FEMS Microbiology Letters, 363, 2016, fnv236

doi: 10.1093/femsle/fnv236Advance Access Publication Date: 14 December 2015Research Letter

RESEARCH LETTER –Taxonomy & Systematics

Aspergillus oerlinghausenensis, a new mould speciesclosely related to A. fumigatusJos Houbraken1,∗, Michael Weig2, Uwe Groß2, Martin Meijer1

and Oliver Bader2

1CBS-KNAW Fungal Biodiversity Centre, Uppsalalaan 8, 3584 CT Utrecht, the Netherlands and 2Institute forMedical Microbiology, University Medical Center Gottingen, Gottingen, Germany∗Corresponding author: CBS-KNAW Fungal Biodiversity Centre, Uppsalalaan 8, 3584 CT Utrecht, the Netherlands. Tel: +31-(0)302122600;Fax: +31-(0)30-2512097; E-mail: j.houbraken@cbs.knaw.nlOne sentence summary: Aspergillus oerlinghausenensis, a new itraconazole-resistant species related to A. fumigatus was described using a combination ofphenotypic characters and multilocus sequencing.Editor: Stefanie Poeggeler

ABSTRACT

Two isolates belonging to Aspergillus section Fumigati were recovered from German soil on itraconazole containing agarmedia. Phylogenetic analysis and phenotypic characterization of both isolates show that they represent a novel speciesnamed Aspergillus oerlinghausenensis (holotype CBS H-22119HT, ex-type CBS 139183T = IBT 33878 = DTO 316-A3). The speciesis phylogenetically related to A. fischeri and A. fumigatus. Aspergillus oerlinghausenensis can be differentiated from A. fischeriby its higher growth rate at 50◦C. Furthermore, A. oerlinghausenensis is protoheterothallic as only the MAT1-1 idiomorph wasdetected, while A. fischeri is homothallic. The species differs from A. fumigatus by a weak sporulation on malt extract agar at25◦C, a floccose colony texture on Czapek yeast extract agar and malt extract agar and subglobose instead of subclavatevesicles. The cyp51A promoter region of A. oerlinghausenensis deviates from the previously reported cyp51A promoter regionsin A. fumigatus and potentially presents a novel azole resistance conferring modification. Due to the close relationship of A.oerlinghausenensis with A. fischeri and A. fumigatus, this species is placed in a good position for comparative studies involvingthese species.

Keywords: phylogeny; taxonomy; Fumigati; itraconazole; thermophiles; cyp51A

INTRODUCTION

Aspergillus section Fumigati (= A. fumigatus group) was intro-duced for thermotolerant and thermophilic species with unise-riate, columnar conidial heads and green-colored conidia (Raperand Fennell 1965). Before changes were made to the current In-ternational Code of Nomenclature for algae, fungi and plants,during the age of dual nomenclature, the species belonging tothis section that could produce a sexual state were classifiedin Neosartorya. Following the proposal of Samson et al. (2014),all species of this section, including the sexual reproducingones, are named under their Aspergillus name. In 2007, Samson

et al. (2007) accepted 33 species in section Fumigati. Since then,the number of species in this section steadily increased andmore recently Sugui et al. (2014) accepted 51 species. The mostwell-known member of this section is A. fumigatus. This speciesretrieved much attention in clinical settings as it can cause awide spectrum of diseases ranging from mild to severe life-threatening symptoms in humans (De Hoog et al. 2000). In foodindustries, other species of this section are of importance be-cause of their ability to spoil pasteurized products (Samsonet al. 2010). An example is the heat-resistant species A. fischeri(synonym Neosartorya fischeri), a sister species of A. fumigatus.

Received: 8 October 2015; Accepted: 10 December 2015C© FEMS 2015. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com

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Actually, in order to gain insight in the virulence and epidemiol-ogy of A. fumigatus, A. fischeri and A. fumigatus were among thefirst genome sequenced taxa in Aspergillus (Fedorova et al. 2008).

