Morpho-molecular characterisation of Arecophila, with A.australis and A.clypeata sp. nov. and A.miscanthi comb. nov.

Three arecophila-like fungal samples were collected on dead culms of gramineous plants in China. Morphological studies of our new collections and the herbarium specimen of Arecophilagulubiicola (generic type) were conducted and the morphological affinity of our new collections with Arecophila was confirmed. Maximum likelihood and Bayesian analyses using combined ITS, LSU, rpb2 and β-tubulin data from our collections revealed the phylogeny of Cainiaceae. The monospecific genus Alishanica (type species Al.miscanthi), which had been accepted in Cainiaceae, is revisited and synonymised under Arecophila. Based on morphology and phylogeny, Arecophilaaustralis sp. nov. and A.clypeata sp. nov. are introduced as new species, while A.miscanthi is a new record for China. All the new collections are illustrated and described. Qi Rui Li, Xu Zhang, Yan Lin, Milan C. Samarakoon, Kevin David Hyde, Xiang Chun Shen, Wan Qing Liao, Anuruddha Karunarathna, Si Han Long, Ying Qian Kang, Ji Chuan Kang.

Introduction

The current study is a part of a series of papers on () from China (Long et al. 2019; Xie et al. 2019, 2020; Pi et al. 2020). K.D. Hyde, which is typified by K.D. Hyde, was introduced by Hyde (1996) with five species. is characterised by immersed, subglobose to lenticular ascomata, peridium with textura angularis cells, non- or poorly-developed clypeus, asci with a wedge-shaped, apical ring, J+ in Melzer’s reagent and 2-celled, brown ascospores with wall striations, surrounded by a mucilaginous sheath. Thanks to subsequently undertaken morphological studies of holotypes, several species have been transferred to from genera such as Ces. & De Not., Arx & E. Müll., Fuckel and Ehrenb. ex Lév. (Hyde 1996; Umali et al. 1999; Wang et al. 2004).

Currently, there are 15 epithets in Index Fungorum (http://www.indexfungorum.org/Names/Names.asp, May 2021), which have been introduced, based on morphology and lack sequence data (e.g. Hyde 1996; Umali et al. 1999; Wang et al. 2004). After searching for in NCBI, there were only five hits of LSU, SSU and metagenomic sequences of and sp. HKUCC 6487 in GenBank.

was introduced as a genus of (Hyde 1996), based on its unitunicate, cylindrical asci with a J+ apical ring and brown, 2-celled ascospores. Kang et al. (1999) reviewed the genus and accepted it in and the occurrence on monocotyledons (palms and bamboo). The single and combined molecular analyses of LSU and SSU genes resulted in grouping with in (Smith et al. 2003). Based on analyses of partial LSU gene sequences, the generic placement of within the has been verified (Jeewon et al. 2003; Senanayake et al. 2015; Hyde et al. 2020; Wijayawardene et al. 2020). However, the available molecular data do not provide strong evidence of the phylogenetic affinity of and related taxa.

During our continuous collecting of xylarialean taxa in China, we found some specimens that share a morphology resembling . In this paper, two new species and a new record of are provided with descriptions and illustrations. Furthermore, is synonymised under , based on morphology and phylogeny.

Materials and methods

Collection, isolation and morphology

Fresh samples were collected in Guizhou and Yunnan Provinces in China during the rainy season and taken to the laboratory in paper bags. Single-spore isolations were obtained following the method described in Chomnunti et al. (2014). The cultures on potato dextrose agar () were transferred to 2 ml screw cap centrifuge tubes filled with 10% glycerol and sterile water to deposit at –20 °C and 4 °C, respectively. Herbarium materials were deposited at the Herbarium of Guizhou Agricultural College () and the Herbarium of Guizhou University (). Cultures were deposited at the Culture Collection of Guizhou University ().

The morphological examination of fresh and herbarium specimens was carried out as described by Hyde (1996). Macro-morphological characters were examined and photographed using a digital camera (Canon 700D) fitted to the Olympus SZ61 stereomicroscope. Materials mounted in water, Melzer’s reagent and Indian ink were examined. At least 30 ascospores, 30 asci and 20 apical rings were measured for each taxa with Tarosoft (R) Image Frame Work (v. 0.9.0.7) and photographed using a digital camera (Nikon 700D) fitted to a light microscope (Nikon Ni).

DNA extraction, polymerase chain reaction (PCR) amplification and sequencing

Total genomic DNA was extracted from fresh mycelium scraped off from pure cultures with the BIOMIGA fungus genomic DNA extraction kit (GD2416) (Wijayawardene et al. 2013) following the manufacturer’s instructions. Primers, LR0R/LR5 (Vilgalys and Hester 1990), ITS4/ITS5 (White et al. 1990), RPB2-5F/RPB2-7cR (Liu et al. 1999), Bt2a/Bt2b and ACT-512F/ACT-783R (Hsieh et al. 2005) were used for amplifying partial large-subunit ribosomal RNA (LSU), internal transcribed spacer (ITS), partial second-largest subunit of the RNA polymerase II (rpb2), β-tubulin (tub) and α-actin gene (Hsieh et al. 2005). The amplification conditions were carried out according to Liu et al. (2011) and Hsieh et al. (2005). Amplified products were examined and sent to the sequencing company, Sangon Biotech, Shanghai, China. The obtained sequences were checked, assembled and uploaded to GenBank.

