683 MycoKeys MycoKeys 121: 341-356 (2025) DOI: 10.3897/mycokeys.121.161062 Research Article Morphology, chemistry, and phylogeny reveal three new species of Lecidea (Lecideaceae) from China Zhaojie Ren'™®, Ruotong Li2®, Chunxiao Wang2®, Junxia Xue2®, Lisong Wang?“®, Xinyu Wang?“®, Lulu Zhang?® 1 Shandong Museum, Jinan, 250014, China 2 College of Geography and Environment, Shandong Normal University, Jinan, 250014, China 3 State Key Laboratory of Phytochemistry and Natural Medicines, Kunming Institute of Botany, CAS, Kunming, 650201, China 4 Yunnan Key Laboratory for Fungal Diversity and Green Development, Kunming Institute of Botany, CAS, Kunming, 650201, China Corresponding authors: Xinyu Wang (wangxinyu@mail.kib.ac.cn); Lulu Zhang (612038@sdnu.edu.cn) OPEN Qaccess This article is part of: Exploring the Hidden Fungal Diversity: Biodiversity, Taxonomy, and Phylogeny of Saprobic Fungi Edited by Samantha C. Karunarathna, Danushka Sandaruwan Tennakoon, Ajay Kumar Gautam Academic editor: Samantha C. Karunarathna Received: 4 June 2025 Accepted: 8 August 2025 Published: 2 September 2025 Citation: Ren Z, Li R, Wang C, Xue J, Wang L, Wang X, Zhang L (2025) Morphology, chemistry, and phylogeny reveal three new species of Lecidea (Lecideaceae) from China. Mycokeys 121: 341-356. https://doi. org/10.3897/mycokeys.121.161062 Copyright: © Zhaojie Ren et al. This is an open access article distributed under terms of the Creative Commons Attribution License (Attribution 4.0 International - CC BY 4.0). Abstract Three new species, Lecidea flavothallia, L. sublaboriosa, and L. tibetica, are reported from China based on morphological, chemical, and molecular characters. Lecidea flavothallia is characterized by an orange thallus and the presence of schizopeltic acid. Lecidea sub- laboriosa is characterized by a sparsely developed thallus, brown hypothecium, and the presence of 4-O-demethylplanaic acid in the apothecia. Lecidea tibetica is characterized by a well-developed thallus, I+ violet medulla, hyaline to pale straw-colored hypothecium, and the presence of 2'-O-methyperlatolic acid as the major secondary metabolite. All of the new species were collected from the Qinghai-Tibetan Plateau in Southwest China. Detailed descriptions, discussions, and figures are provided for each species, along with a key for all known Chinese Lecidea s. str. species. Key words: East Asia, Lecidea s. str., lichen, Qinghai-Tibetan Plateau, taxonomy Introduction Lecidea Ach. (Lecideaceae) was originally described by Acharius (1803). In the early stage, the classification of Lecidea was relatively broad, being characterized by crustose thalli, green algal photobionts, lecideine or biatorine exciples, and colorless ascospores (Spribille and Printzen 2007). With the advancement of research and the development of microscopy, the characteristics of ascospores, apothecia, and ascus apex structures were introduced into the classification, narrowing the scope of Lecidea (Massalongo 1852; Hafellner 1984). Hertel (1967, 1970, 1977a, 1977b, 1987, 1995, 2006) conducted a systematic study on Lecidea, further narrowing the taxonomic category of the genus. The species strictly in line with the definition of Hertel— as lichenologists call Lecidea s. str.—include only approximately 100 species. The main characteristics are thallus crustose to sub-squamulose; ascomata apothecia, lecideoid; asci 8-spored, Lecidea-type; ascospores simple, hyaline, without a perispore; and almost exclusively on rock (Hertel and Printzen 2004). 341 Zhaojie Ren et al.: Three new species of the lichen genus Lecidea from China The Qinghai-Tibetan Plateau is approximately 2.5 million square kilome- ters (26°00'-39°47'N, 73°19'-104°47'E), including all of China’s Tibetan area and parts of Qinghai, Xinjiang, Gansu, Sichuan, and Yunnan provinces. It is the largest plateau in China and the highest in the world, known as the “Roof of the World.” Due to its special climatic, orographic, and geological conditions, the Qinghai-Tibetan Plateau is particularly rich in lichen species, but most of them are insufficiently understood. During our study of the lichen flora of the Qinghai-—Tibetan Plateau, three species of Lecidea were discovered, all of which are new to science. We pres- ent a brief diagnosis, an extended description, and a phylogenetic analysis based on ITS sequence data. Materials and methods Morphology and chemistry The specimens were