Fungal Diversity
DOI 10.1007/s13225-011-0146-5
Phyllosticta—an overview of current status
of species recognition
Saowanee Wikee & Dhanushka Udayanga & Pedro W. Crous & Ekachai Chukeatirote &
Eric H. C. McKenzie & Ali H. Bahkali & DongQin Dai & Kevin D. Hyde
Received: 17 October 2011 / Accepted: 18 October 2011
# Kevin D. Hyde 2011
Abstract Phyllosticta is an important coelomycetous plant
pathogenic genus known to cause leaf spots and various
fruit diseases worldwide on a large range of hosts. Species
recognition in Phyllosticta has historically been based on
morphology, culture characters and host association. Although there have been several taxonomic revisions and
enumerations of species, there is still considerable confusion when identifying taxa. Recent studies based on
molecular data have resolved some cryptic species and
S. Wikee : D. Udayanga : E. Chukeatirote : D. Dai : K. D. Hyde
Institute of Excellence in Fungal Research,
Mae Fah Luang University,
Chiang Rai 57100, Thailand
S. Wikee : D. Udayanga : E. Chukeatirote : D. Dai : K. D. Hyde
School of Science, Mae Fah Luang University,
Chiang Rai 57100, Thailand
P. W. Crous
CBS−KNAW Fungal Biodiversity Centre,
Uppsalalaan 8,
3584 CT Utrecht, The Netherlands
E. H. C. McKenzie
Landcare Research,
Private Bag 92170, Auckland Mail Centre,
Auckland 1142, New Zealand
D. Udayanga
State Key Laboratory of Mycology, Institute of Microbiology,
Chinese Academy of Sciences,
No 3 1st West Beichen Road, Chaoyang District,
Beijing 100101, People’s Republic of China
A. H. Bahkali : K. D. Hyde (*)
College of Science, Botany and Microbiology Department,
King Saud University,
Riyadh, Saudi Arabia
e-mail: kdhyde3@gmail.com
some novel taxa have been discovered. However, compared to
the wide species diversity and taxonomic records, there is a
lack of molecular studies to resolve current names in the
genus. A phylogenetic tree is here generated by combined
gene analysis (ITS, partial actin and partial elongation factor
1α) using a selected set of taxa including type-derived
sequences available in GenBank. Life modes, modal lifecycle
and applications of the genus in biocontrol and metabolite
production are also discussed. We present a selected set of taxa
as an example of resolved and newly described species in the
genus and these are annotated with host range, distribution,
disease symptoms and notes of additional information with
comments where future work is needed.
Keywords Biocontrol . Endophyte . Guignardia . Leaf
spot . Morphology . Molecular phylogeny . Secondary
metabolites
Introduction
The genus Phyllosticta Pers. ex Desm. is a taxonomically
confused group of microfungi comprising mostly important
phytopathogens with a wide host range (van der Aa 1973;
van der Aa and Vanev 2002). Although the generic concept
of Phyllosticta has been refined and species names were
enumerated in a monographic treatment by van der Aa and
Vanev (2002), species recognition still remains problematic
(Hyde et al. 2010a, b; Glienke et al. 2011). Several species
of Phyllosticta have also been reported as endophytes and
saprobes (van der Aa and Vanev 2002; Baayen et al. 2002;
Okane et al. 2003; Wulandari et al. 2009, 2010; Glienke et
al. 2011). Species of Phyllosticta (teleomorph Guignardia
Viala & Ravaz) cause leaf spot symptoms and fruit diseases
on a range of hosts including some economically important
Fungal Diversity
crops and ornamentals such as citrus, banana, apple, grapes,
cranberry, orchids, Ficus sp., Buxus sp. and maple (Uchida
and Aragaki 1980; Paul and Blackburn 1986; Baayen et al.
2002; McManus 1998; Olatinwo et al. 2003; Paul et al.
2005; Liu et al. 2009). Phyllosticta species are also
potential biocontrol agents (Yan et al. 2011), and have
been reported to produce novel bioactive metabolites such
as phyllostine and phyllostoxin (Evidente et al. 2008a).
Molecular data has improved the knowledge of species
relationships and taxonomic classifications in the recent
decade with reference to different complex groups of plant
pathogenic fungi (Crous and Groenewald 2005; Shenoy et
al. 2007; Rossman and Palm-Hernández 2008; Cai et al.
2011; Udayanga et al. 2011). Similarly, Phyllosticta (and its
sexual Guignardia state) is an important genus requiring
modern revisionary treatment employing morphological
characters and a molecular phylogenetic approach, as the
understanding of species is less advanced, but molecular
data are expected to reveal cryptic novel species (Crous and
Groenewald 2005; Hyde et al. 2010a, b). The objectives of
this review are to (1) review the taxonomic and nomenclatural history of Phyllosticta (2) review studies of species
recognition by morphological and molecular phylogenetic
criteria (3) discuss the life styles and biology of species of
the genus (4) summarize the applications of some species in
metabolite production and biocontrol, and (5) provide notes
on selected resolved and recently described species.
History
The genus Phyllosticta Pers. was established by Persoon
(1818) when he introduced the generic name Phyllosticta
for Sphaeria lichenoides DC. Over the past 200 years
numerous species have been added to the genus, often
based on host association so that more than 3,100 names
have been associated with Phyllosticta at various times
(http://www.indexfungorum.org/names/Names.asp: available 21 Sep, 2011). Desmazieres (1847) validated Phyllosticta Pers., and Donk (1968) designated Phyllosticta
convallariae Pers. as the type species. Many of the 3,100
names do not refer to what is now considered to be
Phyllosticta sensu stricto. Initially, many fungi with
unicellular conidia, similar to those of Phoma were named
as either Phoma or Phyllosticta, depending on the location
of conidiomata on the host. Those fruiting on leaves were
described as Phyllosticta while those occurring on other
parts of the plant were placed in Phoma. van der Aa (1973)
provided a key for 46 species of Phyllosticta he accepted,
and this has been widely followed. Furthermore, in their
monographic study, van der Aa and Vanev (2002) accepted
190 species in this genus, while Kirk et al. (2008) estimated
that there are only 92 Phyllosticta species. Since the
revision by van der Aa and Vanev (2002), a further nine
new species have been described (Box 1).
Box 1 History of the study of Phyllosticta
Year
1818
1847
1849
1968
1927
1973
2002
2003-2011
Event
Phyllosticta was introduced
the generic name for
Sphaeria lichenoides DC.
Phyllosticta Pers. was validated
The genus Phyllosticta Pers.
ex Desm typified with
Phyllosticta cruenta
(Kunze ex Fr.) Kickx (1849).
Phyllosticta convallariae Pers.
designated as the type species
A compilation of Phyllosticta
names published
Phyllosticta outline with 46
accepted species
A further revision of the species
described in Phyllosticta with
notes on each of the 191
accepted species
P. ardisiicola Motohashi, I.
Araki & C. Nakash.
P. aspidistricola Motohashi,
I. Araki & C. Nakash.
P. kerriae Motohashi,
I. Araki & C. Nakash.
P. fallopiae Motohashi,
I. Araki & C. Nakash.
P. citriasiana Wulandari,
Crous & Gruyter.
Phyllosticta (Guignardia)
musicola N.F. Wulandari, L.
Cai & K. D. Hyde.
P. bifrenariae O.L. Pereira,
C. Glienke & Crous.
P. brazilianiae D. Stringari,
C. Glienke & Crous.
P. citribraziliensis
C. Glienke & Crous.
Phyllosticta citrichinaensis H.X.
Wang, K.D. Hyde & H.Y. Li
References
Persoon (1818)
Desmazieres
(1847)
Kickx (1849)
Donk (1968)
Petrak and
Sydow, (1927)
Van der Aa
(1973)
Van der Aa and
Vanev (2002)
Motohashi et al.
(2008)
Wulandari et al.
(2009)
Wulandari et al.
(2010)
Glienke et al.
(2011)
Wang et al. 2011
The teleomorph state of Phyllosticta is Guignardia Viala
& Ravaz, represented by the generic type, Guignardia
bidwellii (Ellis) Viala & Ravaz. There are 335 epithets
associated with this genus (http://www.indexfungorum.org/
names/Names.asp) and in a similar fashion to Phyllosticta,
most species where described based on host association.
