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KMID : 1120220130040030133
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Osong Public Health and Research Perspectives
2013 Volume.4 No. 3 p.133 ~ p.139
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Multiplex Real-time Polymerase Chain Reaction Assays for Simultaneous Detection of Vibrio cholerae, Vibrio parahaemolyticus, and Vibrio vulnificus
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Park Jie-Yeun
Jeon Se-Mi
Kim Jun-Young
Park Mi-Sun
Kim Seong-Han
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Abstract
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Objectives: A multiplex real-time polymerase chain reaction (RT-PCR) method was developed for the identification of three Vibrio species: Vibrio cholerae, Vibrio parahaemolyticus, and Vibrio vulnificus.
Methods: Specific primers and probes targeting the hlyA, tlh, and vvhA genes were selected and used for multiplex real-time PCR to confirm the identification of V. cholerae, V. parahaemolyticus, and V. vulnificus, respectively. This method was applied to screen Vibrio species from environmental samples and combining it with a culture-based method, its effectiveness was evaluated in comparison with culture-based methods alone.
Results: Specific PCR fragments were obtained from isolates belonging to the target species, indicating a high specificity of this multiplex real-time PCR. No cross-reactivity with the assay was observed between the tested bacteria. The sensitivity of the multiplex real-time PCR was found to have a lower limit of 104 colony-forming units/reaction for all three Vibrio species. The combination strategy raised the isolation ratio of all three Vibrio species 1.26- to 2.75-fold.
Conclusion: This assay provides a rapid, sensitive, and specific technique to detect these three Vibrio species in the environment.
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KEYWORD
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TaqMan probe multiplex real-time PCR, Vibrio cholerae, Vibrio parahaemolyticus, Vibrio vulnificus
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