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Multiplex polymerase chain reaction-based assay for the specific detection of toxin-producing Vibrio cholerae in fish and fishery products

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Abstract

A multiplex polymerase chain reaction (MPCR)-based assay was developed for the simultaneous detection of Vibrios using the genus-specific RNA polymerase subunit A (rpoA) gene and specific detection of toxin-producing Vibrio cholerae strains using two sets of primer based on cholera toxin subunit A (ctxA) and repeat in toxin subunit A (RtxA)-producing genes. The MPCR method developed is applicable to both the simultaneous and the two-step detection of genus Vibrio total and toxigenic V. cholerae species. This assay was specific as no amplification occurred with the other bacterial pathogens tested. The sensitivity of the assay was tested by artificially spiking the shrimp homogenate with the toxigenic strain of V. cholerae (NICED 16582) in different dilutions. The developed MPCR assay could detect three cells of V. cholerae in 12 h pre-enrichment in APW. The proposed method is rapid, sensitive, and specific for the detection of Vibrio genus as well as toxin-producing V. cholerae strains in environmental samples.

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Acknowledgements

The financial assistance extended by the Department of Biotechnology, Government of India, New Delhi, India for conducting this project work is gratefully acknowledged. The authors wish to thank the dean of the Fisheries College and Research Institute, Tamil Nadu Veterinary and Animal Sciences University, Tuticorin, India for having provided the necessary support for carrying out this work.

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Authors and Affiliations

  1. Department of Fish Processing Technology, Fisheries College and Research Institute, Tamil Nadu Veterinary and Animal Sciences University, Tuticorin, 628 008, India

    Geevaretnam Jeyasekaran, Kannan Thirumalai Raj, Robinson Jeya Shakila, Albin Jemila Thangarani & Durairaj Sukumar

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  1. Geevaretnam Jeyasekaran
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  2. Kannan Thirumalai Raj
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  3. Robinson Jeya Shakila
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Correspondence to Geevaretnam Jeyasekaran.

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Jeyasekaran, G., Thirumalai Raj, K., Jeya Shakila, R. et al. Multiplex polymerase chain reaction-based assay for the specific detection of toxin-producing Vibrio cholerae in fish and fishery products. Appl Microbiol Biotechnol 90, 1111–1118 (2011). https://doi.org/10.1007/s00253-011-3175-9

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  • DOI: https://doi.org/10.1007/s00253-011-3175-9

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