) introduced a new genus Alanomyces, the type of which was collected from soil in India.
Subsequently, Liu et al. (2017) reported crown age of the
order at 114 Mya (73–166) in the Cretaceous with estimates
for stem age at 181 Mya (106–279) in the Jurassic. Phillips
et al. (2019) reported crown age for Aplosporellaceae at 40
Mya in the Paleogene period with estimates for stem age at
94 Mya in the Cretaceous.
Aplosporella Speg.
Aplosporella was introduced by Spegazzini (1880), with
A. chlorostroma as the type species. The genus comprises
plant pathogens, endophytes and saprobes (Jami et al. 2014;
Fan et al. 2015; Ekanayaka et al. 2016; Dou et al. 2017;
Du et al. 2017; Zhu et al. 2018). Ekanayaka et al. (2016)
introduced a new species, A. thailandica from Thailand.
Subsequently, Dou et al. (2017) described a new species
A. macropycnidia from Cerasus yedoensis in China and Du
et al. (2017) introduced a new species, A. ginkgonis isolated
13
Fungal Diversity
Fig. 91 Torula fici (reference
specimen) a Appearance of
colonies on substrate. b, c Conidiophores and conidiogenous
cells. d–f Conidia in chains. g–i
Conidia. Scale bars: a = 500 µm,
d–f = 20 µm, g–i = 10 µm, b,
c = 5 µm
from symptomatic branches of Ginkgo biloba in China. Over
300 epithets are listed in Index Fungorum (2020). Only a
few species have sequence data available in GenBank (Aplosporella africana, A. artocarpi, A. hesperidica, A. javeedii,
A. ginkgonis, A. macropycnidia, A. papillata, A. prunicola,
A. thailandica and A. yalgorensis). In this study, a new species, Aplosporella chromolaenae, and a new record A. hesperidica based on morphology and molecular data, together
with descriptions and illustrations are introduced (Figs. 94,
95). A phylogenetic tree based on combined LSU, ITS and
TEF1 sequence data is presented in Fig. 94.
Aplosporella chromolaenae Mapook & K.D. Hyde, sp. nov.
Index Fungorum number: IF557287, Facesoffungi number: FoF 07829; Fig. 95
Etymology: Name reflects the host genus Chromolaena,
from which this species was isolated.
Holotype: MFLU 20-0298
Saprobic on dead stems of Chromolaena odorata. Sexual
morph: Undetermined. Asexual morph: Conidiostromata
360–430 µm long, 685–780 µm wide ( x̄ = 400 × 735 µm,
n = 10), superficial, coriaceous, gregarious, dark brown to
black. Conidiomata 75–145 μm high × 80–160 µm diam.
( x̄ = 110.5 × 120.5 µm, n = 15), loculate; locules arranged
in rows, clustered, gregarious, with 2–3 locules forming
groups immersed in conidiostroma, globose to subglobose.
Conidiomata wall comprising several layers of thick-walled,
13
dark brown to black cells of textura angularis. Hamathecium comprising 1.5–2.5 µm wide, cylindrical to filiform,
septate, branching paraphyses. Conidiophores reduced to
conidiogenous cells. Conidiogenous cells 4.5–11 × 2.5–4 µm
( x̄ = 7 × 3.5 µm, n = 5), hyaline, holoblastic, oblong to ampuliform. Conidia(13–)16–20 × 8.5–12 µm ( x̄ = 17.5 × 9.8 µm,
n = 25), hyaline when immature, becoming brown to dark
brown when mature, aseptate, ellipsoid or oval to reniform,
thick-walled, with finely verruculose wall observed clearly
in mature conidia.
Culture characteristics: Conidia germinating on MEA
within 24 h. at room temperature and germ tubes produced
from both ends. Colonies on MEA, filamentous, mycelium
slightly raised, moderately fluffy, white aerial hyphae at the
surface, spreading from the center with white in reverse
(Fig. 97a).
Pre-screening for antimicrobial activity: Aplosporella
chromolaenae (MFLUCC 17-1517) showed antimicrobial
activity against E. coli with a 9 mm inhibition zone when
compared to the positive control (9 mm), but no inhibition
of B. subtilis and M. plumbeus.
Material examined: THAILAND, Chiang Rai Province,
Doi Pui, on dead stems of Chromolaena odorata, 7 April
2017, A. Mapook (DP108, MFLU 20-0298, holotype); extype culture MFLUCC 17-1517.
GenBank numbers: LSU: MT214434, ITS: MT214340,
SSU: MT214389
Fungal Diversity
Fig. 92 Torula polyseptata
(new host record) a, b Appearance of colonies on substrate. c–
g Conidiophores and conidiogenous cells. h–m Conidia. Scale
bars: a = 500 µm, b = 200 µm,
d = 20 µm, c, e–m = 10 µm
Fig. 93 Culture characteristics
on MEA: a Torula chromolaenae (MFLUCC 17-1514). b
Torula fici (MFLUCC 17-1494).
c Torula polyseptata (MFLUCC
17-1495)
Notes: In a BLASTn search of NCBI GenBank, the closest
match with the ITS sequence of Aplosporella chromolaenae
(MFLUCC 17-1517, ex-holotype) with 100% similarity was
Aplosporella artocarpi (strain CPC 22791, NR_154688),
while the closest match with the LSU sequence with
99.56% similarity was Aplosporella sp. (strain ICMP 17587,
EU931110). In the present phylogenetic analysis, A. chromolaenae clustered with A. artocarpi (CPC 22791), with
13
Fungal Diversity
Table 19 Synopsis of Torula species with similar morphological features discussed in this study
Species
Conidiophores (µm)
Conidiogenous cells (µm) Catenate conidia (µm)
Septation
References
T. breviconidiophora
(MFLU 18-1693)
T. chiangmaiensis (MFLU
16-2815)
T. chromolaenae
(MFLUCC 17-1514)
T. chromolaenae (MFLU
16-2819)
T. fici (MFLUCC
17-1494)
T. fici (MFLU 16-2817)
T. polyseptata (MFLU
18-1694)
T. polyseptata (MFLUCC
17-1495)
3.5–28 × 3–8
4–7 × 3–7
8–21 × 3.5–7
1–4-septate
Hyde et al. (2019a)
4–12-septate Li et al. (2017)
1–3-septate
This study
8–12.6(−16.1) × 4.5–5.2 3.4–6.5 × 4.8–7.6
4–4.7 × 4–5
3–6 × 3.5–6
(5.4–)25.5–
70(−86.5) × 5.6–7.8
7–18.5 × 4–8
5–6.3 × 3.5–4.6
(3.5–)4.5–5.1 × 4.8–5.6
12.1–16.5 × (3.6–)4.1–5
2–3-septate
Li et al. (2017)
2.5–6.5 × 3.9–5.5
4–7 × 5.5–7
10–25 × 5.5–8
1–5-septate
This study
9.4–12.5 × 3.7–4.5
10–40 × 3.5–8
5.4–8.5 × 5–7.4
4.5–8.5 × 4.5–8
12–20 × 4.6–6.6
10–40 × 3.5–7.5
2–4-septate
2–8-septate
Li et al. (2017)
Hyde et al. (2019a)
(4.5–)10–27 × 3.5–4.5
5.5–7.7 × 4.5–8
15–55 × 6.5–9
1–10-septate This study
bootstrap support (98% ML, Fig. 94). However, A. chromolaenae differs from A. artocarpi in having smaller conidiomata in conidiostromata (75–145 × 80–160 µm vs. (350–
)540–550(–650) × (490–)540–600(–700) µm) and larger
conidiogenous cells (3–5 × 2–4 µm vs. 4.5–11 × 2.5–4 µm)
(Table 20). Therefore, A. chromolaenae is described as
a new species based on phylogeny and morphological
comparison.
Aplosporella hesperidica Speg., Anal. Soc. cient. argent.
13(1): 18 (1882)
Facesoffungi number: FoF 07830; Fig. 96
Saprobic on dead stems of Chromolaena odorata. Sexual morph: Undetermined. Asexual morph: Conidiomata
365–440 µm high × 480–565 µm diam. ( x̄ = 398 × 516 µm,
n = 5), semi-immersed to superficial, erumpent, uniloculate,
coriaceous, solitary or gregarious, globose, dark-brown to
black. Peridium 30–45 µm wide at side, comprising several layers of thick-walled, brown to dark brown cells of
textura globulosa. Conidiophores reduced to conidiogenous
cells. Conidiogenous cells hyaline, holoblastic, oblong or
cylindrical to ampuliform. Conidia16–25(–30) × 9–15 µm
( x̄ = 19 × 11 µm, n = 25), brown to dark brown, aseptate,
ellipsoid to oval, thick-walled, with finely verruculose wall
observed clearly in mature conidia.
Culture characteristics: Conidia germinating on MEA
within 24 h. at room temperature and germ tubes produced
from the apex. Colonies on MEA, filamentous, mycelium
slightly raised, velvety with fluffy, white aerial hyphae at
the surface, spreading from the center with white in reverse
(Fig. 97b).
Pre-screening for antimicrobial activity: Aplosporella
hesperidica (MFLUCC 17-1518) showed antimicrobial
activity against E. coli with a 9 mm inhibition zone when
13
compared to the positive control (9 mm), but no inhibition
of B. subtilis and M. plumbeus.
Known hosts and distribution: Citrus aurantium (Rutaceae) in India (Rao 1969); early stem-end rot of Citrus sinensis (Rutaceae) in Zimbabwe (Yang et al. 2017).
Material examined: THAILAND, Chiang Rai Province,
Doi Pui, on dead stems of Chromolaena odorata, 7 April
2017, A. Mapook (DP110, MFLU 20-0299); living culture
MFLUCC 17-1518 (new host record).
GenBank numbers: LSU: MT214435, ITS: MT214341,
SSU: MT214390
Notes: A phylogenetic analysis shows strain MFLUCC
17-1518 grouped with Aplosporella hesperidica (Fig. 94).
In a BLASTn search of NCBI GenBank, the closest match
of the ITS and the LSU sequences of MFLUCC 17-1518
with 100% (JX681069) and 100% (KX464239) similarity,
respectively, was A. hesperidica. We therefore, identify our
isolates as A. hesperidica based on phylogenetic analyses
and the isolates are introduced here as a new host record
from Chromolaena odorata, collected in Thailand.
Botryosphaeriaceae Theiss. & P. Syd., Annales Mycologici
16 (1–2): 16 (1918)
Botryosphaeriaceae contains numerous plant pathogenic,
saprobic and endophytic species associated with a wide
range of hosts as well as opportunistic human pathogen (de
Hoog et al. 2000; Slippers and Wingfield 2007; Phillips et al.
2013; Mehl et al. 2014; Trakunyingcharoen et al. 2014a;
Doilom et al. 2015). The family was introduced by Theissen
and Sydow (1918) with Botryosphaeria as the type genus.
Divergence time estimates for this family is diverse around
94 Mya in the Cretaceous with crown age of 61 Mya in the
Paleogene period (Phillips et al. 2019).
Fungal Diversity
Fig. 94 Phylogram generated
from maximum likelihood
analysis based on combined
dataset of LSU, ITS and TEF1
sequence data. Twenty-five
strains are included in the
combined sequence analysis,
which comprise 1668 characters with gaps. Tree topology
of the ML analysis was similar
to the BYPP. The best scoring
RAxML tree with a final likelihood value of − 4044.627020
is presented. The matrix had
270 distinct alignment patterns, with 27.23% of undetermined characters or gaps.
Estimated base frequencies
were as follows: A = 0.224904,
C = 0.251747, G = 0.277253,
T = 0.246096; substitution rates:
AC = 2.555314, AG = 3.360132,
AT = 1.917272, CG = 2.128299,
CT = 6.642079, GT = 1.000000;
gamma distribution shape
parameter α = 0.149427. Bootstrap support values for ML
equal to or greater than 60% and
BYPP equal to or greater than
0.90 are given above the nodes.
Newly generated sequences
are in dark red bold and type
species are in bold. Saccharata
proteae (CBS 115206) is used
as outgroup taxon
Dothiorella Sacc.
Dothiorella was introduced by Saccardo (1880) with D.
pyrenophora as the type species. The genus comprises plant
pathogens, endophytes and saprobes of a wide range of hosts
(Crous et al. 2006; Liu et al. 2012; Hyde et al. 2013; Phillips
et al. 2013; Dissanayake et al. 2016; Váczy et al. 2018).Jayasiri et al. (2019) introduced a new species, D. lampangensis
from unidentified decaying fruit in Thailand. Phookamsak
et al. (2019) introduced a new species, D. acericola from
dried twigs of Acer palmatum in China with record of D.
sarmentorum from dead twigs of Platycladus orientalis in
Russia based on both morphology and phylogenetic analyses. We present a new host record for D. dulcispinae isolated
from C. odorata, together with a description and illustrations
(Fig. 99). A phylogenetic tree based on combined ITS and
TEF1 sequence data is presented in Fig. 98.
Dothiorella oblonga F.J.J. Van der Walt, Slippers & G.J.
Marais, in Slippers et al., Persoonia 33: 163 (2014)
Facesoffungi number: FoF 07831; Fig. 99
Saprobic on dead stems of Chromolaena odorata.
Sexual morph: Undetermined. Asexual morph: Conidiomata (110)180–280 µm high × 90–145 µm diam.
( x̄ = 189 × 115 µm, n = 5), semi-immersed, globose to obpyriform, coriaceous, solitary or scattered, appearing as dark
spots. Ostiole papillate. Pycnidial wall 10–25 µm wide,
3–5 layers, brown to dark brown cells of textura angularis.
Conidiophores reduced to conidiogenous cells. Conidiogenous cells 3.5–8 × 2.5–4.5(–6) µm ( x̄ = 6 × 3.5 µm, n = 20),
hyaline, holoblastic, cylindrical. Conidia 17–25 × 8–12.5 µm
( x̄ = 22 × 10.5 µm, n = 30), hyaline to pale brown when
immature, becoming drown to dark brown at maturity, ovoid
or oblong to ellipsoidal, aseptate or uniseptate, constricted at
the septum, moderately thick-walled with granular appearance, straight or slightly curved.
Culture characteristics: Conidia germinating on MEA
within 24 h. at room temperature and germ tubes produced from the apex. Colonies on MEA circular, mycelium
13
Fungal Diversity
Fig. 95 Aplosporella chromolaenae (holotype) a, b Appearance of stromata on substrate.
c Section through stroma. d
Peridium. e Hamathecium. f–h
Conidiogenous cells and developing conidia. i–n Conidia.
Scale bars: a, b = 500 µm,
c = 100 µm, d = 50 µm,
e–n = 10 µm
raised, velvety with moderately fluffy, filiform, white at first,
become olivaceous-grey with age (Fig. 100).
Pre-screening for antimicrobial activity: Dothiorella
oblonga (MFLUCC 17-1498) showed no inhibition of E.
coli, B. subtilis and M. plumbeus.
Known hosts and distribution: Acacia mellifera
(Fabaceae) in South Africa (Slippers et al. 2014)
Material examined: THAILAND, Chiang Rai Province,
Doi Pui, on dead stems of Chromolaena odorata, 2 February
2017, A. Mapook (DP97, MFLU 20-0310); living culture
MFLUCC 17-1498 (new host record).
GenBank numbers: LSU: MT214459, ITS: MT214365,
SSU: MT214411
Notes: A phylogenetic analysis (Fig. 98) shows that
MFLUCC 17-1498 grouped with Dothiorella oblonga,
with bootstrap support (84% ML). In a BLASTn search of
NCBI GenBank, the closest match of the ITS sequence for
MFLUCC 17-1498 is D. oblonga with 99.82% similarity to
13
the strain CBS 121765 (KF766163). We therefore, identify
our isolates as D. oblonga based on phylogenetic analyses
and morphological comparison (Table 21) and the isolates
are introduced here as a new host record from Chromolaena
odorata in Thailand.
Sphaeropsis Sacc.
Sphaeropsis was introduced by Saccardo (1880) with S.
visci as the type species. More than 600 species epithets
are listed in Index Fungorum (2020). Phookamsak et al.
(2019) introduced a new host record of S. eucalypticola from
Bauhinia purpurea in Thailand based on both morphology
and phylogenetic analyses. In this study, a new Sphaeropsis
species is introduced, based on morphology and molecular
data, together with descriptions and illustrations (Fig. 102).
A phylogenetic tree based on combined ITS, LSU, SSU,
TEF1 and TUB2 sequence data is presented in Fig. 101.
Fungal Diversity
Fig. 96 Aplosporella hesperidica (new host record) a
Appearance of conidiomata on
substrate. b Section through
conidioma. c Peridium.
d–g Conidiogenous cells
and developing conidia. h, i
Conidia. Scale bars: a = 500 µm,
b, c = 100 µm, g–i = 20 µm,
d–f = 10 µm
Fig. 97 Culture characteristics on MEA: a Aplosporella chromolaenae (MFLUCC 17-1517). b Aplosporella hesperidica (MFLUCC
17-1518)
Sphaeropsis chromolaenicola Mapook & K.D. Hyde, sp.
nov.
Index Fungorum number: IF557363, Facesoffungi number: FoF 07832; Fig. 102
Etymology: Name reflects the host genus Chromolaena,
on which this species was growing.
Holotype: MFLU 20-0369
Saprobic on dead stems of Chromolaena odorata. Sexual
morph: Ascomata 200–235 µm high × 260–285 µm diam.
( x̄ = 220 × 275 µm, n = 5), immersed to erumpent through
host tissue, globose, coriaceous, solitary or scattered,
appearing as brown to dark brown spots. Ostiole central,
short papillate. Peridium 20–50(–65)µm wide, 3–5 layers,
reddish brown to dark brown cells of textura angularis.
Hamathecium comprising 2.5–4 µm wide, oblong to cylindrical, septate pseudoparaphyses. Asci 90–130 × 34–45 µm
( x̄ = 112 × 37 µm, n = 10), 8-spored, bitunicate, fissitunicate,
cylindric-clavate to clavate, with a short pedicel, straight
to slightly curved, apically rounded with an ocular chamber. Ascospores 28.5–34 × 12–15 µm µm ( x̄ = 30 × 13.5 µm,
n = 25), 2–3-seriate, overlapping in the ascus, hyaline to
yellowish brown when immature, becoming brown to dark
brown at maturity, ellipsoid to ovoid, aseptate, with granular appearance, widest at the center and tapering toward
narrow ends, straight to slightly curved. Asexual morph:
Undetermined.
Culture characteristics: Ascospores germinating on
MEA within 24 h. at room temperature and germ tubes produced from the apex. Colonies on MEA circular, mycelium
velvety, moderately fluffy, filiform, smoke-grey to dark olivaceous at the surface and dark in reverse (Fig. 103).
13
Fungal Diversity
Fig. 98 Phylogram generated
from maximum likelihood analysis based on combined dataset
of ITS and TEF1 sequence data.
Seventy-two strains are included
in the combined sequence analysis, which comprise 888 characters with gaps. Tree topology
of the ML analysis was similar
to the BYPP. The best scoring
RAxML tree with a final likelihood value of − 4583.919557
is presented. The matrix had
382 distinct alignment patterns, with 14.43% of undetermined characters or gaps.
Estimated base frequencies
were as follows: A = 0.212092,
C = 0.286537, G = 0.252906,
T = 0.248465; substitution rates:
AC = 1.390409, AG = 2.605017,
AT = 1.096995, CG = 1.336154,
CT = 5.435452, GT = 1.000000;
gamma distribution shape
parameter α = 0.202628. Bootstrap support values for ML
equal to or greater than 60% and
BYPP equal to or greater than
0.90 are given above the nodes.
Newly generated sequences are
in dark red bold and type species are in bold. Neofusicoccum
luteum (CBS 110299) and N.
luteum (CBS 110497) are used
as outgroup taxa
Pre-screening for antimicrobial activity: Sphaeropsis
chromolaenicola (MFLUCC 17-1499) showed no inhibition
of E. coli, B. subtilis and M. plumbeus.
Material examined: THAILAND, Chiang Rai Province,
Doi Pui, on dead stems of Chromolaena odorata, 2 February 2017, A. Mapook (DP98, MFLU 20-0369, holotype);
ex-type culture MFLUCC 17-1499.
13
GenBank numbers: LSU: MT214460, ITS: MT214366,
SSU: MT214412
Notes: Multigene phylogenetic analyses (Fig. 101) show
that our strain Sphaeropsis chromolaenicola (MFLUCC
17-1499, ex-holotype) clusters with S. eucalypticola clade,
with high bootstrap support (92% ML and 0.99 BYPP). In a
BLASTn search of NCBI GenBank, the closest match of the
ITS sequence for MFLUCC 17-1499 is S. eucalypticola with
Fungal Diversity
Table 20 Synopsis of Aplosporella species with similar morphological features discussed in this study
Species
A. artocarpi (CPC 22791)
Conidiomata (µm)
(350–)540–550(–
650) × (490–)540–
600(–700)
A. chromolaenae (MFLUCC 17-1517) 75–145 × 80–160
A. hesperidica
–
A. hesperidica (MFLUCC 17-1518)
365–440 × 480–565
Conidiogenous cells (µm) Conidia (µm)
Reference
3–5 × 2–4
(17–)18–21(–22) × (9–)10–11 Trakunyingcharoen et al. (2015)
4.5–11 × 2.5–4
–
–
(13–)16–20 × 8.5–12
22–25 × 9–11
16–25(–30) × 9–15
This study
Spegazzini (1882)
This study
Fig. 99 Dothiorella oblonga
(new host record) a, b
Appearance of conidiomata on
substrate. c Section through
conidioma. d Peridium. e–i
Conidiogenous cells and developing conidia. j, k Conidia.
l Conidia with gelatinous
sheath in Indian ink. Scale
bars: a = 500 µm, b = 200 µm,
c = 50 µm, d, e–l = 10 µm
100% similarity to the strain CBS 133993 (MH866075),
while the closest match with the LSU sequence was with
S. porosa (strain CBS 110574, DQ377895) with 99.67%
similarity and closest matches with the SSU sequence were
S. porosa (strain CBS 110496, NG_062740) and S. visci
(strain CBS 100163, EU673177) with 99.82% similarity.
