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Research Article
Ascomycetes from the Qilian Mountains, China – Hypocreales
expand article infoZhao-Qing Zeng‡, Huan-Di Zheng‡, Xin-Cun Wang‡, Sheng-Long Wei§, Wen-Ying Zhuang‡
‡ Chinese Academy of Sciences, Beijing, China
§ Hexi University, Zhangye, China

Corresponding author: Wen-Ying Zhuang ( zhuangwy@im.ac.cn )

Academic editor: Huzefa Raja
© 2020 Zhao-Qing Zeng, Huan-Di Zheng, Xin-Cun Wang, Sheng-Long Wei, Wen-Ying Zhuang.
This is an open access article distributed under the terms of the Creative Commons Attribution License (CC BY 4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
Citation: Zeng Z-Q, Zheng H-D, Wang X-C, Wei S-L, Zhuang W-Y (2020) Ascomycetes from the Qilian Mountains, China – Hypocreales. MycoKeys 71: 119-137. https://doi.org/10.3897/mycokeys.71.55009
Open Access

Abstract

To investigate fungi from the Qilian Mountains in Gansu Province, ascomycetous specimens were collected and hypocrealean fungi were examined. Eighteen species belonging to six genera in the families Hypocreaceae and Nectriaceae were identified, including 11 species of Hypomyces and Trichoderma in Hypocreaceae and seven species of Nectria, Stylonectria, Thelonectria, and Thyronectria in Nectriaceae. Among them, Stylonectria qilianshanensis and Trichoderma gansuanum are new to science. DNA sequence analyses of combined ACL1, ITS, RPB2, and TEF1 regions confirmed their taxonomic placements. Morphological distinctions between the new species and their close relatives are discussed. Hypomyces tremellicola is reported for the first time in China.

Keywords

Biodiversity, Hypocreaceae, Nectriaceae, sequence analysis, taxonomy

Introduction

The Qilian Mountains is located across the northeastern Qinghai and western Gansu provinces (35°50'–39°19'N, 94°10'–103°04'E) at elevations ranging from 4000 to 6000 m. The area has a temperate climate with mean annual precipitation around 400 mm. Forests are composed mainly of mixed broad-leaf and coniferous trees. The Qilian Mountain National Nature Reserve, where ascomycetes were surveyed, is extremely diverse in climate, vegetation, and geographic structure. Previous investigations of fungal resources have mainly focused on Basidiomycota (Gui et al. 2010; Xi et al. 2011). Our understanding of ascomycetes of the region needs to be broadened.

The order Hypocreales includes about 2700 species in 240 genera, which are divided into 12 families (Rossman et al. 1999; Lumbsch and Huhndorf 2007; Kirk et al. 2008; Lombard et al. 2015; Maharachchikumbura et al. 2016; Sun et al. 2017; Zhuang and Zeng 2017). Hypocrealean collections from the Qilian Mountains belong to the families Hypocreaceae and Nectriaceae. These families are ubiquitous in nature and exhibit very high species diversity in temperate and tropical regions (Rossman 1996; Rossman et al. 1999). They are economically important in fields of industry, environment protection, and agriculture. For instance, some species of Trichoderma Pers. play vital roles in production of industrial enzymes and antibiotics (Jangiret et al. 2017), while several species of Hypomyces (Fr.) Tul. & C. Tul. are pathogens of cultivated mushrooms (Tamm and Põldmaa 2013). Some members of Nectria (Fr.) Fr., Thelonectria P. Chaverri & C. Salgado, and Thyronectria Sacc. cause Abies and Rubus cankers (Hirooka et al. 2011, 2012; Salgado-Salazar et al. 2015). Therefore, discovery of fungi in Hypocreales is of theoretical and practical importance. Improvement and updating our knowledge of the group will provide useful information about sustainable utilization and conservation of natural resources.

Studies on fungi of this group in China dates back to 1895 when Trichoderma cornu-damae (Pat.) Z.X. Zhu & W.Y. Zhuang (as Hypocrea cornu-damae Pat.) was reported on rotten wood in Sichuan Province (Patouillard 1895). Early studies were initiated by Teng (1934, 1935, 1936, 1963), and recent studies are summarized by Liang (2007), Zhuang (2013), and Zhuang and Zeng (2017). A survey of ascomycetes in the Qilian Mountains was carried out in 2018. A total of 67 specimens were examined in this study. Eighteen taxa belonging to seven genera were identified, including 11 species of Hypomyces and Trichoderma in Hypocreaceae, and seven of Nectria, Thelonectria, Thyronectria, and Stylonectria Höhn. in Nectriaceae. Stylonectria qilianshanensis and Trichoderma gansuanum are described and illustrated as new species. Hypomyces tremellicola is reported for the first time from China.

