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1 菌物学报 15 May 2013, 32(3): Mycosystema ISSN CN /Q 2013 IMCAS, all rights reserved. A new species of Chlorencoelia from Parque Nacional El Ávila, Venezuela ITURRIAGA Teresa * MARDONES Melissa Laboratorio de Micología, Departamento de Biología de Organismos, Universidad Simón Bolívar, Baruta, Edo. Miranda, Venezuela Abstract: A new species of the genus Chlorencoelia was collected on wood in a tropical cloud forest at El Avila National Park, presently called by its original Indian name Waraira-Repano, located in the north of Caracas in northern Venezuela. The new species can be distinguished from the known species in the genus by its small-sized asci and ascospores, ellipsoid to sigmoid to clavate ascospores, and the presence of a greenish-brown gelatinized layer covering external tomentum hyphae. Key words: discomycetes, neotropics, taxonomy 委内瑞拉产绿散胞盘菌属一新种 ITURRIAGA Teresa * MARDONES Melissa Laboratorio de Micología, Departamento de Biología de Organismos, Universidad Simón Bolívar, Baruta, Edo. Miranda, Venezuela [ 委内瑞拉 ] 摘要 : 从 El Avila 国家公园热带云雾森林中发现了绿散胞盘菌属的一个新种 它以其小型的子囊和子囊孢子 椭圆 -S 型 - 棒状的子囊孢子以及覆盖于外囊盘被菌丝表面的绿褐色胶化层为显著特征, 并与该属已知种相区别 关键词 : 盘菌, 新热带区, 分类 The genus Chlorencoelia J.R. Dixon was erected by Dixon (1975), with C. versiformis (Pers.) J.R. Dixon as the type species. Currently the genus comprises three species: C. indica (K.S. Thind, E.K. Cash & Pr. Singh) W.Y. Zhuang, C. torta (Schwein.) J.R. Dixon and C. versiformis. It is characterized by a stipitated, cupulate to infundibuliform apothecium, gelatinized ectal excipulum of textura angularis to textura globulosa and with hyaline to dark-walled cells which give rise to filamentous or subglobose to clavate tomentum hyphae, pore of the ascus always blue in iodine, and ellipsoid to allantoid hyaline ascospores (Dixon 1975). Members of this genus show morphological similarities with the genus Cordierites Mont., such as the coloration of their apothecia and an ectal excipulum composed of * Corresponding author. Received: , accepted:

2 458 ISSN CN /Q Mycosystema May 15, 2013 Vol.32 No.3 angular cells usually having the hyphae oriented perpendicular to the outer surface (Dixon 1975), but Chlorencoelia differs in having both its cells of the ectal excipulum and hyphae of the medullary excipulum immersed in a gelatinous matrix, ascus apices strongly blue in iodine, yellow-orange to lilaceous-green apothecia lacking of an ionomidotic reaction in KOH (Zhuang 1988). Traditionally Chlorencoelia was considered a member of the family Helotiaceae (Helotiales), subfamily Encoelioideae. Zhuang et al. (2000) based on 18 rdna sequences of six of the 18 genera (Chlorencoelia, Cordierites, Encoelia, Holwaya, Unguiculariopsis and Velutarina) commonly accepted in this subfamily, conclude that the Encoelioideae is not monophyletic and recognized several lineages, as well that Chlorencoelia torta (Schwein.) J.R. Dixon was not closely related to any of the genera used in their study (Zhuang et al. 2000). Recent molecular studies based on three rdna regions (LSU+SSU+5.8S) show that the members of this genus form a clade with another helotiaceous genus Heyderia and members of the family Hemiphacidiaceae: Fabrella tsugae (Farl.) Kirschst., Meria laricis Vuill. and Hemiphacidium longisporum Ziller & A. Funk (Wang et al. 2006b). The family Hemiphacidiaceae contains foliar pathogens producing small, simple apothecia beneath the leaf surface, erumpent apothecia pushing back the host tissue (Korf 1962). In contrast, species of Heyderia and Chlorencoelia produce large apothecia with a well developed excipulum. According to Wang et al. (2006a) the family Hemiphacidiaceae might need to be expanded to include the genera Chlorencoelia and Heyderia. Traditionally, species of Chlorencoelia have been regarded as saprobes, but current studies using molecular analyses suggest that some of these saprophytic fungi probably colonize their hosts as endophytes, and then produce apothecia after the hosts die (Wang et al. 2006a, b). During studies of discomycetes from cloud forests located in El Avila National Park in northern Venezuela, an interesting species of Chlorencoelia was collected on decomposed wood. The species has the morphological characteristics of the genus Chlorencoelia, but does not resemble any known species described in this genus, and is therefore treated here as new. 1 MATERIAL AND METHODS Protocols for morphological examination and sectioning followed those outlined in Iturriaga & Korf (1990). Cotton blue in lactic acid was used to observe structures and tissues in sections and squashed material. Melzer s reagent was used to observe the iodine reaction of ascus apex. 2 TAXONOMY Chlorencoelia ripakorfii Iturr. & Mardones, sp. nov. Figs. 1, 2 MycoBank MB Apothecia superficial, gregarious, discoid to infundibuliform, becoming convex-expanded, unequally sided, stipitated, 2 5mm diam., mm high. Receptacle whitish-yellow, with a pruinose greenish cover. Disc whitish-yellow, lighter than the receptacle. Ectal excipulum in two layers: outer layer composed of two rows of gelatinized dark-walled cells, textura angularis to textura porrecta, in median section 25µm wide at the flanks, 15µm thick at the margin, giving rise to filamentous tomentum hyphae oriented nearly perpendicular to the receptacle surface; inner layer composed of hyaline hyphae in a gel, textura oblita, up to 25µm thick. Tomentum hyphae cylindrical to slightly clavate, septate, µm, cells hyaline to

