Taqman® probe based multiplex RT-PCR for simultaneous detection of Listeria monocytogenes, Salmonella spp. and Shiga toxin-producing Escherichia coli in foods

Three foodborne pathogenic bacteria, Listeria monocytogenes, Salmonella spp. and Shiga toxin-producing Escherichia coli are major contaminants in various foods and often cause food safety problems in public health. Therefore, a simple Taqman® probe based multiplex real-time polymerase chain reaction (mRT-PCR) for fast and reliable presence test to detect those three bacterial targets in different types of foods were developed. In order to improve the effectiveness of the developed method, a culturing step in Simultaneous Enrichment Broth at 37 °C for 18 h, followed by extraction of DNA using a boiling method was included. The developed Taqman® mRT-PCR platform gave all 100% of relative sensitivity, relative specificity and relative accuracy as compared to the conventional method (Bacteriological Analytical Manual) and iQ-check® RT-PCR detection kits. The results showed the developed method could detect as low as 1 CFU of each bacterial target in 25 g of artificially contaminated food samples. For practical food testing, the detection results of the bacterial targets naturally contaminated in 92 food samples by both the developed Taqman® probe mRT-PCR and the conventional method were essentially identical. The developed method is greatly specific, sensitive and effective to detect the three bacteria in foods simultaneously.