Materials and methods

Sampling and morphological studies

Specimens were collected from the Qilian Mountains in Gansu Province, and they are deposited in the Herbarium Mycologicum Academia Sinica (HMAS). Cultures are kept in the State Key Laboratory of Mycology, Institute of Microbiology, Chinese Academy of Sciences. The methods of Luo and Zhuang (2010) and Põldmaa et al. (2019) were followed for morphological observations. The ascomatal wall reactions to 3% potassium hydroxide (KOH) and 100% lactic acid (LA) were tested. To observe microscopic characteristics of perithecial walls, sections were made with a freezing microtome (YD-1508-III, Jinhua, China) at a thickness of 6–8 μm. Cotton blue lactophenol solution and lactic acid solution were used as mounting media for examinations of anatomical structures and measurements of perithecia, asci, and ascospores. Photographs were taken with a Leica DFC450 digital camera (Wetzlar, Germany) attached to a Leica M125 stereomicroscope (Milton Keynes, UK) for gross morphology, and a Zeiss AxioCamMRc 5 digital camera (Jena, Germany) attached to a Zeiss Axio Imager A2 microscope (Göttingen, Germany) for anatomy. Measurements of individual structures were based on n = 30, except as otherwise noted. The culture of Hypomyces tremellicola was isolated from conidia found on the surface of the host. To determine colony features and growth rates, strains were grown on malt extract agar [MEA, 2% (w/v) malt extract+ 2% (w/v) agar] and potato dextrose agar [PDA, 20% (w/v) potato + 2% (w/v) dextrose + 2% (w/v) agar] in 90 mm plastic Petri dishes at 25 °C for 7 d. For observation of conidiophores and conidia, cultures were grown on PDA at 25 °C with alternating periods of light and darkness (12 h/12 h).

DNA extraction, PCR amplification, and sequencing

Genomic DNA was extracted from dry specimens or fresh mycelia following the method of Wang and Zhuang (2004). Four primer pairs, acl1-230up/acl1-1220low (Nowrousian et al. 2000), ITS5/ITS4 (White et al. 1990), RPB2-5f/RPB2-7cR (Liu et al. 1999), EF1-728F/TEF1LLErev (Carbone and Kohn 1999; Jaklitsch et al. 2005) were used to amplify the ACL1, ITS, RPB2, and TEF1 gene regions, respectively. PCR reactions were performed using an ABI 2720 Thermal Cycler (Applied Biosciences, Foster City, USA) with a 25 μl reaction system consisting of 12.5 μl Taq MasterMix, 1 μl each primer (10 μM), 1 μl template DNA, and 9.5 μl ddH2O. DNA sequencing was carried out in both directions on an ABI 3730XL DNA Sequencer (Applied Biosciences, Foster City, USA) based on the procedures detailed in Gräfenhan et al. (2011), Jaklitsch et al. (2005), and Chaverri et al. (2011).

Sequence alignment and phylogenetic analyses

Newly obtained sequences and those retrieved from GenBank are listed in Tables 1 and 2, respectively. The sequences were assembled, aligned and the primer sequences were trimmed using BioEdit 7.0.5 (Hall 1999), and converted to NEXUS files by ClustalX 1.83 (Thompson et al. 1997). The aligned sequences were combined in BioEdit and analyzed with Bayesian inference (BI) and maximum parsimony (MP) methods to determine the phylogenetic positions of the new species. The MP analysis was performed with PAUP 4.0b10 (Swofford 2002) using 1000 replicates of heuristic search with random addition of sequences and subsequent TBR (tree bisection and reconnection) branch swapping. Topological confidence of the resulting trees was tested by maximum parsimony bootstrap proportion (BP) with 1000 replications, each with 10 replicates of random addition of taxa. The Bayesian inference (BI) analysis was conducted by MrBayes 3.1.2 (Ronquist and Huelsenbeck 2003) using a Markov chain Monte Carlo algorithm. MrModeltest v. 2.3 was used to determine the nucleotide substitution models (Nylander 2004). Four Markov chains were run simultaneously for 1000000 generations with the trees sampled every 100 generations. A 50% majority rule consensus tree was computed after excluding the first 2500 trees as ‘burn-in’. Bayesian inference posterior probability (PP) was determined from the remaining trees. Branch support measures were calculated with 1000 bootstrap replicates. Trees were examined by TreeView 1.6.6 (Page 1996). The BIPP greater than 90% and MPBP greater than 70% were shown at the nodes.

Table 1.
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List of Stylonectria species and the relatives, herbarium/strain numbers and GenBank accession numbers of materials used in this study.

