VALIDATION OF A 3-STAGE COMPOUND CONTINUOUS-CULTURE SYSTEM FOR INVESTIGATING THE EFFECT OF RETENTION TIME ON THE ECOLOGY AND METABOLISM OF BACTERIA IN THE HUMAN COLON

A three-stage compound continuous culture system was used to study the effect of retention time (27.1 and 66.7 h) on the catabolism of organ ic carbon and nitrogen sources in mixed populations of human colonic b acteria. The fermentation system was designed to reproduce spatial, te mporal, nutritional, and physicochemical characteristics of the microb iota in the proximal (vessel 1) and distal (vessels 2 and 3) colons, a nd was validated on the basis of chemical and microbiological measurem ents on intestinal contents obtained from human sudden death victims. Results showed that the majority of carbohydrate breakdown and short-c hain fatty acid production occurred in V1. Conversely, dissimilatory a mino acid metabolism, as evidenced by formation of branched-chain fatt y acids and phenolic compounds, occurred primarily in V2 and V3. Ferme ntation of aromatic amino acids was strongly affected by system retent ion lime (R), with concentrations of phenolic metabolites being three times higher in V3, at 66.7 h, compared to 27.1 h. Bacteriological mea surements of intestinal contents, in which nine groups of marker organ isms were studied, showed that, with the exception of bifidobacteria, no major differences in relative bacterial cell numbers were evident i n the proximal and distal colons. These organisms were also studied in the continuous culture system, where marked reductions in Escherichia coli were observed in V2 and V3, especially at R = 27.1 h. Increasing R to 66.7 h reduced numbers of Clostridium perfringens, anaerobic Gra m-positive cocci, and total anaerobe counts. Correlations between in v ivo chemical and bacteriological measurements and data obtained in vit ro demonstrate that the three-stage fermentation system provided a use ful model for studying the physiology and ecology of large intestinal microorganisms under different nutritional and environmental condition s.