Affiliation:
1. Rapid Microbial Detection Facility, Department of Food Science, Louisiana State University Agricultural Center, Baton Rouge, Louisiana 70803
Abstract
ABSTRACT
Vibrio cholerae
is recognized as a leading human waterborne pathogen. Traditional diagnostic testing for
Vibrio
is not always reliable, because this bacterium can enter a viable but nonculturable state. Therefore, nucleic acid-based tests have emerged as a useful alternative to traditional enrichment testing. In this article, a TaqMan PCR assay is presented for quantitative detection of
V. cholerae
in pure cultures, oysters, and synthetic seawater. Primers and probe were designed from the nonclassical hemolysin (
hly
A) sequence of
V. cholerae
strains. This probe was applied to DNA from 60 bacterial strains comprising 21 genera. The TaqMan PCR assay was positive for all of the strains of
V. cholerae
tested and negative for all other species of
Vibrio
tested. In addition, none of the other genera tested was amplified with the TaqMan primers and probe used in this study. The results of the TaqMan PCR with raw oysters and spiked with
V. cholerae
serotypes O1 and O139 were comparable to those of pure cultures. The sensitivity of the assay was in the range of 6 to 8 CFU g
−1
and 10 CFU ml
−1
in spiked raw oyster and synthetic seawater samples, respectively. The total assay could be completed in 3 h. Quantification of the
Vibrio
cells was linear over at least 6 log units. The TaqMan probe and primer set developed in this study can be used as a rapid screening tool for the presence of
V. cholerae
in oysters and seawater without prior isolation and characterization of the bacteria by traditional microbiological methods.
Publisher
American Society for Microbiology
Subject
Ecology,Applied Microbiology and Biotechnology,Food Science,Biotechnology
Cited by
104 articles.
订阅此论文施引文献
订阅此论文施引文献,注册后可以免费订阅5篇论文的施引文献,订阅后可以查看论文全部施引文献