SCFA Profile Research Articles

Multi-Omics Analysis Unravels the Impact of Stool Sample Logistics on Metabolites and Microbial Composition.

Human health and the human microbiome are inevitably intertwined, increasing their relevance in clinical research. However, the collection, transportation and storage of faecal samples may introduce bias due to methodological differences, especially since postal shipping is a common practise in large-scale clinical cohort studies. Using four different Omics layer, we determined the structural (16S rRNA sequencing, cytometric microbiota profiling) and functional integrity (SCFAs, global metabolome) of the microbiota in relation to different easy-to-handle conditions. These conditions were storage at -20 °C, -20 °C as glycerol stock, 4 °C and room temperature with and without oxygen exposure for a maximum of one week. Storage time affected the microbiota on all Omics levels. However, the magnitude was donor-dependent, highlighting the need for purpose-optimized sample collection in clinical multi-donor studies. The effects of oxygen exposure were negligible for all analyses. At ambient temperature, SCFA and compositional profiles were stable for 24 h and 48 h, respectively, while at 4 °C, SCFA profiles were maintained for 48 h. The global metabolome was highly susceptible, already changing at 24 h in non-frozen conditions. Thus, faecal microbiota was best preserved on all levels when transported as a native sample frozen within 24 h, leading to the least biased outcomes in the analysis. We conclude that the immediate freezing of native stool samples for transportation to the lab is best suited for planned multi-Omics analyses that include metabolomics to extend standard sequencing approaches.

The effect of supplemental arginine on the gut microbial homeostasis of broilers during sub-clinical necrotic enteritis challenge.

Necrotic enteritis (NE) is an enteric disease of poultry that alters the structure of the gut microbial community causing dysbiosis. This 28day experiment investigated the effects of 125% and 135% arginine diets on the gut microbial diversity and composition of broilers during a subclinical NE challenge. One hundred and twenty one-day-old chicks were randomly allocated to 4 treatments with six replicates each- Uninfected + Basal, NE + Basal, NE + Arg 125%, and NE + Arg 135% diet groups. NE was induced by inoculating 1 × 104E. maxima sporulated oocysts on day 14 and 1 × 108CFUC. perfringens on days 19, 20, and 21 of age. The NE challenge significantly decreased the number of observed amplicon sequence variants (p = 0.03), the abundance of the phylum Firmicutes (p < 0.01), and the species Mediterraneibacter cottocaccae (p = 0.01) in the ceca of birds on day 21. The NE challenge significantly increased the Bray-Curtis index (p < 0.01), and the abundance of the phylum Bacteroidota (p < 0.01), family Odoribacteraceae (p < 0.01), genus Odoribacter (p < 0.01), and species O. splanchnicus (p = 0.01) on day 21. During NE, the 125% arginine diet restored the abundance of the phylum Bacteroidota (p = 0.03), family Odoribacteraceae (p = 0.03) and Oscillospiraceae (p = 0.03), genus Odoribacter (p = 0.03), and species O. splanchnicus (p = 0.03) and M. cottocaccae (p < 0.01) on day 21. The 135% arginine diet effectively restored the loss in alpha diversity (p = 0.01) caused by NE, the abundance of the phylum Firmicutes (p = 0.01) and Bacteroidota (p < 0.01), family Oscillospiraceae (p = 0.03) and Odoribacteraceae (p < 0.01), genus Odoribacter (p < 0.01), and species O. splanchnicus (p < 0.01) and M. cottocaccae (p < 0.01) on day 21. On day 28, the treatments had a significant effect on the cecal propionate (p = 0.01), butyrate (p = 0.04), and total SCFA (p = 0.04) concentrations. In conclusion, the 125% and 135% arginine diets restored gut microbial composition during a subclinical NE challenge, but not the cecal SCFA profile. Hence, arginine in combination with other feed additives could be used in restoring gut microbial homeostasis during NE in poultry.

