==== Front MycoKeys MycoKeys 11 urn:lsid:arphahub.com:pub:C004A564-9D6A-5F9F-B058-6A3815DFE9C3 urn:lsid:zoobank.org:pub:A4FD9303-3F01-45D9-B438-FCCFE465D2F7 MycoKeys 1314-4057 1314-4049 Pensoft Publishers 10.3897/mycokeys.98.104314 104314 Research Article Acarosporaceae Acarosporales Phylogeny Taxonomy Asia Central Asia China The phylogeny and taxonomy of Glypholecia (Acarosporaceae, lichenized Ascomycota), including a new species from northwestern China Yin An-cheng https://orcid.org/0009-0000-7837-7797 1 Zhong Qiu-yi https://orcid.org/0000-0001-6256-6083 1 Scheidegger Christoph https://orcid.org/0000-0003-3713-5331 2 Jin Ji-zhen https://orcid.org/0009-0005-4351-8162 13 Worthy Fiona R. https://orcid.org/0000-0003-0042-3110 1 Wang Li-song https://orcid.org/0000-0003-3721-5956 1 Wang Xin-yu wangxinyu@mail.kib.ac.cn https://orcid.org/0000-0003-2166-6111 1 1 CAS Key Laboratory for Plant Diversity and Biogeography of East Asia, Kunming Institute of Botany, Chinese Academy of Sciences, Kunming, Yunnan 650201, China 2 Yunnan Key Laboratory for Fungal Diversity and Green Development, Kunming Institute of Botany, Chinese Academy of Sciences, Kunming 650201, China 3 Snow and Landscape Research (WSL), Biodiversity and Conservation Biology, Swiss Federal Institute for Forest, Zürcherstrasse 111, 8903 Birmensdorf, Switzerland 4 Key Laboratory of Plant Stress Research, College of Life Sciences, Shandong Normal University, Jinan, Shandong, 250014, China Corresponding author: Xin-yu Wang (wangxinyu@mail.kib.ac.cn) Academic editor: Gerhard Rambold 2023 21 6 2023 98 153165 0AD41F8C-3CCA-5F6A-B830-E0C02DB3352A31 3 2023 05 6 2023 An-cheng Yin, Qiu-yi Zhong, Christoph Scheidegger, Ji-zhen Jin, Fiona R. Worthy, Li-song Wang, Xin-yu Wang https://creativecommons.org/licenses/by/4.0/ This is an open access article distributed under the terms of the Creative Commons Attribution License (CC BY 4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Abstract Glypholeciaqinghaiensis An C. Yin, Q. Y. Zhong & Li S. Wang is described as new to science. It is characterized by its squamulose thallus, compound apothecia, ellipsoid ascospores, and the presence of rhizines on the lower surface of the thallus. A phylogenetic tree of Glypholecia species was constructed based on nrITS and mtSSU sequences. Two species G.qinghaiensis and G.scabra are confirmed in China. Key words: cosmopolitan lichenized fungi morphological diversity phylogenetic analyses Tibetan Plateau Kunming Institute of Botany, Chinese Academy of Sciences 501100011190 http://doi.org/10.13039/501100011190 Citation Yin A-c, Zhong Q-y, Scheidegger C, Jin J-z, Worthy FR, Wang L-s, Wang X-y (2023) The phylogeny and taxonomy of Glypholecia (Acarosporaceae, lichenized Ascomycota), including a new species from northwestern China. MycoKeys 98: 153–165. https://doi.org/10.3897/mycokeys.98.104314 ==== Body pmcIntroduction Glypholecia Nyl. is a genus of lichenized fungi belonging to Acarosporaceae, Acarosporales, Acarosporomycetidae, Lecanoromycetes, Ascomycota (Nylander 1853; Wijayawardene et al. 2022). The genus Glypholecia is characterized by its squamulose-subfoliose, peltate-subumbilicate thallus, compound small apothecia forming multiple structures, and multi-spored asci, generally exceeding 30 spores per ascus. It is therefore distinct from the genus Acarospora, which has areolate or squamulose thallus, mostly single or several assembled apothecia, and multi-spored asci, generally exceeding 100 spores per ascus (Ajaj et al. 2007; Sohrabi et al. 2019). The first known specimen of the genus now designated Glypholecia, was collected in the summer of 1810, by Balbis G. B., who was working at the Botanical Gardens of Turin, Italy. Balbis sent this lichen specimen to Germany, where it was received by Funck H. C., who labelled the specimen as Balbis. This name was never published. Later, Persoon H. C. examined this same