Rapid Detection of Hypervirulent Serovar 4h Listeria monocytogenes by Multiplex PCR.

Listeria monocytogenes(L. monocytogenes) is a ubiquitous foodborne pathogen that comprises 14 serotypes, of which serovar 4h is a novel serotype recently reported. Serovar 4hL. monocytogenesbelonging to hybrid sub-lineage II exhibit hypervirulent features. Conventional biochemical tests and widely used PCR-based serogrouping schemes could not distinguish serovar 4h strains. In this study, we developed a new multiplex PCR assay for rapid detection of serotype 4hL. monocytogenes. Three primer pairs based on the target genes, LMxysn_1095,lmo1083, andsmcL, were designed. The multiplex PCR results showed that serovar 4h strains could be specifically identified from all tested strains, including variousL. monocytogenesserovars,Listeriaspp., and other species. The detection limits of the multiplex PCR were 291 fg/mu L for genomic DNA and 5.5 x 10(6)CFU/mL for bacterial suspension. Furthermore, pork meat artificially contaminated with serovar 4hL. monocytogenesin a concentration of 1.8 x 10(3)-1.8 x 10(0)CFU/10 g were successfully detected within 10-16 h. These results demonstrate that the multiplex PCR with high specificity and sensitivity is applicable for the rapid detection ofL. monocytogenesserotype 4h strains.