The genus has not been well-studied, and there is no
comprehensive monograph, although individual species or
groups of species have been treated (Hyde 1995; Motohashi
et al. 2009; Wulandari et al. 2011). Most species of
Phyllosticta and Guignardia have been described indepen-
Fungal Diversity
dently from each other, and only a few Phyllosticta species
have been linked to their Guignardia teleomorphs. Consequently, their classification is confusing and the relation
between host range and disease are often poorly understood
(van der Aa 2002)
(2011). Because of this decision we use the name Phyllosticta throughout this review unless we specifically refer to a
Guignardia species. Leptodothiorella, which previously
represented the spermatial state of some Phyllosticta species
(e.g. Leptodothiorella aesculicola (Sacc.) Sivan.), are also
treated as synonyms of Phyllosticta (van der Aa 1973).
Using Phyllosticta versus Guignardia
Morphological characteristics to differentiate species
The name Phyllosticta (asexual state) and Guignardia (sexual
state) have been used separately following the dual classification system used by mycologists over several decades
(Hawksworth 2004; McNeil et al. 2006; Shenoy et al. 2007,
2010). For instance Phyllosticta musarum (Cooke) Aa and
Guignardia musae Racib. are the same biological species but
have different names, P. musarum being the asexual state and
G. musae being the sexual state (van der Aa 1973;
Wulandari et al. 2010). However with the use of molecular
data it is now possible to link asexual and sexual states
(Berbee and Taylor 2001) and the use of the dual
nomenclature system of classification in fungi has become
redundant (Hawksworth 2011). Therefore a single name
should be adopted and there are various views to which
names should be followed, i.e. the oldest, the sexual state
name, the most conserved name, and view maintaining both
names (Berbee and Taylor 2001; Seifert and Rossman 2010;
Hyde et al. 2011). Our view is that we should generally
adopt the oldest name for each genus, which will soon be
enforced in the International Code of Nomenclature for
algae, fungi and plants, but also taking into account which
name is the most important and commonly used. Phyllosticta
Pers. (1818) is a much older name than Guignardia Viala &
Ravaz (1892) and generally Phyllosticta species are known
to cause important diseases (e.g. leaf spot, citrus black spot,
black rot of horse chestnut). There are also many more
species of Phyllosticta than Guignardia. There are exceptions, for example Guignardia candeloflamma K.D. Hyde is
only known in it teleomorph state (Wulandari et al. 2010a),
while banana freckle is caused by both states (Wulandari et
al. 2010b). Because Phyllosticta is the oldest name and
generally more important as the causal agent of disease we
chose to adopt this name and treat all Guignardia species as
synonyms of Phyllosticta, in the sense of Glienke et al.
Phyllosticta pycnidia are usually globose, subglobose or
tympaniform, flattened above, and closely connected with the
subepidermal pseudostroma (Fig. 3a, b). They are mostly
unilocular but occasionally may be multilocular (van der Aa
1973). The conidia are commonly hyaline, one-celled, ovoid,
obovate or ellipsoid, or short cylindrical, seldom pyriform,
globose or subglobose, 10–25 μm long, and usually covered
by a slime layer and bearing a single apical appendage
(Fig. 3c) (van der Aa 1973). Cultural characteristics when
grown on specific media may also be used as differentiating
characters. In the case of P. citricarpa colonies can be
characterised after 14 days at 25°C in the dark on OA as flat,
spreading, olivaceous-grey, becoming pale olivaceous-grey
towards the margin, with sparse to moderate aerial mycelium; surrounded by a diffuse yellow pigment in the agar
medium (Wulandari et al. 2009).
The sexual state Guignardia can be characterized by
erumpent ascomata, which are globose to pyriform in
section, often irregularly shaped, unilocular, and with a
central ostiole forming by dehiscence when mature. The
peridium is thin, comprising a few layers of angular cells.
Asci are 8-spored, bitunicate, clavate to broadly ellipsoid, with a wide, slightly square apex, tapering
gradually to a small pedicel, and with a welldeveloped ocular chamber. Ascospores are ellipsoid to
limoniform, sometimes slightly elongated, aseptate,
hyaline, often guttulate or with a large central guttule,
and some have mucilaginous polar appendages (van der
Aa 1973; Wulandari et al. 2011, Fig. 3d–f).
A spermatial state is often present in the life cycle of
Guignardia species, and readily forms in culture. Spermatia
are cylindrical to dumbbell-shaped with guttules at each
end (Fig. 3g). In the past several spermatial states were
Fig. 1 Phyllosticta sp. on living leaf of jackfruit. a Leaf spots. b–c Lesion on adaxial surface. d Banana freckle disease
Fungal Diversity
Fig. 2 Schematic representation of the life cycle of Phyllosticta and its teleomorph redrawn from: http://www.oardc.ohio-state.edu/fruitpathology/
organic/grape/All-Grapes.html
officially named. For instance, the spermatial state of
Melanops concinna Syd. (= Guignardia concinna (Syd.)
Aa; van der Aa 1973) was described as Leptodothiorella
concinna Sydow (1926).
There have also been several molecular phylogenetic
studies concerning Phyllosticta species that have helped
to facilitate the identification of species and resolution of
species complexes (Baayen et al. 2002; Okane et al.
2003; Motohashi et al. 2009; Wulandari et al. 2009;
Glienke et al. 2011).
Molecular studies advance the understanding
of Phyllosticta
ITS rDNA sequence-based studies in Phyllosticta
The rapid development of molecular phylogenetic tools
have improved our understanding of several other
coelomycetous genera such as Colletotrichum (Cai et
al. 2009; Crouch et al. 2009; b; Hyde et al. 2009),
Phomopsis (Santos and Phillips 2009; Udayanga et al.
2011), Phoma (Aveskamp et al. 2008, 2010; de Gruyter
et al. 2010), Fusicoccum (Crous et al. 2006), Diplodia
(Phillips et al. 2008) and Pestalotiopsis (Liu et al. 2010;
Maharachchikumbura et al. 2011), to name but a few.
ITS rDNA sequences are often used to infer phylogenetic
relationships in many groups of fungi including Phyllosticta (Okane et al. 2003; Motohashi et al. 2009; Wulandari
et al. 2009). Motohashi et al. (2009) evaluated the
phylogenetic relationships among Japanese species of
Phyllosticta sensu stricto and its teleomorph Guignardia
using 18S rDNA sequence data. They observed that
Phyllosticta sensu stricto is a monophyletic clade. In the
same study, ITS and 28S rDNA sequences were used in a
Fungal Diversity
Fig. 3 Comparison of Phyllosticta and Guignardia states. a Vertical
section through pycnidium. b Pycnidial wall with conidiogenous cells.
c Conidia. d Section of ascoma. e Bitunicate and fissitunicate ascus. f
Ascospores. g Leptodothiorella spermatial state. Scale Bars: a, b, d, e
= 50 μm, c, f, g =10 μm
phylogenetic analysis of Phyllosticta strains from various
host plants. Results from this study revealed isolates to
cluster in two subgroups based on molecular data as, (1)
cultures from a wide range of host plants mainly derived
as endophytes from symptomless plants (P. capitalensis
complex, see below) and (2) relatively host-specific
strains (often isolated as foliar pathogens from diverse
plants).
ITS-RFLP and ITS sequence analysis were used to
examine genetic variation of foliar endophytic Phyllosticta
strains from different tropical trees (Pandey et al. 2003).
Although ITS-RFLP failed to infer genetic diversity among
Fungal Diversity
isolates used, the ITS phylogram supported the identity of P.
capitalensis as a common foliar endophyte and pathogen
with wide range of hosts. In a similar study, the diversity of
strains of Guignardia (or Phyllosticta) was evaluated using
rDNA ITS sequence data (Okane et al. 2003). Guignardia
endophyllicola (anamorph Phyllosticta capitalensis) was
shown to have an extensive host range. The taxon was
identified in 53 isolates from the same number of different
plants belonging to 43 genera. Phylogeny based on rDNA
ITS sequence analyses derived from 18 tropical endophytic
strains from different plants confirmed conspecificity of the
Brazilian isolates with Phyllosticta captalensis (as G.
mangiferae) (Rodrigues and Sieber. 2004). In the ITS
sequence comparison, some Guignardia and Phyllosticta
strains from unrelated hosts were more closely related than
other isolates derived from closely related plants. However,
the diversity across the wide range of hosts should be
evaluated by incorporating more genes in analyses and
isolating strains from a wide range of hosts in future studies.