collected on the Qinghai-Tibetan Plateau, China, and are preserved in the Lichen Section of the Kunming Institute of Botany, Chinese Academy of Sciences (KUN). The specimens were examined morphological- ly using a COIC XTL7045B2 dissecting microscope and anatomically with an Olympus CX41 polarizing microscope and photographed under an Olympus SZX16 and BX61 microscope with a DP72 digital camera. The thallus, medulla, exciple, and epihymenium were tested with K (a 10% aqueous solution of po- tassium hydroxide), C (a saturated solution of aqueous sodium hypochlorite), or N (a 50% aqueous solution of nitric acid) for identification. Lugol's iodine solution was employed to determine the amyloid reaction of the medulla and the type of asci. Crystals within the section were examined under polarized light. Lichen substances were identified using standardized thin-layer chroma- tography techniques (TLC) with systems B' (hexane:methy| tert-butyl ether:for- mic acid = 140:72:18) and C (toluene:acetic acid = 200:30) (Orange et al. 2001; Elix 2014). In this study, Lethariella cladonioides (Nyl.) Krog, containing atra- norin and norstictic acid, was used as the partition standard sample. Phylogenetic analyses Total genomic DNA was extracted with a DNAsecure Plant Kit according to the manufacturer's instructions and purified with a PCR quick-spin™ PCR Product Pu- rification Kit. The ITS1-—5.8S-ITS2 regions were amplified ina C1000™ automatic thermocycler using the primers ITS1F (Gardes and Bruns 1993) and ITS4 (White et al. 1990) in a 25 uL volume containing 12.5 pL of 2 x Taq PCR Master Mix (Aidlab) (containing Taq DNA polymerase: 0.1 unit/uL; MgCl,: 4 mM; and dNTPs: 0.4 mM), 8.5 uL of ddH,0, 2 uL of primer (1 pL of a 10 mM solution for each prim- er), and 2 uL of genomic DNA. PCR thermal cycling parameters were as follows: initial denaturation at 94 °C for 10 min, followed by 34 three-step cycles (95 °C for 45 s, 50 °C for 45 s, 72 °C for 1 min 30 s) and a final 10 min extension at 72 °C. PCR products were Sanger sequenced by Sangon Biotechnology Company. All raw sequences were compared to those available in the GenBank da- tabase (http://www.ncbi.nlm.nih.gov/BLAST/) to ensure their reliability. The raw sequences were assembled and edited using SeqMan v. 7.0 (DNASTAR Mycokeys 121: 341-356 (2025), DOI: 10.3897/mycokeys.121.161062 342 Zhaojie Ren et al.: Three new species of the lichen genus Lecidea from China packages). Sequences extracted from the new materials were aligned with additional sequence data from GenBank using an online version of MAFFT v. 7.0.26 (Katoh et al. 2005). The MAFFT algorithm chose Auto (FFT-NS-1, FFT- NS-2, FFT-NS-i, or L-INS-i, depending on the data size). Farnoldia jurana (Schaer.) Hertel was chosen as the outgroup based on previous phylogenetic analyses. Phylogenetic relationships were inferred using maximum likelihood (ML) and Bayesian inference (BI). ML was performed using the CIPRES Science Gateway (http://www.phylo.org/portal2/) (Miller et al. 2010). ML analysis was conducted using RAXML-HPC v. 8.2.12 (Stamatakis 2014), with default parameters as im- plemented on CIPRES, and support values were based on 1,000 non-parametric bootstrap pseudoreplicates. Bayesian analysis was conducted using MrBayes v. 3.2.7 (Ronquist et al. 2012). The best substitution models were estimated us- ing ModelFinder (Kalyaanamoorthy et al. 2017). Based on the results, we used the TIM2 + F + G4 model for nrlTS. Bayesian inference was performed with two parallel Metropolis-coupled Markov chain Monte Carlo runs (one “cold” chain and three heated chains), with trees sampled every 1,000 generations, and the run was automatically stopped when the average standard deviation of split frequencies fell below 0.01. The resulting tree was summarized after discarding the first 25% of samples. Generated phylogenetic trees were visualized using FigTree v. 1.4.2 (Rambaut 2012) and then edited in Adobe Illustrator (Al). Boot- strap support (BS) = 70% and posterior probabilities (PP) = 0.95 were consid- ered significant supporting values and are presented on the tree. Results and discussion The sequence alignment comprised 69 nrlTS sequences (46 downloaded from GenBank and 23 newly generated) of 33 taxa. The final dataset comprised a to- tal of 469 sites. The phylogenetic trees obtained from ML and BI analyses exhib- ited a generally congruent topology; we therefore present