Abstract
Sixty-seven strains, representing 31 Trichoderma and 3 Hypocrea species, were used to study the relationships of species within the genus Trichoderma in this work. All the strains were reexamined to certify the strain correctly identified with morphological characters. The results demonstrated that sixty-two of the examined strains were correctly identified before, and another six strains were proven evidently misidentified. Among the six strains, W10, G39, CCRC 32893, and CCRC33565, regarded as T. koningii before, should be reidentified as T. longibrachiatum, T. reesei, T. pseudokoningii, and T. fasculatum, respectively, and T. lignorum CCRC33613 and T. pseudokoningii CCRC33607 were reassigned as T. asperellum and T. longibrachiatum, respectively. The genetic relatedness of some Trichoderma species shared microscopically similar morphology were verified with molecular approaches including internal transcribed spacer (ITS) sequences of ribosomal DNA, universal primer-polymerase chain reaction (UP-PCR) fingerprinting, and DNA-DNA hybridization. In the analysis of ITS and 5.8S sequences of ribosomal DNA, parsimony analysis demonstrated that species of different sections were distributed in different branches, and species within sections Saturnisporum, Longibrachiatum and Trichoderma were clustered into the same branch, respectively. However, Trichoderma sect. Pachybasium species showed polyphynetic and were clustered into three separated branches. Some species having morphologically similar species revealed showed nearly identical or smaller ITS sequence differences (less than 1-3 bp), they also produced highly similar or identical UP-PCR profiles and high values of DNA complementarity (>70%). In some cases, molecularly and morphologically similar species were found, including T. hamatum and T. pubescens; T. croceum, T. polysporum and T. album; T. crassum and T. flavofuscum; T. strictipile and T. fasciculatum; and T. ghanense and T. paceramosum. Although every species can be differentiated morphologically, there were some species showed highly similar molecular characteristics, indicating that they could be conspecific. However, in some cases (T. longipile, T. crassum and T. flavofuscum; T. fertile and T. minutisporum; T. tomentosum, T. inhamatum and T. harzianum; T. longibrachiatum and T. reesei; T. citrinoviride and T. parceramosum/T. ghanense; T. viride, T. atroviride and T. koningii) there were discriminative patterns of UP-PCR and/or low levels (<50%) of DNA-DNA hybridization even their ITS sequences were similar, suggesting a closely phylogenetic relationship. For intraspecific variation, eight strains of T. koningii could be divided into three groups, termed group I, II, and III, based on different PCR fingerprinting profiles even though all the species showed identical or highly similar (less than 1-3 bp difference) sequences. Among the three groups, strain CCRC33557 in type III were linked with the proposed teleomorph, Hypoccrea ceramica based on morphological and molecular characteristics.