specimen in France, and described it as a new species Urceolariascabra Pers. (Persoon 1810). In 1814, Acharius E. published the lichen species Lecanorarhagadiosa Ach (Acharius 1814). In 1850, Schaerer L. E. added an additional lichen species to this genus: Lecanoragrumulosa Shaer (Schaerer 1850). In 1853, Nylander W. established the monotypic genus Glypholecia, based on the type species Glypholeciacandidissima Nyl. (Nylander 1853). In 1871, Fries T. M. treated these four species names (Urceolariascabra Pers., Lecanorarhagadiosa Ach, Lecanoragrumulosa Shaer and Glypholeciacandidissima Nyl.) as synonyms of Acarosporascabra (Pers.) Th. Fr. (Fries 1871). In 1892, Müller Arg. treated Urceolariascabra and Acarosporascabra as synonyms, and designated them as G.scabra (Pers.) Müll. Arg. (Müller 1892). The subspecies G.scabravar.candidissima (Nyl.) H. Magn. was published; conserving the name G.candidissima Nyl. (Hue 1909; Steiner 1921; Zahlbruckner 1940; Lamb 1963). Recently published phylogenies split Acarosporaceae into 6 main clades, including Acarospora (Westberg et al. 2015). Knudsen split the previous Acarosporaglaucocarpa group into the North American Sarcogynecanadensis-wheeleri clade and the European Acarosporaglaucocarpa group (Knudsen et al. 2020). They placed G.scabra within the Sarcogynecanadensis-wheeleri clade. Accordingly, most lichenologists accept only one species G.scabra (Pers.) Müll. Arg. as belonging to the genus Glypholecia. The distribution of G.scabra is largely disjunct, with intercontinental populations. It occurs on siliceous rocks in desert, alpine regions, including Europe, Africa, Asia, and North America (Magnusson 1940; Thomson 1979, 1984; Ryan 2002). It exhibits extensive morphological diversity across its range. Previously, a second species, G.tibetanica H. Magn., was described as endemic to China. It has only been recorded from Xizang province, China (Zahlbruckner 1933). However, Obermayer (2004) suggested that G.tibetanica might belong to Acarosporanodulosavar.reagens (Zahlbr.) Clauzade and Cl. Roux, which has also been reported from China, based on the K reaction of the cortex. But Magnusson H. appears to have incorrectly reported a ‘C’ reaction (to calcium hypochlorite) as a KOH reaction (potassium hydroxide). According to Zahlbruckner’s (1933) description, the cortex of G.tibetanica should show a reddish C+ reaction due to gyrophoric acid, whereas A.nodulosavar.reagens shows a red K+ reaction caused by norstictic acid (Magnusson 1940; Zahlbruckner 1933; Cao and Wei 2009). Unfortunately, the holotype of G.tibetanica has been lost, so there are no materials available for further research regarding this putative species. During the Second Tibetan Plateau Scientific Expedition and Research Program (STEP), we collected numerous lichen specimens from across northwestern China. The expedition included many field surveys across the type locality of G.tibetanica, during which we discovered some specimens that conformed to Zahlbruckner’s (1932) description. Our new specimens differ from A.nodulosavar.reagens in the characteristics of their upper cortex, hymenium chemical reaction and their number of ascospores. The molecular sequences obtained from these new specimens demonstrate that they belong to the genus Glypholecia, rather than to Acarospora (Knudsen et al. 2020). Of these, some samples had squamulose thalli, compound apothecia and white rhizines. These were confirmed as belonging to a new species of the genus Glypholecia. In this paper, we describe a new species, Glypholeciaqinghaiensis, from northwestern China. Materials and methods Morphological and chemical study We examined materials of Glypholecia from the lichen herbaria of the Kunming Institute of Botany (KUN-L), the College of Life Science and Technology, Xinjiang University (XJU), and the Swiss Federal Institute for Forest, Snow and Landscape Research, Switzerland (WSL). We made morphological observations of the specimens using a Nikon SMZ 745T (Nikon Corp., Tokyo, Japan) dissecting microscope. We cut vertical sections of apothecia and thalli using a razor blade, mounted sections in GAW (glycerol: ethanol: water = 1: 1: 1), then examined them under a Nikon Eclipse 50i stereomicroscope. We measured the average spore size and described sections under both the microscope and stereomicroscope. We photographed all specimens with a Nikon digital camera head DS-Fi2. We identified secondary metabolites by their color reaction coupled with thin-layer chromatography (TLC), using solvent system C (toluene: acetic acid = 85:15), following the methods of Culberson (1970) and Orange et al. (2001). DNA extraction, purification and sequencing We extracted total genomic DNA from 20 specimens (19 from China and one from Switzerland) using the DNAsecure Plant Kit (Tiangen Biotech, Beijing) following the manufacturer’s protocol. We amplified the internal transcribed spacer regions (nrITS) with the primer pairs ITS1F (Gardes and Bruns 1993) and ITS4 (White et al. 1990). We amplified the mitochondrial small subunit (mtSSU) with primer pairs SSU1 and SSU3R (Zoller et al. 1999). We performed PCR amplification with 25 μL volume containing: 12.5 μL 2× MasterMix (0.1 units/μL TaqDNA polymerase, 4 mM MgCl2, and 0.4 nM dNTPs; Aidlab Biotechnologies Co. Ltd), 1 μL of each primer, 9.5 μL of ddH2O, and 1 μL of DNA, following the PCR settings and primer profile of Zhao et al. (2015). Polymerase chain reaction (PCR) products were sequenced by TsingKe Biological Technology company (Kunming, China). Phylogenetic analysis We aligned DNA sequences using the program MAFFT v. 7.107 in GENEIOUS v. 8.0.2, setting the following parameters: algorithm = auto; scoring matrix = 200 PAM / k=2; gap open penalty = 1.53; offset value = 0.123 (Katoh et al. 2005). We conducted single-gene analyses to test for potential incongruence among the two-gene fragments, using maximum likelihood (ML) analyses and Bayesian inference (BI). We generated a matrix of Glypholecia and its related genera using GENEIOUS v. 8.0.2. In addition to the DNA sequences obtained from our own material, we also downloaded all available sequences of Glypholecia from GenBank at the National Center for Biotechnology Information (NCBI, https://www.ncbi.nlm.nih.gov/), and added these to the matrix. We performed ML analyses within RaxML v. 8.2.12 (Stamatakis 2014), using the General Time Reversible model of nucleotide substitution with the gamma model of rate heterogeneity (GTRGAMMA). We selected best partitioning scheme and evolutionary models for two pre-defined partitions using PartitionFinder2 (Lanfear et al. 2016), with greedy algorithm and AICc criterion. We used PhyloSuite (Zhang et al. 2020) inferred Bayesian Inference phylogenies using MrBayes 3.2.6 (Ronquist et al. 2012) under a partition model (2 parallel runs, 10 million generations), for which the initial 25% of sampled data were discarded as burn-in. We inferred support values from the 70% majority-rule tree of all saved trees obtained from 1000 non-parametric bootstrap replicates. We obtained Posterior Probabilities (PPs) from the 95% majority rule consensus tree of all saved trees. We visualized tree files using FigTree 1.4.4. Results In the present study we generated twenty new nrITS and eighteen new mtSSU sequences. We constructed ML and BI topologies based on these nrITS and mtSSU sequences, and nine additional sequences downloaded from GenBank (Table 1). We used Pleopsidium as the outgroup (Crewe et al. 2006; Reeb et al. 2007; Schmull et al. 2011). In the phylogenetic tree, the Glypholecia specimens formed a monophyletic lineage, which was divided into two clades, representing G.scabra in Clade 2 and the new species G.qinghaiensis in Clade 1. The result showed high support for new specimens in this study being