Sequence alignment and phylogenetic analyses

Following the NCBI BLAST results and literature (e.g. Jeewon et al. 2003; Senanayake et al. 2015), relevant sequences from all families of were downloaded from GenBank for the phylogenetic analyses (Table 1). Sequences of each segment were aligned using MAFFT (http://mafft.cbrc.jp/alignment/server/index.html, Katoh and Standley 2019) and improved manually in BioEdit 7.2.3 (Hall 1999). The combined alignment of ITS, LSU, rpb2 and β-tubulin was concatenated from individual datasets. Ambiguously aligned areas of each gene region were excluded and gaps were treated as missing data. The ALTER (http://sing.ei.uvigo.es/ALTER/) phylogeny website tool was used to obtain the phylip file for RAxML analysis and the nexus file for Bayesian analysis (Glez-Peña et al. 2010). Phylogenetic trees were visualised using FigTree v.1.4.0. and processed using Adobe Photoshop CS6 software (Adobe Systems, USA). The alignment for the tree in this paper was uploaded on the website (https://treebase.org/) with submission ID 26613.

Table 1.

Sequences used for phylogenetic analyses in this study.

Species	Strain number	Status	GenBank accession numbers				References
			ITS	LSU	rpb2	β-tubulin
Achaetomiummacrosporum	CBS 532.94	–	KX976574	KX976699	KX976797	KX976915	Wang et al. (2016)
Amphibambusabambusicola	MFLUCC 11-0617	HT	KP744433	KP744474	N/A	N/A	Senanayake et al. (2015)
Amphisphaeriaacericola	MFLU 16-2479	HT	NR_171945	MK640424	N/A	N/A	Senanayake et al. (2019, submitted directly)
Amphisphaeriathailandica	MFLU 18-0794	HT	NR_168783	NG_068588	MK033640	MK033639	Samarakoon et al. (2019)
Amphisphaeriaumbrina	AFTOL-ID 1229	AF009805	N/A	FJ176863	FJ238348	N/A	Schoch (2008, submitted directly)
Apiosporabambusae	ICMP 6889	–	N/A	DQ368630	DQ368649	N/A	Tang et al. (2007)
Apiosporahyphopodii	MFLUCC 15-0003	HT	KR069110	KY356093	N/A	N/A	Dai et al. (2016)
Apiosporasetosa	ICMP 4207	–	N/A	DQ368631	DQ368650	DQ368620	Tang et al. (2007)
Apiosporayunnana	MFLUCC 15-0002	HT	KU940147	NG_057104	KU940177	MK291950	Dai et al. (2017)
Arecophilaaustralis	GZUCC0112	HT	MT742126	MT742133	N/A	MT741734	This study
Arecophilaaustralis	GZUCC0124	–	MT742125	MT742132	N/A	N/A	This study
Arecophilabambusae	HKUCC 4794	–	N/A	AF452038	N/A	N/A	Kang et al. (1999)
Arecophilaclypeata	GZUCC0110	HT	MT742129	MT742136	MT741732	N/A	This study
Arecophilaclypeata	GZUCC0127	–	MT742128	MT742135	N/A	N/A	This study
Arecophilamiscanthi	GZUCC0122	–	MT742127	MT742134	N/A	N/A	This study
Arecophilamiscanthi	MFLU 19-2333	HT	NR_171235	MK503827	N/A	N/A	Hyde et al. (2020)
Arecophila sp.	HKUCC 6487	–	N/A	AF452039	N/A	N/A	Jeewon et al. (2003)
Apiosporayunnana	MFLUCC 15-0002	HT	KU940147	NG_057104	KU940177	MK291950	Dai et al. (2017)
Atrotorquataspartii	MFLUCC 13-0444	HT	N/A	KP325443	N/A	N/A	Thambugala et al. (2015)
Bagadiellalunata	CBS 124762	HT	NR_132832	NG_058637	N/A	N/A	Cheewangkoon et al. (2009)
Barrmaeliarappazii	Cr2 = CBS 142771	HT	MF488989	MF488989	MF488998	MF489017	Voglmayr et al. (2018)
Barrmaeliarhamnicola	BR = CBS 142772	HT	MF488990	MF488990	MF488999	MF489018	Voglmayr et al. (2018)
Bartaliniapondoensis	CMW 31067	–	MH863602	MH875078	MH554904	MH554663	Vu et al. (2019)
Beltraniapseudorhombica	CBS 138003	HT	MH554124	NG_058667	MH555032	N/A	Liu et al. (2019)
Beltraniarhombica	CBS 123.58	T	MH857718	MH868082	MH554899	MH704631	Vu et al. (2019)
Beltraniopsislongiconidiophora	MFLUCC 17-2139	HT	NR_158353	NG_066200	N/A	N/A	Liu et al. (2017)
Biscogniauxianummularia	MUCL 51395	ET	KY610382	KT281894	KY624236	KX271241	Senanayake et al. (2015)
Cainiaanthoxanthis	MFLUCC 15-0539	HT	NR_138407	KR092777	N/A	N/A	Senanayake et al. (2015)
Cainiagraminis	CBS 136.62	–	MH858123	AF431949	N/A	N/A	Vu et al. (2019)
Cainiagraminis	MFLUCC 15-0540	–	KR092793	KR092781	N/A	N/A	Senanayake et al. (2015)
Camilleaobularia	ATCC 28093	–	KY610384	KY610429	KY624238	KX271243	Wendt et al. (2018)
Castanediellaacaciae	CBS 139896	HT	NR_137985	NG_067293	N/A	N/A	Crous et al. (2015)
Castanediellacouratarii	CBS 579.71	HT	NR_145250	NG_066249	N/A	N/A	Vu et al. (2019)
Castanediellaeucalypticola	CPC 26539	HT	KX228266	KX228317	N/A	KX228382	Crous et al. (2013)
Chaetomiumelatum	CBS 374.66	–	KC109758	KC109758	KF001820	KC109776	Wang et al. (2016)
Ciferriascoseafluctuatimura	MFLUCC 15-0541	HT	KR092789	KR092778	N/A	N/A	Senanayake et al. (2015)
Ciferriascosearectimura	MFLUCC 15-0542	HT	NR_153905	KR092776	N/A	N/A	Senanayake et al. (2015)
Clypeophysalosporalatitans	CBS 141463	ET	NR_153929	NG_058958	N/A	N/A	Giraldo et al. (2017)
Coniocessiamaxima	CBS 593.74	HT	NR_137751	MH878275	N/A	N/A	Vu et al. (2019)
Coniocessianodulisporioides	CBS 281.77	IT	MH861061	AJ875224	N/A	N/A	García et al. (2006)
Creosphaeriasassafras	STMA 14087	–	KY610411	KY610468	KY624265	KX271258	Wendt et al. (2018)
Cylindriumaeruginosum	CBS 693.83	–	KM231854	KM231734	KM232430	KM232124	Lombard et al (2014, submitted directly)
Cylindriumgrande	CBS 145655	HT	NR_165557	NG_068656	MK876481	MK876502	Crous et al. (2019)
Cylindriumpurgamentum	CPC 29580	HT	NR_155691	NG_067320	N/A	N/A	Koppel et al. (2017)