Species Herbarium/strain numbers GenBank accession numbers
ACL1 ITS RPB2
Albonectri rigidiuscula (Berk. & Broome) Rossman & Samuels CBS 122570 HQ897896 HQ897815 HQ897760
Clonostachys rosea (Preuss) Mussat CML817/CBS 114056 KX184866 KC806254 DQ522415
Cyanonectria cyanostoma (Sacc. & Flageolet) Samuels & P. Chaverri CBS 101734 HQ897895 FJ474076 HQ897759
Dialonectria episphaeria (Tode) Cooke CBS 125494 HQ897892 HQ897811 HQ897756
Fusarium sambucinum Fuckel CBS 14695 KM231015 KM231813 KM232381
Fusicolla matuoi (Hosoya & Tubaki) Gräfenhan & Seifert CBS 58178 HQ897858 KM231822 HQ897720
Geejayessia cicatricum (Berk.) Schroers CBS 125552 HQ728171 HQ728145 HQ728153
Macroconia leptosphaeriae (Niessl) Gräfenhan & Schroers CBS 100001 HQ897891 HQ897810 HQ897755
Microcera coccophila Desm. CBS 31034 HQ897843 HQ897794 HQ897705
Neocosmospora vasinfecta E.F. Sm. CBS 32554 KM231005 KM231803 KM232370
Stylonectria applanata Höhn. CBS 125489 HQ897875 HQ897805 HQ897739
Stylonectria carpini Gräfenhan DAOM 235819 HQ897909 HQ897823 HQ897773
Stylonectria norvegica Lechat, J. Fourn. & Nordén CBS 139239 – NR154415 –
Stylonectria purtonii (Grev.) Gräfenhan DAOM 235818 HQ897919 HQ897831 HQ897783
Stylonectria qilianshanensis Z.Q. Zeng & W.Y. Zhuang HMAS 255803 MT087289 a MT084413 MT087288
Stylonectria wegeliniana (Rehm) Gräfenhan, Voglmayr & Jaklitsch CBS 125490 HQ897890 KM231817 HQ897754
Trichoderma parareesei Atan., Jaklitsch, Komoń-Zel., C.P. Kubicek & Druzhin. CBS 125925 KJ665112 MH863773 HM182963

a Numbers in bold indicate the newly provided sequences.
Table 2.
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List of Trichoderma species, herbarium/strain numbers, and GenBank accession numbers of specimens used in this study.

Species Herbarium/strain numbers GenBank accession numbers
RPB2 TEF1
Hypocrella discoidea (Berk. & Broome) Sacc. BCC 8237 DQ452461 –
Hypocrella nectrioides Thaxt.
 GJS 8910 DQ522448 –
Trichoderma alutaceum Jaklitsch CBS 120535 FJ179600 FJ179567
CBS 33269 FJ179601 FJ179568
Trichoderma gansuanum Z.Q. Zeng & W.Y. Zhuang HMAS 279687 MT087287 a MT095060
Trichoderma gelatinosum P. Chaverri & Samuels CPK 1618 FJ179604 FJ179569
Trichoderma leucopus Jaklitsch CBS 122495 FJ179606 FJ179570
CBS 122499 FJ179605 FJ179571
Trichoderma lixii (Pat.) P. Chaverri CPK 1934 FJ179608 FJ179573
Trichoderma minutisporum Bissett CBS 121276 FJ179610 FJ179574
Trichoderma nybergianum (T. Ulvinen & H.L. Chamb.) Jaklitsch & Voglmayr CBS 122496 FJ179612 FJ179576
CBS 122500 FJ179611 FJ179575
Trichoderma parapiluliferum (B.S. Lu, Druzhin. & Samuels) Jaklitsch & Voglmayr CBS 20921 FJ179614 FJ179578
Trichoderma pezizoides (Berk. & Broome) Samuels, Jaklitsch & Voglmayr GJS 01257 EU248608 AY937438
Trichoderma piluliferum J. Webster & Rifai CBS 120927 KJ842159 FJ860674
Trichoderma placentula Jaklitsch CBS 120924 FJ179616 FJ179580
Trichoderma polysporum (Link) Rifai CPK 3131 FJ860558 FJ860661
Trichoderma poronioideum (Möller) Samuels GJS 01203 – KP109823
Trichoderma seppoi Jaklitsch CBS 122497 FJ179618 FJ179582
CBS 122498 FJ179617 FJ179581
Trichoderma strictipile Bissett CPK 1601 FJ860594 FJ860704

a Numbers in bold indicate the newly provided sequences.

Results

To determine taxonomic positions of the Hypomyces collections, sequences of ITS and 28S rDNA were searched against the NCBI GenBank database using BLASTN. Sequence comparisons showed that HMAS 247843 shares 99% sequence similarity with H. tremellicola, which confirmed its taxonomic position in the genus.

To place the Stylonectria specime n within a phylogenetic context, sequences of ACL1, ITS, and RPB2 regions from 15 species of the genus and relatives were analyzed using BI and MP methods. Clonostachys rosea (Preuss) Mussat and Trichoderma parareesei Atan., Jaklitsch, Komoń-Zel., C.P. Kubicek & Druzhin. were used as outgroup taxa. The partition homogeneity test (PHT) (P = 0.01) indicated that the individual partitions were not highly incongruent (Cunningham 1997), the three loci were thus combined for phylogenetic analyses. The combined datasets include 2197 characters, of which 964 were constant, 310 were variable and parsimony-uninformative and 923 were parsimony-informative. The MP analysis resulted in two most parsimonious trees (tree length = 4081, CI = 0.5285, HI = 0.4715, RI = 0.4459, RCI = 0.2357) with similar topology. The final matrix was deposited in TreeBASE with accession no. S25189. The BI tree is shown in Figure 1. The MP tree is similar to that of the BI tree in topology. HMAS 255803 was associated with other Stylonectria species forming a highly supported monophyletic group (BIPP/MPBP = 100%/100%), which confirmed its taxonomic position in the genus.