The population structure and phylogenetic relationships
of Guignardia citricarpa (citrus black spot) were investigated by Baayen et al. (2002) using ITS, AFLP and
morphological comparison. The observations supported
the historic distinction between slowly growing pathogenic
isolates and fast growing non-pathogenic isolates, which
proved to belong to P. capitalensis (as G. mangiferae), the
ubiquitous endophyte reported in various studies (Okane et
al. 2003; Rodriguez et al. 2009; Glienke et al. 2011).
Numerous synonyms for P. capitalensis have been used in
earlier studies, reviewed in this section. We have used the
name as it appeared in the original publication, although the
need for careful refinement using the currently accepted
name is recommended.
Studies employing multi-locus analyses in Phyllosticta
Identification of Phyllosticta species is problematic as few
characters are available to differentiate species. Although
ITS sequence data have been widely used for species
discrimination, multi-locus phylogenies might resolve
cryptic species (Wulandari et al. 2009). A combined
phylogenetic analysis based on the rDNA ITS, translation
elongation factor 1 (TEF1), and actin (ACT) genes resolved
three species, namely P. mangiferae, P. citricarpa and a
new species, P. citriasiana (Wulandari et al. 2009), the
latter causing tan spot of Citrus maxima in Asia. Glienke et
al. (2011) investigated the genetic diversity of endophytic
and pathogenic Phyllosticta species, with particular emphasis on Phyllosticta citricarpa and Guignardia mangiferae
occurring on Citrus. Combined DNA sequence analysis
based on rDNA ITS, translation elongation factor 1 (TEF1),
actin (ACT) and glyceraldehyde-3-phosphate dehydrogenase (GPDH) genes resolved nine well-supported clades
related to seven known species and two apparently
undescribed species. They have designated epitypes for P.
citricarpa collected from Australia and Phyllosticta capitalensis collected from Brazil (Glienke et al. 2011). Furthermore, P. brazilinae, P. bifinariae and P. citribraziliensis
were described as novel species based on morphology and a
multilocus phylogeny. The combined gene analysis further
revealed that the allocation of various synonyms for the
endophytic, non-pathogenic isolates occurring on wide
range of hosts would be more correctly referred to as P.
capitalensis. Further work is needed, however, to resolve
whether this taxon is a complex of cryptic species.
Multi-locus phylogeny inferred from available
sequences in GenBank
A selected set of ITS rDNA, ACT, and TEF1 sequences,
including the available ex-type, and ex-epitype materials
were downloaded from GenBank (Table 1). The sequences
were aligned using Bioedit, alignment was optimized
manually and the genes are combined to perform phylogenetic analysis. Parsimony analysis was carried out by PAUP
v. 4.0b10 (Swofford 2002). Ambiguously aligned regions
were excluded from all analyses and the gaps were treated
as missing data. Trees were figured in Treeview (Page
1996). One of the most parsimonious trees generated from
combined gene analysis for 35 strains is provided (Fig. 4).
The phylogenetic tree based on GenBank sequences
including sequences originating from seven ex-type cultures,
and other sequences are identified as appeared in recent
publications. We have used 35 isolates in the multilocus
phylogenetic tree, as there is a lack of sequence data of all
three genes for all known ex-type cultures, some of which
therefore had to be excluded from the analyzed dataset (see
Table 1). However we recommend the improvement of the
multi-locus phylogenetic analysis by using more phylogenetically informative genes, and more ex-type isolates in
future work. Incorporation of more ex-type sequences will
accelerate the accurate identification of other species from
various geographical locations and a wide range of hosts.
Ecology of Phyllosticta species
Phyllosticta species are important plant pathogens and,
although taxa are also commonly identified as endophytes
(Baayen et al. 2002; Rodrigues and Sieber 2004), a few
species have also been reported as saprobes. In some cases
a species may occupy more than one life mode. For
example, P. capitalensis was originally described on
Stanhopea (Orchidaceae) from Brazil as a fungal pathogen.
Recently, Silva et al. (2008) reported that P. capitalensis
Species
Straina
Substrate
Country
Collector(s)
ITS
ACT
TEF1
Guignardia bidwellii
CBS 111645
Parthenocissus quinquefolia
USA
G. Carroll
EU683672
–
EU683653-
Guignardia citricarpa
CBS 102345
Citrus aurantium, lesion on peel
Brazil
–
FJ538311
FJ538427
FJ538369
Guignardia citricarpa
CBS 122482
Citrus sinensis
Zimbabwe
L. Huisman
FJ538317
FJ538433
FJ538375
Guignardia citricarpa
CBS 122384
Citrus limon
South Africa
M. Truter
FJ538316
FJ538432
FJ538374
Guignardia citricarpa (ex-epitype)
CBS 127454
Citrus limon
Australia
S.L. Willingham
JF343583
JF343667
JF343604
Guignardia mangiferae
CBS 115046
Myradcrodruon urundeuva, leaf or bark
Brazil
K.F. Rodriques
FJ538322
FJ538438
FJ538380
Guignardia mangiferae
CBS 115047
Aspidosperma polyneuron, leaf or bark
Brazil
K.F. Rodriques
FJ538323
FJ538439
FJ538381
Guignardia mangiferae
CBS 114751
Spondias mombin, leaf or bark
Brazil
K.F. Rodriques
FJ538349
FJ538465
FJ538407
Guignardia mangiferae
CBS 115049
Bowdichia nitida, leaf or bark
Brazil
K.F. Rodriques
FJ538324
FJ538440
FJ538382
Guignardia mangiferae
IMI 260576
Mangifera indica, leaf endophyte
India
M.V. Leksshmi
JF261459
JF343641
JF261501
Guignardia psidii
CBS 100250
Psidium guajava, fruit
Brazil
C. Glienke
FJ538351
FJ538467
FJ538409
Guignardia vaccinii
CBS 126.22
Oxycocus macrocarpus
U.S.A
–
FJ538353
FJ538469
FJ538411
Phyllosticta bifrenariae(ex-type)
VIC30556; CBS 128855
Bifrenaria harrisoniae, living leaf
Brazil
O. Pereira
JF343565
JF343649
JF343586
Phyllosticta brazilianiae
LGMF 333
Mangifera indica, leaf endophyte
Brazil
C. Glienke
JF343574
JF343658
JF343595
Phyllosticta brazilianiae
LGMF 334
Mangifera indica, leaf endophyte
Brazil
C. Glienke
JF343566
JF343650
JF343587
Phyllosticta brazilianiae(ex-type)
CBS 126270
Mangifera indica, leaf endophyte
Brazil
C. Glienke
JF343572
JF343656
JF343593
Phyllosticta capitalensis
CBS 100175
Citrus sp., healthy leaf
Brazil
C. Glienke
FJ538320
FJ538436
FJ538378
Phyllosticta capitalensis
CBS 123373
Musa paradisiaca
Thailand
N. F. Wulandari
FJ538341
FJ538457
FJ538399
Phyllosticta capitalensis (ex-epitype)
CBS 128856
Stanhopea graveolens
Brazil
O.L. Pereira
JF261465
JF343647
JF261507
Phyllosticta citriasiana
CBS 120488
Citrus maxima
Thailand
J. de Gruyter
FJ538354
FJ538470
FJ538412
Phyllosticta citriasiana
CBS 123393
Citrus maxima
Vietnam
J. de Gruyter
FJ538358
FJ538474
FJ538416
Phyllosticta citriasiana
CBS 123372
Citrus maxima
Vietnam
J. de Gruyter
FJ538357
FJ538473
FJ538415
Phyllosticta citriasiana (ex-type)
CBS 120486
Citrus maxima
Thailand
J. de Gruyter
FJ538360
FJ538476
FJ538418
Phyllosticta citribraziliensis (ex-type)
CBS 100098
Citrus sp., healthy leaves
Brazil
C. Glienke
FJ538352
FJ538468
FJ538410
Phyllosticta citribraziliensis
LGMF09
Citrus sp., healthy leaves
Brazil
C. Glienke
JF261436
JF343618
JF261478
Phyllosticta citricarpa
CBS 122348
Citrus sinensis, lesions on fruit
Zimbabwe
L. Huisman
FJ538315
FJ538431
FJ538373
Phyllosticta citricarpa
CBS 127455
Citrus sinensis
Australia
S.L. Willingham
JF343584
JF343668
JF343605
Phyllosticta citricarpa (ex-epitype)
CBS 127454
Citrus limon
Australia
S.L. Willingham
JF343583
JF343667
JF343604
Phyllosticta cussonia
CPC 14873
Cussonia sp.