Effect of Maternal Egg Yolk Supplementation on Gut Microbiome and SCFA Profiles in Piglets

Effect of Maternal Egg Yolk Supplementation on Gut Microbiome and SCFA Profiles in Piglets

Dynamics of in vitro rumen methane production after nitrate addition

ABSTRACT The present study aimed to assess the dynamics of rumen methane (CH4) production following the addition of NaNO3. This was done using an in vitro rumen fermentation system that ensures continuous gas and methane assessments. Four different levels of NaNO3 were used to get the final nitrate concentrations of 0.5, 1.0, 1.5, and 2.0 mg/ml of rumen fluid. For each dose, corresponding controls contained sodium chloride and urea were realised to ensure comparable levels of sodium and nitrogen. The addition of nitrates had slight effect on the intensity of fermentation because the total gas produced minus CH4 (total methane-free gas) only went down at the highest dose (2.0 mg/ml), and the final concentrations of SCFA were the same at all doses. The most evident effect was a modification of the SCFA profile (low concentrations of propionate and valerate, progressive increments of acetate, and decreases of butyrate) and a reduction in overall CH4 production. The CH4 yield for the 0.5 mg/ml dose was not different from control in the entire fermentation. Yield of the 1.0 mg/ml dose was significantly lower than the control group (p < 0.05) only within the initial 24-h period, and higher dosages (1.5 and 2.0 mg/ml) were lower during the entire fermentation (p < 0.01). Methane yields were well fitted with the Gompertz model, but only the highest level of nitrate inclusion had a significant impact on the majority of model parameters (p < 0.01). The linear regressions between CH4 yields (y) and the amounts of nitrates (x) at progressive fermentation durations (e.g. 6, 12, 24, and 48 h) produced equations with increasing absolute slopes (from −0.069 to −0.517 ml/mg of nitrate). Therefore, nitrate reduced rumen CH4 yield in a dose-dependent manner: the impact of low doses was primarily observed at the initial stages of fermentation, whereas high doses exhibited effectiveness throughout the entire fermentation process. In conclusion, in batch fermentation systems, the dose effect of nitrates on methane yield was time dependent.

P27-018-23 Comparing Maternal Diet and Fecal SCFA Profiles of Infants With and Without Prenatal Opioid Exposure

P27-018-23 Comparing Maternal Diet and Fecal SCFA Profiles of Infants With and Without Prenatal Opioid Exposure

Composition of mucus- and digesta-associated bacteria in growing pigs with and without diarrhea differed according to the presence of colonic inflammation

BackgroundIn the pig production, diarrhea can occur during different growth stages including the period 4–16 weeks post weaning, during which a diarrheal outbreak also termed as colitis-complex diarrhea (CCD) can occur and it is distinct from post-weaning diarrhea (1–2 weeks post weaning). We hypothesized that CCD in growing pigs is associated with changes in colonic microbiota composition and fermentation patterns, and the aim of the present observational study was to identify changes in digesta-associated bacteria (DAB) and mucus-associated bacteria (MAB) in the colon of growing pigs with and without diarrhea. A total number of 30 pigs (8, 11, and 12 weeks of age) were selected; 20 showed clinical signs of diarrhea and 10 appeared healthy. Based on histopathological examination of colonic tissues, 21 pigs were selected for further studies and classified as follows: without diarrhea, no colon inflammation (NoDiar; n = 5), with diarrhea, without colonic inflammation (DiarNoInfl; n = 4), and with diarrhea, with colonic inflammation (DiarInfl; n = 12). Composition (based on 16S rRNA gene amplicon sequencing) and fermentation pattern (short-chain fatty acids; SCFA profile) of the DAB and MAB communities were characterized.ResultsThe DAB showed higher alpha diversity compared to MAB in all pigs, and both DAB and MAB showed lowest alpha diversity in the DiarNoInfl group. Beta diversity was significantly different between DAB and MAB as well as between diarrheal groups in both DAB and MAB. Compared to NoDiar, DiarInfl showed increased abundance of various taxa, incl. certain pathogens, in both digesta and mucus, as well as decreased digesta butyrate concentration. However, DiarNoInfl showed reduced abundance of different genera (mainly Firmicutes) compared to NoDiar, but still lower butyrate concentration.ConclusionDiversity and composition of MAB and DAB changed in diarrheal groups depending on presence/absence of colonic inflammation. We also suggest that DiarNoInfl group was at the earlier stage of diarrhea compared with DiarInfl, with a link to dysbiosis of colonic bacterial composition as well as reduced butyrate concentration, which plays a pivotal role in gut health. This could have led to diarrhea with inflammation due to a dysbiosis, associated with an increase in e.g., Escherichia-Shigella (Proteobacteria), Helicobacter (Campylobacterota), and Bifidobacterium (Actinobacteriota), which may tolerate or utilize oxygen and cause epithelial hypoxia and inflammation. The increased consumption of oxygen in epithelial mucosal layer by infiltrated neutrophils may also have added up to this hypoxia. Overall, the results confirmed that changes in DAB and MAB were associated with CCD and reduced butyrate concentration in digesta. Moreover, DAB might suffice for future community-based studies of CCD.

