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KMID : 0545119950050040181
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Journal of Microbiology and Biotechnology
1995 Volume.5 No. 4 p.181 ~ p.187
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Rapid and Direct Detection of Vibrio vulnificus in Small Octopus (Octopus variabilis) Using Polymerase Chain Reaction
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Lee Jee-Yeon
Choi Sang-Ho
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Abstract
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The cells of Vibrio vulnificus can be induced to the viable but nonculturable (VBNC) state by natural environmental parameters. The V. vulnificus cells in the VBNC state can not be recovered by ordinary laboratory techniques. This nonculturability could often hamper development of effective processing strategies to minimize the number of V. vulnificus in seafoods. Even with V. vulnificus cells in a culturable state, the length of time required to identify the bacteria in contaminated food by phenotyphic characterization may prevent appropriate in-time responses by public health agencies to infections of the bacteria. In the present study, we used polymerase chain reaction (PCR) to develop a rapid and direct detection method for V. vulnificus in small octopus (Octopus variabilis) which is consumed as a raw food in Korea. The region targeted was a 704-base pair (bp) portion of the hemolysin gene, vvhA, of V. vulnificus. The primers designed for PCR amplification were specific for all V. vulnificus sp. tested. Several methods were examined to extract total DNA directly from V. vulnificus seeded into the octopus homogenate and the guanidine isothiocyanate (GITC) method appeared to be most effective. From the octopus homogenate seeded by V. vulnificus at an initial level of 10^2 CFU/㎖ of the homogenate and then incubated for 12 h, the targeted sequence was successfully amplified by PCR and the 704-bp DNA fragment was observed by gel electrophoresis. The total completion of this assay requires less than one day.
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