In the last decade, an increase of azole-resistant A. fumiga-tus strains was observed (Snelders et al. 2008). During the surveyon the occurrence of such azole-resistant A. fumigatus strains inGerman soils (Bader et al. 2015), one sample yielded two colonies,visually appearing as A. fumigatus, but these isolates could notbe identified bymatrix-assisted laser desorption ionization-timeof flight mass spectrometry (MALDI-TOF). Sequencing of the ITSlocus showed that the isolates belonged toAspergillus section Fu-migati, but no significant similarity matches could be retrieved.

Using a polyphasic approach, combining partial β-tubulin(BenA) and calmodulin (CaM) sequence data, macro- and micro-morphology characteristics and temperature-growth data, weshow that the two isolates represent a yet undescribed species,and the name A. oerlinghausenensis sp. nov. is here proposed.

MATERIALS AND METHODS

Strains

Both investigated strains were recovered from a soil samplecollected in Oerlinghausen, Germany as described previously(Bader et al. 2015). In short, the soil was suspended in 0.5%saponin, and after settlement of the debris, the supernatantwas centrifuged. The obtained pellet was resuspended, platedon itraconazole or voriconazole-containing agar and incubatedat 40◦C for at least 3 days. Resulting colonies were spread ontofresh agar, and their drug susceptibility was determined accord-ing to EUCAST rules (EUCAST 2008). The strains were depositedin the CBS-KNAW Fungal Biodiversity Centre (CBS) culture col-lection (Utrecht, the Netherlands) under accession numbers CBS139183 ( = IBT 33878 = DTO 316-A3) and CBS 139184 ( = IBT33877 = DTO 316-A4). The type specimen was placed in the CBSherbarium.

Molecular studies

DNA extraction was performed from 2 to 3 days old cultureson malt extract agar (MEA; Oxoid) using the Ultraclean Micro-bial DNA isolation Kit (Mo-Bio, Solana Beach, USA) and DNAwas stored at –20◦C. The ITS barcode and a part of the CaM andBenA gene was amplified and sequenced according the meth-ods described by Houbraken, Spierenburg and Frisvad (2012). Se-quence contigs were assembled in Seqman v. 10 (DNA-Star Inc.).Primers developed by Barrs et al. (2013) were used to determinethemating type of both isolates. Positive controlswereA. fumiga-tus CBS 127800 (MAT1-1), CBS 127801 (MAT1-2) (both ex-air, Ire-land; O’Gorman, Fuller and Dyer 2009) and A. fischeri CBS 544.65T

(MAT1-1 andMAT1-2). To determine the possible drug resistancemechanism, the cyp51A genewas sequenced as described before(Bader et al. 2015). Newly generated sequences were deposited inGenBank under accession numbers KT359601– KT359609.

Phylogeny

β-tubulin and calmodulin reference sequences of strains be-longing to Aspergillus section Fumigati (Samson et al. 2014; Suguiet al. 2014) were downloaded from GenBank, and the newly gen-erated sequences were added to this data set. The data setswere aligned using the Muscle software. Prior to the analysis,the most suitable substitution model was determined, utilizingthe Akaike Information Criterion (AIC). Statistical support of the

data sets was measured by Maximum Likelihood (ML). Aligningof the sequences and the analyses were performed usingMEGA6(Tamura et al. 2013). Bootstrap values (bs) higher than 70% wereplotted on the phylograms, and both phylograms were rootedwithA. clavatus (NRRL 1). The ITS sequenceswere not used in thephylogenetic analysis and were only examined for their speciesidentification ability.

