Evaluation of different polymerase chain reaction methods for the identification of Vibrio parahaemolyticus strains isolated by cultural methods

• Published in: Journal of AOAC International 2007-11, Vol.90 (6), p.1588-1597
• Main Authors: Croci, L, Suffredini, E, Cozzi, L, Paniconi, M, Ciccaglioni, G, Colombo, M.M
• Format: Article
• Language: English
• Subjects: bacterial contamination; Bacterial Toxins - chemistry; Bacterial Toxins - genetics; Base Sequence; Biochemistry; Biological and medical sciences; chemotaxonomy; Control; Culture Media; DNA, Bacterial - chemistry; DNA, Bacterial - genetics; Electrophoresis, Polyacrylamide Gel; enrichment culture; fish; fish products; Fish Products - microbiology; Fisheries; food contamination; Food industries; food pathogens; Fundamental and applied biological sciences. Psychology; genes; Identification and classification; isolation; Methods; Molecular Sequence Data; molecular systematics; nucleotide sequences; Polymerase chain reaction; Reference Standards; Reverse Transcriptase Polymerase Chain Reaction; serotypes; Vibrio; Vibrio parahaemolyticus; Vibrio parahaemolyticus - chemistry; Vibrio parahaemolyticus - genetics; virulence
• ISSN: 1060-3271; 1944-7922
• DOI: 10.1093/jaoac/90.6.1588

Control of contamination by Vibrio parahaemolyticus in fishery products is often hampered by the lack of standardized methods and by the uncertainty associated with biochemical identification of the isolates. In this study, 5 polymerase chain reaction (PCR) methods for the identification of V. parahaemolyticus to the species level were evaluated by using 25 Vibrio reference strains and 163 isolates from fishery products, environmental sources, and clinical samples. Sequence targets of the methods were toxR, gyrB, and tlh genes (tested with 2 protocols), and the fragment pR72H. Isolate identification was confirmed by sequencing of the 16S rRNA gene and by PCR protocols for the identification of other Vibrio species. The PCR assay targeting the toxR gene achieved the highest performance (100% inclusivity and exclusivity). The 2 PCR protocols based on tlh gene detection, although showing the same inclusivity (100%), differed in the exclusivity (50 and 91%, respectively). Finally, the results provided by the PCR assays targeting the gyrB gene and pR72H fragment were less reliable and, in some cases, difficult to assess. According to the results of this study, the characteristics of accuracy expressed by the toxR identification method make it a suitable candidate as a reference method for the molecular identification of V. parahaemolyticus strains.