The Genus Acervus from Southwestern China and Northern Thailand

Figures & data

Table 1. Primers used for amplification.

Locus	Primer	Primer sequence (5'–3')
LSU [18]	LROR	5'-ACC CGC TGA ACT TAA GC-3'
	LR5	5'-TCC TGA GGG AAA CTT CG-3'
tef1-α [19]	983F	5'-GCY CCY GGH CAY CGT GAY TTY AT-3'
	2218R	5'-AT GAC ACC RAC RGC RAC RGT YTG-3'
rpb2 [20]	fRPB2-5f	5'-GAY GAY MGW GAT CAY TTY GG-3'
	fRPB2-7cR	5'-CCC ATR GCT TGY TTR CCC AT-3'
rpb1 [21]	gRPB1-A	5'-GAK TGT CCK GGW CAT TTT GG -3'
	fRPB1-Crev	5'-CNG CDA TNT CRT TRT CCA TRT A -3'
SSU [22]	NS1	5'-GTA GTC ATA TGC TTG TCT C-3'
	NS4	5'-CTT CCG TCA ATT CCT TTA AG-3'

Table 2. PCR conditions used for amplification.

Procedure	Genes
	LSU	tef1-α	rpb2	rpb1	SSU
Pre-denaturation	94 °C for 5 min
Denaturation	94 °C for 50 s
Annealing	55 °C for 50 s	56 °C for 50 s	55 °C for 2 min	55 °C for 2 min	55 °C for 50 s
Extension	72 °C for 1 min
Final extension	72 °C for 8 min
Cycles	Number of cycles for LSU was 35 and for the rest 40

Table 3. Sequences used in this study.

Species name	Distribution	Voucher	GenBank No.
			LSU	tef1	rpb2	rpb1	SSU
Acervus beijingensis	China, Beijing	HMAS 78150	HM197754	KP993492	KP993484	KP993500	DQ787815
Acervus changchunensis	China, Jilin	HMAS 78146	KP993477	KP993490	KP993482	KP993498	DQ787813
Acervus epispartius	China	HMAS 78149	HM197753	KP993491	KP993483	KP993499	DQ787814
Acervus epispartius	China	HMAS 173242	HM197755	–	–	–	–
Acervus epispartius	USA, New York	s.n. (FH)	DQ220305	–	–	–	–
Acervus epispartius	China, Yunnan	MFLU 20-0257	MT165625	MT336154	MT210807	–	MT165629
Acervus flavidus	Puerto Rico	DHP PR98.2 (FH)	DQ220306	–	–	–	–
Acervus flavidus	China	HKAS 90046	KX765259	KX765252	KX765260	–	–
Acervus flavidus	China, Fujian	HMAS 188443	HM197756	KP993487	KP993479	KP993495	–
Acervus globulosus	China, Yunnan	HKAS 88987	KX765253	KX765255	KX765254	–	–
Acervus globulosus	Thailand, Chiang Mai	MFLU 20-0258	MT165626	MT336155	MT210808	–	MT165630
Acervus heilongjiangensis	China, Heilongjiang	HMAS 271281	KP243162	KP993493	KP993485	KP993501	–
Acervus rufus	China, Yunnan	MFLU 20-0259	MT192219	MT227372	MT210811	MT210810	MT192216
Acervus stipitatus	China, Yunnan	MFLU 16-0607	KX765256	KX765258	KX765257	–	–
Acervus stipitatus	China, Yunnan	MFLU 20-0260	MT165627	MT336156	MT210809	MT210806	MT165631
Monascella botryosa	Spain	CBS 233.85	MH873558	KC109256	JX943831	JX943733	–
Warcupia terrestris	Canada	CBS 891.69	DQ220467	KC109308	–	–	–

Names in red indicate new species, while those in blue indicate new collections in this study.

CBS: Centraalbureau voor Schimmelcultures, Utrecht, The Netherlands; DHP: Donald H. Pfister; HKAS: Herbarium of Cryptogams of Kunming Institute of Botany, Chinese Academy of Sciences. HMAS: Chinese Academy of Sciences, Beijing, China; MFLU: Mae Fah Luang University Herbarium, Chiang Rai, Thailand.

Figure 1. Phylogenetic tree of combined LSU, tef1-α, rpb2, rpb1, and SSU sequence data inferred from 17 taxa and 5257 sites under the GTR (general time reversible) + GAMMA model of nucleotide substitution. Numerical values at the nodes indicate maximum likelihood bootstrap support (blue), maximum parsimony bootstrap support (yellow), and posterior probabilities (black). Only values greater than 70% (maximum likelihood and maximum parsimony) and 0.95 (Bayesian analysis) are indicated. Name in red indicates new species. Names in blue indicate new collections. Tree is artificially rooted to Monascella botryosa (CBS 233.85) and Warcupia terrestris (CBS 891.69) taxa.

Figure 2. Acervus epispartius (HKAS 107301). (a–d) Typical mature specimens. (e) Receptacle surface of pileus in 5% KOH. (f) Asci and paraphyses in 5% KOH. (g, h) Paraphyses in Congo red. (i, j) Asci. (j Asci in Congo red.). (k, l) Apexes of asci. (k Apex in Congo red. l Apex in Cotton blue.). m-p Ascospores. Scale bars: e–f = 50 μm. g–h = 20 μm. i–j = 30 μm. k–p = 5 μm.

Figure 3. Acervus globulosus (MFLU 20-0258). (a–f) Typical mature specimens. (g, h) Receptacle surface of pileus. (i) Asci and paraphyses. (j–l) Asci (l Asci in Melzer’s reagent.). (m–p) Apexes of asci (m–o Apexes in Congo red.). (q–u) Ascospores. Scale bars: c = 1500 μm. d–f = 1000 μm. g–l = 30 μm. m–u = 5 μm.

Table 4. Morphological differences for two collections of Acervus globulosus.

	Ekanayaka et al.’s descriptions	Our descriptions
Medullary excipulum	hyaline of textura intricata cells	hyaline to yellowish of textura angularis to textura globulosa
Paraphyses	hyaline	yellow
Apex of asci	inoperculate	suboperculate
Size of asci	72–96 × 5–6 μm	82 − 100 × 6 − 8 µm

Figure 4. Acervus rufus (HKAS 107302, holotype). (a–c) Typic mature specimens. (d, e) Receptacle surface of pileus. f Hymenium in Congo red. (g) Apex of asci in Congo red. (h) Paraphyses in Congo red. (i–k) Asci (j Asci in Congo red. k Asci in Melzer’s reagent.). (l–o) Ascospores. Scale bars: d–f, i–k = 50 μm. h = 30 μm l = 20 μm. g, m–o = 5 μm.

Figure 5. Acervus stipitatus (HKAS 107302). (a) Habitat. (b, c) Typic mature specimens. (d) Receptacle surface of pileus. (e–g) Paraphyses (g Paraphyses in Congo red.). (h–j) Asci (i-i Asci in Congo red.). (k–m) Apexes of asci. (n–q) Ascospores. Scale bars: d, h–j = 30 μm. e–g = 20 μm. k–m, o–q = 5 μm. n = 15 μm.

Figure 6. Acervus stipitatus (MFLU 16-0607, holotype). (a–e) Herbarium materials. (f) Receptacle surface of pileus. (g–i) Asci (h Asci in Congo red. i Asci in Cotton blue). (k–l) Apexes of asci. (l Apex of asci in Cotton blue). (m–p) Ascospores. Scale bars: d–i = 30 μm. j–l, n–p = 5 μm. m = 15 μm.

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