Two Hypoxylon species from Yunnan Province based on morphological and molecular characters Author Ma, Hai-Xia Institute of Tropical Bioscience and Biotechnology, Chinese Academy of Tropical Agricultural Sciences, Hainan Key Laboratory of Tropical Microbe Resources, Haikou 571101, China Author Qiu, Jun-Zhi College of Life Sciences, State Key Laboratory of Ecological Pest Control for Fujian and Taiwan Crops, Fujian Agriculture and Forestry University, Fuzhou, Fujian 350002, China Author Xu, Biao College of Life Sciences-Province-Ministry Joint Key Laboratory of Protection and Utilization of Biological Resources in Tarim Basin, Tarim University, Aral 843300, China Author Li, Yu Institute of Mycology, Jilin Agricultural University, Changchun 130118, China text Phytotaxa 2018 2018-11-15 376 1 27 36 http://dx.doi.org/10.11646/phytotaxa.376.1.3 journal article 10.11646/phytotaxa.376.1.3 1179-3163 13726618 Hypoxylon baihualingense Hai X. Ma & Yu Li , sp. nov. ( Fig.1 , Fig.2 ) MycoBank: MB 823544 FIGURE 1. Hypoxylon baihualingense (FCATAS477, Holotype). a,b Stromatal habit. c Close-up view of stromatal surface, showing perithecial mounds and ostiolar disks. d Section through stroma, showing perithecia. e Perithecia. f Pigments in KOH. Scale is indicated by bars (a,b = 1 mm; c,d,e = 0.5 mm; f = 0.5 cm). Etymology .—Referring to the type locality Baihualing. Holotype . — CHINA . Yunnan Province , Baoshan , Baihualing , alt. 1260 m , on bark of a fallen branch in evergreen forest with broadleaved trees, 30 November 2015 , Ma Hai X. , H14 ( FCATAS477 ). Teleomorph.— Saprobic on corticated branch. Stromata scattered, glomerate to pulvinate, 2.5–16 mm diam × 1–2.5 mm thick, soft-textured; surface Rust (39) to dark Brick (60), pruinose, with perithecial contours exposed to strongly exposed; dull yellow or rust granules beneath the surface and between the perithecia, with yielding Orange (7), Sienna (8), and Amber (47) pigments in 10% KOH; subperithecial tissue black, brownish black at base, 0.4–1.1 mm thick. Perithecia subglobose, 0.35–0.5 mm diam. Ostioles umbilicate, opening at the centre of discoid area 0.25–0.3 mm diam. Asci with eight ascospores obliquely arranged in uniseriate manner, cylindrical, 135–184 μm total length, the spore-bearing parts 84–109 μm long × 9.2–12 μm broad, the stipes 40–80 μm long, without apical apparatus, not bluing in Melzer’s iodine reagent. Ascospores brown to dark brown, unicellular, ellipsoid-inequilateral, with narrowly rounded ends, smooth, 9.2–15.6 × 5.5–7.5 μm (M = 12.9×6.4 μm, n = 30), with spore-length straight germ slit; perispore dehiscent in 10% KOH, with conspicuous coil-like ornamentation; epispore smooth. Remarks.— Hypoxylon baihualingense is mainly characterized by its ostiolar discs and asci lacking an apical apparatus. Fournier et al . (2010) described two species, H. addis J. Fourn., M. Stadler & U. Lindem and H. gibriacense J. Fourn., M. Stadler & Gardiennet , which have similar ostiolar discs and asci without apical apparatus. This group of Hypoxylon species has clearly discs around the ostioles in stromata, however, they are placed in the genus Hypoxylon rather than Annulohypoxylon , because they have soft-textured stromata, ascospores with transversally dehiscent and striate perispores without a dorsal thickening. The recognition of these taxa in the genus Hypoxylon is also strongly supported by the analyses of ITS rDNA and β- tubulin sequences. FIGURE 2. Hypoxylon baihualingense (FCATAS477, Holotype). a Asci in water. b matured asci in Melzer’s reagent. c immatured asci in Melzer’s reagent. d Ascospore with germ slit in KOH. e Ascospore with germ slit in water. f Ascospore in KOH with dehiscent perispore showing conspicuous ornamentation. g Free perispores in KOH. h Ascospores in water. i Ascospores in KOH with dehiscent perispore. Scale is indicated by bars (a = 20 μm; b,c = 10 μm; d-i = 5 μm). Undoubtedly, H. baihualingense closely resembles H. addis with regard to macro- and micro-morphological characters. H. addis was originally described and illustrated by Fournier et al . (2010) from Ethiopia . The fungus grew on a corticated dry twig of Croton sylvaticus , and was characterized by its glomerate stromata with conspicuous discs around the ostioles, asci lacking apical ring and not bluing in Melzer’s iodine reagent, larger ascospores with inconspicuous ornamentation. However, based on comparisons of the descriptions and illustrations, there were some differences in the two species. The Chinese collections has larger stromata 2.5–16 mm diam × 1–2.5 mm thick, slightly smaller perithecia, and a broad range of ascospores dimensions 9.2–15.6 × 5.5–7.5 μm, slightly smaller mean ascospores dimensions 12.9 × 6.4 μm, with conspicuous coil-like ornamentation, while H. addis has smaller stromata 1–3 mm diam × 1–1.2 mm thick, perithecia 0.5–0.55 mm diam, and slightly larger ascospores 13–16.5 × 6–7.7 μm (M = 14.6 × 7 μm), with striae perispore but inconspicuous ( Fournier et al . 2010 ). The molecular phylogeny did not reveal any contradictions in relation to the morphological data (Kuhner et al . 2014). Hypoxylon baihualingense and H. addis cluster together in a subclade, was statistically supported by the combined datast (ITS +β- tubulin) analysis. The sequence comparison showed that 6bp and 7bp divergences existed in the ITS1 and ITS2 region between the Chinese material (FCATAS477) and the collections from Ethiopia (MUCL 52797) ( Kuhnert et al . 2014 ), and with 2281 max scores and 96% maximal percent identities in β- tubulin sequence, respectively. Therefore, we described the Chinese materials as a different taxon, despite the similar stromatal and teleomorphic morphology of H. addis . They may eventually be revealed by using a phylogenetic multi-gene genealogy approach to be two different taxa that have evolved from a common ancestor, due to long geographic separation. Hypoxylon baihualingense shows morphological similarities to H. gibriacense with orange to sienna pigments in KOH and similar asci, but differs in having Rust (39) or dark Brick (60) instead of Greyish Sepia (106) stromatal surface colour, larger ascospores (9.2–15.6 × 5.5–7.5 vs. 11.5–13 × 6–6.8 μm), slightly smaller subglobose perithecia ( 0.35–0.5 mm diam) instead of ellipsoid to subglobose perithecia ( 0.5–0.6 mm high × 0.4–0.5 mm diam), and slightly larger ostiolar discs (0.25–0.3 vs. ca. 0.2 mm diam) ( Fournier et al . 2010 ). The phylogenetic analysis showed that H. baihualingense from China clustered together with H. gibriacense from France in a clade, but the two species did not have a close relationship in the phylogenetic tree and they are phylogenetically distinct species ( Fig. 5 ). The Chinese collections roughly resembles Hypoxylon shearii Y.M. Ju & J.D. Rogers (Ju & Rogers) from USA in ascospore morphology and the color of its stromatal pigments, but the latter has smaller stromata 0.8–3 mm diam × 0.8–1 mm thick, surface fawn or vinaceous buff when mature, the tissue below the perithecial layer inconspicuous, and larger perithecia 0.4–0.7 mm diam. In addition, the specimens of H. shearii have been only found on wood of Quercus . Unfortunately, the molecular sequences of H. shearii were not available. Material of the type of H. shearii was studied by HPLC and found to contain mitorubrins by Stadler et al . (2008) , we were not able to study the HPLC for the new species. We compared the Chinese collections with H. shearii var. minor F. San Martín et al . (1999) , which was also collected from a branch of Quercus , and it can be separated from the new species by having smaller ascospores 7–8 × 3.5–4 μm. The phylogeny did not reveal any contradictions in relation to the morphological data ( Hsieh et al . 2010 ).