South Africa
P.W. Crous
JF343579
JF343663
JF343600
Phyllosticta cussonia
CPC 14875
Cussonia sp
South Africa
P.W. Crous
JF343578
JF343662
JF343599
Phyllosticta hypoglossi
CBS 101.72
Ruscus aculeatus, living leaf
Italy
W. Gams
FJ538365
FJ538481
FJ538423
Phyllosticta hypoglossi
CBS 434.92
Ruscus aculeatus
Italy
W. Gams
FJ538367
FJ538483
FJ538425
Phyllosticta hypoglossi
CBS 167.85
Ruscus hypoglossum
Italy
W. Gams
FJ538366
FJ538482
FJ538424
Phyllosticta owaniana
CBS 776.97
Brabejum stellatifolium
South Africa
A. den Breeÿen
FJ538368
FJ538484
FJ538426
Phyllosticta spinarum
CBS 292.90
Chamaecyparis pisifera
France
M. Morelet
JF343585
JF343669
JF343606
Phyllosticta spinarum
CBS 937.70
Hedera helix, leaf litter
Italy
W. Gams
FJ538350
FJ538466
FJ538408
a
CBS: Centraalbureau voor Schimmelcultures, Utrecht, The Netherlands; CPC: Culture collection of Pedro Crous, housed at CBS; IMI: International Mycological Institute, CABI-Bioscience,
Egham, Bakeham Lane, UK; LGMF: Culture collection of Laboratory of Genetics of Mycroorganisms, Federal University of parana, Curitiba, Brazil; VIC: Culture Collection of Federal
University of Vicosa, Vicosa, Brazil.
Fungal Diversity
Table 1 Details of Guignardia and Phyllosticta isolates and GenBank accession number of their sequence data
Fungal Diversity
CBS 127454b G. citricarpa
CBS 127454a P. citricarpa
CBS 127455 P. citricarpa
CBS 122384 G. citricarpa
100 CBS 122348 P. citricarpa
CBS 122482 G. citricarpa
CBS102354 G. citricarpa
100
CBS 120488 P. citriasiana
CBS 123393 P. citriasiana
100
CBS 123372 P. citriasiana
CBS 120486 P. citriasiana
100
85
CBS 434.92 P. hypoglossi
CBS 101.72 P. hypoglossi
CBS 167.85 P. hypoglossi
71
100 CPC 14873 P. cussonia
CPC 14875 P. cussonia
85
CBS 126.22 G. vaccinii
100
100 CBS100098 P. citribraziliensis
80
87
LGMF09 P. citribraziliensis
CBS 937.70 P. spinarum
CBS 292.90 P. spinarum
CBS 128855 P. bifrenariae
CBS 115046 G. mangiferae
100
CBS 115049 G. mangiferae
CBS 115047 G. mangiferae
CBS 123373 P. capitalensis
94
98
CBS 100175 P. capitalensis
CBS 128856 P. capitalensis
CBS 114751 G. mangiferae
99
CBS 100250 G. psidii
IMI 260576 G. mangiferae
87
CBS 126270 P. brazilianiae
100
LGMF 374 P. brazilianiae
LGMF 333 P. brazilianiae
CBS776.97 P. owaniana
10
Fungal Diversity
4 Phylogram generated from the parsimony analysis based on
combined rDNA ITS, ACT, TEF1 sequence data for Phyllosticta spp.
downloaded from GenBank. Strictly consensus branches are thickened
and bootstrap support values >70% are shown below or above the
branch. Cultures derived from type specimens are in bold. The tree is
rooted with Phyllosticta owaniana
Fig.
caused disease of leaves and pseudobulbs of Bifrenaria
harrisoniae (Orchidaceae) in Brazil. Phyllosticta capitalensis has also been reported as an endophyte on ericaceous
plants in Japan by Okane et al. (2001), and non-pathogenic
strains have been isolated from Citrus sp. (Table 2) (Baayen
et al. 2002; Glienke et al. 2011).
Host specificity of Phyllosticta species and disease
symptoms
Phyllosticta species may cause leaf spots on many plant
species and it is not clear if they are generalists or hostspecific; this may depend on the particular species or their
life style. It is known that several species that cause
diseases are host, genus or family specific, while endophytes may be generalists. For example, P. sphaeropsoidea
Ellis & Everh. causes leaf blotch disease specific to horse
chestnut in Europe and North America (Hudson 1987). P.
citricarpa causes leaf spot disease of Citrus species, while
P. citriasiana can infect fruits of Citrus maxima (pomeloes),
and causes tan spot and has only been isolated from
pomeloes, but has never been found from lemons, mandarins and oranges, and Phyllosticta musarum is specific on
Musa spp. (Wulandari et al. 2009, 2010, 2011). P. capitalensis is an endophyte of a wide range of hosts (Okane et al.
2003; Baayen et al. 2002).
Knowledge of disease symptoms on hosts are important
for field identification by taxonomists as well as plant
pathologist interested in disease occurrence, management
and distribution. Generally, Phyllosticta species cause
necrotic lesions on leaves, which are characteristically
small, often 1–2 mm in diameter, circular, brown in the
middle and dark brown or sometimes reddish at the margin
(Fig. 1a). One to more than 10 pycnidia are often found in
one lesion (Fig. 1b, c). Pycnidia on leaves are usually black,
globose or subglobose, and semi-immersed (Fig. 1c). After
infection by Phyllosticta the leaf may become dry in the
centre of the lesion, causing the infected tissue to drop out,
forming a hole, and hence this is known as target spot or
shot hole spot. Leaf spots often occur in living leaves in the
late dry and wet seasons or in winter in temperate countries.
There are four types of leaf spot symptoms—hard spot,
false melanose, freckle spot and virulent spot (Kotzé 2000).
In July 1984, Phyllosticta species were the cause of
problems on Muehlenbeckia adpressa in Victoria, Australia. Virtually all mature leaves of plants contained
distinctive necrotic spots for an area of 10 sq. m. Spots
were roughly circular to elliptical in shape and were tan
with a maroon margin (Paul and Blackburn 1986). Freckle
disease occurs on several species and varieties of banana
(Fig. 1d). Characteristic spots (pycnidia and ascomata) form
on fruit, giving the lesion a sandpaper texture. Leaves of
banana will turn yellow when infected with this fungus
(Wulandari et al. 2010).
Phyllosticta species as endophytes
Endophytes are fungi that asymptomatically colonize plant
tissues during some phase of their life cycle (Petrini 1991;
Hyde and Soytong 2008; Saikkonen 2007), but may turn
pathogenic during host senescence (Rodriguez and Redman
2008; Rodriguez et al. 2009). The relationship may be
symbiotic, antagonistic, neutral or mutualistic (Hyde and
Soytong 2008; Aly et al. 2011). Endophytes are horizontally transmitted, and transfer to their host plants via
airborne spores. However, some endophytes may also be
vertically transmitted to the next host plant generations via
seeds (Hartley and Gange 2009). Although the first
discovery of endophytes dates already back to 1904, they
did not receive considerable attention until the recent
recognition of their pharmaceutical and ecological significance (Gunatilaka 2006). Recent development of screening
technologies revealed the great potential of endophytes as a
major source of biologically active compounds (Strobel and
Table 2 Phyllosticta species recorded as endophytes in selected studies
Species
Host
Country
Plant organ
Reference
P. bifrenariae
P. brazilianiae
P. capitalensis
P. ilicina
P. spinarum
Phyllosticta sp.
Phyllosticta sp.
