Corresponding author: Li-Song Wang (
Academic editor: G. Rambold
In this study, the diversity of
Yang M-X, Wang X-Y, Liu D, Zhang Y-Y, Li L-J, Yin A-C, Scheidegger C, Wang L-S (2019) New species and records of
The lichen genus
Regional studies on this genus have been carried out in Australia (
Although many studies have been conducted, few molecular phylogenetic analyses have been completed (
The specimens examined in this study were collected from the Hengduan Mountains region, Taiwan, Zhejiang, Hainan et al. from 1941 to 2016 and deposited in KUN-L (325 specimens) and in the Institute of Microbiology (HMAS-L, 5). Morphological characteristics were studied using a dissecting microscope (Nikon SMZ745T) and a light microscope (Nikon Eclipse Ci-S; Nikon Instruments, Tokyo Japan). Sections were made with a razor blade under a dissecting microscope and anatomical characteristics were examined and measured using a micrometer under light microscopy. Ten measurements each of the thallus, apothecia and ascospore dimensions were taken from a single apothecium per specimen and the ranges of these measurements, from smallest to largest, are presented in this study. The lichen secondary metabolites were analysed using spot reactions and thin-layer chromatography in a solvent C system, according to
Total genomic DNA was extracted from dried herbarium specimens using AxyPrep Multisource Genomic DNA Miniprep Kit 50-prep (Qiagen) according to the manufacturer’s instructions. ITS (nrDNA ITS1-5.8S-ITS2) and mtSSU (mitochondrial small subunit rDNA) were amplified by polymerase chain reactions (PCR) using the primer pairs ITS1F (
Amplifications were performed in a 25 μl volume comprising 12.5 μl of 2× MasterMix (TapDNA Polymerase, 0.1 units/μl; technologies Co. Ltd), 1.0 μl of each primer, 8.5 μl ddH2O and 2 μl DNA. Conditions for the PCR were: initial denaturation at 94 °C for 4 min, 34 cycles at 94 °C for 1 min, 54 °C for 1 min and 72 °C for 1.5 min, with a final extension at 72 °C for 10 min. PCR products were sequenced in an ABI3730X using amplification primers manufactured by Tsingke (Kunming, China).
ITS and mtSSU sequences were assembled with Seqman 7.0 (DNAStar) and manually edited using Mega6. DNA sequences were aligned with MAFFT version 7 with default parameters (
Maximum likelihood (
Nineteen species were recognised, including three that are new to science (i.e.
Specimen information and GenBank accession numbers for taxa used in this study. Newly obtained sequences are shown in bold.
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China: Yunnan | KUN 12-36993 |
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China: Yunnan | KUN 15-48546 |
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Sweden | Wetmore 91254 (UPS) |
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– |
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China: Yunnan | KUN 15-48082 |
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China: Yunnan | KUN14-43258 |
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China: Xizang | KUN 14-46462 |
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China: Xizang | KUN 14-46439 |
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China: Taiwan | KUN 15-49117 |
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China: Taiwan | KUN 15-49153 |
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China: Yunnan | KUN 12-36055 |
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China: Xizang | KUN 14-46410 |
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China: Yunnan | KUN 14-43995 |
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China: Yunnan | KUN 15-48196 |
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China: Yunnan | KUN 12-34386 |
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China: Yunnan | KUN 12-34377 |
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China: Yunnan | KUN 15-49942 |
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China: Yunnan | KUN 15-47417 |
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China: Yunnan | KUN 13-40715 |
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China: Sichuan | KUN 13-39419 |
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China: Taiwan | KUN 15-49457 |
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China: Yunnan | KUN 13-40695 |
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China: Yunnan | KUN 13-40630 |
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China: Yunnan | KUN 13-41372 |
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China: Yunnan | KUN 13-40596 |
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China: Yunnan | KUN 15-47921 |
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China: Yunnan | KUN 14-43730 |
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China: Taiwan | KUN 15-48998 |
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China: Taiwan | KUN 15-49012 |
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USA | NC 27708 |
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Spain | MAF9852 | – |
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China: Yunnan | KUN 14-43569 |
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China: Yunnan | KUN 13-40767 |
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China: Yunnan | KUN 14-43571 |
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China: Yunnan | KUN 13-40706 |
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India | – |
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Spain | MAF 9854 | – |
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Finland | T. Ahti 69359 |
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The ITS and mtSSU datasets were analysed separately and concatenated; both parsimony and Bayesian trees of ITS vs. mtSSU were congruent. A maximum likelihood phylogenetic tree was inferred from the combined dataset of ITS and mtSSU (Fig.