Morphology and physiology

The macro- and micromorphology of the two strains was com-pared with data published in literature. In addition, typicalstrains of A. fumigatus CBS 127800 and CBS 127801, and A. fis-cheri CBS 544.65T and DTO 307-E3 were included in this anal-ysis. For macroscopic analysis, all isolates were inoculated ina three-point pattern on creatine sucrose agar (CREA), Czapekyeast autolysate agar (CYA), CYA supplemented with 5% NaCl(CYAS), dichloran 18% glycerol agar (DG18), MEA, oatmeal agar(OA), Sabouraud dextrose agar (SDA, Difco) and yeast extract su-crose agar (YES). Difco’s yeast extract was used in the agar me-dia. Colony diameters and other macroscopical features wererecorded after 7 days of incubation at 25◦C in darkness. The iso-lates on MEA and OA were incubated up to 10 weeks and fol-lowed in time for potential ascospore formation. Furthermore,growth curves were made on CYA and MEA incubated for 7 daysat 27◦C, 30◦C, 33◦C, 36◦C, 37◦C, 40◦C, 45◦C and 50◦C. For micro-scopic examination, mounts of the strains were made in lacticacid from the MEA colonies according Samson et al. (2014).

RESULTS AND DISCUSSION

Phylogenetic analysis and sequence-basedidentification

The length of the BenA data setwas 423 basepairs (bp) and that ofthe CaM data set 553 bp. The Kimura 2 parameter with gammadistribution (+G) was found to be the most suitable model forboth alignments. The BenA and CaM phylogenies show that theisolates from German soil (CBS 139183 and CBS 139184) are ac-commodated in Aspergillus section Fumigati (Fig. 1). These twoisolates are in both phylograms most closely related to A. fumi-gatus (BenA 83% bs; CaM 83% bs) and basal to these two speciesis A. fischeri (BenA 98% bs; CaM 98% bs).

Nonucleotide differenceswere observed between the twoAs-pergillus strains. These sequences are different from the otherspecies in section Fumigati. The percentage difference at theBenA, CaM and ITS loci were 95.5%, 95.6% and 99.7%, respec-tively, when compared with the type of A. fumigatus (NRRL 163T)and 96.3%, 97.3% and 99.2%, respectively, with A. fischeri NRRL181T. A BLAST analysis with the ITS barcode of these isolateson GenBank didn’t retrieve any 100% similarity matches and thehighest similarities were with A. fumigatus isolates. Based onthis data, it can be concluded that ITS barcoding can be used todifferentiate A. oerlinghausenensis from other species in sectionFumigati.

Morphology and physiology

The two A. oerlinghausenensis isolates are phenotypically simi-lar in their growth rates and sporulation patterns on the usedagar media. The isolates attain a diameter larger than 50 mm in7 days at 25◦C on MEA, CYA, YES and OA and produce green col-ored conidia. Furthermore, the strains produce columnar coni-dial heads, are uniseriate, and have subglobose vesicles. This

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0.01

NRRL 1 Aspergillus clavatus EF669802NRRL 20748 Aspergillus thermomutatus EF669805KACC 41691 Aspergillus assulatus DQ114123CBS 841.96 Aspergillus papuensis AY870738

CCF 4149 Aspergillus brevis pitatus HF933364CCF 4190 Aspergillus conversis HF933363

NRRL 35723 Aspergillus solicola EU220283IBT 16756 Aspergillus galapagensis DQ534145

IFM 57611 Aspergillus shendaweii AB488754CBS 646.95 Aspergillus mul plicatus DQ114129

IFM 57609 Aspergillus tsunodae AB48875599

NRRL 577 Aspergillus unilateralis EF669852IFM 61445 Aspergillus caa ngaensis AB743855

IFM 61342 Aspergillus pernambucoensis AB74385690

CCM 8003 Aspergillus marvanovae HE974387NRRL 179 Aspergillus waksmanii EF669794

IBT 3016 Aspergillus nishimurae DQ534154KACC 42091 Aspergillus turcosus DQ534143

NRRL 4378 Aspergillus auratus EF669835NRRL 4652 Aspergillus stramenius EF669840

100

NRRL 2163 Aspergillus neoglaber EU014107CBM FA-0933 Aspergillus tsurutae AB488760

