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Research on gut microbiota has generally focused on fecal samples, representing luminal content of the large intestine. However, nutrient uptake is restricted to the small intestine. Abundant immune cell populations at this anatomical site combined with diminished mucus secretion and looser junctions (partly to allow for more efficient fluid and nutrient absorption) also results in intimate host-microbe interactions despite more rapid transit. It is thus crucial to dissect key differences in both ecology and physiology between small and large intestine to better leverage the immense potential of human gut microbiota imprinting, including probiotic engraftment at biological sensible niches. Here, we provide a detailed review unfolding how the physiological and anatomical differences between the small and large intestine affect gut microbiota composition, function, and plasticity. This information is key to understanding how gut microbiota manipulation, including probiotic administration, may strain-dependently transform host-microbe interactions at defined locations.

Background: Infusion of short-chain fatty acids (SCFA) to the distal colon beneficially affects human substrate and energy metabolism. Here, we hypothesized that the combination of 2′-fucosyllactose (2′-FL) with resistant starch (RS) increases distal colonic SCFA production and improves metabolic parameters. Methods: In this randomized, crossover study, 10 lean (BMI 20–24.9 kg/m2) and nine men with prediabetes and overweight/obesity (BMI 25–35 kg/m2) were supplemented with either 2′-FL, 2′-FL+RS, or placebo one day before a clinical investigation day (CID). During the CID, blood samples were collected after a overnight fast and after intake of a liquid high-fat mixed meal to determine plasma SCFA (primary outcomes). Secondary outcomes were fasting and postprandial plasma insulin, glucose, free fatty acid (FFA), glucagon-like peptide-1, and peptide YY concentrations. In addition, fecal SCFA and microbiota composition, energy expenditure and substrate oxidation (indirect calorimetry), and breath hydrogen excretion were determined. Results: In lean men, supplementation with 2′-FL increased postprandial plasma acetate (P = 0.017) and fasting H2 excretion (P = 0.041) compared to placebo. Postprandial plasma butyrate concentration increased after 2′-FL and 2′-FL+RS as compared to placebo (P < 0.05) in lean men and men with prediabetes and overweight/obesity. Additionally, 2′-FL+RS decreased fasting and postprandial plasma FFA concentrations compared to placebo (P < 0.05) in lean men. Conclusion: Supplementation of 2′-FL with/without RS the day before investigation increased systemic butyrate concentrations in lean men as well as in men with prediabetes and obesity, while acetate only increased in lean men. The combination of 2′-FL with RS showed a putatively beneficial metabolic effect by lowering plasma FFA in lean men, indicating a phenotype-specific effect. Clinical trial registration: nr. NCT04795804.

Methanogens, reductive acetogens and sulfate-reducing bacteria play an important role in disposing of hydrogen in gut ecosystems. However, how they interact with each other remains largely unknown. This in vitro study cocultured Blautia hydrogenotrophica (reductive acetogen), Desulfovibrio piger (sulfate reducer) and Methanobrevibacter smithii (methanogen). Results revealed that these three species coexisted and did not compete for hydrogen in the early phase of incubations. Sulfate reduction was not affected by B. hydrogenotrophica and M. smithii. D. piger inhibited the growth of B. hydrogenotrophica and M. smithii after 10 h incubations, and the inhibition on M. smithii was associated with increased sulfide concentration. Remarkably, M. smithii growth lag phase was shortened by coculturing with B. hydrogenotrophica and D. piger. Formate was rapidly used by M. smithii under high acetate concentration. Overall, these findings indicated that the interactions of the hydrogenotrophic microbes are condition-dependent, suggesting their interactions may vary in gut ecosystems.

Twelve novel resistant starch type 3 substrates differing in crystal type (A/B), alpha-1,4 glucan chain length (degree of polymerization 14 - 76) and molecular weight distribution (1.08 - 2.11) were fermented in vitro using an inoculum of pooled faeces obtained from four healthy adults. A selection of substrates was also treated with pancreatin to remove an easily digestible fraction, prior to in vitro fermentation using the same pooled adult faecal inoculum., Twelve novel resistant starch type 3 substrates differing in crystal type (A/B), alpha-1,4 glucan chain length (degree of polymerization 14 - 76) and molecular weight distribution (1.08 - 2.11) were fermented in vitro using an inoculum of pooled faeces obtained from four healthy adults. A selection of substrates was also treated with pancreatin to remove an easily digestible fraction, prior to in vitro fermentation using the same pooled adult faecal inoculum.

Bacteriophages (or phages for short) are natural predators of bacteria, and their application at food industry has gain increasingly interested. In order to understand the safety of using Shiga Toxin producing Esherichia coli (STEC) specific phages to the healthy human gut microbiota an in vitro fermentation model that mimics the gut was proposed. Phage Ace, specific STEC phage, and E. coli CECT 5947 were added to the in vitro model as perturbation alone or in combination. Fermentation parameters were monitered, and the final microbiota composition was assessed through shot gun metagenomics. It was shown that the addition of only phage Ace did not result in a perturbation to fermentation, however, the addition of the E. coli strain leaded to differences in the fermentation and microbiota composition. This model was shown to be useful for a first safety screening of phages intended to be used in the biocontrol of food stuff.