Therefore, S. chromolaenicola is described as a new species based on phylogeny with morphological comparison.
The new species, S. chromolaenicola is similar to S. eucalypticola in having cylindric-clavate or clavate asci with
ellipsoidal to ovoid, aseptate ascospores, with wide center
and tapering narrow ends, but S. chromolaenicola has
wider asci (90–130 × 34–45 µm vs. 102 − 175 × 22–32 µm)
and wider ascomata (200–235 × 260–285 µm), while
S. eucalypticola has smaller ascomata in ascostromata
(250–350 × 170 − 250 µm) (Table 22).
Dyfrolomycetales K.L. Pang et al.
Dyfrolomycetales was introduced by Pang et al. (2013)
with a monotypic family Pleurotremataceae (= Dyfrolomycetaceae). We follow the latest treatment and updated
accounts of Dyfrolomycetales in Pang et al. (2013), Norphanphoun et al. (2017), Zhang et al. (2017) and Hyde et al.
(2018).
Pleurotremataceae Walt. Watson (= Dyfrolomycetaceae
K.D. Hyde et al.)*
Pleurotremataceae was introduced by Watson (1929)
and accepted with monotypic genus Pleurotrema within
Chaetosphaeriales (Maharachchikumbura et al. 2015). The
family was excluded from Sordariomycetes based on reexamination of the isotype specimen of Pleurotrema polysemum, which is morphologically similar to Saccardoella and
13
Fungal Diversity
Fig. 100 Culture characteristic on MEA: Dothiorella oblonga
(MFLUCC 17-1498)
Dyfrolomyces in Dothideomycetes (Maharachchikumbura
et al. 2016). Dyfrolomycetaceae was introduced by Pang
et al. (2013) to accommodate the genus Dyfrolomyces with
Fig. 101 Phylogram generated from maximum likelihood
analysis based on combined
dataset of ITS, LSU, SSU,
TEF1 and TUB2 sequence data.
Twelve strains are included in
the combined sequence analysis,
which comprise 3285 characters with gaps. Tree topology
of the ML analysis was similar
to the BYPP. The best scoring
RAxML tree with a final likelihood value of − 6043.146013
is presented. The matrix had
256 distinct alignment patterns, with 17.97% of undetermined characters or gaps.
Estimated base frequencies
were as follows: A = 0.239114,
C = 0.251670, G = 0.275946,
T = 0.233269; substitution rates:
AC = 1.581937, AG = 2.385055,
AT = 0.562202, CG = 1.369785,
CT = 5.675544, GT = 1.000000;
gamma distribution shape
parameter α = 0.020000. Bootstrap support values for ML
equal to or greater than 60% and
BYPP equal to or greater than
0.90 are given above the nodes.
Newly generated sequences
are in dark red bold and type
species are in bold. Barriopsis
fusca (CBS 174.26) and B.
tectonae (MFLUCC 12-0381)
are used as outgroup taxa
13
D. tiomanensis as the type species and three Saccardoella
species, S. rhizophorae, S. mangrovei and S. marinospora
were transferred to Dyfrolomyces. Currently, Dyfrolomycetaceae is a synonymy under Pleurotremataceae (Maharachchikumbura et al. 2016) with three accepted genera,
Dyfrolomyces, Melomastia and Pleurotrema (Pang et al.
2013; Norphanphoun et al. 2017; Zhang et al. 2017; Wijayawardene et al. 2018). Divergence time estimate for this
family and estimated for crown age are in the early Cenozoic Era at 55 Mya (38–74). The family shares a common
ancestry with Palawaniaceae and Muyocopronaceae at 192
Mya (145–243) in the early Jurassic (Mapook et al. 2016c).
Lui et al. (2017) estimated in the late Cretaceous at 76 Mya
(38–125) for crown age and shares the common ancestry
with Acrospermaceae in the Jurassic at 174 Mya (113–243).
However, the families Palawaniaceae and Muyocopronaceae
were not included in this analysis.
Dyfrolomyces K.D. Hyde et al.
Fungal Diversity
Table 21 Synopsis of Dothiorella species with similar morphological features discussed in this study
Species
Conidiomata (µm)
Conidiogenous cells (µm)
Conidia (µm)
References
D. dulcispinae (PREM 60706)
up to 200 wide
–
(14–)16 − 22(–
24) × (6–)7 − 10(–11)
17–25 × 8 − 12.5
Jami et al. (2012)
D. oblonga (MFLUCC 17-1498)
(110)180–280 high × 90–145 3.5–8 × 2.5–4.5(–6)
diam.
D. oblonga (PREM 59628)
840.5 high × up to 550 diam. (6 −)8.5 − 11.5( − 12.5)
× (2.5–)4 − 4.5( − 5.5)
D. thailandica (MFLUCC 11-0438) 400–800 wide, 200–250
2–5.5 × 1.5 − 4.5
high, 250–500 diam.
Dyfrolomyces was introduced by Pang et al. (2013) with
the type species, D. tiomanensis, collected from unidentified
(18.5–)23.5 − 27(–28) ×
(10–)11.5–13(− 15)
15 − 20 × 6.5 − 8
This study
Slippers et al. (2014)
Liu et al. (2012)
mangrove wood, and they transferred three Saccardoella
species (S. rhizophorae, S. mangrovei and S. marinospora)
Fig. 102 Sphaeropsis chromolaenicola (holotype) a, b
Appearance of ascomata on
substrate. c Section through
ascoma. d Ostiole. e Peridium. f Pseudoparaphyses.
g–i Immature and mature asci.
j–o Ascospores (immature and
mature). Scale bars: a = 500 µm,
b = 200 µm, c, g–i = 50 µm, d,
e = 20 µm, f, j–o = 10 µm
13
Fungal Diversity
Fig. 103 Culture characteristic on MEA: Sphaeropsis chromolaenicola (MFLUCC 17-1499)
to Dyfrolomyces. Zhang et al. (2017) introduced two new
species (D. thamplaensis and D. maolanensis) from terrestrial habitats based on morphology together with multigene
analyses and transferred Saccardoella aquatica as D. aquatica, based on its similar morphological characters. Hyde
et al. (2018) introduced a new species, D. phetchaburiensis
from submerged wood of Rhizophora apiculata, based on
combined LSU and SSU sequence data with morphological
comparison. Eight epithets are listed in Index Fungorum
(2020) and most species have been collected from marine
habitats on mangrove wood, as well as in terrestrial (Pang
et al. 2013; Zhang et al. 2017; Hyde et al. 2018). In this
study, a new species Dyfrolomyces chromolaenae, is introduced, together with a description and illustrations, based on
morphology and molecular data (Fig. 105). A phylogenetic
tree based on combined LSU, SSU and TEF1 sequence data
is presented in Fig. 104.
Dyfrolomyces chromolaenae Mapook & K.D. Hyde, sp.
nov.
Index Fungorum number: IF557290, Facesoffungi number: FoF 07833; Fig. 105
Etymology: Name reflects the host genus Chromolaena,
from which this species was isolated.
Holotype: MFLU 20-0311
Saprobic on dead stems of Chromolaena odorata. Sexual
morph: Ascomata 315–380 µm high × 315–400 µm diam.
( x̄ = 340 × 355 µm, n = 5), immersed to erumpent through
host tissue, solitary or scattered, coriaceous to carbonaceous,
without a subiculum. Clypeus extending outwards, thicker
around the papilla. Ostiole central, papillate. Peridium
20–55 µm wide, comprising several layers of pale brown to
brown cells of textura angularis. Hamathecium comprising
1.5–3.5 µm wide, cylindrical to broadly filiform, septate,
branching pseudoparaphyses. Asci 135–160 × 7–8 µm ( x̄
13
= 145 × 7.5 µm, n = 10), 8-spored, cylindrical, short pedicellate, straight or slightly curved, apically rounded, with an
apical ring. Ascospores 29–35 × 4.5–6 µm ( x̄ = 32 × 5.5 µm,
n = 15), uniseriate, hyaline, fusiform, tapering towards narrow ends, 1-septate when immature, becoming 9–11-septate
when mature, straight or slightly curved, without terminal
appendages. Asexual morph: Undetermined.
Culture characteristics: Ascospores germinating on
MEA within 24 h. at room temperature and germ tubes produced from both ends. Colonies on MEA circular, mycelium
slightly raised, entire to slightly filamentous, white aerial
hyphae at the surface, becoming greyish brown and dark
brown in reverse (Fig. 106).
Pre-screening for antimicrobial activity: Dyfrolomyces
chromolaenae (MFLUCC 17-1434) showed antimicrobial
activity against B. subtilis with a 12 mm inhibition zone,
when compared to the positive control (26 mm), but no inhibition of E. coli and M. plumbeus.
Material examined: THAILAND, Chiang Rai Province,
Doi Pui, on dead stems of Chromolaena odorata, 5 August
2015, A. Mapook (DP17, MFLU 20-0311, holotype); extype culture MFLUCC 17-1434.
GenBank numbers: SSU: MT214413, TEF1: MT235800
Notes: In a BLASTn search of NCBI GenBank,
the closest match of the SSU sequence of Dyfrolomyces chromolaenae (MFLUCC 17-1434, ex-holotype) is
D. rhizophorae with 97.18% similarity to the strain JK
5349A (GU479766), while the closest match of the TEF1
sequence with 89.74% similarity was D. tiomanensis (strain
NTOU3636 = MFLUCC13-0440, KC692157). In the present
phylogenetic analysis, D. chromolaenae is found closely
related to D. tiomanensis strain MFLUCC13-0440 (Fig. 104).
However, D. chromolaenae differs from D. tiomanensis in having smaller ascomata (315–380 × 315–400 µm
vs. 565–(615)–667 × 283–(374)–446 µm), smaller asci
(135–160 × 7–8 µm vs. 316–(323)–333 × 12–(16)–17 µm)
and smaller ascospores (29–35 × 4.5–6 µm vs.
69–(74)–82 × 9–(10)–11 µm) with 9–11-septa, while D. tiomanensis has 20–24-septate ascospores (Table 23). A comparison of the TEF1 gene region of D. chromolaenae and D.
tiomanensis reveals 81 base pair differences (10%) across
801 nucleotides. Therefore, D. chromolaenae is described
as a new species based on phylogeny and morphological
comparison.
Muyocopronales Mapook et al.
Muyocopronales was introduced by Mapook et al.
(2016b). The order comprises two families, Muyocopronaceae and Palawaniaceae, based on phylogenetic analyses
and morphological comparison, together with their divergence times for additional evidence to recognize the status
of higher level taxa. See Mapook et al. (2016b, c) for the
details.
Fungal Diversity
Table 22 Synopsis of Sphaeropsis species with similar morphological features discussed in this study
Species
Ascomata (µm)
Asci (µm)
S. chromolaenicola (MFLUCC
17-1499)
S. eucalypticola (MFLUCC
11-0579 = CBS 133993)
S. eucalypticola (MFLUCC
12-0171)
200–235 high × 260–285 diam. 90–130 × 34–45
Ascospores (µm) References
28.5–34 × 12–15 This study
–
(90–)97 − 110(−125) × 28–30 27–35 × 11–14
Liu et al. (2012)
Ascostromata, 250–350
high × 170 − 250 diam.
102 − 175 × 22–32
Phookamsak et al. (2019)
27–33 × 11–14
Muyocopronaceae K.D. Hyde
Muyocopronaceae was introduced by Luttrell (1951)
and was included in the order Hemisphaeriales. The family
was reintroduced by Hyde et al. (2013) as a distinct family
(Dothideomycetes family, incertae sedis) with monotypic
genus Muyocopron. Mapook et al. (2016b) introduced the
new order Muyocopronales to accommodate three new Muyocopron species which were collected in Thailand, based on
phylogenetic analyses and morphological comparison. Crous
et al. (2018a) introduced a new genus, Neocochlearomyces
on leaves of Chromolaena odorata in Thailand and accepted
six genera (Arxiella, Leptodiscella, Mycoleptodiscus, Muyocopron, Neocochlearomyces, Paramycoleptodiscus) in the
family. Hernández-Restrepo et al. (2019) introduced Neomycoleptodiscus as a new genus similar to Mycoleptodiscus in
Muyocopronaceae. Mapook et al. (2020) introduced a new
genus Pseudopalawania in the family with their secondary
metabolites production and biological activity. Divergence
time estimates for this family are crown age of 52 Mya
(38–66) in the Cenozoic Era, Paleogene and shares the most
common ancestor with Palawaniaceae at 172 Mya (130–218)
in Jurassic period (Mapook et al. 2016c). Samarakoon et al.
Fig. 104 Phylogram generated from maximum likelihood analysis
based on combined dataset of LSU, SSU and TEF1 sequence data.
Thirteen strains are included in the combined sequence analysis,
which comprise 2745 characters with gaps. Tree topology of the
ML analysis was similar to the BYPP. The best scoring RAxML tree
with a final likelihood value of − 7474.968962 is presented. The
matrix had 399 distinct alignment patterns, with 27.36% of undetermined characters or gaps. Estimated base frequencies were as follows: A = 0.243143, C = 0.248312, G = 0.294370, T = 0.214174;
substitution rates: AC = 1.048096, AG = 2.972507, AT = 1.232056,
CG = 1.713483, CT = 11.686012, GT = 1.000000; gamma distribution shape parameter α = 0.159409. Bootstrap support values for ML
equal to or greater than 60% and BYPP equal to or greater than 0.90
are given above the nodes. Newly generated sequences are in dark red
bold and type species are in bold. Acrospermum compressum (M151),
A. gramineum (M152) and A. adeanum (M133) are used as outgroup
taxa
13
Fungal Diversity
Fig. 105 Dyfrolomyces
chromolaenae (holotype)
a, b Appearance of ascomata on substrate. c Section
through ascoma. d Peridium.
e Pseudoparaphyses. f–i Asci.
j–o Ascospores. Scale bars:
a = 500 µm, b = 200 µm,
c = 100 µm, f–i = 50 µm,
d = 20 µm, e, j–o = 10 µm
(2019) recorded fossils justification of Muyocopron with two
known fossil species M. mucoris and M. neyveliensis, which
belong to the Cenozoic Era (59–5.33 Mya) and suggested
13
the minimum age at 54 Mya instead of 52 Mya for the future
calibration point of molecular clock analyses.
Fungal Diversity
Fig. 106 Culture characteristic on MEA: Dyfrolomyces chromolaenae (MFLUCC 17-1434)
Muyocopron Speg., Anales de la Sociedad Cientifica Argentina. 12: 113 (1881)
Muyocopron was introduced by Spegazzini (1881b) with
Mu. corrientinum as the type species, collected from dead
leaves of Oncidium sp. in Argentina. The genus is mostly
a saprobe common on the surface of dead aerial twigs,
branches, stems or leaves of a wide variety of plants and
reported from tropical and temperate regions (Taylor and
Hyde 2003; Mapook et al. 2016b). Muyocopron was transferred to Botryosphaeriaceae (von Arx and Müller 1975) as
well as Microthyriaceae (Saccardo 1883; Hawksworth et al.
1995; Lumbsch and Huhndorf 2007, 2010a, b; Kirk et al.
2008), based on morphology. Subsequently, the genus was
excluded from Microthyriaceae and transferred to Dothideomycetes genera, incertae cedis (Wu et al. 2011a, b). Hyde
et al. (2013) reintroduced Muyocorponaceae as a distinct
family with two unidentified Muyocopron species based
on morphological comparison, supported with molecular
studies. Mapook et al. (2016b) introduced the new order
Muyocopronales to accommodate three new Muyocopron
species from northern Thailand (Mu. castanopsis, Mu. dipterocarpi and Mu. lithocarpi), based on a distinct lineage
and morphological comparison with Dyfrolomycetales and
Acrospermales. Tibpromma et al. (2016) also introduced a
new species, Mu. garethjonesii collected from dead leaves
of Pandanus sp. in China. Senwanna et al. (2019) reported
a new host record, Mu. dipterocarpi with a new species,
Mu. heveae from dried twig of Hevea brasiliensis in Thailand. Phookamsak et al. (2019) reported a new host record
of Mu. lithocarpi from dead stems of herbaceous plant in
China and Jayasiri et al. (2019) reported new host record
of Mu. dipterocarpi from twigs and pod of Delonix regia
from Thailand, as well as Mu. lithocarpi on leaves and wild
pods. Hernández-Restrepo et al. (2019) introduced two
new Muyocopron species associated with leaf spots (Mu.
alcornii on Epidendrum sp. in Australia and Mu. zamiae
on Zamia in USA), with four new combinations of Mycoleptodiscus laterale, My. coloratum, My. geniculatum and
My. atromaculans, to the genus Muyocopron. In this study,
Muyocopron chromolaenae is introduced as a new species
and a new host record for Mu. lithocarpi from Thailand,
together with description and illustrations, based on morphology and molecular data (Figs. 108, 109, 110). A phylogenetic tree based on combined LSU, ITS, SSU, TEF1 and
RPB2 sequence data is presented in Fig. 107.
Muyocopron chromolaenae Mapook & K.D. Hyde, sp. nov.
Index Fungorum number: IF557309, Facesoffungi number: FoF 07834; Fig. 108
Etymology: Name reflects the host genus Chromolaena,
from which this species was isolated.
Holotype: MFLU 20-0328
Saprobic on dead stems of Chromolaena odorata. Sexual morph: Ascomata 120–150(–160) µm
high × (315–)370–425(–480) µm diam. ( x̄ = 135 × 390 µm,
n = 10), superficial, coriaceous, solitary or scattered, appearing as circular, scattered, flattened, brown to dark brown
spots, covering the host, without a subiculum, with a poorly
developed basal layer and an irregular margin. Ostiole central. Peridium 10–40(–50) µm wide, widest at the sides,
outer layers comprising dark brown to black pseudoparenchymatous, occluded cells of textura epidermoidea, inner
layers comprising light brown to hyaline cells of textura
epidermoidea. Hamathecium comprising 0.5–1.5 µm wide,
cylindrical to filiform, septate pseudoparaphyses. Asci
40–55 × 10–20 µm ( x̄ = 50 × 16 µm, n = 10), 8-spored, bitunicate, saccate or broadly obpyriform, pedicellate, straight
or slightly curved, with an ocular chamber. Ascospores
15–20 × 6–9 µm ( x̄ = 18 × 7.5 µm, n = 20), irregularly
arranged, overlapping in the ascus, hyaline, broadly fusiform, aseptate, smooth, widest at the center and tapering
towards ends, straight or slightly curved. Asexual morph:
Undetermined.
Table 23 Synopsis of Dyfrolomyces species with similar morphological features discussed in this study
Species
Ascomata (µm)
Asci (µm)
Ascospores (µm)
D. chromolaenae (MFLUCC 17-1434)
D. tiomanensis
(MFLUCC13-0440 = NTOU3636)
315–380 × 315–400
135–160 × 7–8
29–35 × 4.5–6, 9–11-septate
565 − (615) − 667 × 283 316 − (323) − 333 × 1 69 − (74) − 82 × 9 − (10) − 11,
− (374) − 446
2 − (16) − 17
20 − 24-septate
References
This study
Pang et al. (2013)
13
Fungal Diversity
Fig. 107 Phylogram generated
from maximum likelihood analysis based on combined dataset
of LSU, ITS, SSU, TEF1 and
RPB2 sequence data. Seventytwo strains are included in the
combined sequence analysis,
which comprise 5396 characters with gaps. Tree topology
of the ML analysis was similar
to the BYPP. The best scoring
RAxML tree with a final likelihood value of − 31179.160692
is presented. The matrix had
2314 distinct alignment patterns, with 61.16% of undetermined characters or gaps.
Estimated base frequencies
were as follows: A = 0.237959,
C = 0.253544, G = 0.286148,
T = 0.222349; substitution rates:
AC = 1.253242, AG = 2.577505,
AT = 1.560994, CG = 1.058014,
CT = 5.657031, GT = 1.000000;
gamma distribution shape
parameter α = 0.357213. Bootstrap support values for ML
equal to or greater than 60% and
BYPP equal to or greater than
0.90 are given above the nodes.
Newly generated sequences
are in dark red bold and type
species are in bold. Lophium
mytilinum (AFTOL-ID 1609)
and Mytilinidion rhenanum
(CBS 135.45) are used as outgroup taxa
Culture characteristics: Ascospores germinating on
MEA within 24 h. at room temperature and germ tubes produced from the apex. Colonies on MEA circular, initially
mycelium white, slightly raised, velvety with moderately
fluffy, filamentous, cultures grayish to light brown in old,
flattened on surface, brown to dark brown in reverse from the
centre of the colony, pale brown to white margin (Fig. 111a).
13
Pre-screening for antimicrobial activity: Muyocopron
chromolaenae (MFLUCC 17-1513) showed no inhibition of
E. coli, B. subtilis and M. plumbeus.
Material examined: THAILAND, Phrae, Doi Pha Klong,
on dead stems of Chromolaena odorata, 22 September 2016,
A. Mapook (DPKP8, MFLU 20-0328, holotype); ex-type
culture MFLUCC 17-1513.
Fungal Diversity
Fig. 108 Muyocopron chromolaenae (holotype) a, b Superficial ascomata on substrate.
c, d Squash mounts showing
ascomata walls. e Section of
ascoma. f Peridium. g Pseudoparaphyses. h–k. Asci. l–q
Unicellular ascospores. Scale
bars: a = 500 µm, b = 200 µm,
c, e = 50 µm, d, f, h–k = 20 µm,
l–q = 10 µm, g = 5 µm
GenBank numbers: LSU: MT137876, ITS: MT137777,
SSU: MT137881, TEF1: MT136756, RPB2: MT136761
Notes: In a BLASTn search of NCBI GenBank, the
closest match of the SSU sequence of Muyocopron chromolaenae (MFLUCC 17-1513, ex-holotype) was Mu. dipterocarpi with 96.69% similarity to the strain MFLUCC
14-1103(KU726969), while the closest match of the LSU
sequence with 99.06% similarity was Muyocopron geniculatum (strain CBS 721.95, MH874185). On phylogenetic analysis, Mu. chromolaenae forms a separate branch and clusters
with Mu. atromaculans, Mu. geniculatum and Mu. zamiae
(Fig. 107). However, we could not compare the morphological characteristics of those species; Mu.chromolaenae
is found as sexual morph in nature and we could not obtain
its asexual morph in culture, while Mu. atromaculans, Mu.
geniculatum and Mu. zamiae were found as asexual morphs
in nature. A comparison of the ITS (+5.8S) gene region
of Mu. chromolaenae and Mu. geniculatum reveals 37 base
pair differences (6.6%) across 561 nucleotides. Therefore,
Mu.chromolaenae is described here as a new species based
on phylogeny.
