Three-stage continuous culture system with a self-generated anaerobia to study the regionalized metabolism of the human gut microbiota.

The technical and ethical difficulties in studying the gut microbiota in vivo warrant the development and improvement of in vitro systems able to simulate and control the physicochemical factors of the gut biology. Moreover, the functional regionalization of this organ implies a model simulating these differences. Here we propose an improved and alternative three-stage continuous bioreactor called 3S-ECSIM (three-stage Environmental Control System for Intestinal Microbiota) to study the human large intestine. Its main feature compared with other in vitro systems is the anaerobic atmosphere originating directly from the microbiota metabolism, leading to different gas ratios of CO2 and H2 in each compartment. Analyses of the metabolic and microbiological profiles (LC-MS and a phylogenetic microarray) show different profiles together with a maintenance of this differentiation between the three compartments, simulating respectively a proximal, a transversal and a distal colon. Moreover, the last reactor presents a high similarity with the initial fecal sample, at the microbiological diversity level. Based on our results, this in-vitro process improvement is a valuable alternative tool to dynamically study the structure and metabolism of gut microbiota, and its response to nutrients, prebiotics, probiotics, drugs or xenobiotics.