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Redalyc.Delimitation of Some Neotropical Laccate Ganoderma (Ganodermataceae): Molecular Phylogeny and Morphology

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Redalyc.Delimitation of Some Neotropical Laccate Ganoderma (Ganodermataceae): Molecular Phylogeny and Morphology Revista de Biología Tropical ISSN: 0034-7744 [email protected] Universidad de Costa Rica Costa Rica Correia de Lima Júnior, Nelson; Baptista Gibertoni, Tatiana; Malosso, Elaine Delimitation of some neotropical laccate Ganoderma (Ganodermataceae): molecular phylogeny and morphology Revista de Biología Tropical, vol. 62, núm. 3, septiembre, 2014, pp. 1197-1208 Universidad de Costa Rica San Pedro de Montes de Oca, Costa Rica Available in: http://www.redalyc.org/articulo.oa?id=44932441028 How to cite Complete issue Scientific Information System More information about this article Network of Scientific Journals from Latin America, the Caribbean, Spain and Portugal Journal's homepage in redalyc.org Non-profit academic project, developed under the open access initiative Delimitation of some neotropical laccate Ganoderma (Ganodermataceae): molecular phylogeny and morphology Nelson Correia de Lima Júnior, Tatiana Baptista Gibertoni & Elaine Malosso Departamento de Micologia, Programa de Pós-Graduação em Biologia de Fungos, Universidade Federal de Pernambuco, Av. Nelson Chaves s/n, CEP 50760-420, Recife, PE, Brazil; [email protected], [email protected], [email protected] Received 22-XI-2013. Corrected 25-II-2014. Accepted 27-III-2014. Abstract: Ganoderma includes species of great economic and ecological importance, but taxonomists judge the current nomenclatural situation as chaotic and poorly studied in the neotropics. From this perspective, phylogenetic analyses inferred from ribosomal DNA sequences have aided the clarification of the genus status. In this study, 14 specimens of Ganoderma and two of Tomophagus collected in Brazil were used for DNA extraction, amplification and sequencing of the ITS and LSU regions (rDNA). The phylogenetic delimita- tion of six neotropical taxa (G. chalceum, G. multiplicatum, G. orbiforme, G. parvulum, G. aff. oerstedtii and Tomophagus colossus) was determined based on these Brazilian specimens and found to be distinct from the laccate Ganoderma from Asia, Europe, North America and from some specimens from Argentina. Phylogenetic reconstructions confirmed that the laccate Ganoderma is distinct from Tomophagus, although they belong to the same group. The use of taxonomic synonyms Ganoderma subamboinense for G. multiplicatum, G. boninense for G. orbiforme and G. chalceum for G. cupreum was not confirmed. However, Ganoderma parvulum was confirmed as the correct name for specimens called G. stipitatum. Furthermore, the name G. lucidum should be used only for European species. The use of valid published names is proposed according to the specimen geographical distribution, their morphological characteristics and rDNA analysis. Rev. Biol. Trop. 62 (3): 1197- 1208. Epub 2014 September 01. Key words: Agaricomycetes, phylogenetic taxonomy, rDNA sequences, species delimitation, neotropics. Ganoderma P. Karst. (Ganodermataceae, Gugliotta, Abrahão & Gibertoni (2013) listed Agaricomycetes) is one of the largest genera 18 and 28 species, respectively, only in Brazil. of Polyporales and was described by Karsten Being well known as decomposers and (1881) based on Polyporus lucidus (Curtis) pathogens in tropical forests (Zakaria, Ali, Fr. from Europe, a species with a laccate Salleh, & Zakaria, 2009), species of this genus, (shiny varnished looking) surface. The genus is mainly of the G. lucidum complex, produce characterized by double-walled basidiospores bioactive compounds widely studied for pre- venting and relieving human diseases, such with truncate apex and ornamented endospore as several types of tumors and cancers, gas- (Moncalvo & Ryvarden, 1997). The genus tric ulcers, diabetes mellitus, hypertension and includes 80 species of wide geographic dis- viral infections (Zhou et al., 2007). Besides, tribution with several tropical species and some medicinal effects were reported in manu- others restricted to temperate areas (Ryvarden, scripts of the Chinese civilization more than 2000; Kirk, Cannon, Minter, & Stalpers, 2008). 