Muyocopron chromolaenicola Mapook & K.D. Hyde, sp.
nov.
Index Fungorum number: IF557310, Facesoffungi number: FoF 07835; Fig. 109
Etymology: Name reflects the host genus Chromolaena,
on which this species was growing.
Holotype: MFLU 20-0329
Saprobic on dead stems of Chromolaena odorata. Sexual
morph: Ascomata 80–135 µm high × 360–440 µm diam.
( x̄ = 106 × 396 µm, n = 10), superficial, coriaceous, solitary
or scattered, appearing as circular, flattened, dark brown to
black spots, without a subiculum, with a poorly developed
basal layer and an irregular margin. Ostiole central. Peridium 20–35(–50)µm wide, widest at the sides, outer layers
13
Fungal Diversity
Fig. 109 Muyocopron
chromolaenicola (holotype)
a, b Superficial ascomata on
substrate. c, d Squash mounts
showing ascomata walls. e Section of ascoma. f Peridium. g
Pseudoparaphyses. h–k. Asci. l–
q Unicellular ascospores. Scale
bars: a = 1000 µm, b = 500 µm,
c = 100 µm, e = 50 µm, f,
h–k = 20 µm, d, l–q = 10 µm,
g = 5 µm
comprising dark brown to black pseudoparenchymatous,
occluded cells of textura epidermoidea, inner layers comprising light brown cells of textura angularis. Hamathecium comprising (1.5–)2–3.5 µm wide, filiform to cylindrical, septate pseudoparaphyses. Asci 50–80 × 20–30 µm
( x̄ = 63 × 24 µm, n = 20), 8-spored, bitunicate, saccate or
broadly obpyriform, pedicellate, straight or slightly curved,
with a small ocular chamber. Ascospores 14.5–17 × 9–12 µm
( x̄ = 16 × 10.5 µm, n = 20), irregularly arranged, overlapping in the ascus, hyaline, oval to obovoid, with obtuse
ends, aseptate, with granular appearance. Asexual morph:
Undetermined.
Culture characteristics: Ascospores germinating on
MEA within 24 h. at room temperature and germ tubes produced from both ends. Colonies on MEA circular, slightly
raised, initially mycelium white, filamentous, greyish white
in old cultures at the margin, creamy brown in reverse from
the centre of the colony, becoming brown to olivaceous
brown with white margin (Fig. 111b).
13
Pre-screening for antimicrobial activity: Muyocopron
chromolaenicola (MFLUCC 17-1470) showed no inhibition
of E. coli, B. subtilis and M. plumbeus.
Material examined: THAILAND, Mae Hong Son Province, Mae Yen, on dead stems of C. odorata, 25 June 2016,
A. Mapook (MY3, MFLU 20-0329, holotype); ex-type culture MFLUCC 17-1470.
GenBank numbers: LSU: MT137877, ITS: MT137778,
SSU: MT137882, TEF1: MT136757
Notes: In a BLASTn search of NCBI GenBank, the
closest match of the LSU sequence of Muyocopron chromolaenicola (MFLUCC 17-1470, ex-holotype) was Mu.
lithocarpi with 99.76% similarity to the strain MFLUCC
16-0962 (MK348034), while the closest match of the
SSU sequence with 99.62% similarity was Mu. lithocarpi
(strain MFLUCC 14-1106, KU726970). In the present phylogenetic analysis, Mu. chromolaenicola forms
a separate branch as a sister taxon to Mu. lithocarpi and
Mu. heveae (Fig. 107). However, Mu. chromolaenicola
differs from Mu. lithocarpi in having larger ascomata
Fungal Diversity
Fig. 110 Muyocopron
lithocarpi (new host record)
a, b Superficial ascomata on
substrate. c, d Squash mounts
showing ascomata walls. e Section of ascoma. f Peridium. g
Pseudoparaphyses. h–k. Asci. l–
q Unicellular ascospores. Scale
bars: a = 500 µm, b = 200 µm,
c = 100 µm, d–f = 50 µm,
h–k = 20 µm, g, l–q = 10 µm
(80–135 × 360–440 µm vs. 40–70(–120) × 220–320 µm) with
longer asci (50–80 × 20–30 µm vs. 45–65 × (15–)23–28 µm),
and differs from Mu. heveae in having smaller ascomata (80–135 × 360–440 µm vs. 65–180 × 180–620 µm)
and smaller ascospores (14.5–17 × 9–12 µm vs.
(19)22–32(37) × (8)10–14(15) µm) with shorter asci
(50–80 × 20–30 µm vs. (57)77–116(130) × (19)22–31(36)
µm) (Table 24). A comparison of the ITS (+5.8S) gene
region of Mu. chromolaenicola and Mu. lithocarpi reveals
37 base pair differences (6.3%) across 584 nucleotides.
Therefore, Mu. chromolaenicola is described as a new species based on phylogeny and morphological comparison.
Muyocopron lithocarpi Mapook, Boonmee & K.D. Hyde,
Phytotaxa 265(3): 235 (2016)
Facesoffungi number: FoF 01890; Fig. 110
Saprobic on dead stems of Chromolaena odorata. Sexual morph: Ascomata 65–90.5 µm high × 225–330 µm
diam. ( x̄ = 75 × 290 µm, n = 10), superficial, coriaceous,
solitary or scattered with gregarious or confluent, appearing as circular, scattered, flattened, brown to dark brown
spots, covering the host, without a subiculum, with a poorly
developed basal layer and an irregular margin. Ostiole
central. Peridium 15–25(–40) µm wide, widest at the sides,
outer layers comprising dark brown to black pseudoparenchymatous, occluded cells of textura epidermoidea, inner
layers comprising light brown cells of textura angularis.
Hamathecium comprising 1–2 µm wide, cylindrical to filiform, septate pseudoparaphyses. Asci 49–77 × 17–26 µm
( x̄ = 60 × 22.5 µm, n = 25), 8-spored, bitunicate, saccate or
broadly obpyriform, pedicellate, straight or slightly curved,
with small ocular chamber. Ascospores 14–20 × 9–14 µm
( x̄ = 17 × 11 µm, n = 15), irregularly arranged, overlapping in the ascus, hyaline, oval to obovoid with obtuse
ends, aseptate, with granular appearance. Asexual morph:
Undetermined.
Culture characteristics: Ascospores germinating on
MEA within 24 h at room temperature and germ tubes produced from both ends. Colonies on MEA irregular, slightly
raised, initially mycelium white, filamentous, greyish to light
brown in old cultures, flattened on surface, brown to dark
brown in reverse from the centre of the colony, pale brown
to white margin (Fig. 111c).
13
Fungal Diversity
Fig. 111 Culture characteristics on MEA: a Muyocopron
chromolaenae (MFLUCC
17-1513). b Muyocopron
chromolaenicola (MFLUCC
17-1470). c Muyocopron
lithocarpi (MFLUCC 17-1500)
Pre-screening for antimicrobial activity: In this study,
all isolates of Muyocopron lithocarpi showed no inhibition
of E. coli, B. subtilis and M. plumbeus.
Known hosts and distribution: On dead leaves of
Lithocarpus lucidus (Fagaceae) in Thailand (Mapook et al.
2016); on decaying pods of Peltophorum sp. (Fabaceae) in
Thailand and on fallen pod of Cercis chinensis (Fabaceae)
in China (Jayasiri et al. 2019); on dead stems of herbaceous
plant in China (Phookamsak et al. 2019).
Material examined: THAILAND, Chiang Rai Province,
Doi Mae Salong, on dead stems of Chromolaena odorata, 17
May 2016, A. Mapook (DMS2, MFLU 20-0330); living culture MFLUCC 17-1466; Doi Pui, on dead stems of C. odorata, 2 February 2017, A. Mapook (DP99, MFLU 20-0331);
living culture MFLUCC 17-1500 (new host record); Lampang Province, Chae Son, on dead stems of C. odorata, 24
September 2016, A. Mapook (JS1, MFLU 20-0332); living
culture MFLUCC 17-1465.
GenBank numbers: LSU: MT137878, MT137879,
MT137880, ITS: MT137779, MT137780, MT137781,
SSU: MT137883, MT137884, MT137885, TEF1:
MT136758, MT136759, MT136760, RPB2: MT136762
Notes: Multigene phylogenetic analyses (Fig. 107) show
that three strains MFLUCC 17-1466, MFLUCC 17-1500 and
MFLUCC 17-1465 are grouped with Muyocopron lithocarpi
clade. In a BLASTn search of NCBI GenBank, the closest match of the LSU and SSU sequences showed that the
strains are identical to Mu. lithocarpi with 100% similarity.
We therefore, identify our three isolates as Mu. lithocarpi
based on phylogenetic analyses with morphological comparison (Table 24) and the isolates are introduced here as
a new host record from Chromolaena odorata collected in
Thailand.
Patellariales D. Hawksw. & O.E. Erikss.
Patellariales was introduced by Hawksworth and Eriksson (1986) with the monotypic family Patellariaceae. We
follow the latest treatment and updated accounts of Patellariales in Hernández-Restrepo et al. (2016) and Pem et al.
(2018).
Patellariaceae Corda
Patellariaceae was introduced by Corda (1838) with
Patellaria as the type genus and contains species which are
saprobes or weak parasites on wood and bark of various trees
and shrubs. The family was classified in the order Patellariales and 21 genera were accepted in the outline classification
of Ascomycota (Wijayawardene et al. 2018). Members of
Patellariaceae have very few molecular studies. Five genera, Hysteropatella, Yuccamyces, Patellaria, Glyphium and
Holmiella have been reported with available sequence in
GenBank (Boehm et al. 2009b, 2015; Schoch et al. 2009;
Hyde et al. 2013; Yacharoen et al. 2015; Pem et al. 2018).
Divergence time estimates for this family are crown age of
164 Mya (72–283) in the Jurassic period and stem age of
311 Mya (244–407) in the Carboniferous period (Liu et al.
2017).
Table 24 Synopsis of Muyocopron species with similar morphological features discussed in this study
Species
Ascomata (µm)
Peridium (µm) Asci (µm)
Mu. chromolaenicola
(MFLUCC 17-1470)
Mu. heveae (MFLUCC
17-0066)
80–135 × 360–440
20–35(–50)
65–180 × 180–620
12–41
Mu. lithocarpi (MFLUCC 65–90.5 × 225–330
15–25(–40)
17-1500)
Mu. lithocarpi (MFLUCC 40–70(–120) × 220–320 10–20(–28)
14-1106)
13
50–80 × 20–30
Ascospores (µm)
14.5–17 × 9–12, oval to
obovoid
(19)22–32(37) × (8)10–
(57)77–
116(130) × (19)22– 14(15), ellipsoidal
or broad oblong to
31(36)
obovoid
49–77 × 17–26
14–20 × 9–14, oval to
obovoid
45–65 × (15–)23–28 13–18 × 9–11, oval to
obovoid
References
This study
Senwanna et al. (2019)
This study
Mapook et al. (2016b)
Fungal Diversity
Patellaria Fr., Systema Mycologicum 2: 158 (1822)
Patellaria was introduced by Fries (1822) with Patellaria
atrata as the type species. The genus has more than 500
species epithets (Index Fungorum 2019). However, only a
few species have been explored with molecular data (Schoch
et al. 2009; Hernández-Restrepo et al. 2016). HernándezRestrepo et al. (2016) described a new phoma-like fungus,
Patellaria quercus based on 99% similarity of the LSU
sequence with Patellaria cf. atrata. However, further collections are needed to resolve the asexual morphs linked
to Patellariaceae. In this study, Patellaria chromolaenae
is introduced as a new species based on morphology and
molecular data support with a description and illustrations
(Fig. 113). A phylogenetic tree based on LSU, ITS, SSU,
TEF1 and RPB2 sequence data is presented in Fig. 112.
Patellaria chromolaenae Mapook & K.D. Hyde, sp. nov.
Index Fungorum number: IF557350, Facesoffungi number: FoF 07836; Fig. 113
Etymology: Name reflects the host genus Chromolaena,
from which this species was isolated.
Holotype: MFLU 20-0346
Saprobic on dead stems of Chromolaena odorata. Sexual
morph: Ascomata 100–145 µm high × 415–625 µm diam.
(x̅ = 120 × 525 µm, n = 5), apothecial, superficial, circular, flattened with a carbonaceous rim, scattered, brown to
dark brown. Exciple (15–)20–30(–35) µm wide, outer layers comprising dark brown to black cells and inner layers
of hyaline cells arranged in a textura prismatica, continuous to the base (hypothecium). Hamathecium comprising (1.5–)2–2.7 µm wide, cylindrical to filiform, septate,
branching pseudoparaphyses, slightly swollen and rounded
at the apex, forming magenta to dark purple epithecium
Fig. 112 Phylogram generated from maximum likelihood analysis based on combined dataset of LSU, ITS, SSU, TEF1 and RPB2
sequence data. Twenty-three strains are included in the combined
sequence analysis, which comprise 5609 characters with gaps. Tree
topology of the ML analysis was similar to the BYPP. The best scoring RAxML tree with a final likelihood value of − 19011.857286
is presented. The matrix had 1461 distinct alignment patterns, with
57.16% of undetermined characters or gaps. Estimated base frequen-
cies were as follows: A = 0.247075, C = 0.242521, G = 0.279791,
T = 0.230613; substitution rates: AC = 1.589017, AG = 2.732102,
AT = 1.158430, CG = 1.354331, CT = 6.468048, GT = 1.000000;
gamma distribution shape parameter α = 0.258762. Bootstrap support values for ML equal to or greater than 60% and BYPP equal
to or greater than 0.90 are given above the nodes. Newly generated
sequences are in dark red bold and type species are in bold. Botryosphaeria dothidea (CBS 115476) is used as outgroup taxon
13
Fungal Diversity
Fig. 113 Patellaria chromolaenae (holotype) a, b Appearance
of ascomata on substrate. c Section through ascoma. d Exciple.
e Pseudoparaphyses mounted
in lactoglycerol. f Pseudoparaphyses mounted in water. g, h
Asci mounted in lactoglycerol.
i, j Asci mounted in water.
k-p Ascospores. Scale bars:
a, b = 500 µm, c = 100 µm, d,
g–j = 20 µm, e, f, k–q = 10 µm
above the asci when mounted in lactoglycerol and navy
blue epithecium above the asci when mounted in water.
Asci 46–60(–70) × 12–16 µm ( x̄ = 54 × 14 µm, n = 15),
8-spored, bitunicate, fissitunicate, cylindric-clavate, straight
or slightly curved, with a short, bulbous pedicel, apically
rounded. Ascospores 23–32 × 4–5.5 µm ( x̄ = 25.5 × 5 µm,
n = 25), overlapping, 2–3-seriate, hyaline, allantoid to
broadly fusiform, 3–5-septate, straight or slightly curved,
guttulate, without terminal appendages. Asexual morph:
Undetermined.
13
Culture characteristics: Ascospores germinating on
MEA within 24 h at room temperature and germ tubes produced from both ends. Colonies on MEA irregular, mycelium slightly raised, undulate to lobate, brown with white
aerial hyphae spreading from the center of the colony,
becoming smoke-grey at the surface and dark yellowish
brown to brown in reverse (Fig. 114).
Pre-screening for antimicrobial activity: Patellaria
chromolaenae (MFLUCC 17-1479) showed antimicrobial
activity against E. coli with a 13 mm inhibition zone when
Fungal Diversity
compared to the positive control (9 mm), but no inhibition
of B. subtilis and M. plumbeus.
Material examined: THAILAND, Phrae Province, Mae
Kam, on dead stems of Chromolaena odorata, 21 September 2016, A. Mapook (MK1, MFLU 20-0346, holotype);
ex-type culture MFLUCC 17-1479; Lampang Province,
Ngao, on dead stems of C. odorata, 21 September 2016, A.
Mapook (LP4, MFLU 20-0345); living culture MFLUCC
17-1482.
GenBank numbers: LSU: MT214474, MT214475, ITS:
MT214380, MT214381, SSU: MT214425, MT214426,
TEF1: MT235795, MT235796, RPB2: MT235828,
MT235829
Notes: In a BLASTn search of NCBI GenBank, the
closest match of the LSU sequence of Patellaria chromolaenae (MFLUCC 17-1479, ex-holotype) was Patellaria quercus with 90.42% similarity to the strain CPC
27232 (NG_059696). The closest match of the ITS
sequence with 84.07% similarity was P. quercus (strain
BHI-F768exna, MF161322). The closest match of the
SSU sequence with 99.25% similarity was P. cf. atrata
(strain BCC 28877, GU371837). The closest match of the
TEF1 and RPB2 sequences with 91.25% (GU349038) and
88.43% (GU371726) similarity, respectively, was Patellaria atrata strain CBS 958.97. In the present phylogenetic analysis, P. chromolaenae forms a sister taxon to P.
atrata (Fig. 112). However, P. chromolaenae differs from P.
atrata in having smaller ascomata (100–145 × 415–625 µm
vs. 675–1160 × 220–300 µm), smaller asci
(46–60(–70) × 12–16 µm vs. 98–135 × 15–30 µm) and
smaller ascospores (23–32 × 4–5.5 µm vs. 30–45 × 7–10 µm)
that are 3–5-septate, while P. atrata has 5–11-septate
ascospores (Table 25). A comparison of the ITS (+ 5.8S)
gene region of P. chromolaenae and P. atrata reveals 135
base pair differences (21.5%) across 627 nucleotides.
Fig. 114 Culture characteristic on MEA: Patellaria chromolaenae
(MFLUCC 17-1479)
Therefore, P. chromolaenae is described as a new species
based on phylogeny and morphological comparison.
Class Sordariomycetes O.E. Erikss. & Winka
Subclass Diaporthomycetidae Senan. et al.
Diaporthales Nannf.*
Diaporthales was introduced by Nannfeldt (1932). We
follow the latest treatments and updated accounts of Diaporthales in Maharachchikumbura et al. (2016), Senanayake
et al. (2017), Hyde et al. (2019a), Le Dinh et al. (2019) and
Hyde et al. (2020).
Diaporthaceae Höhn. ex Wehm.
Diaporthaceae was introduced by Höhnel (1917) with
Diaporthe as type genus. The family comprises plant pathogens, endophytes or saprobes on terrestrial plants and rarely
on submerged plants (Udayanga et al. 2011; Dai et al. 2014;
Maharachchikumbura et al. 2016; Gao et al. 2017; Senanayake et al. 2017; Thambugala and Hyde 2018). Senanayake et al. (2018) and Hyde et al. (2020) accepted 15 genera
in the family (Apioporthella, Apiosphaeria, Caudospora,
Chaetoconis, Chiangraiomyces, Diaporthe, Hyaliappendispora, Leucodiaporthe, Mazzantia, Ophiodiaporthe, Paradiaporthe, Phaeocytostroma, Phaeodiaporthe, Pustulomyces,
Stenocarpella). Divergence time estimates for this family
have recently reported for crown age of fossil calibration at
61.15 Mya (29.7–89.9) and secondary calibration at 60.63
Mya (34.8–91.7) in the Paleogene period, and the family
shares the most common ancestor with Cytosporaceae at
87.66 Mya for stem age of fossil calibration and 87.20 Mya
for stem age of secondary calibration, in the Cretaceous
(Guterres et al. 2018).
Diaporthe Nitschke.
Diaporthe was introduced by Nitschke (1870) with D.
eres as the type species. The genus comprises important
plant pathogens, endophytes or saprobes from a wide range
of hosts (Udayanga et al. 2011; Gao et al. 2017; Wanasinghe
et al. 2018; Le Dinh et al. 2019). More than 1000 epithets are
listed in Index Fungorum (2020). Wanasinghe et al. (2018)
introduced six taxa which comprised four new host records
and two new species (D. rosae, D. rosicola) from Rosaceae.
Yang et al. (2018a) introduced two new species D. sambucusii and D. schisandrae as Traditional Chinese Medicines
in Northeast China. Yang et al. (2018b) introduced twelve
new Diaporthe species associated with dieback plant diseases in China. Le Dinh et al. (2019) reported D. asparagi
and D. unshiuensis, said to be pathogenic on Asparagus
kiusianus in Japan. In this study, a new Diaporthe species
is introduced, based on morphology and molecular data,
together with descriptions and illustrations (Fig. 116). A
phylogenetic tree based on combined ITS, TEF1 and TUB2
sequence data is presented in Fig. 115.
Diaporthe chromolaenae Mapook & K.D. Hyde, sp. nov.
13
Fungal Diversity
Fig. 115 Phylogram generated from maximum likelihood
analysis based on combined
dataset of ITS, TEF1 and
TUB2 sequence data. Sixtyfour strains are included in the
combined sequence analysis,
which comprise 2191 characters with gaps. Tree topology
of the ML analysis was similar
to the BYPP. The best scoring
RAxML tree with a final likelihood value of − 17612.889312
is presented. The matrix had
1188 distinct alignment patterns, with 31.17% of undetermined characters or gaps.
Estimated base frequencies
were as follows: A = 0.222777,
C = 0.316430, G = 0.233489,
T = 0.227304; substitution rates:
AC = 1.564927, AG = 4.654146,
AT = 1.821542, CG = 1.281990,
CT = 6.081044, GT = 1.000000;
gamma distribution shape
parameter α = 0.316506. Bootstrap support values for ML
equal to or greater than 60% and
BYPP equal to or greater than
0.90 are given above the nodes.
Newly generated sequences
are in dark red bold and type
species are in bold. Cytospora
sacculus (CFCC 89626) is used
as outgroup taxon
Index Fungorum number: IF557294, Facesoffungi number: FoF 07837; Fig. 116
Etymology: Name reflects the host genus Chromolaena,
from which this species was isolated.
Holotype: MFLU 20-0308
13
Saprobic on dead stems of Chromolaena odorata.
