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1 CBS Fungal Biodiversity Centre, Uppsalalaan 8, 3584 CT Utrecht, The
Netherlands
2 Laboratory of Phytopathology, Wageningen University, Binnenhaven 5, 6709
PD Wageningen, The Netherlands
3 Martin-Luther-Universität, Institut für Biologie, Geobotanik und
Botanischer Garten, Neuwerk 21, D-06099 Halle (Saale), Germany
4 Division of Environmental Science & Ecological Engineering, Korea
University, Seoul 136-701, Korea
*
Correspondence: Pedro W. Crous,
p.crous{at}cbs.knaw.nl
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Abstract |
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 TOP Abstract INTRODUCTIONMATERIAlS AND METHODSRESULTSDISCUSSIONReferences |
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Taxonomic novelties: Myrmecridium Arzanlou, W. Gams & Crous, gen. nov., Myrmecridium flexuosum (de Hoog) Arzanlou, W. Gams & Crous, comb. et stat. nov., Myrmecridium schulzeri (Sacc.) Arzanlou, W. Gams & Crous var. schulzeri, comb. nov., Myrmecridium schulzeri var. tritici (M.B. Ellis) Arzanlou, W. Gams & Crous, comb. nov., Periconiella arcuata Arzanlou, S. Lee & Crous, sp. nov., Periconiella levispora Arzanlou, W. Gams & Crous, sp. nov., Pleurothecium obovoideum (Matsush.) Arzanlou & Crous, comb. nov., Pseudovirgaria H.D. Shin, U. Braun, Arzanlou & Crous, gen. nov., Pseudovirgaria hyperparasitica H.D. Shin, U. Braun, Arzanlou & Crous, sp. nov., Radulidium Arzanlou, W. Gams & Crous, gen. nov., Radulidium epichloës (Ellis & Dearn.) Arzanlou, W. Gams & Crous, comb. nov., Radulidium subulatum (de Hoog) Arzanlou, W. Gams & Crous, comb. nov., Ramichloridium australiense Arzanlou & Crous, sp. nov., Ramichloridium biverticillatum Arzanlou & Crous, nom. nov., Ramichloridium brasilianum Arzanlou & Crous, sp. nov., Ramichloridium strelitziae Arzanlou, W. Gams & Crous, sp. nov., Rhinocladiella basitona (de Hoog) Arzanlou & Crous, comb. nov., Rhinocladiella fasciculata (V. Rao & de Hoog) Arzanlou & Crous, comb. nov., Rhinocladiella mackenziei (C.K. Campb. & Al-Hedaithy) Arzanlou & Crous, comb. nov., Rhodoveronaea Arzanlou, W. Gams & Crous, gen. nov., Rhodoveronaea varioseptata Arzanlou, W. Gams & Crous, sp. nov., Thysanorea Arzanlou, W. Gams & Crous, gen. nov., Thysanorea papuana (Aptroot) Arzanlou, W. Gams & Crous, comb. nov., Veronaea japonica Arzanlou, W. Gams & Crous, sp. nov., Veronaeopsis Arzanlou & Crous, gen. nov., Veronaeopsis simplex (Papendorf) Arzanlou & Crous, comb.nov.
Keywords Capnodiales / Chaetothyriales / Mycosphaerella / Periconiella / phylogeny / Rhinocladiella / Veronaea
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INTRODUCTION |
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TOPAbstractINTRODUCTIONMATERIAlS AND METHODSRESULTSDISCUSSIONReferences |
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Periconiella Sacc. (1885) [type species P. velutina (G. Winter) Sacc.] differs from Veronaea Cif. & Montemart. chiefly based on its dark brown, apically branched conidiophores. However, de Hoog (1977) observed numerous specimens of V. musae to exhibit branched conidiophores in culture, as did Stahel (1937) for Ramichloridium musae. De Hoog (1977) subsequently re-introduced Ramichloridium, but typified it with R. apiculatum (J.H. Mill., Giddens & A.A. Foster) de Hoog. He regarded V. musae and P. musae to be conspecific, and applied the name R. musae (Stahel ex M.B. Ellis) de Hoog to both species, regarding Periconiella musae as basionym. The circumscription by de Hoog was based on their similar morphology and ecology. Central in his genus concept was the observed presence of more or less differentiated and pigmented conidiophores, with predominantly aseptate conidia produced on a sympodially proliferating rachis. De Hoog (1977) also used some ecological features as additional characters to discriminate Ramichloridium from other genera, noting, for instance, that species in Ramichloridium were non-pathogenic to humans (de Hoog 1977, Campbell & Al-Hedaithy 1993). This delimitation, however, was not commonly accepted (McGinnis & Schell 1980). De Hoog et al. (1983) further discussed the problematic separation of Ramichloridium from genera such as Rhinocladiella, Veronaea and Cladosporium Link. It was further noted that the main feature to distinguish Ramichloridium from Rhinocladiella, was the presence of exophiala-type budding cells in species of Rhinocladiella (de Hoog 1977, de Hoog et al. 1983, Veerkamp & Gams 1983). The separation of Veronaea from this complex is more problematic, as the circumscriptions provided by Ellis (1976) and Morgan-Jones (1979, 1982) overlap with that of Ramichloridium sensu de Hoog (1977). Cladosporium is more distinct, having very conspicuous, protuberant, darkened and thickened, coronate conidial scars, and catenate conidia (David 1997, Braun et al. 2003, Schubert et al. 2007 - this volume).