Figure 1. 

BI tree generated based on the combined datasets of ACL1, ITS and RPB2 sequences of Stylonectria and its relatives. Supporting values showing at branches: BIPP (left) and MPBP (right). BIPP greater than 90% and MPBP greater than 70% are shown at the nodes.

To place the Trichoderma collections with clavate fruit bodies within a phylogenetic context, the sequences of RPB2 and TEF1 from 15 species of the genus were analyzed using BI and MP. Hypocrella discoidea (Berk. & Broome) Sacc. and Hypocrella nectrioides Thaxt. were used as outgroup taxa. The PHT (P = 0.01) indicated that the individual partitions were not highly incongruent (Cunningham 1997), the two loci were thus combined for phylogenetic analyses. The combined datasets include 1436 characters, of which 688 were constant, 167 were variable and parsimony-uninformative and 581 were parsimony-informative. The MP analysis resulted in one most parsimonious tree (tree length =1828, CI = 0.6510, HI = 0.3490, RI = 0.6706, RCI = 0.4366). The final matrix was deposited in TreeBASE with accession no. S25188. The BI tree is shown in Figure 2, which is similar to the MP tree in topology. Among the investigated species, HMAS 279687 was distinct from but associated with other Trichoderma species forming a highly supported monophyletic group (BIPP/MPBP = 100%/100%), which confirmed its taxonomic position.

Figure 2. 

BI tree generated based on the combined datasets of RPB2 and TEF1 sequences of Trichoderma species. Supporting values showing at branches: BIPP (left) and MPBP (right). BIPP greater than 90% and MPBP greater than 70% are shown at the nodes.

Taxonomy

Stylonectria qilianshanensis Z.Q. Zeng & W.Y. Zhuang, sp. nov.

Fungal Names: FN 570729
Figure 3

Holotype

China. Gansu Province, Wuwei, Chashugou, on decayed fruitbodies of an ascomycete on bark of Picea asperata, 26 August 2018, Z.Q. Zeng, X.C. Wang & H.D. Zheng 12155 (HMAS 255803).

Paratypes

China. Zhangye, Longchanghe, on decayed fruitbodies of an ascomycete on bark of Picea asperata, 24 August 2018, Z.Q. Zeng, X.C. Wang & H.D. Zheng 12016, 12017 (HMAS 255804, 255805); Kangle, on decayed fruitbodies of an ascomycete on bark of Picea asperata, 24 August 2018, Z.Q. Zeng, X.C. Wang & H.D. Zheng 12035, 12036, 12037, 12038 (HMAS 255806, 255807, 255808, 255809); Shandan, Yanzhishan, on decayed fruitbodies of an ascomycete on bark of Picea asperata, 25 August 2018, H.D. Zheng, X.C. Wang & Z.Q. Zeng 12082 (HMAS 255810); Yanzhishan, on decayed fruitbodies of an ascomycete on bark of Picea asperata, 25 August 2018, Z.Q. Zeng, X.C. Wang & H.D. Zheng 12086, 12087, 12088, 12089, 12090 (HMAS 255811, 255812, 255813, 255814, 255815); Wuwei, Chashugou, on decayed fruitbodies of an ascomycete on bark of Picea asperata, 26 August 2018, Z.Q. Zeng, X.C. Wang & H.D. Zheng 12148, 12153, 12156, 12158 (HMAS 255816, 255817, 255818, 279708); Tianzhu, Kelacun, on decayed fruitbodies of an ascomycete on bark of Picea asperata, 27 August 2018, Z.Q. Zeng, X.C. Wang & H.D. Zheng 12229 (HMAS 279709); Haxi, on decayed fruitbodies of an ascomycete on bark of Picea asperata, 28 August 2018, Z.Q. Zeng, X.C. Wang & H.D. Zheng 12276, 12278 (HMAS 255819, 255820).

Etymology

The specific epithet refers to the type locality.

Description

Perithecia gregarious, up to 30 in a group, parasitic on decayed fruitbodies of an ascomycete, deep red to dark red, turning black red in 3% KOH and light yellow in 100% LA, subglobose to globose, not becoming cupulate upon drying, (216–)255–344 × (186–)206–304 μm (n = 12), apex broadly discoid, flattened, 50–70 μm high, 160–220 μm in diameter, slightly constricted below, with a tiny papilla. Perithecial wall of two layers, 25–38 μm thick, outer layer 20–31 μm, of textura angularis, cells 5–8 × 2–4 μm, walls 0.8–1.0 μm thick; inner layer 5–7 μm, of textura prismatica, cells 8–12.5 × 3–5 μm, walls 0.5–0.8 μm thick. Asci clavate, with an apical ring, 8-spored, 55–88 × 5–8(–10) μm. Ascospores ellipsoidal, ends rounded, 1-septate, light brown, smooth, uniseriate, 10–13 × 5–5.5 μm. Asexual state unknown.