Sexual morph: Ascomata 200–240 µm high × 76–95 µm
diam. ( x̄ = 220 × 88 µm, n = 5), immersed to erumpent,
appearing as raised, black spots, or black necks immerging through the host surface, coriaceous, solitary or scattered, subglobose to obpyriform, pale brown to brown.
Fungal Diversity
Table 25 Synopsis of Patellaria species with similar morphological features discussed in this study
Species
Ascomata (µm)
P. atrata (IMI 32777) 675–1160 × 220–300
P.chromolaenae
100–145 × 415–625
(MFLUCC 17-1479)
Exciple (µm)
Asci (µm)
Ascospores (µm)
References
45–76
(15–)20–30(–35)
98–135 × 15–30
46–60(–70) × 12–16
30–45 × 7–10, 5–11-septate
23–32 × 4–5.5, 3–5-septate
Yacharoen et al. (2015)
This study
Ostiole papillate, without periphyses. Peridium 7.5–15 µm
wide, comprising 2–3 layers, pale brown to brown cells
of textura angularis. Hamathecium with paraphyses not
observed. Asci 30–48 × 7.5–15 µm (x̅ = 41.5 × 11 µm,
n = 25), 8-spored, unitunicate, clavate to subclavate, straight
to slightly curved, sessile, with a J- apical ring. Ascospores
9.5–11.5 × 3.5–4.5 µm ( x̄ = 10.5 × 4 µm, n = 25), overlapping, 1–2-seriate, hyaline, ellipsoidal, 1-septate, constricted
at the septum, straight, guttulate when immature, smoothwalled. Asexual morph: Undetermined.
Culture characteristics: Ascospores germinating on
MEA within 24 h. at room temperature and germ tubes produced from the apex. Colonies on MEA irregular, mycelium slightly raised, entire, white aerial hyphae, spreading
from the center at the surface with white-yellow in reverse
(Fig. 117).
Pre-screening for antimicrobial activity: Not tested.
Material examined: THAILAND, Chiang Rai Province,
Doi Pui, on dead stems of Chromolaena odorata, 5 August
2015, A. Mapook (DP21.2, MFLU 20-0308, holotype); extype culture MFLUCC 17-1422.
GenBank numbers: LSU: MT214362, ITS: MT214456,
SSU: MT214408
Notes: In a BLASTn search of NCBI GenBank, the closest match of the ITS sequence of Diaporthe chromolaenae
(MFLUCC 17-1422, ex-holotype) with 100% similarity
was Phomopsis asparagi strain HB3 (JQ613999) and HB2
(JQ613998). The closest match with the LSU sequence
Fig. 116 Diaporthe chromolaenae (holotype) a Appearance of
ascomata on substrate. b Section through ascoma. c Peridium. d–g Immature and mature
asci. h–m Ascospores. Scale
bars: a = 200 µm, b = 50 µm,
c–g = 20 µm, h–m = 5 µm
13
Fungal Diversity
Niessliaceae, Ophiocordycipitaceae, Sarocladiaceae, Stachybotryaceae, Tilachlidiaceae), based on morphology and
molecular data (Hyde et al. 2020). We also follow the latest
treatment and updated accounts of Trichoderma in Chaverri
et al. (2015), Chen and Zhuang (2017), Qin and Zhuang
(2017), Zhang and Zhuang (2017, 2018), Zhu et al. (2017),
du Plessis et al. (2018) and Phookamsak et al. (2019).
Fig. 117 Culture characteristic on MEA: Diaporthe chromolaenae
(MFLUCC 17-1422)
with 99.00% similarity was Diaporthe actinidiae (strain
KFRD-8, KX609783), while the closest match with the SSU
sequences with 99.64% similarity was D. amygdali (strain
MUCC0101, AB454228). In the present phylogenetic analysis, D. chromolaenae is closely related to D. masirevicii
strain LC6740 which is separate from the type of D. masirevicii strain BRIP 57892a (Fig. 115). However, we could not
compare the morphological characteristics of both strains as
the morphology of Diaporthe masirevicii strain LC6740 has
not been reported. Moreover, morphological comparison of
the related taxa, D. kongii (BRIP 54031) and D. masirevicii
(BRIP 57892a) show that both strains differ in morphology
(Table 26). Therefore, a new species is described here, based
on phylogeny.
Subclass Hypocreomycetidae O.E. Erikss. & Winka
Hypocreales Lindau
Hypocreales was introduced by Lindau (1897a). We follow the latest treatments and updated accounts of Hypocreales in Maharachchikumbura et al. (2016), and Hyde
et al. (2019a, 2020). The order comprises 14 accepted
families (Bionectriaceae, Calcarisporiaceae, Clavicipitaceae, Cocoonihabitaceae, Cordycipitaceae, Flammocladiellaceae, Hypocreaceae, Myrotheciomycetaceae, Nectriaceae,
Hypocreaceae De Not.
Hypocreaceae, introduced by De Notaris (1844), presently comprises 17 genera (Arachnocrea, Dialhypocrea,
Escovopsioides, Escovopsis, Hypocreopsis, Hypomyces,
Kiflimonium, Lichenobarya, Mycogone, Protocrea, Rogersonia, Sepedonium, Sphaerostilbella, Sporophagomyces, Stephanoma, Trichoderma, Verticimonosporium) and
accepted in Hyde et al. (2020). Hyde et al. (2017a) reported
the divergence time estimates for Hypocreomycetidae comprises Coronophorales, Clavicipitales, Falcocladiales, Hypocreales, Microascales and Torpedosporales at 171–241 Mya,
during the mid Triassic to mid Jurassic period.
Trichoderma Pers.
Trichoderma was introduced by Persoon (1794) with T.
viride as the type species. The genus contains many species which are commonly found from soil, plant materials,
human and animal clinical specimens, as well as on macrofungi as hyperparasites (Klein and Eveleigh 1998; Rubini
et al. 2005; Sandoval-Denis et al. 2014). Taxonomy of the
T. harzianum species complex has been revised to include
at least 14 species with nine new species based on multi loci
(Chaverri et al. 2015). Bissett et al. (2015) listed 254 species
with available representative sequences. Subsequently, several taxa have been introduced based on molecular and morphology studies. Qin and Zhuang (2017) introduced seven
new species (T. angustum, T. crystalligenum, T. globoides,
T. perviride, T. purpureum, T. tenue, T. viridulum) in the
Harzianum and Strictipile clades. Chen and Zhuang (2017)
introduced seven new species in the Viride clade, isolated
from soil samples in China. Zhu et al. (2017) introduced two
new species, T. fujianense and T. zonatum with green spores,
while Zhang and Zhuang (2017) introduced four new species with hyaline ascospores from southwest China. Zhang
and Zhuang (2018) introduced three new species, T. acremonioides, T. rugosum and T. subalni, and du Plessis et al.
Table 26 Synopsis of Diaporthe species discussed in this study
Species
Conidiomata (µm)
Conidiophores
(µm)
Conidiogenous
cells (µm)
Alpha conidia (µm) Beta conidia (µm)
References
D. kongii (BRIP
54031)
D. masirevicii
(BRIP 57892a)
Up to 2000 diam.
Up to 6 diam.
6–12 × 1.5–4
Up to 250 diam.
20–40 × 1.5–3.5
10–25 × 1.5–3.0
5.5–7(–
7.5) × 2–2.5(–3)
(5.5–)6–7.5(–
8) × 2–3
Thompson et al.
(2011)
Thompson et al.
(2015)
13
13–23 × 1–1.5
15–30 × 1.0–1.5
Fungal Diversity
Fig. 118 Phylogram generated from maximum likelihood
analysis based on combined
dataset of TEF1 and RPB2
sequence data. Ninety-two
strains are included in the
combined sequence analysis,
which comprise 2450 characters
with gaps. The best scoring
RAxML tree with a final likelihood value of − 15383.443045
is presented. The matrix had
1028 distinct alignment patterns, with 44.41% of undetermined characters or gaps.
Estimated base frequencies
were as follows: A = 0.232835,
C = 0.272110, G = 0.242876,
T = 0.252179; substitution rates:
AC = 0.975283, AG = 3.234057,
AT = 0.990042, CG = 0.931920,
CT = 4.895326, GT = 1.000000;
gamma distribution shape
parameter α = 0.323076. Bootstrap support values for ML
equal to or greater than 60% and
BYPP equal to or greater than
0.90 are given above the nodes.
Newly generated sequences
are in dark red bold and type
species are in bold. Nectria
eustromatica (CBS 121896)
and Nectria berolinensis (CBS
127382) are used as outgroup
taxa
(2018) described five new species (T. beinartii, T. caeruleimontis, T. chetii, T. restrictum, T. undulatum). Phookamsak
et al. (2019) introduced three new species, T. koreanum, T.
pinicola and T. rugulosum from Republic of Korea based on
morphology and phylogeny. We describe a new isolate of T.
guizhouense based on phylogeny, together with a description and illustrations (Fig. 119). A phylogenetic tree based
on combined TEF1 and RPB2 sequence data is presented
in Fig. 118.
Trichoderma guizhouense Q.R. Li, McKenzie & Yong
Wang bis, in Li et al., Mycol. Progr. 12(2): 170 (2012)
[2013]
Facesoffungi number: FoF 07838; Fig. 119
13
Fungal Diversity
Fig. 119 Trichoderma
guizhouense (new host record)
a, b Appearance of ascoma
on substrate. c, d Section
through ascoma. e Peridium. f,
g Asci. h–j Ascospores. Scale
bars: a = 500 µm, b = 200 µm,
c = 100 µm, d, f, g = 20 µm,
e = 10 µm, h–j = 5 µm
Saprobic on dead stems of Chromolaena odorata.
Sexual morph: Stromata 375–555 µm long, 585–645 µm
wide. ( x̄ = 480 × 615 µm, n = 5), superficial, solitary, scattered, coriaceous, olivaceous to dark green. Ascomata
perithecial, 130–165(–185) μm high × 70–120 µm diam.
( x̄ = 155 × 90 µm, n = 10), arranged in rows, clustered, gregarious, with 5–6 perithecia forming groups immersed in
stroma, irregular shape. Ostiole central. Peridium 5–10 µm
wide, 4–5 layers, dense, comprising thick-walled, hyaline
to pale brown cells of textura epidermoidea. Hamathecium
with paraphyses not observed. Asci 50–80 × 3.5–5.5 µm
( x̄ = 65 × 4.5 µm, n = 15), appears to contain 16
13
part-ascospores, unitunicate, cylindrical, short pedicellate,
straight or slightly curved, with inconspicuous apical ring.
Ascospores 3–6.5 × 2.5–5.5 µm ( x̄ = 4.2 × 4 µm, n = 40),
uniseriate, hyaline when immature, becoming unicellular,
pale green to green when mature, globose or subglobose to
oblong, verruculose with small guttule. Asexual morph:
Undetermined.
Culture characteristics: Ascospores germinating on
MEA within 24 h at room temperature and germ tubes
produced from both ends. Colonies on MEA, filamentous,
mycelium slightly raised, velvety with fluffy, white aerial
Fungal Diversity
Fig. 120 Culture characteristic on MEA: Trichoderma guizhouense
(MFLUCC 17-1512)
hyphae at the surface, spreading from the center with white
in reverse, filiform margin (Fig. 120).
Pre-screening for antimicrobial activity: Trichoderma
guizhouense (MFLUCC 17-1512) showed antimicrobial
activity against E. coli with a 19 mm inhibition zone and
against M. plumbeus with an 18 mm inhibition zone, observable as partial inhibition, when compared to the positive
control (9 mm and 17 mm, respectively), but no inhibition
of B. subtilis.
Known hosts and distribution: Soil in China (Li et al.
2013); endophyte in stems of Ancistrocladus korupensis
(Ancistrocladaceae) and Cola spp. (Malvaceae) in Cameroon
(Chaverri et al. 2015).
Material examined: THAILAND, Phrae Province, Doi
Pha Klong, on dead stems of Chromolaena odorata, 22 September 2016, A. Mapook (DPKP6, MFLU 20-0381); living
culture MFLUCC 17-1512 (new host record).
GenBank numbers: LSU: MT214480, ITS: MT214386,
SSU: MT214431, TEF1: MT235803
Notes: A phylogenetic analysis shows that the strain
MFLUCC 17-1512 grouped within Trichoderma
guizhouense (Fig. 118). A BLASTn search of the TEF1
sequence showed that the strain is identical to T. pollinicola
with 99.78% similarity while the closest match of the RPB2
sequence with 95.29% similarity was T. lixii (strain GJS
00-18, FJ442750). We therefore, identify our isolates as T.
guizhouense based on phylogenetic analyses and introduced
here as a new host record from Chromolaena odorata collected in Thailand. In this study, the strain was found as the
sexual morph in nature and we could not obtain its asexual
morph in culture.
Stachybotryaceae L. Lombard & Crous
Stachybotryaceae was introduced by Crous et al. (2014)
to accommodate three genera Myrothecium, Peethambara
and Stachybotrys (type genus), which were earlier classified
in the order Hypocreales genera incertae sedis. Subsequently, Lombard et al. (2016) resolved 33 genera in the
family including nine known genera (Albosynnema, Alfaria,
Didymostilbe, Myrothecium, Parasarcopodium, Peethambara, Septomyrothecium, Stachybotrys and Xepicula), and
three reintroduced genera (Melanopsamma, Memnoniella,
Virgatospora). In addition, 13 genera were newly described
for those with myrothecium-like morphology (Albifimbria,
Capitofimbria, Dimorphiseta, Gregatothecium, Inaequalispora, Myxospora, Neomyrothecium, Paramyrothecium,
Parvothecium, Smaragdiniseta, Striaticonidium, Tangerinosporium, Xenomyrothecium) and eight new genera for
those with stachybotrys-like morphology (Achroiostachys,
Brevistachys, Cymostachys, Globobotrys, Grandibotrys,
Kastanostachys, Sirastachys, Striatibotrys). Wijayawardene
et al. (2018) accepted 36 genera in the family including three
additional genera (Alfariacladiella, Koorchalomella, Xepiculopsis). Hyde et al. (2017a) recommended changes within
the Sordariomycetes based on molecular clock evidence
showing that Stachybotryaceae grouped with Nectriaceae
and Niessliaceae which diverged from Clavicipitales at 157
Mya and the family was estimated in late Cretaceous for
stem age at 88 Mya (50–130).
Memnoniella Höhn.
Memnoniella was introduced by Höhnel (1924) and has
been considered as a synonym of Stachybotrys (Wang et al.
2015). However, Memnoniella was reintroduced based on
morphology, with M. echinata (CBS 216.32) designated as
ex-epitype strain, together with multigene analyses using
cmdA, ITS, RPB2, TEF1 and TUB2 (Lombard et al. 2016).
Nineteen epithets are listed in Index Fungorum (2020), with
most species found as saprobic on decaying plant material.
In addition, environmental samples (e.g. soil and air), as well
as from indoor habitats, have also been reported, together
with multigene sequence available in GenBank (Haugland
and Heckman 1998; Lin et al. 2016; Lombard et al. 2016).
We introduce a new species, Memnoniella chromolaenae
based on morphological comparison (Table 27) together
with a description and illustrations (Fig. 122). A phylogenetic tree based on combined LSU, ITS, TEF1, RPB2 and
TUB2 sequence data is presented in Fig. 121.
Memnoniella chromolaenae Mapook & K.D. Hyde, sp. nov.
Index Fungorum number: IF557292, Facesoffungi number: FoF 07839; Fig. 122
Etymology: Name reflects the host genus Chromolaena,
from which this species was isolated.
Holotype:***
Saprobic on dead stems of Chromolaena odorata. Sexual morph: Undetermined. Asexual morph: Hyphomycetous. Colonies effuse on host, dark brown to black, powdery. Mycelium partly immersed to superficial on the host
13
Fungal Diversity
Table 27 Synopsis of Memnoniella species with similar morphological features discussed in this study
Species
Conidiophores (µm)
Conidiogenous cells (µm)
Catenate conidia (µm)
References
M. chromolaenae (MFLUCC 17-1507)
M. longistipitata (ATCC 22699)
40–85 × 2.5–4
(170–)260–460(–
610) × (3.2–)3.6–4.7(–
4.9)
85–180 × 6.5–12
5.5–7.5 × 3–4.5
9.7–10.2 × 4.7–5.5
3–4.5 × 3.5–4.8
5.8–8.5 × 6.3–8.3
This study
Li et al. (2003)
9–14 × 4–6.4
8.5–12 × 4.5–7
Lin et al. (2016)
M. oblongispora (MFLUCC 15-1074)
surface, comprising septate, smooth, and hyaline hyphae.
Conidiophores 40–85 × 2.5–4 µm ( x̄ = 56 × 3 µm, n = 10),
wider at the base, macronematous, mononematous, solitary or in groups, arising from hypha, erect, unbranched,
hyaline at the base becoming dark grey to black towards
the apex, 2–3-septate, smooth to minutely verrucose, thickwalled, bearing 6 conidiogenous cells. Conidiogenous cells
5.5–7.5 × 3–4.5 µm ( x̄ = 6.5 × 3.8 µm, n = 15), phialidic,
Fig. 121 Phylogram generated from maximum likelihood
analysis based on combined
dataset of LSU, ITS, TEF1,
RPB2, TUB2 and cmdA
sequence data. Twenty-nine
strains are included in the
combined sequence analysis,
which comprise 3807 characters with gaps. Tree topology
of the ML analysis was similar
to the BYPP. The best scoring
RAxML tree with a final likelihood value of − 14893.412643
is presented. The matrix had
1078 distinct alignment patterns, with 22.33% of undetermined characters or gaps.
Estimated base frequencies
were as follows: A = 0.235371,
C = 0.268803, G = 0.268291,
T = 0.227535; substitution rates:
AC = 1.011907, AG = 3.371254,
AT = 1.029380, CG = 0.908817,
CT = 7.410720, GT = 1.000000;
gamma distribution shape
parameter α = 0.174666. Bootstrap support values for ML
equal to or greater than 60% and
BYPP equal to or greater than
0.90 are given above the nodes.
Newly generated sequences are
in dark red bold and type species are in bold. Peethambara
sundara (CBS 646.77) is used
as outgroup taxon
13
discrete, clustered at the apex of conidiophores, ellipsoid or
obovoid to clavate or reniform, smooth, subhyaline. Conidia
3–4.5 × 3.5–4.8 µm ( x̄ = 3.5 × 4.5 µm, n = 30), catenate,
acrogenous, globose to subglobose, aseptate, olivaceousgrey to black, smooth, thick-walled, formed in long chains.
Culture characteristics: Conidia germinating on MEA
within 24 h at room temperature and germ tubes produced
from both ends. Colonies on MEA circular, mycelium
Fungal Diversity
crateriform, moderately fluffy, entire to filamentous, cultures white at the surface and pale brown to creamy-white
in reverse appearing as concentric ring pattern with creamywhite at the margin (Fig. 123).
Pre-screening for antimicrobial activity: Memnoniella
chromolaenae (MFLUCC 17-1507) showed antimicrobial
activity against E. coli with an 8 mm inhibition zone when
compared to the positive control (9 mm), but no inhibition
of B. subtilis and M. plumbeus.
Material examined: THAILAND, Chiang Mai Province,
Fah Hom Pok, on dead stems of Chromolaena odorata, 27
September 2016, A. Mapook (FHP11, MFLU 20-0321,
holotype); ex-type culture MFLUCC 17-1507.
GenBank numbers: LSU: MT214465, ITS: MT214371,
SSU: MT214418
Notes: In a BLASTn search of NCBI GenBank, the
closest match of the ITS sequence of Memnoniella chromolaenae (MFLUCC 17-1507, ex-holotype) was Mem.
longistipitata with 99.83% similarity to the strain CBS
136197 (KU846155). The closest match with the LSU
sequence with 99.55% similarity was Mem. echinata (strain
CBS 216.32, MH866746), while the closest match with
the SSU sequence with 99.91% similarity was Stachybotrys echinata (strain UAMH6594, AY489704). In the present phylogenetic analysis, Mem. chromolaenae is closely
related to Mem. longistipitata and Mem. oblongispora
(Fig. 121). A comparison of the ITS (+5.8S) gene region of
Mem. chromolaenae and Mem. longistipitata reveals four
base pair differences (0.73%) across 546 nucleotides, but
we could not compare TEF1 gene region of those closely
related taxa due to the lack of the TEF1 sequence. However, Mem. chromolaenae differs from Mem. longistipitata
and Mem. oblongispora in having shorter conidiophore
(40–85 × 2.5–4 µm vs. (170–)260–460(–610) × (3.2–)3.6–4.
7(–4.9) µm and 85–180 × 6.5–12 µm), smaller conidiogenous
cells (5.5–7.5 × 3–4.5 µm vs. 9.7–10.2 × 4.7–5.5 µm and
9–14 × 4–6.4 µm) and smaller conidia (3–4.5 × 3.5–4.8 µm
vs. 5.8–8.5 × 6.3–8.3 µm and 8.5–12 × 4.5–7 µm) (Table 27).
Therefore, Mem. chromolaenae is described as a new species
based on morphological comparison.
Subclass Xylariomycetidae O.E. Erikss & Winka
Amphisphaeriales D. Hawksw. & O.E. Erikss.*
Amphisphaeriales was introduced by Eriksson and Hawksworth (1986). We follow the latest treatment and updated
accounts of Amphisphaeriales in Hyde et al. (2020) with
14 accepted families (Amphisphaeriaceae, Apiosporaceae,
Beltraniaceae, Clypeophysalosporaceae, Cylindriaceae,
Hansfordiaceae, Hyponectriaceae, Iodosphaeriaceae, Melogrammataceae, Phlogicylindriaceae, Pseudomassariaceae,
Fig. 122 Memnoniella chromolaenae (holotype) a, b Appearance of colonies on substrate.
c-f Conidiophores, conidiogenous cells and conidia. g–h
Conidia in chains. i Conidium.
j Germinated conidia. Scale
bars: a = 500 µm, b = 200 µm, c,
d, g, j = 20 µm, e–f, h = 10 µm,
i = 5 µm
13
Fungal Diversity
Fig. 123 Culture characteristic on MEA: Memnoniella chromolaenae (MFLUCC 17-1507)
Pseudotruncatellaceae, Sporocadaceae, Vialaeaceae).