assigned to Glypholecia (99% ML and 1.00 PP, Fig. 3). Table 1. Specimens and sequences used for phylogenetic analyses. Newly generated sequences are in bold. Taxon Locality Voucher specimens GenBank number (nrITS) GenBank number (mtSSU) References G.qinghaiensis Ningxia, China KUN-L 10-0241 MZ330798 OP749902 – Gansu, China KUN-L 18-58434 MZ330797 OP749903 – Gansu, China KUN-L 18-59534 MZ330793 OP749907 – Qinghai, China KUN-L 20-68255 MZ330789 OP749910 – Xinjiang, China KUN-L 22-71630 OP749916 OP749899 – G.scabra Neimenggu, China XJU 20157514-a MZ330786 – – Gansu, China KUN-L 18-58747 MZ330796 OP749904 – Qinghai, China KUN-L 18-59190 MZ330792 OP749906 – Gansu, China KUN-L 18-58881 MZ330795 OP749905 – Xizang, China KUN-L 19-65418 MZ330791 OP749908 – Xizang, China KUN-L 19-66159 MZ330790 OP749909 – Xinjiang, China KUN-L XY22-856 OP749911 OP749895 – Xinjiang, China KUN-L XY22-856-2 OP749912 OP749896 – Xizang, China KUN-L XY22-584 OP749913 OP749894 – Xinjiang, China KUN-L 22-72868 OP749914 OP749897 – Xinjiang, China KUN-L 22-71693 OP749915 OP749898 – Xizang, China KUN-L 22-71500 OP749917 OP749900 Xizang, China KUN-L 22-71500-2 OP749918 OP749901 – Xizang, China KUN-L 22-71435 OP749919 OP749893 – Canton of Valais, Switzerland Scheideg-10522 MZ330788 – – Unknown AFTOL 1008 HQ650722 – Schmull et al. 2011 Oppland, Norway S. Westberg 08-232 LN810811 LN810936 Westberg et al. 2015 Acarospora sp. Gansu, China Huang Manrong GS157 FJ919810 – Cao and Wei 2009 A.placodiiformis Spain, Madrid Westberg 10-211 LN810795 LN810920 Westberg et al. 2015 A.schleicheri Bouches-du-Rhône, France DUKE Reeb VR5-VII-98/30 DQ525529 – Reeb et al. 2007 Sichuan, China UPS L-070426 LN810800 LN810925 Westberg et al. 2015 Arizona, USA UPS L-162697 LN810801 LN810926 Westberg et al. 2015 Pleopsidiumflavum Steiermark, Austria UPS L-105590 AY853385 AY853336 Crewe et al. 2006 P.chlorophanum Jämtland, Sweden UPS L-179248 LN810813 LN810938 Westberg et al. 2015 Our phylogenetic results indicated that Glypholecia is monophyletic in China. The genus Glypholecia is characterized by its squamulose to crustose thallus, compound apothecia forming multiple structures, multi-spored asci (usually exceeding 30 spores per ascus), spherical small ascospores and C+ red reaction of the cortex due to the presence of gyrophoric acid. Species of Glypholecia were separated into two main clades, as inferred from the phylogenetic tree with strong support. Based on the combination of morphological characters and phylogenetic analysis, we propose a new species in Glypholecia. We divide the specimens of the genus Glypholecia collected from China into two clades, which correspond to differences in the lower surface morphology of the thallus: the specimens in Clade 1 have ellipsoid ascospores (2.5–3 × 4–6.5 μm), and rhizines on the lower surface of the thallus, whereas Clade 2 contains specimens with spherical ascospores (3–4 µm), and a central holdfast, called an umbilicus. Within Clade 2, our samples of G.scabra collected from China were clustered with the European samples, but with some genetic divergence. All Glypholecia specimens which we collected from the type locality of “G.tibetanica” were monophyletic with G.scabra, with a high support value. Within Clade 1, those specimens assigned as G.qinghaiensis lack genetic variability within the gene regions included in this study. This might be due to either the sparse population or the shared geography and environment. All of our G.qinghaiensis specimens were collected from arid to semi-arid areas of northwestern China, usually growing on sandy rock or sandy soil. Our phylogenetic analyses showed that a specimen from Gansu province in China, which was identified as G.scabra by Cao and Wei (2009), was clustered with Acarosporaschleicheri (the type species of Acarospora) in Clade 3. Therefore, this specimen’s previous identification as G.scabra was incorrect. The