only the BI tree, with BS = 70% for the ML analysis and PP = 0.95 for the Bayesian analysis (Fig. 1). Although the results showed that the backbone of our tree is poorly supported, the three new species—Lecidea flavothallia C.X. Wang and Lu L. Zhang, L. sub- laboriosa Z.J. Ren and Lu L. Zhang, and L. tibetica Z.J. Ren, Xin Y. Wang and Lu L. Zhang—formed monotypic lineages with strong support (100% ML, 1.00 PP) and were also supported by morphological and chemical characteristics. Lecidea flavothallia formed a well-supported clade (96/1.00) close to L. diducens Nyl.; however, L. diducens is characterized by a sparsely developed thallus and the presence of 2'-O-methylanziaic acid in the apothecial exciple (Hertel and Print- zen 2004; Fryday et al. 2024). Lecidea sublaboriosa, L. andersonii R. Filson, and L. polypycnidophora U. Rupr. and Turk formed a distinct clade. Species of this clade produce either planaic/4-O-demethylplanaic acids or are acid deficient. Thallus development ranged from scant to largely endolithic; however, each species con- stitutes a monophyletic and strongly supported subclade. Additionally, these spe- cies can be distinguished based on morphological characteristics, which will be further discussed in the taxonomic descriptions. Lecidea tibetica appeared close to L. glacierensis A. Abbas and Mamut, but the two species can be separated by their external morphology and chemistry: L. glacierensis occurs on calcareous rock, has a non-amyloid medulla, smaller apothecia (0.4—1.5 mm), and contains confluentic acid as the major secondary metabolite (Mamut et al. 2022). MycoKeys 121: 341-356 (2025), DOI: 10.3897/mycokeys.121.161062 343 Zhaojie Ren et al.: Three new species of the lichen genus Lecidea from China Lecidea tessellata MW443060 19-64846 (KUN) China Lecidea tessellata KX120213 UR00138 Argentina ; Lecidea tessellata EU263926 GZU Hafellner 49249 Austria Lecidea stratura ON794221 Knudsen 14270 USA 100/1- Lecidea tessellata MW443065 19-65382 ase) China Lecidea tessellata HQ650671 Lay 01-036 99/1 97/1 Ss unknown 96/1 98/0.95, Lecidea flavothallia MW 443057 19-63591 (KUN) 1001 Lecidea flavothallia MW443058* 19-63574 (K' 26/1 Lecidea flavothallia MW 443066 19-63636 (KU. 100/1, Lecidea diducens MT 181002 20181602 (S n Lecidea diducens MT181003 20181709 (SDNU) China 99/1- Lecidea auriculata GU074427 Lund L05184 Sweden 9 Lecidea auriculata MT180996 20141032(SDNU) China Lecidea auriculata MT180994 20141078(SDNU) China Lecidea auriculata KX120212 UR00010 Argentina 100/1- Lecidea lithophila MK 620090 F-LI-1¢ Antarctica Lecidea lithophila MK620247 UR00246 Austria * ) China XxX et 6775 USA Lecidea fuscoatrina MK591838 201710700 (XJU) China Lecidea fuscoatra EU263922 Tuerk 40136 Austria | root Lecidea cancriformis GU074440 LE AQ57501 Antarctica Lecidea cancriformis GU074435 Tuerk 42988 Antarctica Lecidea cancriformis GU074439 MAF-Sancho 2 Antarctica 100/1- Lecidea lapicida KX120201 UR00017 Chile . 99 Lecidea lapicida KX 120200 UR00002 Argentina Lecidea lactea MN483122 STE s.n. (2010) (GZU) USA 98/0.98- Lecidea medusula MK620251 UR00289 Austria 98/1. Lecidea medusula MK620270 US12432a Norway Lecidea medusula MK620130 UR00070 Argentina Lecidea protabacina KY 800506 Muggia KTM-LM144 USA Lecidea confluens EU263921 Tuerk 39641 Austria 100/1- Lecidea violascens MZ922140 Leavitt 19100 USA — Lecidea violascens KY 800509 Rosentreter 18195 (OSC) USA 95/1, Lecidea atrobrunnea MW443067 19-66092 (KUN) China Lecidea atrobrunnea MW 443068 19-63312 (KUN) China Lecidea perlatolica MZ243725 Leavitt 18-5l1la BRY-C USA. 76/0.98|L_ Lecidea atrobrunnea EU259897 Lund, Lich. Groenl. Exsiccati 770 N. greenland loot _, Lecidea syncarpa GU074456 UCR-174803 USA Lecidea syncarpa EU259898 UCR-176128 USA 100/1- Lecidea glacierensis MK590965* 20091647 (XJU) Ch Lecidea tibetica P 3. Lecidea promiscens KX 05 UR00129 Argentina ; Lecidea BOTY rasop hora MW 443063 19-64775.1 (KUN) China Lecidea polypycnidophora MW443064 19-64775.2 (KUN) China Lecidea polypycnidophora MH231423 20167599a (XJU) China Lecidea polypycnidophora MH232960 20167587 (XJU) China Lecidea andersonii GU074454 Tuerk 43022 Antarctica 99/1 79/0.96 _— Lecidea aff. laboriosa MK990115 Leavitt 17524 Cua) USA Lecidea aff. laboriosa MK 990118 Leavitt 18602 (BRY-C) USA 100/1, Lecidea laboriosa MT 181010 20151768 (SDNU) China Lecidea laboriosa MW 443062 19-65582 (KUN) China Lecidea leucothallina MG968261 BG-L-95936 Norway Lecidea