Development of a multiplex real-time PCR assay with an internal amplification control for the detection of total and pathogenic Vibrio parahaemolyticus bacteria in oysters.

Vibrio parahaemolyticus is an estuarine bacterium that is the leading cause of shellfish-associated cases of bacterial gastroenteritis in the United States. Our laboratory developed a real-time multiplex PCR assay for the simultaneous detection of the thermolabile hemolysin (tlh), thermostable direct hemolysin (tdh), and thermostable-related hemolysin (trh) genes of V. parahaemolyticus.

Evaluation of an alkaline phosphatase-labeled oligonucleotide probe for the detection and enumeration of the thermostable-related hemolysin (trh) gene of Vibrio parahaemolyticus.

Reliable methods are needed to detect total and pathogenic Vibrio parahaemolyticus. One marker of V. parahaemolyticus virulence is the thermostable-related hemolysin.

A real-time PCR assay for the rapid determination of 16S rRNA genotype in Vibrio vulnificus.

In a terminal restriction fragment polymorphism (T-RFLP) study, we recently reported a significant association between the type B 16S rRNA gene and clinical strains of Vibrio vulnificus associated with the consumption of raw oysters. In the present study we describe a real-time PCR assay for the rapid determination of the 16S rRNA type of V. vulnificus isolates.

Use of a real time PCR assay for detection of the ctxA gene of Vibrio cholerae in an environmental survey of Mobile Bay.

Toxigenic Vibrio cholerae, the etiological agent of cholera, is a natural inhabitant of the marine environment and causes severe diarrheal disease affecting thousands of people each year in developing countries. It is the subject of extensive testing of shrimp produced and exported from these countries. We report the development of a real time PCR (qPCR) assay to detect the gene encoding cholera toxin, ctxA, found in toxigenic V.

Identification of a protein biomarker unique to the pandemic O3:K6 clone of Vibrio parahaemolyticus.

The present method of characterizing Vibrio parahaemolyticus strains involves serotyping or detection methods based on assessment of the presence or absence of genes thought to be markers of an organism's pathogenicity. It is unclear whether these assays detect all pathogenic V. parahaemolyticus strains since a clear correlation between the presence of a particular gene and the organism's pathogenicity has not yet been observed.

Analysis of Vibrio vulnificus from market oysters and septicemia cases for virulence markers.

Representative encapsulated strains of Vibrio vulnificus from market oysters and oyster-associated primary septicemia cases (25 isolates each) were tested in a blinded fashion for potential virulence markers that may distinguish strains from these two sources. These isolates were analyzed for plasmid content, for the presence of a 460-bp amplicon by randomly amplified polymorphic DNA PCR, and for virulence in subcutaneously (s.c.

Molecular, serological, and virulence characteristics of Vibrio parahaemolyticus isolated from environmental, food, and clinical sources in North America and Asia.

Potential virulence attributes, serotypes, and ribotypes were determined for 178 pathogenic Vibrio parahaemolyticus isolates from clinical, environmental, and food sources on the Pacific, Atlantic, and Gulf Coasts of the United States and from clinical sources in Asia. The food and environmental isolates were generally from oysters, and they were defined as being pathogenic by using DNA probes to detect the presence of the thermostable direct hemolysin (tdh) gene. The clinical isolates from the United States were generally associated with oyster consumption, and most were obtained from outbreaks in Washington, Texas, and New York.

Detection of pathogenic Vibrio parahaemolyticus in oyster enrichments by real time PCR.

A real time polymerase chain reaction (PCR) assay was developed and evaluated to detect the presence of the thermostable direct hemolysin gene (tdh), a current marker of pathogenicity in Vibrio parahaemolyticus. The real time PCR fluorogenic probe and primer set was tested against a panel of numerous strains from 13 different bacterial species. Only V.

Seasonal abundance of total and pathogenic Vibrio parahaemolyticus in Alabama oysters.

Recent Vibrio parahaemolyticus outbreaks associated with consumption of raw shellfish in the United States focused attention on the occurrence of this organism in shellfish. From March 1999 through September 2000, paired oyster samples were collected biweekly from two shellfish-growing areas in Mobile Bay, Ala. The presence and densities of V.

Improved recovery of pathogenic Vibrio parahaemolyticus from oysters using colony hybridization following enrichment.

The traditional streak plating and alternative spread-plating methods were compared for detection of pathogenic Vibrio parahaemolyticus (Vp) in oyster enrichments. We found the alternative method to be more efficient: it was quicker (2d vs. 3d) and had a significantly (p < 0.