Development of a Taqman® Real-Time PCR for the rapid discrimination of the Vibrio splendidus species among the Splendidus clade

The Vibrio splendidus species is ubiquitous in the marine environment but have also been recognized as pathogenic for several aquatic animals. This species belongs to the Splendidus clade which is composed of 16 genetically related species. To our knowledge, there are no available tools able to discriminate the V. splendidus species from the other members of the clade. Thus, we developed a Real Time PCR based on the toxR gene specific to V. splendidus species. Specificity tests were performed on 74 reference strains and 116 field strains and gave 96.05% inclusivity and 100% exclusivity results. The limit of detection of the PCR was determined according to the XP-U47-600-2 AFNOR norm and was estimated at 18 copies genome per reaction. This work describes a reliable, sensible and specific tool to discriminate V. splendidus species from other members of the Splendidus clade and thus could be clearly suitable to perform taxonomic investigations.