BACKGROUND: One of the biggest concerns associated with transgastric surgery is contamination and risk of intra-abdominal infection with microbes introduced from the access route. The purpose of this study was to evaluate the effect of oral decontamination with chlorhexidine on microbial contamination of the endoscope. METHODS: In a prospective, randomized, single-blinded, clinical trial the effect of chlorhexidine mouth rinse was evaluated. As a surrogate for the risk of intra-abdominal contamination during transgastric surgery, microbial contamination of the endoscope during upper endoscopy was examined. Patients referred to upper endoscopy were assessed for eligibility and randomized to either chlorhexidine or no mouth rinse. Culture samples were collected from gastric aspirates and endoscopes. The primary outcome measure was colony forming units (CFU) in the endoscope samples. Secondary outcome measures were species specific effect of chlorhexidine on micro-organisms with abscess forming capabilities and the effect of proton pump inhibitor (PPI) treatment on CFU. RESULTS: Chlorhexidine mouth rinse resulted in a significant reduction of CFU in the endoscope samples (p = 0.001). There was no species specific effect and micro-organisms with abscess forming capabilities were equally present. PPI treatment was associated with significantly higher CFU counts in both the gastric (p = 0.004) and endoscope samples (p = 0.049). CONCLUSIONS: Chlorhexidine mouth rinse was effective in reducing microbial contamination of the endoscope, but micro-organisms with abscess forming capabilities were still present. PPI treatment significantly increased CFU and should be discontinued before transgastric surgery.