Phenotypic study

For cultural characterization, suspensions of conidia were prepared in a semi-solid medium (0.2% agar; 0.05% Tween 80) and inoculated onto phytone yeast extract agar (PYE; Becton, Dickinson & Company, Sparks, MD, USA; Carmichael and Kraus 1959), potato dextrose agar (PDA; Pronadisa, Madrid, Spain; Hawksworth et al. 1995), oatmeal agar (OA; 30g of filtered oat flakes, 15g agar-agar, 1L tap water; Samson et al. 2010), bromocresol purple-milk solids-glucose agar (BCP-MS-G; 80g skim milk powder, 40g glucose, 10mL of 1.6% of bromocresol purple in 95% ethanol, 30g agar-agar,1L tap water; Kane and Smitka 1978), and test opacity tween medium (TOTM; 10g bacteriological peptone, 5g NaCl, 1g CaCl₂, 5mL Tween, 5mL Tween 80, 15g agar-agar, 1L tap water; Slifkin 2000). Colonies were characterized after 14days at 25°C in the dark. Potato dextrose agar (PDA) was used to determine the cardinal temperatures of growth. Colour notations were taken according to Kornerup and Wanscher (1978). Christensen’s urea agar (EMD Millipore, Darmstadt, Germany; Christensen 1946) was inoculated and incubated for 4days at 25°C in the dark to detect the production of urease. Cycloheximide tolerance was tested growing the fungal strains on Sabouraud dextrose agar (SDA; Pronadisa, Spain) supplemented with 0.2% cycloheximide (Sigma, USA) at 30°C for two wk. Fungal tolerance to NaCl was evaluated on SDA adding 3, 10 and 20% w/w NaCl, with the same incubation conditions as previously described. The microscopic structures were characterized and measured from wet mountings of slide cultures, using water and 60% lactic acid. Photo micrographs were taken using a Zeiss Axio-Imager M1 light microscope (Oberkochen, Germany) with a DeltaPix Infinity X digital camera using Nomarski differential interference contrast. The descriptions of the taxonomical novelties were submitted to MycoBank (https://www.mycobank.org/; Crous et al. 2004).

DNA extraction, amplification and sequencing

Total DNA was extracted as previously described (Valenzuela-Lopez et al. 2018), and the following phylogenetic markers were amplified: the internal transcribed spacers (ITS) (ITS5/ITS4 primers; White et al. 1990, and a fragment of the large subunit (LSU) gene (LR0R/LR5 primers; Vilgalys and Hester 1990; Rehner and Samuels 1994) of the nrDNA. Amplicons were sequenced at Macrogen Europe (Macrogen Inc., Madrid, Spain) using the same pair of primers. Consensus sequences were obtained by SeqMan software v. 7 (DNAStar Lasergene, Madison, WI, USA). Sequences generated in this work were deposited in GenBank (Table ​(Table11).

Phylogenetic analysis

A preliminary molecular identification of the isolates was carried out with ITS and LSU nucleotide sequences using BLAST (https://blast.ncbi.nlm.nih.gov/Blast.cgi), and only the sequences of ex-type or reference strains from GenBank were included for identification. A maximum level of identity (MLI)≥98% was used for species-level and<98% for genus-level identification. A maximum-likelihood (ML) and Bayesian-inference (BI) phylogenetic analyses of the concatenated ITS-LSU sequences were performed in order to determine the phylogenetic placement of our clinical strains. Species of the order Arachnomycetales were used as outgroup. The sequence alignments and ML / BI analyses were performed according to Valenzuela-Lopez et al. (2018). The final matrices used for the phylogenetic analysis were deposited in TreeBASE (www.treebase.org; accession number: 25068).

Molecular phylogeny

Our phylogenetic study included 92 sequences corresponding to 75 species with a total of 1213 characters (700 ITS and 513 LSU) including gaps, of which 579 were parsimony informative (402 ITS and 177 LSU). The ML analysis was congruent with that obtained in the BI analysis, both displaying trees with similar topologies. The datasets did not show conflict with the tree topologies for the 70% reciprocal bootstrap trees, which allowed the two genes to be combined for the multi-locus analysis (Fig. 2). (single gene-based phylogenies are as supplemental material Figures S1 and S2).

Open in a separate window

Fig. 2

ML phylogenetic tree based on the analysis of ITS-LSU nucleotide sequences for the 22 clinical fungi from the USA. Bootstrap support values/Bayesian posterior probability scores of 70/0.95 and higher are indicated on the nodes. ^T=ex type. Fully supported branched (100% BS /1 PP) are indicated in bold. Strains identified by us are in bold. Arachnomyces spp. were chosen as out-group. The sequences used in this analysis are in Table ​Table11

Twenty of our strains were placed into a main clade corresponding to the members of the Onygenales (100% BS / 1 PP), while two were placed in the Arachnomycetales (100% BS / 1 PP) (Fig. ​(Fig.2).2). The Onygenales clade was divided into eight clades corresponding to the families Onygenaceae (100% BS / 1 PP), Gymnascaceae (98% BS / 0.99 PP), Nannizziopsiaceae (100% BS / 1 PP), Helicoarthrosporaceae (100% BS / 1 PP), Arthrodermataceae (100% BS / 0.99 PP), Ajellomycetaceae (91% BS / 0.98 PP), Ascosphaeraceae (100% BS / 1 PP), and Spiromastigaceae (91% BS / 0.95 PP), which included a basal terminal branch for Pseudospiromastix tentaculata. Most of our strains (17/22) were distributed into several subclades of the Onygenaceae: 15/22 into Auxarthron/Malbranchea subclade (100% BS / 1 PP), one into a terminal branch (FMR 17683) together Currahmyces indicus (100% BS / 1 PP), and another one (FMR 17684) into a distant, independent terminal branch. One strain (FMR 17692) was placed into the Gymnascaceae, in a terminal branch together with Arthropsis cirrhata (98% BS / 1 PP). The Spiromastigaceae included the last two strains (FMR 17686 and FMR 17696 (CBS 146934)), placed into a terminal branch together Malbranchea gypsea (100% BS / 1 PP).

TAXONOMY

Arachnomyces

Since the strains FMR 17685 and FMR 17691 represented two species of Arachnomyces that were different from the other species of the genus, they are described as new, here.

Arachnomyces bostrychodes Rodr.-Andr., Cano & Stchigel, sp. nov.

(Fig. ​Fig.33)

Open in a separate window

Fig. 3

Arachnomyces bostrychodes CBS 146926 ^T. a Colonies on PYE, PDA and OA after 14 d at 25°C, from left to right (top row, surface; bottom row, reverse). b Detail of the colony on OA. c, d Sinuous, contorted to coiled fertile hyphae. e Arthroconidia. Scale bar=10μm

MycoBank MB 834921

Etymology: From Greek βοστρυχος-, curl, due to the appearance of the reproductive hyphae.

Diagnosis: The phylogenetically closest species to Arachnomyces bostrychodes is A. peruvianum (Fig. ​(Fig.2).2). Nevertheless, A. botrychodes lacks a sexual morph and racket hyphae (both present in A. peruvianum), and produces longer conidia than A. peruvianum (4.0–8.0×1.0–2.0μm vs. 4.0–5.0×1.0–3.0μm); also, A. bostrychodes grows more slowly on OA (13–14mm diam after 2 wk. at 25°C) than A. peruvianum (30mm diam) (Cain 1957; Brasch et al. 2017). Arachnomyces bostrychodes morphologically resembles Arachnomyces gracilis, but the former grows faster, produces more strongly contorted branches and lacks of a sexual morph.

