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Quantitative PCR assay for the simultaneous identification and enumeration of multiple Karenia species

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Abstract

Quantitative PCR (qPCR) is the method of choice for specific detection and quantification of harmful algal bloom (HAB) species. Development of qPCR assay for simultaneous enumeration of species that frequently co-exist in HABs is required. A high sensitivity TaqMan qPCR assay, using probe and primers, located at ITS1–5.8S–ITS2 rDNA region, detecting, specifically, Karenia selliformis, K. bidigitata, and K. mikimotoi, was designed. ITS1–5.8S–ITS2 rDNA region copy numbers per Karenia cell genome were estimated to 217.697 ± 67.904, allowing cell quantification. An application of the designed methodology in field samples has been conducted, and it showed high sensitivity (detection of around 10−1 cell/100 mg of bivalve mollusk tissue, equivalent to about 20 copies of the target sequence). We suggest that the optimized method could contribute to early detection of three closely related Karenia species in seafood cultivating areas to promote control quality, guarantee a fast and effective intervention, and improve public health prevention.

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Acknowledgments

This study was performed in the framework of the PRF-2017-D1P2 Project (2018–2020) with the financial support of Tunisian Ministry of Higher Eduction and Scientific Research.

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Authors and Affiliations

  1. Laboratoire de Génie Enzymatique et Microbiologie, Equipe Biotechnologie des Algues, Ecole Nationale d’Ingénieurs de Sfax, Université de Sfax, Sfax, Tunisia

    Jihen Elleuch, Mohamed Barkallah & Slim Abdelkafi

  2. Cawthron Institute, 98 Halifax Street East, Private Bag 2, Nelson, 7042, New Zealand

    Kirsty F. Smith

  3. Veterinary Research Center of Sfax, Sfax, Tunisia

    Idriss Ben Neila

  4. Laboratory of Plant Biotechnology Applied to the Improvement of Cultures, Faculty of Sciences of Sfax, University of Sfax, Sfax, Tunisia

    Imen Fendri

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  1. Jihen Elleuch
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Correspondence to Jihen Elleuch.

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Elleuch, J., Barkallah, M., Smith, K.F. et al. Quantitative PCR assay for the simultaneous identification and enumeration of multiple Karenia species. Environ Sci Pollut Res 27, 36889–36899 (2020). https://doi.org/10.1007/s11356-020-09739-4

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  • DOI: https://doi.org/10.1007/s11356-020-09739-4

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