Furthermore, we follow the latest treatments and updated
accounts of the genus Arthrinium in Crous and Groenewald
(2013), Wang et al. (2018), Jiang et al. (2018), Pintos et al.
(2019) and Hyde et al. (2020).
Apiosporaceae K.D. Hyde et al.
Apiosporaceae was introduced by Hyde et al. (1998) to
accommodate the genera Appendicospora and Apiospora,
which had been earlier assigned to Lasiosphaeriaceae by
Barr (1990) based on morphology, with Apiospora as the
type genus. Crous and Groenewald (2013) synonymized
Apiospora, Pteroconium and Cordella under the genus
Arthrinium. Six genera, Appendicospora, Arthrinium, Dictyoarthrinium, Endocalyx, Scyphospora and Spegazzinia
were previously accepted in Outline of Ascomycota 2017
(Wijayawardene et al. 2018). However, Tanaka et al. (2015)
and Jayasiri et al. (2019) placed Spegazzinia in Didymosphaeriaceae based on molecular data. Scyphospora was synonymized under the genus Arthrinium (Nag Raj 1974; Kirk
1986; Hyde et al. 2020), and Nigrospora was transferred to
Apiosporaceae based on morphology and phylogeny (Wang
et al. 2017). Recently, Hyde et al. (2020) placed Appendicospora, Arthrinium, Dictyoarthrinium, Endocalyx and Nigrospora in the family. Divergence time estimates for the family
have been reported based on molecular clock evidence. The
family was estimated for stem age at 69 Mya (50–130) in the
late Cretaceous (Hyde et al. 2017a).
Arthrinium Kunze, in Kunze & Schmidt, Mykologische
Hefte (Leipzig) 1: 9 (1817)
Arthrinium is an ecologically diverse genus found on
various substrates and associated with various plants as
endophytes, saprobes, and plant pathogens on some important ornamentals (Chen et al. 2014; Li et al. 2016), and also
causes cutaneous infections in humans (Rai 1989; Zhao et al.
13
1990; de Hoog et al. 2000; Crous et al. 2013; Wang et al.
2018). The genus was introduced by Schmidt and Kunze
(1817) with A. caricicola as the type species. Crous and
Groenewald (2013) described eight new species (A. hydei, A.
kogelbergense, A. malaysianum, A. ovatum, A. phragmites,
A. pseudospegazzinii, A. pseudosinense, A. xenocordella)
based on phylogeny and morphology. Wang et al. (2018)
introduced eight new species (A. bambusae, A. camelliaesinensis, A. dichotomanthi, A. guizhouense, A. jiangxiense,
A. obovatum, A. pseudoparenchymaticum, A. subroseum)
from China and reported the major host plant families of
Arthrinium species were Poaceae and Cyperaceae. Jiang
et al. (2018) introduced two new species, A. qinlingense
and A. gaoyouense from China. Subsequently, Pintos et al.
(2019) introduced six new species (A. balearicum, A. descalsii, A. esporlense, A. ibericum, A. italicum, A. piptatheri)
from Europe. We introduce a new species Arthrinium chromolaenae based on phylogeny and morphological comparison (Table 28) together with a description and illustrations (Fig. 125). A phylogenetic tree based on combined
ITS, LSU, TEF1, and TUB2 sequence data is presented in
Fig. 124.
Arthrinium chromolaenae Mapook & K.D. Hyde, sp. nov.
Index Fungorum number: IF557288, Facesoffungi number: FoF 07840; Fig. 125
Etymology: Name reflects the host genus Chromolaena,
from which this species was isolated.
Holotype: MFLU 20-0300
Saprobic on dead stems of Chromolaena odorata. Sexual morph: Undetermined. Asexual morph: Colonies
on host surface, black, powdery. Mycelium consisting of
smooth, hyaline, branched, septate, hyphae. Conidiophores
reduced to conidiogenous cells, grouped together to form
sporodochia. Conidiogenous cells basauxic, 6.5–12 µm
high × 1–2 µm diam. ( x̄ = 8.5 × 1.5 µm, n = 5), aggregated in clusters on hyphae, hyaline, smooth, elongated,
broadly filiform to ampulliform. Conidia 4–6 × 4.5–6.5 µm
( x̄ = 5 × 5.5 µm, n = 35), in surface view, irregular arrangement in pseudo-chains, pale brown to brown, smooth, globose to subglobose, lenticular in side view, with pale equatorial slit.
Culture characteristics: Conidia germinating on MEA
within 24 h at room temperature, germ tubes produced from
conidia. Colonies on MEA circular, white, flat, spreading,
with fluffy aerial mycelium, filiform, mycelia not tightly
attached to the surface (Fig. 126).
Pre-screening for antimicrobial activity: Arthrinium
chromolaenae (MFLUCC 17-1505) showed antimicrobial
activity against E. coli with an 8 mm inhibition zone when
compared to the positive control (9 mm), but no inhibition
of B. subtilis and M. plumbeus.
Fungal Diversity
Table 28 Synopsis of Arthrinium species with similar morphological features discussed in this study
Species
Conidiophores (µm)
Conidiogenous cells (µm)
Conidia (µm)
References
A. chromolaenae (MFLUCC
17-1505)
A. euphorbiae (IMI 110788)
–
6.5–12 × 1–2
4–6 × 4.5–6.5
This study
–
–
A. italicum (CBS 145138)
A. thailandicum (MFLUCC
15–0202)
A. vietnamense (IMI
99670= A. malaysianum)
10–50 × 1–3
5.5–11 × 3–4.5
(3–)4–7(–9) × (1.5–)2–3(–5)
11.5–39 × 2–3.5
(4–)4.7(–5.5) in surface view, Ellis (1965), Crous et al.
(2013)
(3–)3.2(–4) in side view
4–6 × 3–4
Pintos et al. (2019)
5–9 × 5–8
Dai et al. (2017)
–
4–7 × 3–5
Material examined: THAILAND, Chiang Mai Province,
Fah hom pok, on dead stems of Chromolaena odorata, 27
September 2016, A. Mapook (FHP9, MFLU 20-0300, holotype); ex-type culture MFLUCC 17-1505.
GenBank numbers: LSU: MT214436, ITS: MT214342,
TEF1: MT235802
Notes: In a BLASTn search of NCBI GenBank, the closest
match of the ITS sequence of A. chromolaenae (MFLUCC
17-1505, ex-holotype) was A. malaysianum with 97.38%
similarity to the strain CBS 251.29 (KF144897). The closest
match with the LSU sequence with 100% similarity was A.
malaysianum (strain CBS 102053, NG_042780), while the
closest match with the TEF sequence with 99.06% similarity
was Arthrinium sp. (strain HKU41, LC439290). Crous et al.
(2013) introduced A. malaysianum as a new species based
on phylogeny and morphologically comparing with a closely
related strain, A. euphorbiae. They mentioned that A. malaysianum differs from A. euphorbiae in having slightly longer
conidia. Wang et al. (2017) demonstrated that the ex-type
strain of A. vietnamensis and A. malaysianum are conspecific, supported by morphological comparison and proposed
use of the older name, A. vietnamensis. Phylogenetic analysis showed that A. chromolaenae is closely related to the
clade comprising, A. euphorbiae and A. vietnamense (= A.
malaysianum) (Fig. 124). However, A. chromolaenae differs
from A. vietnamense in having long and narrow conidiogenous cells (6.5–12 × 1–2 µm vs. 4–7 × 3–5 µm) (Table 28).
A comparison of the ITS (+ 5.8S) gene region of A. chromolaenae and A. vietnamense reveals 16 base pair differences
(2.62%) across 610 nucleotides. Therefore, A. chromolaenae
is described as a new species based on phylogeny and morphological comparison.
Xylariales Nannf.
Xylariales was introduced by Nannfeldt (1932). We follow the latest treatment and updated accounts of Xylariales in Hyde et al. (2020) with 15 accepted families
(Barrmaeliaceae, Cainiaceae, Clypeosphaeriaceae, Coniocessiaceae, Diatrypaceae, Graphostromataceae, Hansfordiaceae, Hypoxylaceae, Induratiaceae, Lopadostomataceae,
5–6 in surface view, 3–4 in
side view
Crous et al. (2013), Wang
et al. (2017)
Microdochiaceae, Polystigmataceae, Requienellaceae,
Xylariaceae, Zygosporiaceae). Furthermore, we follow
the latest treatments and updated accounts of the family
Cainiaceae in Liu et al. (2015), Maharachchikumbura et al.
(2015, 2016), Senanayake et al. (2015), Wijayawardene et al.
(2017a, 2018) and Hyde et al. (2020).
Cainiaceae J.C. Krug
Cainiaceae was introduced by Krug (1977) to accommodate the generic type, Cainia, which is characterized by
unique apical rings in the asci and longitudinal germ slit in
the ascospores. Kang et al. (1999) revived the family and
accepted five genera, Atrotorquata, Arecophila, Cainia,
Ceriophora and Reticulosphaeria based on morphology.
Subsequently, several molecular and morphological studies have confirmed the placement of these genera with five
accepted genera in Cainiaceae (Amphibambusa, Arecophila, Atrotorquata, Cainia, Seynesia) (Jeewon et al. 2003;
Smith et al. 2003; Liu et al. 2015; Maharachchikumbura
et al. 2015, 2016; Senanayake et al. 2015; Wijayawardene
et al. 2017a, 2018), while Reticulosphaeria was placed in
Amphisphaeriales genera incertae sedis (Senanayake et al.
2015). The family has been placed within the Xylariales
(Maharachchikumbura et al. 2015, 2016; Senanayake et al.
2015; Jaklitsch et al. 2016), while divergence time estimates
suggested that the family share the most common ancestor
with Xylariales, with a divergence time of ca 128 Mya (Hyde
et al. 2017a). Hyde et al. (2020) introduced a new genus
Alishanica from Taiwan in the family and placed six genera, Alishanica, Amphibambusa, Arecophila, Atrotorquata,
Cainia and Seynesia in Cainiaceae within Xylariales.
Longiappendispora Mapook & K.D. Hyde, gen. nov.
Index Fungorum number: IF557336, Facesoffungi number: FoF 07841
Etymology: The generic epithet reflects the ascospores
with long bristle-like polar appendages.
Saprobic on dead stems. Sexual morph: Ascomata
immersed beneath clypeus, unilocular, globose to subglobose, coriaceous, solitary or scattered, or sometimes
13
Fungal Diversity
Fig. 124 Phylogram generated from maximum likelihood
analysis based on combined
dataset of ITS, LSU, TEF1, and
TUB2 sequence data. Eightysix strains are included in the
combined sequence analysis,
which comprise 3780 characters
with gaps. The best scoring
RAxML tree with a final likelihood value of − 25718.058495
is presented. The matrix had
1793 distinct alignment patterns, with 39.70% of undetermined characters or gaps.
Estimated base frequencies
were as follows: A = 0.240172,
C = 0.253238, G = 0.253613,
T = 0.252978; substitution rates:
AC = 1.178756, AG = 2.769752,
AT = 1.160953, CG = 0.997632,
CT = 4.711267, GT = 1.000000;
gamma distribution shape
parameter α = 0.246871. Bootstrap support values for ML
equal to or greater than 60%
and BYPP equal to or greater
than 0.90 are given at the nodes.
Newly generated sequences
are in dark red bold and type
species are in bold. Seiridium
phylicae (CPC 19965) is used
as outgroup taxon
13
Fungal Diversity
Fig. 125 Arthrinium chromolaenae (holotype) a, b Appearance of colonies on substrate.
c–g Conidia and conidiogenous
cells. h–k Conidia. Scale
bars: a = 500 µm, b = 100 µm,
h = 20 µm, c–g, i–k = 5 µm
Fig. 126 Culture characteristic on MEA: Arthrinium chromolaenae
(MFLUCC 17-1505)
gregarious. Ostiole central. Peridium comprising 2–3 layers,
pale brown to hyaline cells of textura angularis. Hamathecium composed of cylindrical to broadly filiform, septate,
guttulate, hyaline paraphyses, embedded in a gelatinous
matrix. Asci 8-spored, unitunicate, cylindrical to broadly
filiform, short-pedicellate, straight or slightly curved, with
an apical ring, apically rounded. Ascospores uniseriate, hyaline when immature, becoming brown to dark brown when
mature, 1-septate, ellipsoid to broadly fusiform, tapering
towards narrow ends, constricted at the septum, straight or
slightly curved, guttulate at both cells, with longitudinal
striations and bristle-like polar appendages from both ends,
without a gelatinous sheath. Asexual morph: Undetermined.
Type species: Longiappendispora chromolaenae Mapook
& K.D. Hyde
Notes: A phylogenetic analysis based on combined
dataset of the ITS and LSU sequence data show that
Longiappendispora chromolaenae groups with a new
genus, Alishanica miscanthii (FU 31025), with low bootstrap support in Cainiaceae and clusters with Cainia
clade (0.93 BYPP, Fig. 127). However, Longiappendispora species differ from Alishanica by larger ascomata
(500–580 × 360–505 µm vs. 277–272 × 288–285 µm), a thin
peridium wall [(15–)20–30 µm vs. 52–60 µm], narrower asci
(140–230 × 13–20 µm vs. 173–179 × 23–31 µm) and smaller
ascospores (28.5–43 × 9.5–12 µm vs. 59–62 × 19–21 µm)
with bristle-like polar appendages on both ends of the
ascospores and without a gelatinous sheath (Table 29).
Therefore, we introduce Longiappendispora as a new genus
with a new species L. chromolaenae, based on morphological comparison with phylogenetic analyses. A comparison
of the ITS (+5.8S) gene region of L. chromolaenae and Alishanica miscanthii reveals 78 base pair differences (17.9%)
across 435 nucleotides.
Longiappendispora chromolaenae Mapook & K.D. Hyde,
sp. nov.
Index Fungorum number: IF557337, Facesoffungi number: FoF 07842; Fig. 128
13
Fungal Diversity
Fig. 127 Phylogram generated
from maximum likelihood analysis based on combined dataset
of ITS and LSU sequence data.
Fourteen strains are included in
the combined sequence analysis,
which comprise 1506 characters
with gaps. The best scoring
RAxML tree with a final likelihood value of − 4717.509202
is presented. The matrix had
329 distinct alignment patterns, with 22.49% of undetermined characters or gaps.
Estimated base frequencies
were as follows: A = 0.255033,
C = 0.233739, G = 0.280548,
T = 0.230680; substitution rates:
AC = 2.006227, AG = 2.877479,
AT = 3.211990, CG = 2.743197,
CT = 8.119404, GT = 1.000000;
gamma distribution shape
parameter α = 0.202825. Bootstrap support values for ML
equal to or greater than 60% and
BYPP equal to or greater than
0.90 are given above the nodes.
Newly generated sequences are
in dark red bold and type species are in bold. Entosordaria
perfidiosa (CBS 142773) is
used as outgroup taxon
Table 29 Morphological features of species in Cainiaceae discussed in this study
Species
Asocomata (µm)
Peridium (µm) Asci (µm)
Longiappendispora chromolaenae (MFLUCC
17-1485)
500–580 high × 360–
505 diam.
(15–)20–30
Alishanica miscanthii 277–272 high × 288–
(FU 31025)
285 diam.
13
52–60
Ascospores (µm) Morphology of
ascospores
References
This study
140–230 × 13–20 28.5–43 × 9.5–12 With longitudinal
striations and
bristle-like polar
appendages from
both ends, without
gelatinous sheath
Hyde et al. (2020)
173–179 × 23–31 59–62 × 19–21
With striations, surrounded by a thick,
hyaline mucilaginous sheath sub
globose, parallel to
the margin of the
spore
Fungal Diversity
Fig. 128 Longiappendispora
chromolaenae (holotype) a,
b Appearance of ascomata on
substrate. c Section through
ascoma. d Ostiole. e Peridium.
f Pseudoparaphyses. g–h Asci.
i–o Ascospores. Scale bars:
a = 500 µm, b, c = 200 µm, d,
g–i = 50 µm, j–o = 20 µm, e,
f = 5 µm
Etymology: Name reflects the host genus Chromolaena,
from which this species was isolated.
Holotype: MFLU 20-0320
Saprobic on dead stems of Chromolaena odorata. Sexual morph: Ascomata 500–580 µm high × 360–505 µm
diam. ( x̄ = 535 × 440 µm, n = 5), immersed beneath clypeus,
unilocular, globose to subglobose, coriaceous, solitary or
scattered, sometimes gregarious. Ostiole central. Peridium
(15–)20–30 µm wide, comprising 2–3 layers, pale brown
to hyaline cells of textura angularis. Hamathecium comprising 2–4 µm wide, cylindrical to broadly filiform, septate, guttulate, hyaline paraphyses, embedded in a gelatinous matrix. Asci 140–230 × 13–20 µm ( x̄ = 195 × 15.5 µm,
n = 15), 8-spored, unitunicate, cylindrical to broadly filiform,
short pedicellate, straight or slightly curved, with an apical
ring, apically rounded. Ascospores 28.5–43 × 9.5–12 µm
( x̄ = 33 × 11 µm, n = 30), uniseriate, hyaline when immature, becoming brown to dark brown when mature, 1-septate,
13
Fungal Diversity
name and fungal classification are used according to Index
Fungorum (2020), outline of Ascomycota (Wijayawardene
et al. 2018), Notes for genera: Ascomycota (Wijayawardene
et al. 2017b) and recent relevant literature.
Phylum Ascomycota Caval.-Sm.
Class Dothideomycetes sensu O.E. Erikss & Winka
Subclass Dothideomycetidae P.M. Kirk et al.
Capnodiales Woron.
Capnodiaceae (Sacc.) Höhn. ex Theiss.
1. Capnodium sp.
Venezuela (Urtiaga 1986)
Fig. 129 Culture characteristic on MEA: Longiappendispora chromolaenae (MFLUCC 17-1485)
ellipsoid to broadly fusiform, tapering towards narrow ends,
constricted at the septum, straight or slightly curved, guttulate at both cells, with longitudinal striations and bristlelike polar appendages from both ends, without a gelatinous
sheath. Asexual morph: Undetermined.
Culture characteristics: Ascospores germinating on
MEA within 24 h. at room temperature and germ tubes produced from both ends. Colonies on MEA circular, mycelium raised and velvety with moderately fluffy, filamentous,
white aerial hyphae at the surface and orange in reverse with
white-yellow at the margin (Fig. 129).
Pre-screening for antimicrobial activity: Longiappendispora chromolaenae (MFLUCC 17-1485) showed
antimicrobial activity against E. coli with a 9 mm inhibition
zone when compared to the positive control (9 mm), but no
inhibition of B. subtilis and M. plumbeus.
Material examined: THAILAND, Chiang Rai Province,
Doi Pui, on dead stems of Chromolaena odorata, 2 February 2017, A. Mapook (DP83, MFLU 20-0320, holotype);
ex-type culture MFLUCC 17-1485.
GenBank numbers: LSU: MT214464, ITS: MT214370,
SSU: MT214417
Notes: In a BLASTn search of NCBI GenBank, the closest
match of the ITS sequence of Longiappendispora chromolaenae (MFLUCC 17-1485, ex-holotype) with 84.88% similarity
was Cainia desmazieri (strain CBS 137.62, MH858124), while
the closest match with the LSU sequence with 96.99% similarity was Cainia graminis (strain CBS 136.62, AF431949).
Checklist of fungi associated with Chromolaena odorata
(= Eupatorium odoratum)
The checklist of fungi associated with C. odorata is based
on the USDA Systematic Mycology and Microbiology Laboratory (SMML) database (Farr and Rossman 2020), together
with relevant literature and the author’s studies. The current
13
Cladosporiaceae Nann.
2. Cladosporium sp.
West Indies (Minter et al. 2001)
3. Cladosporium eupatorii Cif., Sydowia 8(1–6): 251
(1954) (= Hormodendrum eupatorii)
Dominican Republic (Ciferri 1961; Barreto and Evans
1994)
Mycosphaerellaceae Lindau
4.
Cercospora sp.
Cambodia (Litzenberger et al. 1994), Malaysia
(Johnston 1960; Williams and Liu 1976), Nepal (Barreto and Evans 1994), Thailand (Puckdeedindan 1966;
Nguanhom et al. 2015), Venezuela (Urtiaga 2004)
5. Micronematomyces caribensis (Crous & den Breeÿen)
U. Braun, C. Nakash., Videira & Crous, in Videira
et al., Stud. Mycol. 87: 337 (2017)
Cuba, Jamaica (den Breeyen et al. 2006; Videira
et al. 2017)
6. Micronematomyces chromolaenae (Crous & den
Breeÿen) U. Braun, C. Nakash., Videira & Crous, in
Videira et al. Stud. Mycol. 87: 337 (2017)
Mexico (den Breeyen et al. 2006; Videira et al.
2017)
7. Passalora capsicicola (Vassiljevsky) U. Braun & F.O.
Freire, Cryptog. Mycol. 23(4): 299 (2003) [2002]
= Phaeoramularia capsicicola (Vassiljevsky)
Deighton, in Ellis, Trans. Br. mycol. Soc. 67(1): 140
(1976)
Jamaica, USA (Videira et al. 2017)
8. Passalora sp.
Jamaica, USA (den Breeyen et al. 2006)
9. Phloeospora sp.
Venezuela (Urtiaga 1986; Barreto and Evans 1994)
10. Pseudocercospora aciculina (Chupp) U. Braun &
Crous, in Crous & Braun, CBS Diversity Ser. (Utrecht)
1: 42 (2003)
Fungal Diversity
11.
12.
13.
14.
15.
16.
17.
= Cercospora aciculina Chupp, Monograph of Cercospora: 118 (1954)
Cambodia (Litzenberger el al. 1962), Nigeria (Fajola
1978; Barreto and Evans 1994)
Pseudocercospora ageratoides (Ellis & Everh.) Y.L.
Guo, Acta Mycol. Sin. 12(4): 271 (1993)
= Cercospora ageratoides Ellis & Everh., J. Mycol.
