[Quantitative detection of Vibrio parahaemolyticus by real-time TaqMan PCR]

Vibrio parahaemolyticus is one of important human food pathogens. Traditional diagnostic tests for V. parahaemolyticus are laborious and always present false negative results. Therefore, it is important to develop a nucleic acid-based test for quantitative detection of V. parahaemolyticus. A TaqMan PCR assay was presented for quantitative detection of V. parahaemolyticus in pure cultures and oysters. The primers and probe were designed according to the gyrase B gene (gyrB) sequence of V. parahaemolyticus strains. Amplification of DNAs from 12 bacterial strains comprising 9 genera showed that all of the strains of V. parahaemolyticus tested (n = 4) were positive and all other species of strains tested (n = 8) were negative. The results of the TaqMan PCR with raw oysters inoculated with V. parahaemolyticus were comparable to those of pure cultures. The sensitivity of the assay was 1 CFU PCR Mixture(-1) and 10 CFU PCR Mixture(-1) in pure culture and inoculated raw oyster, respectively. The correlation rate was 0.99 (gamma2 = 0.99). The assay could be completed within 1h. The Real-time PCR can be used as a rapid screening tool for the presence of V. parahaemolyticus in seafood without prior isolation and characterization of the bacteria by traditional microbiological methods.