To date 26 species have been named in Ramichloridium; they not only differ in morphology, but also in life style. Ramichloridium mackenziei C.K. Campb. & Al-Hedaithy is a serious human pathogen, causing cerebral phaeohyphomycosis (Al-Hedaithy et al. 1988, Campbell & Al-Hedaithy 1993), whereas R. musae causes tropical speckle disease on members of the Musaceae (Stahel 1937, Jones 2000). Another plant-pathogenic species, R. pini de Hoog & Rahman, causes a needle disease on Pinus contorta (de Hoog et al. 1983). Other clinically relevant species of Ramichloridium are R. basitonum de Hoog and occasionally R. schulzeri (Sacc.) de Hoog, while the remaining species tend to be common soil saprobes.
No teleomorph has thus far been linked to species of Ramichloridium. The main question that remains is whether shared morphology among the species in this genus reflects common ancestry (Seifert 1993, Untereiner & Naveau 1999). To delineate anamorphic genera adequately, morphology and conidial ontogeny alone are no longer satisfactory (Crous et al. 2006a, b), and DNA data provide additional characters to help delineate species and genera (Taylor et al. 2000, Mostert et al. 2006, Zipfel et al. 2006). The aim of the present study was to integrate morphological and cultural features with DNA sequence data to resolve the species concepts and generic limits of the taxa currently placed in Periconiella, Ramichloridium, Rhinocladiella and Veronaea, and to resolve the status of several new cultures that were isolated during the course of this study.
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MATERIAlS AND METHODS |
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TOPAbstractINTRODUCTIONMATERIAlS AND METHODSRESULTSDISCUSSIONReferences |
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DNA extraction, amplification and sequence analysis
Genomic DNA was extracted from colonies grown on 2 % malt extract agar
(MEA, Difco) (Gams et al.
2007) using the FastDNA kit (BIO101, Carlsbad, CA, U.S.A.). The
primers ITS1 and ITS4 (White et
al. 1990) were used to amplify the internal transcribed
spacer region (ITS) of the nuclear ribosomal RNA operon, including: the 3' end
of the 18S rRNA gene, the first internal transcribed spacer region (ITS1), the
5.8S rRNA gene, the second internal transcribed spacer region (ITS2) and the
5' end of 28S rRNA gene. Part of the large subunit 28S rRNA (LSU) gene was
amplified with primers LR0R (Rehner &
Samuels 1994) and LR5
(Vilgalys & Hester 1990).
The ITS region was sequenced only for those isolates for which these data were
not available. The ITS analyses confirmed the proposed classification based on
LSU analysis for each major clade and are not presented here in detail; but
the sequences are deposited in GenBank where applicable. The PCR reaction was
performed in a mixture with 0.5 units Taq polymerase (Bioline,
London, U.K.), 1x PCR buffer, 0.5 mM MgCl2, 0.2 mM of each
dNTP, 5 pmol of each primer, approximately 10-15 ng of fungal genomic DNA,
with the total volume adjusted to 25 µL with sterile water. Reactions were
performed on a GeneAmp PCR System 9700 (Applied Biosystems, Foster City, CA)
with cycling conditions consisting of 5 min at 96 °C for primary
denaturation, followed by 36 cycles at 96 °C (30 s), 52 °C (30 s), and
72 °C (60 s), with a final 7 min extension step at 72 °C to complete
the reaction. The amplicons were sequenced using BigDye Terminator v. 3.1
(Applied Biosystems, Foster City, CA) or DYEnamicET Terminator (Amersham
Biosciences, Freiburg, Germany) Cycle Sequencing Kits and analysed on an ABI
Prism 3700 (Applied Biosystems, Foster City, CA) under conditions recommended
by the manufacturer. Newly generated sequences were subjected to a Blast
search of the NCBI databases, sequences with high similarity were downloaded
from GenBank and comparisons were made based on the alignment of the obtained
sequences. Sequences from GenBank were also selected for similar taxa. The LSU
tree was rooted using sequences of Athelia epiphylla Pers. and
Paullicorticium ansatum Liberta as outgroups. Phylogenetic analysis
was performed with PAUP (Phylogenetic Analysis Using Parsimony) v. 4.0b10
(Swofford 2003), using the
neighbour-joining algorithm with the uncorrected ("p"), the Kimura
2-parameter and the HKY85 substitution models. Alignment gaps longer than 10
bases were coded as single events for the phylogenetic analyses; the remaining
gaps were treated as missing data. Any ties were broken randomly when
encountered. Phylogenetic relationships were also inferred with the parsimony
algorithm using the heuristic search option with simple taxon additions and
tree bisection and reconstruction (TBR) as the branch-swapping algorithm;
alignment gaps were treated as a fifth character state and all characters were
unordered and of equal weight. Branches of zero length were collapsed and all
multiple, equally parsimonious trees were saved. Only the first 5 000 equally
most parsimonious trees were saved. Other measures calculated included tree
length, consistency index, retention index and rescaled consistency index (TL,
CI, RI and RC, respectively). The robustness of the obtained trees was
evaluated by 1 000 bootstrap replications. Bayesian analysis was performed
following the methods of Crous et al.
(2006c). The best nucleotide
substitution model was determined using MrModeltest v. 2.2
(Nylander 2004).
MRBAYES v. 3.1.2
(Ronquist & Huelsenbeck
2003) was used to perform phylogenetic analyses, using a general
time-reversible (GTR) substitution model with inverse gamma rates, dirichlet
base frequencies and the temp value set to 0.5. New sequences were lodged with
NCBI's GenBank (Table 1) and
the alignment and trees with TreeBASE
(www.treebase.org).