leucothallina MG968257 O-L-165806 Norway Lecidea toensbergii MG973073 O:L 181870 Norwa ; Lecidea obluridata GU074423 Tuerk 42730 Austria Lecidea silacea GU074424 Tuerk 42156 Austria Lecidea silacea HQ650629 AFTOL-ID 1368 unknown Farnoldia jurana EU263920 Herbarium Tuerk 39660 Austria 0.04 Figure 1. Phylogenetic tree constructed via Bayesian inference (BI) analysis of Lecidea species based on the concate- nated nrlTS dataset. Bootstrap support values = 70 for ML and posterior probabilities = 0.95 (second value) for Bayesian methods are indicated above the branches. Newly obtained sequences in this study are in bold; * represents type mate- rial. The new species are highlighted with colored backgrounds. Taxonomy Lecidea flavothallia C.X. Wang & Lu L. Zhang, sp. nov. MycoBank No: 838415 Fig. 2 Diagnosis. Similar to Lecidea truckeei but with orange thallus and shorter conidia ([6-]7-9[-10.5] x 1 um vs. [10-]11.5-15[-18] x 1.2 um). Type. CHINA, * Tibet Prov., Lazi Co., Xigin vil., 29°04'24.14"N, 87°58'35.83'E, alt. 4492 m, on soil, 18 Jul. 2019, L.S. Wang et al. 19-63574 (KUN, holotype; GenBank mw443058). MycoKkeys 121: 341-356 (2025), DOI: 10.3897/mycokeys.121.161062 344 Zhaojie Ren et al.: Three new species of the lichen genus Lecidea from China Etymology. The epithet “flavothallia” refers to the orange thallus. Description. Thallus: areolate, flat to bullate or lobate; prothallus: indistin- guishable; areoles: dispersed, flat to convex, up to 2 mm in diam.; regular to irregular, angular to round in outline, with a whitish margin, esorediate; upper surface: shiny, (usually) yellowish orange to orange, occasionally with white or gray patches; cortex: 110-160(—220) um thick; medulla: white, I-; algal layer: (60-)70-100(-150) um thick; photobiont trebouxioid, cells (6-)7—10.5(-13) um diam. Apothecia: abundant, subimmersed to sessile, lecideine, (0.6—)1.0- 1.5(-2.0) mm diam.,; disc: black, flat to convex, epruinose to faintly white pru- inose; proper margin: black, prominent when young, occasionally becoming ex- cluded in old apothecia. In section: exciple: black-brown outside, unpigmented inside, with small crystals (POL+) dissolving in N and insoluble in K; epihymeni- um: black-green to olive-green (N+ red-brown with a faint purple, K-), 10-15 um thick; hymenium: hyaline or slight green, 40-60 um tall; paraphyses: simple, occasionally scarcely branched and anastomosing; subhymenium: hyaline, 15-20 um thick; hypothecium: brown, red-brown to dark brown; asci: clavate, Lecidea-type, 8-spored; ascospores: hyaline, simple, ellipsoid, (7—)8-9(-12) x 3.5-5 um, length-width index: 1.8-2.2(-—2.5) (n = 20). Conidiomata: immersed, graphidoid; conidia: bacilliform, (6-)7-—9(-10.5) x 1 um. Chemistry. Cortex and medulla K+ yellow, C-, KC-; schizopeltic acid detect- ed by TLC. Additional specimens examined. CHINA * Tibet Prov., Lazi Co., Xiqin vil., 29°04'18.92"N, 87°58'37.49"E, alt. 4536 m, on soil, 18 Jul. 2019, L.S. Wang et al. 19-63591 (KUN); 19-65264 (KUN). * Angren Co., Kerangla vil., 29°19'01.67'N, 87°01'58.50"E, alt. 4530 m, on siliceous rock, 19 Jul. 2019, L.S. Wang et al. 19- 63636 (KUN). * Dingri Co., Zhaguo vil., 28°35'09.72'N, 87°03'44.35’E, alt. 4316 m, on siliceous rock, 16 Jun. 2022, L.S. Wang et al. 22-71230 (KUN). * Jilong Co., Zheba vil., 29°17'25.50"N, 85°14'52.99"E, alt. 4555 m, on siliceous rock, 17 Jun. 2022, L.S. Wang et al. 22-71328 (KUN); * 29°17'24.62'"N, 85°14'53.17’E, alt. 4571 m, on siliceous rock, 17 Jun. 2022, L.S. Wang et al. 22-71332 (KUN). * Yunnan Prov., Deqin Co., Baima Snow Mountain, 28°20'513'N, 99° 03'984’E, alt. 4320 m, on soil, 28 Aug. 2006, L.S. Wang, Oh Soon-Ok & Niu Dong-ling. 06-26744 (KUN). Distribution. Currently, this species is only found on the Qinghai-Tibetan Pla- teau of China, growing on exposed rocks and occasionally on alpine meadow permafrost soil, between altitudes of 4,000 and 5,000 m. Discussion. The species has the general appearance of a member of the Lecidea atrobrunnea group, but the |- medulla and short conidia indicate oth- erwise. Molecular data indicate that it belongs to the L. auriculata group, with which it shares the dark hypothecium, low hymenium, narrow ascospores, and graphidioid conidiomata, but that it differs from other members of this group by the atrobrunnea-type thallus containing schizopeltic acid. Other Lecidea spe- cies with schizopeltic acid include L. cinerata Zahlbr., L. hassei