Characterization of diet-dependent temporal changes in circulating short-chain fatty acid concentrations: A randomized crossover dietary trial

Production of SCFAs from food is a complex and dynamic saccharolytic fermentation process mediated by both human and gut microbial factors. Knowledge of SCFA production and of the relation between SCFA profiles and dietary patterns is lacking. Temporal changes in SCFA concentrations in response to 2 contrasting diets were investigated using a novel GC-MS method. Samples were obtained from a randomized, controlled, crossover trial designed to characterize the metabolic response to 4 diets. Participants (n = 19) undertook these diets during an inpatient stay (of 72 h). Serum samples were collected 2 h after breakfast (AB), after lunch (AL), and after dinner (AD) on day 3, and a fasting sample (FA) was obtained on day 4. The 24-h urine samples were collected on day 3. In this substudy, samples from the 2 extreme diets representing a diet with high adherence to WHO healthy eating recommendations and a typical Western diet were analyzed using a bespoke GC-MS method developed to detect and quantify 10 SCFAs and precursors in serum and urine samples. Considerable interindividual variation in serum SCFA concentrations was observed across all time points, and temporal fluctuations were observed for both diets. Although the sample collection timing exerted a greater magnitude of effect on circulating SCFA concentrations, the unhealthy diet was associated with a lower concentration of acetic acid (FA: coefficient: -17.0; SE: 5.8; P-trend = 0.00615), 2-methylbutyric acid (AL: coefficient: -0.1; SE: 0.028; P-trend = 4.13 × 10-4 and AD: coefficient: -0.1; SE: 0.028; P-trend = 2.28 × 10-3), and 2-hydroxybutyric acid (FA: coefficient: -15.8; SE: 5.11; P-trend: 4.09 × 10-3). In contrast, lactic acid was significantly higher in the unhealthy diet (AL: coefficient: 750.2; SE: 315.2; P-trend = 0.024 and AD: coefficient: 1219.3; SE: 322.6; P-trend: 8.28 × 10-4). The GC-MS method allowed robust mapping of diurnal patterns in SCFA concentrations, which were affected by diet, and highlighted the importance of standardizing the timing of SCFA measurements in dietary studies. This trial was registered on the NIHR UK clinical trial gateway and with ISRCTN as ISRCTN43087333.

Evaluation of Stool Short Chain Fatty Acids Profiles in the First Year of Life With Childhood Atopy-Related Outcomes.