5(2): 71 (1889)
Cote d’Ivoire (Yen 1974)
Pseudocercospora convoluta (Crous & den Breeÿen)
U. Braun, C. Nakash., Videira & Crous, in Videira
et al., Stud. Mycol. 87: 312 (2017)
= Passalora convoluta Crous & den Breeÿen,
in Breeÿen, Groenewald, Verkley & Crous, Fungal
Divers. 23: 96 (2006)
Costa Rica (den Breeyen et al. 2006; Videira et al.
2017)
Pseudocercospora eupatoriella Crous & den Breeÿen,
in Breeÿen, Groenewald, Verkley & Crous, Fungal
Divers. 23: 101 (2006)
Jamaica (den Breeyen et al. 2006; Guatimosim et al.
2016), USA (den Breeyen et al. 2006)
Pseudocercospora eupatorii (Peck) U. Braun & R.F.
Castañeda, Cryptog. bot. 2(2-3): 293 (1991)
= Cercospora eupatorii Peck, Ann. Rep. N.Y. St.
Mus. nat. Hist. 33: 29 (1883) [1880]
Cambodia, Cote d’Ivoire, Cuba, Hawaii, India,
Nepal, North America (Govindu et al. 1970; Yen
1974; Urtiaga 1986; Barreto and Evans 1994), Andaman Islands (Hosagoudar and Mathew 2000), India
(Kamal 2010), Thailand (Meeboon et al. 2007)
Pseudocercospora eupatorii-formosani U. Braun &
Bagyan., Sydowia 51(1): 8 (1999)
Bangladesh (Barreto and Evans 1994), Brunei
Darussalam (Peregrine and Ahmad 1982; Barreto
and Evans 1994; Braun and Sivapalan 1999), China
(Roux et al. 1997; Liu and Guo 1998; Guo 1999a, b;
Zhuang 2001), India (Barreto and Evans 1994), Laos
(Phengsintham et al. 2010), Malaysia (Barreto and
Evans 1994), Myanmar (Thaung 1984; Barreto and
Evans 1994), Taiwan (Kirschner and Chen 2007),
Thailand (Barreto and Evans 1994; Phengsintham et al.
2013), Venezuela (Urtiaga and Braun 2013)
Pseudocercospora sp.
Guatemala (Videira et al. 2017), Mexico (den
Breeyen et al. 2006)
Ragnhildiana perfoliati (Ellis & Everh.) U. Braun,
C. Nakash., Videira & Crous, in Videira, Groenewald,
Nakashima, Braun, Barreto, de Wit & Crous, Studies
in Mycology 87: 345 (2017)
= Mycovellosiella perfoliati (Ellis & Everh.) U.
Braun, C. Nakash., Videira & Crous 1960
18.
19.
20.
21.
22.
23.
24.
25.
= Passalora perfoliati (Ellis & Everh.) U. Braun &
Crous, in Crous & Braun, CBS Diversity Ser. (Utrecht)
1: 314 (2003)
Brazil (Barreto and Evans 1994), Laos (Videira et al.
2017), West Indies (Minter et al. 2001), Jamaica (den
Breeyen et al. 2006), Taiwan (Kirschner and Chen
2007)
Ramularia eupatorii J.M. Yen, Cahiers Pacif. 13: 272
(1969) (= Mycosphaerella eupatorii)
Malaysia (Yen 1969; Barreto and Evans 1994)
Ramularia tungurahuana Petr., Sydowia 4(1-6): 507
(1950) (= Mycosphaerella tungurahuana)
Malaysia (Yen 1969)
Septoria chromolaenae Crous & den Breeÿen, in
Breeÿen et al., Fungal Divers 23: 102 (2006)
Cuba (Verkley et al. 2013), Jamaica (den Breeyen
et al. 2006)
Septoria ekmaniana Petr. & Cif., Annls mycol.
30(3/4): 300 (1932)
Dominican Republic (Ciferri 1961; Barreto and
Evans 1994), Mexico (den Breeyen et al. 2006), West
Indies (Minter et al. 2001)
Septoria eupatorii Roberge ex Desm., Annls Sci. Nat.,
Bot., sér. 3 20: 90 (1853)
Venezuela (Urtiaga 1986; Barreto and Evans 1994)
Septoria sp.
Guam (Russo 1985; Barreto and Evans 1994)
Myriangiales Starbäck
Myriangiaceae Nyl.
Anhellia nigra (Viégas) Arx, Persoonia 2(4): 434
(1963)
Rhytidhysteron bruguierae Dayarathne, in Dayarathne
et al., Mycosphere 11(1): 20 (2020)
= Tubercularia nigra F. Stevens, Annls mycol.
28(5/6): 371 (1930)
Brazil (Barreto and Evans 1994), West Indies
(Minter et al. 2001)
Subclass Pleosporomycetidae C.L. Schoch et al.
Hysteriales Lindau
Hysteriaceae Chevall.
Thailand (Dayarathne et al. 2020; This study)
26. Rhytidhysteron chromolaenae Mapook & K.D. Hyde
Thailand (This study)
Pleosporales Luttrell ex M.E. Barr
Acrocalymmaceae Crous & Trakun.
13
Fungal Diversity
27. Acrocalymma medicaginis Alcorn & J.A.G. Irwin,
Trans. Br. mycol. Soc. 88(2): 163 (1987)
Thailand (This study)
44. Hermatomyces chromolaenae Jun F. Li, Mapook &
K.D. Hyde, in Tibpromma et al., Fungal Divers 83: 47
(2017)
Thailand (Tibpromma et al. 2017)
Corynesporascaceae Sivan.
Lophiostomataceae Sacc.
28. Corynespora cassiicola (Berk. & M.A. Curtis) C.T.
Wei, Mycol. Pap. 34: 5 (1950)
Palau (Dixon et al. 2009)
Didymellaceae Gruyter et al.
29. Didymella chromolaenae Mapook & K.D. Hyde
Thailand (This study)
30. Nothophoma chromolaenae Mapook & K.D. Hyde
Thailand (This study)
45. Flabellascoma minimum A. Hashim., K. Hiray. &
Kaz. Tanaka, in Hashimoto et al., Studies in Mycology 90: 169 (2018)
Thailand (This study)
46. Pseudocapulatispora longiappendiculata Mapook &
K.D. Hyde
Thailand (This study)
Melanommataceae G. Winter (= Pseudodidymellaceae A.
Hashim. & Kaz. Tanaka)
Didymosphaeriaceae Munk
31. Chromolaenicola chiangraiensis Mapook & K.D.
Hyde
Thailand (This study)
32. Chromolaenicola lampangensis Mapook & K.D.
Hyde
Thailand (This study)
33. Chromolaenicola nanensis Mapook & K.D. Hyde
Thailand (This study)
34. Chromolaenicola thailandensis Mapook & K.D. Hyde
Thailand (This study)
35. Didymosphaeria sp.
India (Barreto and Evans 1994)
36. Montagnula chiangraiensis Mapook & K.D. Hyde
Thailand (This study)
37. Montagnula chromolaenae Mapook & K.D. Hyde
Thailand (This study)
38. Montagnula chromolaenicola Mapook & K.D. Hyde
Thailand (This study)
39. Montagnula thailandica Mapook & K.D. Hyde
Thailand (This study)
40. Pseudopithomyces palmicola J.F. Li, Ariyaw. & K.D.
Hyde, in Ariyawansa et al., Fungal Divers. 75: 41
(2015)
Thailand (This study)
41. Tremateia chiangraiensis Mapook & K.D. Hyde
Thailand (This study)
42. Tremateia chromolaenae Mapook & K.D. Hyde
Thailand (This study)
43. Tremateia thailandensis Mapook & K.D. Hyde
Thailand (This study)
47. Byssosphaeria schiedermayeriana (Fuckel) M.E. Barr,
Mycotaxon 20(1): 34 (1984)
= Herpotrichia schiedermayeriana Fuckel, Jb. nassau. Ver. Naturk. 27-28: 27 (1874) [1873-74]
Brazil (Barreto and Evans 1994)
Nigrogranaceae Jaklitsch & Voglmayr
48. Nigrograna chromolaenae Mapook & K.D. Hyde
Thailand (This study)
Neomassarinaceae Mapook & K.D. Hyde
49. Neomassarina chromolaenae Mapook & K.D. Hyde
Thailand (This study)
50. Neomassarina thailandica Phook., Jayasiri & K.D.
Hyde, in Hyde et al., Fungal Divers. 80: 138 (2016)
Thailand (This study)
Neopyrenochaetaceae Valenzuela-Lopez et al.
51. Neopyrenochaeta chiangraiensis Mapook & K.D.
Hyde
Thailand (This study)
52. Neopyrenochaeta chromolaenae Mapook & K.D.
Hyde
Thailand (This study)
53. Neopyrenochaeta thailandica Mapook & K.D. Hyde
Thailand (This study)
54. Neopyrenochaeta triseptatispora Mapook & K.D.
Hyde
Thailand (This study)
Hermatomycetaceae Locq.
Phaeosphaeriaceae M.E. Barr
13
Fungal Diversity
55. Leptospora chromolaenae Mapook & K.D. Hyde
Thailand (This study)
56. Leptospora phraeana Mapook & K.D. Hyde
Thailand (This study)
57. Leptospora thailandica Phukhams. & K.D. Hyde, in
Hyde et al., Fungal Divers. 80: 100 (2016)
Thailand (This study)
58. Murichromolaenicola chiangraiensis Mapook & K.D.
Hyde
Thailand (This study)
59. Murichromolaenicola chromolaenae Mapook & K.D.
Hyde
Thailand (This study)
60. Neoophiobolus chromolaenae Mapook & K.D. Hyde
Thailand (This study)
61. Ophiobolus ipohensis J.M. Yen, Cahiers Pacif. 13: 277
(1969)
Malaysia (Yen 1969; Barreto and Evans 1994)
62. Ophiosphaerella eupatorii J.M. Yen, Cahiers Pacif.
13: 278 (1969)
Malaysia (Yen 1969; Barreto and Evans 1994)
63. Paraleptospora chromolaenae Mapook & K.D. Hyde
Thailand (This study)
64. Paraleptospora chromolaenicola Mapook & K.D.
Hyde
Thailand (This study)
65. Phaeosphaeria eupatoriicola J.M. Yen, Cahiers Pacif.
13: 279 (1969)
Malaysia (Yen 1969; Barreto and Evans 1994)
66. Pseudoophiosphaerella huishuiensis J.F. Zhang, J.K.
Liu & Z.Y. Liu, in Zhang, Liu, Jeewon, Wanasinghe &
Liu, Mycosphere 8(1): 207 (2019)
Thailand (This study)
67. Pseudostaurosphaeria chromolaenae Mapook & K.D.
Hyde
Thailand (This study)
68. Pseudostaurosphaeria chromolaenicola Mapook &
K.D. Hyde
Thailand (This study)
69. Setophoma chromolaenae Quaedvl., Verkley, R.W.
Barreto & Crous, Studies in Mycology 75: 373 (2013)
Brazil (Quaedvlieg et al. 2013)
70. Yunnanensis chromolaenae Mapook & K.D. Hyde
Thailand (This study)
Pleosporaceae Nitschke
71. Alternaria zinniae M.B. Ellis, Mycol. Pap. 131: 22
(1972)
Brazil (Barreto and Evans 1994)
Pleosporales genera incertae sedis
72. Scolecobasidium eupatorii Y.L. Guo, Acta Mycol. Sin.
12(4): 271 (1993)
China (Zhuang 2001)
Pyrenochaetopsidaceae Valenzuela-Lopez et al.
73. Pyrenochaetopsis chromolaenae Mapook & K.D.
Hyde
Thailand (This study)
Roussoellaceae J.K. Liu et al.
74. Pseudoroussoella chromolaenae Mapook & K.D.
Hyde
Thailand (This study)
75. Pseudoroussoella elaeicola (Konta & K.D. Hyde)
Mapook & K.D. Hyde
Thailand (This study)
76. Setoarthopyrenia chromolaenae Mapook & K.D.
Hyde
Thailand (This study)
77. Xenoroussoella triseptata Mapook & K.D. Hyde
Thailand (This study)
Thyridariaceae Q. Tian & K.D. Hyde
78. Chromolaenomyces appendiculatus Mapook & K.D.
Hyde
Thailand (This study)
79. Pseudothyridariella chromolaenae Mapook & K.D.
Hyde
Thailand (This study)
Torulaceae Corda
80. Torula chromolaenae JF. Li, Phook., Mapook & K.D.
Hyde, in Li et al., Mycol. Progr. 16(4): 454 (2017)
Thailand (Li et al. 2017; This study)
81. Torula fici Crous, in Crous et al., IMA Fungus 6(1):
192 (2015)
Thailand (Li et al. 2017; This study)
82. Torula polyseptata C.G. Lin & K.D. Hyde, in Hyde
et al., Fungal Divers. 96: 71 (2019)
Thailand (Hyde et al. 2019a; This study)
Dothideomycetes orders incertae sedis
Botryosphaeriales C.L. Schoch et al.
Aplosporellaceae Slippers et al.
83. Aplosporella chromolaenae Mapook & K.D. Hyde
Thailand (This study)
84. Aplosporella hesperidica Speg., Anal. Soc. cient.
argent. 13(1): 18 (1882)
13
Fungal Diversity
Thailand (This study)
Botryosphaeriaceae Theiss. & P. Syd., Annales Mycologici
16 (1-2): 16 (1918)
85. Dothiorella oblonga F.J.J. Van der Walt, Slippers
& G.J. Marais, in Slippers et al., Persoonia 33: 163
(2014)
Thailand (This study)
86. Sphaeropsis chromolaenicola Mapook & K.D. Hyde
Thailand (This study)
94. Patellaria chromolaenae Mapook & K.D. Hyde
Thailand (This study)
Dothideomycetes families incertae sedis
Dysrhynchisceae Boonmee & K.D. Hyde*
95. Dysrhynchis citricola Bat. & Peres, Nova Hedwigia
2(4): 469 (1960)
Dominican Republic (Ciferri 1961; Barreto and
Evans 1994)
Dothideomycetes genera incertae sedis
Phyllostictaceae Fr.
87. Asteromella eupatoriicola (Kabát & Bubák) H. Ruppr.
1958
= Phyllosticta eupatoriicola Kabát & Bubák, Hedwigia 46: 288 (1907)
Malaysia, South America (Barreto and Evans 1994),
Puerto Rico (Stevenson 1975), Virgin Islands (Stevenson 1975), West Indies (Minter et al. 2001)
88. Phyllosticta eupatorii Punith., Mycol. Pap. 136: 21
(1974)
Sri Lanka (Barreto and Evans 1994)
Dyfrolomycetales K.L. Pang et al.
Pleurotremataceae Walt. Watson (= Dyfrolomycetaceae
K.D. Hyde et al.)*
89. Dyfrolomyces chromolaenae Mapook & K.D. Hyde
Thailand (This study)
Muyocopronales Mapook et al.
Muyocopronaceae K.D. Hyde
90. Muyocopron chromolaenae Mapook & K.D. Hyde
Thailand (This study)
91. Muyocopron chromolaenicola Mapook & K.D. Hyde
Thailand (This study)
92. Muyocopron lithocarpi Mapook, Boonmee & K.D.
Hyde, Phytotaxa 265(3): 235 (2016), new host record
Thailand (This study)
93. Neocochlearomyces chromolaenae Pinruan, Sommai,
Suetrong, J.Z. Groenew. & Crous, in Crous et al., Persoonia 41: 381 (2018)
Thailand (Crous et al. 2018)
Patellariales D. Hawksw. & O.E. Erikss.
Patellariaceae Corda
13
96. Ampullifera sp.
West Indies (Minter et al. 2001)
97. Melioliphila melioloides (Speg.) Piroz., Kew Bull.
31(3): 596 (1977)
West Indies (Minter et al. 2001)
98. Phaeoramularia eupatorii-odorati (J.M. Yen) X.J. Liu
& Y.L. Guo, Acta phytopath. sin. 12(no. 4): 7 (1982)
= Mycovellosiella eupatorii-odorati (J.M. Yen) J.M.
Yen, Bull. trimest. Soc. mycol. Fr. 97(3): 131 (1981)
China (Liu and Guo 1988; Morgan-Jones 1997;
Roux et al. 1997; Zhuang 2001; Guo and Liu 2003),
Malaysia (Yen JM 1968; Barreto and Evans 1994)
Class Eurotiomycetes Tehler ex O.E. Eriksson & K. Winka
Subclass Chaetothyriomycetidae Doweld
Chaetothyriales M.E. Barr
Chaetothyriaceae Hansf. ex M.E. Barr
99. Chaetothyrium dominicanum Cif., Sydowia 10(1-6):
133 (1957) [1956]
= Sphaerochaetia dominicana (Cif.) Bat. & Cif.,
Beih. Sydowia 3: 27 (1962)
Dominican Republic (Ciferri 1961; Batista and
Ciferri 1962; Barreto and Evans 1994), on living leaf
of C. odorata (= E. odoratum)
Class Leotiomycetes O.E. Erikss. & Winka
Helotiales Nannf
Ploettnerulaceae Kirschst
100. Pyrenopeziza sp. (= Cylindrosporium sp.)
Cambodia (Barreto and Evans 1994)
Class Sordariomycetes O.E. Erikss. & Winka
Subclass Diaporthomycetidae Senan. et al.
Diaporthales Nannf.*
Diaporthaceae Höhn. ex Wehm.
Fungal Diversity
101. Diaporthe chromolaenae Mapook & K.D. Hyde
Thailand (This study)
102. Diaporthe eupatoriicola Petr., Annls mycol. 20(3/4):
147 (1922)
Sri Lanka (Barreto and Evans 1994)
Diaporthomycetidae families incertae sedis
Sporidesmiaceae Fr.
103. Sporidesmium sp.
West Indies (Minter et al. 2001)
Subclass Hypocreomycetidae O.E. Erikss. & Winka
Glomerellales Chadef. ex Réblová et al.
Glomerellaceae Locq. ex Seifert & W. Gams
104. Colletotrichum gloeosporioides (Penz.) Penz. & Sacc.,
Atti Inst. Veneto Sci. lett., ed Arti, Sér. 6 2: 670 (1884)
Cuba (Urtiaga 1986; Barreto and Evans 1994), Sri
Lanka (Barreto and Evans 1994), Venezuela (Urtiaga
2004)
105. Colletotrichum sp.
: Cambodia - (Barreto and Evans 1994), Venezuela
- (Urtiaga 2004)
Hypocreales Lindau
Cordycipitaceae Kreisel ex G.M. Sung et al.
106. Akanthomyces lecanii (Zimm.) Spatafora, Kepler &
B. Shrestha, in Kepler et al. 2017
= Verticillium lecanii (Zimm.) Viégas, Revista Inst.
Café São Paulo 14: 754 (1939)
West Indies (Minter et al. 2001)
Hypocreaceae De Not.
107. Acrostalagmus albus Preuss, Linnaea 24: 126 (1851)
South America (Viegas 1961)
108. Acrostalagmus aphidum Oudem., Beih. Botan. Centralbl., Abt. B 11: 537 (1902)
Puerto Rico, Virgin Islands (Stevenson 1975); West
Indies (Minter et al. 2001)
109. Trichoderma guizhouense Q.R. Li, McKenzie & Yong
Wang bis, in Li et al., Mycol. Progr. 12(2): 170 (2012)
[2013]
Thailand (This study)
111. Fusarium fujikuroi Nirenberg, Mitt. biol. BundAnst.
Ld- u. Forstw. 169: 32 (1976)
= Fusarium moniliforme J. Sheld., Nebraska Agric.
Exp. Stat. Rep. 17: 23 (1904)
Nigeria (Richardson 1990)
112. Fusarium pallidoroseum (Cooke) Sacc., Syll. fung.
(Abellini) 4: 720 (1886)
Nigeria (Richardson 1990)
113. Fusarium solani (Mart.) Sacc., Michelia 2(no. 7): 296
(1881)
Nigeria (Richardson 1990)
Stachybotryaceae Lombard & Crous
114. Memnoniella chromolaenae Mapook & K.D. Hyde
Thailand (This study)
Subclass Sordariomycetidae O.E. Erikss & Winka (= Meliolomycetidae P.M. Kirk & K.D. Hyde)*
Meliolales Gäum. ex D. Hawksw. & O.E. Erikss.
Meliolaceae G.W. Martin ex Hansf.
115. Appendiculella sororcula (Speg.) Hansf., Beih.
Sydowia 2: 615 (1961)
= Irene sororcula (Speg.) F. Stevens, Annls mycol.
25(5/6): 423 (1927)
Dominican Republic (Ciferri 1961), Puerto Rico,
Venezuela (Hansford 1949), Trinidad and Tobago
(Baker and Dale 1951)
116. Meliola sp.
India (Barreto and Evans 1994)
117. Ophiociliomyces bauhiniae Bat. & I.H. Lima, Anais
Soc. Biol. Pernambuco 13(2): 30 (1955)
Brazil (Barreto and Evans 1994), Cambodia
- (Litzenberger et al. 1994)
Subclass Xylariomycetidae O.E. Erikss & Winka
Amphisphaeriales D. Hawksw. & O.E. Erikss.*
Apiosporaceae K.D. Hyde et al.
118. Arthrinium chromolaenae Mapook & K.D. Hyde
Thailand (This study)
Xylariales Nannf.
Cainiaceae J.C. Krug
Nectriaceae Tul. & C. Tul.
119. Longiappendispora chromolaenae Mapook & K.D.
Hyde
Thailand (This study)
110. Fusarium culmorum (Wm.G. Sm.) Sacc., Syll. fung.
(Abellini) 10: 726 (1892)
Nigeria (Richardson 1990)
Sordariomycetes genera incertae sedis
13
Fungal Diversity
120. Pleurophragmium capense (Thüm.) S. Hughes, Can.
J. Bot. 36: 796 (1958)
= Spiropes capensis (Thüm.) M.B. Ellis, Mycol.
