Defect WO3-carbon hybrid nanosheets (D-WO3@C), synthesized by a facile hydrothermal method, is utilized as an effective visible-light driven photocatalyst for sustainable inactivation of bacteria. Detailed characterization through SEM-EDS, XRD, FT-IR, HRTEM, and XPS confirmed the formation of the nanohybrids. As a result, the as-synthesized D-WO3@C exhibited the best sterilization efficiency among all of the WO3 photocatalysts, and 7 log inactivation of E. coli or S. aureus was obtained after treating samples for 45 min with 1000 mu g/mL D-WO3@C. It was found that D-WO3@C exhibited enhanced visible-light-driven photocatalytic sterilization in killing bacteria compared with D-WO3 and P-WO3. The results show that the oxygen vacancies and carbon matrix were able to promote photocatalytic performance by triggering a faster interfacial transfer and a higher efficient separation of electron-hole pairs, which most likely is the reason behind the improved bactericidal effect. Furthermore, fluorescent-based cell viability/death tests and SEM technology were employed to demonstrate the lethal effect as well as the integrity of bacterial membranes during photocatalytic process. Photoelectrochemical techniques and UV-visible diffuse reflectance revealed the efficient separation of electron-hole pairs between D-WO3 and carbon matrix. More importantly, after being tested in four cycles, D-WO3@C showed good photocatalytic performance due to its chemical stability. The excellent properties associated with the novel D-WO3/C indicated that they could be considered as a promising light-visible driven photocatalyst to remove, in practice, microbial contamination of water.