Hypomyces pseudolactifluorum sp. nov. (Hypocreales: Hypocreaceae) on Russula sp. from Yunnan, PR China.

BACKGROUND: Hypomyces is a large genus of fungicolous fungi, parasitising the fruiting bodies of Agaricales, Boletales, Helotiales, Pezizales and Polyporales. Hypomyces currently comprises of 147 species widely distributed in Australia, China, France, Germany, Italy, Japan, North America, Sri Lanka, Thailand and UK. Amongst them, 28 species have been recorded in China. NEW INFORMATION: Hypomyces pseudolactifluorum sp. nov., growing on the fruiting bodies of Russula sp. in subsect. Lactarioideae and collected from Yunnan, China, is described with illustrations and molecular phylogenetic data (combined ITS, LSU, TEF1-α and RPB2 sequence dataset). The new species is characterised by semi-immersed to immersed perithecia and fusiform, apiculate and verrucose ascospores. We also review the species diversity of the genus Hypomyces in China. Feng-ming Yu, Ruvishika S. Jayawardena, Jianwei Liu, Kevin D. Hyde, Qi Zhao.

Introduction

Fungicolous fungi are a large and diverse ecological group, currently containing more than 1500 taxa distributed in many lineages across the fungal kingdom (Põldmaa 2011, Sun et al. 2019a). (Fr.) Tul. & C. Tul. is an important genus of fungicolous fungi and placed in the family (, , ) (Hyde et al. 2020). was originally introduced as a subgenus of Fr. (Fries 1825) and then Tulasne and Tulasne (1860) revised it to a genus and designated (Schwein.) Tul. & C. Tul. from the USA as its type. parasitises the fruiting bodies of , , , and (Rossman et al. 1999, Tamm and Põldmaa 2013, Sun et al. 2019a). is characterised by: superficial or immersed, spherical to ovate, pyriform, papillate and yellow, orange, tawny red or green perithecia in a subiculum; 8-spored, subcylindrical to cylindrical and with a thickened apical asci; and ellipsoid, lanceolate, fusiform to navicular, 0-1-septate or rarely 3-septate, hyaline, spinulose or verrucose and smooth-walled ascospores (Rossman et al. 1999, Zeng and Zhuang 2015). Its allied genera include Nees, Link, Link and Wallr (Wijayawardene et al. 2017) and its asexual morphs are -, -, -, - or -like (Jaklitsch et al. 2006, Hyde et al. 2020). currently comprises of 147 species in Species Fungorum (http://www.speciesfungorum.org/, accessed in April 2020) and is widely distributed in Australia, China, France, Germany, Italy, Japan, North America, Sri Lanka, Thailand and UK (Zhuang et al. 2012, Rossman et al. 2013, Zeng and Zhuang 2016, Zare and Gams 2016, Lechat et al. 2017, Wei and Kirschner 2017, Sun et al. 2019a, Sun et al. 2019b, Zeng and Zhuang 2019). Amongst them, 28 species have been reported in China (Table 1).

Table 1.

Species diversity of the genus in China (29 species in total).