Pap. 114: 5 (1968)
West Indies (Minter et al. 2001)
Ascomycota unclassified
Ascomycota genera incertae sedis
121. Alysidiopsis yunnanensis Y.L. Guo & X.J. Liu, Acta
Mycol. Sin. 11(3): 213 (1992)
China (Zhuang 2001), on leaves of C. odorata (= E.
odoratum)
122. Hansfordia pulvinata (Berk. & M.A. Curtis) S.
Hughes 1958
= Dicyma pulvinata (Berk. & M.A. Curtis) Arx
1981
West Indies (Minter et al. 2001)
123. Redbia trichomambusta R.W. Barreto, in Barreto &
Evans, Mycol. Res. 98(10): 1111 (1994)
Brazil (Barreto and Evans 1994)
124. Redbia sp.
West Indies (Minter et al. 2001)
Phylum Basidiomycota R.T. Moore
Class Agaricomycetes Doweld
Cantharellales Gäum.
Ceratobasidiaceae G.W. Martin
Colombia (Chardon and Toro 1930; Kern et al.
1933b), Venezuela (Chardon and Toro 1934; Kern
et al. 1934)
Pucciniales genera incertae sedis
129. Uredo bullula F. Kern, Mycologia 20(2): 77 (1928)
Dominican Republic (Kern 1930; Kern et al. 1933a;
Ciferri 1961; Barreto and Evans 1994), West Indies
(Minter et al. 2001)
Pucciniosiraceae Cummins & Y. Hirats.
130. Cionothrix praelonga (G. Winter) Arthur, N. Amer.
Fl. (New York) 7(2): 124 (1907)
= Cronartium praelongum G. Winter, Hedwigia 26:
24 (1887)
Dominican Republic, Mexico, Trinidad and Tobago,
Venezuela (Arthur JC 1922; Baker and Dale 1951;
Barreto and Evans 1994), Colombia (Buritica and
Pardo Cardona 1996; Pardo Cardona 1998), Costa
Rica (Arthur 1918a; Berndt 2004) Guatemala (Arthur
1918b), Mexico (Gallegos and Cummins 1981), Panama (Piepenbring 2006), West Indies (Minter et al.
2001)
Discussion
125. Rhizoctonia solani J.G. Kühn, Ann. Sper. agr., N.S.:
224 (1858)
= Thanatephorus cucumeris (A.B. Frank) Donk,
Reinwardtia 3: 376 (1956)
Taiwan (Sawada 1931)
Class Pucciniomycetes Bauer et al.
Pucciniales Clem. & Shear
Coleosporiaceae Dietel
126. Coleosporium eupatorii Arthur, Bull. Torrey bot. Club
33: 31 (1906)
China (Tai 1979; Barreto and Evans 1994; Zhuang
2005)
127. Coleosporium steviae Arthur, Bot. Gaz. 40: 197 (1905)
China (Barreto and Evans 1994), Mexico (Gallegos
and Cummins 1981)
Cronartiaceae Dietel
128. Cronartium praelongum G. Winter, Hedwigia 26: 24
(1887)
13
Taxonomic and phylogeny of fungi associated
with Chromolaena odorata
Chromolaena odorata (Siam weed) grows well in sunny,
open areas (abandoned fields, disturbed forests, pastures,
roadsides), as well as partial shade, but does not grow in
shade (Zachariades et al. 2009). In this study, we describe
the saprobic fungi associated with C. odorata, which were
mainly collected in northern Thailand. Our study included
morphological comparison and multigene analyses. A total
of 253 fungal specimens were collected. Eighty-eight fungal collections were successfully isolated into culture and
identified to 77 taxa distributed in ten orders and 23 families. These included one new family, 11 new genera, 43 new
species, 11 new host records, three new combinations, and
two reference specimens belonging to Dothideomycetes.
Pleosporales was the dominant order with most taxa in
the families Didymosphaeriaceae and Phaeosphaeriaceae
(Figs. 130, 131, 132). One new genus, four new species,
and one new host record of taxa belonging to Sordariomycetes were also identified, with Hypocreales being the
dominant order (Figs. 130, 133, 134). The result indicates
that Siam weed harbors a high fungal diversity, especially
Fungal Diversity
Fig. 130 Treatment of fungal
taxa from C. odorata
Fig. 131 Families in each order
of Dothideomycetes associated
with C. odorata
of Dothideomycetes (Schoch et al. 2009; Hyde et al. 2013;
Jayasiri et al. 2019).
Numerous studies have reported the capability of Dothideomycetes to survive extreme environmental conditions,
such as extreme temperatures, severe droughts, and solar
radiation (Ohm et al. 2012; Murgia et al. 2018). Murgia et al.
(2018) studied the biodiversity of fungi in hot desert sands
and reported that the most abundant fungal Phyla identified
in all the samples was Ascomycota and Dothideomycetes
were the dominant group. This group therefore plays an
important role in the desert ecosystem, as well as severe
conditions on rock surfaces (Ruibal et al. 2009). In this
study, Dothideomycetes were the dominant group of saprobic fungi on Siam weed in northern Thailand. Collections
were mostly from roadsides, abandoned fields, and disturbed
forests in sunny and open areas. However, only 35% (88/253)
of the saprobic fungi we collected from Siam weed were
successfully cultured. Another 65% of taxa contained dormant spores which did not germinate, and some collections
were partially or completely discharged with dry or empty
ascomata. This may be due to unfavorable nutritional and
environmental factors, such as temperature, humidity, as
well as UV light, all of which may reduce the capability of
spore germination (Gottlieb 1950; Ayerst 1969; Osman and
13
Fungal Diversity
Fig. 132 Numbers of taxa in each family of Dothideomycetes associated with C. odorata
Fig. 133 Families in each order
of Sordariomycetes associated
with C. odorata
13
Fungal Diversity
Fig. 134 Numbers of taxa in each family of Sordariomycetes associated with C. odorata
Valadon 1981; Fourtouni et al. 1998; Mitakakis et al. 2003;
Dagno et al. 2011; Wenneker et al. 2013).
Forty percent (25/62) of the saprobic taxa identified in
this study had spores with appendages or mucilaginous
sheaths. Such propagules have also been reported in terrestrial ascomycetes, and frequently occur in aquatic fungi,
especially marine species, for spore dispersal and attachment
to substrata (Hyde and Jones 1988; Hyde and Goh 2003;
Jones 2006; Hyde et al. 2016a; Jones et al. 2019). Moreover,
77.5% (48/62) of the taxa had spore pigmentation, which is
divided into 55% (34/62) of dark pigmentation and 22.5%
(14/62) with light pigmentation, while 22.5% (14/62) of the
spores were hyaline. We assume that appendages, mucilaginous sheaths, and pigmentation of the spores serve to
protect the propagules from drought and UV light, as well
as other unfavorable conditions. In addition, saprobic fungi
associated with C. odorata in our study were mainly sexual
morphs, which is related to sexual reproduction. This is
likely to result in evolution of species for survival in adverse
environmental competition and better competition with other
species. Evolution occurs due to genetic complementation
provided by the combination of different parents’ genomes
that has benefits to repair DNA damage and increase genetic
variability with the evolution rate of sexual species (Maynard Smith 1978; Colegrave 2012; Wallen and Perlin 2018).
A comparison of fungi on Siam weed and other
hosts
Of the 62 species identified in this study as saprobes on
Siam weed, only 15 are previously known, while the other
47 species are new to science. Of these 15 species, we found
that 13 species are previously known from unrelated hosts,
which comprised eight known species reported from Thailand, while five known species have been reported from
elsewhere, and the other two species (Torula chromolaenae and Torula fici) are previously known from Siam weed
(Table 30). The previously known reported species were
mainly found in northern Thailand, such as Leptospora
thailandica, Muyocopron lithocarpi, Pseudopithomyces
palmicola, Pseudoroussoella elaeicola, Torula polyseptata,
Neomassarina thailandica and Flabellascoma minimum,
while some have been reported in the adjacent area, China
(Table 30). Furthermore, we found some known species
occurred in different habitats. Rhytidhysteron bruguierae
was collected from submerged branches of Bruguiera sp. in
a marine habitat and also from terrestrial habitats on a dead
stem of Siam weed in this study. The strain from submerged
habitats was collected from Phetchaburi Province in western
Thailand (Dayarathne et al. 2020), while our strains from
terrestrial habitats were collected from several provinces in
13
Fungal Diversity
northern Thailand (Chiang Rai, Phrae, Chiang Mai and Mae
Hong Son).
Why are the fungi on Siam weed and other hosts
so different?
Some studies have focused on microfungi associated with
selected groups of plants in both Thailand and China and
from different climates. Doilom et al. (2017) showed that
Dothideomycetes were a dominant group on Tectona grandis
(teak; Lamiaceae) in northern Thailand with collecting sites
in six provinces (Phayao, Chiang Mai, Chiang Rai, Sukhothai, Phrae, Uttaradit), while Dai et al. (2017) found a
similar number of Dothideomycetes and Sordariomycetes on
bamboo (Poaceae) from northeastern Thailand (Mukdahan
province), northern Thailand (Phayao, Chiang Rai, Chiang
Mai, Uttaradit and Phitsanulok province), southern Thailand
(Phangnga and Krabi province), and southern China. Tibpromma et al. (2018) reported on the fungi on Pandanaceae
from northern Thailand (Chiang Rai and Chiang Mai provinces), eastern Thailand (Chonburi Province), western Thailand (Prachuabkhirikhan Province), and southern Thailand
(Krabi, Phangnga, Nakhonsithummarat, Phatthalung and
Phuket provinces), as well as in southern China. In general,
the fungal communities differed, depending on the host,
with low overlap. This indicates that if we study other unrelated hosts, we should expect to find a high, unique diversity
(Hyde et al. 2018a, b, 2019a, b).
In this study, fungi on Siam weed were mainly collected
in northern Thailand (Chiang Mai, Chiang Rai, Lampang,
Mae Hong Son, Nan, and Phrae provinces), as well as in
western Thailand (Phetchaburi Province). We found three
overlapping species (Pseudopithomyces palmicola, Torula
chromolaenae and T. fici) which were previously reported
from Pandanus in Tibpromma et al. (2018); however, we
could not find any overlapping species with teak (Doilom
et al. 2017) and bamboo (Dai et al. 2017). Although collecting sites are in the same provinces, fungi are likely to
differ on the unrelated hosts. In addition, we also compared
existing species to microfungi on Tamarix (Tamaricaceae)
from Italy and Russia in Thambugala et al. (2017a), on grass
(Poaceae) from China, Italy, Russia and Thailand in Thambugala et al. (2017b), as well as Rosaceae, which were collected from Thailand, China, England, India, Italy, Russia,
Sweden, and Wales in Wanasinghe et al. (2018); however,
there were no overlapping species. This indicates that fungi
may be typically different in each country, and each study
site, and on each unrelated host. It has been postulated that
the differences in the dominant taxa occurring on unrelated
plant species may be due to factors, such as moisture content,
temperature, pH, nutrients, organic content in each substrate,
decay period, fungal adaptation, competitive interactions
among the fungal group, as well as plant and soil chemistry,
13
which are likely to influence diversity, species, and the community of fungi (Venugopal et al. 2016, 2017). This was
supported by Boonyuen et al. (2014) where high humidity
influenced the number of fungal species compared to other
sites with low humidity. Therefore, in areas with similar or
relatively similar environmental conditions we should expect
to find numerous overlapping species. However, this is not
the case, as shown with studies on communities on bamboo,
Pandanaceae, and teak and thus host dependence is probably
causing speciation to be more strongly shaped rather than
climate (Ezard et al. 2011). The saprobic fungal communities on bamboo, grass, Pandanaceae, Rosaceae and teak
have been shown to have very few overlapping species. Thus
herein, we studied the fungi on Siam weed and also found
few overlapping species with those on the aforementioned
hosts. The reason for the uniqueness of taxa on any host
may be due to them having endophytic lifestyles (Promputtha et al. 2007). Endophytes are likely to be a host-specific
and close relationship with their particular host (Wong and
Hyde 2001; Chauhan et al. 2019). However, most endophytes recovered to date are not unique to their hosts (Guo
et al. 2001; Wang et al. 2005; Huang et al. 2009; Sun et al.
2011), but this is probably because techniques isolate only
fast growing taxa (Guo et al. 2000, 2001; Promputtha et al.
2007).
Life modes of fungi on Siam weed
Fungi are able to survive in various substrates in varied
habitats (Cai et al. 2006; Kodsueb et al. 2016), and they
can also change their lifestyle. Some endophytic fungi have
parasitic or pathogenic modes and this is thought to occur
when environmental and physiological conditions become
suitable, such as the host becoming stressed (Strobel 2018).
Endophytes may also be mutualistic and protect the host
plant from diseases or herbivores (Carroll 1988; Fróhlich
et al. 2000; Chaverri and Samuels 2013). Several studies
have postulated that endophytes which live within plants are
the initial colonizers of dead leaves and twigs from the plant
and they can be saprotrophic when the host senescence or
when the plant part dies (Promputtha et al. 2007, 2010; Purahong and Hyde 2011; Sun et al. 2011; Jeewon et al. 2013).
Some of the previously known species were found to
have saprotrophic lifestyles in our study; however, they
have also been reported to occupy other life modes in plants
(Table 30). For example, three fungal species found as
saprobes on Siam weed, have been reported as pathogenic
on various hosts. For example, Acrocalymma medicaginis
causes crown rot disease of Medicago sativa (Fabaceae) in
Australia (Alcorn and Irwin 1987; Trakunyingcharoen et al.
2014), Aplosporella hesperidica causes early stem-end rot of
Citrus sinensis (Rutaceae) in Zimbabwe (Yang et al. 2017),
and Pseudopithomyces palmicola causes chlorotic and/or
Fungal Diversity
Table 30 Species found on Siam weed with their previous known host
Species
Host
Host family
Origin
Lifestyle
Reference
Acrocalymma medicaginis
Crown rot of Medicago
sativa
Fabaceae
Australia
Pathogen
Asteraceae
Chiang Rai; Thailand
Saprobe
(Alcorn and Irwin 1987),
(Trakunyingcharoen
et al. 2014)
This study
Rutaceae
Rutaceae
India
Zimbabwe
Saprobe
Pathogen
Rao (1969)
Yang et al. (2017)
Asteraceae
Chiang Rai; Thailand
Saprobe
This study
Fabaceae
Asteraceae
Namibia; South Africa
Chiang Rai; Thailand
Uncertain
Saprobe
Slippers et al. (2014)
This study
Arecaceae
Taiwan
Saprobe
Hashimoto et al. (2018)
Fabaceae
Taiwan
Saprobe
Hashimoto et al. (2018)
Fabaceae
Lampang; Thailand
Saprobe
Jayasiri et al. (2019)
Asteraceae
Nan; Thailand
Saprobe
This study
Verbenaceae
Chiang Rai; Thailand
Saprobe
Hyde et al. (2016a, b)
Asteraceae
Chiang Rai; Thailand
Saprobe
This study
Fagaceae
Chiang Rai; Thailand
Saprobe
Fabaceae
Chiang Rai; Thailand
Saprobe
Mapook et al. (2016a,
b, c)
Jayasiri et al. (2019)
Fabaceae
Guizhou; China
Saprobe
Jayasiri et al. (2019)
Unknown
Yunnan; China
Saprobe
Phookamsak et al. (2019)
Asteraceae
Saprobe
This study
Asparagaceae
Chiang Rai, Lampang;
Thailand
Chiang Mai; Thailand
Saprobe
Hyde et al. (2016a, b)
Asteraceae
Chiang Rai; Thailand
Saprobe
This study
Unknown
China
Saprobe
Zhang et al. (2019)
Asteraceae
Saprobe
This study
Vitaceae
Chiang Rai, Chiang
Mai, Mae Hong Son;
Thailand
Italy
Fabaceae
Unknown
Weak Pathogen Liu et al. (2018)
Poaceae
Unknown
Weak Pathogen Liu et al. (2018)
Rosaceae
Unknown
Weak Pathogen Liu et al. (2018)
Dead stems of Chromolaena odorata
Aplosporella hesperidica Citrus aurantium
Early stem-end rot of
Citrus sinensis
Dead stems of C.
odorata
Dothiorella oblonga
Acacia mellifera
Dead stems of C.
odorata
Flabellascoma minimum On petioles of Arenga
engleri
On pods of Bauhinia
purpurea
On pods of Leucaena
leucocephala
Dead stems of C.
odorata
Leptospora thailandica On dead branches of
Duranta sp.
Dead stems of C.
odorata
Muyocopron lithocarpi On dead leaves of
Lithocarpus lucidus
On decaying pods of
Peltophorum sp.
On fallen pod of Cercis
chinensis
On dead stems of herbaceous plant
Dead stems of C.
odorata
Neomassarina thailanOn dead bract-like leaves
dica
from flower stalk of
Agave angustifolia
Dead stems of C.
odorata
Pseudoophiosphaerella On dead culms of unihuishuiensis
dentified herbaceous
plant
Dead stems of C.
odorata
Pseudopithomyces
palmicola
On leaves lesions of Vitis
vinifera
On leaves lesions of
Phaseolus vulgaris
On leaves lesions of Poa
annua
On leaves lesions of
Fragaria sp.
Weak Pathogen Liu et al. (2018)
13
Fungal Diversity
Table 30 (continued)
Species
Pseudoroussoella
elaeicola
Rhytidhysteron bruguierae
Torula chromolaenae
Torula fici
Torula polyseptata
Trichoderma
guizhouense
Host
Host family
Origin
Lifestyle
Reference
On dead leaves of Pandanus amaryllifolius
On dead leaves of unidentified grass species
On leaves of Acoelorrhaphe wrightii
Dead stems of C.
odorata
On dead petiole of Elaeis
guineensis
Dead stems of C.
odorata
Submerged branches of
Bruguiera sp.
Dead stems of C.
odorata
Pandanaceae
Chiang Rai; Thailand
Saprobe
Tibpromma et al. (2018)
Unknown
Yunnan; China
Saprobe
Hyde et al. (2017b)
Arecaceae
Chiang Rai; Thailand
Saprobe
Ariyawansa et al. (2015)
Asteraceae
This study
Arecaceae
Chiang Rai, Chiang Mai; Saprobe
Thailand
Chiang Rai; Thailand
Saprobe
Asteraceae
Lampang; Thailand
Saprobe
This study
Rhizophoraceae
Phetchaburi; Thailand
Saprobe
Dayarathne et al. (2020)
Asteraceae
This study
Dead stem of Chromolaena odorata
Dead leaf of Pandanus
tectorius
Dead stems of C.
odorata
On dead stems of Chromolaena odorata
On decaying cone of
Magnolia grandiflora
On decaying fruit pericarp of Garcinia sp.
On dead leaf of Pandanus sp.
On submerged decaying
wood
Ficus religiosa
Dead stems of C.
odorata
On submerged decaying
wood
Dead stems of C.
odorata
Soil
Asteraceae
Chiang Rai, Chiang Mai, Saprobe
Mae Hong Son, Phrae;
Thailand
Chiang Mai; Thailand
Saprobe
Pandanaceae
Yunnan; China
Tibpromma et al. (2018)
Asteraceae
This study
Asteraceae
Chiang Rai, Chiang Mai; Saprobe
Thailand
Chiang Rai; Thailand
Saprobe
Magnoliaceae
Yunnan; China
Saprobe
Jayasiri et al. (2019)
Clusiaceae
Ranong; Thailand
Saprobe
Jayasiri et al. (2019)
Pandanaceae
Chiang Mai; Thailand
Saprobe
Tibpromma et al. (2018)
Unknown
China
Saprobe
Su et al. (2018)
Moraceae
Asteraceae
Cuba
Chiang Rai; Thailand
Unknown
Saprobe
Crous et al. (2015a)
This study
Unknown
Chiang Rai; Thailand
Saprobe
Hyde et al. (2019a)
Asteraceae
Chiang Rai; Thailand
Saprobe
This study
–
China
Saprobe
Li et al. (2013)
Endophyte in stems of
Ancistrocladus korupensis
Endophyte in stems of
Cola spp.
Dead stems of C.
odorata
Ancistrocladaceae Cameroon
Endophyte
Chaverri et al. (2015)
Malvaceae
Cameroon
Endophyte
Chaverri et al. (2015)
Asteraceae
Phrae; Thailand
Saprobe
This study
necrotic symptoms on leaves of Vitis vinifera (Vitaceae),
Phaseolus vulgaris (Fabaceae), Fragaria sp. (Rosaceae) and
Poa annua (Poaceae) in Italy (Liu et al. 2018). In addition,
Trichoderma guizhouense was initially found in soil polluted
13
Saprobe
Phookamsak et al. (2019)
Li et al. (2017)
Li et al. (2017)
by heavy metals in China, and has also been reported as
endophytic in stems of Ancistrocladus korupensis (Ancistrocladaceae) and Cola spp. (Malvaceae) in Cameroon (Li
et al. 2013; Chaverri et al. 2015), while in this study it was
Fungal Diversity
reported as a saprobic on Siam weed. This indicates that
some of the fungi on Siam weed are likely to have the ability to be endophytes or plant pathogen in other hosts, while
most of the previously known species had saprobic lifestyles
on Siam weed.
Where did the fungi on Siam weed come from?
Co-introduction and host-jumping are hypotheses generally used to explain the fungi occurring in invasive plants
(Shipunov et al. 2008; Burgess et al. 2016; Truter et al.
2017). In co-introduction, the plant colonizes a new invasive range accompanied by associated microorganisms from
the native range. In the case of fungi, however, this could
only occur if the new invasive range was adjacent to native
range and spores could be wind-dispersed or the fungi are
seed-borne. In host-jumping, the invasive plants colonizes
a new invasive range without any associated fungi from the
native range. It then obtains newly associated fungi from
surrounding plants in the invasive range. In the case of Siam
weed, whose origins are the Americas, it is likely that cointroduction also occurred, as the weed was dispersed by
seed (Scott et al. 1998; Setter and Campbell 2002; Zachariades et al. 2009). Esuruoso (1971) reported seed-borne fungi
from Siam weed, were found as important pathogens of the
various food crops in Nigeria. However, there are no further
studies on seed-borne fungi of Siam weed and the distances
involved should prohibit dispersal by wind. Therefore, we
suggest that Siam weed gained its associated fungi via hostjumping rather than co-introduction. A few associated fungi
may have been dispersed by seeds when it colonized a new
area (co-introduction hypothesis), but probably most newly
associated fungi were acquired from the surrounding hosts
(host-jumping hypothesis).
