Multigene phylogeny and morphology reveal Ophiocordyceps hydrangea sp. nov. and Ophiocordyceps bidoupensis sp. nov. (Ophiocordycipitaceae) Author Zou, Weiqiu Yunnan Herbal Laboratory, College of Ecology and Environmental Sciences, Yunnan University, Kunming 650504, Yunnan, China Author Tang, Dexiang Yunnan Herbal Laboratory, College of Ecology and Environmental Sciences, Yunnan University, Kunming 650504, Yunnan, China Author Xu, Zhihong Yunnan Herbal Laboratory, College of Ecology and Environmental Sciences, Yunnan University, Kunming 650504, Yunnan, China Author Huang, Ou Yunnan Herbal Laboratory, College of Ecology and Environmental Sciences, Yunnan University, Kunming 650504, Yunnan, China Author Wang, Yuanbing Yunnan Herbal Laboratory, College of Ecology and Environmental Sciences, Yunnan University, Kunming 650504, Yunnan, China Author Tran, Ngoc-Lan School of Life Science, Yunnan University, Kunming 650504, Yunnan, China Author Yu, Hong https://orcid.org/0000-0002-2149-5714 Yunnan Herbal Laboratory, College of Ecology and Environmental Sciences, Yunnan University, Kunming 650504, Yunnan, China hongyu@ynu.edu.cn text MycoKeys 2022 2022-08-30 92 109 130 http://dx.doi.org/10.3897/mycokeys.92.86160 journal article http://dx.doi.org/10.3897/mycokeys.92.86160 1314-4049-92-109 934FF488EC855941A7EB52D0A4E15A1E Ophiocordyceps hydrangea H. Yu, W.Q. Zou & D.X. Tang sp. nov. Fig. 2 Etymology. Hydrangea, referred to the top of the stroma similar to Ophiocordyceps hydrangea . Holotype. China, Yunnan Province, Jinghong City, Nabanhe National Nature Reserve, 22°8'21.32"N , 100°42'18.35"E , alt. 612 m, on cicada nymphs ( Cicadidae , Hemiptera ). The material was found in the soil of an evergreen broad-leaved forest, 18 August 2020, H. Yu (YHH 20081, holotype; YFCC 8834, ex-holotype culture). Figure 2. Ophiocordyceps hydrangea A, B fungus on a cicada nymph C, D colony on PDA medium E conidiophores, conidiogenous cells and conidia F-J conidiogenous cells and conidia K conidia. Scale bars: 1 cm ( A, B ); 2 cm ( C, D ); 10 µm ( E, F, G, I, J ); 5 µm ( H, K ). Sexual morph. The stroma was grown from the head of the host cicada nymph, solitary, the top of the stroma similar to Ophiocordyceps hydrangea , pale pink, 1.6-6.4 cm long. Sexual morph was not observed. Asexual morph. The colony grew slowly on PDA medium. Cultured at 25 °C for about 12 weeks, the diameter of the colony was 25-28 mm, pale pink, the edge white, hard texture. The back of the colony was white to brown. Surface hyphae rough, hyaline, septate. Conidiophores were cylindrical. Conidiogenous cells were solitary or whorled, ampuliform, smooth-walled, forming on conidiophores or colonies, hyaline, with swollen base, and slender top, 10.6-17.6 µm long, 2.9-4.3 µm wide at the swollen base, and 1.1-2.2 µm wide at the slender top. Conidia hyaline, ovoid or long oval, solitary, 6.8-10.1 x 3.3-4.5 µm . Host. Cicada nymph ( Cicadidae , Hemiptera ). Habitat. In the soil of an evergreen broad-leaved forest. Distribution. China. Other material examined. China , Yunnan Province , Jinghong City , Nabanhe National Nature Reserve , 22°8'21.32"N , 100°42'18.35"E , alt. 612 m , on cicada nymphs ( Cicadidae , Hemiptera ) was found in the soil an evergreen broad-leaved forest, 18 August 2020 , H. Yu (YFCC 8832, YFCC 8833) . Notes. Phylogenetic analyses showed that O. hydrangea clustered with O. sobolifera , O. longissima , and O. yakusimensis of the O. sobolifera clade (Fig. 1 ). Their hosts were cicada nymphs compared to other species of the O. sobolifera clade (Table 2 ). Ophiocordyceps hydrangea was well supported by BI and ML results, forming a separate subclade with O. sobolifera , O. longissima , and O. yakusimensis . The macro-morphology of O. hydrangea was clearly different from O. sobolifera , O. longissima , O. khonkaenensis , and O. yakusimensis . The stroma of O. hydrangea grew from the head of the host cicada nymph, solitary, and the top of the stroma was like a pale pink Ophiocordyceps hydrangea . Table 2. Morphological comparisons of two new species and related species.

