Combined Loop-Mediated Isothermal Amplification Assays for Rapid Detection and One-Step Differentiation of and in Meat Products.

A loop-mediated isothermal amplification (LAMP) assay system was established, allowing gene-based simultaneous detection of and in enriched meat products. Additionally, one-step differentiation of target species on agar plates was enabled by gene- and gene-based duplex LAMP. Both the and - LAMP assays amplified the target sequences in all 62 and 27 strains used for determining inclusivity and revealed 100% exclusivity toward 85 tested non-target species. Throughout the entire experiments, and strains were 100% distinguishable by melting curves of and LAMP products. After 24-h enrichment, the LAMP assay reliably detected initial inoculation levels of 10-100 CFU/g in artificially contaminated minced meat. Investigation of naturally contaminated meat samples revealed a diagnostic accuracy of 95% toward real-time PCR and 94.1% toward the standard culture method applying the 24-h incubation period. Diagnostic sensitivity and specificity, and positive and negative predictive values were 89.8, 100, 100, and 91.2%, respectively, when measured against real-time PCR, and 89.6, 98.1, 97.7, and 91.2%, respectively, when measured against the standard culture method. After 48-h enrichment, the detection limit of the LAMP assay improved to initial inoculation levels of 1-10 CFU/g in artificially contaminated minced meat. Applying the 48-h incubation period on naturally contaminated meat samples resulted in 100% concordant results between LAMP, real-time PCR, and the standard culture method. The established LAMP assay system was proved to be suitable for rapid meat sample screening. Furthermore, it constitutes a promising tool for investigating other sources and could therefore make a valuable contribution to protect consumers from foodborne illness.