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 Spin and the MagMax protocols were both tested.
Methods: The Rapid Spin protocol includes a single enrichment step in Half-Fraser broth, while the MagMax protocol requires an additional sub-culture in Fraser 1. After DNA extraction, the MicroSeq® L. monocytogenes Real Time PCR is run with the 7500 Fast Automate.
Results: 407 food and environmental samples were analyzed for relative accuracy, sensitivity and specificity study. The results demonstrate equivalent performances between the real-time PCR method and the ISO 11290-1 methods. Depending on the tested (matrix/strain) pairs, the relative detection limits of the real-time PCR method vary from 0.3 et 1.6 CFU/25 g, those of the ISO standard vary from 0.2 to 1.1 CFU/25 g. The selectivity and specificity of the alternative method was assessed by testing 50 target strains and 30 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 real-time PCR method precision is equivalent to the ISO 11290-1 standard one.
Significance: The real-time PCR method is a reliable alternative method for L. monocytogenes detection in food and environmental samples, and offers important economic savings by reducing time to result and handling time.