Phylogenetic relationships and taxonomic placement based on rDNA, histone H4 and beta-tubulin genes of endophytic fungi isolated from Tripterygium wilfordii Hook F

Abstract

Tripterygium wilfordii Hook f. (TwHf) is a traditional Chinese medicinal plant has long been used to treat various autoimmune disorders since ancient times. TwHf is reported hosting abundant endophytic fungi, some of which may produce bioactive metabolites possessing immunosuppressive effects. Hence, effective isolation, systematics and phylogenetic analyses of these endophytic fungi are indispensable prerequisite requirements for possible discovery of new therapeutic drugs. The objectives of this project are mainly to (1) isolate endophytic fungi from various parts of TwHf; (2) conduct molecular characterizations of ribosomal DNA (ITS1-5.8S-ITS2), histone H4 and beta-tubulin genes; (3) construct phylogenetic relationships and study taxonomic placement of endophytic fungi; (4) infer usefulness of each gene in probing evolutionary relationships or phylogenetics; and (5) verify usefulness of histone H4 gene for phylogenetics and taxonomic identification among closely related fungal species. In this study, 265 fungal isolates clustered into 85 morphotypes were isolated from TwHf. Genera Glomerella (32%), Colletotrichum (29%), Diaporthe (13%) and Phomopsis (9%) are the most abundant fungal endophytes isolated. Genera Colletotrichum and Glomerella infect prevalently in xylems whereas Diaporthe and Phomopsis exist mainly in leaves of TwHf. The greatest species diversity is in flowers where four fungal genera are co-existed in a relatively small habitat. This is an indication that fungal endophytes may utilize vertical transmission to increase likelihood of propagation. Taxonomic placement analyses reveal that fungal endophytes may possibly evolve from soil fungi. Phylogenetic analyses reveal that genera Colletotrichum and Glomerella as well as Diaporthe and Phomopsis are distinctive closely related siblings. However, genera Colletotrichum and Glomerella evolved six lineages earlier than Diaporthe and Phomopsis siblings. Phylogenetic analyses based on ITS of rDNA, histone H4 and partial beta-tubulin genes were conducted, trees using single and multi-gene genealogies were constructed by Neighbor-Joining, Maximum Parsimony, Maximum Likelihood and Bayesian analyses. Phylogenetic analyses reveal that histone H4 coding sequences alone are too conserved for ordinary phylogeny. However, it yields excellent phylogeny resolution especially in intra-species when introns are included. Beta-tubulin gene contains fewer parsimony-informative characters than ITS of rDNA in resolving intra-species phylogeny. Multi-gene based phylogenetic tree shows great improvement on taxonomic resolution. Thus, combined gene datasets may be a good alternative to complement insufficiency of individual gene to obtain more reliable phylogeny.

Date of Award	16 Jul 2012
Original language	English
Awarding Institution	City University of Hong Kong
Supervisor	Hon Yeung CHEUNG (Supervisor)