Detection of virulence-associated genes and epidemic clone markers in Listeria monocytogenes isolates from PDO Gorgonzola cheese

Fifty-three Listeria monocytogenes isolates obtained from Gorgonzola cheese and previously characterized with biochemical typing, serotyping and pulsed field gel electrophoresis (PFGE), were analyzed in this study. Seven virulence-associated genes were selected (actA, inlC, inlJ, plcA, prfA, hlyA and iap) and their presence was investigated using PCR. All virulence-associated genes were detected in 51 isolates. One isolate did not show amplification of the inlC gene and one other isolate, previously mis-identified as L. monocytogenes probably due to atypical phenotypes, resulted negative by PCR for all virulence genes and was identified as Listeria innocua by 16S rRNA gene sequencing analysis. A multiplex PCR assay was used to evaluate the presence of markers specific for epidemic clones (ECs) of L. monocytogenes. The marker specific for the recently identified epidemic clone V (ECV) was detected in 38 of 43 (88%) of serotype 1/2a isolates. These findings suggest that Gorgonzola cheese can represent a significant source of L. monocytogenes and potential health risk for listeriosis as almost all isolates (94%) could be potentially virulent and that 38 (~72%) were presumptive positive ECV. PCR screening for both virulence-associated genes and EC markers may be useful for rapidly evaluating the epidemic potential and public health risk posed by L. monocytogenes in PDO Gorgonzola cheese and other dairy products.