Daldiniaconcentrica	CBS 113277	–	AY616683	KT281895	KY624243	KC977274	Senanayake et al. (2015)
Delonicicolasiamense	MFLUCC 15-0670	HT	MF167586	NG_059172	MF158346	N/A	Perera et al. (2017)
Diatrypepalmicola	MFLUCC 11-0018	–	KP744439	KP744481	N/A	N/A	Liu et al. (2015)
Diatrypewhitmanensis	ATCC MYA-4417	–	FJ746656	FJ430587	N/A	N/A	Igo et al. (2009, direct submission)
Entosordariaperfidiosa	EPE = CBS 142773	ET	MF488993	MF488993	MF489003	MF489021	Voglmayr et al. (2018)
Entosordariaquercina	RQ = CBS 142774	HT	MF488994	MF488994	MF489004	MF489022	Voglmayr et al. (2018)
Eutypaflavovirens	MFLUCC 13-0625	–	KR092798	KR092774	N/A	N/A	Senanayake et al. (2015)
Eutypalaevata	CBS 291.87	–	HM164737	N/A	HM164805	HM164771	Trouillas and Gubler (2010)
Eutypalata	CBS 208.87	NT	MH862066	MH873755	KF453595	DQ006969	Vu et al. (2019)
Furfurellanigrescens	CBS 143622	HT	MK527844	MK527844	MK523275	MK523332	Voglmayr et al. (2019)
Furfurellastromatica	CBS 144409	HT	NR_164062	MK527846	MK523277	MK523334	Voglmayr et al. (2019)
Graphostromaplatystomum	AFTOL-ID 1249	HT	HG934115	DQ836906	DQ836893	HG934108	Zhang et al. (2006)
Hyponectriabuxi	UME 31430	–	-	AY083834	N/A	N/A	Smith et al. (2002, submitted directly)
Hypoxylonfragiforme	MUCL51264	ET	KM186294	KM186295	KM186296	KM186293	Daranagama et al. (2015)
Iodosphaeriahonghensis	MFLU 19-0719	HT	MK737501	MK722172	MK791287	N/A	Marasinghe et al. (2019)
Iodosphaeriatongrenensis	MFLU 15-0393	HT	KR095282	KR095283	N/A	N/A	Li et al. (2015)
Jackrogersellamultiformis	CBS 119016	ET	KC477234	KT281893	KY624290	KX271262	Wendt et al. (2018)
Kretzschmariadeusta	CBS 163.93	–	KC477237	KT281896	KY624227	KX271251	Senanayake et al. (2015)
Lepteutypafuckelii	CBS 140409	NT	NR_154123	KT949902	MK523280	MK523337	Jaklitsch et al. (2016)
Leptosilliapistaciae	CBS 128196	HT	NR_160064	MH798901	MH791334	MH791335	Voglmayr et al. (2019)
Leptosilliawienkampii	CBS 143630	ET	NR_164067	MK527865	MK523297	MK523353	Voglmayr et al. (2019)
Longiappendisporachromolaenae	MFLUCC 17-1485	HT	NR_169723	NG_068714	N/A	N/A	Mapook et al. (2020)
Lopadostomaamericanum	LG8	HT	KC774568	KC774568	KC774525	N/A	Jaklitsch et al. (2014)
Lopadostomadryophilum	LG21	ET	KC774570	KC774570	KC774526	MF489023	Jaklitsch et al. (2014)
Lopadostomafagi	LF1	HT	KC774575	KC774575	KC774531	N/A	Jaklitsch et al. (2014)
Lopadostomaquercicola	LG27	HT	KC774610	KC774610	KC774558	N/A	Jaklitsch et al. (2014)
Lopadostomaturgidum	LT2	ET	KC774618	KC774618	KC774563	MF489024	Jaklitsch et al. (2014)
Melogrammacampylosporum	MBU	–	JF440978	JF440978	N/A	N/A	Jaklitsch and Voglmayr (2012)
Neophysalosporaeucalypti	CBS 111123	–	KP031107	KP031109	N/A	N/A	Crous et al. (2014)
Neophysalosporaeucalypti	CBS 138864	HT	KP004462	MH878627	N/A	N/A	Crous et al. (2014)
Oxydothismetroxylicola	MFLUCC 15-0281	HT	KY206774	KY206763	KY206781	N/A	Konta et al. (2016)
Oxydothispalmicola	MFLUCC 15-0806	HT	KY206776	KY206765	KY206782	N/A	Konta et al. (2016)
Oxydothisphoenicis	MFLUCC 18-0269	HT	MK088065	MK088061	N/A	N/A	Hyde et al. (2020)
Phlogicylindriumuniforme	CBS 131312	HT	JQ044426	JQ044445	MH554910	MH704634	Crous et al. (2011)
Podosordariatulasnei	CBS 128.80	–	KT281902	KT281897	N/A	N/A	Senanayake et al. (2015)
Poroniapunctata	CBS 656.78	HT	KT281904	KY610496	KY624278	KX271281	Wendt et al. (2018)
Pseudomassariachondrospora	MFLUCC 15-0545	–	KR092790	KR092779	N/A	N/A	Senanayake et al. (2015)
Pseudomassariasepincoliformis	CBS 129022	–	JF440984	JF440984	N/A	N/A	Jaklitsch and Voglmayr (2012)
Pseudosporidesmiumknawiae	CBS 123529	HT	MH863299	MH874823	N/A	N/A	Crous et al. (2017, submitted directly)
Pseudosporidesmiumlambertiae	CBS 143169	HT	NR_156656	NG_058506	N/A	N/A	Crous et al. (2017)
Pseudotruncatellaarezzoensis	MFLUCC 14-0988	HT	NR_157489	NG_070426	N/A	N/A	Perera et al. (2018)
Pseudotruncatellabolusanthi	CBS 145532	HT	NR_165575	MK876448	N/A	N/A	Crous et al. (2019)
Robillardaroystoneae	CBS 115445	HT	NR_145251	NG_069287	MH554880	KR873317	Liu et al. (2019)
Sarcoxyloncompunctum	CBS 359.61	–	MH858083	KY610462	KY624230	KX271255	Wendt et al.(2018)
Seiridiummarginatum	CBS 140403	ET	NR_156602	MH554223	LT853149	MT853249	Liu et al. (2019)
Seynesiaerumpens	SMH 1291	–	N/A	AF279410	AY641073	N/A	Bhattacharya et al. (2000)
Sordariafimicola	CBS 723.96	–	MH862606	MH874231	DQ368647	DQ368618	Vu et al. (2019)
Sporocadusrotundatus	CBS 616.83	HT	NR_161091	NG_069584	MH554974	MH554737	Liu et al. (2019)
Subsessilaturbinata	MFLUCC 15-0831	HT	NR_148122	NG_059724	N/A	N/A	Lin et al. (2017)
Vialaeainsculpta	DAOM 240257	–	JX139726	JX139726	N/A	N/A	Hambleton et al. (2010, submitted directly)
Vialaeamangiferae	MFLUCC 12-0808	HT	NR_171903	NG_073594	N/A	N/A	Senanayake et al. (2021, submitted directly)
Vialaeaminutella	BRIP 56959	–	KC181926	KC181924	N/A	N/A	McTaggart et al. (2013)
Xyladictyochaetalusitanica	CBS 143502	–	MH107926	MH107972	N/A	MH108053	Crous et al. (2013)
Xylariahypoxylon	CBS 122620	ET	KY610407	KY610495	KY624231	KX271279	Wendt et al. (2018)
Xylariaobovata	MFLUCC 13-0115	–	KR049088	KR049089	N/A	N/A	Wendt et al. (2018)
Xylariapolymorpha	MUCL 49884	–	KY610408	KT281899	KY624288	KX271280	Wendt et al. (2018)