new species Taxon classification Fungi Acarosporales Acarosporaceae  Glypholecia qinghaiensis An C. Yin, Q. Y. Zhong & Li S. Wang, sp. nov. 556592F6-76BF-5278-AAED-14F04F53D61D 839606 Fig. 1 青海聚盘衣 Remark. Resembles G.scabra, has abundant and compound apothecia, but differs in having ellipsoid ascospores, rhizines, and a different molecular fingerprint (based on nrITS and mtSSU data). Type. China, Qinghai Prov., Haixi Mongolian and Tibetan Autonomous Prefecture, Dulan Co., 3066 m, on sandy soil, 2020-09-15, Wang Lisong et al. 20-68255 (holotype – KUN). Thallus squamulose to squamulose-subfoliose, peltate-subumbilicate, up to 6 mm in diam., 6.5–7 mm thick, with margins sometimes rolling under; surface: upper surface white, pale brown to brown, cracked and wrinkled, usually warty, partly pruinose or occasionally densely pruinose at margins; lower surface white, gray to pale brown, rough, wrinkled, rhizines, umbilicate, 3–6 mm long, attached with a broad; upper cortex: paraplectenchymatous, pale brown, containing calcium oxalate crystals, 30–70 μm thick; medulla: pale, containing calcium oxalate crystals, 125–250 μm thick; hyphae loose, arachnoid, 2.5–3 μm. Apothecia very common, lecanorine, punctiform when young, but later becoming compound reddish brown to dark brown, becoming lower than the thallus surface when mature; disc with cracks and sometimes tuberculous; paraphyses septate, 2–2.5 μm in diam.; margins concolorous with the thallus; asci: clavate, c. (30–)50-spored; ascospores: ellipsoid, 4–6.5 × 2.5–3 μm, hyaline, thick wall. Pycnidia: rare, flask-shaped. Conidia bacilliform, c. 2–3 × 1 µm. Chemistry. Hymenium: I+ blue; cortex and medulla: K–, C+ red, KC+ red, P–; secondary metabolites: gyrophoric acid. Ecology and distribution. Usually on sandy rocks or rarely on soil over rocks; so far only known from Gansu, Ningxia, Qinghai and Xizang provinces in China; growing in arid to semi-arid areas; distributed from 1600 to 4700 m altitude. Etymology. The epithet “qinghaiensis” refers to the holotype locality of the species. Notes. Glypholeciaqinghaiensis can be distinguished from G.scabra by having ellipsoid ascospores and rhizines. Phylogenetic analysis in this study supports the separate classification of these two species. This species has only been reported from northwestern China, including Gansu, Ningxia, Qinghai and Xizang provinces. Specimens examined. China (stored in KUN). Gansu Prov.: Jiuquan City, Yumen City, Yuerhong Vil., 3044 m, on rock, 2018-05-27, Wang Lisong et al. 18-59534; Subei Mongolian Autonomous Co., Suyan Line, 2376m, on soil over rock, 2018-05-23, Wang Lisong et al. 18-58434; Ningxia Prov.: Zhongwei Co., Suwumuyang Site, 1611 m, on rock, 2010-09-19, Niu Dong-Ling et al. 10-0241. Qinghai Prov.: Dulan Co., 3066 m, on sandy rock, 2020-09-15, Wang Lisong et al. 20-68255; Xinjiang Prov.: Wuqia Co., Fossil hill, 2559 m, on sandy rock, 2022-06-26, Wang Lisong et al. 22-71630. 10.3897/mycokeys.98.104314.figure1 D0B75ECF-0DCE-5367-8D9A-A3DA974192A3 Figure 1. Morphology and anatomy of GlypholeciaqinghaiensisA upper surface of the thallus B section of the thallus and apothecia (Lugol’s) C paraphyses and asci D ascospores. Scale bars: 1 mm (A); 50 μm (B); 10 μm (C); 5 μm (D). https://binary.pensoft.net/fig/866963 Species of Glypholecia reported in China Taxon classification Fungi Acarosporales Acarosporaceae  Glypholecia scabra (Pers.) Müll. Arg., Hedwigia 31: 156, 1892. 3103D9F7-EC69-5422-90C5-18AAB1A456DD ≡ Urceolariascabra Pers., Ann. Wetter. Gesellsch. Ges. Naturk. 