International Journal of Systematic and Evolutionary Microbiology (2014), 64, 3046–3052 DOI 10.1099/ijs.0.062380-0

Basidioascus persicus sp. nov., a yeast-like species of the order Geminibasidiales isolated from soil

Shaghayegh Nasr,1,2 Mohammad Reza Soudi,1 Seyyedeh Maryam Zamanzadeh Nasrabadi,1 Mahdi Moshtaghi Nikou,2 Ali Hatef Salmanian3 and Hai D. T. Nguyen4

Correspondence 1National Laboratory of Industrial Microbiology, Department of Biology, Faculty of Sciences, Mohammad Reza Soudi Alzahra University, Tehran, Iran [email protected] 2Microorganism Bank, Iranian Biological Resource Center (IBRC), ACECR, Tehran, Iran 3Department of Agricultural Biotechnology, National Institute of Genetic Engineering and Biotechnology, Tehran, Iran 4Department of Biology, University of Ottawa, Ottawa, Ontario, Canada K1N 6N5

A novel species of basidiomycetes was isolated from kitchen garden soil in Shahryar city, Tehran province, Iran. Molecular and conventional methods were employed to identify and classify this single isolate. Morphologically, the isolate was considered yeast-like with hyaline and oval cells reproducing by monopolar budding, forming ballistoconidia, hyphae, arthroconidia and didymospores. Basidia and basidiospores resembling those produced by Basidioascus species were observed. Sequencing and Bayesian phylogenetic analysis of rRNA genes and the internal transcribed spacer region revealed its sister relationship to described species of the genus Basidioascus. Assimilation and fermentation tests, cell-wall carbohydrate analysis and enzyme activity tests were performed to provide insight into the metabolism of the isolate. Based on morphology, physiology and phylogeny of rRNA gene sequences, the isolate was shown to represent a novel species of the genus Basidioascus, described as Basidioascus persicus sp. nov. (holotype IBRC P1010180T5ex-type IBRC M30078T5isotype CBS 12808T). The MycoBank number of the novel species is MB 804703. An emended description of the genus Basidioascus is also provided.

Introduction Debaryomyces, Hansenula, Candida, Brettanomyces, Rhodo- Fungi, including yeasts, occur in a wide range of soil types torula, Cryptococcus, Trichosporon and Torulopsis (Bresolin (Anderson & Cairney, 2004; Jones & Pang, 2012; Carvalho et al., 2010; Kurtzman et al., 2011; Carvalho et al., 2013). et al., 2013) and abiotic factors such as temperature, There are, at the time of writing, around 1500 yeast species humidity and chemical composition influence their described (Kurtzman, 2011). The doubling time of species diversity (Botha 2011; Yurkov et al., 2012). Soil yeasts are discovery is about 10 years (Boekhout et al., 2012) so our important; for example, they contribute to essential knowledge of yeast biodiversity is rapidly advancing. However, ecological processes such as the mineralization of organic even at the current rate of discovery, our knowledge of the fungal biodiversity on Earth, which includes the yeasts, is material and dissipation of carbon and energy through deficient with an estimated 5–10 % of fungi known to science the soil ecosystem (Botha, 2011). Many yeast genera currently (Blackwell 2011). have been isolated from soils, such as species of Kluyvero- myces, Lipomyces, Schwanniomyces, Schizoblastosporium, A fungal order called Geminibasidiales was taxonomically defined recently (Nguyen et al., 2013). The Geminibasidiales Abbreviation: ITS, internal transcribed spacer. comprises two genera (Geminibasidium and Basidioascus). The GenBank/EMBL/DDBJ accession numbers for the 28S rRNA and together they form a sister group to the order gene (D1/D2 domains), ITS and 18S rRNA gene region sequences of Wallemiales based on phylogeny of rRNA gene sequences Basidioascus persicus sp. nov.5ex-type IBRC M30078T5isotype CBS (Nguyen et al., 2013). These two orders are grouped under 12808T are KC751416, KC751417 and KC751418, respectively. the class Wallemiomycetes and they represent a basal lineage One supplementary table and two supplementary figures are available to the major fungal subdivision Agaricomycotina. Species with the online version of this paper. of the genera Basidioascus and Geminibasidium are hyphal,