Note. Type specimens are labelled with HT (holotype), ET (epitype) and IT (isotype), T (Type). N/A: not available.

Maximum likelihood (ML) analysis was performed on the CIPRES Science Gateway v.3.3 (http://www.phylo.org/portal2; Miller et al. 2010) using RAxML v.8.2.8 as part of the ‘RAxML-HPC BlackBox’ tool (Stamatakis et al. 2008). All free model parameters were estimated by RAxML with ML estimates of 25 per-site rate categories. GTRGAMMA + I model was chosen for RAxML, based on the result of MrModeltest 2.2. The best-scoring tree was selected with a final likelihood value of –10720.566919.

A Bayesian analysis (BY) was performed using MrBayes v.3.2.2 (Ronquist et al. 2012). The best-fit model was selected with MrModeltest 2.2 (Nylander 2004). Posterior probabilities (PP) (Rannala and Yang 1996) were determined by Markov Chain Monte Carlo sampling (MCMC) (Ronquist and Huelsenbeck 2003). Six simultaneous Markov chains were initially run for 30 × 106 generations and for every 1000th generation, a tree was sampled (resulting in 30,000 total trees). The MCMC heated chain was set with a ‘temperature’ value of 0.15. All sampled topologies beneath the asymptote (20%) were discarded. The remaining 24,000 trees were used to calculate the posterior probability (PP) values in the majority rule consensus tree (Liu et al. 2011).

Results

Phylogenetic analyses

The resulted trees from ML and BY were similar in topology. is a monophyletic group (Fig. 1) with 100%/1.00 (PP/BS) support. species form two clades. Clade 1 consists of (≡ ), and , with high statistical support (100%/1.00 PP). In Clade 2, (HKUCC 4794) and sp. (HKUCC 6487) display a close relationship with .