2: 10, 1810. Type. Monte Cenisio [in Alps between France and Italy], s. d., Balbis s. n. (not seen). Glypholeciascabra is characterized by its squamulose thallus, abundant and compound apothecia, and umbilicate lower surface, as shown in Fig. 2A–F, J–L. In the Qinghai-Tibetan Plateau, these specimens have an umbilicus at the center of the lower surface, formed by fasciculate white rhizines, which are single or branched, dense or loose. It has a global distribution (see citations above). Within China it has been reported from Gansu (Magnusson 1940; Cao and Wei 2009), Xinjiang (Wang 1985; Abbas et al. 1993; Abbas and Wu 19941998), Ningxia (Liu and Wei 2013) and Xizang (Wei and Jiang 1986) provinces. Delimiting species boundaries can be complicated by the potential role of the environment in shaping morphology. Our phylogenetic study showed that a specimen previously collected and reported from Gansu (GenBank number: FJ919810) did not belong to the genus Glypholecia, but should rather be placed within Acarospora. We also report a new record for G.scabra in Neimenggu province, northern China. For further synonyms and detailed descriptions of G.scabra, see Thomson (1979) and Ryan (2002). 10.3897/mycokeys.98.104314.figure2 56EE6E03-2126-5FF4-A3BA-2BEB039BF0A6 Figure 2. Morphological diversity within the genus GlypholeciaA–FGlypholeciascabra. Differing morphology putatively caused by different habitat conditions, e.g., high or low temperature, aridity, different altitudes (1300–5100 m) G–IG.tibetanica. Collected from type locality, thallus with numerous pycnidia. J-F Umbilicus at the lower surface, formed by fasciculate white rhizines A, J Wang Lisong KUN 18-58925 B Wang Lisong KUN 18-58814-b C Wang Lisong KUN 18-58820 D Wang Lisong KUN 19-65418 E, K Wang Lisong KUN 18-59346 F Wang Lisong KUN 18-58747 G Wang Xinyu KUN XY22-854 H Wang Lisong KUN 22-71500 I, L Wang Xinyu KUN XY22-856. Scale bars: 1 mm. https://binary.pensoft.net/fig/866965 10.3897/mycokeys.98.104314.figure3 8509E05D-618E-52F8-8A30-4CD64229EF0C Figure 3. Maximum likelihood (ML) phylogeny of the genus Glypholecia and related species of Acarosporaceae, based on nrITS and mtSSU sequences. ML bootstrap value ≥ 70% and posterior probabilities ≥ 0.95 from the Bayesian analysis are displayed adjacent to nodes. https://binary.pensoft.net/fig/866964 Specimens examined. China. Gansu Prov. (stored in KUN): Zhangye City, Sunan Yugur Autonomous Co., on the way from Sunan to Qilian, hinterland of Qilian Mt., 3958 m, on rock, 2018-05-30, Wang Lisong et al. 18-58881; near Binggoudanxia Geopark, 1984 m, on rock, 2018-05-29, Wang Lisong et al. 18-58747. Qinghai Prov. (stored in KUN): Hainan Tibetan Autonomous Region, Gonghe Co., Heimahe Vil., 3429 m, on rock, 2018-05-19, Wang Lisong et al. 18-59190; Xining City, Huangyuan Co., on the way from Xining to Qinghai Lake, 2476 m, on rock, 2018-05-18, Wang Lisong et al. 18-59094. Neimenggu Prov. (stored in XJU): Alashan League, 1342 m, 2015-08-16, Hurnisa Xahidin 20157514-a. Xinjiang Prov. (stored in XJU): eastern Tianshan Mt., Miquan Tree Farm, 1959 m, 2015-06-28, Hurnisa Xahidin 20155538. Xizang Prov. (stored in KUN): Dingjie Co., Riwu Town, 4848 m, on rock, 2019-07-28, Wang Lisong et al. 19-66159; Cuoqin Co., 5015 m, on rock, 2019-07-20, Wang Lisong et al. 19-65418. Switzerland (stored by C. Scheidegger). Canton of Valais: Evolène, Mount Le Tsaté., 2492 m, on calcareous rocks, s. d., C. Scheidegger Scheideg-10522. Taxon classification Fungi Acarosporales Acarosporaceae  Glypholecia tibetanica H. Magn., Feddes Repert. Spec. Nov. Regni veg. 31: 24, 1932. 514C3496-C736-581F-908E-C8E20FE58924 Type. Aksai-Chin-Plateau, [in Xizang Prov., China], (in Botanischer Garten Zürich, holotype, lost). These specimens were collected by Walter Bosshard in 1927 in Ritu County, Xizang Province, and then reported by H. Magnusson as a new species G.tibetanica, characterized by its squamulose thallus, C+ red cortex, abundant black pycnidia and cylindroid conidia, 3.5–4.5 × 1 μm, but the cited type specimen was sterile (Zahlbruckner 1933). Its characteristic of abundant pycnidia is not seen in G.qinghaiensis. Obermayer (2004) suggested that G.tibetanica might belong to Acarosporanodulosavar.reagens. Alternatively, because the cortex of