Species of
Phylogenetic relationship of
Nineteen
CHINA, YUNNAN PROVINCE, Nujiang Perf., Chide Vil., 1916 m elevation,
Thallus 5–9 cm wide, attached to closely adnate. Lobes radiating, plane to convex, but often slightly concave towards the tips, (0.5) 1–3 (4) mm wide, subround at the apices. Upper surface white-grey to celadon, sparsely pruinose at the lobe tips or epruinose, isidia and soredia absent. Medulla pale yellow above, white below. Lower surface black in the centre, paler towards the margin; rhizines dense, furcate. Apothecia common, (0.5) 0.8–1.5 (2) mm wide, constricted at base, plane to possibly convex; margin black. Hymenium height 80–120 μm; hypothecium light brown to brown, internal stipe K– pale yellow to yellow; spores brown, two-celled, 18–20 × 6–8 μm. Upper cortex K+ yellowish, UV–; medulla K–, C–; containing atranorin, chloroatranorin (minor), zeorin and unknown terpenes.
Growing on bark of
The epithet
This species resembles
CHINA: SICHUAN PROVINCE: Muli Co., 2850 m elev., on
CHINA, YUNNAN PROVINCE, Nujiang Pref., Dizhengdang Vil., 1858 m elevation,
Thallus corticolous, 4–9 cm wide, firmly to loosely adnate to substrate. Lobes linear, compact, imbricate to discrete, (0.5) 1–2.5 mm wide, upper cortex plane but often slightly concave towards the tips; pseudocyphellae linear, marginal; upper surface grey to greyish-green, lower-side black; rhizines dense, squarrosely branched. Soralia marginal, labriform; soredia grey to bluish-grey, powdery to granular. Medulla pale yellow. Dactyls and isidia absent. Apothecia absent. Upper cortex K+ yellowish, UV–; medulla K–, C–; containing chloroatranorin (minor) and unknown terpenes.
Growing on bark of
The epithet
CHINA: SICHUAN PROVINCE: Dukou Co., Yanbian Vil., Shibao Mt., 2900 m elev., 29 Jun 1983,
CHINA, YUNNAN PROVINCE, Yongren Co., Lagu Vil., 1050 m elevation,
Thallus saxicolous, up to 7 cm in diam., closely appressed to the substrate. Lobes radiating, irregularly branched, plane to slightly concave, 0.2–1.0 mm wide, subround to truncate at the apices. Upper surface pale grey to yellowish-grey, sparsely pruinose at the lobe tips or epruinose. Lower surface brownish-black, rarely pale brown, rhizines indistinct, sparse to moderately abundant, brownish-black to black. Isidia and soredia absent. Medulla pale yellow in the upper part, white in the lower part. Apothecia abundant, 0.2–0.8 mm wide, constricted at base, plane to possibly convex; margin black. Hymenium height 80–120 μm; hypothecium light brown to brown, internal stipe white; spore brown with two cells, 10–15 × 4–7 μm. Upper cortex K+ yellowish, UV+ yellow; medulla K–, C–; containing lichexanthone, chloroatranorin (minor), zeorin and unknown terpenes (detected by TLC).