IntroductionShort chain fatty acids (SCFAs) are the main intestinal intermediate and end products of metabolism of dietary fibers/polyphenols by the gut microbiota. The aim of this study was to evaluate the biological implication of stool SCFA profiles determined in the first year of life on the clinical presentation of allergic outcomes in childhood.MethodsFrom the Growing Up in Singapore Toward healthy Outcomes (GUSTO) cohort, a sub-cohort of 75 participants was recruited. Scheduled questionnaire data was collected for cumulative prevalence of physician-diagnosed eczema, wheezing with the use of nebuliser, and allergen sensitization till the age of 8 years. Stool samples collected at week 3 and months 3, 6 and 12 were quantitated for 9 SCFAs using LC/MS/MS. SCFA data were grouped into lower (below the 25th) and higher (above the 75th percentiles) categories. Generalized Linear Mixed Models was employed to analyse longitudinal association between SCFAs and atopy-related outcomes.ResultsChildren with lower stool butyric acid levels (≤25th percentile) over the first 3 time points had higher odds ratio (OR) for wheezing (adjOR = 14.6), eczema (adjOR = 13.2), food sensitization (adjOR = 12.3) and combined outcomes of both wheezing and eczema (adjOR = 22.6) till age 8 years, compared to those with higher levels (≥75 percentile). Additionally, lower longitudinal levels of propionic acid (≤25th percentile) over 4 time points in first year of life was associated with recurrent wheezing (≥2 episodes) till 8 years (adjOR = 7.4) (adj p < 0.05).ConclusionOur results suggest that relatively low levels of gut SCFAs in early life are associated with increased susceptibility to atopic-related outcomes in childhood.

In vitro fermentability of a broad range of natural ingredients by fecal microbiota from lean and obese individuals: potential health benefits

The prevalence of obesity and related complications is continuously increasing while the gut microbiota might have a significant role to address this challenge. In this context, the food industry generates large amounts of residues that could be likely revalorised as functional ingredients. Hence, we evaluated the fermentability of food skins, husks, shells, trimming residues, mosses and mushrooms, which were subjected to in vitro fermentation with faecal microbiota from lean and obese adults. We demonstrated for the first time that pumpkin skin is highly fermented by human faecal microbiota showing pH-lowering effects and promoting gas and SCFA production. Furthermore, brewers’ spent grain generated an inulin-like SCFA profile after microbial fermentation, whereas Irish moss, plum skin, quinoa husk and mushrooms, including Armillaria mellea and Boletus edulis, showed high fermentation rates. Remarkably, although propionate production was significantly higher in obese individuals, the fermentability of the ingredients was similar between lean and obese conditions.

Solid State Fermentation as a Tool to Stabilize and Improve Nutritive Value of Fruit and Vegetable Discards: Effect on Nutritional Composition, In Vitro Ruminal Fermentation and Organic Matter Digestibility.

Simple SummaryA huge quantity of fruits and vegetables are wasted every year, having a negative impact in both the economy and the environment. Valorizing them as animals’ feeds would contribute to reduce feeding cost and, at the same time, would be in the interest of prevention of resource wastage and better economy of the processing plants. The aim of this study was, on the one side, to transform fruit and vegetable discards using solid state fermentation (SSF) to a stabilized product enriched in protein and, on the other side, to evaluate its suitability for ruminants feeding by determining the in vitro organic matter digestibility, fermentation characteristics and methane production of the control and the fermented product. As a result, it was found that SSF reduced the organic matter and reducing sugar content of the fermented product, while crude protein and fiber fractions were increased. In conclusion, SSF led to a stabilized feed ingredient enriched in protein, but at the expense of digestibility reduction. This research aimed to evaluate in vitro organic matter digestibility, fermentation characteristics and methane production of fruit and vegetable discards processed by solid state fermentation (SSF) by Rhizopus sp. Mixtures were composed of approximately 28% citric fruits, 35% other fruits and 37% vegetables. Fruit and vegetables were processed and fermented to obtain a stabilized product. Nutritional characterization and in vitro ruminal fermentation tests were performed to determine the effect of fungal bioconversion on digestibility, end products and gas production kinetics. Results indicate that SSF reduced organic matter and reducing sugars, while it increased crude protein and neutral detergent fiber, acid detergent fiber and neutral detergent insoluble protein. The in vitro gas production showed that SSF led to a reduction of the organic matter digestibility (p < 0.001), short chain fatty acids (SCFA; p = 0.003) and CH4 (p = 0.002). SSF reduced the gas production from the insoluble fraction (p = 0.001), without modifying the production rate (p = 0.676) or the lag time (p = 0.574). Regarding SCFA profile, SSF increased acetic (p = 0.020) and decreased propionic (p = 0.004) and butyric (p = 0.006) acids proportions, increasing acetic to propionic (p = 0.008) and acetic plus butyric to propionic (p = 0.011) ratios. SSF succeeded in obtaining a stabilized material enriched in protein, but at the expense of a reduction of protein availability and organic matter digestibility. These changes should be considered before including them in a ruminant’s rations.