Zahlbr., L. man- nii Tuck., and L. truckeei Herre, but L. cinerata has larger ascospores (13-16 x 4.2-6.3 um vs. [7-]8-9[-12] x 3.5-5 um) and a C+ red exciple (lecanoric acid vs. schizopeltic acid); L. hassei has an endolithic thallus and a brownish opaque exciple; L. mannii has a black prothallus, larger ascospores (10-15 x 5-6.7 um vs. [7-]8-9[-12] x 3.5-5 um), and gyrophoric acid (C+ red); and L. truckeei has a dark reddish brown thallus and an I+ intensely violet medulla (Herre 1911; Hertel and Printzen 2004; Lendemer and Knudsen 2007; Fryday MycoKeys 121: 341-356 (2025), DOI: 10.3897/mycokeys.121.161062 345 Zhaojie Ren et al.: Three new species of the lichen genus Lecidea from China Wy ft ve f - ¥ ne 10 um aoe a yy be Figure 2. Lecidea flavothallia. A. Habitat and thallus (KUN 19-63574, holotype). (B-G KUN 19-63636): B. Apothecia; C. Apothecium section; D. Ascospores; E. Paraphyses; F. Ascus; G. Amyloid reaction of ascus. Scale bars: 500 um (B); 50 um (C); 10 um (D, E, F, G). and Hertel 2014). Additionally, Lecidea flavothallia and L. poeltii Hertel share similar characteristics, both possessing a shiny, well-developed thallus, a green epihymenium, a low hymenium, and a dark hypothecium, with distributions in the Himalayan region. However, L. poeltii differs in having a brown thallus, 1+ medulla, and larger mature spores (11-16 x 4.5-7 um vs. [7-]8-9[-12] x 3.5-—5 um) (Hertel 1975, 1977a). MycoKeys 121: 341-356 (2025), DOI: 10.3897/mycokeys.121.161062 346 Zhaojie Ren et al.: Three new species of the lichen genus Lecidea from China Lecidea sublaboriosa Z.J. Ren & Lu L. Zhang, sp. nov. MycoBank No: 859348 Fig. 3 Diagnosis. Similar to Lecidea laboriosa but with a brown to dark brown hypoth- ecium and shorter ascospores ([6—]7-8.5[-10] x 2.5-3.5[-4] um vs. [6-]8- 12[-16] x [2-]2.5-4[-5] um). Type. CHINA * Tibet Prov., Angren Co., Sangsang vil., 29°18'59.01'N, 87°02'02.97'E, alt. 4498 m, on rock, 19 Jul. 2019, L.S. Wang et al. 19-65307 (KUN, holotype; GenBank PV698372). Etymology. The specific epithet refers to the new species’ similarity to Lecidea laboriosa. Description. Thallus: endolithic to scarcely epilithic; prothallus: absent; sur- face: if present, whitish to cream-colored, rough, esorediate; medulla: white, I- or partially weakly I+ violet; algal layer: not obvious; photobiont trebouxioid, cells (9-)10-13(-15) um diam. Apothecia: abundant, sessile, lecideine, 0.65-1.5mm diam.; disc: black, flat to convex, epruinose to faintly white pruinose; proper mar- gin: black, regular or corrugated, occasionally becoming excluded in old apoth- ecia. In section: exciple: black-brown outside, hyaline to light brown inside, with small crystals (POL+) dissolving in N and insoluble in K; epihymenium: black- green (N+ violaceous, K—), 7.5—12.5 um thick; hymenium: hyaline or slight blue- green, 30-40 um tall; paraphyses: simple, unbranched, slightly enlarged at the apex, apical cells 2.5-3(—4) um; subhymenium: hyaline to light brown, 12-20 um thick; hypothecium: brown to dark brown; asci: clavate, Lecidea-type, 8-spored; ascospores: hyaline, simple, ellipsoid, (6-—)7—8.5(—10) x 2.5-3.5(-4) pm, length- width index: 1.8-2.5(—3.2) (n = 20). Conidiomata: not seen. Chemistry. Cortex and medulla K-, C-, KC-; 4-O-demethylplanaic acid de- tected by TLC. Additional specimens examined. CHINA «+ Tibet Prov., Zhongba Co., Larang Vil., 29°47'20.00"N, 83°54'12.10"E, alt. 4652 m, on rock, 25 Jul. 2019, L.S. Wang et al. 19-64947 (KUN); 19-65613 (KUN). * Dazi Dist., Bangdui Vil., 29°45'44.00"N, 91°25'29.28"E, alt. 3698 m, on siliceous rock, 16 Jul. 2019, L.S. Wang et al. 19-64619 (KUN). Distribution. This species is found in the plateau sub-frigid, semi-arid climate zone of China, growing on exposed siliceous rocks. Discussion. We have conducted multiple phylogenetic tree reconstructions, in which Lecidea sublaboriosa is placed as a sister species to L. laboriosa Mull. Arg. is either weakly supported or unsupported. Despite this phylogenetic un- certainty, L. sublaboriosa exhibits a high degree of morphological and chemical similarity with L. laboriosa, including an endolithic to scarcely epilithic thallus, a green epihymenium, a low hymenium, and the presence of 4-O-demethylplana- ic acid. Additionally, it is morphologically similar to L. andersonii, L. auriculata Th. Fr., L. diducens, and some specimens of L. polypycnidophora, in that they all either lack an epilithic thallus or, if