Research on the Genetic Polymorphism and Function of inlA with Premature Stop Codons in Listeria monocytogenes

<i>Listeria monocytogenes</i> is a Gram-positive bacterial species that causes listeriosis, a major foodborne disease worldwide. The virulence factor <i>inlA</i> facilitates the invasion of <i>L. monocytogenes</i> into intestinal epithelial cells expressing E-cadherin receptors. Naturally occurring premature stop codon (PMSC) mutations in <i>inlA</i> have been shown to result in the production of truncated proteins associated with attenuated virulence. Moreover, different <i>L. monocytogenes</i> strains contain distinct <i>inlA</i> variants. In this study, we first characterized <i>inlA</i> in 546 <i>L. monocytogenes</i> strains isolated from various foods in Shanghai. The results showed that 36.1% (95% Confidence Interval: 32.0~40.2%) of the food isolates harbored <i>inlA</i> with PMSC, which was found to be associated with clonal complex (CC) types, with the highest proportions observed in CC9 and CC121. To investigate the function of <i>inlA</i>, we first used the dominant CC87 isolated from patients as the test strain and constructed an <i>inlA-deleted</i> strain via homologous recombination. Resistance tests and virulence tests showed that while <i>inlA</i> did not affect the resistance of <i>L. monocytogenes</i>, it significantly influenced cell adhesion and invasiveness. To further explore the function of <i>inlA</i>, we performed virulence tests on five CC-type strains carrying <i>inlA</i> with PMSC and their corresponding strains with intact <i>inlA</i>. We found that the virulence of <i>L. monocytogenes</i> strains carrying <i>inlA</i> or <i>inlA</i> with PMSC was associated with their CC type. Our preliminary results showed that premature termination of <i>inlA</i> did not significantly affect the adhesion and invasion abilities of low-virulence CC-type <i>L. monocytogenes</i> strains in Caco-2 cells, but substantially promoted those of high-virulence strains such as CC8 and CC7. In summary, this study preliminarily evaluated the effects of <i>inlA</i> integrity and PMSC mutation variation on the virulence of <i>L. monocytogenes</i>, providing a foundation for further research on <i>inlA</i>-related pathogenic mechanisms.