Distribution

China.

Notes

Stylonectria qilianshanensis is morphologically similar to S. wegeliniana in having the perithecia with a broad, discoid apex, clavate asci with an apical ring, and ellipsoidal ascospores with rounded ends (Petch 1938). However, S. qilianshanensis differs in having smaller asci [55–88 × 5–8(–10) μm vs 90–100 × 9–10 μm] and ascospores (10–13 × 5–5.5 μm vs 10–18 × 6–9 μm) (Petch 1938). Sequence comparisons indicate that ITS of S. qilianshanensis differs from that of S. wegeliniana by 20 bp in a total length of 576 bp; ACL1 and RPB2 of the former differ from those of the latter by 84 bp and 38 bp, respectively among 722 bp and of 869 bp in length. The asexual state of the fungus remains unknown until culture is available.

Figure 3. 

Stylonectria qilianshanensis a–c ascomata on natural substrate d–f median section through perithecium g–l ascus with ascospores m–q ascospores. From HMAS 255803. Scale bars: 1 mm (a–c); 50 μm (d–f); 10 μm (g–q).

Trichoderma gansuanum Z.Q. Zeng & W.Y. Zhuang, sp. nov.

Fungal Names: FN 570730
Figure 4

Holotype

China. Gansu Province, Shandan, Yanzhishan, on mossy humus, 25 August 2018, Z.Q. Zeng, H.D. Zheng & X.C. Wang 12100 (HMAS 279687).

Paratypes

China. Gansu Province, Shandan, Yanzhishan, on mossy humus, 25 August 2018, Z.Q. Zeng, H.D. Zheng 12043, 12044 (HMAS 279684, 279685), on mossy humus, 25 August 2018, X.Z. Liu & Z.Q. Zeng 12045 (HMAS 279686); Wuwei, Chashugou, on mossy humus, 26 August 2018, Z.Q. Zeng, H.D. Zheng & X.C. Wang 12104, 12105 (HMAS 279688, 279689); Wuwei, Xiama, on mossy humus, 26 August 2018, Z.Q. Zeng, H.D. Zheng & X.C. Wang 12162, 12163 (HMAS 279690, 279691).

Etymology

The specific epithet refers to the type locality.

Description

Stromata simple, rare dichotomously branched, clavate, 20–54 mm long. Fertile part clavate, 5–18 mm long, 1.2–4 mm wide at apex, only slightly broader than stipe, distinctly laterally compressed or longitudinally furrowed, gradually tapered downwards, reddish brown to brownish orange, KOH+; sterile part 15–36 mm long, 1–3 mm wide, beige to cream KOH+. Stromatal surface slight tuberculate from papillate perithecial elevations. Ostiolar openings visible, 30–58 μm high. In section, cortical tissue of textura angularis,15–35 μm thick, cells hyaline to light yellow, 5–15 × 2–3 μm; subcortical tissue of textura angularis, 8–28 μm thick, cell hyaline to light yellow, 8–15 × 3–5 μm; subperithecial tissue of textura epidermoidea, rare textura angularis, cells hyaline to light yellow. Perithecia globose to subglobose, 196–206 × 167–235 μm; peridium 8–18 μm thick at flanks, 9–20 μm thick at the base. Papilla prominent, blunt or truncate, brown, 15–35 μm high, 18–43 μm wide at the base. Asci subcylindrical, 50–80 × 3–4 μm. Part-ascospores green, spinulose, dimorphic, distal cells globose, rarely ellipsoidal, 2.5–4 × 2.5–4 μm; proximal cells ellipsoidal to oblong, 3–5.5 × 2.5–3 μm. Asexual state unknown.

Figure 4. 

Trichoderma gansuanum a fresh stroma b dry stroma c stromatal surface d color of stroma after rehydration e median section through stromata f cortical tissue in section g subperithecial tissues in section h perithecia in section i ascus with ascospores j–m ascus with part-ascospores a from HMAS 279684, b–m from HMAS 279687. Scale bars: 1 cm (a); 1 mm (b–d); 50 μm (e–f); 10 μm (g–m).

Distribution

China.