Figure 1.

Phylogenetic tree, based on a combined ITS, LSU, rpb2 and β-tubulin gene dataset. Numbers close to each node represent Maximum Likelihood bootstrap values (≥ 75%) and Bayesian posterior probabilities (≥ 0.95). The hyphen (“–”) means a value lower than 75% (BS) or 0.95 (PP). New taxa are marked in red. Type materials are marked with T after the strains. The tree is rooted to (CBS 532.94), (CBS 374.66) and (CBS 723.96).

Taxonomy

K.D. Hyde, Nova Hedwigia 63(1–2): 82 (1996)

F28DD8D5-5910-5F1B-BF13-11C2653DC3B0

27653

≡

Sexual morph.

Ascomata immersed, raised, blackened areas on the host surface, a central erumpent, short, cone-shaped or umbilicate papilla, subglobose to lenticular in vertical section. present or not, comprising host cells and intracellular brown hyphae. Peridium comprising several layers of angular cells. Paraphyses hypha-like, filamentous, septate, hyaline. Asci 8-spored, unitunicate, cylindrical, with an apical ring bluing in Melzer’s reagent or not. Ascospores ellipsoidal, 2-celled, constricted at the septum, brown, with longitudinal striations or a verrucose wall and surrounded by a wide mucilaginous sheath (Hyde 1996).

Asexual morph.

Undetermined.

Q.R. Li, J.C. Kang & K.D. Hyde sp. nov.

BFDFCDD6-E7CD-5553-AB3D-217FDC12ED2B

836166

Fig. 2

Figure 2.

(holotype) A material B ascoma on the surface of host C section of ascoma D peridium E paraphyses F, G ascus apex with a J+, apical ring (stained in Melzer’s reagent) H–K asci with ascospores L–O ascospores surrounded by a wide mucilaginous sheath (O stained in India ink). Scale bars: 300 μm (B); 50 μm (C); 5 μm (D–O).

Diagnosis.

differs from similar species by its dimension of ascospores (22.5–29 × 8–11 µm) covered by striations and ascomata with a disc area surrounding the ostioles.

China, Guizhou Province, Guiyang City, Forest Park of Guiyang (), on dead culm of (Cav.) Steud., 15 March 2014, Q.R. Li, GZ58 (GZUH0112, , ex-type: GZUCC0112; GACP QR0152, ).

Additional sequences.

ACT: MT741737

Etymology.

In reference to the host, (Cav.) Steud. australis

Description.

Saprobic on dead culm of gramineous host. Sexual morph: Ascomata 420–560 × 290–380 µm (x̄= 495 × 325 µm, n = 10), immersed under a clypeus, solitary, slightly raised, blackened, dome-shaped areas, scattered or gregarious, globose to subglobose, with a central, erumpent, cone-shaped papilla in vertical section. black, comprising host cells and intracellular brown hyphae. papillate, black. Peridium 15–25 µm (x̄= 21 µm, n = 15) wide, comprising several layers, outer layer brown, thick-walled angular cells, inner layer hyaline. Paraphyses 3.3–5 μm (x̄= 3.5 µm, n = 15) wide, hyaline, unbranched, septate. Asci 140–230 × 15.5–24 µm (x̄= 183.5 × 19 µm, n = 30), 8-spored, unitunicate, long-cylindrical, short-pedicellate, apically rounded, with a 4–5 × 2.5–3 μm (x̄= 4.5 × 2.7 μm, n = 20), trapezoidal, J+, apical ring. Ascospores 22.5–29 × 8–11 µm (x̄= 25.5 × 9 µm, n = 30), overlapping uniseriate, 2-celled, light brown to brown, equilateral ellipsoidal, constricted at the septum, longitudinal with sulcate striations, along the entire spore length, surrounded by a mucilaginous sheath, lacking germ slits and appendages. Asexual morph: undetermined.

Culture characteristics.

Colonies on PDA, reached 3 cm diam. after one week at 25 °C, white, cottony, flat, low, dense, with slightly wavy margin.

Known distribution.

China

Additional material examined.

China, Guizhou Province, Guiyang City, Leigongshan National Nature Reserve (), on dead culm of an unidentified gramineous plant, 13 June 2015, Q.R. Li, GY67 (GACP QR0124, GZUH 0136; living cultures, GZUCC0124).

Notes.

resembles (Ellis & Martin) K.D. Hyde and K.D. Hyde (Hyde 1996). However, has white ring surrounding ostioles of ascomata, narrower ascospores (17–26 × 7–9.5 µm vs. 22.5–29 × 8–11 µm), smaller asci and apical ring (3.2 × 2.4 µm vs. 4.5 × 2.7 μm) compared to (Hyde 1996). differs from in lacking clypeus, ascospores covered by verrucose ornamentation and surrounding by a mucilaginous sheath attached at the poles. Molecular phylogeny, based on combined ITS, LSU, rpb2 and β-tubulin sequences, shows that clusters as a distinctive clade in (Clade 1). Based on its distinct morphology and phylogeny, is introduced as a new species. Here, we need to explain the name of . Although Index Fungorum (02/07/2022) shows that the current name of is (Ellis & G. Martin) K.D. Hyde & Aptroot, we have not found relevant literature. Hyde (1996) renamed Eltis & G. Martin and as synonyms of (Ellis & G. Martin) K.D. Hyde. was erected with the unitunicate asci with a blue-staining ring (Hyde 1996) which is clearly inconsistent with the morphological features of Sacc. Therefore, we still compare with the original description of in this article.

