Abstract:[Objective] Vibrio vulnificus is a pathogen with the highest mortality in Vibrio genus. However, currently there is lacking knowledge of virulence-associated factors across the whole genome wide for this species. By using bacterial genome-wide association study (BGWAS), we investigated the genes that are related with different sources (clinical/environment) of V. vulnificus isolates, aiming to provide further understanding on genetic factors that potentially influence its pathogenicity. [Methods] In total 260 V. vulnificus genomes were analyzed, including 139 newly sequenced in this study and 121 published genomes. We identified the genomes variations, including SNPs and accessory genes from the dataset, and then by using software pyseer, we determined the associations between sources of isolates and SNPs, genes, and Kmers, separately. We annotated the function of the statistically significant genes and inferred their possible influence on virulence of the bacteria. [Results] We found that 11 genes to be significantly correlated with clinical isolates of V. vulnificus, among which 9 were newly identified in this study.[Conclusion] By using methods of population genomics and statistical genetics, we screened the potential virulence-associated factors of V. vulnificus across the whole genome. Our findings provided new targets on vaccine designing and drug development, and would help to get further insight on pathogenic mechanism of this pathogen.