2 000 years ago (Hong & Jung, 2004; Seo Ryvarden (2004) reported the presence of 20 & Kirk, 2000). species in the neotropics, although Torres-Tor- Although of significant ecological and res, Guzmán-Dávalos & Gugliotta (2012) and biotechnological importance, the taxonomy Rev. Biol. Trop. (Int. J. Trop. Biol. ISSN-0034-7744) Vol. 62 (3): 1197-1208, September 2014 1197 of laccate Ganoderma has been questioned sections of dried material were mounted in 5% in recent years and poorly studied in the KOH to ensure rehydration. Melzer’s reagent neotropics. Currently, taxonomists consider was used to test the amyloid reaction of the the nomenclatural situation as chaotic and cuticle hyphae. Our data were then compared suggest the necessity for a global revision to the available literature (Bresadola, 1911; due to the existence of multiple names for Gottlieb & Wright, 1999; Núñez & Ryvarden, single species (Ryvarden, 1991; Ryvarden, 2000; Ryvarden, 2000; Ryvarden, 2004; Welti 2004; Moncalvo & Ryvarden, 1997; Postnova & Courtecuisse, 2010; Tham et al., 2012, & Skolotneva, 2010). Torres-Torres et al., 2012) and discussed in Usually, different morphological charac- the text. All studied specimens were deposited teristics of the basidiospores (dimensions) and in the herbarium Pe. Camille Torrend (URM), basidiomata (thickness of the cuticle and pres- Department of Mycology, Universidade Fed- ence or absence of resinaceous deposits in eral de Pernambuco, Brazil. the context) are widely used in an attempt to identify laccate species of Ganoderma (Seo Genomic DNA extraction, polymerase & Kirk, 2000; Ryvarden, 2004; Torres-Tor- chain reaction and sequencing: DNA was res & Guzmán-Dávalos, 2012; Torres-Torres, extracted using fragments of basidiomata (30- Guzmán-Dávalos, & Gugliotta, 2012). How- 50mg) ground with a pestle in a porcelain ever, the use of molecular data, especially for mortar containing liquid nitrogen. The result- phylogenetic studies based on ribosomal DNA ing powder was transferred to a tube contain- sequences, combined with morphological stud- ing 700μL of extraction buffer [CTAB 2%, ies has helped many authors to clarify the status 100mM Tris-HCl pH8, 1.4M NaCl, 20mM of the genus (Moncalvo, Wang, & Hseu, 1995a; EDTA, 1% PVP (Rogers & Bendich, 1985)] Moncalvo, Wang, & Hseu, 1995b; Gottlieb, and incubated at 65ºC for 30-40 min. DNA was Ferrer, & Wright, 2000; Smith & Sivasitham- purified with 700μL of chloroform-isoamyl param, 2000; Hong & Jung, 2004; Kaliyape- alcohol (24:1), gently precipitated in 600μL of rumal & Kalaichelvan, 2008; Cao, Wu, & Dai, isopropanol and washed with 1mL of ethanol. 2012; Yang & Feng, 2013). Finally, the pellet was suspended in 50μL of In Brazil, there are only a few studies ultrapure water (Góes-Neto, Loguercio-Leite, regarding Ganoderma (Torrend, 1920; Loguer- & Guerrero, 2005). The reaction mix and cio-Leite, Groposo, & Halmenschlager, 2005; parameters for PCR amplification of the full Torres-Torres et al., 2012) and none of these ITS regions was according to Smith & Sivasi- include a phylogenetic analysis. Thus, the aim thamparam (2000) using the primers ITS1 and of the present study was to determine phyloge- ITS4 (White, Bruns, Lee, & Taylor, 1990). For netic relationships of the laccate Ganoderma LSU rDNA region, the amplification was per- based on sequence variation of the Internal formed with parameters and reagent concentra- Transcribed Spacer (ITS) and Large subunit tions following Góes-Neto, Loguercio-Leite & (LSU) rDNA and to delimit the species’ occur- Guerrero (2005) using the primer pair LR0R rence within Brazilian territory. and reverse LR7 (Moncalvo, Lutzoni, Rehner, Johnson, & Vilgalys, 2000). Negative controls MATERIAL AND METHODS containing all components of the reaction mix, except DNA, were used in each procedure to Morphological analysis: Thirty-one lac- detect possible contamination. The amplifica- cate specimens of Ganoderma and four of tion products of the sixteen laccate specimens Tomophagus colossus (Fr.) Murrill were used (Table 1) were purified using the PureLink in this study (Table 1). To observe the char- PCR Purification Kit (Invitrogen) and the puri- acteristics of the basidiospores, the hyphal fied products were sequenced at the Human system and the cuticle hyphae, free-hand thin Genome Research Center of the Universidade 1198 Rev. Biol. Trop. (Int. J. Trop. Biol. ISSN-0034-7744) Vol. 62 (3): 1197-1208, September 2014 TABLE 1 List of specimens collected from different areas in Brazil Geographic origin Number of deposit GenBank accession Species Locality (city, State) in the Herbarium number (ITS/LSU) Ganoderma chalceum Sítio Carro Quebrado (Triunfo, Pernambuco) URM 80457 JX310812/JX310826 G. chalceum Parque Estadual de Dois Irmãos (Recife, Pernambuco) URM 81393 * G. chalceum Parque Estadual de Dois Irmãos (Recife, Pernambuco) URM 82131 * G. multiplicatum Mineradora Millennium (Mataraca, Paraíba) URM 83346 JX310823/ JX310837 G. multiplicatum Parque Nacional Municipal de Porto Velho (Porto Velho, Rondônia) URM 81081 * G. orbiforme Parque Estadual de Dois Irmãos (Recife, Pernambuco) URM 83332 JX310813/ JX310827 G. orbiforme Parque Estadual de Dois Irmãos (Recife, Pernambuco) URM 83334 JX310814/ JX310828 G. orbiforme Parque Estadual de Dois Irmãos (Recife, Pernambuco) URM 83335 JX310815/ JX310829 G. orbiforme Parque Estadual de Dois Irmãos (Recife, Pernambuco) URM 83336 JX310816/ JX310830 G. orbiforme Parque Estadual de Dois Irmãos (Recife, Pernambuco) URM 83337 * G. orbiforme Parque Estadual de Dois
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