Notes

Among the known stipitate species of Trichoderma, T. gansuanum resembles T. nybergianum in habitat and having simple, clavate, erect stromata, cylindrical asci and disarticulating ascospores (Chamberlain et al. 2004). Trichoderma gansuanum differs from the latter in shorter ascomatal stipe (20–54 mm vs 22–220 mm long), smaller perithecia (196–206 × 167–235 μm vs 180–450 × 65–315 µm), asci (50–80 × 3–4 μm vs 63–130 × 3.2–7.5 μm), and part-ascospores (distal: 2.5–4 × 2.5–4 μm vs 3–6 × 3–4.5 μm; proximal: 3–5.5 × 2.5–3 μm vs 3–6 × 2.5–4.5 μm) (Chamberlain et al. 2004). Sequence comparisons reveal that there are 25 bp and 16 bp divergences between the two species in the regions of RPB2 and TEF1. Both morphological and molecular data support distinguish them at the species level. The asexual state of T. gansuanum remains unknown until a culture is available.

Hypomyces cervinigenus Rogerson & Simms, Mycologia 63: 418, 1971.

Specimens examined

China, Gansu Province, Wuwei, Tianzhu, on Helvella sp., 26 August 2018, X.C. Wang, Z.Q. Zeng & H.D. Zheng 12103 (HMAS 279612); Wuwei, Tianzhu, on Helvella sp., 27 August 2018, Z.Q. Zeng, H.D. Zheng & X.C. Wang 12210, 12221, 12225 (HMAS 279667, 279668, 279669).

Distribution

Canada, China, United States.

Hypomyces chrysospermus Tul. & C. Tul., Annls Sci. Nat., Bot., Sér. 4, 13: 16, 1860.

Specimens examined

China, Gansu Province, Tianzhu, Zhuchacun, on Boletus sp., 27 August 2018, Z.Q. Zeng, H.D. Zheng & X.C. Wang 12206 (HMAS 279670); Tianzhu, Kelacun, on Boletus sp., 27 August 2018, Z.Q. Zeng, H.D. Zheng & X.C. Wang 12220 (HMAS 279671).

Distribution

Australia, Belgium, Canada, China, French, Japan, New Zealand, United Kingdom, United States.

Hypomyces lateritius (Fr.) Tul. & C. Tul., Annls Sci. Nat., Bot., Sér. 4, 13: 11, 1860.

Specimens examined

China, Gansu Province, Shandan, Yanzhishan, on Lactarius sp., 25 August 2018, Z.Q. Zeng, H.D. Zheng & X.C. Wang 12060, 12061 (HMAS 254608, 254609); Tianzhu, Kelacun, on Lactarius sp., 27 August 2018, Z.Q. Zeng, H.D. Zheng & X.C. Wang 12216, 12217, 12218, 12219 (HMAS 279672, 279673, 279674, 279675).

Distribution

Austria, Belgium, Canada, China, Czech, Denmark, Finland, France, Germany, Italy, Mexico, New Zealand, Russia, Sweden, United Kingdom, United States.

Hypomyces perniciosus Magnus, Bot. Ztg. 46: 394, 1888.

Specimen examined

China, Gansu Province, Tianzhu, Kelacun, on Agaricus sp., 27 August 2018, Z.Q. Zeng, H.D. Zheng & X.C. Wang 12222 (HMAS 279705).

Sequences

ITS (MT676396) and LSU (MT669266).

Distribution

China, France, Germany, United Kingdom.

Hypomyces rosellus (Alb. & Schwein.) Tul. & C. Tul., Annls Sci. Nat., Bot., Sér. 4 13: 12, 1860.

Specimen examined

China, Gansu Province, Shandan, Yanzhishan, on Lactarius sp., 25 August 2018, Z.Q. Zeng, H.D. Zheng & X.C. Wang 12056 (HMAS 279706).

Sequences

ITS (MT676395).

Distribution

China, Estonia, Poland, Ukraine, United States.

Hypomyces stephanomatis Rogerson & Samuels, Mycologia 77: 775, 1985.

Specimens examined

China, Gansu Province, Shandan, Yanzhishan, on Humaria sp., 25 August 2018, Z.Q. Zeng, H.D. Zheng & X.C. Wang 12063 (HMAS 279676); Tianzhu, Zhuchacun, on Humaria sp., 27 August 2018, Z.Q. Zeng, H.D. Zheng & X.C. Wang 12205, 12207, 12208, 12209, 12211 (HMAS 279677, 279678, 279679, 279680, 279681); Tianzhu, Kelacun, on Humaria sp., 27 August 2018, Z.Q. Zeng, H.D. Zheng & X.C. Wang 12223, 12226 (HMAS 279682, 279683).

Distribution

Canada, China, Germany, United States.

Hypomyces tremellicola (Ellis & Everh.) Rogerson, Mem. N. Y. Bot. Gdn. 26(3): 20, 1976.

Specimen examined

China, Gansu Province, Zhangye, Minyue, on (?) Agaricus sp., August 2018, C.H. Dong & S.J. Li 12287 (HMAS 247843).

Sequences

ITS (MT084414) and LSU (MT078664).