3 ITURRIAGA Teresa & MARDONES Melissa / A new species of Chlorencoelia from Parque Nacional El Ávila, Venezuela 459 Fig. 1 Chlorencoelia ripakorfii (holotype). A, B: Fresh apothecia growing on dead wood; C: Longitudinal median section through an apothecium; D: Tomentum hyphae arising from the outer ectal excipulum, covered by a gelatinized matter; E: Detail of a tomentose hypha; F: Ascus with eight ascospores; G: Seven Ascospores. Scale bars: A=0.2mm; B=0.5mm; C,D=100μm; E G=10μm. Fig. 2 Chlorencoelia ripakorfii (holotype). A: Ascus with eight ascospores; B: Ascospores of different shapes, drawn from spores outside the asci; C: Longitudinal median section showing to mentum hyphae, and both layers of the ectal excipulum. Bars: A,B=10µm; C=100μm. Mycosystema

4 460 ISSN CN /Q Mycosystema May 15, 2013 Vol.32 No.3 pale-brown thick-walled, containing a green pigment and covered by a greenish-brown gelatinized layer. Medullary excipulum of hyaline to pale brown gelatinized hyphae, textura oblita, 45µm thick. Subhymenium not observed. Hymenium hyaline to pale brown, 60 70µm high. Asci hyaline, cylindrical to clavate, with long tapering stalks, ascus pore J+, (42 ) [ x 54.6 m 55.0 s5.8 n 22]µm 3 4 [ x 3.9 m 4.0 s0.2 n22]µm, 8-spored. Ascospores uniseriate, hyaline, ellipsoid, sigmoid or pyriform, with one end broader than the other, non-septate, with one to two guttules, covered by a thin gelatinized sheath, 5 8 [ x 6.5 m 7.0 s 0.8 n 30]µm 1,5 3 [ x 2.6 m 3.0 s 0.5 n 30]µm. Paraphyses hyaline, filiform, rounded at the apex, flexuous, aseptate, 1 1.5µm wide, exceeding the asci up to 7.0µm. Holotype: Venezuela, Distrito Capital, El Avila National Park, cloud forest close to Humboldt Peak N, W, 2,000m, on dead wood, 02 May 2007, M. Mardones (AV-120), VEN Etymology: The epithet ripakorfii is dedicated to honor Richard Paul Korf, a leader in Discomycete Systematics and, PhD thesis Major Professor of T. Iturriaga, to be published in an invited contribution for a special issue dedicated to him in the Chinese journal Mycosystema (will be published in May 2013) on the occasion of his 88th birthday, when under the Chinese tradition he will become a Mi-shou. Key to the species of Chlorencoelia 1a. Filamentous tomentum hyphae arising from cells of the outer ectal excipulum 2 1b. Subglobose to clavate to angular tomentum hyphae arising from cells of the outer ectal excipulum 3 2a. Tomentum hyphae of 5 7µm diam., containing green pigments, without a gelatinous gel covering; ascospores oblong to allantoid, µm C. versiformis 2b. Tomentum hyphae of 2 4µm diam., without green pigments, covered by a greenish-brown gelatinized matter; ascospores allantoids to sigmoid to pyriform, with one end broader than the other, µm C. ripakorfii 3a. Tomentum hyphae subglobose to angular, brown, without green granules; ascospores with two large guttules C. indica 3b. Tomentum hyphae subglobose to clavate, hyaline, containing green granules; ascospores non-guttulate C. torta 3 DISCUSSION Chlorencoelia ripakorfii is easily distinguished from other species in the genus by small sized asci and ascospores, sigmoid to clavate to pyriform ascospores, and having tomentum hyphae covered by a greenish-brown gelatinized matrix. This combination of characters does not occur in any of the published species in the genus. Morphological similarities between C. ripakorfii and C. versiformis are the presence of filamentous tomentum hyphae and a hyaline medullary excipulum. However, C. ripakorfii can be distinguished from C. versiformis by its smaller asci and ascospores and its ascospore shape. Furthermore, in the new species, tomentum hyphae are longer, lacking green granules and are covered