one is present, it is poorly developed. All of the above species have been detected in the Qinghai-Tibetan Plateau; however, they differ in that Lecidea andersonii has larger ascospores ([8-]9-12 x 3.5-4.4 um vs. [6-]7-8.5[-10] x 2.5-3.5[-4] um) and an I+ violet medulla; L. auriculata has an I+ violet medulla and contains confluentic acid; L. didu- cens has a C+ red exciple and contains 2’-O-methylanziaic acid; L. laboriosa MycoKeys 121: 341-356 (2025), DOI: 10.3897/mycokeys.121.161062 347 Zhaojie Ren et al.: Three new species of the lichen genus Lecidea from China >. Figure 3. Lecidea sublaboriosa (KUN 19-65307, holotype). A. Thallus; B. Apothecia; C. Apothecium section; D. Para- physes; E. Ascus; F. Amyloid reaction of ascus; G. Ascospores. Scale bars: 1 mm (A); 200 um (B); 50 um (C); 10 pm (D, E, F, G). has larger ascospores ([6—]8—12[-16] x [2-]2.5-4[-5] um vs. [6-]7-8.5[-10] x 2.5-3.5[-4] um) and hyaline to light brown hypothecium; L. polypycnidophora has broader ascospores (6-8 x 3-4.5 um vs. [6-]7-8.5[-10] x 2.5-3.5[—4] pm) and abundant pycnidia (Inoue 1991; Hertel and Printzen 2004; Ruprecht et al. 2010; Fryday et al. 2024). MycoKeys 121: 341-356 (2025), DOI: 10.3897/mycokeys.121.161062 348 Zhaojie Ren et al.: Three new species of the lichen genus Lecidea from China Lecidea tibetica Z.J. Ren, Xin Y. Wang & Lu L. Zhang, sp. nov. MycoBank No: 859349 Fig. 4 Diagnosis. Lecidea tibetica is characterized by a well-developed thallus, I+ vi- olet medulla, hyaline to pale straw-colored hypothecium, and the presence of 2-O-methyperlatolic acid as the major secondary metabolite. Type. CHINA * Tibet Prov., Chayu Co., Zhuwagen Town Mont. Pass, 28°43'17.75'N, 97°42'21.01"E, alt. 4685 m, on siliceous rock, 13 Jul. 2019, X.Y. Wang et al. XY 19-659 (KUN, holotype; GenBank PV698370). Etymology. The epithet “tibetica” refers to the type location, Tibet Province, China. Description. Thallus: crustose, areolate to rimose-areolate, flat to bullate, thick (up to 1.1 mm); prothallus: black, obvious at the margin of the thalli; are- oles: dispersed, irregular, flat to convex, sometimes expand into a tuberous structure; surface: gray to bluish gray or whitish gray (the shady side), esore- diate; cortex: 12-20 um thick; medulla: white, I+ violet; algal layer: 50-100 um thick; photobiont trebouxioid, cells (5.5—-)7.5-13 um diam. Apothecia: black, immersed to subimmersed, lecideine, up to 4.5 mm in diam.; disc: black, flat to slightly convex, epruinose, often irregularly cracked in old apothecia; proper margin: dull, often dark grey, persistent, sometimes undulate. In section: exci- ple: dark brown outside, unpigmented inside, with large crystals insoluble in K and N; epihymenium: olive-brown (N+ orange-brown, K-), 15-25 um thick; hymenium: hyaline, 70-85 um tall; paraphyses: simple, occasionally scarce- ly branched and anastomosing; subhymenium: hyaline, poorly distinguishable from the hymenium; hypothecium: hyaline to pale straw colored, c. 200 um; asci: clavate, Lecidea-type or Porpidia-type (occasionally observed in imma- ture asci), 8-spored; ascospores: hyaline, simple, ellipsoid, 11.5-13 x 6-8 pm, length-width index: 1.6—-2 (n = 20). Conidiomata: immersed, graphidoid; conid- ia: bacilliform, (14—)15-18(-19) x 1 um. Chemistry. Cortex and medulla K+ yellow, C-, KC-; thallus UV-; 2’-O-methy- perlatolic acid and confluentic acid (trace) were detected by TLC. Additional specimens examined. CHINA * Tibet Prov., Chayu Co., Yixiula Puerto, 28°44'16.62'"N, 97°42'7.98'E, alt. 4492 m, on siliceous rock, 25 Sep. 2014, L.S. Wang et al. 14-46952 (KUN). Distribution. This species is associated with Rhizocarpon sp. and has been documented so far only in the Tibet Province of China, where it grows on non-calcareous rocks in high-altitude areas. Discussion. Lecidea tibetica is morphologically similar to L. confluens (We- ber) Ach., L. tessellata Florke, and Porpidia speirea (Ach.) Kremp., as they all have a well-developed and grayish thallus, I+ violet medulla, and immersed to sub-immersed apothecia. However, among the latter three species, all ex- hibit smaller apothecia (generally less than 2 mm), conidia measuring 8-13 x 1-1.3 um (vs. [14-]15-18[-19] x 1 um), and contain confluentic acid as the primary secondary metabolite. Specifically, Lecidea confluens has a brown to dark brown hypothecium, while L. tessellata lacks conspicuous apothecial mar- gins. Porpidia speirea can be