Description

On MEA, colony radius 33 mm after 7d at 25 °C, velvet, surface white, reverse light brown; aerial hyphae white. On PDA, colony radius 20 mm after 7d at 25 °C, floccose, surface grey white, reverse light sienna; aerial hyphae white. Simple branches of aerial hyphae terminating in 1–2-verticillate conidiophores with terminal whorl of 2–4 phialides. Phialides subulate, tapering towards apex, smooth, 8–25 × 1.5–2 μm. Conidia ellipsoidal to rod-shaped, aseptate, hyaline, smooth, 2.5–8 × 1–3 μm. Chlamydospores globose, hyaline, smooth, 5–8 μm in diameter, rare ellipsoidal, 6–12 × 5–10 μm, formed singly or in chains in intercalary position.

Distribution

Canada, China, Germany, New Zealand, The Netherlands, United States, Venezuela.

Notes

Hypomyces tremellicola is a new record for China. This species was originally described as Hypocrea tremellicola Ellis & Everh. (Ellis and Everhart 1892), and later transferred to Hypocreopsis P. Karst. (Seaver 1910) and Nectriopsis Maire (Gams and Zaayen 1982). Samuels (1976) redescribed the species and assigned it to Hypomyces. It usually grows on Crepidotus spp., and less frequently on Polyporus spp. and Pleurotus spp. The shape and size of conidia and chlamydospores of the Chinese material match well with the description provided by Zare and Gams (2019). Sequence comparisons showed that 4 bp and 1 bp divergences existed in ITS and 28S rDNA between the Chinese material (HMAS 247843) and a collection from Germany (CBS 441.65). We treat them as infraspecific variations.

Nectria asiatica Hirooka, Rossman & P. Chaverri, Stud. Mycol. 68: 44, 2011.

Specimen examined

China, Gansu Province, Tianzhu, Zhuchacun, on rotten twigs, 27 August 2018, Z.Q. Zeng, H.D. Zheng & X.C. Wang 12214 (HMAS 254610).

Distribution

China, Japan.

Nectria berberidicola Hirooka, Lechat, Rossman & P. Chaverri, in Hirooka, Rossman, Samuels, Lechat & Chaverri, Stud. Mycol. 71: 48, 2012.

Specimens examined

China, Gansu Province, Shandan, Yanzhishan, on rotten twigs, 25 August 2018, Z.Q. Zeng, H.D. Zheng & X.C. Wang 12084 (HMAS 279707); Tianzhu, Zhuchacun, on rotten twigs of Berberis sp., 27 August 2018, Z.Q. Zeng, H.D. Zheng & X.C. Wang 12212 (HMAS 254611); Tianzhu, Haxia, on rotten twigs of Berberis sp., 28 August 2018, Z.Q. Zeng, H.D. Zheng & X.C. Wang 12279, 12280, 12281 (HMAS 254612, 254613, 255801).

Sequences

ITS from HMAS 254613 (MT676394).

Distribution

China, France.

Nectria nigrescens Cooke, Grevillea 7(no. 42): 50, 1878.

Specimen examined

China, Gansu Province, Shandan, Yanzhishan, on rotten twigs of broadleaf tree, 25 August 2018, Z.Q. Zeng, H.D. Zheng & X.C. Wang 12085 (HMAS 255802).

Sequences

ITS (MT676393) and LSU (MT669265).

Distribution

Canada, China, France, Germany, United Kingdom, United States.

Thelonectria discophora (Mont.) P. Chaverri & C. Salgado sensu lato, in Chaverri, Salgado, Hirooka, Rossman & Samuels, Stud. Mycol. 68: 76, 2011.

Specimen examined

China, Gansu Province, Wuwei, Chashugou, on rotten twigs, 26 August 2018, Z.Q. Zeng, H.D. Zheng & X.C. Wang 12149 (HMAS 279710).

Distribution

Chile, China, United Kingdom.

Thyronectria rosellinii (Carestia) Jaklitsch & Voglmayr, Persoonia 33: 204, 2014.

Specimens examined

China, Gansu Province, Wuwei, Chashugou, on rotten twigs, 26 August 2018, Z.Q. Zeng, H.D. Zheng & X.C. Wang 12150, 12151, 12154 (HMAS 279711, 255821, 255822).

Distribution

Canada, China, France, Germany, Italy, Japan, United States.

Thyronectria zangii (Z.Q. Zeng & W.Y. Zhuang) Voglmayr & Jaklitsch, in Voglmayr, Akulov & Jaklitsch, Mycol. Progr. 15: 934, 2016.

Specimens examined

China, Gansu Province, Wuwei, Chengshanqizu, on rotten twigs of Populus sp., 28 August 2018, Z.Q. Zeng, H.D. Zheng & X.C. Wang 12285, 12286 (HMAS 255823, 255824).

Sequences

ITS from HMAS 255824 (MT676392).

Distribution

China.

Trichoderma alutaceum Jaklitsch, Fungal Diversity 48: 69, 2011.

Specimen examined

China, Gansu Province, Zhangye, Dayekou, on mossy ground under Picea asperata, 3 September 1958, Q.M. Ma 890 (HMAS 23955).

