Background: Decontaminating urinary catheters prior to disposal or potential reuse is essential for mitigating environmental hazards and reducing the spread of multidrug-resistant (MDR) pathogens. Objective: This study aimed to evaluate the efficacy of gamma irradiation in decontaminating urinary catheters by inactivating MDR uropathogenic bacteria and disrupting their associated biofilms. Methodology: Fifty strains bacteria were isolated from urine samples obtained from catheterized patients and four bacterial isolates were recorded with 100% resistance against all tested antibiotics. Among them, two isolates were selected and identified as strong biofilm producers, Proteus mirabilis and Staphylococcus haemolyticus. The effect of different doses of Gamma radiation on the viability of the drug resistant strains and biofilm formation were studied. The gamma irradiation technique was applied to decontaminate artificially infectious urinary catheters. Results: The study showed that a dose-dependent reduction in bacterial survival and biofilm formation was observed. The lethal gamma doses for P. mirabilis and S. haemolyticus were 0.5 kGy and 1.5 kGy, respectively. Conclusion: Gamma irradiation is an effective strategy for decontaminating urinary catheters and eliminating MDR uropathogenic bacteria, cause biofilm degradation and offering a potential eco-friendly solution for managing infectious medical waste.