distinguished by the presence of prominent dark parathecial margins, halonate ascospores, and a dark brown hypothecium. MycoKeys 121: 341-356 (2025), DOI: 10.3897/mycokeys.121.161062 349 Zhaojie Ren et al.: Three new species of the lichen genus Lecidea from China A 4 | _ =a 0.5 mm diam., the largest ones up to 4 mm; medulla I+ VIGIS Tet Load un coat sent An Aaland nines ome fea saan Lecidea promiscens - Most apothecia < 0.5 mm diam., the largest ones up to 1 mm; medulla I- hale rorrh i. x ea ebb Ae, IR aah he BARN ere VR: lx eae Ripe RE oe Lecidea promixta 5 — Stictic acid is a major lichen substance (norstictic acid lacking or minor STICTIGACID-CHEMOSYNALOME) ai4- icercecescbareereccosrecereces4urennons Lecidea lapicida = »Stictic acid-or norstictic acid not-deteCtables......c.iccscsececsaesecteedeereasserrceasts 6 6 2-O-methylanziaic present in apothecia; excipulum C+ red...............ccee hace acrid ME et aMR ed enltet We Ras Arana ais Mica Rese Be EU Racene caine, AAR AO Lecidea diducens - 2-O-methylanziaic not detectable in apothecia; excipulum C-................. 7 7 Confluentic acid is the main lichen Substance................c.ccccecsseceetseeeneeens 8 - 4-0-demethylplanaic is the main lichen substance ................cccceeesseeeees 9 8 Ascospores small (6-11 x 2.5-3.5 um); excipulum rather broad, often be- COMMMEG MOXOS 5 od Feet Peaches are ae ceecrenanBecete be trccceh aaa Lecidea auriculata - Ascospores larger (6.5-12 x 3.3—4.5 um); excipulum narrower, usually not HMA KSGYHOXUOUSE, oie) cners sue erneleleannelb dM recensreennnce sie Lecidea promiscens 9 Méedulla I+ violet, excipulum rather broad in young apothecia.................... balkeed sd sdtthestpstadd sid ddswauhttesisas hisned bya virtenetsnshaed hasetirnsses.s Mist usanseehee Lecidea andersonii - Medulla I- or I+ pale violet, excipulum normal-sized in young apothecia.... 10 MycoKeys 121: 341-356 (2025), DOI: 10.3897/mycokeys.121.161062 35] Zhaojie Ren et al.: Three new species of the lichen genus Lecidea from China 10 29 Ascospores small (7-8.5 x 2.5-3.5 pm), hypothecium brown to dark B16 cd puck Ral ed oe Leet RE a Wee ae denen NL met ei Lecidea sublaboriosa Ascospores larger (8-12 x 2.5-4 um), hypothecium hyaline to light O/Ce 10) Rot vO.) x el EUR A. It. As ani RGAE E Ls PURULENT IN. Rh ciiBind Lecidea laboriosa Schizopeltic acid detected, thallus yellowish orange to orange .................. pl Notaries a en Te Lecidea flavothallia SChIZOPEHIG a CIOINIOt CELECKEG Fina. .ivact. sete eostensavesiec deta exenbedeatezehocedencnavest 12 Stictic acid or hypostictic acid detected.................... Lecidea protabacina Stictic acid and hypostictic acid not detectable ........... ee eceeeeeeees 13 NOrstHevioacid GeteCteds 2a has, Screens poe ee 14 NOrSHETIO-ACIEMOLGCTSOLA 6 x cecexsaves-qtes cies crteseceuspoecomneeetnnssetegers secures 15 Thallus thin or moderately thick, areolate plane, white to bluish-gray......... Lea ret ont ty Botan tect Seyi e, Maas tance oa hi tates eM esha che ee Lecidea lactea Thallus crustose, bullate to subsquamulose, pale yellowish brown to more rarely dark brown (atrobunnea-type)............cccccseeeeeeees Lecidea syncarpa Gyropnoric acid detected DY TLCS scidecsssanecssssa dred dansuateyeasansavedimaanwercaras seed des 16 Gyrophoric acid not detectable Dy TLC .....0......cccscccccesesssscresssccesrressrecsseees 19 Ascospores usually longer than 10 UM uuu... cee ccesccceesseeceesseeceesseeeeens 17 Ascospores usually shorter than 10 UM ......... ccc eccsscceesseeceesseeeeesseeeeens 18 Thallus at the margins starting as confluent areoles, surface grey-brown NGC OMTMN sh os Aas arisen bre gh SOBVEN ecada gale toad Manat het fete Lecidea fuscoatra Thallus continuous at the margins, rimose to areolate towards the center, STAC OMEN arsed etre trey Meneses tone at aren: Lecidea grisella Thallus thin (ca 0.1 mm thick), shiny; apothecia margin persistent ............ AI erytiges oe NUS Rory de, WOMAN is ln AE AR pe ee SOEs op Lecidea siderolithica Thallus thick (up to 5 mm thick), dull; old apothecia without conspicuous margin and often irregularly cracked .............cccceceeseeeeees Lecidea sinensis 4-0-demethylplanaic acid detected by TLC ........... ccc cescecsestesestecssteeessnees 20 4-0-demethylplanaic acid not detectable by TLC............ cc eeccceeseeeeneees 21 Apothecia black or pruinose, discs turn brown on wetting, epithecium brown, hypothecium Colorless. ...........c.ccccsscceessceeesseeeees Lecidea lithophila Apothecia black, not pruinose; epithecium dark green, hypothecium brown; PY CIMGIArADUNMOAM Le >, 5.1.'dens casos sasenthaneaumesue Coaches dest Lecidea polypycnidophora Perlatolic acid detected by TLC; thallus pale to medium brown to (more rarely) dark brown in the centel.............c.ccccccecsecseeeeees Lecidea perlatolica Perlatolic acid not detectable by TLC ............ cece ccccccssssesssecsseecsssesssseeesees 22 2'-O-methyperlatolic acid is the main lichen substance; apothecia large, up to 4.5 mm in diam.; hypothecium hyaline to pale straw colored; medulla I+ NOC es Ad. sect, Bn Racse bei c RE Rel ante BOR Nast «cinta Reh hrs Lecidea tibetica Confluentic acid is the main lichen Substance..............cccccceseestecereeeseees 23 Thallus brown, with an epinecral layer (atrobrunnea-type)..............0.08 24 Thallus white to grey, lacking an epinecral layer ............ ccc eceeeeeeeteeeeees 25 Ascospores small (7.5-10 x 2.5—5 um), hypothecium hyaline to brown... chante enhanc its ccs bindete he eRe te A Aetret AEE Reich ark tane eee ae Lecidea atrobrunnea Ascospores larger (10-17 x 4.5-7.5 um), hypothecium dark brown to al- MOST DIACK ...1s-recnressvcepsenarrcnveldivercconmeverssitdaredtyaehiewepannessve Lecidea fuscoatrina Apothecia margin very thin or disappearing, hypothecium hyaline to very pale green or pale yellowish Drown .............::cceseeeeseeeees Lecidea tessellata Apothecia margin usually persistent, hypothecium brownish.................. 26 MycoKeys 121: 341-356 (2025), DOI: 10.3897/mycokeys.121.161062 352 Zhaojie Ren et al.: Three new species of the lichen genus Lecidea from China 26 Ascospores 6.5-7.5 um wide, medulla I+ violet............ Lecidea confluens - Ascospores 7-9 um wide, medulla I- ............. eee Lecidea glacierensis Acknowledgements The authors thank John Elix (Department of Chemistry, Faculty of Sciences, The Australian National University, Canberra, ACT 0200, Australia), Dr. J.C. Lendemer (Institute of Systematic Botany, New York Botanical Garden, Bronx, New York, USA), and Dr. Alan M. Fryday (Michigan State University, USA) for providing suggestions and useful literature during the study. We also thank Dr. Reyim Mamut (College of Life Science and Technology, Xinjiang University, China) for lending us the type materials. Additional information Conflict of interest The authors have declared that no competing interests exist. Ethical statement No ethical statement was reported. Use of Al No use of Al was reported. Funding This research was funded by the National Natural Science Foundation of China (Nos. 32170002, 31750001), Yunnan Fundamental Research Projects (202401AT070196), the Sec- ond Tibetan Plateau Scientific Expedition and Research Program (STEP) (2019QZKK0503), and the Yunnan Young and Elite Talents Project (YNWR-QNBJ-2020-224). Author contributions Methodology: Lulu Zhang, Zhaojie Ren; software: Junxia Xue; formal analysis: Lulu Zhang, Zhaojie Ren, Chunxiao Wang, Junxia Xue; investigation: Lisong Wang, Xinyu Wang; data curation: Chunxiao Wang, Ruotong Li; writing—original draft preparation: Lulu Zhang, Zhaojie Ren, Ruotong Li, Junxia Xue; writing—review and editing: Lulu Zhang, Junxia Xue, Xinyu Wang; project administration: Lulu Zhang; funding acquisition: Lulu Zhang, Xinyu Wang. All authors have read and agreed to the published version of the manuscript. Author ORCIDs Zhaojie Ren ® https://orcid.org/0000-0003-3206-1484 Ruotong Li © https://orcid.org/0009-0003-9428-3055 Chunxiao Wang © https://orcid.org/0009-0009-2094-4854 Junxia Xue © https://orcid.org/0000-0002-6387-0148 Lisong Wang ® https://orcid.org/0000-0003-3721-5956 Xinyu Wang ® hitps://orcid.org/0000-0003-21 66-6111 Lulu Zhang © https://orcid.org/0000-0001-8011-4451 Data availability All of the data that support the findings of this study are available in the main text. MycoKeys 121: 341-356 (2025), DOI: 10.3897/mycokeys.121.161062 oie Zhaojie Ren et al.: Three new species of the lichen genus Lecidea from China References Acharius E (1803) Methodus Qua Omnis Detectos Lichenes. Stockholm, 393 pp. Elix JA (2014) A Catalogue of Standardized Chromatographic Data and Biosynthet- ic Relationships for Lichen Substances. Third Edition. Published by the author, Canberra. Fryday AM, Hertel H (2014) A contribution to the family Lecideaceae s. lat. 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