Exopolysaccharides From Lactobacillus paracasei Isolated From Kefir as Potential Bioactive Compounds for Microbiota Modulation.

Microbiota coexists in true symbiosis with the host playing pivotal roles as a key element for well-being and health. Exopolysaccharides from lactic acid bacteria are an alternative as novel potential prebiotics that increase microbiota diversity. Considering this, the aim of the present work was to evaluate the capacity of the EPS produced by two L. paracasei strains isolated from kefir grains, to be metabolized in vitro by fecal microbiota producing short chain fatty acids. For this purpose, fecal samples from healthy children were inoculated in a basal medium with EPS and incubated in anaerobiosis at 37°C for 24, 48, and 72 h. DGGE profiles and the production of SCFA after fermentation were analyzed. Additionally, three selected samples were sequenced by mass sequencing analysis using Ion Torrent PGM. EPS produced by L. paracasei CIDCA 8339 (EPS8339) and CIDCA 83124 (EPS83124) are metabolized by fecal microbiota producing a significant increase in SCFA. EPS8339 fermentation led to an increment of propionate and butyrate, while fermentation of EPS83124 increased butyrate levels. Both EPS led to a profile of SCFA different from the ones obtained with inulin or glucose fermentation. DGGE profiles of 72 h fermentation demonstrated that both EPS showed a different band profile when compared to the controls; EPS profiles grouped in a cluster that have only 65% similarity with glucose or inulin profiles. Mass sequencing analysis demonstrated that the fermentation of EPS8339 leads to an increase in the proportion of the genera Victivallis, Acidaminococcus and Comamonas and a significant drop in the proportion of enterobacteria. In the same direction, the fermentation of EPS83124 also resulted in a marked reduction of Enterobacteriaceae with a significant increase in the genus Comamonas. It was observed that the changes in fecal microbiota and SCFA profile exerted by both polymers are different probably due to differences in their structural characteristics. It can be concluded that EPS synthesized by both L. paracasei strains, could be potentially used as bioactive compound that modify the microbiota increasing the production of propionic and butyric acid, two metabolites highly associated with beneficial effects both at the gastrointestinal and extra-intestinal level.

The Fermentation Process Improves the Nutritional Value of Rapeseed Cake for Turkeys—Effects on Performance, Gut Bacterial Population and Its Fermentative Activity

Simple SummaryRapeseed cake (RC) could be valuable raw material in turkey diets, but its wide use is limited by the presence of anti-nutritional factors that are detrimental to gut function. The fermentation process contributes to the degradation of over 80% of carbohydrates, 30% of lignin, and 45% of total glucosinolates in RC, which are harmful to birds. Our research showed that fermentation can improve the nutritional value of RC, enabling good performance and maintainenance of a healthy gut in birds. Therefore, FRC appears to be highly promising in commercial turkey nutrition.This experiment investigated the potential inclusion of fermented rapeseed cake (FRC) in turkey diets. The turkeys received diets either not supplemented (C) or supplemented with raw rapeseed cake (RRC) or FRC at 150 g/kg diet. In comparison with RRC, turkeys receiving FRC achieved significantly higher final BW comparable with that noted in the control group. The dietary inclusion of FRC increased the concentrations of propionic and valeric acid in the cecal digesta compared with the control group, and increased the proportion of butyric acid in SCFA profile compared with RRC group. The activities of glycolytic bacterial enzymes in the cecal digesta, were lowest in turkeys fed FRC. Experimental diets did not cause a shift in the relative abundances of the main bacterial phyla or orders in the cecal digesta. FRC increased the abundance of Bacteroidaceae at the family level, but decreased the abundance of Lactobacillus at the genus level compared with birds fed RRC. In conclusion, the dietary inclusion of FRC at 150 g/kg did not compromise bird performance, did not excessively stimulate bacterial activity, and did not cause shifts in the bacterial composition in the cecum. Actually, FCR exerted several beneficial effects that contributed to maintaining gut health in turkeys, which points to its advantage over RRC.

