BackgroundInfection prevention and control (IPC) is important to prevent the spread of multidrug resistant organisms (MDROs). We aimed to enhance and explore the implementation of preventing and controlling MDROs using audit and feedback and the benchmark. MethodsThis quasi-experimental design was conducted in three hospitals from 1st March 2018 to 30th September 2019. A multimodal intervention treated audit and feedback and benchmark as key components for MDROs IPC was conducted in Wuhan, China. A checklist of 40 implementation indicators based on IPC measures was formed to guide the audit twice a week. Immediate feedback was verbally given after each audit on the spot, and written feedbacks containing benchmark and individual implementation levels were delivered monthly or quarterly in three hospitals. The intervention effect was evaluated by Chi-square and Poisson segmented linear regression. Impacts of implementation of indicators on the incidence were modeled by mixed-effect regressions. ResultsThe incidence of nosocomial MDROs decreased by 19.39%, 20.55%, and 24.03% in A, B, and C hospital, respectively. The lowest implementation compliance of indicators was the use of personal protective equipment of doctors (50.24%). The highest was isolated warning signs of nurses (96.46%). The implementation on hand hygiene of doctors (Coef. = -27.87, p=0.001) and nurses (Coef. = -35.44, p=0.001), clean of surrounding instruments and bed unit (Coef. = -4.84, p=0.030), education to patients and relatives (Coef. = -59.51, p=0.031), and sending of specimen inspection timely (Coef. = -9.95, p<0.001) were negatively associated with the incidence of nosocomial MDROs infection. Conclusions The multimodal intervention by strengthening the implementation of audit and feedback and the benchmark is feasible and effective in China. The checklist is an effective and practical tool to measure the level of implementation. Education to patients and relatives, hand hygiene, clean of surrounding instruments and bed unit, and sending of specimen inspection timely are especially crucial.