Objective To investigate the occurrence of healthcare-associated infection(HAI) caused by multidrug-resistant organisms (MDROs) and economic burden due to MDRO HAI in an intensive care unit (ICU). Methods MDRO HAI occurred in patients in the general ICU of a hospital from 2016 to 2017 was investigated retrospectively, infection rate, infection site, and drug-resistant bacteria were analyzed, difference in length of hospital stay and hospitalization expenses between patients with MDRO HAI and patients without infection were compared by matched case-control study. Results MDRO HAI rate in patients in general ICU in 2016-2017 was 7.5%, MDRO HAI rate in 2017 was lower than that in 2016 (4.5% vs 10.7%). The main MDRO HAI was lower respiratory tract infection (non-ventilator-associated pneumonia), ventilator-associated pneumonia,and surgical site infection, accounting for 50.0%, 16.9%, and 10.8% respectively. The main MDROs were carbapenem-resistant Acinetobacter baumannii (CRAB), methicillin-resistant Staphylococcus aureus (MRSA) and carbapenem-resistant Pseudomonas aeruginosa (CRPA), accounting for 42.9%, 24.8%, and 24.0% respectively. The length of hospital stay of patients with MDRO infection were 26.0 days longer than patients without infection, and total hospitalization expenses increased by 116 147.0 yuan (all P<0.001). Patients with CRPA infection had the longest length of hospital stay and the highest hospitalization expenses. Conclusion MDRO HAI in general ICU patients can increase length of hospital stay and hospitalization expenses, and bring huge economic burden to patients, effective measures for prevention and control of MDRO infection should be taken to standardize the rational use of antimicrobial agents.