Morphology
Cultural growth rates and morphology were recorded from colonies grown on
MEA for 2 wk at 24 °C in the dark, and colony colours were determined by
reference to the colour charts of Rayner
(1970). Microscopic
observations were made from colonies cultivated on MEA and OA (oatmeal agar,
Gams et al. 2007),
using a slide culture technique. Slide cultures were set up in Petri dishes
containing 2 mL of sterile water, into which a U-shaped glass rod was placed,
extending above the water surface. A block of freshly growing fungal colony,
approx. 1 cm square was placed onto a sterile microscope slide, covered with a
somewhat larger, sterile glass cover slip, and incubated in the moist chamber.
Fungal sporulation was monitored over time, and when optimal, images were
captured by means of a Nikon camera system (Digital Sight DS-5M, Nikon
Corporation, Japan). Structures were mounted in lactic acid, and 30
measurements (x 1 000 magnification) determined wherever possible, with
the extremes of spore measurements given in parentheses.
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RESULTS |
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TOPAbstractINTRODUCTIONMATERIAlS AND METHODSRESULTSDISCUSSIONReferences |
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Taxonomy
The species previously described in Ramichloridium share some
morphological features, including erect, pigmented, more or less
differentiated conidiophores, sympodially proliferating conidiogenous cells
and predominantly aseptate conidia. Other than conidial morphology, features
of the conidiogenous apparatus that appear to be more phylogenetically
informative include pigmentation of vegetative hyphae, conidiophores and
conidia, denticle density on the rachis, and structure of the scars. By
integrating these data with the molecular data set, more natural genera are
delineated, which are discussed below.
Key to ramichloridium-like genera
Capnodiales (Mycosphaerellaceae, Teratosphaeriaceae)
The type species of Ramichloridium, R. apiculatum, together with
R. indicum cluster as a sister group to the Dissoconium de
Hoog, Oorschot & Hijwegen clade in the Mycosphaerellaceae. Some
other Ramichloridium species, including R. musae, R.
biverticillatum Arzanlou & Crous, R. pini and R.
cerophilum, are also allied with members of the
Mycosphaerellaceae. Three additional new species are introduced for
Ramichloridium isolates from Musa banksii, Strelitzia
nicolai, and forest soil. Periconiella velutina, the type
species of Periconiella, which also resides in the
Mycosphaerellaceae, is morphologically sufficiently distinct to
retain its generic status. Two new species of Periconiella are
introduced for isolates obtained from Turpinia pomifera and
Ischyrolepis subverticellata in South Africa. Zasmidium
cellare (Pers.) Fr., the type species of Zasmidium (Pers.) Fr.,
is also shown to cluster within the Mycosphaerellaceae.
Periconiella Sacc., in Sacc. & Berlese, Atti Ist. Veneto Sci., Ser. 6, 3: 727. 1885.
In vitro: Colonies with entire margin; aerial mycelium rather compact, raised, velvety, olivaceous-grey; reverse olivaceous-black. Submerged hyphae verrucose, hyaline, thin-walled, 1-3 µm wide; aerial hyphae subhyaline, later becoming dark brown, thick-walled, smooth. Conidiophores arising vertically from creeping hyphae, straight or flexuose, up to 260 µm long, dark brown at the base, paler towards the apex, thick-walled; in the upper part bearing short branches. Conidiogenous cells terminally integrated, polyblastic, smooth or verrucose, subcylindrical, mostly not or barely geniculate-sinuous, variable in length, subhyaline, later becoming pale brown, fertile part as wide as the basal part, proliferating sympodially, sometimes becoming septate and forming a short, straight rachis with pigmented, slightly thickened and hardly prominent, more or less flat scars. Conidia solitary, occasionally in short chains, 0-multi-septate, subhyaline to rather pale olivaceous or olivaceous-brown, smooth to verrucose, globose, ellipsoidal to obovoid or obclavate, with a slightly darkened and thickened hilum; conidial secession schizolytic.
Type species: P. velutina (G. Winter) Sacc., Miscell. mycol. 2: 17. 1884.
Notes: Periconiella is distinct from other ramichloridium-like genera by its conidiophores that are prominently branched in the upper part, and by its darkened, thickened conidial scars, that are more or less flat and non-prominent. Although conidiophores are also branched in the upper part in Thysanorea Arzanlou, W. Gams & Crous, the branching pattern in the latter genus is different from that of Periconiella. Thysanorea has a complex head consisting of up to six levels of branches, while in Periconiella the branching is limited, with mainly primary and secondary branches. Furthermore, Thysanorea is characterised by having dimorphic conidiophores and more or less prominent denticle-like conidiogenous loci.
Periconiella velutina (G. Winter) Sacc., Miscell. mycol. 2: 17. 1884. Fig. 3.
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Basionym: Periconia velutina G. Winter, Hedwigia 23: 174. 1884.
In vitro: Submerged hyphae verrucose, hyaline, thin-walled, 1-3 µm wide; aerial hyphae subhyaline, later becoming dark brown, thick-walled, smooth. Conidiophores arising vertically from creeping hyphae, straight or flexuose, up to 260 µm long, dark brown at the base, paler towards the apex, thick-walled; in the upper part bearing short branches, 10-35 µm long. Conidiogenous cells mostly terminally integrated, sometimes discrete, smooth or verrucose, cylindrical, variable in length, subhyaline, later becoming pale brown, fertile part as wide as the basal part, proliferating sympodially, sometimes becoming septate and forming a short, straight rachis with pigmented, slightly thickened and hardly prominent, more or less flat scars, less than 1 µm diam. Conidia 0(-1)-septate, subhyaline, thin-walled, verrucose or smooth, globose, ellipsoidal to obovoid, (7-)8-9(-11) x (2.5-)3(-4) µm, with a slightly darkened and thickened hilum, 1.5-2 µm diam.
Cultural characteristics: Colonies on MEA slow-growing, reaching 4 mm diam after 14 d at 24 °C, with entire margin; aerial mycelium rather compact, raised, velvety, olivaceous-grey; reverse olivaceous-black.
