Identification of species of Tilletiopsis using random amplified polymorphic DNA analysis
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| Authors: | Urquhart, EJ; Sun, LJ; Punja, ZK |
| Year: | 1997 |
| Journal: | Canadian Journal of Plant Pathology-Revue Canadienne de Phytopathologie 19: 380-389 |
| Title: | Identification of species of Tilletiopsis using random amplified polymorphic DNA analysis |
| Abstract: | Tilletiopsis is a genus of dimorphic yeast-like fungi containing six defined species, several of which have shown marked potential as biological control agents of a number of powdery mildew diseases. Identification of isolates of Tilletiopsis is currently based on colony and spore morphology, patterns of carbon and nitrogen utilization, and differences in guanine c cytosine ratios. Since these identification criteria can be time-consuming the use of RAPD analysis of PCR-generated DNA using random primers was evaluated as a method for species identification. Isolates of three Tilletiopsis species from British Columbia - T. minor, T. pallescens and T. washingtonensis - as well as type cultures of these and two additional species, T. albescens and T. fulvescens, were included in this study. The isolates originated from diverse plant hosts and geographical areas. Two isolates of Pseudozyma flocculosa, a biological control agent of rose powdery mildew, were included as an outgroup. The morphological characteristics of the colonies of these isolates are described. Following amplification of DNA using four RAPD primers, the data were subjected to UPGMA cluster analysis. Each of the known Tilletiopsis species could be clearly separated from one another, forming five distinct groups. T. albescens and T. pallescens were grouped closely together, while T. minor and T. washingtonensis formed separate clusters. T.fulvescens grouped closest to T. washingtonensis. The greatest intraspecific variation was observed among isolates of T. washingtonensis, followed by T. minor. Three isolates of Tilletiopsis sp. that were morphologically different from the known species, and which formed a separate cluster, may represent an undescribed species. Isolates of P. flocculosa were readily distinguished from the Tilletiopsis species (similarity coefficient of 0.6) and also formed a distinct group. An isolate tentatively identified as a Tilletiopsis sp. was found to be a Pseudozyma sp. by RAPD analysis. The RAPD-PCR technique resolved the identity of several anomalous isolates of Tilletiopsis that were difficult to identify using current morphological criteria. In addition, unique DNA fingerprints were generated for some isolates that may be useful for monitoring the distribution and spread of specific isolates. The RAPD results indicated that a high degree of genetic variation exists within the genus Tilletiopsis. |
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