Orchidaceae
Anacardiaceae
Various hosts, woody plant
Quercus ilex (Fagaceae)
Platycladus orientalis (Cupressaceae)
Ginkgo biloba (Ginkgoaceae)
Abies grandis (Pinaceae)
Brazil
Brazil
New Zealand
Switzerland
USA
Japan
USA
Leaf and bulb
Leaf and fruit
Leave and fruit
Leaves or needles
Leaves
Leaf, petiole, twigs
Leaves or needles
Glienke et al. (2011)
Glienke et al. (2011)
Baayen et al. (2002), Glienke et al. (2011)
Collado et al. (1996)
Wijeratne et al. (2008)
Thongsandee et al. (2011)
Carroll and Carroll (1978)
Fungal Diversity
Daisy 2003; Huang et al. 2009; Xu et al. 2010; Tan and Zou
2001). Investigations related to endophytic microorganisms
in plants and especially tropical hosts have increased, due
to the significance of using endophytes in biological control
and the discovery of biologically active compounds
(Wijeratne et al. 2008; Le Calvé et al. 2011).
Although Phyllosticta species have been reported as
endophytes there are relatively few reports of Phyllosticta
species being recorded as endophytes in recent studies. In
two volumes of the journal Fungal Diversity (Volume 41,
2010, Volume 47, 2011) there were 13 manuscripts devoted
to biodiversity of fungal endophytes and only one (Lin et
al. 2010) reported an endophytic Guignardia species.
Phyllosticta capitalensis however has commonly been
recorded as an endophyte in several studies (Baayen et al.
2002; Glienke et al. 2011, Okane et al. 2001; Okane et al.
2003) and was reported as an endophyte on more than 20
hosts in eight countries (Wulandari et al. 2010; Glienke et
al. 2011). Therefore the species is thought to be one of the
most common endophytic species of Phyllosticta (Glienke
et al. 2011). There are few records of other Phyllosticta
species recorded as endophytes and they are usually listed
as unidentified Phyllosticta sp. (Pandey et al. 2003). Some
Phyllosticta species reported as endophytes are listed in
Table 2.
Phyllosticta species as saprobes
Most fungi have the ability to grow as saprobes, and
degrade organic material from dead plant material as a food
source. Plant pathogenic fungi can often survive as saprobes between growing seasons (Trigiano et al. 2004). For
example, Phyllosticta carpogena and P. ericae occurred as
saprobes on Rubus sp. (Rosaceae) and Erica carnea
(Ericaceae), respectively (van der Aa and Vanev 2002)
(Table 3).
Life cycle
Concepts concerning life cycles of plant pathogens may
have significant practical consequences for plant patholo-
gists and taxonomists. Herein, we provide a schematic
diagram of the life cycle of a typical species of Phyllosticta
(Phyllosticta ampelicida, Fig. 2). After infection by the
Phyllosticta or Guignardia propagules, pycnidia and/or
ascomata develop under the leaf tissue, and produce leaf
spots on the host. During the wet season conidia and
ascospores, and sometimes the spermatial stage are present.
Subsequently spores are released and ejected from the
pycnidia and ascomata. The spores are carried by rain and
wind to other leaves and young fruits. Germ tubes develop
from spores and grow into leaves and develop within the
plant tissue. The disease spreads by transmission by warm
wind and rain during the wet season.
Secondary metabolites from species of Phyllosticta
Fungi are well-known as a good source of important
metabolites, some of which are useful in the pharmocological industry and agriculture (Pearce 1997, Smith and
Casey 2008, Aly et al. 2010; Xu et al. 2010; Udayanga et
al. 2011). Both novel and previously known metabolites
have been isolated from species of Phyllosticta (Tables 4,
5). Metabolites produced by Phyllosticta species include
phyllostin and phyllostoxin. Phyllostictines A–D that were
isolated from P. cirsii (Evidente et al. 2008b). Phytotoxins,
including phyllosinol, brefeldin, and PM-toxin (Sakamura
et al. 1969; Sakai et al. 1970) were extracted from
Phyllosticta maydis (Comstock et al. 1973) and Phyllosticta
medicaginis (Entwistle et al. 1974), respectively. Phyllostictines A–D have been tested with on five cancer cell lines
which displayed growth-inhibitory activity (Le Calvé et al.
2011). In addition, five new metabolites were isolated
from P. spinarum, reported by Wijeratne et al. (2008)
namely (+)-(5S,10S)-4′hydroxymethylcyclozonarone, 3ketotauranin, 3-hydroxytauranin, 12-hydroxytauranin,
phyllospinarone.
Taxol was initially known as a phytochemical derived
from the bark of Taxus brevifolia (Western Yew) and is an
expensive and important diterpenoid anti-cancer intensive
treatment drug used against breast, ovarian and lung
cancers (Wani et al. 1971). It has been reported that the
molecule has anti-tumour activity in several experimental
Table 3 Phyllosticta species recorded as saprobes in selected studies
Species
P.
P.
P.
P.
acetosellae A.L. Sm. & Ramsb.
capitalensis
cocoicola
pyrolae Ellis & Everh.
Host
Country
Plant organ
References
Rumicis acetosellae
Magnolia liliifera
Palm
Pyrola rotundiforia
England
Thailand
Europe
America
Leaves
Senescent leaves
Leaves
Leaves
Smith and Ramsbottom (1913)
Okane et al. (2003)
Punithalingam (1974), Taylor and Hyde (2003)
Ellis and Everhart (1889)
Fungal Diversity
Table 4 Metabolites produced by Phyllosticta species
Compound
Properties
Name of taxa as in publication
References
Befeldin
Phyllosinal
Bioactive metabolite
Bioactive metabolite
P. medicaginis
Phyllosticta. sp., P. maydis
Phyllostictine
Phyllostin
Mycoherbicide
Anti-microbial, anti-cancer
P. cirsii
P. cirsii
Phyllostoxin
PM-toxin
Tauranine
Taxol
(+)-(5S,10S)-4′ hydroxymethylcyclozonarone
3-ketotauranin
3-hydroxytauranin
12-hydroxytauranin
Phyllospinarone
Mycoherbicide
Mycoherbicide
Anti-cancer activity
Anti-cancer activity
Inhibition of cell proliferation
Inhibition of cell proliferation
Inhibition of cell proliferation
Inhibition of cell proliferation
Inhibition of cell proliferation
P.
P.
P.
P.
P.
P.
P.
P.
P.
Entwistle et al. (1974)
Sakamura et al.(1969),
Sakai et al. (1970)
Evidente et al. (2008a)
Evidente et al. (2008a),
Le Calvé et al. (2011)
Evidente et al. (2008a)
Comstock et al. (1973)
Wijeratne et al. (2008)
Kumaran et al. (2009a)
Wijeratne et al. (2008)
Wijeratne et al. (2008)
Wijeratne et al. (2008)
Wijeratne et al. (2008)
Wijeratne et al. (2008)
trials. Taxol is produced by various fungal strains of
Pestalotiopsis (Strobel et al. 1996, 1997), Phomopsis
(Kumaran and Hur 2009) and Phyllosticta (Kumaran et al.
2008a, b, 2009a) in culture media under various conditions.
Taxol has been reported from P. citricarpa, from Citrus
medica and P. dioscoreae from Hibiscus rosa-sinensis
(Kumaran et al. 2008a, 2009b). Species of Phyllosticta are
therefore potential sources for discovery of pharmaceutical,
medical and agricultural novel compounds.
cirsii
maydis
spinarum
tabernaemontanae
spinarum
spinarum
spinarum
spinarum
spinarum
that had high affect on motility of the second stage juveniles
of Meloidogyne incognita and has potential in parasitic
nematode control (Yan et al. 2011). P. cirsii, a pathogen
isolated from diseased leaves of Cirsium arvense has been
evaluated as a potential biocontrol agent of this noxious
perennial weed, also produces different phytotoxic metabolites with potential herbicidal activity when grown in liquid
cultures (Evidente et al. 2008a). The metabolites reported are
Phyllostictines A–D which are potential mycoherbicides
(Berestetskiy et al. 2008; Evidente et al. 2008a,b).