3046 062380 G 2014 IUMS Printed in Great Britain Basidioascus persicus sp. nov. heat-resistant and xerotolerant. and they have been isolated assimilation ability of the isolate was determined by employing tubes from soil (Matsushima, 2001; Nguyen et al., 2013a). Only containing 5 ml liquid yeast nitrogen base medium (Sigma-Aldrich) two species of the genus Basidioascus, Basidioascus undulatus and a particular carbon compound (50 mM) as a sole source of energy. The tubes were incubated at 28 uC for 4 weeks and the results and Basidioascus magus, have been described and the internal from growth tests were read spectrophotometrically at 600 nm. transcribed spacer (ITS) phylogeny with environmental sequences hinted at additional uncultured species in this Cell-wall carbohydrate analysis. To analyse the carbohydrate genus (Nguyen et al., 2013a). composition of the cell wall, 50 mg dried cells cultivated from yeast extract peptone glucose broth [YPD: 1 % yeast extract, 2 % peptone ] In the present study, an unknown fungal isolate was found and 2 % glucose (all w/v) , were hydrolysed with 1 ml of 1.0 M H2SO4 from soil in Shahryar city, Tehran province, Iran. Based on at 100 uC for 2 h, then cooled and the pH adjusted to 5.5 with a morphology, physiology and phylogeny of rRNA gene saturated solution of Ba(OH)2. The precipitate was removed by sequences, this isolate is shown to represent a novel species centrifugation and the supernatant was dried in a rotary evaporator m m of the genus Basidioascus. and resolved in 50 l distilled water. A 1 l aliquot of this solution was applied to a cellulose TLC plate and developed with a solvent system containing n-butanol/water/piridine/toluene (10 : 6 : 6 : 1, by vol.) for 4 h. Sugars were visualized by an aniline phthalate solution Methods and compared with a 1 ml aliquot of 1 % (w/v) standard solutions of high purity sugar mixtures (Komagata & Suzuki, 1987; Kurtzman Isolation. A kitchen garden soil sample from Shahryar city was et al., 2011). transferred to the National Laboratory of Industrial Microbiology at Alzahra University. Dilutions of the sample were directly plated on Coenzyme Q. Intact cells were cultivated in YPD broth on a Rose Bengal agar (Himedia) and incubated at 28 uC for 72 h. Fungal rotary shaker at 150 r.p.m., 28 uC for 72 h and were harvested by colonies were subcultured and purified on yeast peptone glucose centrifugation. To identify coenzyme Q, these cells were washed three agar [YPG; 5 g yeast extract, 10 g peptone, 20 g glucose and 20 g agar times with distilled water and freeze-dried. Purification of coenzyme (all Merck), 1000 ml distilled water] at 28 uC. An unknown isolate T Q was performed by preparative TLC (silica gel 60 F254, glass plates) designated IBRC M30078 was chosen for subsequent studies using benzene for development. The dark band of coenzyme Q described below. observed under UV light was scraped off and extracted with 2 ml acetone (Mikita & Yamada, 1999). Coenzyme Q isoprenologues were Morphological and physiological studies. To assess colony identified using HPLC as described previously (Limtong et al., 2007). morphology and sizes, isolate IBRC M30078T was grown on YPG agar, 2 % malt extract agar (MEA; Merck), dichloran 18 % glycerol Enzymic profiling. To test for degradation of starch, protein agar (DG18; Merck) and malt yeast 40 % sucrose agar (M40Y; (casein), lipids (Tweens 20 and 80), phospholipids (egg yolk), gelatin, Nguyen et al., 2013a). To test for forcibly discharged basidia or DNA, xylan, CM-cellulose and guaiacol, the procedures from Brizzio basidiospores, the isolate was grown on corn meal agar (CMA; et al. (2007) and Li et al. (2008) were employed. Petri dishes Merck). The dish was then inverted over another Petri dish of containing each substrate were incubated at 28 uC. 5 % MEA containing a sterile slide, and incubated at 28 uC. After 21 days of incubation in the inverted position, the MEA dish was checked for structures shot down from the colony grown on CMA. Results and discussion Molecular studies. The ITS region of the rRNA gene was amplified by PCR using the primer pair ITS1 (forward) (59-TCCGTAGGTGA- Morphological and physiological studies ACCTGCGG-39) and ITS4 (reverse) (59-TCCTCCGCTTATTGAT- ATGC-39) (White et al., 1990). The D1/D2 domain of the LSU was An unknown fungal isolate was found in the kitchen amplified by PCR with primers NL1 (forward) (59-GCATATCAAT- garden soil sample from Shahryar city. In an attempt to AAGCGGAGGAAAAG-39) and NL4 (reverse) (59-GGTCCGTGTTT- identify this unknown isolate, morphological, physiological CAAGACG G-39) (Lin et al., 1995; Kurtzman & Robnett, 1998). The and phylogenetic studies were performed initially. SSU rRNA gene was amplified using the combination of primers NS1 (forward) (59-GTAGTCATATGCTTGTCTC-39) and NS8 (reverse) The isolate produced cream-coloured colonies on MEA (59-TCCGCAGGTTCACCTACGGA-39) (White et al., 1990). These and M40Y (Fig. 1a, c), white hyphal colonies on YPG genes were sequenced directly from PCR products using the same (Fig. 1b), and olive-coloured colonies on DG18 (Fig. 1d). primers. Sequences were aligned with MAFFT software (Katoh et al., Arthroconidia (Fig. 1e) and hyaline oval cells that multi- 2005) and phylogenetic analysis was carried out with MrBayes version plied by mono-polar budding (Fig. 1f) were observed. 3.2 (Ronquist & Huelsenbeck, 2003) following the method described Additionally, the isolate produced didymospores on YPG by Nguyen et al. (2013a). and DG18 medium after 48 h of incubation at 28 uC (Fig. Fermentation and assimilation. The standard laboratory methods 1g) and ballistoconidia on PDA after at least 5 days of of Barnett et al. (2000) and Kurtzman et al. (2011) for identifying incubation at the same temperature (Fig.1h–j). Based on yeasts were used for determination of phenotypic and physiological our inverted Petri dish test for forcibly discharged spores, characteristics of isolate IBRC M30078T. Fermentation ability was single colonies, shot from the top dish, were observed determined using Wickerham (1951) basal medium containing 4.5 g 21 21 on the bottom dish, indicating that forcible discharge yeast extract l and 7.5 g peptone l . Four millilitres of the stock occurred. The ballistoconidium is produced on a short solution (50 mg/75 ml) of Bromothymol blue was used as indicator and Durham tubes were used for detecting carbon dioxide pro- sterigma (Fig. 1h) attached to a dark oval basal cell and duction. The sugars used in fermentation tests were at 2 % (w/v), enlarged over the time. Their forcible discharge left a scar on except for raffinose, which was at 4 % (w/v). The tubes were inspected the mother oval cell (Fig. 1k). Germination of ballistoco- at frequent intervals for accumulation of gas for up to 2 weeks. The nidia was observed (Fig. 1m). Sexual reproduction was http://ijs.sgmjournals.org 3047 S. Nasr and others