NRRL 2154 Aspergillus quadricinctus EF669806CBS 253.94 Aspergillus primulinus DQ53415898

98

NRRL 2439 Aspergillus brevipes EF669812NRRL 4021 Aspergillus duricaulis EF66982777

NRRL 2392 Aspergillus australensis EF669811CBS 294.93 Aspergillus hiratsukae AF057324

NRRL 20549 Aspergillus spathulatus EF669803NRRL 5534 Aspergillus fennelliae AF057320

NRRL 32571 Aspergillus otanii EF669818IFM 57847 Aspergillus huiyaniae AB787219

CBS 652.73 Aspergillus den culatus DQ114125NRRL 4179 Aspergillus ferenczii EF669833IFM 53598 Aspergillus sublevisporus AB48875996

99

98

CBS 101754 Aspergillus delicatus DQ114124NRRL 4584 Aspergillus tatenoi EF669838

87

NRRL 2244 Aspergillus aureolus EF669808CBM FA-0702 Aspergillus udagawae AF132226

89

KUFC 6349 Aspergillus siamensis AB646989CCF 4417 Aspergillus wyomingensis HF933359NIH AV1 Aspergillus pseudoviridinutans KJ914690

CM 3147 Aspergillus parafelis KJ914692CBS 130245 Aspergillus felis JX021700CM 6087 Aspergillus pseudofelis KJ914697

9990

81

IMI 062875 Aspergillus viridinutans AF134779IFM 61334 Aspergillus arcoverdensis AB818845

79

KACC 41659 Aspergillus coreanus AY870758CBM FA-884 Aspergillus takakii AB787221

KACC 41657 Aspergillus laciniosus AY870756CBM FA-0690 Aspergillus paulistensis AF132231

NRRL 5034 Aspergillus spinosus EF669844CBM FA-0672 Aspergillus botucatensis AF132225

92

IBT 12703 Aspergillus fumiga affinis DQ094885IBT 16806 Aspergillus novofumigatus DQ094886

97

IFM 42277 Aspergillus fumisynnematus AB248076NRRL 35552 Aspergillus lentulus EF669825

99

NRRL 181 Aspergillus fischeri EF669796CBS 139183 Aspergillus oerlinghausenensis KT359603CBS 139184 Aspergillus oerlinghausenensis KT359604

99

NRRL 163 Aspergillus fumigatus EF669791NRRL 5109 Aspergillus fumigatus EF669845

10083

98

83

76

81

BenA

0.1

NRRL 1 Aspergillus clavatus EF669871NRRL 20549 Aspergillus spathulatus EF669872

NRRL 181 Aspergillus fischeri EF669865CBS 139183 Aspergillus oerlinghausenensis KT359605CBS 139184 Aspergillus oerlinghausenensis KT359606100

NRRL 163 Aspergillus fumigatus EF669860NRRL 5109 Aspergillus fumigatus EF669915

10083

98

IFM 42277 Aspergillus fumisynnematus AB259968NRRL 35552 Aspergillus lentulus EF669895

99

IBT 12703 Aspergillus fumiga affinis DQ094891IBT 16806 Aspergillus novofumigatus DQ094893

72

KACC 41657 Aspergillus laciniosus AY870716CBM FA-884 Aspergillus takakii AB787566CBM FA-0690 Aspergillus paulistensis AB48876699

97

KACC 41659 Aspergillus coreanus AY870718NRRL 5034 Aspergillus spinosus EF669914CBM FA-0672 Aspergillus botucatensis AY87072398

100

IMI 367415 Aspergillus viridinutans DQ534162IFM 61334 Aspergillus arcoverdensis AB818856

CCF 4417 Aspergillus wyomingensis HF933397KUFC 6397 Aspergillus siamensis AB776704NRRL 2244 Aspergillus aureolus EF669877