Specimens examined: South Africa, Cape Town, on Brabejum stellatifolium, P. MacOwan, herb. G. Winter (B), lectotype selected here; Cape Town, on leaves of Brabejum stellatifolium (= B. stellatum), P. Mac-Owan, PAD, F42165, F462166, isolectotypes; Stellenbosch, Jonkershoek Nature Reserve, on Brabejum stellatifolium, 21 Jan. 1999, J.E. Taylor, epitype designated here CBS H-15612, cultures ex-epitype CBS 101948-101950.
Periconiella arcuata Arzanlou, S. Lee & Crous, sp. nov. MycoBank MB504547. Figs 4, 7A.
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Ab aliis speciebus Periconiellae conidiis obclavatis, rectis vel curvatis, (30-)53-61(-79) x (3-)5(-7) µm, distinguenda.
Submerged hyphae smooth, hyaline, thin-walled, 2 µm wide; aerial hyphae pale brown, smooth or verrucose, slightly narrower. Conidiophores arising vertically from creeping hyphae, straight or flexuose, up to 300 µm long, dark brown at the base, paler towards the apex, thick-walled; loosely branched in the upper part, bearing short branches. Conidiogenous cells integrated, cylindrical, variable in length, 20-50 µm long, subhyaline, later becoming pale brown, fertile part as wide as the basal part, proliferating sympodially, forming a geniculate conidium-bearing rachis with pigmented and thickened, prominent, cone-shaped scars, 1 µm diam. Conidia formed singly, obclavate, straight or mostly curved, 0(-4)-septate, coarsely verrucose, pale olive, thin-walled, tapering towards the apex, (30-)53-61(-79) x (3-)5(-7) µm, with a narrowly truncate base and a darkened, hardly thickened hilum, 2 µm diam; microcyclic conidiation observed in culture.
Cultural characteristics: Colonies on MEA reaching 12 mm diam after 14 d at 24 °C, with entire, smooth, sharp margin; mycelium compacted, becoming hairy, colonies up to 1 mm high; surface olivaceous to olivaceous-grey, reverse dark grey-olivaceous to olivaceous-black.
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Periconiella levispora Arzanlou, W. Gams & Crous, sp. nov. MycoBank MB504546. Figs 5-6B.
Etymology: (Latin) levis = smooth.
A simili Periconiella velutina conidiis levibus et maioribus, ad 23 µm longis distinguenda.
In vitro: Submerged hyphae smooth, hyaline, thin-walled, 2-2.5 µm wide; aerial hyphae subhyaline, later becoming dark brown, thick-walled, smooth. Conidiophores arising vertically from creeping aerial hyphae, dark brown at the base, paler towards the apex, thick-walled; in the upper part bearing several short branches, up to 120 µm long. Conidiogenous cells integrated, occasionally discrete, cylindrical, variable in length, 10-20 µm long, subhyaline, later becoming pale brown, fertile part as wide as the basal part, proliferating sympodially, forming a short rachis with pigmented and slightly thickened, somewhat protruding scars, less than 1 µm diam. Conidia solitary, 0(-2)-septate, smooth, pale olivaceous, cylindrical, ellipsoidal, pyriform to clavate, (7-)11-14(-23) x (3-)4-5(-6) µm, with a truncate base and a darkened, slightly thickened hilum, 2 µm diam.
Cultural characteristics: Colonies on MEA slow-growing, reaching 5 mm diam after 14 d at 24 °C, with entire margin; aerial mycelium compact, raised, velvety, olivaceous-grey; reverse olivaceous-black.
Specimen examined: Sri Lanka, Hakgala Botanic Gardens, on dead leaves of Turpinia pomifera, Jan. 1973, W. Gams, holotype CBS H-15611, culture ex-type CBS 873.73.
Ramichloridium Stahel ex de Hoog, Stud. Mycol. 15: 59. 1977.
In vitro: Colonies flat to raised, with entire margin; surface olivaceous-green to olivaceous-black. Mycelium consisting of submerged and aerial hyphae; submerged hyphae hyaline to subhyaline, thin-walled, aerial hyphae smooth or verrucose. Conidiophores straight, unbranched, rarely branched, thick-walled, dark brown (darker than the subtending hyphae), continuous or with several additional thin septa. Conidiogenous cells integrated, terminal, polyblastic, smooth, thick-walled, golden-brown, apical part subhyaline, with sympodial proliferation, straight or flexuose, geniculate or nodose, with conspicuous conidiogenous loci; scars crowded or scattered, unthickened, unpigmented to faintly pigmented, or slightly prominent denticles. Conidia solitary, 0-1-septate, subhyaline to pale brown, smooth to coarsely verrucose, rather thin-walled, obovate, obconical or globose to ellipsoidal, fusiform, with a somewhat prominent, slightly pigmented hilum; conidial secession schizolytic.
Type species: R. apiculatum (J.H. Mill., Giddens & A.A. Foster) de Hoog, Stud. Mycol. 15: 69. 1977.
Ramichloridium apiculatum (J.H. Mill., Giddens & A.A. Foster) de Hoog, Stud. Mycol. 15: 69. 1977. Fig. 8.
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Veronaea apiculata (J.H. Mill., Giddens & A.A. Foster)
M.B. Ellis, in Ellis, More Dematiaceous Hyphomycetes: 209. 1976. [non Rhinocladiella apiculata Matsush., in Matsushima, Icon. Microfung. Mats. lect.: 122. 1975].
[non Chloridium indicum Subram., Proc. Indian Acad. Sci., Sect. B, 42: 286. 1955].
