To explore the bactericidal effect and mechanism of curcumin-mediated photodynamic inactivation (PDI) against Shewanella putrefaciens, this study explored the influence of curcumin concentration, light time and incubation time on the bactericidal effect. Meanwhile, the underlying mechanism and major action sites were elucidated from the aspects of intracellular oxidative stress, bacterial structure, endogenous enzyme activity and macromolecules damage. The results showed that curcumin concentration, light time and incubation time all significantly affected the bactericidal effect of PDI. The total bacterial count decreased by 4.30 (lg(CFU/mL)) with a bactericidal rate of 99.99% after 1 min of incubation with 7.5 μmol/L and 5 min of light exposure. It was found that the bacterium gradually transformed from sublethal to dead cells, the cell membrane permeability increased, and the size of bacterial cells and its internal complexity first increased and then decreased during PDI. PDI caused an increase in the level of reactive oxygen species (ROS) inside the bacteria cells, triggered lipid peroxidation of the cell membrane, which disrupted cell wall/membrane integrity and led to leakage of intracellular contents. The accumulation of ROS during PDI led to the collapse of the oxidative defense system while causing disruption to the energy metabolism system, oxidative DNA breakage and protein degradation, and the accumulation of damage ultimately led to bacterial multi-target death. In summary, curcumin-mediated PDI is potentially applicable to aquatic product preservation and the control of foodborne contamination of S. putrefaciens.