G.tibetanica differs in having a C+ red reaction (versus A.nodulosa has C–, K+ red), it might belong to a different species. Therefore, we thoroughly sampled specimens of the genus Glypholecia at the type locality of G.tibetanica. These new specimens have umbilicate, upper cortex paraplectenchymatous, C+ red, KC+ red, K–, P– in the medulla and contain gyrophoric acid. In contrast, A.nodulosa has few rhizines, upper cortex scleroplectenchymatous, C–, K+ yellow turning red, P+ orange-yellow in the medulla, and contains norstictic acid. We found some Glypholecia specimens with numerous pycnidia, as shown in Fig. 2G–H, but rarely with apothecia. Their lower surface usually had fasciculate rhizines aggregated into an umbilicate. Although morphological characters, including the shape (bacilliform), size (3.5–4.5 × 1 µm) of the conidia, and spot reaction (cortex and medulla C+ red) are consistent with those of Zahlbruckner’s G.tibetanica, the molecular data show that these specimens instead belong to G.scabra. In 2019, we also searched the herbarium of the Zurich Botanical Garden for the holotype of G.tibetanica which had been deposited by Walter Bosshard (Zahlbruckner 1933). The holotype could not be located. Therefore, we propose that G.tibetanica could potentially be treated as a synonym of G.scabra. Further research is required to determine their synonymy. At present, as the holotype could not be examined, the species name G.tibetanica should be maintained. Supplementary Material XML Treatment for Glypholecia qinghaiensis XML Treatment for Glypholecia scabra XML Treatment for Glypholecia tibetanica Acknowledgements The authors are very grateful to Hurnisa Xahidin (Xinjiang University) for providing sequence data for this study. This research was supported by the Second Tibetan Plateau Scientific Expedition and Research Program (STEP) (2019QZKK0503), grants from the Flora Lichenum Sinicorum (31750001), Youth Innovation Promotion Association CAS (2020388), Yunnan Young and Elite Talents Project and National Natural Science Foundation of China (Nos. 31970022, 31670028, 32060001). Additional information Conflict of interest The authors have declared that no competing interests exist. Ethical statement No ethical statement was reported. Funding This research was funding from the Flora Lichenum Sinicorum (31750001), Youth Innovation Promo­tion Association CAS (2020388), Yunnan Young and Elite Talents Project and Na­tional Natural Science Foundation of China (Nos. 31970022, 31670028, 32060001), Second Tibetan Plateau Scientific Expedition and Research Program (STEP) (2019QZKK0503). Author contributions All authors contributed to this work. Author ORCIDs An-cheng Yin https://orcid.org/0009-0000-7837-7797 Qiu-yi Zhong https://orcid.org/0000-0001-6256-6083 Christoph Scheidegger https://orcid.org/0000-0003-3713-5331 Ji-zhen Jin https://orcid.org/0009-0005-4351-8162 Fiona R. Worthy https://orcid.org/0000-0003-0042-3110 Li-song Wang https://orcid.org/0000-0003-3721-5956 Xin-yu Wang https://orcid.org/0000-0003-2166-6111 Data availability All of the data that support the findings of this study are available in the main text or Supplementary Information. ==== Refs References Abbas A Wu JN (1994) The lichens from Tielimaiti Pass Kuqa County, Xinjiang, China. Ganhanqu Yanjiu 11 (4 ): 19–23. Abbas A Wu JN (1998) Lichens of Xinjiang. Urumqi: Sci-Tec and Hygiene Publishing House of Xinjiang, 177 pp. Abbas A Jiang YC Wu JN (1993) Lichens from the Tianshan Mountains, Xinjiang, China. Journal of Nanjing Normal University-Natural Science 16 : 74–82. Acharius E (1814) Synopsis methodica lichenum, 440 pp. Ajaj A El-Assfouri A Ouazzani Touhami A Benkirane R Fennane M Douira A (2007) Inventaire de la collection des lichens et champignons lichénicoles de l’Herbier national “RAB” de l’Institut Scientifique (Rabat, Maroc). 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