Detection of Listeria monocytogenes in food, equivalent to EN ISO 11290-1 or ISO 10560, by a three-days polymerase chain reaction-based method

Abstract

A method is presented for the detection of Listeria monocytogenes in food, which produces definitive results on the third day after the sample collection. The method is equivalent to EN ISO 11290-1 or ISO 10560 in terms of the same detection limit of 10 0 cfu per 25 or 10 g and 100% relative accuracy. The version alternative to EN ISO 11290-1 begins with a primary enrichment in half Fraser broth (24 h), secondary enrichment in Fraser broth (24 h) and post-enrichment in brain heart infusion broth (5 h). The version alternative to ISO 10560 begins with enrichment in Listeria enrichment broth (48 h) and post-enrichment in tryptone soya broth (5 h). These steps are followed by bacterial cell lysis by boiling, PCR oriented to inlB gene using a mimic internal control, and agarose gel electrophoresis. At the evaluation of the method on model food samples artificially contaminated with decimal dilutions of a L. monocytogenes culture (cheese, smoked fish, ready-to-eat meat products; 21 samples altogether), a detection limit of 10 0 cfu per 25 or 10 g was determined. Dead L. monocytogenes cells did not cause false positivity, as determined using model food samples artificially contaminated with decimal dilutions of dead L. monocytogenes cells. At the evaluation of the method on naturally contaminated food samples (same as above, 140 samples altogether) identical results (8 positives) as with the reference method were obtained.