(a) (h) (q) orange to brown colour (Fig. 1p, q). Sometimes basidia lost their cytoplasm and collapsed (Fig. 1r). Only one mature basidiospore was observed per basidium. A similar matura- (b) (r) tion process was observed in existing species of the genus Basidioascus. The yeast cells produced both pseudohyphae (Fig. 1s) and septate hyphae (Fig. 1t). No clamp connections (i) could be seen. (c) Species of the genus Basidioascus are known to be (s) osmotolerant so physiological tests were performed on (j) the isolate. The isolate grew in 5–10 % NaCl, in 50–60 % (d) glucose and also on M40Y and DG18 agar, similarly to other species of the genus Basidioascus, particularly (k) Basidioascus undulatus (Nguyen et al., 2013a). According (e) to Nguyen et al. (2013), species of the genus Basidioascus are considered mesophilic and their optimum growth (l) temperature is 25–30 uC; however, Basidioascus undulatus is able to grow at 37 uC. Isolate IBRC M30078T had a similar optimal growth temperature but it was not able to grow at 37 or 40 uC. (f) (m) (t) Molecular studies Amplification of the SSU, ITS and LSU loci followed by (n) sequencing and phylogenetic analysis were performed to classify and identify the novel isolate. Amplicons of 1686, 651 and 600 bp were obtained for the SSU, ITS and D1/D2 (g) (o) region of the LSU, respectively. Searches via the BLAST program were performed with these sequences with the top five BLAST hits listed in Table S1 (available in the online (p) Supplementary Material). All BLAST matches belonged to fungi isolated from soil. However, the closest identifiable match was Basidioascus, supporting the morphological and physiological observations described above.