Taxa names	Hosts	Distribution	References
Hypomyces amaniticola	Amanita sp.	China (Yunnan)	Zeng and Zhuang 2016
H. aurantius	Agaricus bisporus, Polyporales (Cymatoderma sp., Laetiporus sulphureus, Panellus sp., Polyporus picipes), Stereum sp.	China (Anhui, Fujian, Guangxi, Hainan, Hebei, Hunan, Jiangsu, Jiangxi, Shanghai, Sichuan, Zhejiang), New Zealand, USA	Chen and Fu 1989, Põldmaa 2011, Luo and Zhuang 2012
H. aureonitens	Phlebia tremellosa, Polyporus sp.	China (Fujian, Guangxi), Europe	Teng 1963, Sun et al. 2019a
H. chlorinigenus	Agaricaceae, Boletaceae	Belgium, China (Taiwan), Guyana; Indonesia, New Zealand, USA	Rogerson and Samuels 1989, Zeng and Zhuang 2016
H. chrysospermus	Boletus sp., Hemileccinum impolitum, Suillus americanus, Russula sp.	China (Fujian, Jiling, Nanjing), Russia	Ma 2008, Luo and Zhuang 2012
H. completiopsis	Boletus sp.	China (Yunnan)	Zeng and Zhuang 2016
H. fistulina	Fistulina sp.	China (Guangxi)	Sun et al. 2019b
H. hubeiensis	Agaricus sp.	China (Hubei)	Zeng and Zhuang 2019
H. hyalinus	Agaricales (Amanita sp.), Polyporales	Canada, China (Jiangsu), Japan, USA	Teng 1934, Teng 1963, Rogerson and Samuels 1994
H. lateritius	Lactarius camphoratus, L. chelidonium, L. controversus, L. deliciosus, L. sanguifluus, L. thejogalus, L. trivialis, Lactarius sp.	Canada, China (Tibet), Europe, Japan, Mexico, New Zealand, USA	Rogerson and Samuels 1994, Luo and Zhuang 2012
H. luteovirens	Russula atropurpurea, R. rosea, R. sanguinaria, Russula sp.	Canada, China (Inner Mongolia), Europe, Japan, Russia, USA	Rogerson and Samuels 1994, Ma 2008
H. macrosporus	Russulaceae	China (Hubei), Mexico, USA	Rogerson and Samuels 1994, Luo and Zhuang 2012
H. microspermus	Boletaceae, Boletus sp., Imleria badia, Xanthoconium affine, Xerocomellus chrysenteron, Xerocomus sp.	Canada, China (Fujian, Guizhou, Hainan, Hubei, Jilin, Taiwan, Yunan), Indonesia, New Zealand, USA	Rogerson and Samuels 1989, Zeng and Zhuang 2016
H. mycophilus	Auricularia sp., Bulgari sp., Marasmius sp., Polyporus sp., Trametes versicolor	China (Guangdong), USA	Rogerson and Samuels 1993, Zeng et al. 2017
H. ochraceus	Decaying leaves, wood and fungi (e.g. Russula sp.)	China (Guangxi, Yunnan), Europe, USA	Teng 1963, Sun et al. 2019a
H. orthosporus	Polyporales	China (Tibet), Estonia, Finland, The Netherlands	Põldmaa 1996, Zeng and Zhuang 2019
H. papulasporae	Geoglossum difforme, G. fallax, G. glabrum, G. nigritum, G. simile, Glutinoglossum glutinosum, Trichoglossum hirsutum, T. walteri	China, USA, New Zealand	Rogerson and Samuels 1985, Sun et al. 2019a
H. polyporinus	Auricularia auricula-judae, Polyporales, Trametes versicolor, T. pubescens, Polyporus sp.	Canada, China (Guangxi), USA	Teng 1963, Rogerson and Samuels 1993
H. pseudolactifluorum sp. nov.	Russula sp.	China (Yunnan)	This study
H. rosellus	Agaricus bisporus, Armillaria sp., Hydnellum sp., Hyphoderma sp., Mycena sp., Polyporus sp., Russula sp., Trichaptum sp.	China (Gansu), Europe, Iran, Japan, Korea, USA	Tamm and Põldmaa 2013, Sun et al. 2019b
H. semicircularis	Ganoderma sichuanense, Microporus xanthopus	Cuba, China	Wei and Kirschner 2017, Sun et al. 2019a
H. sibirinae	Aphyllophorales, Boletus sp., Polyporales	China (Hunan), Indonesia, USA	Samuels et al. 1990, Zeng et al. 2017, Sun et al. 2019a
H. sinicus	Schizophyllum sp.	China (Anhui)	Zhuang et al. 2012
H. stephanomatis	Humaria hemisphaerica, Humaria sp.	Canada, China (Hubei), Germany, USA	Rogerson and Samuels 1985,Zeng and Zhuang 2016
H. subiculosus	Polyporaceae (Microporus affinis, Trametes versicolo)	China (Anhui, Beijing, Guangxi, Zhejiang), Cuba, Japan	Rogerson and Samuels 1993, Luo and Zhuang 2012
H. succineus	Pholiota sp.	China (Taiwan), USA	Rogerson and Samuels 1994, Zeng and Zhuang 2016
H. tegillum	Aphyllophorales, Polyporales	Brazil, China (Guangxi, Yunnan), Panama, USA	Rogerson and Samuels 1993, Luo and Zhuang 2012
H. triseptatus	Bark or associated with an ascomycete; Pyrenomycete	China (Hunan, Guangdong), Gabon	Rossman and Rogersson 1981, Zeng et al. 2017
H. yunnanensis	Boletus sp.	China (Yunnan)	Zeng and Zhuang 2016

Fungicolous fungi play important roles in the processes of the growths and degradations of their hosts. With the rapid development of mushroom industries, the fungicolous fungi on mushrooms have received more and more attention (Hyde et al. 2019). In this paper, we introduce a new member of fungicolous fungi, sp. nov., on the fruiting bodies of sp., collected from Yunnan Province, China. At the same time, we review the species diversity of the genus in China.

