Sensitive and specific detection of Xanthomonas campestris pv. zinniae by PCR using pathovar-specific primers

A sensitive and specific assay was developed to detect bacterial leaf and flower spot disease on zinnia caused by Xanthomonas campestris pv. zinniae (Xcz). A draft genome sequence was determined for Xcz strain E4, isolated from zinnia in Jilin province, China. Comparative genomics analyses were utilized to identify a sequence specific to Xcz in the unique gene 0834. This sequence region was amplified as 1044 bp DNA fragment using designed primers 0834F and 0834R. Using this primer set, specific PCR products were only amplified from DNA from Xcz strains and not from other pathovars of X. campestris, other species of Xanthomonas, or from other genera such as Pseudomonas and Bacillus. The limit of PCR detection in pure culture suspension was approximately 1.7 × 103 CFU/ml per reaction. Specific amplification of the fragments for Xcz was sensitive relatively to the technique used, detecting as low as 0.1 pg template DNA. The PCR technique was also applied to detect the Xcz pathogen in naturally infected leaves and seeds of zinnia.