AbstractAeromonas veronii (A. veronii) is an opportunistic pathogen of fish–human–livestock, which poses a threat to the development of aquaculture. Based on our previous studies on proteomics and genomics, we found out that the aodp gene may be related to the virulence of A. veronii TH0426. However, aodp gene encodes a hypothetical protein with an unknown function, and its role in A. veronii TH0426 is not clear. Here, we first constructed a mutant strain (△‐aodp) to investigate the functional role of aodp in A. veronii TH0426. Compared with the wild strain A. veronii TH0426, the growth rate of strain △‐aodp was slower and was resistant to neomycin and kanamycin, but sensitive to cephalexin. The swimming and swarming ability of △‐aodp strain decreased, and the pathogenicity to mice decreased by 15.84‐fold. Besides, the activity of caspase‐3 in EPCs infected with △‐aodp strain was 1.49‐fold lower than that of the wild strain. We examined 20 factors closely related to A. veronii virulence, among them 17 genes were down‐regulated as a result of aodp deficiency. This study laid a foundation for further studies on the pathogenesis of A. veronii.