Q.R. Li, J.C. Kang & K.D. Hyde sp. nov.

B31AA394-95A5-5A80-9A0D-796D0C51AC6E

836167

Fig. 3

Figure 3.

(holotype) A material B ascomata on the surface of host C, D section of ascomata E peridium F, G ascus apex with a J+, apical ring (stained in Melzer’s reagent) H–K asci with ascospores L–O ascospores. Scale bars: 500 μm (B, C); 100 μm (D); 10 μm (E, H–K); 5 μm (F, G, L–O).

differs from similar species by its ascomata with clypeus and ascospores (18.5–22.5 × 6.5–9 µm).

China, Yunnan Province, Kunming City, Kunming Botanical Garden (), on dead culm of gramineous plant, 20 March 2014, Q.R. Li, kib21 (: GZUH0110; : GACP QR0173; ex-type living cultures: GZUCC0110).

In reference to the clypeus.

Saprobic on dead stem of a gramineous. Sexual morph: Ascomata 367–448 × 278–363 µm (x̄= 403 × 323 µm, n = 8), immersed under a black clypeus, solitary, slightly raised, dome-shaped areas, scattered or gregarious, subglobose to globose, with a central, erumpent, cone-shaped papilla, in vertical section. papillate on the centre, black. Peridium 15–30 µm (x̄= 25 µm, n = 10) wide, comprising several layers, outer layer brown, thick-walled angular cells, inner layer hyaline. Paraphyses 3–5 μm (x̄= 4 µm, n =15) wide, hyaline, unbranched, septate. Asci 180–245 × 10.5–14.5 µm (x̄= 215.5 × 12 µm, n=20), 8-spored, unitunicate, long-cylindrical, short-pedicellate, apically rounded, with a square-shaped, J+, apical ring, 3–4 × 3–4 μm. Ascospores 18.5–22.5 × 6.5–9 µm (x̄= 20.5 × 7.5 µm, n = 30), overlapping uniseriate, 2-celled, light brown to brown, equilateral ellipsoidal, constricted at the septum, longitudinal, sulcate along the entire spore length, faint, surrounded by a mucilaginous sheath, lacking germ slits and appendages. Asexual morph: undetermined.

Colonies on PDA, reached 3 cm diam. after one week at 25 °C, white, cottony, flat, low, dense, with slightly wavy margin; fructifications were not observed in culture.

China

China, Guizhou Province, Buyi and Miao Autonomous Prefecture in southern Guizhou Province, Maolan National Nature Reserve (), on dead culm of an unidentified gramineous plant, 12 June 2015, Q.R. Li, GZ120 (GACP QR0129; GZUH0127; living cultures, GZUCC0127).

ACT: MT741737

has long and weakly striate ascospores similar to (Rehm) Umali & K.D. Hyde, (Ellis & G. Martin) K.D. Hyde and (Hyde 1996; Umali et al. 1999). However, does not have a prominent clypeus and has longer and fusiform ascospores. differs from by the ascomata without a central papilla surrounded by a circle of white tissue, further in having ascospores with wide sheaths (Hyde 1996). is similar to which, however, has narrower ascospores (19–22.5 × 5.5–7 µm) covered by the strong striations and has ascomata without a central papilla surrounded by a black corolla protuberance (Umali et al. 1999).

K.D. Hyde, Nova Hedwigia 63(1–2): 91 (1996)

E533ED2A-0E76-55FA-A447-14E32DF15B0E

416041

Fig. 4

Figure 4.

(BRIP 23002a, holotype) A, B herbarium material with label C ascomata on the host D, E sections of ascomata F paraphyses G–J asci K peridium L, M wedge-shaped, J+ apical ring bluing in Melzer’s reagent N–Q ascospores. Scale bars: 50 μm (D, E); 5 μm (F–Q).

Saprobic on dead trunk of (Becc.) Becc. Sexual morph: Ascomata 290–400 × 140–190 μm (x̄ = 336 × 167 µm, n = 8), immersed under a clypeus, solitary or clustered, in vertical section, lenticular, with a central ostiole. raised, oval, blackened areas on the host surface, dome-shaped, well-developed and black. Peridium 25–35 μm wide, dense, compressed layers of brown-walled, angular cells, tightly adhered to the host tissues. Paraphyses 2–2.5 μm wide, filamentous, hyaline, septate, branched, tapering distally. Asci 107–145 × 11–13.5 μm (x̄ = 114.3 × 12.4 μm, n = 15), 8-spored, unitunicate, cylindrical, short-pedicellate, apically rounded, wedge-shaped, J+, subapical ring, 3–4 × 1–2 μm (x̄ =3.5 × 1.5 μm, n = 15). Ascospores 14.5–18.5 × 6–9 μm (x̄ = 17.4 × 6.5 μm, n = 25), overlapping uniseriate, ellipsoidal, brown, 2-celled, septate at the centre, constricted at the septum, longitudinal, sulcate striations along the entire spore length, surrounded by a mucilaginous sheath. Asexual morph: Undetermined.

Material examined.

