The SureFast® Listeria 3plex ONE Kit and the SureFast® PREP Bacteria Kit in a Variety of Foods and Environmental Samples: Multi-Laboratory Validation, First Action 2025.04

BACKGROUND: The SureFast® Listeria 3plex ONE Kit uses the real-time polymerase chain reaction (PCR) technique for multiplex detection of Listeria species and Listeria monocytogenes in foods and environmental samples. The SureFast® Listeria 3plex ONE Kit can also be used with the SureFast® PREP Bacteria Kit to provide a second DNA extraction option and can be analyzed on multiple thermocyclers. OBJECTIVE: The kit was evaluated using 25 g test portions in an unpaired study design for a single matrix, ready-to-eat smoked deli turkey as part of a harmonized validation protocol with MicroVal Certification. METHODS: The matrix was compared to the International Organization for Standardization Technical (ISO) 11290-1:21017 reference standard. 17 participants from 16 laboratories, located within Austria, Germany, Italy, Spain, and the United States were solicited for the multi-laboratory validation (MLV), with only 10 participants submitting data. Three levels of contamination were evaluated: an uninoculated control level (0 colony forming units (CFU)/test portion), a low inoculum level (0.2-2 CFU/test portion) and a high inoculum level (2-10 CFU/test portion). Statistical analysis was conducted according to the Probability of Detection (POD) statistical model. RESULTS: Results obtained for the low inoculum level test portions produced a dLPOD value with 95% confidence interval of -0.01 (-0.04, 0.02). The dLPOD results indicate equivalence between the candidate method and reference method for the matrix evaluated and the method demonstrated acceptable multi-laboratory reproducibility as determined in the MLV. False positive and false negative rates were determined for the matrix and produced values of 0.0% and 0.8% respectively. CONCLUSIONS: Based on the data generated, the method demonstrated equivalence to the ISO reference method in a MLV study. HIGHLIGHTS: The SureFast ONE 3plex method is a rapid and reliable alternative for the multiplex detection of Listeria species and Listeria monocytogenes in a broad range of foods and environmental surfaces.