Studies On Species Diversity Of Rhytismatales In South China And Phylogenetics Of This Order Based On Two-gene Sequences

This study includes two parts. The species diversity of Rhytismatales from South Chinawas researched systematically and comprehensively in the aspects of morphological andanatomical characters, ontogeny, geographic distribution and ecology using moderntechniques for fungal taxonomy in the first part. Based on rDNA-ITS, nLSU rDNAsequences and their joint sequences, phylogeny of Rhytismatales members from nativeplants was researched using molecular biotechnology and the relationship of interfamily,intergenera, interspecies and intraspecies were explored by establishing phylogenetic treesin the second part.More than70specimens which were collected from South China and deposited inthe Reference Collection of Forest Fungi of Anhui Agricultural University, China (AAUF)were examined. According to the taxonomic principles by Darker (1967), Cannon&Minter (1983,1986), Johnston (1986), Spooner (1991), Kirk et al.(2001),1family9genera29species of Rhytismatales were identified. Among them, Lophodermiumcathayae, Terriera simplex and T. trachelosperi were new species; Lophomerumrhododendri was a new record to China; Terriera brevis was a new record to Chinesemainland. Morphologic descriptions and point and line integrated illustrations ormicrograph were given for3new species; illustrations of asci, ascospores and paraphyseswere provided for2newly recorded species; and simple descriptions and discussions ofmorphologic characteristics were given for other species. Keys were compiled foridentification of all species.The reasons of species diversity of Rhytismatales were analyzed in3aspects: host,the characteristics of ontogeny and distributions. The results showed that: the host rangesof Rhytismatales were broad that involved many kinds of plants. Among these plants,gymnosperm was predominant, especially Pinus sp. And in this study,9species ofRhytismatales were collected on pines. However, there were some species having hostspecificity, for example L. cathayae. In a sense of fungal conservation, though L. cathayaecaused the needle blight of Cathaya argyrophylla, this pathogen would disappear followingthe extinction of the “living fossil”, which would be the loss of fungus resource in theworld. Meanwhile, geography had a great effect on the phylogeny of Rhytismatales. Mostof the species being collected in South China had a wide distribution, but there were alsosome species only being seen in South China, for example, L. cathayae and L. guangxiense. In addition, Rhytismataceae was dominating family in this area; and Lophodermium,Coccomyces and Terriera were dominating genera. The identifications of intergenera orinterspecies were showed based on the features of external morphology and internalstructures of ascoma, such as shapes of ascoma, asci, ascospore, paraphyses tips, texturalstructures between covering and basal stroma, absence or presence of lip cells, shapes ofconidioma and conidia, quantity of zone lines and so on.In the second part, the phylogeny of Rhytismatales from various areas of China，especially Yangtze River and its south region was studied. Genomic DNA from isolates oftested strains was extracted from mycelia by the method of CTAB. ITS and28S regionswere amplified by the polymerase chain reaction (PCR). The phylogenetic trees wereestablished based on the sequences of rDNA-ITS, nLSU rDNA and ITS+28S rDNA,selecting Leotia lubrica as outgroup. The classification status of L. cathayae wasconfirmed based on some sequences of rDNA-ITS. The results showed that: species in thesame family or genus and the same species were usually clustered into a group with strongbootstraps, which verified the modern techniques based on phenotypic traits for fungaltaxonomy had important significance and high reference value to the classification ofRhytismatales. However, some species being placed in different genera, even differentfamily with different morphology could also be clustered together, which showed that thetaxonomic results based on classical classification may be limited, and must be combinedwith molecular biotechnology.From the phylogenetic trees, we know that host, geography, altitude, climate andenvironment were related to the relationship of Rhytismatales. Rhytismataceae on conifersand broad leaves and Cudoniaceae on duff were separated into two clades, especially in thetree based on ITS+28S rDNA sequences. In28S-rDNA tree, species from abroad wereclustered together. In rDNA-ITS tree, one group included R. members, of which beyond95%from Shennongjia. In the two-gene phylogenetic tree, Hypoderma rubi fromShennongjia (Hubei Province) couldn’t be clustered with the same species fromHuangshan Scenic Spot and Tiantangzhai (Anhui Province). As a result, it’s not enough tostudy the phylogeny of fungus only by the comprehensive analysis based on two-ormulti-gene. The scientific and reasonable results will be achieved if combining molecularbiotechnology with phenotypic traits of fungi.