In vitro: Submerged hyphae hyaline to subhyaline, thin-walled, 1-2.5 µm wide; aerial hyphae slightly darker, smooth-walled. Conidiophores generally arising at right angles from creeping aerial hyphae, straight, unbranched, thick-walled, dark brown, continuous or with 1-2(-3) additional thin septa, up to 100 µm long; intercalary cells 10-28 µm long. Conidiogenous cells integrated, terminal, smooth, thick-walled, golden-brown, straight, cylindrical, 25-37(-47) x 2-3.5 µm; proliferating sympodially, resulting in a straight rachis with conspicuous conidiogenous loci; scars prominent, crowded, slightly pigmented, less than 1 µm diam. Conidia solitary, obovate to obconical, pale brown, finely verrucose, (3-)5-5.5(-7.5) x (2-)2.5-3(-4) µm, hilum conspicuous, slightly pigmented, about 1 µm diam.
Cultural characteristics: Colonies on MEA reaching 35 mm diam after 14 d at 24 °C; minimum temperature for growth above 6 °C, optimum 24 °C, maximum 30 °C. Colonies raised, velvety, dense, with entire margin; surface olivaceous-green, reverse olivaceous-black, often with a diffusing citron-yellow pigment.
Specimens examined: Pakistan, Lahore, from soil, A. Kamal, CBS 400.76 = IMI 088021. South Africa, from preserved Cucumis sativus in 8-oxyquinoline sulphate, M.C. Papendorf, CBS 390.67; Potchefstroom, from Aloe sp., M.C. Papendorf, CBS 391.67. U.S.A., Georgia, isolated from forest soil, CBS 156.59 = ATCC 13211 = IMI 100716 = QM 7716, ex-type culture.
Ramichloridium australiense Arzanlou & Crous, sp. nov. MycoBank MB504548. Figs 9-10A.
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Etymology: Named after its country of origin, Australia.
Ab aliis speciebus Ramichloridii conidiophoris ex hyphis verrucosis, crassitunicatis ortis distinguendum.
In vitro: Submerged hyphae hyaline, smooth, thin-walled, 1-2 µm wide; aerial hyphae pale brown, warted. Conidiophores arising vertically and clearly differentiated from creeping aerial hyphae, up to 400 µm tall, with several additional thin septa; intercalary cells, 8-40 x 2-5 µm, from the broadest part at the base tapering towards the apex, subhyaline, later becoming pale brown and warted in the lower part. Subtending hyphae thick-walled, warted. Conidiogenous cells integrated, terminal, 10-18 µm long, proliferating sympodially, giving rise to a short rachis with conspicuous conidiogenous loci; scars slightly thickened and darkened, about 1 µm diam. Conidia solitary, aseptate, thin-walled, smooth, subhyaline, subcylindrical to obclavate, (10-)12-15(-23) x 2.5-3 µm, with a truncate base and a slightly darkened and thickened hilum, 1.5-2 µm diam, rarely fusing at the basal part.
Cultural characteristics: Colonies on MEA rather slow growing, reaching 8 mm diam after 14 d at 24 °C, with entire, smooth margin; mycelium flat, olivaceous-grey, becoming granular, with gelatinous droplets at the margin developing with aging; reverse pale olivaceous-grey.
Specimen examined: Australia, Queensland, Mount Lewis, Mount Lewis Road, 16°34'47.2" S, 145°19'7" E, 538 m alt., on Musa banksii leaf, Aug. 2006, P.W. Crous and B. Summerell, holotype CBS H-19928, culture ex-type CBS 121710.
Ramichloridium musae (Stahel ex M.B. Ellis) de Hoog, Stud. Mycol. 15: 62. 1977. Fig. 11.
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Chloridium musae Stahel, Trop. Agric., Trinidad 14: 43.
1937 (nom. inval. Art. 36). Misapplied name: Chloridium indicum Subram., sensu Batista & Vital, Anais Soc. Biol. Pernambuco 15: 379. 1957.
In vitro: Submerged hyphae smooth, hyaline, thin-walled, 1-2 µm wide; aerial hyphae subhyaline, smooth. Conidiophores arising vertically and mostly sharply differentiated from creeping aerial hyphae, golden-brown; unbranched, rarely branched in the upper part, up to 250 µm tall, with up to 6 additional thin septa, cells 23-40 x 2-2.5 µm, basal cell occasionally inflated. Conidiogenous cells terminally integrated, cylindrical, variable in length, 10-40 µm long, golden-brown near the base, subhyaline to pale brown near the end, fertile part as wide as the basal part, later also becoming septate; rachis elongating sympodially, 2-2.5 µm wide, with hardly prominent, scattered, slightly pigmented scars, about 0.5 µm diam. Conidia solitary, aseptate, hyaline to subhyaline, ellipsoidal, (4-)7-8(-12) x 2-3 µm, smooth or verruculose, thin-walled, with slightly darkened hilum, about 1 µm diam.
Cultural characteristics: Colonies on MEA slow-growing, reaching 27 mm diam after 14 d at 24 °C, with entire, smooth, sharp margin; mycelium mostly submerged, some floccose to lanose aerial mycelium in the olivaceous-grey centre, becoming pale pinkish olivaceous towards the margin; reverse pale orange.
Specimens examined: cameroon, from Musa sapientum, J.E. Heron, CBS 169.61 = ATCC 15681 = IMI 079492 = DAOM 84655 = MUCL 2689; from Musa sapientum, J. Brun, CBS 190.63 = MUCL 9557. Surinam, Paramaribo, from Musa sapientum leaf, G. Stahel, CBS 365.36 = JCM 6973 = MUCL 9556, ex-type strain of Chloridium musae; from Musa sapientum, G. Stahel, CBS 365.36; dried culture preserved as CBS H-19933.
