Development of multiplex PCR for the specific detection of Xanthomonas campestris pv. campestris in cabbage and correlation with disease incidence

Black rot, the most serious disease of crucifers especially Brassica oleracea var. capitata (Cabbage) causes huge yield losses. Black rot is a seed-borne disease and the assessment of seeds for the pathogen is very crucial. The cabbage seeds were tested for X. campestris pv. campestris infection by biochemical and PCR analysis. The seed samples were subjected to PCR analysis with three primers, out of which one of the pairs of primers specific for hrpF gene could detect the black rot pathogen. The current study is the first of its kind wherein the locally available cabbage cultivars were assessed using molecular assays and the widely grown cultivars in the study area were checked for the correlation between laboratory and field performances. The aim of the study was to use the available specific and sensitive PCR-based procedures for routine detection of X. campestris pv. campestris in cabbage seeds. Multiplex PCR was standardized for the simultaneous detection of pathogen and Brassica species, internal transcribed spacer (ITS) in infected seed and leaf material. The multiplex PCR fingerprints were obtained which amplified a 619 bp fragment of the hrpF gene from X. campestris pv. campestris and a 360 bp section of the internal transcribed spacer region from Brassica sp. The assay also readily detected X. campestris pv. campestris infections in diseased plants and from bacterial colonies isolated on selective media, and was more sensitive and specific than traditional plating methods. Thus by the use of multiplex PCR, determination of the threshold level for bacterial detection in cabbage seeds was possible