Objective To investigate the continuous use time of water injection bottles. Methods Five sets of water injection bottles in the endoscopy center of a tertiary first-class hospital in Chongqing was monitored through prospective study, sterile water was injected into the water injection bottles from July 2019 to November 2019, water was collected for microbiological test on the 1st, 2nd, 3rd, 4th and 5th days after diagnosis and treatment every day, bacterial species and qualified rate of bacterial colony count in water injection bottle was monitored. Results Qualified rates of bacterial colony count on the 1st, 2nd, 3rd, 4th, and 5th days were 96.0%, 82.0%, 76.0%, 70.0%, and 38.0% respectively, difference was statistically significant (P<0.001). A total of 69 strains of bacteria were isolated, all were opportunistic pathogens, among which 29 strains were Ralstonia mannitolilytica and 10 strains were Pseudomonas aeruginosa. Conclusion Medical institutions should pay more attention to the microbial contamination of water for endoscopic diagnosis and treatment, water injection bottles have become a potential source of water contamination, in order to reduce potential risk of endoscopic diagnosis and treatment, continuous use of water injection bottles should not exceed one day.