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Notes on Eight Threatened Species of Lichens in Japan

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Bull. Natl. Mus. Nat. Sci., Ser. B, 37(2), pp. 55–61, May 22, 2011 Notes on Eight Threatened Species of Lichens in Japan Yoshihito Ohmura Department of Botany, National Museum of Nature and Science, Amakubo 4–1–1, Tsukuba, 305–0005 Japan E-mail: [email protected] (Received 17 February 2011; accepted 23 March 2011) Abstract New localities and/or molecular data are provided for the following eight threatened species of lichens in Japan: Chiodecton congestulum, Flavopunctelia soredica, Icmadophila japon- ica, Lethariella togashii, Pseudocyphellaria argyracea, Thysanothecium scutellatum, Usnea fil- ipendula, and U. nipparensis. ITS rDNA sequences were successfully obtained from all species except for C. congestulum, and were compared with the known sequences retrieved from GenBank. Key words : distribution, ITS rDNA, lichenized Ascomycota, Red List. situation of each species as well as molecular Introduction data. The current Red List of Japanese lichens is comprised of 152 taxa: 152 species, 1 sub- Materials and Methods species, and 3 varieties (Ministry of the Environ- ment, 2007; Inoue et al., 2010). Molecular data All collections examined are stored at the of these lichens are of potentially great value for National Museum of Nature and Science, Tokyo taxonomic, phylogenetic, phytogeographic, and (TNS) unless otherwise indicated. Two speci- ecological purposes. mens of Chiodecton congestulum are stored at During recent field surveys in Japan, fresh ma- Uppsala University (UPS). terials of eight threatened species were collected Lichen substances were examined by thin layer in 2009–2010 from both new and already known chromatography (TLC) (Culberson and Johnson, localities, and the sequences of internal tran- 1982). Only the solvent B system was used here. scribed spacer of ribosomal DNA (ITS rDNA) Materials for molecular analysis and the asso- were successfully obtained from all extracted ciated GenBank accession numbers are shown in DNA samples except for Chiodecton congestu- Table 1. DNA was extracted from 5–10 mg of lum. The purpose of this study is to provide in- lichen thalli by using a FastDNA SPIN Kit (MP formation of new localities, and/or the present Biomedicals) following the method of Ohmura et Table 1. Specimen vouchers for DNA analysis and the GenBank accession numbers obtained in this study Species Collection number of Y. Ohmura (in TNS) GenBank accession no. Flavopunctelia soredica 7164 AB623069 Icmadophila japonica 6762 AB623070 Lethariella togashii 6735 AB623071 Pseudocyphellaria argyracea 6418 AB623072 Thysanothecium scutellatum 6773 AB623073 Usnea filipendula 6771 AB623074 U. nipparensis 6282 AB623075 56 Yoshihito Ohmura al. (2006). Specimen examined. Japan. Honshu. Prov. PCR amplification of the ITS rDNA region Mikawa (Pref. Aichi): Mt. Horaiji, Kadoya, (including ITS1, 5.8S rDNA, and ITS2) was per- Horai-cho, Minamishitara-gun, on rock, eleva- formed using ITS1F (Gardes and Bruns, 1993) as tion about 315 m, 5 December 1996, Y. Ohmura the 5Ј primer and LR1 (Vilgalys and Hester, 2840. Prov. Aki (Pref. Hiroshima): Mt. Misen, 1990) as the 3Ј primer. PCR reactions were per- Miyajima Island, Hatsukaichi-city, on rock (gran- formed according to the method of Ohmura et al. ite), elevation 259 m, 12 January 2010, M. Kono (2006) by using PuReTaq Ready-To-go PCR (41 and 42), Y. Ohmura & Y. Koda. Shikoku. beads (GE Healthcare) and a thermal cycler Prov. Tosa (Pref. Kochi): Nagatani Shrine, Naga- (Gene Amp PCR system 9700, Applied Biosys- sawa, Tsuno-mura Village, on coniferous tree, tems). PCR products were purified using an elevation 590–610 m, 30 October 2006, G. Thor ExoSAP-IT (GE Healthcare). Sequencing was 21515 (UPS, det. G. Thor, not seen); ditto, on carried out using a BigDye Terminator ver. 3.1 stump, G. Thor 21516 (UPS, det. G. Thor, not Cycle Sequencing Kit on an ABI Prism 3130x seen). genetic analyzer (Applied Biosystems) according to the manufacturer’s instructions. DNA