CBM FA-0702 Aspergillus udagawae AB74856678

85

NIH AV1 Aspergillus pseudoviridinutans KJ914708CBS 458.75 Aspergillus sp. HG426048

100

CM 3147 Aspergillus parafelis KJ914702CBS 130245 Aspergillus felis JX021715CM 6087 Aspergillus pseudofelis KJ91470591

96

96

91

95

NRRL 20748 Aspergillus thermomutatus EF669874CBS 101754 Aspergillus delicatus DQ114132

NRRL 4584 Aspergillus tatenoi EF66990895

NRRL 4021 Aspergillus duricaulis EF669897NRRL 2439 Aspergillus brevipes EF669881

CBS 253.94 Aspergillus primulinus DQ534170NRRL 2154 Aspergillus quadricinctus EF669875

CBM FA-0933 Aspergillus tsurutae AB488768

100

78

NRRL 5534 Aspergillus fennelliae EF669920NRRL 32571 Aspergillus otanii EF669888

99

IFM 57847 Aspergillus huiyaniae AB787564CBS 652.73 Aspergillus den culatus DQ114133

NRRL 4179 Aspergillus ferenczii EF669903IFM 53598 Aspergillus sublevisporus AB48876791

88

99

CBS 294.93 Aspergillus hiratsukae AY870699IFM 61445 Aspergillus caa ngaensis AB743861NRRL 577 Aspergillus unilateralis EF669923

CBS 646.95 Aspergillus mul plicatus DQ114137IFM 57609 Aspergillus tsunodae AB488763

IBT 27911 Aspergillus assulatusCCM 8003 Aspergillus marvanovae HE974389KACC 42091 Aspergillus turcosus DQ53414877

IFM 54133 Aspergillus nishimurae HE974392NRRL 179 Aspergillus waksmanii EF669863

82

76

NRRL 2163 Aspergillus neoglaber EU014120IFM 57611 Aspergillus shendaweii AB488762

IBT 16756 Aspergillus galapagensis DQ534151NRRL 4378 Aspergillus auratus EF669905NRRL 4652 Aspergillus stramenius EF669910

88

IFM 61342 Aspergillus pernambucoensis AB743862NRRL 2392 Aspergillus australensis EF669880NRRL 35723 Aspergillus solicola EU220284

CBS 841.96 Aspergillus papuensis AY870697CCF 4149 Aspergillus brevis pitatus HF933388

CCF 4190 Aspergillus conversis HF93338799

CaM

Figure 1. ML based on partial β-tubulin (left) and calmodulin (right) sequences, showing the phylogenetic relationship among members of Aspergillus section Fumigati.Isolates of the newly described species A. oerlinghausenensis are presented in bold font. Bootstrap percentages of the ML analysis are presented at the nodes and thoselower than 70% are not shown. The bars indicate the number of substitutions per site and the phylogram is rooted with A. clavatus NRRL 1.

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combination of characteristics confidentially places these iso-lates in section Fumigati, and this placement is confirmed bymolecular phylogenetic data. The isolates are phylogeneticallyrelated to A. fischeri and A. fumigatus. The ability to grow at 50◦Cis a distinguishing feature for A. fumigatus, and this characteris used to differentiate it from other clinical section Fumigatispecies (Balajee et al. 2007). Interestingly, A. oerlinghausenensisshares the ability to grow at 50◦CwithA. fumigatus andA. fischeri.On the other hand, it differs from A. fumigatus by a weak sporu-lation onMEA at 25◦C, a floccose colony texture on CYA andMEAand subglobose instead of subclavate vesicles. The phylogeneti-cally related species A. fischeri grows slower at 50◦C (5–25 mm atCYA, 7 days) and readily produces a sexual state on oatmeal andmalt extract agar. No sexual state is observed in A. oerlinghause-nensis cultures.