Papua New Guinea, Central Province, , on dead trunk of (Becc.) Becc. (), May 1992, K.D. Hyde, (BRIP 23002a, ).

has deeply immersed, subglobose to lenticular ascomata with a small or lacking clypeus, cylindrical, short-pedicellate asci with a wedge-shaped, conical, apical ring and ellipsoidal, brown ascospores with wall striations and surrounded by a mucilaginous sheath (Hyde 1996). has been introduced as a monospecific genus with the type species Karun. et al. on dead sheaths of () from Taiwan (Hyde et al. 2020). We re-examined both and herbarium specimens and observed that they are congeneric. has characters that immersed ascoma under a clypeus, unitunicate, cylindrical asci with a J+ apical ring and brown, 2-celled ascospores with longitudinal wall striations and a mucilaginous sheath which are consistent with the generic characteristics of . The phylogeny of was mainly considered by the (HKUCC 4794) sequences (Hyde et al. 2020). However, HKUCC 4794 is not the type material of and cannot be used to represent . In our phylogeny, HKUCC 4794 forms a distinct clade (Fig. 1; Clade 2) from the representing the clade. Based on morphology and phylogeny, we synonymise under and is accepted as an species. Furthermore, needs to be recollected and provided with the phylogenetic affinity in future studies.

(Karun., C.H. Kuo & K.D. Hyde) Q.R Li & J.C. Kang comb. nov.

555587DE-0701-5A9E-B0BC-CD2BF216FBA8

839706

≡

Description

(MFLU 19-2333).Saprobic on dead sheaths of (). Sexual morph: Ascomata 272–277 × 283–296 µm (x̄ = 275 × 291.5 µm, n = 8), immersed beneath blackened aggregated clypeus of the surface of dead sheath, loosely aggregated or rarely solitary; dark brown to black, globose to subglobose, slightly depressed, uniloculate. Ostiole 92–110 μm long, 52–56 μm diameter (x̄ = 101 × 54 μm, n = 5), centrally erumpent, with periphyses, surrounded by distinct shiny black flanges, the tissue spreading down along the papilla. Peridium 51–60 μm wide, comprising 4–5 cell layers of thin-walled, brown cells of textura angularis, inwardly lighter. Paraphyses filamentous, distinctly septate, embedded in a hyaline gelatinous matrix. Asci 147–189 × 10–13 μm (x̄ = 167 × 11 μm, n = 30), 8-spored, unitunicate, cylindrical, short pedicellate, slightly truncate at the apex, with a wedge-shaped J+, subapical ring, 3.5–4 µm broad, 2–2.5 µm high. Ascospores 20–24 × 6–8 μm (x̄ = 22 × 7 μm, n = 40), overlapping, uniseriate, ellipsoidal, slightly tapering at the ends, equally 2-celled and guttulate at both cells, constricted at the septum, brown with striations, surrounded by a thick, hyaline mucilaginous sheath, subglobose, parallel to the margin of the spore. Asexual morph: Undetermined.

China, Taiwan, Chiayi Province, Ali Mountain, Kwang Hwa, on dead sheaths of (), 5 May 2018, A. Karunarathna, AKTW 44 (MFLU 19-2333, )

Additional material.

China, Yunnan Province, Kunming City, Kunming Botanical Garden (), on dead culm of monocotyledon, 20 March 2014, Q.R. Li, GZ43 (GZUH0122, GACP QR0201; living cultures, GZUCC0122).

Note.

The characteristics of the holotype specimen (≡ ) were revised, re-measured and described. is similar to and (Wang et al. 2004, Hyde et al. 2020). However, no clypeus was observed for . has larger ascomata (480–560 × 280–320 μm) with a central papilla surrounded by a circle of white tissue (Hyde 1996) which differs from those of . One new collection (GZUH0122, Fig. 5) shows the same traits of (MFLU 19-2333) in having immersed ascomata with clypeus, a wedge-shaped J+, ascus subapical ring, same dimensions of ascospores and here we provide it as a new geographical record from China.

Figure 5.

(GZUH0122) A, B ascomata on the surface of host C paraphyses and asci D section of ascoma E peridium F, G apical rings H–K asci with ascospores M–P ascospores. Scale bars: 50 μm (C, D); 10 μm (E, F–K); 5 μm (L–P).

Discussion

shares similar morphology to , and in having immersed ascomata and 2-celled ascospores (Hyde 1996). , , and are accepted in with newly-introduced genera, such as and (Mapook et al. 2020). has similar characteristics to in its occurrence on monocotyledons, having asci with J+, apical rings and brown 2-celled ascospores (Kohlmeyer and Volkmann-Kohlmeyer 1993). The ascospores of are provided with several longitudinal germ slits and differ from those of , where the ascospores are provided with ridges or a verrucose wall and lack germ slits. produces ascospores that are smooth-walled and surrounded by mucilaginous sheaths that are drawn out at the poles with germ slits, which differ from (Hyde 1995). possesses hyaline ascospores pointed at both ends, which differs from that of (Liu et al. 2015). The phylogenetic tree (Fig. 1) displays that (≡ ) clusters in the group with high support values (100%/1.00 PP). possesses ascospores with longitudinal striations and bristle-like polar appendages at both ends, without a gelatinous sheath, which differentiates it from other genera in . Ascospores of are provided with several longitudinal germ slits and differ from those of (Kohlmeyer and Volkmann-Kohlmeyer 1993). At present, 16 species have been described and a summary of each species are given in the Table 2.

Table 2.

Synopsis of the species of .

