Design and testing of multiplex-PCR primers for detection of bacterial spot of tomato

The subject of this work is to develop multiplex-PCR assay for specific detection of plant pathogenic bacteria of Xanthomonas genus causing bacterial spot of tomato. PCR primers for detection of groups A (X. euvesicatoria), B (X. vesicatoria), C (X. perforans) and D (X. gardneri) were developed based on the DNA sequences obtained by sequencing and from the GenBank database (NCBI). Four primer pairs - Xe_shotgun_104, Xe_shotgun_1819, Xv_atpD_403, Xp_efP_202 were designed and subsequently thoroughly tested and optimized for parallel detection of these bacteria. Specificity of the primers was tested on a large complex of bacterial strains pathogenic to tomato and related crops. Following the protocol described above X. vesicatoria, X. euvesicatoria, X. perforans and X. gardneri can be quickly and reliably identified in a single multiplex-PCR assay