Résumé
Introduction: Understanding the genomic diversity and population structure of pathogenic bacteria is crucial for elucidating their evolutionary trajectories, ecological niches and virulent potential. Vibrio aestuarianus cardii, a recently described subspecies, has caused recurring mortality events in cockle (Cerastoderma edule) populations along the French Channel coast since 2012. However, its ecological dynamics, genetic diversity, and virulence mechanisms remain poorly characterized. Methodology: We conducted a two-year ecological and genomic study at two sites in France (Bay of Somme and Bay of Authie) monitoring natural cockle beds to assess environmental conditions favoring disease emergence. 383 strains of V. aestuarianus cardii were isolated from cockle spat, adult cockles, mussels, and sediments. 40 of these strains were subjected to whole-genome sequencing and comparative genomic analyses to investigate population structure, recombination patterns, and potential virulence factors. Virulence assays were performed to evaluate strain pathogenicity in vivo, and three distinct genogroups were described based on genetic clustering and virulence profiles. We selected three representative strains per genogroup to conduct in vitro assays aimed at elucidating mechanisms underlying their differential virulence. These assays assessed bacterial multiplication in the presence of cockle hemocytes, bacterial cytotoxicity towards hemocytes, and bacterial gene expression (focusing on toxin-encoding genes) in the presence and absence of hemocytes. Results: Environmental monitoring identified specific thermal windows permissive to V. aestuarianus cardii proliferation in cockle populations, with cockle spat emerging as a probable reservoir. Comparative genomics revealed three genogroups associated with cockles, two exhibiting high virulence and one exhibiting low virulence in vivo. Frequent genetic exchanges within genogroups contributed to differentiation and acquisition of virulence factors, notably Type I and Type VI Secretion Systems. In vitro assays demonstrated that high-virulence strains exhibited enhanced multiplication and cytotoxicity in the presence of cockle hemocytes compared to lowvirulence strains. Gene expression analyses revealed differential regulation of toxin-related genes, which showed distinct genomic profiles, during interaction with hemocytes. Conclusions: Our study advances understanding of the ecological niche, population structure, and virulence mechanisms of V. aestuarianus cardii. The identification of three genogroups with distinct virulence profiles, coupled with mechanistic insights from in vitro assays, highlights the complex interplay between bacterial genetic makeup and host interactions. Our findings underscore the role of temperature and cockle spat reservoirs in disease dynamics and suggest that horizontal gene transfer facilitates virulence evolution. This work provides a foundation for improved risk assessment and management strategies for mitigating V. aestuarianus cardii-associated disease outbreaks in cockle fisheries.