Type: USA: Texas: from a human scalp, 2008, N. Wiederhold (CBS H-24452 – holotype; CBS 146926=FMR 17685=UTHSCSA DI18-91 – ex-type cultures; LSU/ITS sequences GenBank LR701766/LR701765).

Description: Vegetative hyphae hyaline, septate, branched, smooth- and thin-walled, 1.0–2.0μm wide. Fertile hyphae well-differentiated, arising as lateral branches from the vegetative hyphae, successively branching to form dense clusters, arcuate, sinuous, contorted or tightly curled, 1.0–2.0μm wide, forming randomly intercalary and terminally arthroconidia. Conidia enteroarthric, hyaline, one-celled, smooth-walled, cylindrical, barrel-shaped, and finger-like-shaped when terminal, 4.0–8.0×1.0–2.0μm, mostly curved and truncated at one or (mostly) both ends, separated from the fertile hyphae by rhexolysis. Chlamydospores, racquet hyphae, setae, and sexual morph not observed.

Culture characteristics: Colonies on PYE reaching 19–20mm diam after 2 wk. at 25°C, elevated, cottony, margins regular, white (5A1), sporulation absent; reverse light orange (5A4). Colonies on PDA reaching 11–12mm diam after 2 wk. at 25°C, elevated, velvety with floccose patches, margins regular, yellowish white (4A2), sporulation abundant; reverse greyish yellow (4B6). Colonies on PDA reaching 13–14mm diam after 2 wk. at 30°C, slightly elevated, velvety to floccose, regular margins, white (4A1), sporulation sparse; reverse, greyish yellow (4B6). Colonies on OA researching 13–14mm diam after 2 wk. at 25°C, flattened, smooth and granulose, irregular margins, yellowish white (2A2) at centre and light yellow (2A5) at edge, sporulation abundant. Exudate and diffusible pigment absent.

Minimum, optimal and maximum temperature of growth (on PDA): 10°C, 30°C, and 37°C, respectively. Non-haemolytic. Casein not hydrolysed. Not inhibited by cycloheximide. Urease and esterase (TOTM) tests positive. Growth occurs at NaCl 10% w/w, but not at 20% w/w.

Arachnomyces graciliformis Rodr.-Andr., Stchigel & Cano, sp. nov.

(Fig. ​(Fig.44)

Open in a separate window

Fig. 4

Arachnomyces graciliformis CBS 146927 ^T. a Colonies on PYE, PDA and OA after 14 d at 25°C, from left to right (top row, surface; bottom row, reverse). b Detail of the colony on OA. c–e Contorted, apically coiled fertile hyphae bearing arthroconidia. Scale bar=10μm

MycoBank MB 834923

Etymology: Recalling the morphological similarity with Arachnomyces gracilis.

Diagnosis: Arachnomyces graciliformis is phylogenetically close to A. glareosus and to A. minimus (Fig. ​(Fig.2).2). These three species form a clade together with A. nodosetosus and A. jinanicus (84 BS / 1 PP). Unlike A. glareosus and A. minimus, A. graciliformis does not produces racquet hyphae nor sexual morph (Gibas et al. 2004) but produces longer conidia than A. glareosus (4.0–10.0×1.5–2.0μm vs. 2.5–4.5×1.5–2.0μm), which are not produced by A. minimus. Arachnomyces graciliformis morphologically resembles A. gracilis, but the former grows more slowly, produces more twisted fertile branches and does not form a sexual morph (Udagawa and Uchiyama 1999).

Type: USA: Massachusetts: from an animal’s bone, 2012, N. Wiederhold (CBS H-24453 – holotype; CBS 146927=FMR 17691=UTHSCSA DI18-97 – ex-type cultures; LSU/ITS sequences GenBank LR743668/LR743667).

Description: Vegetative hyphae hyaline, septate, branched, smooth- and thin-walled, 1.0–2.0μm wide. Fertile hyphae well-differentiated, arising as lateral branches from the vegetative hyphae, branching repeatedly, sinuous to arcuate or apically coiled, 1.5–2.0μm wide, forming randomly intercalary and terminally arthroconidia. Conidia enteroarthric, hyaline, unicellular, smooth- and thin-walled, cylindrical or finger-like-shaped when terminal, 4.0–10.0×1.5–2.0μm, mostly curved, detached from the fertile hyphae by rhexolysis. Chlamydospores, racquet hyphae, setae, and sexual morph not observed.

Culture characteristics: Colonies on PYE reaching 12–13mm diam after 2 wk. at 25°C, elevated, velvety to floccose, margins regular, slightly furrowed, yellowish white (3A2), sporulation absent; reverse greyish orange (5B3). Colonies on PDA reaching 9–10mm diam after 2 wk. at 25°C, slightly elevated, velvety to floccose, margins regular, slightly furrowed, yellowish white (1A2), sporulation absent; reverse greyish yellow (4B3). Colonies on PDA reaching 3–4mm diam after 2 wk. at 30°C, slightly elevated, velvety to floccose, margins regular, slightly furrowed, yellowish white (1A2), sporulation absent; reverse, greyish yellow (4B3). Colonies on OA researching 6–7mm diam after 2 wk. at 25°C, flattened, velvety and granulose, margins irregular, pale yellow (4A3), sporulation absent (conidia appear after 5–6 wk. incubation). Exudate and diffusible pigment absent. Minimum, optimal and maximum temperature of growth (on PDA): 10°C, 25°C, and 30°C, respectively. Non-haemolytic. Casein not hydrolysed. Not inhibited by cycloheximide. Urease and esterase tests positive. Growth occurs at NaCl 10% w/w, but not at 20% w/w.

Currahmyces

Due to the strain FMR 17683 being placed into a terminal branch of Onygenaceae together with Currahmyces indicus (Sharma and Shouche 2019), and because they differ molecularly and phenotypically, we erect the new species Currahmyces sparsispora.

Currahmyces sparsispora Rodr.-Andr., Cano & Stchigel, sp. nov.

(Fig. ​Fig.55)

Open in a separate window

Fig. 5

Currahmyces sparsispora CBS 146929 ^T. a Colonies on PYE, PDA and OA after 14 d at 25°C, from left to right (top row, surface; bottom row, reverse). b Detail of the colony on OA. c–d Intercalary arthroconidia along the fertile hyphae. Scale bar=10μm

MycoBank MB 835692

Etymology: From Latin sparsa-, splashed, −sporarum, spore, due to the disposition of the conidia along the hyphae.

Diagnosis: Currahmyces sparsispora is phylogenetically close to C. indicus; however, they can be differentiated because the former has broader hyphae (1.5–2.0μm vs. 0.7–1.1μm) and lacks a sexual morph (typical gymnothecial ascomata are produced on hair-baited soil plates by C. indicus).

Type: USA: Florida: from human sputum, 2007, N. Wiederhold (CBS H-24455 – holotype; CBS 146929=FMR 17683=UTHSCSA DI18-89 – ex-type cultures; LSU/ITS sequences GenBank LR723273/LR723273).