Phyllosticta in biocontrol
Need for epitypification of Phyllosticta species
Biocontrol is “the control of unwanted organisms such as
weeds by the use of other organisms, as through the use of
organisms that are natural predators, parasites, or pathogens
(http://www.answers.com/topic/biological-pest-control#ixz
z1YSqa0e00)” Fungi are commonly used as biological
control agents (Charudattan and Dinoor 2000, Mortensen
1998; Trujillo 2005; Rosskopf et al. 2000). Phyllosticta
species may have potential for use as biocontrol agents
although there are presently few examples (Table 5). A strain
of Phyllosticta (Ph511) was shown to produce compounds
An epitype is a fresh specimen (usually with accompanying
culture) selected to serve as a representative type when such
authentic material has been recollected, and confirmed to
represent the same species as the original type material (Phillips
et al. 2006; Hyde et al. 2010a, b; Abd-Elsalam et al. 2010).
This practice enables mycologists to link older names to DNA
data derived from fresh collections. The sole purpose of
epitypification, is to move mycology into the culture and
DNA era. When an epitype is designated the original material
Table 5 Phyllosticta species used in biocontrol
Species
Host
Used against
References
Phyllostictine A−D
Cirsium arvense (weed)
Evidente et al. (2008b)
Phyllostin (8-hydroxy-3-methylCirsium arvense
Tuzi et al. (2010)
2-oxo-2,3,4a, 5,8,8a-hexahydrobenzo[1,4]dioxine-6-carboxylic
acid methyl ester)
Curcumis sativus Ph511
Meloidogyne incognita root knot nematode Yan et al. (2011)
Phyllosticta cirsii Cirsium arvense
Phyllosticta sp.
Cirsium arvense
Phyllosticta sp.
Compound name
Fungal Diversity
that the epitype supports must be explicitly cited. Several
authors have discussed the urgent need for epitypication in
plant pathogenic genera (Verkley et al. 2004; Crous 2005;
Crous et al. 2007, Shenoy et al. 2007; Hyde et al. 2010a, b;
Cai et al. 2011). Since few species of Phyllosticta have been
epitypied or have ex-type cultures available it is imperative
that pathogenic species are recollected and epitypified as has
been done for other plant pathogenic genera.
Notes on selected species of Phyllosticta
Molecular data has to date proven to be inadequate in resolving
many species in the genus Phyllosticta (Wulandari et al. 2009).
Either there are very few species of Phyllosticta with some
having a very wide host range, or the genes that we are
presently using do not resolve species complexes. Differentiation of the 192 species accepted by van der Aa (1973) and
van der Aa and Vanev (2002) was based on morphological
data with often minor differences, and molecular evidence is
not available to support this differentiation. Most species also
lack living cultures and their uniqueness cannot be confirmed.
For this reason it is not possible to list which species can be
stated as currently in use in Phyllosticta, as has been done for
Cochliobolus (Manamgoda et al. 2011), Colletotrichum (Hyde
et al. 2009), Fusarium (Summerell et al. 2010; Summerell and
Leslie 2011) and Phomopsis (Udayanga et al. 2011).
Below we discuss alphabetically selected Phyllosticta
names which includes the generic type, an earlier name for
the generic type, plant pathogens and endophytes resolved
using molecular data, all taxa introduced since van der Aa
and Vanev (2002) and some other taxa that we believe
warrant discussion and is partly based on the most recent
literature. This cannot be considered as a thorough account
of the generic species, but provides a starting point towards
establishing the number of acceptable species in future
revisionary treatments of Phyllosticta as phylogenetic data
accumulates and helps to resolve the species.
The account of selected species names provides authorities and publication details as appeared in Index Fungorum
(http://www.indexfungorum.org/names/Names.asp). Synonyms are not given as these can be searched for in Index
Fungorum. The teleomorph is given where known. This is
not a rigorous list as it is impossible to verify at this stage
whether collections of each taxon on a host are correctly
identified. We have annotated the notes with host range,
symptoms and known distribution, and additional notes on
pathological, taxonomic and phylogenetic research. Also
the additional notes emphasize the need for molecular data
in future studies. We recommend that other resolved species
are added to this selected list based on future studies of
Phyllosticta employing molecular and morphological data
as has been done in Colletotrichum (Phoulivong 2011).
When referring to Phyllosticta species one also has to
consider the teleomorph Guignardia. However, although
there is molecular data for this teleomorphic genus, it is
mostly for the endophyte incorrectly identified as G.
mangiferae (e.g. P. capitalensis). We therefore do not
discuss Guignardia species below. However, Guignardia
psidii Ullasa & Rawal, was shown to be distinct in the
molecular analysis of Wang et al. (2011). This strain (CBS
100250) was isolated from a fruit of Psidium guajava
collected from Sao Paulo, Brazil. The Phyllosticta state is
unknown. Several new species of Guignardia have also
been described in recent years (e.g. G. musicola N.F.
Wulandari, L. Cai & K.D Hyde, G. bispora N.F. Wulandari
& K.D. Hyde, Wulandari et al. 2010, 2011) and these also
need recollecting and sequencing to establish their relationships with species of Phyllosticta and whether they can be
considered as distinct species.
Phyllosticta ampelicida (Engelm.) Aa, Stud. Mycol. 5:
28 (1973)
Teleomorph: Guignardia bidwellii (Ellis) Viala & Ravaz,
Bull. Soc. mycol. Fr. 8:63 (1892)
Hosts: Vitis spp., Ampelopsis spp., Cissus spp., Parthenocissus spp. (Vitaceae).
Disease symptoms: Black rot
Distribution: Asia, Canada, South America, UK and USA.
Notes: This Phyllosticta species is linked to the generic
type of Guignardia and has a Leptodothiorella spermatial
state. The black rot fungus can infect all parts of the vine,
although the most significant losses are caused by berry
infection (Reddick 1911, Miller 1968; Kuo and Hoch 1996;
Wilcox 2003; Ellis et al. 2004). In warm humid climates,
susceptible varieties can experience complete loss if the
pathogen is left uncontrolled (Hoover et al. 2011).
Sequences of reference isolates of this species are available
in GenBank but it has not yet been epitypified.
Phyllosticta ardisiicola Motohashi, I. Araki & C.
Nakash., Mycoscience 49 (2008)
Hosts: Ardisia crenata (Myrsinaceae)
Disease symptoms: Leaf spot
Distribution: Japan
Note: This species was introduced as new species based on
its morphological differences with taxa from related hosts
(Motohashi et al. 2008). Molecular data should be included in
future work to confirm the status with closely related species.
Phyllosticta aspidistricola Motohashi, I. Araki & C.
Nakash., Mycoscience 49: 138–146 (2008)
Hosts: Aspidistra elatior (Liliaceae)
Disease symptoms: Leaf spot
Distribution: Japan
Note: This species were introduced as new species base on
its morphological differences with taxa from related hosts
(Motohashi et al. 2008). Molecular data should be included in
future work to confirm the status with closely related species.
Fungal Diversity
Phyllosticta beaumarisii A.P. Paul & M.D. Blackburn,
Australas. Pl. Path. 15: 41 (1986)
Spermatial state: Leptodothiorella sp.
Hosts: Muehlenbeckia adpressa (Nyctaginaceae)
Disease symptoms: leaf spots with distinctive necrotic
lesions
Distribution: Australia
Note: The disease is prevalent during autumn and winter.
Mature diseased leaves age and abscise when a new flush
of growth occurs in spring. Pathogenicity testing has shown
that P. beaumarisii is the causal agent of the disease as
compared to other species of Phyllosticta associated with
the host (Paul and Blackburn 1986). Although Yip (1987)
have provided the full description and illustration of the
species, molecular data is needed in future studies to
confirm its status as distinct species.
Phyllosticta bifinariae O.L. Pereira, C. Glienke &
Crous, Persoonia 26: 52 (2011)
Hosts: Bifrenaria harrisoniae (Orchidaceae)
Disease symptoms: Leaf spot
Distribution: Brazil
Notes: This isolate was originally thought to be representative of P. capitalensis but was found to be ecologically and
phylogenetically distinct and a pathogen of Bifrenaria
harrisoniae (Glienke et al. 2011).
Phyllosticta brazilianiae D. Stringari, C. Glingke &
Crous, Persoonia 26: 47–56 (2011)
Hosts: Mangifera indica (Anacardiaceae)
Disease symptoms: Symptomless endophyte
Distribution: Brazil
Note: The species is ecologically distinct from P.
anacardiacearum being an endophyte, and failing to induce
leaf spots despite repeated inoculation on mango (Glienke
et al. 2011). Molecular data has also shown that it is distinct
from other closely related species.
