Involvement of the Commensal Organism Bacillus subtilis in the Pathogenesis of Anastomotic Leak.

Background: It is now well established that microbes play a key and causative role in the pathogenesis of anastomotic leak. Yet, in patients, determining whether a cultured pathogen retrieved from an anastomotic leak site is a cause or a consequence of the complication remains a challenge. The aim of this study was to test a methodology to invoke causality between a retrieved microbe from a leak site and its role in anastomotic leak. Methods: The commensal organism Bacillus subtilis was isolated from an esophagojejunostomy leak site in a 35-year-old patient with a CDH1 mutation after a prophylactic gastrectomy whose body mass index (BMI) was 35 kg/m2. The organism was screened for its ability to degrade collagen, shift human recombinant matrix metalloprotease-9 (MMP9) to its active form, and induce a clinical anastomotic leak when introduced to anastomotic tissues of mice fed their standard diet (SD) of chow or an obesogenic Western-type diet (WD). Results: The Bacillus subtilis strain retrieved from the anastomotic leak site displayed a high degree of collagenolytic activity and was able to activate human MMP9 consistent with other pathogens expressing this characteristic "leak phenotype." Exposure of the Bacillus subtilis to the anastomotic tissues of obese mice fed a WD led to dehiscence of the anastomosis, abscess formation with peritonitis, and mortality in 50% of mice (3/6). When anastomotic healing was evaluated by a validated anastomotic healing score (AHS), substantially worse healing was observed (i.e., higher AHS) in WD-fed mice exposed to Bacillus subtilis compared to SD-fed mice (analysis of variance [ANOVA], p = 0.0006). Conclusions: Microbial strains obtained from patients' anastomotic leak sites can be evaluated for their pathogenic in the leak process by assessing their ability to produce collagenase, activate MMP9 and cause clinical leaks in mice fed a WD. These studies may aid in identifying those bacterial strains that play a causal role in patients with an anastomotic leak.