POLYSACCHARIDE DEGRADATION BY HUMAN INTESTINAL BACTERIA DURING GROWTHUNDER MULTI-SUBSTRATE LIMITING CONDITIONS IN A 3-STAGE CONTINUOUS-CULTURE SYSTEM

Human faecal microorganisms were grown on mixtures of chemically diver se polymerised C-sources (starch, pectin, xylan, mucin, arabinogalacta n, inulin, guar gum) in a three-stage continuous culture model of the colon. The effects of retention time (R=27.1 h. R=66.7 h) on bacterial populations, their expression of hydrolytic enzymes involved in subst rate depolymerisation, carbohydrate utilisation and short chain fatty acid formation were investigated. Eleven bacterial marker groups were studied in the fermenters, Strictly anaerobic bacteria predominated in cluding bacteroides, bifidobacteria, clostridia and anaerobic Gram-pos itive cocci. Changing system retention time from 27.1 to 66.7 h did no t significantly affect the predominant bacterial populations in V1, ho wever; enterobacterial cell numbers increased in V3, while saccharolyt ic anaerobe recoveries declined markedly, reflecting their greater dep endence on polymerised carbon sources in the fermentation system. The majority of polysaccharide degrading activity in the colon model was c ell-associated, under all culture conditions. Increasing R from 27.1 t o 66.7 h did not substantially change overall polysaccharidase (amylas e, polygalacturonanase, alpha-xylanase, arabinogalactanase, galactoman nanase) profiles, however, synthesis of some glycosidases was enhanced (e.g. alpha-glucosidase, N-acetyl-beta-glucosaminidase, neuraminidase ), whereas reduced expression of other enzymes such as beta-galactosid ase, N-acetyl-alpha-galactosaminidase, alpha-fucosidase and alpha-arab inofuranosidase occurred. These observations demonstrate that cataboli te regulation is an important control process in the colonic microbiot a, with respect to the induction and repression of enzyme synthesis, a nd that substrate availability plays a major role in regulating bacter ial metabolism. Measurements of carbohydrate utilisation demonstrated that while all polysaccharides in the feed medium were digested extens ively by bacteria growing in the fermentation system, specific rates o f carbohydrate utilisation were maximal at R = 27.1 h. These data also provided evidence of bacterial substrate preferences in the colon mod el, particularly in relation to xylan and inulin digestion, demonstrat ing that catabolite regulatory mechanisms were also involved in contro lling the assimilation of carbohydrate in the microbiota. Short chain fatty acid measurements showed that fermentation was more efficient at R = 27.1 h compared to R = 66.7 h, with putative conversion of carboh ydrate to short chain fatty acids being approximately 60% and 40%, res pectively. This was probably due to increased maintenance energy requi rements at low bacterial growth rates. Differences were also observed with respect to short chain fatty acid molar ratios, with more propion ate, branched chain and longer chain fatty acid production at R = 66.7 h, demonstrating the increasing importance oi amino acid fermentation under these culture conditions. (C) 1998 Federation of European Micro biological Societies. Published by Elsevier Science B.V. All rights re served.