P315 Short-chain fatty acid profile in inflammatory bowel disease

Abstract Background Short-chain fatty acids (SCFAs) are products of colonic bacterial degradation of dietary fibre. They are important in the colon, affecting the morphology and function of colonocytes. SCFAs consist of a molecule with one to six carbons, of which acetate, propionate and butyrate are the most abundant. In recent decades, it has become apparent that SCFAs can play a key role in the prevention and treatment of metabolic syndrome, intestinal disorders and certain cancers. Crohn’s disease (CD) and ulcerative colitis (UC) are characterised by recurrent chronic intestinal inflammation, probably due to an inadequate immune response coupled with intestinal microbiota imbalance. The aim of this study was to evaluate the profile of SCFA in patients with UC and CD, and compared with non-ill individuals. Methods The individuals were divided into three groups: RCU, CD and control, the faeces were donated by them, and the SCFAs were measured by chromatographic analysis using a Thermo Scientific GC-MS coupled to a Thermo ISQ 230ST mass detector. All results are expressed as mean ± SEM. Results It was possible to observe a different SCFAs profile between individuals with CD and UC and control where acetate and propionate levels in patients with UC and CD were higher than in non-sick individuals and butyrate with lower levels in individuals with CD and RCU (Graph 1). SCFAs have anti-inflammatory capabilities and also a preferred energy source for colon epithelial cells, as well as lowering the pH of the colon and inhibiting the growth of pathogenic organisms. Dysbiosis decreases butyrate concentrations, which may result in nutrient deficiency at the epithelial level, altering immune responses, as well as acting directly as an anti-inflammatory agent by disabling the NFκB pathway, with a consequent decrease in inflammatory cytokine synthesis. Conclusion Thus, the reported results have implications for various physiological and pathological conditions in inflammatory bowel diseases, especially with respect to butyrate and the production of inflammatory mediators, and partly explain the beneficial effects attributed to this fatty acid in the treatment of inflammatory and inflammatory diseases support the realisation of new studies aimed at the development of therapeutic alternatives to the use of conventional anti-inflammatory drugs.

Prebiotics Fructo-, Galacto-, and Mannan-Oligosaccharide Do Not Protect against 5-Fluorouracil–Induced Intestinal Mucositis in Rats

Prebiotics selectively stimulate the growth of beneficial bacteria within the gastrointestinal tract, and have been investigated in human and animal studies for their capacity to improve intestinal health. We investigated the prebiotics fructo-oligosaccharide (FOS), galacto-oligosaccharide (GOS), and mannan-oligosaccharide (MOS) for their potential to alleviate intestinal damage in rats. Female Dark Agouti rats (6-8 wk old, 110-150 g) were allocated to 1 of the following treatment groups (n=8/group): saline/water, saline/FOS, saline/GOS, saline/MOS, 5-fluorouracil (5FU)/water, 5FU/FOS, 5FU/GOS, and 5FU/MOS. Rats were pretreated with either 5% GOS, MOS, or FOS or vehicle (water) from day -12 to day 0. On day 0, rats received a single intraperitoneal injection of saline or 5FU. Metabolic data were recorded daily and all rats were killed on day 3. Histopathology was quantified in hematoxylin and eosin-stained sections. Intestinal sucrase and myeloperoxidase activity were quantified by biochemical assay. Fecal SCFAs-acetic, propionic, and butyric acid-were also measured. Statistical analysis was by repeated-measures, 2-factor ANOVA or Kruskal-Wallis and Mann-Whitney U test; P<0.05 was considered statistically significant. Body weight was significantly decreased in all treatment groups after 5FU injection, with no change in body weight observed in any prebiotic treatment group. Total food intake was lower by ≥7% in the GOS treatment group pre-5FU than in all other groups (P<0.05). Ileal villus height was 18% higher in GOS-treated rats pre-5FU than in respective water controls (P<0.05). Jejunal and ileal villus height and crypt depth were significantly decreased in all treatment groups after 5FU injection, with no prebiotic effect observed. SCFAs were differentially increased in prebiotic treatment groups compared with water-only controls (P<0.05). FOS, GOS, and MOS have differential effects in modifying small intestinal pathology and SCFA profiles in rats with healthy and damaged small intestinal mucosa.

