Genetic differentiation of Phoma ligulicola isolates from pyrethrum in Tasmania

Phoma ligulicola is the most severe fungal pathogen affecting pyrethrum crops in Tasmania, Australia. It causes the disease, ray blight, which is associated with necrotic lesions on leaves, at nodes on stems, and a failure of the flower to develop causing a decrease in yield and pyrethrin content in the flowers. The internal transcribed spacer regions (ITS1 and ITS 2) of the ribosomal DNA genes from 11 isolates of P. ligulicola were amplified using the polymerase chain reaction (PCR) and sequenced. Phylogenetic trees were produced by combining the sequences of the ITS regions and comparison to the published sequences of other Phoma spp. and other closely related fungi. Isolates were differentiated into three subgroups, P1, P2, and P3 through phylogenetic analysis. The closest match to the ITS sequences of other fungi were Phoma herbarum (86.5 – 91.5% homology) and Didymella bryoniae (88.2 to 93.6% homology). Primers were designed to the ITS regions that could be used to detect all three subgroups. Further work will concentrate on the biological implications of this molecular diversity and the development of a nested PCR test for the simultaneous characterisation of the three subgroups.