SYBR Green I RT-PCR Assay for Quantitative Detection of Salmonella in Irrigation Water

  Objective  A SYBR Green I-based quantitative RT-PCR (qPCR) method was developed for detecting salmonella in irrigation water.

  Method   For the methodology development a pair of primers were designed based on the sequences of the invasion protein A gene (invA) of Salmonella. qPCR reaction conditions were optimized, the assay tested for specificity and sensitivity, and a standard curve of amplification constructed. Test results on a specimen of contaminated irrigation water using the qPCR assay were compared for detection accuracy with those obtained from the national standard method.

  Result  The newly developed qPCR assay showed a minimum detection limit of 1×10⁻¹pg·μL⁻¹ and free of interference from genomic nucleic acids of non-target microbes. The constructed linear standard curve between 2×10⁰cfu·mL⁻¹ and 2×10⁵ cfu·mL⁻¹ had a high correlation coefficient of 0.999 6. The assay demonstrated same accuracy as did the national standard method in detecting salmonella in irrigation water.

  Conclusion  The newly established qPCR assay could be adequately applied for salmonella detection in agriculture irrigation water.