Materials and methods

Collections and Morphology

specimens, including their host mushrooms, were collected in an evergreen broad-leaved forest in Baihualing, Baoshan, Yunnan Province, China. The specimens, as well as collected host mushrooms, were placed on a piece of aluminium foil at first, then rolled the paper into a cylinder, twisted at the ends for sealing and lastly taken back to the laboratory for study (McKnight and McKnight 1997). Colour codes were recorded following those of Kornerup and Wanscher (1978). A Nikon Coolpix P510 camera was used to take photos in the wild. Dried specimens were observed and photographed using an Olympus SZ61 stereomicroscope and a Nikon ECLIPSE Ni compound microscope fitted with a Canon EOS 600D digital camera. Measurements were made using the Tarosoft® Image Frame Work programme v.0.9.7. The colour change of the perithecial wall was tested using 5% potassium hydroxide (KOH). Type specimens are deposited at the Herbarium of Mae Fah Luang University, Thailand (MFLU) and the Herbarium of Cryptogams Kunming Institute of Botany, Chinese Academy of Sciences, PR China (HKAS).

DNA extraction, PCR amplification and sequencing

The genomic DNA was extracted from the dried materials using the CTAB method (Doyle 1987). Tissues from the ascocarps of parasitic fungi and fruiting bodies of the host mushrooms were used to extract DNA, respectively. Primer pairs ITS1F/ITS4 (White et al. 1990), LR0R/LR5 (Rehner and Samuels 1994, Vilgalys and Hester 1990), TEF1-α 983f/TEF1-α 2218r (Carbone and Kohn 1999, Rehner and Buckley 2005) and RPB2-5f/RPB2-7cR (Liu et al. 1999) were used for amplification of the ITS, LSU, TEF1-α and RPB2 gene regions.

PCR was performed in a 25 μl reaction volume: 12.5 μl Taq PCR Master Mix (Abmgood, Richmond, BC, Canada), 1 μl forward primer, 1 μl reverse primer, 1 μl DNA template and 9.5 μl ddH2O. For ITS and LSU, PCR reaction conditions are: 8 min at 94ºC, followed by 30 s at 94ºC, 30 s at 52ºC and 1 min at 72ºC for 35 cycles and a final extension of 10 min at 72ºC. PCR reaction conditions of TEF1-α and RPB2 are: 8 min at 94ºC, followed by 1 min at 95ºC, 45 s at 59ºC for RPB2/55ºC for TEF1-α and 1 min at 72ºC for 35 cycles and a final extension of 10 min at 72ºC. The PCR products were detected using agarose gel electrophoresis and, in the gel documentation system, clear bands were observed. Sequencing was performed by Sangon Biotech (Shanghai) Co. Ltd., PR China; partial impure products were purified using the Cycle-pure-kit (Omega, America) and then cloned into pClone007 Simple vector (TSV-007S from Beijing TsingKe Biotech). Twenty clones of PCR products of each gene were sequenced using the universal primer pairs M13-47/M13-48.

Sequence alignment and phylogenetic analyses

The parasitic fungus: sp. nov.

Molecular phylogenetic trees were constructed using our sequencing results of sp. nov. and the voucher sequences of their allies obtained from NCBI GenBank (Table 2). Two species of , (DAOM 167057) and (CBS 119325) were used as outgroup taxa. All sequences were assembled and aligned using MAFFT v6.8 (Katoh et al. 2005) and manually edited via BioEdit version 7.0.9 (Hall 1999). Four sequence matrices of ITS, LSU, TEF1-α and RPB2 genes, respectively, were compiled. The optimal substitution model for each gene dataset was determined using jModelTest2 under the Akaike information criterion (AIC) (Darriba et al. 2012). The results indicated that the GTR+I+G model (-lnL = 8658.2624) is optimal for the ITS dataset, as well as the TIM1+I+G model (-lnL = 4392.5417) for LSU, the TrN+I+G model (-lnL = 5751.4959) for TEF1-α and the model SYM+I+G (-lnL = 6419.6669) for RPB2, respectively. Using the aligned sequence matrices, a combined gene sequence dataset (ITS, LSU, TEF1-α and RPB2, orderly) was assembled and aligned and was finally deposited in TreeBASE database (http://purl.org/phylo/treebase/phylows/study/TB2:S26593?x-access-code=152eadfc2292343af7627cfad5c2946c&format=html).

Table 2.

Voucher information and GenBank accession numbers for samples appearing in the phylogenetic tree. Our sequencing results are displayed in bold. (Label T indicate the sequences from ex-type strains.)