Distribution

Austria, China, Denmark, Finland, Germany, Japan, Sweden, United Kingdom, United States.

Trichoderma paraviridescens Jaklitsch, Samuels & Voglmayr, Persoonia 31: 128, 2013.

Specimen examined

China, Gansu Province, Tianzhu, Haxi, on rotten twigs, 28 August 2018, Z.Q. Zeng, H.D. Zheng & X.C. Wang 12282 (HMAS 255825).

Sequences

ITS (MT676391) and TEF1 (MT674562).

Distribution

Austria, China, Colombia, France, Germany, Greece, Iran, Italy, Japan, Mexico, Spain, Switzerland, United States.

Trichoderma virens (J.H. Mill., Giddens & A.A. Foster) Arx, Beih. Nova Hedwigia 87: 288, 1987.

Specimen examined

China, Gansu Province, Yongchang, in soil, April 2016, K. Chen TC896.

Distribution

China, United States.

Discussion

The genus Stylonectria, typified by S. applanata, was established as a monotypic genus by Höhnel (1915). It was included in Nectria by Booth (1959) and then treated as a synonym of Cosmospora Rabenh. by Rossman et al. (1999). Gräfenhan et al. (2011) resurrected the generic name Stylonectria and accepted four species. Lechat et al. (2015) described S. norvegica Lechat, J. Fourn. & Nordén from Norway. The morphological features of S. qilianshanensis, such as red perithecia growing on other ascomycetes, with flattened discoid apices, and two-layered wall bearing thick-walled outer cells, fit well the generic concept defined by Gräfenhan et al. (2011), which was confirmed by sequence analyses of the ACL1, ITS, and RPB2 regions. Stylonectria qilianshanensis is associated with S. wegeliniana (BIPP/MPBP = 100%/76%). Stylonectria purtonii [as Cosmospora purtonii (Grev.) Rossman & Samuels] on carbonaceous pyrenomycetes was the only species of the genus reported from China (Zhang and Zhuang 2003).

For some nectriaceous fungi, substrate type was considered of taxonomic importance (Zeng and Zhuang 2016). Species of Stylonectria are fungicolous and sometimes host-specific. For example, S. applanata is known only from Melogramma bulliardii Tul. & C. Tul. on Corylus avellana, S. carpini is restricted to a pyrenomycete on Carpinus, and S. wegeliniana colonizes solely on Hapalocystis bicaudata Fuckel on Ulmus glabra (Gräfenhan et al. 2011; Lechat et al. 2015). Similarly, S. qilianshanensis occurs on decayed fruit bodies of an ascomycete on the bark of Picea asperata. However, S. norvegica and S. purtonii have slightly wider host ranges. The former occurs on pyrenomycetes on Betulaceae and Fagaceae, while the latter is found on pyrenomycetes on coniferous trees. Stylonectria is currently a poorly known genus. Further investigations may provide useful information about its selection of hosts or substrates.

Gross morphology like stroma, ascospore features and asexual states were stressed for generic delimitations of the hypocrealean fungi. Species of Hypocreaceae possessing clavate to cylindrical, fleshy, bright-colored stromata were previously accommodated in Podostroma P. Karst. (Karsten 1892; Rossman et al. 1999). The accumulated molecular evidence argued that shape of fruit body is not phylogenetically distinctive within genus. Chamberlain et al. (2004) then synonymized Podostroma with Hypocrea Fr. (= Trichoderma). The diagnostic characteristics for Trichoderma include substrate, fruit body gross morphology, anatomy, extent of fertile region, surface pigmentation of stromata, and ascospore shape, size and ornamentation (Chamberlain et al. 2004). The genera Aphysiostroma Barrasa, A.T. Martinez & G. Moreno, Pseudohypocrea Yoshim. Doi, and Sarawakus Lloyd, which possess gliocladium-, trichoderma- and verticillim-like asexual states, were also synonymized with Trichoderma (Jaklitsch et al. 2014; Jaklitsch and Voglmayr 2015; Zeng and Zhuang 2017). The taxonomic position of T. gansuanum is confirmed by the combined sequence analyses of RPB2 and TEF1 regions and morphological characteristics, such as the stipitate, clavate, upright stromata, cylindrical asci, and disarticulating ascospores. A few stipitate species of Trichoderma have not been cultured or linked to asexual states (Chamberlain et al. 2004). Knowing the whole fungus is surely our future goal.

Acknowledgements

The authors thank Prof. Cai-Hong Dong and Drs Xin-Zhan Liu and Jun-Min Liang of Institute of Microbiology, CAS, for providing strain or collecting specimens jointly for this study. This work was supported by the National Natural Science Foundation of China (no. 31750001, 31870012, 31570018), Frontier Key Program of Chinese Academy of Sciences (no. QYZDY-SSW-SMC029) and Ecological restoration in the habitats of rare and endangered plants and animals as well as fungi protection and utilization in the Qilian Mountains (2018–20).

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