Purpose: An independent study was conducted, to validate this new method in comparison to the ISO 11290-1 reference method, as part of the NF Validation approval process and according to the ISO 16140 standard. The rapid and automated protocols were both tested.
Methods: The rapid protocol includes a single enrichment step in Half-Fraser broth, while the automated protocol requires an additional sub-culture in Fraser 1. After DNA extraction, the new real time PCR is run with the 7500 Fast Automate.
Results: 398 food and environmental samples were analyzed for relative accuracy, sensitivity and specificity. The results demonstrate equivalent performances between the new real-tme PCR and the ISO 11290-1 methods. The relative detection limits of the new method vary from 0.3 to 1.6 CFU/25 g, those of the ISO standard vary from 0.2 to 1.2 CFU/25 g. The selectivity and specificity of the alternative method was assessed by testing 55 target strains and 32 non-target strains. The alternative method was also evaluated in a ring trial involving 17 laboratories. The results of the calculated accuracy, accordance, concordance, and odds ratio clearly show that the alternative precision is equivalent to the ISO 11290-1 standard one.
Significance: The proposed method is a reliable alternative method for Listeria spp detection in food and environmental samples, and offers important economic savings by reducing time to result and handling time.