Species	Host	stromata	Perithecia	Asci	Ascospores	Conidiogenous cells	Conidia	References
O. bidoupensis	Larva of Elateridae ( Coleoptera )	Solitary, solid, cylindrical, yellow, 11.8-22.5 cm long.	Immersed, pyriform to lanceolate, brown-yellow, 213.4-405.9 x 74.8-192.4 μm .	Hyaline, slender, 116.1-192.7 x 4.8-7.5 μm .	Hyaline, filiform, multi-septate.	Cone, hyaline, septate, smooth-walled, forming on hyphae, with a hypertrophic base, tapering abruptly into a thin neck, smooth-walled, 13.8-46.4 x 0.42-5.13 μm .	Oval or briolette, hyaline, smooth-walled, 2.24-3.61 x 1.49-2.70 μm .	This study
O. brunneipunctata	Larva of Elateridae ( Coleoptera )	Solitary, rarely up to 3, simple, 25-90 mm high.	Immersed, perithecioid, brown, ovate to pyriform, brown-walled, 270-335 x 110-160 μm .	Hyaline, cylindric, capitate, 8-spored, 280-295 x 6-7 μm .	Hyaline, filiform, multiseptate breaking into 64 part spores, 4-6 x 1-1.5 μm .	Monophialidic, rarely polyphialidic, hyaline, smooth, 5.5-7.5 x 2.5-3.0 μm at the base, up to 15 x 0.5 μm above.	Hyaline, aseptate, smooth, spherical 1.5-2.5 μm diam., enveloped by a mucous sheath.	Hywel-Jones 1995b ; Luangsa-ard et al. 2008
O. cossidarum	Larva of Cossidae ( Lepidoptera )	Solitary, simple, 40-70 mm high.	Immersed, red, ovate to phialide, red-walled, 355-454 x 136-171 μm .	Hyaline, cylindrical, 8-spored with a thickened apex, 174-221 x 5.7-7 μm .	Hyaline, fifiliform, multiseptate,131-153 x 1.8-2.2 μm , breaking into 32 part-spores.	-	-	Hyde et al. 2017
O. furcatosubulata	Larva of Elateridae ( Coleoptera )	Single, solid, yellow to brown, 40-80 mm long, 1.5-2.2 mm wide.	Immersed, long ovoid or pyriform, 289.6-405.8 x 87.0-159.2 µm .	Hyaline, cylindrical, 138.8-202.5 x 4.3-6.0 μm .	Hyaline, filiform, multi-septate, finally breaking into secondary ascospores, 3.7-5.3 x 1.3-2.0 μm .	Polyphialidic, forming on conidiophores or side branches, hyaline, with a slender or subulate base, tapering gradually, smooth-walled or verruculose, 3.5-15.8 x 0.9-1.7 μm .	Solitary, aseptate, smooth-walled, broadly ellipsoid or ellipsoid, 1.5-2.5 x 1.2-1.9 μm .	Wang et al. 2021a
O. houaynhangensis	Larva of Coleoptera	Solitary, cylindrical, cream, up to 11 cm long and 1.5-2.5 mm in width.	Completely immersed, obclavate, 300-450 x 80-170 µm .	Cylindrical, 100-250 x 4-7.5 µm .	Hyaline, cylindrical, breaking into 32 small truncate part-spores, 4-7 x 1-2 µm .	Monophialidic, phialides flasked-shaped with long necks, up to 30 µm long and 2-4 µm in breadth; phialide necks up to 18 μm long and 0.5 µm in breadth.	Hyaline, smooth, spherical, 2-3 µm .	Crous et al. 2018
