Real-time PCR (qPCR) assays cannot distinguish between DNA from dead and live cells

Highlights ► Real-time PCR (qPCR) assays cannot distinguish between DNA from dead and live cells ► DNA binding molecules allow specific detection of viable cells ► PMA-qPCR was able to specifically detect viable cells in pure cultures and food samples ► Power ultrasound has been successfully applied for microbial decontamination ► PMA-qPCR results correlate precisely with plate counts of ultrasound treated water samples. Application of propidium monoazide-qPCR to evaluate the ultrasonic inactivation of E. coli O157:H7 in fresh-cut vegetable wash water P. Elizaquívela, G. Sánchezb, M.V. Selmac, R. Aznara, b, ,  doi:10.1016/j.fm.2011.10.008 | How to Cite or Link Using DOI   Permissions & Reprints Purchase a Department of Microbiology and Ecology, University of Valencia, Burjassot, Valencia, Spain b Department of Biotechnology, Institute of Agrochemistry and Food Technology (IATA-CSIC). Burjassot, Valencia, Spain c Research Group on Quality, Safety and Bioactivity of Plant Foods, Department of Food Science and Technology, CEBAS-CSIC, Murcia, Spain Received 18 April 2011; revised 23 September 2011; Accepted 5 October 2011. Available online 13 October 2011. Abstract The efficacy of sanitizing technologies in produce or in vegetable wash water is generally evaluated by plate count in selective media. This procedure is time consuming and can lead to misinterpretations because environmental conditions and sanitizing processes may affect bacterial growth or culturable capability. Thus, the aim of this study was to determine the applicability of a propidium monoazide real-time PCR (PMA-qPCR) method to monitor the inactivation by ultrasound treatment of foodborne bacteria in fresh-cut vegetable wash water. To this aim, lettuce wash water was artificially inoculated with E. coli O157:H7 (106 CFU/mL) and treated by means of a continuous ultrasonic irradiation with a power density of 0.280 kW/L. Quantification data obtained by PMA-qPCR and plate counts were statistically similar during the viability reduction of 99.996% which corresponds to 4.4 log reductions. Further reductions of E. coli O157:H7 were not detected by the PMA-qPCR method due to the limit of detection of this technique (20 CFU/mL). Inactivation data obtained by both techniques successfully fitted a linear model, giving no significant differences in kinetic parameters. These results indicate that the PMA-qPCR method is a suitable technique for evaluating ultrasonic disinfection of vegetable wash water, being able to distinguish between live and dead bacteria.