Genetic and Phenotypic Characterization of Listeria monocytogenes Strains with Attenuated Virulence

Epidemiological studies have provided considerable evidence that Listeria monocytogenes strains differ in their ability and relative likelihood to cause human disease. Multiple mutations leading to premature stop codons (PMSCs) have been identified in the key L. monocytogenes virulence gene inlA, which codes for a surface protein that mediates the entry of L. monocytogenes into host cells. Mutations leading to premature stop codons in inlA are associated with a reduced invasion phenotype in vitro, supporting the hypothesis that these mutations may also be responsible for attenuated human virulence. In order to further define the frequency of L. monocytogenes strains with reduced invasion capability, 207 isolates had previously been screened for ability to invade Caco-2 cells (Nightingale, unpublished). The inlA gene was sequenced for 27 isolates that showed attenuated ability to invade the epithelial cell line. Sequencing revealed mutations leading to PMSCs in 12 of the 27 isolates, including 2 novel mutations (PMSC types 5 and 7) and 3 previously reported mutations (PMSC types 1, 4 and 6). All but one of the isolates with confirmed PMSC mutations in inlA were isolated from food products or food processing plants. Our data support the hypothesis that L. monocytogenes isolates associated with food have attenuated invasion due to the presence of inlA PMSCs. Our data further suggest that sequence variation in inlA is an important factor that can be used to define L. monocytogenes that have a reduced ability to cause human disease. Since some strains with PMSCs in inlA were common in foods and environmental samples and were found in high levels in foods, it was hypothesized that these mutations may provide selective advantage, possibly outside a human host. Isogenic inlA PMSC strains were thus tested for their growth in rich medium and their resistance against common sanitizing agents. The results indicate that there is little difference in growth trends or sanitizer resistance between isogenic strains with and without inlA PMSC. It thus appears that inlA PMSCs do not enhance the ability of L. monocytogenes to grow in rich media or to be resistant to sanitizing agents