O. langbianensis	Larva of Coleoptera	Solitary, rarely branched, 40-100 mm long.	Immersed, ovate or pyriform, 260-400 x 100-190 µm .	Cylindrical, with thickened cap, 200-250 x 5.0-6.0 μm .	Fliform, multiseptate, articulated in long-chain afer discharging, sometimes breaking into 1-celled part spores, 5-7.5 x 1.3-2 µm .	Divergent.	Chains, elliptical.	Lao et al. 2021
O. sobolifera	Cicada nymph ( Cicadidae , Hemiptera )	Commonly single, rarely fasciculated by twos or threes, arising from head among polster, clavate or cylindric 2-8 cm long, 2-6 mm thick, become hollow after maturity.	Rectangularly immersed, ampullaceous 500-600 x 220-260 μm , with somewhat long neck, ostiola somewhat prominent, walls hyaline 8-16 μm thick.	Cylindric, 400-470 x 5.6-6.3 μm .	Finally breaking into secondary ascospores, truncate at both ends, 6-12 x 1.0-1.3 μm .	-	Terminal or lateral, ellpsoid or fusiformed, hyaline, 6.5-10.5 x 2.5-4.0 μm .	Kobayasi and Shimizu 1963
O. yakusimensis	Cicada nymph ( Cicadidae , Hemiptera )	Very long attaining 14 cm, arising from the apical part between eyes.	Wholly embedded , narrow ovoid or almost naviculate, 740-800 x 170-230 μm , without protruding ostiola, neck almost destitute, wall 21-23 μm thick, composed of very thin cells.	270-310 x 5 μm .	Finally breaking into secondary ascospores, long cylindrical, somewhat attenuated on both sides, terminally truncate, 10-15 x 1 μm .	-	-	Kobayasi and Shimizu 1963
Ophiocordyceps longissima	Cicada nymph ( Cicadidae , Homoptera )	5-20 cm long, some times much longer.	Ovoid to long ovoid, with a short neck, 440-590 x 130-300 µm .	190-350 x 5-6 µm .	-	-	-	Sung et al. 2011
O. khonkaenensis	Cicada nymph ( Hemiptera )	Variable in number, solitary to three, 20-30 mm long and 2-3 mm in breath.	Immersed, flask shaped, 590-700 x 200-300 µm .	Cylindrical, 237.5-337.5 x 5-6 µm .	Filiform, 300-360 x 1-1.5 µm readily breaking into 32 part-spores, 7-13 x 1-1.5 µm .	Phialidic, hirsutella-like, 5.5-11 x 2-3 µm .	Hyaline, fusiform, smoothwalled, 3-5.5 x 1-3 µm .	Crous et al. 2019
O. hydrangea	Cicada nymph ( Cicadidae , Hemiptera )	Solitary, the top of the stroma similar to Ophiocordyceps hydrangea , pale pink,1.6-6.4 cm long.	-	-	-	Solitary or whorled, ampuliform, smooth-walled, forming on conidiophores or colonies, hyaline, with swollen base, and slender top, 10.6-17.6 µm long, 2.9-4.3 µm wide at the swollen base, and 1.1-2.2 µm wide at the slender top.	Hyaline, ovoid or long oval, solitary, 6.8-10.1 x 3.3-4.5 µm .	This study