Description: Vegetative hyphae septate, hyaline, smooth- and thin-walled, mostly straight, rarely branched, 1.5–2.0μm wide. Fertile hyphae undifferentiated from the vegetative hyphae. Conidia enteroarthric, hyaline, unicellular, smooth- and thin-walled, disposed relatively far from each other along the fertile hyphae, separated by 1–2 evanescent connective cells, cylindrical to slightly barrel-shaped, 3.0–12.0×1.0–2.0μm, separated by rhexolysis. Chlamydospores, racquet hyphae, setae, and sexual morph not observed.

Culture characteristics: Colonies on PYE reaching 27–28mm diam after 2 wk. at 25°C, slightly elevated, velvety to floccose, margins regular, pale orange (5A3) at centre and white (5A1) at edge, sporulation sparse; reverse orange (5A6). Colonies on PDA reaching 23–24mm diam after 2 wk. at 25°C, slightly elevated, velvety, margins regular, light orange (5A5) at centre and orange white (5A2) at edge, sporulation sparse; reverse deep orange (6A8). Colonies on PDA reaching 30–31mm diam after 2 wk. at 30°C, slightly elevated, velvety, slightly furrowed, margins regular, orange (5A6), sporulation sparse; reverse brownish orange (6C8). Colonies on OA reaching 20–21mm diam after 2 wk. at 25°C, slightly elevated, velvety, margins regular, orange white (5A2) at centre and white (5A1) at edge, sporulation sparse. Exudate and diffusible pigment absent in all culture media tested. Minimum, optimal and maximum temperature of growth on PDA: 10°C, 30°C, and 37°C, respectively. Haemolytic. Casein not hydrolysed. Not inhibited by cycloheximide. Urease and esterase tests positive. Growth occurs at NaCl 3% w/w and 10% w/w, but not at 20% w/w.

Malbranchea

An emended description of the genus Malbranchea is provided as follows:

Malbranchea Sacc., Michelia 2(no. 8): 639 (1882).

MycoBank MB 8833.

Description: Vegetative hyphae septate, hyaline, smooth- and thin-walled, straight or branched. Asexual morph consisting in undifferentiated fertile hyphae, and/or well-differentiated lateral branches, curved or not, which form randomly or basipetally terminal and intercalary arthroconidia. Conidia enteroarthric, rarely holoarthric, unicellular, hyaline, smooth- and thin-walled, mostly cylindrical, barrel-shaped, or irregularly shaped, detached from the fertile hyphae by rhexolysis. Sexual morph (when present) consisting in ascomata formed by of an anastomosing network of orange to brown, ornamented or not thick-walled hyphae (gymnothecia), bearing elongate appendages and/or spine projections, within there are small, evanescent, inflated asci which forms eight globose to oblate ascospores, whose cell wall is ornamented with a (coarse or thin) reticulate pattern. Species homothallic or heterothallic, thermotolerant or thermophilic, keratinolytic, chitinolytic or cellulolytic.

Taking into account that Auxarthron and Malbranchea are congeneric, as has been shown in previous studies (Sigler et al. 2002; Sarrocco et al. 2015) and here (Fig. ​(Fig.2),2), and that Malbranchea (Saccardo 1882) has historical priority (Turland et al. 2018) over Auxarthron (Orr et al. 1963), we transfer the species of Auxarthron to Malbranchea as follows:

Malbranchea californiensis (G.F. Orr & Kuehn) Rodr.-Andr., Stchigel & Cano, comb. nov.

MycoBank MB 835229

Basionym: Auxarthron californiense G.F. Orr & Kuehn, Can. J. Bot. 41: 1442 (1963).

Synonym: Gymnoascus californiensis (G.F. Orr & Kuehn) Apinis, Mycol. Pap. 96: 12 (1964).

Malbranchea chinensis (Z.F. Zhang & L. Cai) Rodr.-Andr., Cano & Stchigel, comb. nov. MycoBank MB 839604

Basionym: Auxarthron chinense Z.F. Zhang & L. Cai, Fungal Divers. 106: 55 (2020).

Malbranchea chlamydospora (M. Solé et al.) Rodr.-Andr., Cano & Stchigel, comb. nov.

MycoBank MB 835230

Basionym: Auxarthron chlamydosporum M. Solé, et al., Stud. Mycol. 47: 108 (2002).

Malbranchea compacta (G.F. Orr & Plunkett) Rodr.-Andr., Cano & Stchigel, comb. nov.

MycoBank MB 835231

Basionym: Auxarthron compactum G.F. Orr & Plunkett, Can. J. Bot. 41: 1453 (1963).

Malbranchea concentrica (M. Solé et al.) Rodr.-Andr., Stchigel & Cano, comb. nov.

MycoBank MB 835232

Basionym: Auxarthron concentricum M. Solé et al., Stud. Mycol. 47: 106 (2002).

Malbranchea conjugata (Kuehn) Rodr.-Andr., Cano & Stchigel, comb. nov.

MycoBank MB 835233

Basionym: Myxotrichum conjugatum Kuehn, Mycologia 47: 883 (1956) [“1955”].

Malbranchea guangxiensis (Z.F. Zhang & L. Cai) Rodr.-Andr., Cano & Stchigel, comb. nov.

MycoBank MB 839605

Basionym: Auxarthron guangxiense Z.F. Zhang & L. Cai, Fungal Divers. 106: 57 (2020).

Synonym: Auxarthron conjugatum (Kuehn) G.F. Orr & Kuehn, Mycotaxon 24: 148 (1985).

Malbranchea longispora (Stchigel et al.) Rodr.-Andr., Stchigel & Cano, comb. nov.

MycoBank MB 835235

Basionym: Auxarthron longisporum Stchigel et al., Persoonia 31: 267 (2013).

Malbranchea ostraviensis (Hubka et al.) Rodr.-Andr., Cano & Stchigel, comb. nov.

MycoBank MB 835236

Basionym: Auxarthron ostraviense Hubka et al., Med. Mycol. 50: 619 (2012).

Malbranchea pseudauxarthron (G.F. Orr & Kuehn) Rodr.-Andr., Stchigel & Cano, comb. nov.

MycoBank MB835237

Basionym: Auxarthron pseudauxarthron G.F. Orr & Kuehn, Mycologia 64: 67 (1972).

Malbranchea umbrina (Boud.) Rodr.-Andr., Cano & Stchigel, comb. nov.

MycoBank MB 835238

Basionym: Gymnoascus umbrinus Boud., Bull. Soc. mycol. Fr. 8: 43 (1892).

Synonyms: Auxarthron brunneum (Rostr.) G.F. Orr & Kuehn, Can. J. Bot.41: 1446 (1963).

Auxarthron umbrinum (Boud.) G.F. Orr & Plunkett, Can. J. Bot. 41: 1449 (1963).

Auxarthron thaxteri (Kuehn) G.F. Orr & Kuehn, Mycologia 63: 200 (1971).

Gymnoascus subumbrinus A.L. Sm. & Ramsb., Trans. Br. Mycol. Soc. 5: 424 (1917) [“1916”].

Gymnoascus umbrinus var. thaxteri (Kuehn) Apinis, Mycol. Pap. 96: 14 (1964).

Myxotrichum brunneum Rostr., Bot. Tidsskr. 19: 216 (1895).

Myxotrichum thaxteri Kuehn, Mycologia 47: 878 (1956) [“1955”].

Malbranchea zuffiana (Morini) Rodr.-Andr., Stchigel & Cano, comb. nov.

MycoBank MB 835239

Basionym: Gymnoascus zuffianus Morini, Mem. R. Accad. Sci. Ist. Bologna, ser. 4 10: 205 (1889).