Fig. 1. Morphology of Basidioascus persicus IBRC M30078T. Bayesian phylogenetic analyses were performed to assess Colonies on MEA (a), YPG (b), M40Y (c) and DG18 (d) after the relationship of the novel isolate to fungi related to incubation for 1 week at 28 6C. Arthroconidia (e). Oval cells (f). Basidioascus. The resulting rRNA gene phylogeny (Fig. 2) Didymospores (g). Ballistoconidia forming on oval cells (h–j). showed that the isolate grouped together with strains Oval cell with an ejected ballistoconidium leaving a scar (k). of species of the genus Basidioascus with a posterior Ballistoconidia (l). Germination of ballistoconidia (m). Developing probability of 1.00. The ITS phylogeny (Fig. 3) also basidium with a basidiospore attached on a sterigma (n–r). confirmed this result and showed a sister group relation- Pseudohyphae (s). Septate hyphae with arrows pointing to the ship to Basidioascus undulatus and Basidioascus magus septum (t). Bar,10 mm. Media: YPG (e–g), PDA (h–m), PDA and with a posterior probability of 0.99. Strain MD76-1BY MEA (n–t). (GenBank accession nos. EU626069 and EU626068, other- wise not formally described), previously isolated from soil in China, represented the same species or a very closely related species to our isolate from Iran (Table S1 and observed after 14 days of incubation at 28 uC on PDA and Fig. 3) and was misidentified as representing Cryptococcus MEA (Fig. 1n–r). The probasidium was recognized by at sp. Furthermore, another uncultured strain from soil in least one basal lateral projection (Fig. 1n–p), a character France (GenBank accession no. FN397319) was closely that was previously observed in species of the genera related to the new isolate. The ITS sequences from these Geminibasidium and Basidioascus, and a basidiospore was fungi isolated in China and France form a strongly attached to it by a long tubular sterigma. The basal lateral supported clade with strain IBRC M30078T from Iran, projection is a small protrusion, resembling clamp connec- supporting the existence of a third but undescribed species tions, but not actually attached to the subtending cell, that of Basidioascus. The existence of these sequences suggests is found near the base of the basidium in species of the that this novel species of the genus Basidioascus was genus Basidioascus (Nguyen et al., 2013a). Over time, previously found, but not identified nor described the basidiospore became granular and verrucose with an previously.

3048 International Journal of Systematic and Evolutionary Microbiology 64 Basidioascus persicus sp. nov.

Glomus mosseae UT101 1.00 Saccharomyces cerevisiae ALI308, CECT11761 & D4* Schizosaccharomyces pombe NRRL Y-12796 & 972* Ascomycota Exobasidium rhododendri CBS 101457 Sympodiomycopsis paphiopedili IAM13459 Rhamphospora nymphaea CBS 72.38 Malassezia pachydermatis CBS 1879 Ustilago tritici CBS 669.70 Schizonella melanogramma CBS 174.42 1.00 Ustilaginomycotina Cintractia limitata HAJB 10488 Melanotaenium endogenum CBS 481.91 Thecaphora spilanthis JAG 53 Tilletiaria anomala CBS 436.72 Agaricus bisporus RWK 1885 Bolbitius vitellinus MTS 5020 Aureoboletus thibetanus HKAS 41151 Boletellus projectellus MB03-118 Ganoderma tsugae ZWsn Polyporus squamosus AFTOL-ID 704 Hydnellum geogenium PBM 2382 Polyozellus multiplex PBM 2412 americanum PBM 2498 Lactarius deceptivus PBM 2462 0.85 Fomitiporia mediterranea 3/22-7 Hydnochaete duportii CBS 939.96 Trechispora alnicola CBS 577.83 1.00 Exidia uvapsassa AFTOL-ID 461 1.00 Sebacina incrustans PBM 2709 & MW 573* Phallus hadriani KH11092003-1 Calocera cornea GEL 5359 0.75 Dacryopinax spathularia GEL 5052 Dacrymycetes Cryptococcus humicola PYCC3387T Cryptococcus sp. CBS 681.93 1.00 1.00 Tremella aurantia E6123 Filobasidium globisporum CBS 7642 & JCM 10633* Filobasidium uniguttulatum JCM 3685 & WH84* Tremellomycetes 1.00 Cystofilobasidium capitaum CBS 6358 & IAM 13521* Itersonilia perplexans CBS 286.50 Mrakia frigida CBS 5266 1.00 T 1.00 Wallemia ichthyophaga CBS 116629 & CBS 113033 * 1.00 Wallemia muriae CBS 116628T & CBS 110583* Wallemiales Wallemia sebi EXF483 & EXF757* T 1.00 Basidioascus undulatus DAOM 241956 Basidioascus undulatus DAOM 241961 1.00 Basidioascus magus DAOM 241948T Basidioascus persicus IBRC M30078T Geminibasidiales 0.98 Geminibasidium donsium DAOM 241966T 1.00 Geminibasidium donsium DAOM 241967 Wallemiomycetes Geminibasidium hirsutum DAOM 241969T Microbotryum violaceum CBS 438.34 Leucosporidium scottii CBS 614 0.74 Rhodosporidium toruloides PYCC4416 Tritirachium sp. CBS 265.96 Sakaguchia dacryoidea PYCC 4491 Occultifur externus CBS 8732 Chrysomyxa arctostaphyli CFB 22246 Pucciniomycotina Puccinia poarum PBM 2567 & clone 19* 1.00 Auriculoscypha anacardiicola AFTOL-ID 1885 Platygloea disciformis IFO 32431 Kondoa malvinella CBS 6082 Sterigmatomyces halophilus CBS 4609 Entorrhiza aff. fineranae PDD70949