Galacto-oligosaccharides (GOS) and 2′-fucosyllactose (2′-FL) are non-digestible carbohydrates (NDCs) that are often added to infant formula to replace the functionalities of human milk oligosaccharides (HMOs). It is not known if combining GOS and 2′-FL will affect their fermentation kinetics and subsequent immune-modulatory effects such as AhR-receptor stimulation. Here, we used an in vitro set-up for the fermentation of 2′-FL and GOS, either individually or combined, by fecal microbiota of 8-week-old infants. We found that GOS was fermented two times faster by the infant fecal microbiota when combined with 2′-FL, while the combination of GOS and 2′-FL did not result in a complete degradation of 2′-FL. Fermentation of both GOS and 2′-FL increased the relative abundance of Bifidobacterium, which coincided with the production of acetate and lactate. Digesta of the fermentations influenced dendritic cell cytokine secretion differently under normal conditions and in the presence of the AhR-receptor blocker CH223191. We show that, combining GOS and 2′-FL accelerates GOS fermentation by the infant fecal microbiota of 8-week-old infants. In addition, we show that the fermentation digesta of GOS and 2′-FL, either fermented individually or combined, can attenuate DC cytokine responses in a similar and in an AhR-receptor dependent way.

Detrimental consequences of antibiotic treatment may include long-lasting disruption of the gut microbiota. Previous studies found no negative effects of antibiotics on metabolic health, although individualized responses were observed. Here, we aimed to investigate the subject-specific response to vancomycin use in tissue-specific insulin sensitivity by stratifying individuals based on the presence of antibiotic resistance genes (ARGs) or opportunistic pathogens (OPs) in the baseline fecal microbiota. Quantitative Polymerase Chain Reaction (qPCR) was used to detect ARGs and OPs in DNA isolated from fecal samples of 56 males with overweight/obesity (Body Mass Index: 25–35 kg/m2) and impaired glucose metabolism (fasting plasma glucose ≥5.6 mmol/L and/or 2-hour glucose 7.8–11.1 mmol/L). A two-step hyperinsulinemic-euglycemic clamp was performed to determine tissue-specific insulin sensitivity. Abdominal subcutaneous adipose tissue (AT) gene expression was assessed using Affymetrix microarray. Gut microbial composition was determined using the Human Intestinal Tract Chip (HITChip) microarray. At baseline, the vancomycin resistance gene vanB was present in 60% of our population. In individuals that were vanB-negative at baseline, AT insulin sensitivity (insulin-mediated suppression of plasma free fatty acids) improved during vancomycin use, while it decreased among vanB-positive individuals (% change post versus baseline: 14.1 ± 5.6 vs. −6.7 ± 7.5% (p = .042)). The vancomycin-induced increase in AT insulin sensitivity was accompanied by downregulation of inflammatory pathways and enrichment of extracellular matrix remodeling pathways in AT. In the vanB-positive group, well-known vanB-carrying bacteria, Enterococcus and Streptococcus, expanded in the gut microbiome. In conclusion, microbiome composition and adipose tissue biology were differentially affected by vancomycin treatment based on fecal vanB carriage.

Infusions of the short-chain fatty acid (SCFA) acetate in the distal colon improved metabolic parameters in men. Here, we hypothesized that combining rapidly and slowly fermentable fibers will enhance distal colonic acetate production and improve metabolic health. In vitro cultivation studies in a validated model of the colon were used to identify fiber mixtures that yielded high distal colonic acetate production. Subsequently, in two randomized crossover studies, lean and prediabetic overweight/obese men were included. In one study, participants received supplements of either long-chain inulin+resistant starch (INU+RS), INU or maltodextrin (PLA) the day prior to a clinical investigation day (CID). The second trial studied beta glucan+RS (BG+RS) versus BG and PLA. During each CID, breath hydrogen, indirect calorimetry, plasma metabolites/hormones were assessed during fasting and postprandial conditions. Additionally, fecal microbiota composition and SCFA were determined. In prediabetic men, INU+RS increased plasma acetate compared to INU or PLA (P < .05), but did not affect metabolic parameters. In lean men, INU+RS increased breath hydrogen and fasting plasma butyrate, which was accompanied by increased energy expenditure, carbohydrate oxidation and PYY and decreased postprandial glucose concentrations (all P < .05) compared to PLA. BG+RS increased plasma butyrate compared to PLA (P < .05) in prediabetic individuals, but did not affect other fermentation/metabolic markers in both phenotypes. Fiber-induced shifts in fecal microbiota were individual-specific and more pronounced with INU+RS versus BG+RS. Administration of INU+RS (not BG+RS) the day prior to investigation improved metabolic parameters in lean but not in prediabetic individuals, demonstrating that effects were phenotype- and fiber-specific. Further research should study whether longer-term supplementation periods are required to elicit beneficial metabolic health in prediabetic individuals. Trial registr

Hydrogenotrophic microbes, primarily including the three functional groups methanogens, sulfate-reducing bacteria, and reductive acetogens, use hydrogen as an energy source and play an important role in maintaining the hydrogen balance in gut ecosystems. A distorted hydrogen balance has been associated with irritable bowel syndrome (IBS). However, the role of hydrogenotrophic microbes in overall microbiota composition and function remains largely unknown. This study aims to assess the distribution and stability of hydrogenotrophic functional groups in healthy adults (HAs) and IBS patients and their association with overall microbiota composition and IBS symptoms. A two-time-point study with 4 weeks in between was performed with 27 HAs and 55 IBS patients included. Our observations revealed that methanogens showed a bimodal distribution across samples. A high-level methanogen microbiota was consistently associated with higher alpha diversity, and its composition was significantly different from that of individuals with a low-level methanogen microbiota. In general, these associations were more pronounced in IBS patients than in HAs. The differences in the copy numbers of genes indicative of total bacteria and acetogens between HAs and IBS patients and their correlations with IBS symptom severity, anxiety, depression, and quality of life (QoL) were sampling time dependent. Hydrogenotrophic functional groups did not show negative abundance correlations with each other in HAs and IBS patients. These findings suggest that methanogen levels in the gut have a pronounced association with microbiota alpha diversity and composition, and the interactions between hydrogenotrophic functional groups are complex in gut ecosystems.