Synonym: Auxarthron zuffianum (Morini) G.F. Orr & Kuehn, Can. J. Bot. 41: 1445 (1963).

We also update the Malbranchea species names listed below:

Malbranchea albolutea Sigler & J.W. Carmich., Mycotaxon 4: 416 (1976).

Synonym: Auxarthron alboluteum Sigler et al., Stud. Mycol. 47: 118 (2002).

Malbranchea filamentosa Sigler & J.W. Carmich., Mycotaxon 15: 468 (1982).

Synonym: Auxarthron filamentosum Sigler et al., Stud. Mycol. 47: 116 (2002).

Because in a BLAST search using the ITS and LSU nucleotide sequences from the ex-type strains, Malbranchea circinata and M. flavorosea match with taxa in the family Myxotrichaceae, both those species are excluded to the genus.

After examination of the lectotype of Auxarthron indicum (Patil and Pawar 1987, as “indica”), we concluded that this fungus must be excluded from Malbranchea because its sexual morph differs mainly from all species described for the former genus. Whereas Auxarthron indicum produces smooth-walled ellipsoidal ascospores and gymnothecial ascomata lacking of true appendages, in Malbranchea spp. the ascospores are globose and mostly ornamented, and the ascomata have appendages. Based on the fact that there is no type strain of this species available we consider it as of uncertain application.

Despite the strain FMR 17681 being placed phylogenetically close to Malbranchea ostraviense and M. umbrina, it differs genetically and phenotypically from both species, therefore we describe the new species Malbranchea gymnoascoides as follows:

Malbranchea gymnoascoides Rodr.-Andr., Stchigel & Cano, sp. nov.

(Fig. ​Fig.66)

Open in a separate window

Fig. 6

Malbranchea gymnoascoides CBS 146930 ^T. a Colonies on PYE, PDA and OA after 14 d at 25°C, from left to right (top row, surface; bottom row, reverse). b Detail of the colony on OA. c–d Young and mature ascomata. e Young ascus on fertile hyphae. f Peridial spine-like appendage. g Intercalary arthroconidia along the fertile hyphae. Scale bar=10μm

MycoBank MB 835212

Etymology: As the ascomata are morphologically like those of Gymnoascus reessii.

Diagnosis: Malbranchea gymnoascoides is phylogenetically close to M. ostraviensis and M. umbrina (Fig. ​(Fig.2).2). Nevertheless, M. gymnoascoides produces smaller ascomata (to 250μm diam in M. gymnoascoides vs. to 450 and to 600μm diam in both, M. ostraviensis and M. umbrina, respectively) (Orr et al. 1963; Hubka et al. 2013). Also, the peridial appendages of M. gymnoascoides are longer than those of M. umbrina (250–400μm vs. 5–72μm), but shorter than those of M. ostraviensis (350–600μm long). The ascospores of M. gymnoascoides are like those of M. ostraviensis (smooth-walled under the bright field microscope, oblate to globose, 2.5–3.5μm diam), whereas those of M. umbrina are lenticular and measure 2.8–4.0×2.1–2.6μm. Moreover, the arthroconidia of M. gymnoascoides are larger than those of M. umbrina (6.0–10.0×1.5–2.0μm and 2.6–7.0×1.4μm, respectively). Malbranchea ostraviensis also produces a pinkish to red diffusible pigment on MEA, PDA and SDA, a feature not observed in M. gymnoascoides nor in M. umbrina. Both Malbranchea gymnoascoides as well as of M. umbrina can grow slowly at 35°C, whereas the maximum temperature of growth for M. ostraviensis is of 32°C.

Type: USA: Texas: from human bronchial washing specimen, 2005, N. Wiederhold (CBS H-24456 – holotype; CBS 146930=FMR 17681=UTHSCSA DI18-87 – ex-type cultures; LSU/ITS sequences GenBank LR701758/LR701757).

Description: Vegetative hyphae septate, hyaline, smooth- and thin-walled, mostly straight, rarely branched, 1.5–2.5μm wide. Asexual morph consisting in undifferentiated fertile hyphae which form randomly intercalary and terminally arthroconidia. Conidia enteroarthric, unicellular, hyaline, smooth- and thin-walled, mostly barrel-shaped, sometimes cylindrical or irregularly-shaped, 6.0–10.0×1.5–2.0μm, detached by rhexolysis. Ascomata gymnothecial, solitary or in clusters, hyaline at first, becoming orange brown with the age, globose or nearly so, 130–250μm diam excluding the appendages, which cover entirely the surface. Peridial hyphae septate, orange brown, branching and anastomosing to form a reticulate network, asperulate, very thick-walled, 3.5–5.5μm wide, fragmenting by the septa when ageing, with lateral appendages. Appendages 0–1-septate, orange brown, asperulate, thick-walled, progressively tapering towards the apex, apex sinuous, 250–400μm long, connected by basal knuckle joints. Asci 8-spored, globose or nearly so, 4–7μm diam, soon deliquescent. Ascospores unicellular, hyaline at first, yellowish in mass when mature, smooth-walled under bright field microscope, globose, 2.5–3.5μm diam.

Culture characteristics: Colonies on PYE reaching 46–47mm diam after 2 wk. at 25°C, slightly elevated, velvety to floccose, margins regular, pale orange (5A3) at centre and white (5A1) at edge, sporulation sparse; reverse orange (5A6). Colonies on PDA reaching 36–37mm diam after 2 wk. at 25°C, slightly elevated, velvety, margins regular, light orange (5A5) at centre and orange white (5A2) at edge, sporulation sparse; reverse deep orange (6A8). Colonies on PDA reaching 31–32mm diam after 2 wk. at 30°C, slightly elevated, velvety, margins regular, slightly furrowed, orange (5A6), sporulation sparse; reverse brownish orange (6C8). Colonies on OA reaching 21–22mm diam after 2 wk. at 25°C, slightly elevated, velvety, margins regular, orange white (5A2) at centre and white (5A1) at edge, sporulation sparse. Exudate and diffusible pigment absent in all culture media tested. Minimum, optimal and maximum temperature of growth on PDA: 10°C, 25°C, and 35°C, respectively. Non-haemolytic. Casein hydrolysed without pH change. Not inhibited by cycloheximide. Urease and esterase tests positive. Growth occurs at NaCl 10% w/w, but not at 20% w/w.

Despite the strain FMR 17695 being phylogenetically close to Malbranchea longispora, it differs phylogenetically and morphologically from it. Consequently, we describe the new species Malbranchea multiseptata.

Malbranchea multiseptata Rodr.-Andr., Cano & Stchigel, sp. nov.

(Fig. ​(Fig.77)

Open in a separate window

Fig. 7

Malbranchea multiseptata CBS 146931 ^T. a Colonies on PYE, PDA and OA after 14 d at 25°C, from left to right (top row, surface; bottom row, reverse). b Detail of the colony on PDA. c–d Highly septate fertile hyphae and arthroconidia. Scale bar=10μm

MycoBank MB 835213

Etymology: From Latin multi-, many, and –septatae, septa, because the vegetative hyphae are multiseptate.

Diagnosis: Malbranchea multiseptata is phylogenetically linked to M. longispora. Nevertheless, M. multiseptata does not form chlamydospores nor a sexual morph as in M. longispora (Crous et al. 2013). Also, M. multiseptata produces shorter conidia (3.0–9.0×1.5–2.0μm) than those of M. longispora (4.0–24.0×1.0–5.5μm).

