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Molecular identification and multilocus phylogeny of Ophiosphaerella species associated with spring dead spot of bermudagrass

Abstract:

Spring dead spot (SDS) is a devastating disease of bermudagrass [Cynodon dactylon (L.) Pers.], a widely used turfgrass in the transition zone of the United States. The fungi causing SDS have been identified as belonging to three species of the genus Ophiosphaerella based on cultural characters and the morphology of seldom encountered pseudothecial stages. The three species [O. herpotricha (Fr.) Walker, O. korrae (Walker & Smith) Shoemaker & Babc., and O. narmari (Walker & Smith) Wetzel, Hulburt & Tisserat] usually form only sterile mycelia in culture, preventing accurate morphological identification. Molecular identification has been hampered by a lack of available sequences, the presence of primer-disrupting introns in the ribosomal small subunit (SSU), and uncertain species differentiation due to interspecies similarity of available ribosomal sequences. In this study, sequences of the ribosomal DNA (SSU–internal transcribed spacer–partial large subunit), translation elongation factor 1-α (EF1α) and RNA polymerase II second largest subunit (RPB2) genes from 79 isolates of SDS-associated Ophiosphaerella spp. from 27 locations in North America and Australia were used to resolve the phylogeny of Ophiosphaerella in the Pleosporales and to determine an informative locus (“barcode”) for species identification. Single-locus and multilocus phylogenies group isolates of SDS-associated Ophiosphaerella in monophyletic clades corresponding to the species O. herpotricha, O. korrae, and O. narmari form a monophyletic lineage with the Asian O. agrostidis (Dern, Câmara, O’Neill, Berkum & Palm) isolates within Phaeosphaeriaceae. This lineage is distinct from the European O. herpotricha isolates, which group in a separate clade in Phaeosphaeriaceae. The inferred polyphyly of the genus Ophiosphaerella leaves the generic placement of SDS-associated species uncertain. Based on the high interspecific and low intraspecific variability of EF1α, this gene should prove useful as a barcode for species identification of SDS-associated Ophiosphaerella.