Alleviating the concentration of hazardous bacteria by photocatalysts is an efficient way for treating ballast water. In this work, we present the synthesis of in-situ Bi vacancies on Bi/CuBi2O4, which is assisted by H2O2 for Fenton-like photocatalytic potential in deactivating marine microorganisms in natural seawater (Yellow Sea, China), the optimal Bi vacancy content (3BCBO) is determined and exhibits superior photocatalytic inactivation performance, additional H2O2 activation enhances the production of •OH for comprehensive oxidation, with •OH confirmed as the dominant reactive species. Density functional theory (DFT) calculations highlight the synergistic of vacancies and metallic Bi play a pivotal role in decreasing the energy barrier of H2O2 adsorption for •OH improving production, promotes efficient separation of charge carriers and utilization of visible light, the co-effect of metallic Bi nanoparticles and bismuth defection mechanisms effectively inhibit the madness thriving of marine microorganisms, this work offers an ideal and promising perspective for ballast water treatment on ships.