Type: USA: Texas: from human bronchial washing specimen, 2014, N. Wiederhold (CBS H-24457 – holotype; CBS 146931=FMR 17695=UTHSCSA DI18-101 – ex-type cultures; LSU/ITS sequences GenBank LR701760/LR701759).

Description: Vegetative hyphae hyaline, smooth- and thin-walled straight to sinuous, sparsely branched, 1.0–2.0μm wide, becoming highly septate with the age, septa thickened. Fertile hyphae arising as lateral branches (sometimes opposite each other) from the vegetative hyphae, unbranched, straight or slightly sinuous, 1.5–2.0μm wide, forming randomly intercalary and terminally arthroconidia. Conidia enteroarthric, unicellular, hyaline, smooth- and thin-walled, separated by evanescent connective cells, cylindrical, 3.0–9.0×1.5–2.0μm, rounded at the end when terminal, rhexolytic secession. Chlamydospores, racquet hyphae, setae, and sexual morph not observed.

Culture characteristics: Colonies on PYE reaching 35–36mm diam after 2 wk. at 25°C, elevated, velvety to floccose, margins regular, white (5A1), sporulation sparse; reverse greyish yellow (4B4). Colonies on PDA reaching 34–35mm diam after 2 wk. at 25°C, slightly elevated, velvety to floccose, margins regular, white (5A1), sporulation absent; reverse yellowish white (3A2). Colonies on PDA reaching 27–28mm diam after 2 wk. at 30°C, slightly elevated, velvety to floccose, margins regular, white (5A1), sporulation absent; reverse pale yellow (3A3). Colonies on OA researching 37–38mm diam after 2 wk. at 25°C, flattened, barely perceptible growth, not distinguishable colour, sporulation sparse. Exudate and diffusible pigment absent in all culture media tested. Minimum, optimal and maximum temperature of growth on PDA: 10°C, 25°C, and 35°C, respectively. Haemolytic. Casein hydrolyzed without pH change. Not inhibited by cycloheximide. Urease positive. Growth occurs at NaCl 3% w/w, but not at 10%w/w. Neither grow on TOTM.

Because the strain FMR 17680 was placed phylogenetically close to Malbranchea filamentosa but in a separate terminal branch, and because both differ morphologically and genotypically, the new species Malbranchea stricta is also described.

Malbranchea stricta Rodr.-Andr., Stchigel & Cano, sp. nov.

(Fig. ​Fig.88)

Open in a separate window

Fig. 8

Malbranchea stricta CBS 146932 ^T. a Colonies on PYE, PDA and OA after 14 d at 25°C, from left to right (top row, surface; bottom row, reverse). b Detail of the colony on OA. c–e Alternate arthroconidia on primary hyphae and lateral branches. Scale bar=10μm

MycoBank MB 835219

Etymology: Latin stricta, strict, due to the production of the typical reproductive structures of the genus.

Diagnosis: Malbranchea stricta is phylogenetically close to M. filamentosa. Also, both species lack a sexual morph (Sigler et al. 2002). However, M. filamentosa produces more regularly shaped conidia than M. stricta, and forms thick-walled brown setae, structures absent in M. stricta.

Type: USA: Florida: human nail, 2003, N. Wiederhold (CBS H-24458 – holotype; CBS 146932=FMR 17680=UTHSCSA DI18-86 – ex-type cultures; LSU/ITS sequences GenBank LR701639/LR701638).

Description: Vegetative hyphae hyaline, smooth- and thin-walled, straight to sinuous, sparsely branched, 1.5–2.0μm wide. Fertile hyphae well-developed, arising as lateral branches from the vegetative hyphae, mostly unbranched, right or slightly sinuous, contorted or arcuate at the end, up to 25μm long, 1.5–2.0μm wide, or developing at the extremes of the vegetative hyphae, in both cases forming arthroconidia randomly intercalary and terminally. Arthroconidia enteroarthric, hyaline, becoming yellowish with the age, barrel-shaped, “T”-shaped, “Y”-shaped, finger-shaped or irregularly-shaped, 2.0–6.0×1.0–2.0μm, with rhexolytic secession. Chlamydospores, racquet hyphae, and sexual morph not observed.

Culture characteristics: Colonies on PYE reaching 32–33mm diam after 2 wk. at 25°C, flattened, velvety, regular margins, furrowed, white (4A1), sporulation sparse; reverse pale orange (5A3). Colonies on PDA reaching 20–21mm diam after 2 wk. at 25°C, slightly elevated, velvety to floccose, regular margins, white (3A1), sporulation abundant; reverse pale yellow (4A3). Colonies on PDA reaching 20–21mm diam after 2 wk. at 30°C, slightly elevated, velvety to floccose, margins regular, white (3A1), sporulation abundant; reverse yellowish brown (5E8) at centre and greyish yellow (4B5) at the margins. Colonies on OA researching 16–17mm diam after 2 wk. at 25°C, flattened, granulose, white (3A1), margins regular, sporulation sparse. Exudate and diffusible pigment absent. Minimum, optimum and maximum temperature of growth on PDA: 10°C, 30°C, and 37°C, respectively. Colonies haemolytic (on BA), and casein hydrolyzed without pH changes at 25°C (on BCP-MS-G). Not inhibited by cycloheximide. Urease and esterase tests positive. Growth occurs at NaCl 10% w/w, but not at 20% w/w.

Pseudoarthropsis

Since the strain FMR 17692 was placed in the same terminal clade as Arthropsis cirrhata, while the type species of the genus (Arthropsis truncata) is phylogenetically distant (in Sordariales; Giraldo et al. 2013), we erect the new genus Pseudoarthropsis for A. cirrhata, and the new species Pseudoarthropsis crassispora.

Pseudoarthropsis Stchigel, Rodr.-Andr. & Cano, gen. nov.

MycoBank MB 834925

Etymology: From Greek ψευδής-, resembling, because the morphological semblance to Arthropsis.

Diagnosis: Mycelium composed by hyaline to orange, septate hyphae. Conidiophores consisting of fertile lateral branches and a portion of the main subtending hypha, with all these structures disintegrating into yellowish orange, thin-walled, cylindrical to cuboid enteroarthric conidia, or into hyaline, thick-walled, ellipsoidal, globose to barrel-shaped holoarthric conidia.

Type species: Pseudoarthropsis cirrhata (Oorschot & de Hoog) Stchigel, Rodr.-Andr. & Cano 2021.

Pseudoarthropsis cirrhata (Oorschot & de Hoog) Stchigel, Rodr.-Andr. & Cano, comb. nov. MycoBank MB 834928

Basionym: Arthropsis cirrhata Oorschot & de Hoog, Mycotaxon 20: 130 (1984).

Description: Vegetative hyphae septate, pale yellowish orange, smooth- and thin-walled, dichotomously branched, 2–3μm wide. Fertile hyphae well differentiated, arising at right angles as recurved lateral branches of the vegetative hyphae, forming septa basipetally to produce chains of enteroarthric conidia. Arthroconidia yellowish orange, smooth- and thin-walled, cylindrical to cuboid, often broader than long, 2.5–4.0×2–3μm, truncated at both ends, separated by trapezoid connectives, secession rhexolytic. Colonies on PYE reaching 4–5mm diam after 10 d at 25°C, powdery, fealty, slightly raised, orange (5A7), pale orange (5A5) at centre; reverse brownish orange (7C8), diffusible pigment brown.

