Development of species-specific primers for detection of xanthomonas campestris pv. campestris causing black rot of crucifers

Abstract

Aim: Black rot of crucifers caused by Xanthomonas campestris pv. campestris (Pammel) Dowson (Xcc) is a major seed-borne disease. The present study aimed to develop a rapid diagnostic protocol for the specific and sensitive detection of this pathogen. Methodology: Aspecific primerset was designed based on rpf gene and optimization of PCR condition was done for specific detection of Xcc. Sensitivity of PCR for primer set was then determ ined by diluting the Xcc DNAandcells. Results: Specific primer set was able to amplify a specific band of 304 bp inall 11 isolates of Xcc but failed to amplify other Xanthomonas species and one each of Ralstonia solanacearum. Erwinia caratovora subsp. caratovora. Bacillus subtilis. Pseudomonas fluorescens and P. aeruginosa. The primer set was highly sensitiveas it was able to detect 10 pg pi'1 bacterial DNAand up to 3x103CFU ml1 corresponding to 12 viable cells of Xcc which were used as template for PCR reaction. Interpretation : The results suggest that develope d PCR primers are highly specific and sensitive and it can be used to detect the pathogen at an early stage of infection for disease management.