Sulfide- and nitrite-dependent nitric oxide production in the intestinal tract

In the gut ecosystem, nitric oxide (NO) has been described to have damaging effects on the energy metabolism of colonocytes. Described mechanisms of NO production are microbial reduction of nitrate via nitrite to NO and conversion of l-arginine by NO synthase. The aim of this study was to investigate whether dietary compounds can stimulate the production of NO by representative cultures of the human intestinal microbiota and whether this correlates to other processes in the intestinal tract. We have found that the addition of a reduced sulfur compound, i.e. cysteine, contributed to NO formation. This increase was ascribed to higher sulfide concentrations generated from cysteine that in turn promoted the chemical conversion of nitrite to NO. The NO release from nitrite was of the order of 4 parts per thousand at most. Overall, it was shown that two independent biological processes contribute to the chemical formation of NO in the intestinal tract: (i) the production of sulfide by fermentation of sulfur containing amino acids or reduction of sulfate by sulfate reducing bacteria, and (ii) the reduction of nitrate to nitrite. Our results indicate that dietary thiol compounds in combination with nitrate may contribute to colonocytes damaging processes by promoting NO formation

Arabinoxylans, inulin and Lactobacillus reuteri 1063 repress the adherent-invasive Escherichia coli from mucus in a musosa-comprising gut model

The microbiota that colonises the intestinal mucus may particularly affect human health given its proximity to the epithelium. For instance, the presence of the adherent-invasive Escherichia coli (AIEC) in this mucosal microbiota has been correlated with Crohn's disease. Using short-term screening assays and a novel long-term dynamic gut model, which comprises a simulated mucosal environment (M-SHIME), we investigated how (potential) pro-and prebiotics may repress colonisation of AIEC from mucus. Despite that during the short-term screening assays, some of the investigated Lactobacillus strains adhered strongly to mucins, none of them competed with AIEC for mucin-adhesion. In contrast, AIEC survival and growth during co-culture batch incubations was decreased by Lactobacillus rhamnosus GG and L. reuteri 1063, which correlated with (undissociated) lactic acid and reuterin levels. Regarding the prebiotics, long-chain arabinoxylans (LC-AX) lowered the initial mucin-adhesion of AIEC, while both inulin (IN) and galacto-oligosaccharides (GOS) limited AIEC survival and growth during batch incubations. L. reuteri 1063, LC-AX and IN were thus retained for a long-term study with the M-SHIME. All treatments repressed AIEC from mucus without affecting AIEC numbers in the luminal content. As a possible explanation, L. reuteri 1063 treatment increased lactobacilli levels in mucus, while LC-AX and IN additionally increased mucosal bifidobacteria levels, thus leading to antimicrobial effects against AIEC in mucus. Overall, this study shows that pro-and prebiotics can beneficially modulate the in vitro mucosal microbiota, thus limiting occurrence of opportunistic pathogens among those mucosal microbes which may directly interact with the host given their proximity to the epithelium

Nitric Oxide Production by the Human Intestinal Microbiota by Dissimilatory Nitrate Reduction to Ammonium

The free radical nitric oxide (NO) is an important signaling molecule in the gastrointestinal tract. Besides eukaryotic cells, gut microorganisms are also capable of producing NO. However, the exact mechanism of NO production by the gut microorganisms is unknown. Microbial NO production was examined under in vitro conditions simulating the gastrointestinal ecosystem using L-arginine or nitrate as substrates. L-arginine did not influence the microbial NO production. However, NO concentrations in the order of 90 ng NO-N per L feed medium were produced by the fecal microbiota from nitrate. 15N tracer experiments showed that nitrate was mainly reduced to ammonium by the dissimilatory nitrate reduction to ammonium (DNRA) pathway. To our knowledge, this is the first study showing that gastrointestinal microbiota can generate substantial amounts of NO by DNRA and not by the generally accepted denitrification or L-arginine pathway. Further work is needed to elucidate the exact role between NO produced by the gastrointestinal microbiota and host cells

Impact of carbohydrate substrate complexity on the diversity of the human colonic microbiota.

The diversity of the colonic microbial community has been linked with health in adults and diet composition is one possible determinant of diversity. We used carefully controlled conditions in vitro to determine how the complexity and multiplicity of growth substrates influence species diversity of the human colonic microbiota. In each experiment, five parallel anaerobic fermenters that received identical faecal inocula were supplied continuously with single carbohydrates (either arabinoxylan-oligosaccharides (AXOS), pectin or inulin) or with a '3-mix' of all three carbohydrates, or with a '6-mix' that additionally contained resistant starch, β-glucan and galactomannan as energy sources. Inulin supported less microbial diversity over the first 6 d than the other two single substrates or the 3- and 6-mixes, showing that substrate complexity is key to influencing microbiota diversity. The communities enriched in these fermenters did not differ greatly at the phylum and family level, but were markedly different at the species level. Certain species were promoted by single substrates, whilst others (such as Bacteroides ovatus, LEfSe P = 0.001) showed significantly greater success with the mixed substrate. The complex polysaccharides such as pectin and arabinoxylan-oligosaccharides promoted greater diversity than simple homopolymers, such as inulin. These findings suggest that dietary strategies intended to achieve health benefits by increasing gut microbiota diversity should employ complex non-digestible substrates and substrate mixtures