Type: The Netherlands: from a wall near Schiphol, 1984, C.A.N. van Oorschot (CBS 628.83).

Pseudoarthropsis crassispora Rodr.-Andr., Stchigel & Cano, sp. nov.

(Fig. ​Fig.99)

Open in a separate window

Fig. 9

Pseudoarthropsis crassispora CBS 146928 ^T. a Colonies on PYE, PDA and OA after 14 d at 25°C, from left to right (top row, surface; bottom row, reverse). b Detail of the colony on OA. c–e Bi- to trichotomously-branched fertile hyphae. f A large chain of holoarthric conidia. Scale bar=10μm

MycoBank MB 834930

Etymology: From Latin crassus-, thick, and -sporarum, spore, because of the thick wall of the conidia.

Diagnosis: Pseudoarthropsis crassispora is phylogenetically close to P. cirrhata. Nevertheless, the former produces holoarthric conidia, while they are enteroarthric in the latter. Also, the conidia of P. crassispora are ellipsoidal, globose or broadly barrel-shaped, while these are cylindrical to cuboid (often wider than they are long) in P. cirrhata (van Oorschot and de Hoog 1984). Moreover, the conidia are bigger in P. crassispora than in P. cirrhata (4.5–5.5×2.5–3.5μm vs. 2.5–4.0×2.0–3.0μm). Also, P. crassispora grows faster than P. cirrhata (on PYE at 25°C), and the maximum temperature of growth is at 37°C and 30°C, respectively.

Type: USA: Minnesota: from a human bronchial washing specimen, 2012, N. Wiederhold (CBS H-24454 – holotype; CBS 146928=FMR 17692=UTHSCSA DI18-98 – ex-type cultures; LSU/ITS sequences GenBank LR701763/LR701764).

Description: Vegetative hyphae septate, hyaline, smooth- and thin-walled, mostly straight, occasionally branched, 1.5–2.0μm wide. Fertile hyphae well-differentiated, arising as lateral branches of the vegetative hyphae, hyaline, septate, smooth- and thin-walled, erect, simple or branched up to 3 times at the apex, stipe 10–20×1.5–2.0μm, branches 10–70×1.5–2.0μm, forming septa basipetally to produce chains of arthroconidia. Conidia holoarthric, unicellular, hyaline, smooth- and thick-walled, ellipsoidal, globose or barrel-shaped, transiently presents as bi-cellular conidia, 2.5–3.5×4.5–5.5μm, in chains of up to 20, separate from the fertile hyphae by schizolysis, rarely by rhexolysis. Chlamydospores, racquet hyphae, setae, and sexual morph not observed.

Culture characteristics: Colonies on PYE reaching 13–14mm diam after 2 wk. at 25°C, slightly elevated, velvety, margins regular, furrowed, yellowish white (3A2) and yellowish grey (4B2) at centre, sporulation abundant; reverse pale yellow (4A3. Colonies on PDA reaching 14–15mm diam after 2 wk. at 25°C, flattened, velvety, margins regular, greenish white (30A2) and pastel green (30A4) at centre, sporulation abundant; reverse pastel yellow (3A4). Colonies on PDA reaching 15–16mm diam after 2 wk. at 30°C, slightly elevated, velvety, margins regular, furrowed, yellowish white (3A2), sporulation sparse; reverse yellow (3A6), with a scarce production of yellowish diffusible pigment. Colonies on OA researching 10–11mm diam after 2 wk. at 25°C, flattened, velvety to floccose, margins irregular, greenish white (30A2) and pale green (28A3) at centre, sporulation abundant. Exudate and diffusible pigment absent, except on PDA. Minimum, optimal and maximum temperature of growth on PDA: 10°C, 30°C, and 37°C, respectively. Non-haemolytic. Casein hydrolyzed without pH change. Not inhibited by cycloheximide. Urease and esterase tests positive. The fungus grows up to NaCl 10% w/w, but not at 20% w/w.

Pseudomalbranchea

Despite the strain FMR 17684 being placed phylogenetically in Onygenaceae, it is paraphyletic described as the type species of the new genus Pseudomalbranchea.

Pseudomalbranchea Rodr.-Andr., Cano & Stchigel, gen. nov.

MycoBank MB 835220

Etymology: Recalling the morphological similarity with Malbranchea.

Diagnosis: Arthroconidia one-celled, intercalary disposed along unbranched vegetative hyphae, mostly enteroarthric, occasionally holoarthric, cylindrical but becoming globose with the age.

Type species: Pseudomalbranchea gemmata Rodr.-Andr., Cano & Stchigel 2021

Description: Mycelium sparse, composed of hyaline, smooth- and thin-walled septate hyphae. Asexual morph consisting of mostly enteroarthric, occasionally holoarthric, conidia, intercalary disposed along unbranched vegetative hyphae, solitary or in short chains, with rhexolytic or rarely schizolytic secession. Arthroconidia one-celled, hyaline, smooth- and thick-walled, cylindrical but becoming globose with the age. Chlamydospores, racquet hyphae and sexual morph not observed.

Pseudomalbranchea gemmata Rodr.-Andr., Cano & Stchigel, sp. nov.

(Fig. ​(Fig.1010)

Open in a separate window

Fig. 10

Pseudomalbranchea gemmata CBS 146933 ^T. a Colonies on PYE, PDA and OA after 14 d at 25°C, from left to right (top row, surface; bottom row, reverse). b Detail of the colony on OA. c–d Large, intercalary, irregularly-shaped arthroconidia disposed singly or in chains along the fertile hyphae. Scale bar=10μm

MycoBank MB 835221

Etymology: From the Latin gemmatum, jewelled, because the swollen conidia disposed in chains.

Diagnosis: Pseudomalbranchea gemmata is phylogenetically close to Uncinocarpus reesii and Amauroascus volatilis-patellis. However, it does not produce a sexual morph and it differs from U. reessi and A. volatilis-patellis by the production of longer arthroconidia (4.0–11.0×2.0–3.5μm in P. gemmata vs. 3.5–6.0×2.5–3μm in U. reessi, and 4.0–5.4×2.0–3.0 in A. volatilis-patellis; Orr and Kuehn 1972, Sigler and Carmichael 1976, Currah 1985). As well as A. volatilis-patellis, P. gemmata lacks appendages, which are present and similar to the asexual morph in U. reessi (Currah 1985).

Type: USA: Florida: from human bronchial washing specimen, 2014, N. Wiederhold (CBS H-24459 – holotype, CBS 146933=FMR 17684=UTHSCSA DI18-90 – ex-type cultures; LSU/ITS sequences GenBank LR701762/LR701761).

Description: Mycelium sparse, composed of hyaline, smooth- and thin-walled, sparsely septate hyphae, 1.0–2.0μm wide. Conidia enteroarthric (occasionally holoarthric), intercalary disposed along unbranched vegetative hyphae, one-celled, solitary or in short chains of up to 7, one-celled, hyaline, smooth- and thick-walled, cylindrical but becoming globose with the age, 4.0–11.0×2.0–3.5μm, liberated from the fertile hyphae by rhexolysis (rarely by schizolysis). Chlamydospores, racquet hyphae and sexual morph not observed.

