Objective To understand the clinical characteristics, antimicrobial resistance and influencing factors of healthcare-associated infection (HAI) as well as carbapenem-resistant Gram-negative bacterial (CR-GNB) infection after cardiac valve surgery. Methods Clinical data of patients after cardiac valve surgery in a hospital from January 2018 to October 2021 were collected retrospectively and divided into the CR-GNB group and the non-CR-GNB group according to whether CR-GNB infection had occurred after surgery, non-infected patients in the same period were selected, and paired in a 2:1 ratio with different infection groups (CR-GNB group and non-CR-GNB group) based on age and gender, the influencing factors for infection were analyzed, and the differences in clinical characteristics among groups were compared. Results Among the 11 120 patients underwent cardiac valve surgery, 296 cases (2.7%) developed HAI, including 170 cases of Gram-negative bacterial infection and 126 cases of Gram-positive bacterial infection. Among patients with Gram-negative bacterial infection, 62 cases (36.5%) were in the CR-GNB group, and 108 cases (63.5%) in the non-CR-GNB group. 112 CR-GNB strains were isolated from patients in the CR-GNB group, most of which were Acinetobacter baumannii (n=48, 42.9%), showing high resistance rates to several commonly used antimicrobial agents, except amikacin (12.5%) and polymyxin (5.3%). Comparing among groups, the durations of surgery, ICU stay, and extracorporeal circulation were longer in the CR-GNB group than in the non-CR-GNB group and in the non-infected group, and the non-CR-GNB group than the non-infected group. The proportion of antimicrobial use before infection was higher (all P < 0.05). Multivariate analysis showed that durations of operation, ICU stay and extracorporeal circulation were independent risk factors for CR-GNB infection in patients after cardiac valve surgery (all P < 0.05). The all-cause fatalities of patients in the CR-GNB and non-CR-GNB groups (31.8%, 54/170) were higher than that in the non-infected group (5.0%, 17/340; P < 0.05). Conclusion It is very important to strengthen perioperative management during cardiac surgery. Optimizing surgical operation and using antimicrobial agents rationally can reduce the occurrence of drug-resistant bacteria and improve the prognosis of patients.