0.05

Fig. 2. Bayesian ribosomal (SSU+5.8S+LSU) phylogeny showing the relationship of isolate IBRC M30078T to other fungi in the and the Wallemiomycetes. Posterior probabilities are displayed at nodes of the tree. Asterisks indicate SSU and LSU sequences that were combined belong to different strains of the same species. Bar, 0.05 expected changes per site.

Fermentation, carbohydrate analysis, coenzyme produced hyphal colonies only on YPG, this novel species Q, enzymic profiling should be treated as a yeast-like basidiomycete. While Basidioascus undulatus and Basidioascus magus Yeasts vary in their ability to ferment sugars. In contrast are the only two known species of the genus Basidioascus, to ascomycetes, basidiomycete yeasts are generally less our study introduces a novel species of this genus with fermentative and less copiotrophic (Kurtzman et al., 2011). substantiating morphological and molecular phylogenetic Species of the genera Kluyveromyces, Saccharomyces and evidence. Morphologically, the isolate behaved more like Zygosaccharomyces, for example, ferment glucose and some a yeast in our extensive observations. As the genus other sugars vigorously, whereas few basidiomycetous yeasts Basidioascus is composed of hyphal fungi and our isolate such as Bandoniozyma, Mrakia, Mrakiella, Phaffia and http://ijs.sgmjournals.org 3049 S. Nasr and others

Wallemia muriae CBS 116628T (AY302534) Wallemia sebi EXF483 (AY328917)

T 0.99 Geminibasidium hirsutum DAOM 241969 (JX242880) Geminibasidium donsium DAOM 241966T (JX242877) Basidioascus persicus IBRC M30078T (KC751417) 1.00 Uncultured from France (FN397319) Cryptococcus sp. MD76-1BY from China (EU626069) 1.00 Basidioascus magus DAOM 241954 (JX242857) Basidioascus magus DAOM 241955 (JX242861) Basidioascus magus DAOM 241949 (JX242868)

Basidioascus magus DAOM 241948T (JX242869) 0.1 Basidioascus magus DAOM 241951 (JX242871) 1.00 Basidioascus sp. DAOM 241960 (JX242860) Basidioascus sp. DAOM 241957 (JX242858) Basidioascus sp. DAOM 241965 (JX242867)

Uncultured fungus clone RFLP52 from Australia (DQ388859) Basidioascus undulatus DAOM 241958 (JX242859) Basidioascus undulatus DAOM 241961 (JX242862)

Basidioascus undulatus DAOM 241956T (JX242863) Basidioascus undulatus DAOM 229809 (JX242882) 0.97 Basidioascus undulatus DAOM 241962 (JX242864) Basidioascus undulatus DAOM 241963 (JX242865) Basidioascus undulatus DAOM 241964 (JX242866) Basidioascus undulatus DAOM 241959 (JX242875) Basidioascus undulatus DAOM 242122 (JX242876)

Fig. 3. Bayesian ITS phylogeny showing the relationship of isolate IBRC M30078T to existing species in the class Wallemiomycetes including uncultured sequences. Posterior probabilities are displayed at nodes of the tree. Bar, 0.1 expected changes per site.

