Direct Detection of Vibrio vulnificus, Vibrio parahaemolyticus, and Vibrio alginolyticus from Clinical and Environmental Samples by a Multiplex Touchdown Polymerase Chain Reaction Assay.

Background:Vibrio vulnificus, V. alginolyticus, and V. parahaemolyticus are commonly and opportunistically pathogenic to humans. Methods: In this study, a novel multiple touchdown polymerase chain reaction method (MT-PCR) was developed to benefit rapid and simultaneous detection of the presence of the three Vibrio species from the enriched clinical and environmental samples. Results: The method showed a sensitivity of 10(4) colony forming units (CFU)/mL for V. vulnificus, 10(3) CFU/mL for V. parahaemolyticus and V. alginolyticus, and a specificity of 100% for all the three Vibrio species. All strains of the three Vibrio species were detected in the spiked samples artificially contaminated with reference strains and were identified directly from the enriched clinical and environmental samples within three hours by this MT-PCR assay. All the corresponding bacteria were isolated from these enriched samples in 48 hours by standard microbiologic procedures. Conclusions: This MT-PCR method, which can detect V. vulnificus, V. parahaemolyticus, and V. alginolyticus directly and simultaneously, was rapid, sensitive, specific, and can be used in clinical diagnostics, food industry studies, and risk assessment of environment.