In this study, the previously known species were mostly
reported from other hosts in Thailand and elsewhere, while
there is no report in closely related plants, such as other
Asteraceae. Although we have little evidence, we suggest
that the saprobic fungi found on Siam weed in this study, are
likely to have jumped host from unrelated adjacent plants,
rather than arrived by co-introduction. Alternatively, they
may occur on other Asteraceae hosts in Thailand, but there
have been very few studies of the fungal diversity of this
host.
Did the fungi on Siam weed jump hosts?
The term host-jumping to unrelated or distantly hosts
was discussed in Roy (2001) and seems to have occurred
here. There is some evidence that the fungi on Siam weed
jumped from other hosts. Muyocopron lithocarpi was previously found in Thailand from the host family Fabaceae
on Lithocarpus lucidus and Peltophorum sp. (Mapook et al.
2016b, Jayasiri et al. 2019) and Cercis chinensis in China,
while it was found on Siam weed in this study. It is likely
that Muyocopron lithocarpi jumped from Fabaceae or some
other host to Siam weed (Asteraceae). Muyocopron dipterocarpi was introduced as saprobe from dried twigs of
Dipterocarpus tuberculatus (Dipterocarpaceae) in northern
Thailand, and did not produce specialized infection structures (Mapook et al. 2016b). The taxon was subsequently
found on a dried twig of a rubber tree, Hevea brasiliensis
(Euphorbiaceae). The species produces appressoria from
germinating ascospores, indicating an endophytic lifestyle
(Konta et al. 2016; Senwanna et al. 2019). This species, is
therefore, likely to have jumped host from some other local
host, as both Siam weed and rubber are not native plants
(Senwanna et al. 2019). Muyocopron heveae was also collected from rubber trees, and produced hyaline appressoria
(Senwanna et al. 2019). This suggests that Muyocopron species are likely to have the ability to be endophytic or pathogenic and have jumped to unrelated hosts.
Most pathogens (e.g. Colletotrichum species) produce
appressoria, so that they can invade the host and some species are thought to be host-specific (Sutton 1968; Zhou and
Hyde 2001; Nesher et al. 2008; Yan et al. 2018). Many saprobes also produce appressoria (Konta et al. 2016; Phukhamsakda et al. 2016; Senwanna et al. 2019; Hyde et al. 2020),
so that they can also infect the host. Like pathogens, the
endophytes will have built up a relationship with the host
over an evolutionary timescale so that they can infect the
host without being killed and also without causing symptoms. For example, the palm and Pandanaceae genera Linocarpon and Oxydothis, the majority of which are saprobes,
have been shown to produce appressoria, indicating that
they colonize living plants (Konta et al. 2016). The saprobic species, Sparticola junci, grows on Spartium junceum
(Fabaceae) and also produce appressoria, which suggests
that also have endophytic lifestyles (Phukhamsakda et al.
2016).
Fungi have the ability to exhibit more than one lifestyle
in a different host. Although they are the same species, the
ability to produce appressoria is important for living host
colonization. Appressoria formation is related to surface
signals with cell division and cell wall modifications in the
early germination stages of pathogenic conditions, followed
by cAMP with other additional pathways, while the appressoria formation is not needed for a saprobic lifestyle and its
germination pathways is a reversed process (Barhoom and
Sharon 2004). There was no evidence for appressoria formation in this study, as all taxa germinated without forming
specialized infection structures. Most of the taxa on Siam
weed are therefore likely to have jumped host from other
plants and are unlikely to be a specialist to this weed.
13
Fungal Diversity
Evolution of Asteraceae
Asteraceae, a large and widespread flowering plant (Angiospermae) family which accommodates Chromolaena odorata (Jeffrey 2007) evolved in the late Cretaceous at 85.9
Mya (82.3-91.5) with a stem age at 88–89 Mya. This dating
is based on the oldest fossils of pollen grains (Barreda et al.
2015). According to the study of Barreda et al. (2015), the
majority of the subfamilies in the Asteraceae clade diverged
during the warmest Earth’s temperature, which started during the late Paleocene to early Eocene Epoch in the Cenozoic Era. Climatic change may have been associated with the
diversification of Asteraceae and some Asteraceae members
probably survived across the Cretaceous-Tertiary (K-T) mass
extinction event (approximately 66 Mya), due to their adaptation and tolerance to extreme environmental, as well as
ecological conditions (McFadyen 1996; Chauhan and Johnson 2008; Koutika and Rainey 2010; Barreda et al. 2015).
For example, Siam weed has been reported as adapted for
acidic soils (Koutika and Rainey 2010) and can tolerate
harmful heavy metals, such as cadmium and zinc (Agunbiade and Fawale 2009; Okoronkwo et al. 2014; Ruangdech
et al. 2017), as well as improving soil contaminated by used
oil (Anolifo and Vwioko 2001; Atagana 2011; Ikhajiagbe
and Akindolor 2016), which may also affect the microbial
community, especially soil microorganisms.
Does evolution drive the diversity of saprobic fungi
associated with Chromolaena odorata?
The divergence time estimates of each fungal family identified in this study, were obtained from previous studies
(Mapook et al. 2016c; Hyde et al. 2017a; Hongsanan et al.
2017; Liu et al. 2017; Guterres et al. 2018; Soleimani et al.
2018; Phillips et al. 2019; Samarakoon et al. 2019). The
divergence times for 19 of the 23 families are recorded in
Tables 31 and 32. Divergence time estimates for stem ages
show that one family diverged in the Carboniferous (5%),
one in the Triassic (5%), two in the Jurassic (10%), and 15
(79%) in the Cretaceous Period (Fig. 135), while the results
of divergence time estimates for crown age show that the
fungal family associated with C. odorata in this study mostly
diverged in the Paleogene period (Fig. 136).
Although most fungal families coincided with the
expected origin of the host family (Asteraceae), all species
identified are probably not specific to Siam weed. Perhaps
these taxa evolved with other Asteraceae present in Thailand. Four fungal families found on Siam weed, e.g. Pleurotremataceae (174 Mya) and Muyocopronaceae (172 Mya),
evolved earlier than Asteraceae in the Jurassic, Hysteriaceae
(219 Mya) evolved in the Triassic, and Patellariaceae (311
Mya) evolved in the Carboniferous (Table 31). Asteraceae
evolved 85.9 Mya in the late Cretaceous. Fungi that evolved
13
before the Asteraceae may have jumped from hosts or have
evolved with the ancestors of the family. The fungal families that evolved earlier than Asteraceae produce ascomata
with heavy pigmentation in the thick peridium wall, which
may protect them from drought and UV light and appear to
have wide host ranges (Gueidan et al. 2015; Ekanayaka et al.
2017; Hernández-Restrepo et al. 2019; Jayasiri et al. 2019).
Antimicrobial activity of fungi on Siam weed
In this study, we provide evidence of the antimicrobial
activity of fungi associated with Chromolaena odorata
(Table 33). Forty (65%) out of 62 species showed potential
antimicrobial activity, with 12 species showing antimicrobial
activity against Mucor plumbeus (filamentous fungus), 12
species against Escherichia coli (Gram-negative bacterium),
eight species against both M. plumbeus and Bacillus subtilis (Gram-positive bacterium), three species against bacteria
and filamentous fungus, two species against only B. subtilis,
as well as two species against both E. coli and M. plumbeus,
and one species against both B. subtilis and E. coli. In contrast, 18 (29%) of the 62 species did not show antimicrobial
activity and four species were not tested (Figs. 137, 138).
This infers that more than half of the fungal species associated with C. odorata have antimicrobial potential and could
be promising sources for the discovery of novel bioactive
compounds.
Among the previously known species in this study, 12 of
the 15 lack previously reported antimicrobial activity. For
example, our study showed for the first time that Pseudopithomyces palmicola, Neomassarina thailandica, Torula
fici, T. polyseptata and Rhytidhysteron bruguierae showed
antimicrobial activity against M. plumbeus and Leptospora
thailandica, Pseudoophiosphaerella huishuiensis and Torula chromolaenae showed antimicrobial activity against all
tested organisms. Furthermore, Acrocalymma medicaginis
and Flabellascoma minimum showed antimicrobial activity
against B. subtilis and M. plumbeus. Aplosporella hesperidica and Pseudoroussoella elaeicola showed antimicrobial
activity against E. coli, while Dothiorella oblonga and Muyocopron lithocarpi showed no antimicrobial activity. Trichoderma guizhouense, which had previously been reported
with antifungal activity against Fusarium oxysporum f. sp.
cubense causing banana wilt disease (Zhang et al. 2016),
also showed antimicrobial activity against E. coli and M.
plumbeus in this study. However, research on novel bioactive
compounds and other chemical discoveries from these new
fungal taxa is in its infancy.
Fungal Diversity
Table 31 Divergence time estimates of the Dothideomycetes families recorded in this study
Subclass
Order
Pleosporomycetidae Hysteriales
Dothideomycetes
orders incertae
sedis
Family
Divergence times (crown
age)
Hysteriaceae
149 Mya (90–213), late
Jurassic
Pleosporales
Acrocalymmaceae
23 Mya (8–44), the Neogene period
Didymellaceae
63 Mya (35–97), Paleogene period
Didymosphaeriaceae
72 Mya (48–101), Cretaceous
Lophiostomataceae
–
Nigrogranaceae
72 Mya (44–124), Cretaceous
Neopyrenochaetaceae –
Phaeosphaeriaceae
75 Mya (46–102), Cretaceous
Pyrenochaetopsidaceae –
Roussoellaceae
62 Mya (34–91), Cenozoic Era (Paleogene
period)
Thyridariaceae
15 Mya (3–38), Cenozoic
Era (Neogene period)
Torulaceae
15 Mya (4–34), Cenozoic
Era (Neogene period)
Neomassarinaceae
–
Botryosphaeriales Aplosporellaceae
40 Mya, Cenozoic Era,
(Paleogene period)
Botryosphaeriaceae
Dyfrolomycetales Pleurotremataceae
Muyocopronales
Muyocopronaceae
Patellariales
Patellariaceae
61 Mya, Cenozoic Era,
(Paleogene period)
76 Mya (38–125), late
Cretaceous
52 Mya (38–66), Cenozoic Era, (Paleogene
period)
164 Mya (72–283),
Jurassic
Divergence times (stem
age)
Reference
219 Mya (161–282), late
Triassic
114 Mya (71–156),
Cretaceous
115 Mya (84–149),
Cretaceous
109 Mya (83–139),
Cretaceous
–
131 Mya (86–180),
Cretaceous
–
99 Mya (73–129), Cretaceous
–
77 Mya (44–110), late
Cretaceous
Liu et al. (2017)
95 Mya (60–138), Cretaceous
140 Mya (95–188), early
Cretaceous
–
94 Mya, Cretaceous
Liu et al. (2017)
–
Phillips et al. (2019)
94 Mya, Cretaceous
Phillips et al. (2019)
174 Mya (113–243),
Jurassic
172 Mya (130–218),
Jurassic
Lui et al. (2017)
311 Mya (244–407),
Carboniferous period
Liu et al. (2017)
Liu et al. (2017)
Liu et al. (2017)
Liu et al. (2017)
–
Liu et al. (2017)
–
Liu et al. (2017)
–
Liu et al. (2017)
Liu et al. (2017)
Mapook et al. (2016c)
Table 32 Divergence time estimates of the Sordariomycetes families recorded in this study
Subclass
Order
Diaporthomycetidae Diaporthales
Family
Diaporthaceae
Divergence times (crown
age)
61.15 Mya (29.7–89.9),
Cenozoic Era, (Paleogene
period)
Hypocreomycetidae Hypocreales
Hypocreaceae
Cenozoic Era, (Paleogene
period)
Stachybotryaceae Cenozoic Era, (Paleogene
period)
Xylariomycetidae
Amphisphaeriales Apiosporaceae
Cenozoic Era, (Paleogene
period)
Xylariales
Cainiaceae
Cenozoic Era, (Paleogene
period)
Divergence times (stem age) Reference
87.66 Mya, Cretaceous
Guterres et al. (2018)
Cretaceous
Hyde et al. (2017a)
88 Mya (50–130), Cretaceous
69 Mya (50–130), late
Cretaceous
128 Mya, Cretaceous
Hyde et al. (2017a)
Hyde et al. (2017a)
Hyde et al. (2017a)
13
Fungal Diversity
Fig. 135 Fungal families
recorded on Siam weed with
stem age divergence estimates
Checklist of fungi associated with Chromolaena
odorata
There are 68 fungal species previously reported from Siam
weed, based on the USDA Systematic Mycology and Microbiology Laboratory (SMML) database (Farr and Rossman
2020), together with relevant literature. These 68 species
were recorded from Asia (45), Americas (37), West Indies
(36), Africa (7), and Oceania (1). Several taxa are known
from more than one location, while only seven species were
recorded from Thailand (Cercospora sp., Hermatomyces
chromolaenae, Neocochlearomyces chromolaenae, Pseudocercospora eupatorii, P. eupatorii-formosani, Torula
chromolaenae, and T. fici).
Fig. 136 Fungal families
recorded on Siam weed with
crown age divergence estimates
13
The 130 taxa recorded from C. odorata, including the 62
species listed in this study are distributed in 20 orders, 48
families and 85 genera and are divided among (i) Ascomycota: four classes, 18 orders, 43 families, 80 genera and 110
species identified with 13 unidentified species (Fig. 139),
and (ii) Basidiomycota: two classes, two orders, five families, five genera, with six species identified (Fig. 140). Siam
weed harbors a higher diversity, especially of Dothideomycetes with 75% of fungal taxa (98/130) from 30 families
with Pleosporales being the dominant order (Figs. 139, 141)
and 15% (20/130) of fungal taxa are belonging to Sordariomycetes with Hypocreales is dominant orders (Fig. 142).
The most commonly reported genus on Siam weed is
Pseudocercospora with six known species including Pseudocercospora aciculina, P. ageratoides, P. convoluta, P.
Fungal Diversity
Table 33 Preliminary screening
of Siam weed fungi for
antimicrobial activity against
Bacillus subtilis, Escherichia
coli and Mucor plumbeus with
diameter of inhibition zone
Species name
B. subtilis (mm)
E. coli (mm)
M.
plumbeus
(mm)
Acrocalymma medicaginis
Aplosporella chromolaenae
Aplosporella hesperidica
Arthrinium chromolaenae
Chromolaenicola chiangraiensis
Chromolaenicola lampangensis
Chromolaenicola nanensis
Chromolaenicola thailandensis
Chromolaenomyces appendiculatus
Diaporthe chromolaenae
Didymella chromolaenae
Dothiorella oblonga
Dyfrolomyces chromolaenae
Flabellascoma minimum
Leptospora chromolaenae
Leptospora phraeana
Leptospora thailandica
Longiappendispora chromolaenae
Memnoniella chromolaenae
Montagnula chiangraiensis
Montagnula chromolaenae
Montagnula chromolaenicola
Montagnula thailandica
Murichromolaenicola chiangraiensis
Murichromolaenicola chromolaenae
Muyocopron chromolaenae
Muyocopron chromolaenicola
Muyocopron lithocarpi
Neomassarina chromolaenae
Neomassarina thailandica
Neoophiobolus chromolaenae
Neopyrenochaeta chiangraiensis
Neopyrenochaeta chromolaenae
Neopyrenochaeta thailandica
Neopyrenochaeta triseptatispora
Nigrograna chromolaenae
Nothophoma chromolaenae
Paraleptospora chromolaenae
Paraleptospora chromolaenicola
Patellaria chromolaenae
Pseudocapulatispora longiappendiculata
Pseudoophiosphaerella huishuiensis
Pseudopithomyces palmicola
Pseudoroussoella chromolaenae
Pseudoroussoella elaeicola
Pseudostaurosphaeria chromolaenae
Pseudostaurosphaeria chromolaenicola
Pseudothyridariella chromolaenae
Pyrenochaetopsis chromolaenae
12
–
–
–
No inhibition
–
–
No inhibition
20
–
No inhibition
No inhibition
12
15
13
–
13
–
–
8
–
No inhibition
No inhibition
–
–
No inhibition
No inhibition
No inhibition
–
–
No inhibition
–
13
No inhibition
16
–
No inhibition
–
–
–
7
10
–
No inhibition
–
10
–
–
–
–
9
9
8
13
–
–
–
–
–
14
12
–
13
–
–
–
–
8
9
8
–
–
–
17
20
22
24
–
–
11
18
17
–
11
–
–
30
10
–
–
–
24
11
8
12
13
–
8
–
–
–
–
23
10
10
10
–
–
–
9
–
25
18
23
–
13
Fungal Diversity
Table 33 (continued)
Fig. 137 Fungal species on
Siam weed with potential for
antimicrobial activity
Fig. 138 Numbers of fungal
species on Siam weed with
potential antimicrobial activity
against: G + Gram-positive
bacteria, G − Gram-negative
bacteria, F filamentous fungus,
A active with all microbial
13
Species name
B. subtilis (mm)
E. coli (mm)
M.
plumbeus
(mm)
Rhytidhysteron bruguierae
Rhytidhysteron chromolaenae
Setoarthopyrenia chromolaenae
Sphaeropsis chromolaenicola
Torula chromolaenae
Torula fici
Torula polyseptata
Tremateia chiangraiensis
Tremateia chromolaenae
Tremateia thailandensis
Trichoderma guizhouense
Xenoroussoella triseptata
Yunnanensis chromolaenae
–
–
16
No inhibition
9
–
–
No inhibition
No inhibition
No inhibition
–
No inhibition
–
–
–
–
18.5
20
–
8
–
–
18
13
12
19
18
8
35
Fungal Diversity
Fig. 139 Checklist of fungi
associated with C. odorata in
each class of Ascomycota
Fig. 140 Checklist of fungi
associated with C. odorata in
each class of Basidiomycota
13
Fungal Diversity
Fig. 141 Fungal taxa associated
with C. odorata in each order of
Dothideomycetes
Fig. 142 Fungal taxa associated
with C. odorata in each order of
Sordariomycetes
eupatoriella, P. eupatorii, P. eupatorii-formosani, and
Pseudocercospora sp. The genus is generally found as
plant pathogens from leaf lesions, and belongs to the family
Mycosphaerellaceae (Den Breeyen et al. 2006). Most previous studies on Siam weed focused on fungal pathogens from
symptomatic leaves, to assess the potential for biological
control (Barreto and Evans 1994), while our study focused
on saprobic fungi on dead stems. Thus, we only found two
overlapping species, Torula chromolaenae, and T. fici, which
have been previously recorded from dead stems of Siam
weed in Thailand as saprotrophs. The use of culture-independent approaches based on Next Generation Sequencing
13
techniques may further increase understanding of the Siam
weed mycobiome and community structure, together with
corresponding environmental factors and functional groups.
Acknowledgements The Sanger sequencing cost was funded by personal research budgets of W. Purahong and T. Wubet from the UFZHelmholtz Centre for Environmental Research. Ausana Mapook
was financially supported by Research and Researchers for Industry
Program (RRI) PHD57I0012, Thailand and the German Academic
Exchange Service (DAAD) for a joint TRF-DAAD (PPP 2017–2018)
academic exchange grant to Kevin D. Hyde and Marc Stadler. Kevin
D. Hyde thanks to the 2019 high-end foreign expert introduction plan
to Kunming Institute of Botany (granted by the Ministry of Science
and Technology of the People’s Republic of China, Grant Number
Fungal Diversity
G20190139006), Thailand Research grants entitled Biodiversity, phylogeny and role of fungal endophytes on above parts of Rhizophora
apiculata and Nypa fruticans (Grant No: RSA5980068), the future
of specialist fungi in a changing climate: baseline data for generalist
and specialist fungi associated with ants, Rhododendron species and
Dracaena species (Grant No: DBG6080013), Impact of climate change
on fungal diversity and biogeography in the Greater Mekong Subregion
(Grant No: RDG6130001). Ausana Mapook also thanks to Shaun Pennycook, Katalee Jariyavidyanont, Dhanushka N. Wanasinghe, Ishani
D. Goonasekara, Chayanard Phukhamsakda, Saowaluck Tibpromma,
Sirinapa Konta and Chanokned Senwanna for their valuable suggestions and help.
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Affiliations
Ausana Mapook1,2,3,4 · Kevin D. Hyde1,3,5 · Eric H. C. McKenzie6 · E. B. Gareth Jones7 · D. Jayarama Bhat8 ·
Rajesh Jeewon9 · Marc Stadler4 · Milan C. Samarakoon3 · Maitree Malaithong2 · Benjawan Tanunchai2 ·
François Buscot2,11 · Tesfaye Wubet2,10,11 · Witoon Purahong2
1
6
Manaaki Whenua-Landcare Research, Private Bag 92170,
Auckland, New Zealand
7
Department of Soil Ecology, UFZ-Helmholtz Centre
for Environmental Research, Theodor-Lieser-Str. 4,
06120 Halle (Saale), Germany
Department of Botany and Microbiology, College of Science,
King Saud University, P.O Box 2455, Riyadh 11451,
Kingdom of Saudi Arabia
8
3
No. 128/1-J, Azad Co-Op Housing Society, Curca,
Goa Velha 403108, India
Center of Excellence in Fungal Research, Mae Fah Luang
University, Chiang Rai 57100, Thailand
9
4
Department of Health Sciences, Faculty of Science,
University of Mauritius, Reduit 80837, Mauritius
Department Microbial Drugs, Helmholtz Centre for Infection
Research, and German Centre for Infection Research (DZIF),
Partner Site Hannover-Braunschweig, Inhoffenstrasse 7,
38124 Brunswick, Germany
10
Present Address: Department of Community Ecology,
UFZ-Helmholtz Centre for Environmental Research,
Theodor-Lieser-Str. 4, 06120 Halle (Saale), Germany
5
Institute of Plant Health, Zhongkai University of Agriculture
and Engineering, Haizhu District, Guangzhou 510225,
People’s Republic of China
11
German Centre for Integrative Biodiversity Research (iDiv),
Halle-Jena-Leipzig, Deutscher Platz 5e, 04103 Leipzig,
Germany
2
Key Laboratory for Plant Biodiversity and Biogeography
of East Asia (KLPB), Kunming Institute of Botany, Chinese
Academy of Science Kunming, Kunming 650201, Yunnan,
People’s Republic of China
13