Taxa names	Specimen/Strain number	GenBank accession numbers				References
		ITS	LSU	TEF1-α	RPB2
Cladobotryum amazonense	CBS 470.80	MH861285	MH873051	/	/	Vu et al. 2019
C. apiculatum	CBS 174.56 T	NR_159770	MH869109	/	/	Vu et al. 2019
C. asterophorum	CBS 676.77 T	FN859395	MH872869	FN868712	FN868649	Põldmaa 2011
C. croceum	CBS 231.95	MH862511	MH874154	/	/	Vu et al. 2019
C. cubitense	CBS 416.85	FN859396	/	FN868713	FN868650	Põldmaa 2011
	G.A. m643.w	FN859397	/	FN868714	FN868651	Põldmaa 2011
C. indoafrum	TFC 201295	FN859403	FN859403	FN868721	FN868657	Põldmaa 2011
C. multiseptatum	CBS 472.71 T	FN859405	MH871991	FN868723	FN868659	Põldmaa 2011
C. obconicum	CBS 528.81	MH861373	MH873126	/	/	Vu et al. 2019
C. paravirescens	TFC 97-23 T	FN859406	FN859406	FN868724	FN868660	Põldmaa 2011
C. penicillatum	CBS 407.80 T	FN859407	MH873046	FN868725	FN868661	Põldmaa 2011
C. protrusum	CBS 118999	FN859408	FN859408	FN868726	FN868662	Põldmaa 2011
C. purpureum	CBS 154.78 T	FN859415	/	FN868733	FN868669	Põldmaa 2011
C. rubrobrunnescens	CBS 176.92 T	FN859416	MH874016	FN868734	FN868670	Põldmaa 2011
Cladobotryum sp.	FSU 5046	FN859421	/	FN868739	FN868675	Põldmaa 2011
C. stereicola	CBS 457.71 T	MH860217	MH871984	/	/	Vu et al. 2019
C. tchimbelense	TFC 201146 T	FN859419	FN859419	FN868737	FN868673	Põldmaa 2011
C. tenue	CBS 152.92 T	FN859420	FN859420	FN868738	FN868674	Põldmaa 2011
Hypomyces aconidialis	TFC 201334 T	FN859457	FN859457	FN868775	FN868711	Põldmaa 2011
	TFC 201215	FN859456	FN859456	FN868774	FN868710	Põldmaa 2011
H. albidus	CBS 460.71	MH860220	MH871987	/	/	Vu et al. 2019
H. armeniacus	TFC 02-86/2 T	FN859424	FN859424	FN868742	FN868678	Põldmaa 2011
H. aurantius	TFC 95-171	FN859425	FN859425	FN868743	FN868679	Põldmaa 2011
H. australasiaticus	TFC 03-8 T	FN859428	FN859428	FN868746	FN868681	Põldmaa 2011
	TFC 99-95	FN859427	/	FN868745	FN868680	Põldmaa 2011
H. australis	TFC 2007-18	AM779860	AM779860	FN868747	/	Põldmaa 2011
H. boletiphagus	MFLU 17-1391	MH459152	MH459168	/	MH464785	Sun et al. 2019b
H. boletus	MFLU 17-1392	MH459153	MH459170	/	MH464787	Sun et al. 2019b
H. chlorinigenus	KSH511	KT946843	/	KU041505	KU041493	Otto et al. 2016
	KSH512 T	KT946844	/	KU041506	KU041494	Otto et al. 2016
H. completus	KSH411 (S172)	KT946842	/	KU041504	KU041492	Otto et al. 2016
	KSH410 (S171) T	KT946841	/	KU041503	KU041491	Otto et al. 2016
H. corticiicola	CBS 137.71 T	MH860037	MH871817	/	/	Vu et al. 2019
H. dactylarioides	CBS 141.78 T	FN859429	MH872879	FN868748	FN868683	Põldmaa 2011