5 ITURRIAGA Teresa & MARDONES Melissa / A new species of Chlorencoelia from Parque Nacional El Ávila, Venezuela 461 by a greenish-brown gelatinized matter. Tomentum hyphae that arise from the ectal excipulum are a diagnostic character in the classification of Chlorencoelia. Among the four species of the genus, two have filamentous tomentum hyphae (C. ripakorfii and C. versiformis) while the other two are subglobose to clavate to angular (C. indica and C. torta). These tomentum hyphae have been reported as containing a green pigment in C. versiformis and the new species, or green granules in C. torta, but the presence of a greenish-brown gelatinized matrix has not been previously reported for any species of the genus. This gelatinous matter is apparently hydrophobic while it disappears under cotton blue stain and Meltzer s reagent. Acknowledgements: We are grateful to FONACIT, (Fondo Nacional de Ciencia, Innovacion y Tecnología) Caracas-Venezuela for funding the project No S : Sistemática Molecular y Morfológica de Hongos Ascomycetes y Basidiomycetes presentes en las Sucesiones Fúngicas Descomponedoras de Madera en Bosques Tropicales under the direction of TI which permitted research in a forest located in Luepa, Parque Nacional Canaima, as well as collaborating in the pursual of several of the projects being undertaken in the Mycology Lab at Simón Bolívar University (USB) during that same period. We also extend our thanks to the Decanato de Estudios de Postgrado of USB for financial support. We would like to acknowledge the support of INPARQUES in giving permits to collect at El Avila National Park, for facilities during field work, and to Jesús Hernández and Carlos Rondón for field assistance. MM would like to acknowledge the Department of Organisms Biology and the Decanato de Investigacion y Desarrollo of Simón Bolívar University, as well as the Ministerio de Ciencia y Tecnología from Costa Rica for financial support during her Master Degree in Venezuela. [REFERENCES] Dixon JR, Chlorosplenium and its segregates II. The genera Chlorociboria and Chlorencoelia. Mycotaxon, 1: Iturriaga T, Korf RP, A monograph of the discomycete genus Strossmayeria (Leotiaceae), with comments on its anamorph Pseudospiropes (Dematiaceae). Mycotaxon, 36: Korf RP, A synopsis of the Hemiphacidiaceae, a family of the Helotiales (Discomycetes) causing needle blights of conifers. Mycologia, 54: Thind KS, Cash EK, Singh P, The Helotiales of the Mussoorie Hills-II. Mycologia, 51: Wang Z, Binder M, Schoch CL, Johnston PR, Spatafora JW, Hibbett DS, 2006a. Evolution of helotialean fungi (Leotiomycetes, Pezizomycotina): a nuclear rdna phylogeny. Molecular Phylogenetic and Evolution, 41: Wang Z, Johnston PR, Takamatsu S, Spatafora JW, Hibbett DS, 2006b. Toward a phylogenetic classification of the Leotiomycetes based on rdna data. Mycologia, 98: Zhuang WY, Studies on some discomycete genera with an ionomidotic reaction: Ionomidotis, Poloniodiscus, Cordierites, Phyllomyces, and Ameghiniella. Mycotaxon, 31: Zhuang WY, Yu ZH, Wu WP, Langue C, Fouret N, Preliminary notes on the phylogenetic relationships in the Encoelioideae inferred from 18S rdna sequences. Mycosystema, 19: Mycosystema

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