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A filtration-based real-time PCR method for the quantitative detection of viable Salmonella enterica and Listeria monocytogenes in food samples

D'Urso, Oscar Fernando, Poltronieri, Palmiro, Marsigliante, Santo, Storelli, Carlo, Hernández, Marta, Rodríguez-Lázaro, David
Food microbiology 2009 v.26 no.3 pp. 311-316
foods, isolation, bacterial contamination, new methods, filtration, polymerase chain reaction, quantitative analysis, microbial detection, molecular epidemiology, Salmonella, Salmonella enterica, Listeria monocytogenes, viability, food pathogens, predictive microbiology, linear models, discriminant analysis
We developed a novel filtration-based method that can eliminate dead or severally damaged Salmonella enterica and Listeria monocytogenes in food samples. This new method can recover all viable bacteria in less than 30 min, and can be coupled with a subsequent bacterial DNA extraction and real-time PCR. No statically significant differences (p < 0.01) were found between real-time PCR results obtained separately from S. enterica and L. monocytogenes when different ratios of living and dead cells were used. The analytical sensitivity in both cases was 1 genome equivalent (GE), and the quantification was linear (R2 > 0.9969) over a 5-log dynamic range with PCR efficiencies >0.9754. When compared with the standard microbiological methods for the detection of these foodborne pathogens, the relative accuracy was excellent ranging from 95.72% to 104.48%. Finally, we applied the pre-treatment method to the direct detection of viable forms of these foodborne pathogens in food samples using yogurt as a model, the results being similar to those obtained using pure cultures.