H. fistulina	HMAS 279800 T	MH459154	MH459171	MH464781	/	Sun et al. 2019b
H. gabonensis	TFC 201156 T	FN859430	FN859430	FN868749	FN868684	Põldmaa 2011
H. heterosporus	CBS 719.88 T	FN859398	MH873844	FN868716	FN868653	Põldmaa 2011
H. khaoyaiensis	G.J.S. 01-304 T	FN859431	AJ583483	FN868750	FN868685	Põldmaa 2011
H. lactifluorum	TAAM 170476 T	FN859432	EU710768	FN868751	EU710773	Põldmaa 2011
H. laeticolor	JCM 10758 T	LC228655	LC228712	/	/	Sun et al. 2019b
H. luteovirens	CBS 128483	MH864958	MH876402	/	/	Vu et al. 2019
H. mycophilus	CBS 175.56	MH857567	MH869110	/	/	Vu et al. 2019
H. odoratus	G.A. m329	FN859434	FN859434	FN868753	FN868688	Põldmaa 2011
	TFC 200887	FN859439	/	FN868757	FN868693	Põldmaa 2011
H. orthosporus	10736	MK478468	MN044763	MK484609	/	Zeng and Zhuang 2019
H. penicillatus	NBRC 100524	LC146740	LC146740	/	/	Sun et al. 2019b
H. pseudolactifluorum sp.nov.	MFLU 20-0265 T	MT260402	MT260399	MT259361	MT259359	This study
	MFLU 20-0266	MT260403	MT260400	MT259362	MT259360	This study
H. polyporinus	ATCC 76479	AF543771	AF543793	AF543784	/	Currie et al. 2003
H. pseudocorticiicola	JCM 12654 T	LC228663	LC228721	/	/	Sun et al. 2019b
H. rosellus	TFC 201071	FN859443	FN859443	FN868762	FN868697	Põldmaa 2011
H. samuelsii	CBS 536.88	FN859444	/	FN868763	FN868698	Põldmaa 2011
	TFC 2007-23	FN859451	FN859451	FN868769	FN868705	Põldmaa 2011
H. semicircularis	CBS 705.88 T	FN859417	MH873843	FN868735	FN868671	Põldmaa 2011
H. semitranslucens	CBS 458.71	MH860218	MH871985	/	/	Vu et al. 2019
	CBS 821.70	MH859960	MH871759	/	/	Vu et al. 2019
H. sibirinae	CBS 744.88	MH862151	AJ459304	/	/	Vu et al. 2019
H. sinicus	HMAS 251317 T	NR_156252	MN044986	MK484610	/	Zhuang et al. 2012
H. stephanomatis	G.J.S. 88-50	/	AF160243	AF534632	AF545566	Põldmaa et al. 2000
H. subiculosus	TFC 97.166	FN859452	/	FN868770	FN868706	Põldmaa 2011
H. tubariicola	CBS 115.79 T	KU382164	MH872953	/	/	Vu et al. 2019
	CBS 225.84	KU382162	KU382220	/	/	Zare and Gams 2016
H. virescens	G.A. i1906 T	FN859454	/	FN868772	FN868708	Põldmaa 2011
	G.A. i1899	FN859453	/	FN868771	FN868707	Põldmaa 2011
Sepedonium ampullosporum	CBS 392.52 T	MH857094	MH868629	/	/	Vu et al. 2019
S. chalcipori	CBS 278.92	MH862358	MH874023	/	/	Vu et al. 2019
	CBS 148.92 T	MH862347	MH874014	/	/	Vu et al. 2019
S. tulasneanum	CBS 940.69	MH859489	MH871270	/	/	Vu et al. 2019
Trichoderma hamatum	DAOM 167057 T	EU280124	HM466686	AF534620	AF545548	Hoyos-Carvajal et al. 2009
T. viride	CBS 119325 T	DQ677655	/	DQ672615	EU711362	Jaklitsch et al. 2006

