Genomic Features and Antimicrobial Susceptibility of Listeria innocua Isolated from Raw Drinking Milk in Poland

<i>Listeria innocua</i> is a bacterium frequently detected in food and food production plants (FPPs). Understanding the heterogeneity of <i>L. innocua</i> food isolates is essential for predicting potential food safety threats and developing preventive and control measures. This study aimed to characterize <i>L. innocua</i> isolated from raw drinking milk by investigating the genomic features related to virulence, antimicrobial resistance, and persistence using whole-genome sequencing (WGS), along with phenotypic antimicrobial susceptibility testing using the disk diffusion method. All ten isolates analyzed in this study belonged to sequence type (ST) 492 and were distantly related to the reference strain. A total of 80 virulence-associated genes were identified, including the complete <i>Listeria</i> Pathogenicity Islands-3 (LIPI-3) and LIPI-4 clusters typically found in virulent <i>L. monocytogenes</i> clones, as well as 66 additional genes involved in adhesion, invasion, motility, post-translational modification, regulation, immune modulation, and stress survival. Stress survival islet 2 (SSI-2) and genes encoding the <i>Clp</i> protease complex (<i>clpC</i>, <i>clpE</i>, <i>clpP</i>), which support both persistence and virulence, were also detected, whereas LIPI-1 and internalin genes were not detected. The antimicrobial resistance determinants included <i>fosX</i>, <i>lin</i>, <i>norB</i>, <i>sul</i>, and three multidrug efflux pumps (<i>lde</i>, <i>mdrL</i> and <i>mdrM</i>). Mobile genetic elements (plasmids, prophages, or transposons) were not detected. All isolates were phenotypically susceptible to benzylpenicillin, ampicillin, meropenem, erythromycin, and trimethoprim-sulfamethoxazole. These findings underscore the importance of ongoing genomic surveillance of <i>L. innocua</i> in food environments and highlight the need to assess the potential risk posed by specific lineages, such as ST492, to food safety.