Detection of enteropathogenic Vibrio parahaemolyticus, Vibrio cholerae and Vibrio vulnificus: performance of real-time PCR kits in an interlaboratory study

Bacteria belonging to the genus Vibrio are very common to marine and estuarine environments and are found in association with marine plants and animals. Vibrio parahaemolyticus strains that produce thermostable direct hemolysin (TDH) and TDH-related hemolysin (TRH), toxigenic strains of Vibrio cholerae belonging to the serogroups O1 and O139, and Vibrio vulnificus are regarded as important food-borne pathogens, which represent a serious and growing public health hazard. In this study, we established and validated real-time PCR assays for the detection of enteropathogenic Vibrio strains. In a first step, seafood is investigated for the presence of the three Vibrio species. In case of detection of V. cholerae or V. parahaemolyticus, samples are tested for the presence of the cholera toxin gene (ctxA) or tdh/trh genes, respectively, in a second step. All PCR analyses were performed with the same cycling program. Primer/probe sets were thoroughly tested for limit of detection, inclusivity, exclusivity and performance in the matrix. In an interlaboratory study, kits based on these primer/probe sets were successfully tested with cultural and DNA samples.