Maximum Likelihood (ML) analysis was performed using IQ-Tree (Nguyen et al. 2014, Chernomor et al. 2016) with the computing models listed above and a bootstrap test of 1000 replicates. Bayesian Inference (BI) analysis was carried out using MrBayes v3.2.6 (Ronquist et al. 2012). The TIM1 and TrN substitution models were replaced by the GTR model (Huelsenbeck and Rannala 2004). Four simultaneous Markov Chain Monte Carlo (MCMC) chains were run for random trees of 10,000,000 generations and were sampled by every 100 generations. The computing was stopped when the standard deviation of the split frequencies fell below 0.01 and ESS values > 200. Subsequently, phylogenetic trees were summarised and posterior probabilities (PP) were performed using MCMC by discarding the first 25% generations as “burn-in” (Huelsenbeck and Ronquist 2001). Gaps were treated as missing data. Phylogenetic trees were viewed in FigTree v.1.4.2 (http://tree.bio.ed.ac.uk/software/figtree).

The host mushroom:

Voucher sequences (ITS gene) for phylogenetic analyses of the host mushroom and its allies were obtained from our sequencing results and GenBank databases (Li et al. 2020) (Table 3). Five species of , R. . . . and were selected as the outgroup taxa. Sequence alignment and phylogenetic analyses followed those of the parasitic fungus above. ML analysis was performed using IQ-Tree with TVM+I+G model (-lnL = 5298.7964) (Nguyen et al. 2014, Chernomor et al. 2016). The ITS sequence matrix of the host mushroom and its allies were deposited in the TreeBASE database (http://purl.org/phylo/treebase/phylows/study/TB2:S26693?x-access-code=2e445b17aebe1f93266051a8920ae62f&format=html).

Table 3.

Voucher information and GenBank accession numbers for samples appearing in the phylogenetic tree. Our sequencing results are displayed in bold.

Taxa names	Specimen/Strain number	GenBank accession	References
Russula acrifolia	TUB UE12.09.2003-3	DQ421998	Eberhardt 2002
R. adusta	PC 547RUS27	AY061652	Miller and Buyck 2002
R. aff. chloroides	FH 12273	KT934015	Looney et al. 2016
R. brevipes	SMI329	FJ845429	Kranabetter et al. 2009
R. brevipes	JS160927-01	MG407682	GenBank
R. brevipes	TENN 070667	KY848511	Looney et al. 2018
R. brevipes var. acrior	JMP 0058	EU819422	Palmer et al. 2008
R. byssina	HGAS-MF 009907	MN648951	Li et al. 2020
R. byssina	HGAS-MF009921	MN648949	Li et al. 2020
R. byssina	HGAS-MF 009913	MN648950	Li et al. 2020
R. cascadensis	UBC F30189	KX812838	Bazzicalupo 2018
R. cascadensis	UBC F19691	HM240541	Buyck et al. 2017
R. cf. angustispora	PC BB2004-252	EU598152	GenBank
R. cf. brevipes	F 28785	MH718203	GenBank
R. cf. brevipes	F CDW47	GQ166868	GenBank
R. cf. brevipes	GO 2009-276	KC152212	GenBank
R. cf. delica	UBC F30260	KX812852	Bazzicalupo 2018
R. chloroides	PC 205RUS24	AY061663	Miller and Buyck 2002
R. chloroides	UBC F20353	KC581331	GenBank
R. chloroides	RUS-12091401	KF432954	Wisitrassameewong et al. 2014
R. cremicolor	HGAS-MF 009901	MN648955	Li et al. 2020
R. cremicolor	HGAS-MF 009908	MN648952	Li et al. 2020
R. cremicolor	HGAS-MF 009912	MN648953	Li et al. 2020
R. cremicolor	HGAS-MF 009919	MN648954	Li et al. 2020
R. delica	hue22 (TUB)	AF418605	Eberhardt 2002
R. delica	FH 12-272	KF432955	Wisitrassameewong et al. 2014
R. delica	HA 2015-004	KX263000	Aghajani et al. 2017
R. delica	PC 496RUS26	AY061671	Miller and Buyck 2002
R. delica	TUB hue22	AF418605	Eberhardt 2002
R. delica	UBC F30263	KX812842	Bazzicalupo 2018
R. delica	RMUKK 37	KX267630	GenBank
R. delica	KA 12-1327	KR673555	Kim et al. 2015
R. delica	HMJAU 32182	KX094989	Liu et al. 2017
R. eccentrica	HCCN 23685	KC699778	Park et al. 2014
R. japonica	MHHNU 31049	MK167414	Chen and Zhang 2019
R. japonica	HGAS-MF 009923	MN648957	Li et al. 2020
R. japonica	HGAS-MF 009915	MN648956	Li et al. 2020
R. leucocarpa	HGAS-MF 009910	MN648948	Li et al. 2020
R. leucocarpa	HGAS-MF 009916	MN648947	Li et al. 2020
R. littoralis	PC 1222IS87	AY061702	Miller and Buyck 2002
R. marangania	MEL 2293694	EU019930	Lebel and Tonkin 2007
R. nigricans	TUB fo46761	AF418607	Eberhardt 2002
R. pallidospora	PC 2-1221IS85	AY061701	Miller and Buyck 2002
R. pumicoidea	MEL T-14771	EU019931	Lebel and Tonkin 2007
R. sinuata	MEL H4755	EU019943	Lebel and Tonkin 2007
R. subnigricans	MHHNU ZP6932	EF534351	Yin et al. 2008
R. vesicatoria	PC 0124666	KY800359	Buyck et al. 2017
Russula sp.	MFLU 20-0265 (host)	MT755627	In this study

