Development of simple sequence repeat markers for the classification of the clubroot pathogen Plasmodiophora brassicae

Clubroot disease caused by Plasmodiophora brassicae is one of the most serious diseases in cruciferous crops. To classify isolates, we developed simple sequence repeat (SSR) markers for P. brassicae. Twenty-four Japanese isolates were used in this study: 12 isolates of an unknown pathotype from the Kyoto Prefecture, as well as 12 isolates of known pathotypes, including three single-spore lines. From the 12 isolates from Kyoto Prefecture, 11 were classified into either pathotype 2 (three isolates) or 4 (eight isolates). We designed 23 SSR markers based on the P. brassicae genome, of which 11 markers from intergenic regions showed polymorphisms in the 24 isolates. Many haploid isolates belonging to pathotypes 2 and 4 were monomorphic, and typical alleles were detected in some isolates not belonging to pathotype 4. Two bands were detected for eight SSR loci in five isolates, indicating that different genotypes were mixed in these isolates. We constructed a phylogram based on the 11 polymorphic SSRs. Pathotypes 2 and 4 formed a cluster, from which pathotypes 3 and 1 were successively placed. These results strongly suggest a close genetic relationship between isolates in pathotypes 2 and 4, consistent with our finding that isolates in these two pathotypes were found at one collection site. In combination with pathotype classification and other marker systems, the SSR markers can be used for more detailed analyses to improve the control of clubroot disease.