Phyllosticta capitalensis Henn., Hedwigia 48: 13 (1908)
Hosts: Phyllosticta capitalensis was originally described
on Stanhopea (Orchidaceae) from Brazil by Hennings
(1908) although now thought to occur in wide range of
hosts.
Disease symptoms: leaf spots (when cause disease)
Distribution: Worldwide
Note: Phyllosticta capitalensis is the most recently
proposed name for the entities that were formally incorrectly referred to as Guignardia mangiferae (Baayen et al.
2002; Glienke et al. 2011). The taxon is frequently isolated
as an endophyte and has a wide host range and geographic
distribution. Okane et al. (2001) identified an endophytic
Phyllosticta strain in ericaceous plants from Japan, as
Phyllosticta capitalensis, describing the teleomorph as a
new species, G. endophyllicola. Baayen et al. (2002)
recognized the common endophytic species associated with
a wide host range of plants based on ITS sequence
similarities, which was similar to G. endophyllicola in
morphology. Although several names were available for
this species, they opted to call the species G. mangiferae (a pathogen on Mangifera indica (Anacardiaceae)
in India), while the anamorph was referred to as P.
capitalensis. Although no clear argument was presented
for choosing the name G. mangiferae for this fungus, the
choice of the anamorph name was based on the fact that
two isolates from Orchidaceae (CBS 398.80, CBS 226.77)
clustered in the same clade in their study. A comprehensive study of endophytic and pathogenic Phyllosticta
species on Citrus was carried out by Glienke et al.
(2011). Their combined phylogenetic tree revealed the P.
capitalensis sensu lato clade to be genetically distinct
from a reference isolate of G. mangiferae isolated from
India. Several names were available for this clade, the
oldest being P. capitalensis. Glienke et al. (2011)
therefore, suggested that endophytic, non-pathogenic isolates occurring on a wide host range would be more
correctly referred to as P. capitalensis. However, more
genes need to be analyzed to fully resolve the morphological variation still observed within this clade.
Phyllosticta citriasiana Wulandari, Crous & Gruyter,
Fungal Diversity 34: 31 (2009).
Hosts: Citrus maxima (Rutaceae)
Disease symptoms: tan spots (produces shallow lesions
with a small central grey to tan crater usually with dark
brown margin on fruits)
Distribution: Asia (China, Thailand, Vietnam)
Notes: The tan spot symptom usually appears after the
fruit has started to ripen and sometimes it can occur after
harvest. Combined gene analysis, morphological and
culture based characters were employed to distinguish the
species from P. citricarpa and other species considered
(Wulandari et al. 2009). Recent studies on endophytic and
pathogenic species of Phyllosticta from citrus in different
regions of the world shows that the morphological, cultural
and biochemical characters for species were consistent with
the results of phylogenetic analysis of related taxa (Glienke
et al. 2011; Wang et al. 2011). A specific primer pair Pca8/
ITS4 was also designed and selected, and a PCR protocol
was used to detect P. citriasiana in recent study (Wang et
al. 2011).
Phyllosticta citribraziliensis C. Glienke & Crous, Persoonia 26: 54 (2011)
Hosts: Citrus limon (Rutaceae)
Disease symptoms: Symptomless endophyte
Distribution: Brazil
Notes: This species is closely related to P. spinarum but
phylogenetically distinct. Also P citribraziliensis is morphologically distinguished in having larger conidia, a thick
mucilaginous sheath surrounding its conidia and branched
conidiophores.
Fungal Diversity
Phyllosticta citricarpa (McAlpine) van der Aa, Stud.
Mycol. 5: 40 (1973)
Teleomorph: Guignardia citricarpa Kiely
Hosts: Citrus aurantius, C. limon, C. delicoisa, C.
reticulata, C. sinensis (Rutaceae)
Disease symptoms: Black spot of citrus, foliar and fruit
diseases, premature fruit drop
Distribution: Asia, Africa, Australia, USA (Florida)
Notes: P. citricarpa causes foliar and fruit disease of
Citrus spp. G. citricarpa (anamorph P. citricarpa) which
causes Citrus Black Spot is regulated as a quarantine pest in
the European Union and the USA (Wang et al. 2011). This
pathogen can infect the rind of Citrus fruit causing disease
lesions (Kiely 1948a). Serious infection near the pedicel of
the developing fruit possibly will lead to premature fruit
drop (Baayen et al. 2002). The first report of Black spot on
Citrus orchards was near Sydney, Australia and it was
described as Phoma citricarpa McAlpine (McAlpine 1899).
The teleomorph was described as Guignardia citricarpa
Kiely (Kiely 1948b). van der Aa (1973) classified the
anamorph as Phyllosticta citricarpa (McAlpine) Van der
Aa. The species was recollected from Australia and an
epitype was designated and the distinctiveness from P.
citriasiana was confirmed (Glienke et al. 2011).
Phyllosticta citrichinaensis H.X. Wang, K.D. Hyde &
H.Y. Li, Fungal Diversity (2011)
Hosts: Citrus spp. (Rutaceae)
Distribution: China
Disease symptoms: small grey, red-brown or brown
spots and freckles on leaves, melanose like black spots on
fruits
Notes: This taxon has been isolated as an endophyte and
is also weak pathogen (Wang et al. 2011). P. citrichinaensis
differs from the other four Phyllosticta species associated
with citrus in its morphological, cultural and biochemical
characteristics.
Phyllosticta convallariae Pers., Traité sur les Champignons Comestibles (Paris): 148 (1818)
Hosts: Polygonatum spp., Convallaria (Convallariaceae),
Maianthemum (Liliaceae).
Disease symptoms: Red leaf spot
Distribution: Asia, Europe
Note: This is the generic type of Phyllosticta and has
been clearly designated in Donk (1968). This species
causes reddish-brown leaf spots on its host and has a
Leptodothiorella spermatial state (http://www.uni-graz.at/
∼oberma/fungi-of-austria/phyllosticta-convallariae.html;
available online 19 Sep, 2011) and needs recollecting and
epitypifying. An earlier name was found in Phyllosticta
cruenta (van der Aa 1973) but this needs confirmation
based on sequencing collections from the original host.
Phyllosticta cruenta (Fr.) J.J. Kickx, Fl. Crypt. Flandres
1: 412 (1867)
Teleomorph: Guignardia reticulata (DC.: Fr.) Aa
Hosts: Polygonatum spp., Convallaria (Convallariaceae),
Maianthemum (Liliaceae).
Disease symptoms: Leaf spot
Distribution: Asia, Europe
Notes: P. cruenta is the earlier name for type of
Phyllosticta convallariae and in older literature, it was
considered to be an intermediate form between Phyllostictina and Dothiorella (van der Aa and Vanev 2002). This
taxon should be recollected and epitypified as its distinctiveness from P. convallariae needs confirmation.
Phyllosticta cussoniae Cejp, Bothalia 10(2): 341 (1971)
Teleomorph: Guignardia cussonia Crous
Host: Cussonia spp.
Disease symptoms: On leaves causing a prominent leaf
spot.
Distribution: South Africa
Notes: In the phylogenic tree presented in Glienke et al.
(2011) the isolates of this species clusters in a distinct clade
and appears to represent a distinct taxa. Representative
isolates were obtained from South Africa by P.W. Crous
and designated as epitype (Glienke et al. in prep.).
Phyllosticta dioscoreae Cooke, Grevillea 6(no. 40): 136
(1878)
Teleomorph: Guignardia dioscoreae A.K. Pande, Sydowia
22(5–6): 367 (1969) [1968]
Host: Dioscorea spp. (Dioscoreaceae)
Disease symptoms: Leaf spot
Distribution: Africa (South Africa), Asia, Australia,
South America (Brazil), USA.
Notes: An isolate identified as Phyllosticta dioscoreae
from Hibiscus rosa-sinensis has been reported to produce
the anti-cancer compound taxol (Kumaran et al. 2009b), but
the species was identified on basis of morphological
characters and therefore needs confirmation with molecular
data. This species commonly causes leaf spots on Dioscorea spp. with its Guignardia state usually being
produced (N. Wulandari, pers. comm.) and should be
recollected and epitypified to establish if it is a distinct
species.
