AN OPTIMIZED EMA-RAPD-PCR FOR A RELIABLE DETECTION OF VIABLE SALMONELLA SPP. IN CHICKEN PRODUCTS

This study aimed to develop the reliable technique called ethidium bromide monoazide-random amplified polymorphic DNA-PCR (EMA-RAPD-PCR) for detection of only viable Salmonella cells due to PCR cannot distinguish DNA from viable and dead cells. In EMA-RAPD-PCR, EMA was used to intercalate the DNA obtained from 1.2x10(6) cells of viable and heat-killed Salmonella Typhimurium and Salmonella Enteritidis. The optimized conditions of EMA treatment for an effective prevention of DNA amplification from dead cells by RAPD-PCR were as follows: the minimum amount of 3g/mL EMA; the suitable light exposure time of at least 5min; and the optimum light exposure distance of 20cm. To improve a reliability in specific detection of viable Salmonella cells in food samples, use of an optimized 20-h preenrichment in nutrient broth followed by EMA-RAPD-PCR could inhibit the DNA amplification of the dead cells in all artificially and naturally Salmonella-contaminated chicken products tested. Practical ApplicationsThe developed Ethidium Bromide Monoazide-Random Amplified Polymorphic DNA-Polymerase Chain Reaction (EMA-RAPD-PCR) involving 20-h culturing in preenrichment medium is an efficient, reliable and economical procedure to detect only viable Salmonella spp. in food samples. This technique has demonstrated effectiveness in preventing the DNA amplification of dead Salmonella cells. Moreover, the EMA-RAPD-PCR has the potential for use as a rapid, simple and reliable monitoring tool of Salmonella spp. prevalence along the food production chain.