In vitro prebiotic potential of agricultural by-products on intestinal fermentation, gut barrier and inflammatory status of piglets.

The inclusion of fibre-rich ingredients in diets is one possible strategy to enhance intestinal fermentation and positively impact gut ecology, barrier and immunity. Nowadays, inulin-type fructans are used as prebiotics in the feed of piglets to manipulate gut ecology for health purposes. Likewise, some by-products could be considered as sustainable and inexpensive ingredients to reduce gut disorders at weaning. In the present study, chicory root and pulp, citrus pulp, rye bran and soya hulls were tested in a three-step in vitro model of the piglet's gastro-intestinal tract combining a pepsin-pancreatin hydrolysis (digestion), a dialysis step using cellulose membranes (absorption) and a colonic batch fermentation (fermentation). The fermentation kinetics, SCFA and microbiota profiles in the fermentation broth were assessed as indicators of prebiotic activity and compared with the ones of inulin. The immunomodulatory effects of fermentation supernatant (FS) were investigated in cultured intestinal porcine epithelial cells (IPEC-J2) by high-throughput quantitative PCR. Chicory root displayed a rapid and extensive fermentation and induced the second highest butyrate ratio after inulin. Citrus pulp demonstrated high acetate ratios and induced elevated Clostridium clusters IV and XIVa levels. Chicory root and pulp FS promoted the intestinal barrier integrity with up-regulated tight and adherens junction gene expressions in comparison with inulin FS. Chicory pulp FS exerted anti-inflammatory effects in cultured IPEC-J2. The novel approach combining an in vitro fermentation model with IPEC-J2 cells highlighted that both chicory root and pulp appear to be promising ingredients and should be considered to promote intestinal health at weaning.

In-Vitro Prebiotic Analysis of Microbiota Accessible Pectic Polysaccharides.

Pectin is a diverse polysaccharide comprising of polygalacturonic acid, rhamnogalactouronan, and neutral polysaccharides (arabinan and arabinogalactan) as side chains or branches. They are resistant to salivary amylase and gastric juice. In the present study the prebiotic potentials of different pectic derived substrates were performed by using intestinal cultures and measuring the growth, change in pH and short chain fatty acid, (SCFA) production. Arabinan, arabinogalactan, and rhamnogalacturonan were fermented by probiotic bacteria such as Bifidobacterium breve, Bi. longum, Bi. bifidum, and Bacteroides thetaiotaomicron (commensal bacteria). While, polygalacturonic acid was fermented by Ba. fragilis, Ba. thetaiotaomicron, and Ba. uniformis. All the screened bacteria significantly decreased the pH from 7 to 5 (pH difference of ≈ 2), which clearly indicated that the above bacteria produced the enzymes necessary for the digestion of the pectic derived polysaccharides. The SCFA profiles of the above screened bacteria clearly demonstrated the production of lactate, acetate and propionate which are the key metabolites involved in maintaining gut health and prevention of several intestinal diseases. Thus, pectic polysaccharides hold potential application in food industry as prebiotic ingredients or dietary fibers.

