Phylogenetic Relationships Among Colletotrichum Pathogens of Strawberry and Design of PCR Primers for their Identification

Isolates of Colletotrichum acutatum, C. fragariae and C. gloeosporioides pathogenic to strawberry plants were examined by sequence analysis of the 5.8S-ITS region. Phylogenetic relationships among isolates of Colletotrichum are, for the most part, congruent with the molecular groups established in earlier works. 5.8S-ITS sequence analysis showed a high level of genetic divergence within C. acutatum. Isolates of this species clustered into two very distinct clusters with further subdivision. The divergences between C. fragariae and C. gloeosporioides were too low to distinguish themas separate species. On the basis of the sequence data, specific primers were designed both to identify isolates belonging to the genus Colletotrichum, and to distinguish isolates of the species C. acutatum. The specificity of these primers was validated by testing a wide range of strawberry isolates of Colletotrichum, non-strawberry isolates of Colletotrichum and other fungi used as controls. Although the 5.8S-ITS sequences were not polymorphic enough to allow the construction of C. gloeosporioides-specific primers, specific PCR amplification followed by an MvnI digestion provides a tool to specifically identify strawberry isolates of C. gloeosporioides.