Growing on rocks around 1050–1650 m elevation in secondary forests in a dry to semi-arid environment; known only from Yunnan.
The epithet
CHINA: YUNNAN PROVNCE: Yongsheng Co., Dongjiang of Renhe Town, 1130 m elev.,
Upper surface white to whitish-grey or grey-brown; isidia and soredia absent; medulla orange-yellow to orange; lower surface black in the centre, paler towards the margin; apothecia common, (0.3) 0.5–1.0 (1.5) mm wide; internal stipe upper part orange, K+ purple, P–; lower part yellow or much paler than upper part, K–, P–. Upper cortex K–, UV+ yellow, medulla K– or K+ pale red, C–, P– or P+ orange; containing lichexanthone (major), triterpenes, unknown pigment (minor) (detected by TLC).
Growing on bark of
CHINA: SICHUAN PROVINCE: Miyi Co., Malong north slope, 2100 m elev., on
Thallus grey-white, soredia and isidia absent; medulla yellow to orange-yellow; apothecia common, laminal, constricted at base, up to 2 mm in diam.; internal stipe colourless, K–, hypothecium 50–80 μm thick, spores brown, 15–25 × 6–9 μm. Upper cortex K+ yellow, UV–, medulla K–, C–, P–; containing atranorin (major), +/– zeroin, triterpense.
Growing on bark of
CHINA: SICHUAN PROVINCE: Dukou City, Shibao Mt., 2800 m elev., 29 Jun 1983,
Growing on bark of
There is some confusion in the classification of
CHINA: SICHUAN PROVINCE: Dukou Co., Dabaoding, 1950 m elev., 21 Jun 1983,
1 | Thallus UV+, lichexanthone present |
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– | Thallus UV–, lichexanthone absent |
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2 | Thallus with vegetative propagules |
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– | Thallus lacking vegetative propagules |
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3 | Thallus with soredia |
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– | Thallus with isidia |
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4 | Medulla yellow |
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– | Medulla white |
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5 | Medulla yellow; isidia dactyliform |
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– | Medulla white; isidia cylindrical |
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6 | Norstictic acid present |
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– | Norstictic acid absent |
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7 | Atranorin present |
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– | Atranorin absent |
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8 | Medulla pale yellow to yellow |
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– | Medulla white |
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9 | Internal stipe of apothecia absent or indistinct |
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– | Internal stipe of apothecia well developed |
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10 | Norstictic acid present, as well as other triterpenes |
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– | Norstictic acid absent |
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11 | Internal stipe of apothecia brown, K+ red violet |
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– | Internal stipe of apothecia white, K– |
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12 | Thallus with soralia |
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– | Thallus lacking vegetative propagules |
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13 | Medulla white, soralia laminal; norstictic acid present |
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Medulla yellow; soralia marginal; norstictic acid absent |
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14 | Atranorin absent; soralia labriform |
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– | Atranorin present; soralia granular, laminal or orbicular |
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15 | Lobe margin without pseudocyphellae; soredia yellow |
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– | Lobe margin with intermittent pseudocyphellae; soredia grey to bluish-grey |
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16 | Medulla yellow |
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– | Medulla white |
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17 | Internal stipe of apothecia colourless |
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– | Internal stipe of apothecia brown or yellow |
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18 | Internal stipe brown, K+ red violet |
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– | Internal stipe of apothecia pale yellow; upper medulla yellow, lower medulla white |
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The authors are grateful to Dr. Melissa Dawes for the scientific editing, as well as Prof. Bernard Goffinet from the University of Connecticut and Prof. James R. Shevock from the California Academy of Sciences for guidance and help with writing in English. Sincere thanks are extended to Institute of Microbiology, Chinese Academy of Sciences for the loan of specimens. This study was supported by grants from the National Natural Science Foundation of China (No. 31370069, 31400022, 31670028, 31750001), the Second Tibetan Plateau Scientific Expedition and the China Scholarship Council.