Objective To identify the priority disease diagnosis-related groups (DRGs) of healthcare-associated infection (HAI) prevention and control among orthopedic patients aged≥15 years old, and provide an evaluation framework and evidence-based basis for precise prevention. Methods Based on DRG-related methods and indicators, this study evaluated various DRGs of the discharged patients from orthopedic department in a tertiary first-class hospital in 2024 from three dimensions (quality and safety, service capability, and service efficiency) through DRG stratification, case-mix index (CMI) standardization, and additional cost algorithms methods, high-priority populations for HAI prevention and control were then identified. Results A total of 36 084 patients were included in the study, with an overall HAI incidence of 0.46% (n=165). HAI concentrated in 22.76% (33/145) of DRGs. The main infection sites were surgical sites (n=58, 32.95%), respiratory system (n=51, 28.99%), and urinary system (n=25,14.20%), accounting for 76.14% (134/176) of the total HAI cases. From the quality and safety dimension, the top three DRGs with the highest standardized incidences of infection were identified as ZC11(1.48%), ZJ11(0.63%), and ZJ15(0.61%). From the service capability dimension, the vulnerable groups with lower technical difficulty but higher incidences of infection were identified (ZJ15[0.61%], IE21[0.53%], IE39[0.45%]). From the service efficiency dimension, the groups with the heaviest time and economic burden caused by HAI were both identified as IB19 (8 days and 24 200 Yuan). Conclusion It is recommended to incorporate key DRGs into targeted surveillance and conduct precise prevention and control. Based on DRG multidimensional evaluation, key populations for orthopedic infection prevention and control can be accurately identified, which provide scientific basis for resource allocation and optimization of prevention measures.