Ramichloridium biverticillatum Arzanlou & Crous, nom. nov. MycoBank MB504549. Fig. 12.
Basionym: Periconiella musae Stahel ex M.B. Ellis, Mycol. Pap. 111: 5. 1967.
[non Ramichloridium musae (Stahel ex M.B. Ellis) de Hoog, 1977].
Ramichloridium musae Stahel, Trop. Agric., Trinidad 14:
43. 1937 (nom. inval. Art. 36). Etymology: Named after its biverticillate conidiophores.
In vitro: Submerged hyphae smooth, hyaline, thin-walled, 1-2 µm wide; aerial hyphae subhyaline, smooth, slightly darker. Conidiophores arising vertically from creeping aerial hyphae, pale brown, profusely branched, biverticillate, with up to three levels of main branches; branches tapering distally, 2-3 µm wide at the base, approx. 2 µm wide in the upper part, up to 250 µm long. Conidiogenous cells terminally integrated, cylindrical, variable in length, 15-50 µm long, rachis straight or geniculate, pale brown, as wide as the basal part; elongating sympodially, forming a rachis with crowded, slightly darkened and thickened minute scars, less than 0.5 µm wide. Conidia solitary, aseptate, hyaline to subhyaline, dacryoid to pyriform, (2-)3-4(-6) x (1.5-)2(-2.5) µm, smooth, thin-walled, with an inconspicuous hilum.
Cultural characteristics: Colonies on MEA slow-growing, reaching 16 mm diam after 14 d at 24 °C, with entire, smooth, sharp margin, rather compact, velvety; surface vinaceous-buff to olivaceous-buff; reverse buff.
Specimen examined: Surinam, from Musa sapientum, Aug. 1936, G. Stahel, CBS 335.36.
Notes: Ramichloridium biverticillatum is a new name based on Periconiella musae. The species is distinct from R. musae because of its profusely branched conidiophores, and conidia that are smaller (2-5 x 1.5-2.5 µm) than those of R. musae (5-11 x 2-3 µm).
Ramichloridium brasilianum Arzanlou & Crous, sp. nov. MycoBank MB504550. Figs 10B, 13.
Etymology: Named after its country of origin, Brazil.
A simili Ramichloridio cerophilo conidiis minoribus, ad 8 µm longis, et conidiis secundariis absentibus distinguendum.
In vitro: Submerged hyphae pale olivaceous, smooth or slightly rough, 1.5-2 µm wide; aerial hyphae olivaceous, smooth or rough, narrower and darker than the submerged hyphae. Conidiophores unbranched, arising vertically from creeping aerial hyphae, straight or flexuose, dark brown, with up to 10 additional septa, thick-walled, cylindrical, 2-2.5 µm wide and up to 70 µm long. Conidiogenous cells integrated, terminal, 10-30 µm long, proliferating sympodially, giving rise to a long, straight rachis with crowded, slightly darkened minute scars, about 0.5 µm diam. Conidia solitary, obovoid to fusiform with the widest part below the middle, thin-walled, verruculose, aseptate, pale brown, slightly rounded at the apex, truncate at the base, (4-)5-6(-8.5) x 2-2.5(-3) µm, with a slightly thickened and darkened hilum, 1-1.5 µm diam.
Cultural characteristics: Colonies on MEA slow-growing, reaching 6 mm diam after 14 d at 24 °C, velvety to hairy, colonies with entire margin, surface dark olivaceous-grey; black gelatinous exudate droplets produced on OA.
Specimen examined: Brazil, São Paulo, Peruibe, Jureia Ecological Reserve, forest soil, Jan. 1991, D. Attili, holotype CBS H-19929, culture ex-type CBS 283.92.
Ramichloridium cerophilum (Tubaki) de Hoog, Stud. Mycol. 15: 74. 1977. Fig. 14.
Basionym: Acrotheca cerophila Tubaki, J. Hattori Bot. Lab. 20: 143. 1958.
Cladosporium cerophilum (Tubaki) Matsush., in Matsushima,
Icon. Microfung. Matsush. lect. (Kobe): 34. 1975. In vitro: Submerged hyphae pale olivaceous-brown, smooth or slightly rough, 1.5-3 µm wide; aerial hyphae olivaceous-brown, smooth or slightly rough, somewhat narrower and darker than the submerged hyphae. Conidiophores unbranched, arising vertically from creeping aerial hyphae, dark brown, thick-walled, smooth or verruculose, hardly tapering towards the apex, 2-3 µm wide, up to 50 µm long, with up to 3 additional septa. Conidiogenous cells integrated, terminal, proliferating sympodially, rachis short and straight, with crowded, prominent, pigmented unthickened scars, minute, approx. 0.5 µm diam. Conidia solitary, fusiform to clavate, thin-walled, smooth, 0(-1)-septate, subhyaline, (4-)6-7(-11) x (2-) 2.5(-3) µm, with a conspicuous hilum, about 0.5 µm diam, slightly raised with an inconspicuous marginal frill, somehow resembling those of Cladosporium. Conidia sometimes producing 1-3(-4) short secondary conidia.
Cultural characteristics: Colonies on MEA rather slow-growing, reaching 12 mm diam after 14 d at 24 °C, velvety to hairy, with entire margin; surface dark olivaceous-grey, with black gelatinous exudate droplets on OA.
Specimen examined: Japan, isolated from Sasa sp., K. Tubaki, CBS 103.59, ex-type.
Notes: Phylogenetically, this species together with Ramichloridium apiculatum and R. musae cluster within the Mycosphaerellaceae clade. Ramichloridium cerophilum can be distinguished from its relatives by the production of secondary conidia and its distinct conidial hila.
