Objective To explore the potential influencing factors of post-operative healthcare-associated infection (HAI) in colorectal cancer patients, as well as the mediating effect relationship between the influencing factors and the diagnosis-intervention packet (DIP) payment differentials. Methods Medical data of patients who underwent colorectal cancer surgery in a tertiary first-class cancer hospital in Guangzhou were retrospectively analyzed. According to HAI status, patients were divided into infection group and non-infection group. Baseline demographic information and differences in DIP payment differentials between two groups of patients were compared by rank sum test or chi-square test. The influence of each potential factor on the occurrence of HAI was analyzed by logistic regression. Mediating analysis was preformed by bootstrap method, and mediating effect of HAI and total hospitalization days on DIP payment differentials was evaluated. Results A total of 350 patients were included in analysis, 50 were in the infection group and 300 in the non-infection group. The incidence of HAI was 14.29%. Logistic regression analysis result showed that risk of HAI in patients with central venous catheterization ≥10.00 days was 13.558 times higher than that <10.00 days (P<0.001); risk of HAI in patients with urinary catheterization ≥3.00 days was 2.388 times higher than that <3.00 days (P=0.022). There were all statistically significant differences in DIP payment differentials among patients with different ages, prognostic nutritional index (PNI), HAI status, total length of hospitalization stay, duration of surgery, central venous catheterization days, and catheterization days (all P<0.05). The mediating analysis results showed that the occurrence of HAI resulted a change in DIP payment di-fferentials by affecting the total number of hospitalization days. The mediating effect value of total hospitalization days was 0.038, accounting for 35.68% of the total effect. Conclusion Medical institutions should pay attention to HAI resulting from prolonged central venous and urinary catheterization in patients underwent surgery for colorectal cancer, reducing the total length of hospital stay, thus reducing the overruns associated with the increased DIP payment differentials.