Search

A high-density phylogenetic microarray targeting small subunit rRNA (SSU rRNA) sequences of over 1000 microbial phylotypes of the human gastrointestinal tract, the HITChip, was used to assess the impact of faecal inoculum preparation and operation conditions on an in vitro model of the human large intestine (TIM-2). This revealed that propagation of mixed faecal donations for the production of standardized inocula has only a limited effect on the microbiota composition, with slight changes observed mainly within the Firmicutes. Adversely, significant shifts in several major groups of intestinal microbiota were observed after inoculation of the in vitro model. Hierarchical cluster analysis was able to show that samples taken throughout the inoculum preparation grouped with microbiota profiles observed for faecal samples of healthy adults. In contrast, the TIM-2 microbiota was distinct. While members of the Bacteroidetes and some groups within the Bacilli were increased in TIM-2 microbiota, a strong reduction in the relative abundance of other microbial groups, including Bifidobacterium spp., Streptococcus spp., and Clostridium clusters IV and XIVa, was observed. The changes detected with the HITChip could be confirmed using denaturing gradient gel electrophoresis (DGGE) of SSU rRNA amplicons. DOI 10.1099/mic.0.042044-0

A validated in vitro model of the large intestine ( TIM-2), set up with human or pig faeces, was used to evaluate the impact of potentially probiotic Lactobacillus amylovorus DSM 16698, administered alone (i), in the presence of prebiotic galactooligosaccharides ( GOS) (ii), and co-administered with probiotic Bifidobacterium animalis ssp. lactis Bb-12 ( Bb-12) (iii) on GOS degradation, microbial growth ( L. amylovorus, lactobacilli, bifidobacteria and total bacteria) and metabolite production. High performance anion exchange chromatography revealed that GOS degradation was more pronounced in TIM-2 inoculated with pig faeces than with human faeces. Denaturing gradient gel electrophoresis profiling of PCR-amplified 16S r RNA genes detected a more complex Lactobacillus spp. community in pig faecal material than in human faecal inoculum. According to 16S r RNA gene-targeted q PCR, GOS stimulated the growth of lactobacilli and bifidobacteria in faecal material from both materials. The cumulative production of short chain fatty acids and ammonia was higher ( P < 0.05) for pig than for human faeces. However, lactate accumulation was higher ( P < 0.05) in the human model and increased after co-administration with GOS and Bb-12. This study reinforced the notion that differences in microbiota composition between target host organisms need to be considered when animal data are extrapolated to human, as is often done with pre- and probiotic intervention studies. [ABSTRACT FROM AUTHOR]

Galacto-oligosaccharides (GOS) are considered to be prebiotic, altliough the contribution of specific members of the microbiota to GOS fermentation and the exact microbial metabolites that are produced upon GOS fermentation are largely unknown. We aimed to determine this using uniformly 13C-labeled GOS. The normal (control) medium and unlabeled or 13C-labeled GOS was added to a dynamic, validated, in vitro model of the large-intestine containing an aduit-type microbiota. Liquid-chromatography MS was used to measure the incorporation of C label into metabolites. 16S-rRNA stable isotope probing coupled to a phylogenetic micro-array was used to determine label incorporation in microbial biomass. The priman/ members within the complex microbiota that were directly involved in GOS fermentation were shown to be Bifidobacterium longum, B. bifidum, B. catenulatum, Lactobacillus gasseh, and L. salivarius, in line with the prebiotic effect of GOS, although some other species incorporated label also. GOS fermentation led to an increase in acetate (+49%) and lactate (+23%) compared with the control. Total organic acid production was 8.50 and 7.52 mmol/g of carbohydrate fed for the GOS and control experiments, respectively. At the same time, the cumulative production of putrefactive metabolites (branched-chain fatty acids and ammonia) was reduced by 55%. Cross-feeding of metabolites from primary GOS fermenters to other members of the microbiota was observed. Our findings support a prebiotic role for GOS and its potential to act as a synbiotic in combination with certain probiotic strains. [ABSTRACT FROM AUTHOR]

This study introduces a stable-isotope metabolic approach employing [U-13C]glucose that, as a novelty, allows selective profiling of the human intestinal microbial metabolic products of carbohydrate food components, as well as the measurement of the kinetics of their formation pathways, in a single experiment. A well-established, validated in vitro model of human intestinal fermentation was inoculated with standardized gastrointestinal microbiota from volunteers. After culture stabilization, [U-13C]glucose was added as an isotopically labeled metabolic precursor. System lumen and dialysate samples were taken at regular intervals. Metabolite concentrations and isotopic labeling were determined by NMR, GC, and enzymatic methods. The main microbial metabolites were lactate, acetate, butyrate, formate, ethanol, and glycerol. They together accounted for a 13C recovery rate as high as 91.2%. Using an NMR chemical shift prediction approach, several minor products that showed 13C incorporation were identified as organic acids, amino acids, and various alcohols. Using computer modeling of the 12C contents and 13C labeling kinetics, the metabolic fluxes in the gut microbial pathways for synthesis of lactate, formate, acetate, and butyrate were determined separately for glucose and unlabeled background substrates. This novel approach enables the study of the modulation of human intestinal function by single nutrients, providing a new rational basis for achieving control of the short-chain fatty acids profile by manipulating substrate and microbiota composition in a purposeful manner. Copyright © 2009 John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]

The article discusses a study which determined the effect of maize-based fibers on the activity and composition of microbiota in the colon. Results showed that by microbial metabolites analysis, an increase in short-chain fatty acids for all fibers, including different butyrate production levels for each fiber, were identified. All the fibers reduced toxic metabolites and showed increased numbers of bifidobacteria and Lactobacillus species. The study concluded that all fibers displayed prebiotic activity with regard to more beneficial microbes.

This study introduces a stable-isotope metabolic approach employing [U-(13)C]glucose that, as a novelty, allows selective profiling of the human intestinal microbial metabolic products of carbohydrate food components, as well as the measurement of the kinetics of their formation pathways, in a single experiment. A well-established, validated in vitro model of human intestinal fermentation was inoculated with standardized gastrointestinal microbiota from volunteers. After culture stabilization, [U-(13)C]glucose was added as an isotopically labeled metabolic precursor. System lumen and dialysate samples were taken at regular intervals. Metabolite concentrations and isotopic labeling were determined by NMR, GC, and enzymatic methods. The main microbial metabolites were lactate, acetate, butyrate, formate, ethanol, and glycerol. They together accounted for a (13)C recovery rate as high as 91.2%. Using an NMR chemical shift prediction approach, several minor products that showed (13)C incorporation were identified as organic acids, amino acids, and various alcohols. Using computer modeling of the (12)C contents and (13)C labeling kinetics, the metabolic fluxes in the gut microbial pathways for synthesis of lactate, formate, acetate, and butyrate were determined separately for glucose and unlabeled background substrates. This novel approach enables the study of the modulation of human intestinal function by single nutrients, providing a new rational basis for achieving control of the short-chain fatty acids profile by manipulating substrate and microbiota composition in a purposeful manner.