Xanthophyllomyces ferment glucose weakly (Kurtzman et al., The carbohydrate components of the cell wall were studied. 2011; Valente et al., 2012). The fermentation ability of the The TLC chromatogram from the whole-cell hydrolysate of novel isolate was studied according the standard methods sugars (Fig. S2) indicated a Tremella type with glucose being of Kurtzman et al. (2011) and Barnett et al. (2000). The predominant, and with xylose, mannose and galactose fermentation result is based on the time and the amount of present in trace amounts (Do¨rfler, 1990). Extracellular gas accumulated. Based on the indicator changes from green amyloid compounds were not found in either solid or liquid to yellow, consumption of sugars was observed after 24 h medium. These results suggested placement of the novel but the accumulation of gas was seen after 6 days. However, species near the Tremellales, consistent with phylogenetic further incubation for up to 2 weeks did not result in analysis. further gas production except for sucrose (Fig. S1). The yeasts and yeast-like fungi have coenzyme Q, which is a Fermentation is considered weakly positive when less than naturally occurring coenzyme formed from the conjugation one-third of the insert Durham tubes is filled with gas, of a benzoquinone ring with a hydrophobic isoprenoid whereas it is considered positive when more than one-third chain of varying chain length, depending on the species is filled with gas. Therefore, the fermentation of D-glucose, (Kurtzman et al., 2011). The number of isoprene units in maltose, melezitose, trehalose and raffinose was recorded the side chains of coenzyme Q differs from Q5 to Q10 as weakly positive and the fermentation of sucrose was among fungal taxa (Olsen, 1990; Kurtzman et al., 2011). In considered positive. general, species of a single genus have the same Q-system,

