INVESTIGATIONS OF BIFIDOBACTERIAL ECOLOGY AND OLIGOSACCHARIDE METABOLISM IN A 3-STAGE COMPOUND CONTINUOUS-CULTURE SYSTEM

Background: Several different types of in vitro fermentation systems a re currently employed to investigate pro- and prebiotic activities in the human large intestinal microbiota, ranging from simple batch cultu res, with or without stirring and pH control, to more complex models i nvolving pH controlled single and multiple-component continuous cultur e systems. Methods: In this investigation, we used a three-stage conti nuous culture model to study the activities of colonic bacteria. This fermentation system reproduces several of the nutritional and environm ental characteristics of the proximal large intestine (vessel 1) and t he distal colon (vessels 2 and 3), and was validated using bacteriolog ical, metabolic and chemical measurements made with intestinal materia l obtained from different regions of the large bowel. In this paper, w e report studies on prospective probiotic effects of Bifidobacterium l ongum NCFB 2259 in relation to other bacterial populations, production of tyrosine and phenylalanine metabolites, and bacterial synthesis of enzymes involved in the formation of putatively genotoxic metabolites , including beta-glucosidase (GS), arylsulphatase (AS), beta-glucuroni dase (GN), nitroreductase (NR) and azoreductase (AR). Results: Bacteri al activities at mio different retention times were studied (31.1 and 68.4 h), which correspond to large intestinal transit times. At R = 31 .4 h, significant probiotic effects were observed with respect to redu ctions in GS and GN, upon adding B. longum. However, despite the fact that this organism does not ferment aromatic amino acids or produce si gnificant amounts of genotoxic enzymes, dysbiotic manifestations occur red in that both NR synthesis and dissimilatory tyrosine metabolism we re stimulated. In contrast, at R = 68.4 h, GS formation increased betw een five and 20-fold, while GN and NR activities increased by a factor of two after adding the bifidobacterium. These data are reviewed in r elation to potential health hazards that may be encountered with long- term probiotic administration. In the prebiotic experiments, the three -stage fermentation system was operated at R = 65 h. Oligofructose was added to V1 to give an initial concentration of 30 grams per litre, w hen the system was in steady state, to study its effects on a number o f experimental parameters including bifidogenicity, bacterial growth, fermentation product formation and mutagenicity. After addition of the oligosaccharide, a multiplicity of effects were observed in V1, where synthesis of NR and AR, bifidobacterial populations and overall ferme ntation processes were stimulated, although these influences progressi vely diminished in V2 and V3. Conclusions: These studies indicate that bacterial metabolism and putative beneficial consequences associated with the breakdown of readily fermentable prebiotics in the large inte stine may in some circumstances be spatially and temporally limited to the proximal bowel.