Mating type analysis

Only the MAT1-1 idiomorph was detected in A. oerlinghausenen-sis during the mating type PCR analysis. The PCR ampliconswere sequenced and a homology search on GenBank confirmedtheir designation to MAT1-1. This data show that A. oerling-hausenensis is protoheterothallic. This term was introduced forasexual species where genetic evidence, such as the presenceof complementary MAT loci, indicates the presence of a sex-ual cycle (Houbraken and Dyer 2014). The A. oerlinghausenensiscultures were inoculated on oatmeal agar and incubated up to10 weeks. No sexual state was detected in the cultures; how-ever, hyphal coils resembling ascoma initials were observed bylight microscopy. Although it is possible that only isolates witha MAT1-1 signature are present in nature (clonal distribution),heterothallism is also frequently occurring in section Fumigati(Houbraken and Dyer 2014). When strains with a MAT1-2 signa-ture are isolated, then experiments can then be performed toshow whether this species has a fully functional heterothallicmating system.

Ecology

BLAST searches with BenA, CaM and ITS sequences of A. oerling-hausenensis on GenBank, and the internal sequence database atCBS-KNAW Fungal Biodiversity Centre did not retrieve any highsimilarity matches. This suggests that A. oerlinghausenensis is anuncommon species and is therefore probably not relevant as ahuman pathogen. However, this would yet have to be confirmedexperimentally e.g. in a mouse model system. This species isphylogenetically closely related to A. fumigatus and shares sev-eral characters with this species such as growth at 50◦C and a(proto)heterothallic mating system. This places A. oerlinghause-nensis in a good position as a potential non-pathogenic speciesfor comparative virulence studies with A. fumigatus. The inves-tigated strains were isolated from azole containing agar media,and it is therefore unknown whether this resistance phenotypeis due to mutations in the cyp51A gene, or if it is an intrinsic fea-

Table 1. Overview of MIC values of A. oerlinghausenensis and re-lated species (ITZ = itraconazole; VRZ = voriconazole; PSZ =posaconazole).

ITZ VRZ PSZ

A. fumigatus DSM 819 0.5 0.25 0.25A. fumigatus CBS 487.65 1 0.25 0.25A. fischeri NRRL 181 1 1 0.5A. oerlinghausenensis CBS 139183 >4 2 0.5A. oerlinghausenensis CBS 139184 >4 2 0.5

ture of A. oerlinghausenensis. Other species, such as A. lentulus,are intrinsically resistant against azoles while A. fumigatus azoleresistance has been defined as secondary or acquired (Melladoet al. 2011). The isolates of A. oerlinghausenensis investigated herewere confirmed to be resistant against azoles by the EUCASTmi-crodilution method (Table 1). The predicted Cyp51A protein se-quence (KT359609) did not yield any amino acid substitutionsparticularly suspicious for azole resistance. Comparison of thecyp51A promotors of azole susceptible A. fumigatus and A. fis-cheri strains showed a 6 bp insertion in the cyp51A promoter ofA. oerlinghausenensis preceding the region duplicated in A. fumi-gatus TR34-isolates (Fig. 2). Potentially, this might explain theobserved susceptibility profile; however, the possibility that an-other resistance pathway is present should also be consideredas azole resistant A. fumigatus isolates with a wild-type versionof the cyp51A can readily be found (Bueid et al. 2010; Bader et al.2013, 2015).

Taxonomy

Based on the data presented above, we show that CBS 139183and CBS 139184 are well separated from all other species in sec-tion Fumigati and describe A. oerlinghausenensis here as a newspecies. We choose to describe this species under the Aspergillusname following the concept of Samson et al. (2014).

Aspergillus oerlinghausenensis Bader &Houbraken, sp. nov.My-cobank MB813868 (Fig. 3).

Etymology: named after Oerlinghausen, Germany, the collec-tion location of the type strain.