3050 International Journal of Systematic and Evolutionary Microbiology 64 Basidioascus persicus sp. nov. but exceptions exist in both the Ascomycota and the their cytoplasm and collapse as the attached basidiospore Basidiomycota (Olsen, 1990). The major coenzyme Q in matures (Fig. 1r). Pseudohyphae (Fig. 1s) and septate isolate IBRC M30078T was Q6. hyphae (Fig. 1t) are also observed. No clamp connections are seen. Growth is observed in vitamin-free medium, in 0.1 Determination of enzymic activity profiles in novel micro- and 0.01 % cycloheximide, in 5 and 10 % NaCl, and in 50 organisms can reveal their potential value for biotechno- and 60 % glucose. Growth is positive at 28, 30 and 35 uC, logical application as enzyme producers. In that regard, but negative at 37 and 40 uC in YPG medium. No growth is enzymic activity of isolate IBRC M30078T was studied, and recorded in 16 % NaCl or in the presence of 1 % acetic acid. the preliminary screening revealed DNase and laccase Urease activity and diazonium blue B reaction are positive. activity. Starch-like compounds are not produced. The major coenzyme Q is Q6. Laccase and DNase enzyme activity are Novel yeast-like species of Basidioascus detected. Negative enzyme activity for amylase, protease, lipase, lecithinase, gelatinase, xylanase and cellulase. Positive Morphological, physiological and phylogenetic evidence for fermentation of sucrose and weakly positive for suggests that isolate IBRC M30078T represents a novel fermentation of D-glucose, maltose, melezitose, trehalose species of the genus Basidioascus. The substrate assimilation and raffinose. D-Galactose, D-arabinose, cellobiose, myo- and fermentation results, cell-wall carbohydrate analysis inositol, D-xylose, D-ribose, L-arabinose, L-rhamnose, D- and coenzyme Q determination, as well as enzyme activity arabinitol, L-arabinitol, lactose, succinate, citrate, DL-lactate, tests gave further insight into the metabolism of this novel gluconic acid, starch, inulin, D-mannitol, salicin, methanol, species and these data could serve as a starting point ethanol and n-hexadecane are not fermented. Positive result for comparison studies of other species of Basidioascus. in tests for assimilation of D-glucose, sucrose, maltose, The results supported the identification of isolate IBRC T melezitose, trehalose, raffinose, sucrose, inulin, ethanol, D- M30078 as representing a novel species of the genus ribose (weak), D-xylose (weak), cellobiose, citrate, gluconic Basidioascus, described as Basidioascus persicus sp. nov. acid, mannitol, D-arabinitol, L-arabinose (weak) and Furthermore, the genus description of Basidioascus was arbutin, but negative result for assimilation of D-galactose, emended slightly to accommodate Basidioascus persicus. D-arabinose, L-rhamnose, lactose, myo-inositol, starch, succinate, DL-lactate, salicin, methanol and n-hexadecane. Description of Basidioascus persicus S. Nasr, Ethylamine hydrochloride, creatine, L-lysine, cadaverine M. R. Soudi & H. D. T. Nguyen sp. nov. and potassium nitrate are assimilated, but glucosamine, sodium nitrite, creatinine and imidazole are not. Basidioascus persicus (per9si.cus. L. masc. adj. persicus of T Persia, present Iran, referring to the location where this A dried holotype (IBRC P1010180 ) and an ex-type (IBRC T species was isolated). M30078 ) culture were deposited at the Iranian Biological Resource Center, Tehran, Iran. The dried holotype was MycoBank number: MB 804703 created from a culture of the ex-type grown on YPG dried Colony diameters (mm) after 7 days at 28 uC: MEA, 11–17 under the biological safety cabinet for 4 days in January (mean514.0, n59); YPG, 18 (mean517.8, n59); M40Y, 2014. An isotype strain was deposited at the Centraalbureau voor Schimmelcultures, Fungal Biodiversity Centre, Utrecht, 13–15 (mean514.0, n59); DG18, 10 (mean510.2, n59). T The isolate produces cream-coloured yeast colonies on the Netherlands, as CBS 12808 . The type strain was isolated MEA (Fig. 1a), white hyphal colonies with a beige centre on from the soil of a kitchen garden in Shahryar city, Tehran YPG at 28 uC (Fig. 1b), cream-coloured yeast colonies with province, Iran. a yellow centre on M40Y (Fig. 1c) and olive-coloured yeast colonies on DG18 (Fig. 1d). Forms arthroconidia 7.0– Emended description of Basidioascus Matsush., 13.062.5–4.5 mm (mean±SE510.0±0.463.4±0.1 mm) Matsushima Mycological Memoirs 10:98 (2003) (Fig. 1e), cells that are hyaline and oval, 5.5–12.063.5– [2001], emend H.D.T. Nguyen 5.5 mm (mean±SE58.3±0.464.7±0.1 mm) (Fig. 1f) and MycoBank number: MB28558 didymospores 10.0–17.564.5–7.5 mm(mean±SE513.3± 0.465.8±0.2 mm) (Fig. 1g). Oval cells reproduce by mono- Basidia are produced singly or in clusters, and have polar budding and sometimes produce ballistoconidia 3.5– granular cellular contents and one or more basal lateral 7.563.0–7.0 mm(mean±SE55.1±0.364.7±0.3 mm) attached projections. One sterigma, and in one species more than by short sterigmata (Fig. 1h) that may be forcibly ejected one sterigma, arises randomly over the apical two-thirds of leaving a scar on the mother oval cell (Fig. 1k). Basidia are the surface of the basidium. Only one mature basidiospore 7.0–14.063.0–6.0 mm(mean±SE58.7±0.464.8±0.1 mm) develops per basidium. Basidiospores are symmetrical on (Fig. 1n–r), with one or sometimes more than one basal sterigma, hyaline or brown when mature. Basidia lose their lateral projection (Fig. 1n–p), and are seen with one cytoplasm and sometimes collapse as basidiospores mature. basidiospore attached by a long sterigma. Basidiospores Arthroconidia are produced in two species and didymos- are globose, orange to brown, verrucose (Fig. 1q) and 2.0– pores are observed in one species. Species are xerotolerant 6.0 mm in diameter (mean±SE53.8±0.2 mm). Basidia lose and two species are heat-resistant. Analyses of SSU, http://ijs.sgmjournals.org 3051 S. Nasr and others

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