Listeria monocytogenes causes listeriosis, a severe foodborne disease with high mortality. Contamination with it poses significant risks to food safety and public health. Notably, genetic characteristic differences exist between strains causing human infections and those found in routine food inspections. This study examined the genotypic factors influencing the pathogenicity of L. monocytogenes, focusing on virulence gene profiles and key integrity genes like inlA to explain these divergences. The dataset included 958 strains isolated from human, food, and environmental samples. Whole-genome sequencing identified virulence genes, and principal component analysis (PCA) examined 92 virulence genes and inlA integrity to uncover potentially pathogenic patterns. The results highlight differences in virulence characteristics between strains of different origins. The integrity of inlA and genes such as inlD, inlG, and inlL were pivotal to pathogenicity. Strains with premature stop codons (PMSCs) in inlA, associated with reduced virulence, accounted for a low percentage of human cases but over 30% of food isolates. Sequence types (STs) like ST121, ST580, and ST199 showed unique profiles, while ST9, dominant in food, occasionally caused human cases, posing risks to vulnerable individuals. This research highlights the complexity of the pathogenicity of L. monocytogenes and emphasizes the importance of genomic surveillance for effective risk assessment.