Species	Host	Clypeus	Ascomata	Asci	Ascal ring	Ascospores	Distribution
A.australis	Phragmitesaustralis	Present	420–560 × 290–380 µm, globose to subglobose	140–230 × 15.5–24 µm	4–5 × 2.5–3 μm, trapezoidal, J+	22.5–29 × 8–11 µm, wall striate, mucilaginous sheath	China (Guizhou)
A.bambusae	Bambusa sp.	Absent	500–560 × 294–350 µm, globose to subglobose	132.5–140 × 7.5–8 µm	2.5–3 µm in diam., ca. 2.5 µm high, wedge-shaped, J+	19–22.5 × 5.5–7 µm, slightly tapering at the ends, wall striate, mucilaginous sheath	Hong Kong
A.calamicola	Calamus sp.	Absent	520 × 390 µm, subglobose	160–190 × 14–20 µm	4-4.8 µm diam., 3.2-4 µm high, wedge-shaped, J+	24–33 × 5.5–9 µm, wall striate, verrucose, mucilaginous sheath	Brunel, Indonesia
A.chamaeropis	Chamaeropshumilis	Minute	400–700 × 300–400 µm, subglobose	150–190 × 9–10 µm	3.5–4.5 diam., 1.5–2 µm high, wedge-shaped, J+	15–23 × 5.5–7 µm, wall striate, covered by pronounced verrucose ornamentation, mucilaginous sheath	Spain
A.coronata	Gigantochloascribneriana, Bambusa sp.	Present	90–100 × 42–105 µm, subglobose or ellipsoidal	132.5–157.5 × 7.5–9 µm	3.5–4 µm in diam., 2–2.5 µm high, wedge-shaped, J+, with a faint canal leading to the apex.	29–31 × 5–5.5 µm, wall faint striate, mucilaginous sheath	Philippines, Hong Kong
A.clypeata	A unknown gramineous plant	Present	367‒448 × 278‒363 µm, subglobose to globose	180–245 × 10.5–14.5 µm	3–4 × 3–4 μm, square-shaped, J+	18.5–22.5 × 6.5–9 µm, wall striate, mucilaginous sheath	China (Guizhou)
A.deutziae	Deutziaestamineae	Absent	400–600 µm diam., globose	180–240 × 16–19 µm	3.5–4. 5 µm diam., 1.5–2 µm high, wedge-shaped, J+	26–32 × 11–13 µm, wall striate	India
A.eugeissonae	Eugeissonatristis	Absent	460–520 × 180–260 µm, Subglobose or ellipsoidal	175–220 × 11–16.5 µm	3–4 µm diam., 1.5–2.0 µm high, discoid, J+	25-40 × 6.5–9 µm, wall weakly striate, verrucose, mucilaginous sheath	Malaysia
A.foveata	Nolinae sp.	Present	300–400 ×400–500 µm, globose or ovoid	130–150 × 14–15 µm	3–4 µm wide, 4–5 µm high, tubular, J+	16–20 × 8–10 µm, wall striate, foveate, surface aspect of numerous warts	USA
A.gulubiicola	Gulubiacostate	Present	290–400 × 140–190 μm, subglobose or lenticular	107–145 × 11–13.5 μm	3.2–4 µm diam., 2.4–3.2 µm high, cylindrical, J+	14.5–18.5 × 6–9 μm with a minutely verrucose wall, mucilaginous sheath	Papua New Guinea
A.miscanthi	Miscanthussinensis	Present	283–296 × 272–277 µm, globose to subglobose	147–189 × 10–13 μm	3.5–4 µm broad, 2–2.5 µm high, wedge-shaped, J+	20–24 × 6–8 μm, wall striate, mucilaginous sheath.	China (Taiwan, Yunnan)
A.muroiana	Phyllostachysbambusoides	Absent	350–460 × 320–400 µm, globose	125–165 × 10–12 µm	3.5–4 µm diam., 2–2.5 µm high, wedge-shaped, J+	20–25 × 6–7.5 µm, wall finely striate, mucilaginous sheath	Japan
A.notabilis	Calamus, Bamboo	Present	400 × 360 µm, subglobose	180–220 × 11–14 µm	4–4.45 µm diam., 3–4.5 µm high, wedge-shaped, J+	20–26 × 6–8 µm, wall striate, finely verrucose, mucilaginous sheath	Brunei, Hong Kong, Indonesia
A.nypae	Nypafruticans	Absent	400–500 µm diam., subglobose	140–205 × 11–13 µm	4.5 µm diam., 2.5–4 µm high, wedge-shaped, J+	19–26 × 7–8 µm, wall striate, mucilaginous sheath	Malaysia
A.saccharicola	Sacchariofficinarum	Absent	420–525 × 350–420 µm high	140–16 × 7–10 µm	Not blued by Melzer’s reagent	20–24 × 6–8 µm, wall smooth or striated	Jamaica
A.serrulata	Korthalsia sp., Sabal sp., Serenoa sp.	Present	480–560 × 280–320 µm, conical with flattened base	110–112 × 10–12 µm,	3.2 µm diam., 2.4 µm high, wedge-shaped, J+	17–26 × 7–9.5 µm, wall striate, mucilaginous sheath	Brunei, USA, Florida

The combined ITS, LSU, rpb2 and β-tubulin phylogeny (Fig. 1) showed two clades of as Clade 1 and Clade 2. The differs from and (see above). The sequence from the holotype of is noticeably clustered with spp. in (Fig. 1). However, showed a close affinity with spp. in , based on analysis of the combined LSU and ITS sequence alignment in Senanayake et al. (2015). has similar characteristics to and other genera of (Hyde 1996). Hence, there should be more evidence to reassess in the future. The unitunicate asci with a J+ apical ring in Melzer’s regent and brown ascospores covered with longitudinal wall striations, without germ slits can clearly distinguish from its similar genera. In addition, a table including synopsis of the species of is provided.