Culture characteristics: Colonies on PYE reaching 22–23mm diam after 2 wk. at 25°C, slightly elevated, velvety, margins regular, pale yellow (3A3), sporulation sparse; reverse brown (6E6). Colonies on PDA reaching 24–25mm diam after 2 wk. at 25°C, slightly elevated, velvety, margins regular, pale yellow (3A3), sporulation sparse; reverse light yellow (4A5). Colonies on PDA reaching 25–26mm diam after 2 wk. at 30°C, flattened, radially folded, velvety, margins regular, pale yellow (3A3), sporulation sparse; reverse light yellow (4A5). Colonies on OA reaching 28–29mm diam after 2 wk. at 25°C, flattened, velvety to granulose, irregular margins, white (6A1), sporulation sparse. Exudate and diffusible pigment lacking. Minimum, optimum and maximum temperature of growth on PDA: 10°C, 30°C, and 37°C, respectively. Colonies haemolytic, casein not hydrolyzed. The fungus was not inhibited by cycloheximide. Urease and esterase tests positive. Growth occurs at NaCl 3% w/w, but not higher concentration.

Spiromastigoides

Because strains FMR 17686 and FMR 17696 were placed together in a terminal branch close to the ex-type strain of M. gypsea in the Spiromastigaceae clade (Fig. ​(Fig.2),2), M. gypsea is combined into Spiromastigoides and these two strains are described as the new species S. geomycoides.

Spiromastigoides geomycoides Stchigel, Rodr.-Andr. & Cano, sp. nov.

(Fig. ​Fig.1111)

Open in a separate window

Fig. 11

Spiromastigoides geomycoides CBS 146934 ^T. a Colonies on PYE, PDA and OA after 14 d at 25°C, from left to right (top row, surface; bottom row, reverse). b Detail of the colony on OA. c Fertile lateral branches mimicking Geomyces spp. conidiophores. d–e Fertile hyphae with intercalary, barrel-shaped arthroconidia. f Morphological diversity of arthroconidia. Scale bar=10μm

MycoBank MB 835222

Etymology: From the production of conidiophores morphologically similar to those of the genus Geomyces.

Diagnosis: Spiromastigoides geomycoides is phylogenetically close to S. gypsea. However, it produces smaller conidia (1.5–2.5×1.0–2.0μm) than S. gypsea [(2.5)3–6(9)×2–2.5μm; Sigler and Carmichael 1976]. Also, S. geomycoides grows faster than S. gypsea on PYE at 35°C.

Type:USA: Illinois: from a human foot skin, 2014, N. Wiederhold (CBS H-24460 – holotype, CBS 146934=FMR 17696=UTHSCSA DI18-102 – ex-type cultures; LSU/ITS sequences GenBank LR701768/LR701768).

Description: Mycelium abundant, composed of hyaline, smooth- and thin-walled, septate, branched, 1.0–2.0μm wide hyphae, septa thickened with age. Fertile hyphae arising as lateral branches, straight or slightly curved, unbranched or, rarely, with a branching pattern similar to that of the conidiophores of Geomyces, septate, hyaline, smooth- and thin-walled, producing intercalary and terminally arthroconidia separated by 1–2 empty intermediary cells. Conidia enteroarthic, unicellular, hyaline, mostly barrel-shaped, less frequently “T”-shaped or cylindrical, 1.5–2.5×1.0–2.0μm, rhexolytic dehiscence. Chlamydospores, racquet hyphae and sexual morph not observed.

Culture characteristics: Colonies on PYE reaching 24–25mm diam after 2 wk. at 25°C, flattened, velvety, furrowed, regular margins, white (4A1), abundant sporulation; reverse, pale orange (5A3). Colonies on PDA reaching 26–27mm diam after 2 wk. at 25°C, flattened, velvety, regular margins, white (4A1), abundant sporulation; reverse, yellowish white (4A2). Colonies on PDA reaching more than 90mm diam after 2 wk. at 30°C, flattened, velvety, regular margins, yellowish white (4A2), sporulation absent; reverse, pale yellow (4A3). Colonies on OA researching 20–21mm diam after 2 wk. at 25°C, flattened, granulose, regular margins, white (4A1), abundant sporulation. Exudate and diffusible pigment absent in all culture media tested. Minimum, optimum and maximum temperature of growth on PDA: 5°C, 30°C, and 37°C, respectively. Colonies non-haemolytic. Casein not hydrolyzed. Resistant to cycloheximide. Urease negative and esterase positive. Growth occurs at NaCl 10% w/w, but not at 20% w/w.

Other specimen examined: USA: Minnesota: from blood, 2009, N. Wiederhold (FMR 17686).

Spiromastigoides gypsea (Sigler & Carmichael) Stchigel, Rodr.-Andr. & Cano, comb. nov.

MycoBank MB 835228

Basionym: Malbranchea gypsea Sigler & Carmichael, Mycotaxon 4: 455 (1976).

Description (adapted from the original description): Arthroconidia produced intercalary or terminally along straight primary hyphae, or on short or long lateral branches, separated each one by one or more alternate empty cells, or, rarely, formed immediately adjacent to each other. Arthroconidia unicellular, hyaline, smooth- and thin-walled, cylindrical or slightly barrel-shaped, (2.5) 3–6 (9)×2–2.5μm, slightly broader than the interconnecting cells. No sexual morph obtained by matting. Colonies on PYE reaching 17–39mm after three wk. at room temperature, chalky white to creamy white, downy to velvety, slightly raised, surface folded to convoluted, umbonated at centre, reverse buff. Optimum temperature of growth 25–30°C. Maximum temperature of growth 37°C (but strain depending).

KEYS

Key to Arachnomyces species

Adapted from Sun et al. (2019).

1 Homothallic; asexual morph present or not.................................................................................................................................. 2

Heterothallic; asexual morph present ................................................................................................................................................. 6

2(1) Peridial setae coiled or circinate; asexual morph absent....................................................................................................... 3

Peridial setae straight, tapering towards the apex; asexual morph arthroconidia ……………................................. gracilis

3(2) Peridial setae slightly nodose; ascospores mostly <3.5μm diam ………………….…...................................................... 4

Peridial setae smooth-walled; ascospores mostly >3.5μm diam................................................................................................. 5

4(3) Ascospores smooth-walled............................................................................................................................................. minimus

Ascospores echinulate.......................................................................................................................................................... peruvianus

5(3) Ascomata 100–300μm diam............................................................................................................................................. nitidus

Ascomata 500–700μm diam.............................................................................................................................................. sulphureus

6(1) Arthroconidia alternate................................................................................................................................................................... 7

Arthroconidia in persistent chains..................................................................................................................................................... 12

7(6) Arthroconidia cylindrical or barrel-shaped; sclerotia present............................................................................................. 8

Arthroconidia distinct; sclerotia absent.............................................................................................................................................. 9

8(7) Colonies becoming greyish brown, not growing at 35°C..................................................................................... glareosus

Colonies white to pale brown, growing at 35°C........................................................................................................... scleroticus

9(7) Arthroconidia subglobose to pyriform..................................................................................................................................... 10

Arthroconidia cylindrical to finger-like-shaped.............................................................................................................................. 11

10(9) Arthroconidia smooth-walled to finely asperulate; setae (produced on the vegetative mycelium) smooth-walled to slightly nodose............................................................................................................................... kanei

Mature arthroconidia coarsely verrucose; setae (produced on the vegetative mycelium) strongly nodose............................................................................................................................................................. pilosus

11(9) Fertile hyphae successively branching to form dense clusters, arcuate, sinuous, contorted or tightly curled................................................................................................................ bostrychodes

Fertile hyphae branching but not in clusters; branches only apically coiled.......................................................................... graciliformis

12(6) Setae (produced on the vegetative mycelium) strongly nodose, circinate or loosely coiled at the apex................................................................................................................................................ nodosetosus

Setae (produced on the vegetative mycelium) strongly nodose, tip straight................................................................... jinanicus

Key to Malbranchea species

Adapted from Sigler and Carmichael (1976), Solé et al. (2002), and Hubka et al. (2013).