Phyllosticta fallopiae Motohashi, I. Araki & C. Nakash.,
Mycoscience 49 (2008)
Hosts: Fallopia japonica
Disease symptoms: leaf spots
Distribution: Japan
Note: This species were introduced as new species base
on its morphological differences with taxa from related
hosts therefore molecular data are needed in future studies
to establish its uniqueness (Motohashi et al. 2008).
Phyllosticta hypoglossi (Mont.) Allesch., Rabenh.
Krypt.-Fl., Edn 2 (Leipzig) 1(6): 163 (1898)
Hosts: Living leaves and dead cladodes Ruscus spp.
(Liliaceae)
Fungal Diversity
Disease symptoms: Occurring on living and dead
cladodes and stems, though distinct spots not reported,
Distribution: Asia, Europe (France, Italy, Portugal,
Turkey, Ukraine)
Notes: The spermatial state is a Leptodothiorella sp. The
taxon was originally described from France, and the
sequenced isolates used by Glienke et al. (2011) collected
from Italy could potentially be used for epitypification.
Phyllosticta kerriae Motohashi, I. Araki & C. Nakash.,
Mycoscience 49 (2008)
Teleomorph: Unknown
Hosts: Kerria japonica
Disease symptoms: leaf spots
Distribution: Japan
Note: This species were introduced as new species base
on its morphological differences with taxa from related
hosts therefore the molecular based re evaluation is needed
to establish its uniqueness (Motohashi et al. 2008).
Phyllosticta minima (Berk. & M.A. Curtis) Underw. &
Earle, Bulletin of the Alabama Agricultural Experiment
Station 80: 168 (1897)
Hosts: Acer spp. (Maples) (Sapindaceae)
Disease symptoms: Leaf spot (black or purple eye spot).
Distribution: Asia (China), North America
Notes: This pathogen has been reported as a common
causative agent of ornamental Acer spp. (http://www.unce.
unr.edu/publications/files/ho/2005/fs0547.pdf). Phyllosticta
gallarum has been recorded causing similar leaf spots on
Caragana spp. (http://www.unl.edu/nac/diseasetrees/chap4.
pdf). Recollection and taxonomic re evaluation is needed to
confirm if this species can be differentiated from closely
related species.
Phyllosticta musarum (Cooke) Aa, Stud. Mycol. 5: 72
(1973)
Teleomorph: Guignardia musae Racib.
Hosts: Musa spp. (Musaceae).
Disease symptoms: Leaf spot, fruit spot, banana freckle,
banana black spot
Distribution: Widespread
Notes: The pathogen (Phyllosticta musarum or the
sexual state Guignardia) infects leaves and also affects the
external appearance of the fruit, decreasing its quality and
marketability. Severe infections of the disease may cause
premature death of the older leaves on some banana
cultivars. Preinfection studies shows that the pathogen
seems to penetrate directly through the epidermal cuticle
layer of the host by forming appressoria and infection pegs
(Pu et al. 2008). Wulandari et al. (2010) investigated the
problem of the occurrence of the species epithet (“musae”)
on separate occasions related to sexual state based on
herbarium specimens and fresh collectios and distingushed
there different species including taxonomic novelties.
However the Phyllosticta/Guidnardia species from banana
needs to re evaluated based on morphological and molecular approach in future studies.
Phyllosticta owaniana G. Winter, Hedwigia 24: 31
(1885)
Host: Brabejum stellatifolium (Proteaceae)
Disease symptoms: leaf spot
Distribution: South Africa
Notes: P. telopeae Yip has been reported from Telopea
speciosissima (Proteaceae) and is distingushed from P.
owaniana by its larger conidia and much longer appendages. Both of the species being accepted in van der Aa and
Vanev (2002). The sequences of the type of P. owaniana
has been used in phylogenetic analysis but as the outgroup
which shows the species to be significantly different from
other Phyllosticta species. An epitype will be designated
based on fresh collections (Glienke et al., in prep).
Phyllosticta solitaria Ellis & Everh., Proc. Acad. nat.
Sci. Philad. 47: 430 (1895)
Hosts: Malus spp., Crataegus spp. (Rosaceae). Pyrus
spp.
Disease symptoms: Leaf spot, fruit blotch, twig canker.
Distribution: Asia (India, China), Africa (Zimbabwe,
South Africa), Europe (Greece), North America (USA),
South America (Brazil).
Notes: Although the teleomorph is unknown, Guba (1925)
have noticed the fructification on fallen leaves in spring (van
der Aa and Vanev 2002). P. solitaria causes a serious
blotching of apples which reduces fruit quality and also
known as quarantine pest by EPPO (http://www.eppo.org/
QUARANTINE/fungi/Phyllosticta_solitaria/PHYSSL_ds.
pdf). The ability of the fungus to withstand long periods of
cold storage should be noted in quarantine purposes. Reevaluation of the pathogen and epitypification is needed in
future studies.
Phyllosticta sphaeropsoidea Ellis & Everh., Bull. Torrey
bot. Club 10(7): 97 (1883)
Teleomorph: Guignardia aesculi (Peck) V.B. Stewart
Hosts: Aesculus spp. (Hippocastanaceae).
Disease symptoms: leaf blotch (disease known as
buckeye blotch or horse chestnut blotch), black rot, brown
leaf margin and necrotic tissue
Distribution: Asia, Europe, North America
Notes: Guignardia aesculi (sexual stage), initiates leaf
infections in early spring, while P. sphaeropsoidea (asexual
stage) perpetuates infections during the summer. Infections
from both stages combine to cause horse chestnut leaf
blotch (Gillman 2005; Pastricakova 2004). Recollection
from various host species of Aesculus and various geographical locations are needed to establish the uniqueness
of the taxa with molecular data.
Phyllosticta spinarum (Died.) Nag Raj & M. Morelet,
Bull. Soc. Sci. nat. Arch. Toulon et du Var 34(219): 12
(1978)
Fungal Diversity
Hosts: Juniperus sp, Chamaecyparis pisifera, Platycladus orientalis (Cupressaceae), Hedera helix (Araliaceae)
Disease symptoms: none reported, presumed endophyte
Distribution: Europe (Germany, France, Italy), USA
Notes: This was originally described from Juniperus sp.
in Germany while the isolates sequenced in Glienke et al.
(2011) were from Chamaecyparis pisifera and Hedera helix
(from France and Italy). The endophytic isolate putatively
identified as P. spinarum from Platycladus orientalis is
known to produce novel secondary metabolites (Wijeratne
et al. 2008).
Phyllosticta vaccinii Earle, Bull. Torrey bot. Club 24: 31
(1897)
Teleomorph: Guignardia vaccinii Shear
Hosts: Vaccinium spp. (Ericaceae).
Disease symptoms: Blast or blight of flowers and young
fruits; early rot of fruits in storage
Distribution: Asia (China), North America
Notes: Weidemann et al. (1982) listed the anamorph of
Guignardia vaccinii as Phyllosticta elongata, but van der
Aa (2002) lists both Phyllosticta elongata and Phyllosticta
vaccinii as anamorphs. A strain of this species
(CBS165.86) has been sequenced by Duong (2008) and
appears to be a distinct species in phylogenetic analysis.
However, future work is needed to establish the uniqueness of taxa.
Concluding remarks
Species recognition criteria in Phyllosticta and the sexual
state Guignardia have evolved from morphological criteria
to phylogenetic species concepts that involves DNA
sequence data derived from type material. However as
compared to the number of accepted species in previous
revisionary treatments, there is a lack of molecular-based
studies considering the wide range of hosts and various
geographic locations. We therefore recommend the recollection of taxa, and epitypification where possible. The
number of resolved names in the genus would be updated
with ongoing projects and the sexual states should be linked
accordingly to establish operational biological species.
Acknowledgements We acknowledge The Royal Golden Ph.D.
Jubilee Program grant No. Ph.D./0198/2552 in 2.B.M.F./52/A.1.N.XX
for the first author to carry out a PhD on the taxonomy and
phylogeny of the genus Phyllosticta. Dhanushka Udayanga thanks
Chinese Academy of Sciences, Beijing and Mushroom Research
Foundation, Thailand for a postgraduate scholarship. The National
Research Council of Thailand awarded grant No 54201020004 to
study the genus Phyllosticta in Thailand. The Global Research
Network for Fungal Biology and King Saud University are also
thanked for support.
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