strands Flavopunctelia soredica (Nyl.) Hale, Mycotax- were assembled and manually corrected using on, 20: 682 (1984). ATGC ver. 6.03 (Genetyx). The obtained se- Flavopunctelia soredica is a well-known fo- quences in this study were compared with the liose species of Parmeliaceae and is easily distin- known sequences of related species registered in guished from similar species on the basis of the GenBank by using BLAST, a homology search presence of soralia along the margin of lobes and program (Altschul et al., 1997). usnic and lecanoric acids as major substances in the cortex and medulla respectively. Although F. soredica is widely distributed in the Northern Results and Discussion Hemisphere, its presence is restricted in central Chiodecton congestulum Nyl., Bull. Soc. Linn. Honshu, Japan, where it has been found on barks Normandie, sér. 2, 2: 106 (1868). of Betula, Castanea, Chamaecyparis, Larix, Chiodecton congestulum was well revised tax- Pinus, Prunus, and Zelkova (Asahina, 1941; onomically by Thor (2002). This species has Kurokawa and Kashiwadani, 1988; Saitama been reported from India, Fiji, Australia, Museum of Natural History, 1988; Kashiwadani, Malaysia, and Japan (Thor, 2007). In Japan, it 1991; Harada et al., 2003; Morita and Harada, has been known from western Honshu, Kyushu, 2006). Additional specimens were collected from Ogasawara Islands, and Yaeyama Islands (Thor, Arakoda (Saku-city) at 730 m alt. and Sugadaira 2002). New localities were found at Mt. Horaiji (Ueda-city) at 1577 m alt. in Nagano Prefecture. in Aichi Prefecture and Nagatani Shrine in Kochi They were found on barks of Larix and Quercus. Prefecture. Although the known localities are restricted to a Fresh materials of this species were also col- narrow region in Japan, F. soredica appears to lected from Miyajima Island in Hiroshima Pre- grow well in these conditions. fecture. Although DNA was extracted from the ITS rDNA sequences were successfully ob- specimens, the ITS rDNA sequence could not be tained from two specimens that were both col- obtained. lected from the same locality of Sugadaira but As reported by Thor (2002), this species grows from different trees: Betula ermanii and Quercus on rocks and tree barks. The specimens collected crispula. Each ITS rDNA sequence is 495 bp in from Aichi and Hiroshima Prefectures were saxi- length, and there are no differences between colous, and those collected from Kochi Prefec- them. The known F. soredica sequence from New ture were corticolous. York, U.S.A. [GenBank accession no. AY773128 Eight Threatened Lichens in Japan 57 (Thell et al., 2005)] is different in only two Bot., 23: 95 (1976). residues from Japanese materials (identityϭ Lethariella togashii was revised taxonomically 493/495 bp, 99%). by Asahina (1952), Krog (1976), and Obermayer Specimens examined. Japan. Honshu. Prov. (1997). The distribution of this species is restrict- Shinano (Pref. Nagano): along the route from ed to Far East Russia and Japan (Skirina, 2006). Sugadaira Pasture to Mt. Azumaya, Ueda-city, on In Japan, it has been collected from Hokkaido bark of Quercus crispula, elevation 1577 m, 30 and central Honshu (e.g. around Lake Yamanaka, May 2010, Y. Ohmura (7119 and 7164) & A. Mt. Fuji), but the habitats in the latter locality Okamura; ditto, on bark of Betula ermanii, Y. were heavily damaged and the populations are Ohmura (7121 and 7167) & A. Okamura; Arako- almost disappeared (Kashiwadani and Inoue, da, Saku-city, on bark of Larix leptolepis, eleva- 2000). tion about 730 m, 17 July 1972, H. Kashiwadani The ITS rDNA sequence was successfully 10332. obtained from one specimen collected at Mt. O-akan, Hokkadio. The sequence of the ITS Icmadophila japonica (Zahlbr.) Rambold & rDNA region is 501 bp in length. According to Hertel, Biblioth. Lichenol., 53: 230 (1993). the BLAST result, the top scores of known relat- ϭ Glossodium japonicum Zahlbr., Bot. Mag. ed species were obtained with Letharia Tokyo, 41: 336 (1927). columbiana (Nutt.) J.W. Thomson [AF115762 Icmadophila japonica is a well-known species (Thell et al., unpublished); identityϭ434/461 