Persistent microbial contamination of medical implant surfaces is becoming a serious threat to public health. This is principally due to antibiotic-resistant bacterial strains and the formation of bacterial biofilms. The development of novel antibacterial materials that will effectively fight both Gram-positive and Gram-negative bacteria and prevent biofilm formation represents a big challenge for researchers in the last few decades. In the present work, we report an antibacterial hydrophobic carbon quantum dots/polyurethane nanocomposite (hCQD-PU), with enhanced antibacterial properties induced by pre-treatment with gamma-irradiation. Hydrophobic quantum dots (hCQDs), which are capable of generating reactive oxygen species (ROS) upon irradiation with low-power blue light (470 nm), have been integrated into the polyurethane (PU) polymer matrix to form a photoactive nanocomposite. To modify its physical and chemical properties and improve its antibacterial efficacy, various doses of gamma irradiation (1, 10, and 200 kGy) in the air environment were applied to the formed nanocomposite. Gamma-irradiation pre-treatment significantly influenced the rise in ROS production, therefore, the prooxidative activity under the blue-light illumination of hCQD-PU was also significantly improved. The best antibacterial activity was demonstrated by the hCQD-PU nanocomposite irradiated with a dose of 200 kGy, with the complete eradication of Gram-positive Staphylococcus aureus (S. aureus) and Gram-negative Escherichia coli (E. coli) bacteria after 15 min of exposure to the blue lamp. © 2021 Elsevier Ltd