Taxon treatments

F. M. Yu, Q. Zhao & K. D. Hyde sp. nov.

1119A7CE-9897-55CC-94D7-5CF3C180F567

Type status: Holotype. Occurrence: catalogNumber: MFLU 20-0265; recordedBy: Jian-Wei Liu; lifeStage: Telemorph; Taxon: scientificName: ; Location: country: China; stateProvince: Yunnan; locality: Baoshan, Longyang, Baihualing; verbatimElevation: 2094m; locationRemarks: label transliteration: "Yunnan, Baoshan, Longyang, Baihualing, on ; verbatimCoordinates: ; decimalLatitude: 25.2989; decimalLongitude: 98.7845; georeferenceProtocol: label; Identification: identifiedBy: Feng-Ming Yu; dateIdentified: 2019

Type status: Paratype. Occurrence: catalogNumber: MFLU 20-0266; recordedBy: Jian-Wei Liu; lifeStage: Telemorph; Taxon: scientificName: ; Location: country: China; stateProvince: Yunnan; locality: Baoshan, Longyang, Baihualing; verbatimElevation: 2094m; locationRemarks: label transliteration: "Yunnan, Baoshan, Longyang, Baihualing, on ; verbatimCoordinates: ; decimalLatitude: 25.2989; decimalLongitude: 98.7845; georeferenceProtocol: label; Identification: identifiedBy: Feng-Ming Yu; dateIdentified: 2019

Type status: Isotype. Occurrence: catalogNumber: HKAS 107300; recordedBy: Jian-Wei Liu; lifeStage: Telemorph; Taxon: scientificName: ; Location: country: China; stateProvince: Yunnan; locality: Baoshan, Longyang, Baihualing; verbatimElevation: 2094m; locationRemarks: label transliteration: "Yunnan, Baoshan, Longyang, Baihualing, on ; verbatimCoordinates: ; decimalLatitude: 25.2989; decimalLongitude: 98.7845; georeferenceProtocol: label; Identification: identifiedBy: Feng-Ming Yu; dateIdentified: 2019

Description

Index Fungorum number: IF557817

Sexual morph. Subiculum light yellow (4A4–5) when fresh and pale orange, light orange to brownish-orange (5A3–4, 5C4, 6C6) after being dried, usually covering the pileus, stipe and deformed gills of the host mushroom. Perithecia aggregated, semi-immersed to immersed in subiculum, except for their erumpent papilla, yellowish-brown to dark brown (5E6, 6E6, 6F6–8), pyriform to subglobose, 262–484 × 136–284 μm; perithecial wall 12–25 μm thick, single-layer, cells 9–22 × 4–8 μm. Papilla prominent, 129–177 μm high, at base 135–284 μm wide. Asci 8-spored, cylindrical, 147–222 × 4–9 μm; apex thickened, 4.9–6.0 wide and 2.5-3.0 μm high. Ascospores uniseriate and with ends overlapping, fusiform, 30–38 × 6–8 μm, single-septate, septum median and with dense verrucae and prominently apiculate, apiculi 4.5–8.0 μm long, straight or curved. Asexual morph: unknown. (Fig. 1)

Figure 1.

sp. nov.. a: The host mushroom ( sp.); b-e: Perithecia embedded in subiculum effused over the substratum; d-e: Median sections of an ascoma; f: Section of peridium; g-l: Asci with ascospores; m-t: Ascospores. Scale bars: a = 5 cm; b = 1 mm; c = 200 μm; d, e = 100 μm; f, g= 50 μm; h - l = 20 μm; m - t = 10 μm.

Diagnosis

The new species is similar to on and spp. from North America (Rogerson and Samuels 1994), but has smaller perithecia and shorter asci. The main differences of the two species are compared in Table 4.

Table 4.

Main differences between and sp. nov..

	H. lactifluorum(Rogerson and Samuels 1994)	H. pseudolactifluorum
Subiculum	Pale yellowish-orange to bright orange (young), in age becoming deep red, reddish-purple to very dark purple (old), occasionally fading to pink, turning purple in 3% KOH.	Light yellow (4A4–5) when fresh, and pale orange to light orange to brownish-orange (5A3–4, 5C4, 6C6) after being dried, KOH (-).
Perithecia	Ovate to obpyriform, deep orange to reddish-purple, 400–600 × 200–450 μm	Pyriform to subglobose, yellowish-brown to dark brown (5E6, 6E6, 6F6–8), 262–484 × 136–284 μm
Embedded type	Immersed except for papilla	Semi-immersed to immersed except for papilla
Papilla	Averaging 120 μm high, 120 μm wide	129–177 μm high and 135 –284 μm wide at base
Asci	Long cylindrical, 200–260 × 5–10 μm	Cylindrical, 147–222 × 4–8.5 μm
Ascospores	Fusiform, 1-septate, 35–40 × 4.5–7 μm	Fusiform, 1-septate, 30–38 × 5.5–8 μm
Apiculi	4.5–7.5 μm long	4–6 μm long
Hosts	Russula and Lactarius spp.	Russula sp.
Distribution	North America	P.R. China (Yunnan)

Etymology

Referring to the most closely-related species,

Distribution

PR CHINA (Yunnan).