1 Homothallic species.............................................................................................................................................................................. 2

Heterothallic species............................................................................................................................................................................... 13

2(1) Peridial appendages longer than 150μm long.......................................................................................................................... 3

Peridial appendages shorter or absent................................................................................................................................................. 8

3(2) Appendages 350–600μm in length; diffusible pigment pinkish to reddish; not growing at 35°C …............ ostraviensis

Above features not combined................................................................................................................................................................ 4

4(3) Ascospores smooth-walled under bright field microscope....................................................................... gymnoascoides

Ascospores reticulate................................................................................................................................................................................ 5

5(4) Peridial cells short, 4–12μm in length; peridial projections with truncate ends.......................................... compacta

Peridial cells longer; peridial projections with mostly acute ends............................................................................................... 6

6(5) Ascospores usually exceeding 4μm diam......................................................................................................... californiensis

Ascospores ≤4μm diam........................................................................................................................................................................... 7

7(6) Species growing at 37°C................................................................................................................................................ conjugata

No growth at 37°C..................................................................................................................................................................... umbrina

8(2) Asexual morph not produced......................................................................................... guangxiensis / pseudauxarthron

Malbranchea-like asexual morph present........................................................................................................................................... 9

9(8) Ascomata with spine-like peridial projections, 27–40μm in length.................................................................... zuffiana

Ascomata without peridial projections............................................................................................................................................. 10

10(9) Colonies on PDA brown................................................................................................................................................. kuehnii

Colonies on PDA otherwise................................................................................................................................................................. 11

11(10) Peridial hyphae smooth-walled........................................................................................................................... concentrica

Peridial hyphae strongly ornamented; chlamydospores present ….......................................................................................... 12

12(11) Arthroconidia 2–10×2.5–3.5μm; growing above 30°C …................................................................. chlamydospora

Arthroconidia 4–24×1.0–5.5μm; not growing above 30°C..................................................................................... longispora

13(1) Fertile hyphae arcuate or curved............................................................................................................................................. 14

Fertile hyphae straight to sinuous, branched or not..................................................................................................................... 21

14(13) Fertile hyphae coiled................................................................................................................................................................. 15

Fertile hyphae curved or arcuate........................................................................................................................................................ 16

15(14) Thermophilic; conidia 2.5–4.5μm wide......................................................................................................... cinnamomea

Not thermophilic; conidia narrower.................................................................................................................................... pulchella

16(14) Colonies orange.......................................................................................................................................................................... 17

Colonies different.................................................................................................................................................................................... 18

17(16) Aleuroconidia laterally or terminally dispersed.................................................................................... chrysosporoidea

Aleuroconidia absent............................................................................................................................................................. aurantiaca

18(16) Colonies golden yellow, exudate brown, diffusible pigment yellow........................................................ graminicola

Above features not combined..................................................... .........................................................................................................19

19(18) Sexual morph produced by in vitro mating of compatible strains................................................................ albolutea

Sexual morph not formed..................................................................................................................................................................... 20

20(19) Thick-walled brown setae produced on OA from the vegetative mycelium......................................... filamentosa

Setae not produced....................................................................................................................................................................... arcuata

21(13) Fertile hyphae unbranched or scarcely branched............................................................................................................. 22

Fertile hyphae branched........................................................................................................................................................................ 24

22(21) Arthroconidia cylindrical; becoming many septate with the age............................................................ multiseptata

Arthroconidia barrel-shaped, “T”-shaped, “Y”-shaped, finger-shaped or more irregular, mostly unicellular.............. 23

23(22) Arthroconidia barrel-shaped, 4–8×2–3.5μm; racquet hyphae present...................................................... chinensis

Arthroconidia barrel-shaped, “T”-shaped, “Y”-shaped, finger-shaped or more irregular, 2–6×1–2μm; racquet hyphae absent....................................................................................................................................................................... stricta

24(21) Fertile hyphae branching acutely, displaying a tree-like appearance.......................................................... dendritica

Fertile hyphae branching pattern otherwise.................................................................................................................................... 25

25(24) Fertile hyphae repeatedly branched, in dense tufts...................................................................................... flocciformis

Fertile hyphae more restrictedly branched.......................................................................................................................................26

26(25) Colonies buff or tan............................................................................................................................................................. fulva

Colonies lemon yellow...................................................................................................................................................................... flava

Key to Spiromastigoides species

Adapted from Hirooka et al. (2016).

1 Homothallic............................................................................................................................................................................................. 2

Heterothallic................................................................................................................................................................................................ 6

2(1) Ascospores globose to subglobose, reticulate.................................................................................................. sphaerospora

Ascospores oblate, equatorial thickening present or not............................................................................................................... 3

3(2) Ascospores with equatorial thickening...................................................................................................................................... 4

Ascospores without such equatorial thickening............................................................................................................................... 5

4(3) Ascomata appendages straight or slightly undulate; ascospores yellow, smooth-walled under LM, pitted under SEM........................................................................................................................................................................ alatospora

Ascomata appendages slightly undulate or wavy; ascospores pale yellowish brown, minutely punctate under SEM........................................................................................................................................... saturnispora

5(3) Ascospores punctate, sometimes with a few fine grooves in the polar region, 2.5–2.9×2.0–2.5μm...................................................................................................................................................................... warcupii

Ascospores lens-shaped, regularly pitted, 3.0×2.0μm................................................................................................ sugiyamae

6(1) Asexual morph chrysosporium-like; sterile ascomata present............................................................................. asexualis

Asexual morph distinct............................................................................................................................................................................ 7

7(6) Asexual morph malbranchea-like............. .................................................................................................................................. 8

Conidiophores well-developed......... .................................................................................................................................................. 11

8(7) Fertile hyphae straight, branched...................................................................................................................................... gypsea

Fertile hyphae curved............................................................................................................................................................................... 9

9(8) Fertile hyphae successively branched to form sporodochia-like structures........................................................... albida

Fertile hyphae unbranched or scarcely branched.......................................................................................................................... 10

10(9) Fertile hyphae unbranched or sparsely branched, curved, > to 28μm long; chlamydospores present...................... curvata

Fertile hyphae unbranched, slightly curved, > to 15μm long; chlamydospores absent......................................... minimus

11(7) Conidiophores unbranched or scarcely branched......................................................................................... geomycoides

Conidiophores branched several times............................................................................................................................................. 12

12(11) Conidiophores > to 300μm in length, verticillate............................................................................................ kosraensis

Conidiophores 100–150μm in length, with pyramidal or bush-like branching................................................................... 13

13(12) Conidiophores > to 150μm long, with pyramidal branching.................................................................... pyramidalis

Conidiophores > to 100μm long, with bush-like branching............................................................................................... frutex

We are indebted to Phil Hoddy for editing and proofreading the final text. We also thank Patricia Navarro Rodríguez for support in the antifungal susceptibility testing.