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Vibrio vulnificus is a pathogenic bacterium that causes disease in marine fish, affecting fish farming and human health worldwide. In May 2021, in the Bohai Bay region, a disease broke out in commercially farmed pearl gentian grouper (♀Epinephelus fuscoguttatus × ♂Epinephelus lanceolatus), causing huge economic losses. The diseased fish had skin lesions, water accumulation in their abdomens, and showed tissue and organ damage. V. vulnificus biotype 2 has been reported in eels and other marine fish, but it is less reported in pearl gentian grouper. In this study, the pathogenic strain isolated from diseased fish was identified as V. vulnificus EPL 0201 biotype 2 on the basis of physiological and biochemical characteristics and the results of 16S rRNA gene and gyrB sequencing, virulence gene detection, and recursive infection experiments. To gain a comprehensive understanding of the pathogenicity and drug resistance of this strain, whole-genome sequencing was performed. Whole-genome analysis showed that the gene map of this strain was complete. The Virulence Factor Database annotation results showed that this strain had the key virulence factor genes vvhA and rtxA, which cause host disease. In addition, this strain had genes conferring resistance against cephalosporins, aminoglycosides, tetracyclines, and sulfonamides. Antimicrobial susceptibility testing confirmed the presence of these resistance genes identified in the genome. The results of this study show that V. vulnificus EPL 0201 biotype 2 is a multi-drug resistant strain with high pathogenicity. [ABSTRACT FROM AUTHOR]