bp, of lichens under the name of Glossodium japon- 94%], L. vulpina (L.) Hue [GQ398408 (Alter- icum Zahlbr. by Japanese lichenologists (see mann et al., unpublished); identityϭ428/456, Rambold et al., 1993). This species generally 93%], Lethariella cashmeriana Krog. grows on decayed wood of coniferous trees in an [DQ980014 (Crespo et al., 2007); identi- old-growth forest, and its distribution is restricted tyϭ521/457, 92%], and L. sernanderi (Motyka) to Far East Russia and Japan. In Japan, it grows Obermayer [AF297744 (Kroken, unpublished); widely from Hokkaido to Kyushu (Sato, 1967), identityϭ420/459, 91%]. Although large-scale but the populations in Japan are decreasing phylogenetic analysis based on three ribosomal according to recent field surveys (Inoue et al., DNA regions and RPB1 gene supports a mono- 2010). phyletic clade consisting of Letharia columbiana ITS rDNA sequence was successfully obtained and Lethariella cashmeriana (Crespo et al., from one specimen collected from Mt. O-akan, 2007), a narrower scale analysis has not yet been Hokkaido. The sequence of ITS rDNA region is performed to confirm generic independence be- 466 bp in length. No reference ITS rDNA tween them. The genus Letharia is morphologi- sequence of Icmadophila has been registered cally separated from Lethariella mainly by the in GenBank. lacerate axis. The yellow-colored thallus due to Further molecular phylogenetic analysis based vulpinic acid is distinct in Letharia spp., but this on ITS rDNA sequences and other appropriate substance is also present as a major substance in markers is needed to examine the independence Lethariella togashii. The delimitation and inde- of Glossodium and Icmadophila (s. str.) since pendence between these genera should be revised morphology of the ascomata is quite different in the further researches. between them. Specimen examined. Japan. Hokkaido. Prov. Specimens examined. Japan. Hokkaido. Prov. Kushiro: Mt. O-akan, on bark of Picea glehnii, Kushiro: Mt. O-akan, on decayed stump, eleva- elevation 728 m, 10 August 2009, Y. Ohmura tion 494 m, 10 August 2009, Y. Ohmura 6762. 6735.
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    Mycologia, 98(6), 2006, pp. 1088–1103. # 2006 by The Mycological Society of America, Lawrence, KS 66044-8897 New insights into classification and evolution of the Lecanoromycetes (Pezizomycotina, Ascomycota) from phylogenetic analyses of three ribosomal RNA- and two protein-coding genes Jolanta Miadlikowska1 Soili Stenroos Frank Kauff Botanical Museum, Finnish Museum of Natural Vale´rie Hofstetter History, University of Helsinki, P.O. Box 7, FI-00014 Emily Fraker Finland Department of Biology, Duke University, Durham, Irwin Brodo North Carolina 27708-0338 Canadian Museum of Nature, P.O. Box 3443, Station Martin Grube D, Ottawa, Ontario, K1P 6P4 Canada Josef Hafellner Gary B. Perlmutter Institut fu¨ r Botanik, Karl-Franzens-Universita¨t, North Carolina Botanical Garden, University of North Holteigasse 6, A-8010, Graz, Austria Carolina at Chapel Hill, CB 3375, Totten Center, Chapel Hill, North Carolina 27599-3375 Vale´rie Reeb Brendan P. Hodkinson Damien Ertz Department of Biology, Duke University, Durham, National Botanic Garden of Belgium, Department of North Carolina 27708-0338 Bryophytes-Thallophytes, Domaine de Bouchout, B-1860 Meise, Belgium Martin Kukwa Department of Plant Taxonomy and Nature Paul Diederich Conservation, Gdansk University, A. Legionow 9, Muse´e national d’histoire naturelle, 25 rue Munster, 80-441 Gdansk, Poland L-2160 Luxembourg, Luxembourg Robert Lu¨cking James C. Lendemer Field Museum of Natural History, 1400 South Lake Department of Botany, Academy of Natural Sciences of Shore Drive, Chicago, Illinois 60605-2496 Philadelphia, 1900 Benjamin Franklin Parkway, Philadelphia, Pennsylvania 19103 Geir Hestmark Department of Biology, University of Oslo, P.O. Box Philip May 1066 Blindern, NO-0316 Oslo, Norway Farlow Herbarium, Harvard University, 22 Divinity Avenue, Cambridge, Massachusetts 02138 Monica Garcia Otalora A´ rea de Biodiversidad y Conservacio´n, ESCET, Conrad L.