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Basionym: Chloridium indicum Subram., Proc. Indian Acad. Sci., Sect. B, 42: 286. 1955 [non Rhinocladiella indica Agarwal, Lloydia 32: 388. 1969].
Veronaea indica (Subram.) M.B. Ellis, in Ellis, More
Dematiaceous Hyphomycetes: 209. 1976. In vitro: Submerged hyphae smooth, thin-walled, hyaline, 1-2.5 µm wide, with thin septa; aerial hyphae coarsely verrucose, olivaceous-green, rather thick-walled, 2-2.5 µm wide, with thin septa. Conidiophores arising vertically from creeping hyphae at right angles, straight, unbranched, thick-walled, smooth, dark brown, with up to 10 thin septa, up to 250 µm long, 2-4 µm wide, often with inflated basal cells. Conidiogenous cells terminally integrated, up to 165 µm long, smooth, dark brown, sympodially proliferating, rachis straight or flexuose, geniculate or nodose, subhyaline; scars thickened and darkened, clustered at nodes, approx. 0.5 µm diam. Microcyclic conidiation observed in culture. Conidia solitary, (0-)1-septate, not constricted at the septum, subhyaline to pale brown, smooth or coarsely verrucose, rather thin-walled, broadly ellipsoidal to globose, (5-)7-8(-10) x (4-)6-6.5(-9) µm, with truncate base; hilum conspicuous, slightly darkened, not thickened, about 1 µm diam.
Cultural characteristics: Colonies on MEA reaching 35 mm diam after 14 d at 24 °C. Colonies velvety, rather compact, slightly elevated, with entire, smooth, whitish margin, dark olivaceous-green in the central part.
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Ramichloridium pini de Hoog & Rahman, Trans. Brit. Mycol. Soc. 81: 485. 1983.
Specimen examined: U.K., Scotland, Old Aberdeen, branch of Pinus contorta (Pinaceae), 1982, M.A. Rahman, ex-type strain, CBS 461.82 = MUCL 28942.
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Ramichloridium strelitziae Arzanlou, W. Gams & Crous, sp. nov. MycoBank MB504551. Figs 16-17A.
Etymology: Named after its host, Strelitzia.
Ab aliis speciebus Ramichloridii conidiophoris brevibus, ad 40 µm longis, et cicatricibus rotundis, paulo protrudentibus distinguendum.
In vitro: Submerged hyphae smooth, hyaline, thin-walled, 2-2.5 µm wide; aerial hyphae pale brown, verrucose. Conidiophores arising vertically from creeping aerial hyphae, clearly differentiated from the vegetative hyphae, subhyaline, later becoming pale brown, thick-walled, smooth or verruculose, with 1-3 additional septa; up to 40 µm long and 2 µm wide. Conidiogenous cells integrated, terminal, cylindrical, variable in length, 10-35 µm long, subhyaline, later turning pale brown, fertile part as wide as the basal part, proliferating sympodially, forming a straight rachis with slightly thickened and darkened, circular, somewhat protruding scars, approx. 0.5 µm diam. Conidia solitary, aseptate, smooth or verruculose, subhyaline, oblong, ellipsoidal to clavate, (3-)4-5(-5.5) x (1-)2(-2.5) µm, with truncate base and unthickened, non-pigmented hilum.
Cultural characteristics: Colonies on MEA slow-growing, reaching 5 mm diam after 14 d at 24 °C, with entire margin; aerial mycelium rather compact, raised, dense, olivaceous-grey; reverse olivaceous-black.
Specimen examined: South Africa, KwaZulu-Natal, Durban, near Réunion, on leaves of Strelitzia nicolai, 5 Feb. 2005, W. Gams & H. Glen, CBS-H 19776, holotype, culture ex-type CBS 121711.
Zasmidium Fr., Summa Veg. Scand. 2: 407. 1849.
In vitro: Submerged hyphae smooth, thin-walled, hyaline, with thin septa; aerial hyphae coarsely verrucose, olivaceous-green, thick-walled, with thin septa. Conidiophores not differentiated from vegetative hyphae, often reduced to conidiogenous cells. Conidiogenous cells integrated, predominantly terminal, sometimes lateral, arising from aerial hyphae, cylindrical, pale brown; polyblastic, proliferating sympodially producing crowded, conspicuously pigmented, almost flat, darkened, somewhat refractive scars. Conidia in short chains, cylindrical to fusiform, verrucose, obovate to obconical, pale brown, base truncate, with a conspicuous, slightly pigmented, thickened and refractive hilum. Primary conidia sometimes larger, subhyaline, verrucose or smooth-walled, 0-4-septate, variable in length, fusiform to cylindrical; conidial secession schizolytic.
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Zasmidium cellare (Pers. : Fr.) Fr., Summa Veg. Scand. 2: 407. 1849. Fig. 18.
Basionym: Racodium cellare Pers., Neues Mag. Bot. 1: 123. 1794.
Antennaria cellaris (Pers. : Fr.) Fr., Syst. Mycol. 3:
229. 1829. Cladosporium cellare (Pers. : Fr.) Schanderl, Zentralbl.
