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A Small In Vitro Fermentation Model for Screening the Gut Microbiota Effects of Different Fiber Preparations.

ABSTRACT: The development of prebiotic fibers requires fast high-throughput screening of their effects on the gut microbiota. We demonstrated the applicability of a mictotiter plate in the in vitro fermentation models for the screening of potentially-prebiotic dietary fibers. The effects of seven rye bran-, oat- and linseed-derived fiber preparations on the human fecal microbiota composition and short-chain fatty acid production were studied. The model was also used to study whether fibers can alleviate the harmful effects of amoxicillin-clavulanate on the microbiota. The antibiotic induced a shift in the bacterial community in the absence of fibers by decreasing the relative amounts of Bifidobacteriaceae, Bacteroidaceae, Prevotellaceae, Lachnospiraceae and Ruminococcaceae, and increasing proteobacterial Sutterilaceae levels from 1% to 11% of the total microbiota. The fermentation of rye bran, enzymatically treated rye bran, its insoluble fraction, soluble oat fiber and a mixture of rye fiber:soluble oat fiber:linseed resulted in a significant increase in butyrate production and a bifidogenic effect in the absence of the antibiotic. These fibers were also able to counteract the negative effects of the antibiotic and prevent the decrease in the relative amount of bifidobacteria. Insoluble and soluble rye bran fractions and soluble oat fiber were the best for controlling the level of proteobacteria at the level below 2%.

SUBMITTER: Tsitko I

PROVIDER: S-EPMC6514940 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Driven by the growing recognition of the influence of the gut microbiota (GM) on human health and disease, there is a rapidly increasing interest in understanding how dietary components, pharmaceuticals and pre- and probiotics influence GM. In vitro colon models represent an attractive tool for this purpose. With the dual objective of facilitating the investigation of rare and expensive compounds, as well as an increased throughput, we have developed a prototype in vitro parallel gut microbial fermentation screening tool with a working volume of only 5 ml consisting of five parallel reactor units that can be expanded with multiples of five to increase throughput. This allows e.g., the investigation of interpersonal variations in gut microbial dynamics and the acquisition of larger data sets with enhanced statistical inference. The functionality of the in vitro colon model, Copenhagen MiniGut (CoMiniGut) was first demonstrated in experiments with two common prebiotics using the oligosaccharide inulin and the disaccharide lactulose at 1% (w/v). We then investigated fermentation of the scarce and expensive human milk oligosaccharides (HMOs) 3-Fucosyllactose, 3-Sialyllactose, 6-Sialyllactose and the more common Fructooligosaccharide in fermentations with infant gut microbial communities. Investigations of microbial community composition dynamics in the CoMiniGut reactors by MiSeq-based 16S rRNA gene amplicon high throughput sequencing showed excellent experimental reproducibility and allowed us to extract significant differences in gut microbial composition after 24 h of fermentation for all investigated substrates and fecal donors. Furthermore, short chain fatty acids (SCFAs) were quantified for all treatments and donors. Fermentations with inulin and lactulose showed that inulin leads to a microbiota dominated by obligate anaerobes, with high relative abundance of Bacteroidetes, while the more easily fermented lactulose leads to higher relative abundance of Proteobacteria. The subsequent study on the influence of HMOs on two infant GM communities, revealed the strongest bifidogenic effect for 3'SL for both infants. Inter-individual differences of infant GM, especially with regards to the occurrence of Bacteroidetes and differences in bifidobacterial species composition, correlated with varying degrees of HMO utilization foremost of 6'SL and 3'FL, indicating species and strain related differences in HMO utilization which was also reflected in SCFAs concentrations, with 3'SL and 6'SL resulting in significantly higher butyrate production compared to 3'FL. In conclusion, the increased throughput of CoMiniGut strengthens experimental conclusions through elimination of statistical interferences originating from low number of repetitions. Its small working volume moreover allows the investigation of rare and expensive bioactives.

Project description:Vitamin B12 (cobalamin) is present in the human lower gastrointestinal tract either coming from the unabsorbed dietary fraction or from in situ production of the gut microbiota. However, it is unclear whether the gut microbial communities need exogenous B12 for growth and metabolism, or whether B12 in low and high levels could affect gut community composition and metabolite production. Here, we investigated in vitro B12 production of human fecal microbiota and the effects of different levels of B12 (as cyanocobalamin) on composition and activity. Eight fecal communities from healthy human adults distributed over three enterotypes, dominated by Firmicutes (n = 5), Bacteroides (n = 1) or Prevotella (n = 2) were used to perform batch fermentations in Macfarlane medium supplemented with low B12 medium (Control, 5 ng/ml, within the tested fecal range), no B12 addition (NB12), and high B12 addition (ExtraB12, 2500 ng/ml). The microbiota community composition (qPCR, 16S rRNA metabarcoding), metabolic activity (HPLC-RI), and B12 levels (UHPLC-DAD) were measured after 24 h incubation at 37°C under strict anaerobic conditions. All fecal microbial communities produced B12 in the NB12 condition after 24 h, in the range from 152 ± 4 to 564 ± 25 ng/ml. None of the B12 treatments had an impact on total bacterial growth, community richness, diversity and total metabolite production, compared to the low B12 control. However, a significant increase of propionate was measured in ExtraB12 compared to NB12. Most taxonomic and metabolite changes compared to control incubations were donor-dependent, implying donor-microbiota-specific changes upon B12 treatments. Our in vitro data suggest that healthy human adult gut microbial communities have the capacity to produce B12 at levels fulfilling their own requirements, independently of the initial B12 content tested in the donor's feces. Further, supplementation of exogenous dietary B12 may have limited impact on the healthy human gut microbial community composition and function.

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A Small In Vitro Fermentation Model for Screening the Gut Microbiota Effects of Different Fiber Preparations.

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