Quantitative real-time PCR may be a rapid and automated procedure for detection of bacterial pathogens from food samples. Nevertheless, when testing the effects of antimicrobials on the viability of bacterial pathogens in foods, we found that DNA from dead cells interfered greatly in the detection of viable Listeria monocytogenes after treatment with the broad-spectrum bacteriocin enterocin AS-48. To overcome this problem, a quantitative real-time PCR (qRT-PCR) assay based on bacterial mRNA was adapted to quantify viable L. monocytogenes in food after bacteriocin treatments. The procedure allowed a better and faster estimation of viable cells compared to PALCAM viable cell counts when the threshold level was 2 log units/g of food, while PALCAM viable count allowed detection of one log unit/g. This procedure may be useful to verify the efficacy of bacteriocins against L. monocytogenes in foods.