Bakteriol., 2. Abt., 94: 117. 1936. Rhinocladiella cellaris (Pers. : Fr.) M.B. Ellis, in
Ellis, Dematiaceous Hyphomycetes: 248. 1971. In vitro: Submerged hyphae smooth, thin-walled, hyaline, 2-3 µm wide, with thin septa; aerial hyphae coarsely verrucose, olivaceous-green, rather thick-walled, 2-2.5 µm wide, with thin septa. Conidiophores not differentiated from vegetative hyphae, often reduced to conidiogenous cells. Conidiogenous cells integrated, predominantly terminal, sometimes lateral, arising from aerial hyphae, cylindrical, 20-60 µm long and 2-2.5 µm wide, pale brown, proliferating sympodially producing crowded, conspicuously pigmented scars that are thickened and refractive, about 1 µm diam. Conidia cylindrical to fusiform, verrucose, obovate to obconical, pale brown, with truncate base, (6-)9-14(-27) x 2-2.5 µm, with a conspicuous, slightly pigmented, refractive hilum, approx. 1 µm diam. Primary conidia sometimes subhyaline, verrucose or smooth-walled, thin-walled, 0-1(-4)-septate, variable in length, fusiform to cylindrical.
Cultural characteristics: Colonies reaching 7 mm diam after 14 d at 24 °C. Colonies velvety, rather compact, slightly elevated with entire margin; surface dark olivaceous-green in the central part, margin smooth, whitish.
Specimen examined: Wall in wine cellar, Jun. 1936, H. Schanderl, ATCC 36951 = IFO 4862 = IMI 044943 = LCP 52.402 = LSHB BB274 = MUCL 10089 = CBS 146.36.
Notes: The name Racodium Fr., typified by Ra. rupestre Pers. : Fr., has been conserved over the older one by Persoon, with Ra. cellare as type species. De Hoog (1979) defended the use of Zasmidium in its place for the well-known wine-cellar fungus.
Morphologically Zasmidium resembles Stenella Syd., and both reside in the Capnodiales, though the type of Stenella, S. araguata Syd., clusters in the Teratosphaeriaceae, and the type of Zasmidium, Z. cellare, in the Mycosphaerellaceae. When accepting anamorph genera as polyphyletic within an order, preference would be given to the well-known name Stenella over the less known Zasmidium, even though the latter name is older. Further studies are required, however, to clarify if all stenella-like taxa should be accommodated in a single genus, Stenella. If this is indeed the case, a new combination for Zasmidium cellare will be proposed in Stenella, and the latter genus will have to be conserved over Zasmidium.
Chaetothyriales (Herpotrichiellaceae)
The four "Ramichloridium" species residing in the
Chaetothyriales clade do not differ sufficiently in morphology to
separate them from Rhinocladiella (type Rh. atrovirens).
Because of the pale brown conidiophores, conidiogenous cells with crowded,
slightly prominent scars and the occasional presence of an Exophiala
J.W. Carmich. synanamorph, Rhinocladiella is a suitable genus to
accommodate them. These four species chiefly differ from
Ramichloridium in the morphology of their conidial apparatus, which
is clearly differentiated from the vegetative hyphae. The appropriate
combinations are therefore introduced for Ramichloridium anceps, R.
mackenziei, R. fasciculatum and R. basitonum.
The genus Veronaea (type species: V. botryosa) also resides in the Chaetothyriales clade. Veronaea can be distinguished from Rhinocladiella by the absence of exophiala-type budding cells and its predominantly 1-septate conidia. Furthermore, the conidiogenous loci in Veronaea are rather flat, barely prominent.
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In vitro: Colonies dark olivaceous-brown, slow-growing, almost moist. Submerged hyphae hyaline to pale olivaceous, smooth; aerial hyphae, if present, more darkly pigmented. Exophiala-type budding cells usually present in culture. Conidial apparatus usually branched, olivaceous-brown, consisting of either slightly differentiated tips of ascending hyphae or septate, markedly differentiated conidiophores. Conidiogenous cells intercalary or terminal, polyblastic, cylindrical to acicular, with a sympodially proliferating, subdenticulate rachis; scars unthickened, non-pigmented to somewhat darkened-refractive. Conidia solitary, hyaline to subhyaline, aseptate, thin-walled, smooth, subglobose, with a slightly pigmented hilum; conidial secession schizolytic.
Type species: Rh. atrovirens Nannf., Svenska Skogsvårdsfören. Tidskr. 32: 461. 1934.
Rhinocladiella anceps (Sacc. & Ellis) S. Hughes, Canad. J. Bot. 36: 801. 1958. Fig. 19.
Basionym: Sporotrichum anceps Sacc. & Ellis, Michelia 2: 576. 1882.
Misapplied name: Chloridium minus Corda sensu Mangenot, Rev. Mycol. (Paris) 18: 137. 1953.
In vitro: Submerged hyphae subhyaline, smooth, thick-walled, 2-2.5 µm wide; aerial hyphae pale brown. Swollen germinating cells often present on MEA, giving rise to an Exophiala synanamorph. Conidiophores slightly differentiated from vegetative hyphae, arising from prostrate aerial hyphae, consisting of either unbranched or loosely branched stalks, thick-walled, golden to dark-brown, up to 350 µm tall, which may have up to 15 thin, additional septa, intercalary cells 9-14 µm long. Conidiogenous cells terminal, rarely lateral, cylindrical, occasionally intercalary, variable in length, smooth, golden to dark brown at the base, paler toward the apex, later becoming inconspicuously septate, fertile part as wide as the basal part, 15-40 x 1.5-2 µm; with crowded, slightly prominent, unpigmented, conidium-bearing denticles, about 0.5 µm diam. Conidia solitary, subhyaline, thin-walled, smooth, subglobose to ellipsoidal, 2.5-4 x 2-2.5 µm, with a less conspicuous, slightly darkened hilum, less than 0.5 µm diam.
Cultural characteristics: Colonies on MEA reaching 6-12 mm diam after 14 d at 24 °C, with entire, smooth, sharp margin; mycelium powdery, becoming hairy at centre; olivaceous-green to brown, reverse dark-olivaceous.
Specimens examined: Canada<