Diagnostic characters: this species is phylogenetically relatedtoA. fischeri andA. fumigatus.Aspergillus oerlinghausenensis growswell on MEA and CYA incubated at 50◦C, is weakly sporulatingon MEA and CYA at 25◦C, and is strongly sporulating at highertemperatures. The species has subglobose vesicles.

In: subgenus Fumigati section Fumigati.Typus: ex-soil, Oerlinghausen, Germany (lat. 51.950950; long.

8.658570), I. Szymczak (CBS H-22119 – holotypus, cultures ex-type: CBS 139183 = IBT 33878 = DTO 316-A3 = Isolate 227).

Description: colony diam, 7 days, 25◦C (mm): CYA 50–55; MEA>60; YES >60; DG18 40–45; CYAS 18–22; OA >60; SDA >60; CREA35–40, poor growth, no acid production. Other incubation tem-peratures: CYA 30◦C >60; CYA 37◦C >60; CYA 45◦C >60; CYA 50◦C

Figure 2. Comparison of the cyp51A promoters of azole susceptible A. fumigatus and A. fischeri strains with those of A. oerlinghausenensis. The underlined nucleotidesare duplicated in resistant TR34-isolates of A. fumigatus.

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Figure 3. Aspergillus oerlinghausenensis CBS 139183. (A) Colonies grown for 7 days, from left to right (top row) CYA, YES, MEA, SDA; bottom row CYA reverse, YES reverse,

DG18, CREA. (B–F) Conidiophores. (G) Hyphal coils. (H) Conidia.

30–40; MEA 45◦C >60; MEA 50◦C 32–37; optimum growth temper-ature 37◦C, maximum > 50◦C.

Macromorphology: CYA 25◦C, 7 days: colony surface floccosewhen sporulating, mostly consisting of white sterile mycelia,pale grayish green when sporulating; soluble pigment absent;exudate mostly absent, sometimes present in center as smallclear droplets; reverse brown to orange–brown in center, palebrown near margins. MEA 25◦C, 7 days: colony surface floccose,mycelium white, conidia blue green; exudate absent; reversebrown. SDA 25◦C, 7 days: clony surface floccose, mostly consist-ing of white sterile mycelium, pale green in sporulating parts;soluble pigments absent; exudate absent; reverse pale brown.YES 25◦C, 7 days: colony surface floccose in sporulating iso-

lates, mostly consists of white sterile mycelia, dull gray–greenwhen sporulating; soluble pigment absent; exudate absent, re-verse yellow to olive brown. DG18 25◦C, 7 days: colony surfacestrongly floccose in center, strong sporulation, blue green; sol-uble pigment absent; exudate absent, reverse white. OA 25◦C, 7days: colony surface floccose when sporulating, mainly consist-ing of white mycelium, sporulation sparse, pale green; solublepigment absent; exudate absent; reverse white.

Micromorphology: Hyphal coils, resembling initials observedafter 7 days of incubation; ascomata not observed on MEA andOA after 10 weeks of incubation at 25◦C, 30◦C and 37◦C. Conidialheads columnar, uniseriate. Stipes hyaline, smooth walled, 100–30 × 5.0–7.0 μm. Vesicles subglobose, up to 30 μm in diameter

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when fully grown, smaller (<15 μm) when young. Phialides am-pulliform, 6.5–8.5 × 2.0–3.0 μm, covering ∼50%–75% of the head.Conidia subglobose to broadly ellipsoid, smooth to finely rough-ened, 2.0–3.0 × 2.5–3.0 μm; average width/length = 0.89, n = 53;Hulle cells absent.

ACKNOWLEDGEMENTS

Uwe Braun is acknowledged for his suggestion on the newspecies name, and Rob Samson is thanked for his review of themanuscript.

FUNDING

This work was supported in part by a research grant from PfizerGermany [grant no. WI186302 to O.B.].

Conflict of interest. None declared.

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