Host

On the fruiting bodies of sp. that grew on the humus layer in an evergreen broad-leaved forest of a rainforest. The host mushrooms: basidiocarps medium-sized and infundibuliform, pilei 63−77 mm in diameter. As serious degradation has occurred, the colour and other characters of the host mushrooms cannot be determined. Molecular phylogenetic evidence indicates it is a species.

Notes

Only sexual morph had been discovered on the hosts ( sp.) of the new species.

Analysis

Phylogenetic analyses

Parasitic fungus: sp. nov

The combined ITS+LSU+TEF1-α+RPB2 sequence dataset (excluding the outgroup taxa) contains 3,262 characters (709 for ITS, 893 for LSU, 921 for TEF1-α and 739 for RPB2) from 56 species and two species. Amongst them, 2,246 characters are constant, 209 variable characters are parsimony-uninformative and 807 characters are parsimony-informative. The ML and BI analyses resulted in trees with similar topology and support values and the ML tree is shown in Fig. 2.

Figure 2.

ML tree of sp. nov. and its allies generated from a combined ITS, LSU,TEF1-α and RPB2 gene sequence dataset. Supporting values of MLBP (left, greater than 75%) and BIBP (right, greater than 0.9) are shown at the nodes, respectively. The new species is marked in red.

In the phylogenetic tree, the parasitic fungi MFLU 20-0265 and MFLU 20-0266 are clustered together and formed a distinct lineage with the same branch length and strong supportive values (MLBP = 100%, BIPP = 1), which support them to be conspecific. The parasitic fungi are closely related and they form a sister clade also with strong supportive values (MLBP = 100%, BIPP = 1). Comparing the gene sequences of the two species, there are 25 bp (4.3%) differences across 582 bp in ITS, 28 bp (3.2%) differences across 870 bp in LSU, 24 bp (2.6%) differences across 921 bp in TEF1-α and 24 bp (3.2%) differences across 739 bp in RPB2 (Suppl. material 1). Following the recommendations from Jeewon and Hyde (2016), we assign the parasitic fungi as sp. nov.

The host mushroom:

According to the ITS phylogenetic tree of the host mushroom and its allies, the host mushroom (MFLU 20-0265) is clustered together with (HGAS-MF 009910 and HGAS-MF 009916) (MLBP = 100%) in subsect. . However, their ITS sequences have 24 bp (3.5%) differences across 694 bp, which indicated they may be two distinct species. Due to lack of sufficient morphological evidence, the host mushroom was temporarily identified as sp. (Fig. 3).

Figure 3.

ML tree of sp. (in red) and it allies inferred from the ITS sequence dataset. Five species of were used as the outgroup taxa. Supporting values of MLBP (greater than 75%) are shown at the nodes.

Discussion

Zeng and Zhuang (2016) described on sp. and and on sp., also from China. Though with similar colour and shapes of perithecia, the host of sp. nov. is decidedly different from those of these three species. Furthermore, sp. nov. (KOH-) has smaller perithecia and larger ascospores than those of (KOH+) and sp. nov. has larger perithecia, asci and ascospores than those of (KOH+) and (KOH-). Unfortunately, these three species all lack molecular data.

With the rapid development of mushroom industries, fungal pathogens on mushrooms have received more and more attention (Hyde et al. 2019). The fungicolous fungi is an important group of mushroom pathogens. Many species, for example, , , , etc., have all been recorded as the causes of Cobweb or Web bubble disease which seriously influence mushroom industries (Fletcher and Gaze 2007, Carrasco et al. 2017, Zhang et al. 2017, Zhang et al. 2017). is the largest subgenus in agaric with approximately 800 species (Li et al. 2020) and many species are important edible mushrooms. Since growing on sp., sp. nov., as well as from North America (Rogerson and Samuels 1994), could be one of the potential pathogens of some species in Asia.

sp. nov. (: ) on sp. from Yunnan, PR China

word

Sequence differences of ITS, LSU, TEF1-α and RPB2 genes between (TAAM 170476) and sp. nov.. The locus’ numbers refer to the nucleotide positions of the gene sequences of from GenBank. Gap is replaced by ‘-’.

File: oo_438866.docx