The Role of Vagal Free Fatty Acid Receptor 3 (FFAR3) in FMT Improvement of Glucose Homeostasis

More than 100 million Americans are obese, representing over 35% of the Unites States population. Western diet (WD) is a major cause of the obesity epidemic, and it increases prevalence of many other comorbidities, particularly Type II Diabetes. The western diet consisting of high saturated fat, high cholesterol, high sugar, and heavily processed foods robustly disrupts the composition of the gut bacteria. The microbiome ferments non‐digestible fibers to release short chain fatty acids (SCFA's) which serve as major energy substrates as well as potent signaling molecules through free fatty acid receptors 2 (FFAR2) and 3 (FFAR3). Gut dysbiosis has been associated with changes in SCFA levels that correlate with obesity in humans and mice. The SCFA's propionate and butyrate bind FFAR3 with high affinity, and FFAR3 signaling has been shown to modulate energy expenditure through sympathetic nervous system activation. The role of FFAR3 in the parasympathetic nervous system control of glucose homeostasis is currently unexplored. The vagus nerve primarily consists of sensory afferents which relay specific nutrient information from the periphery to the brain. Several studies have shown that disruption of the vagus nerve eliminates effects of the microbiome and SCFA's altering metabolism, yet the molecular mechanisms of this communication remain largely unknown. We have found that western diet alters SCFA profiles, which have been shown to modulate glucose homoeostasis and energy expenditure in a vagus nerve dependent manner. Furthermore, FFAR3 expression was reduced in the cell bodies of the vagus nerve after WD feeding. We confirmed previous findings that fecal microbiome transplantation (FMT) from lean mice reversed some WD‐induced changes to bacterial composition, plasma SCFA's, and glucose homeostasis. Our data suggest that altered FFAR3 signaling in vagal afferent neurons may contribute to some of the metabolic changes seen upon western diet feeding, and that gut microbes transplanted from lean to obese mice may improve glucose homeostasis through SCFA's binding FFAR3 in the vagus nerve.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Sialyllactose and Galactooligosaccharides Promote Epithelial Barrier Functioning and Distinctly Modulate Microbiota Composition and Short Chain Fatty Acid Production In Vitro.

Human milk oligosaccharides (HMO) and prebiotic oligosaccharides are proposed to confer several health benefits to the infant. They shape the microbiota, have anti-inflammatory properties, and support epithelial barrier functioning. However, in order to select the best oligosaccharides for inclusion in infant formulas, there is a need to increase our understanding of the specific effects of HMO and prebiotics on the host immune system. Therefore, we investigated the effects of the HMO sialyllactose (SL), and galactooligosaccharides (GOS) on epithelial barrier functioning, microbiota composition, and SCFA production. The effect of GOS and SL on epithelial barrier functioning and microbiota composition was investigated using in vitro models. Epithelial barrier function was investigated by transcriptome analysis of fully polarized Caco-2 cells exposed for 6 h to SL or GOS. In addition, epithelial cell growth, alkaline phosphatase production, and re-epithelization was studied. Further, we investigated the effect of SL and GOS on microbiota composition and SCFA production using in vitro fecal batch cultures. Transcriptome analysis showed that SL and GOS both induced pathways that regulate cell cycle control. This gene-expression profile translated to a phenotype of halted proliferation and included the induction of alkaline phosphatase activity, a marker of epithelial cell differentiation. SL and GOS also promoted re-epithelialization in an in vitro epithelial wound repair assay. SL and GOS did show distinct modulation of microbiota composition, promoting the outgrowth of Bacteroides and bifidobacteria, respectively, which resulted in distinct changes in SCFA production profiles. Our results show that SL and GOS can both modulate epithelial barrier function by inducing differentiation and epithelial wound repair, but differentially promote the growth of specific genera in the microbiota, which is associated with differential changes in SCFA profiles.

Understanding the prebiotic potential of different dietary fibers using an in vitro continuous adult fermentation model (PolyFermS)

Consumption of fermentable dietary fibers (DFs), which can induce growth and/or activity of specific beneficial populations, is suggested a promising strategy to modulate the gut microbiota and restore health in microbiota-linked diseases. Until today, inulin and fructo-oligosaccharides (FOS) are the best studied DFs, while little is known about the gut microbiota-modulating effects of β-glucan, α-galactooligosaccharide (α-GOS) and xylo-oligosaccharide (XOS). Here, we used three continuous in vitro fermentation PolyFermS model to study the modulating effect of these DFs on two distinct human adult proximal colon microbiota, independently from the host. Supplementation of DFs, equivalent to a 9 g daily intake, induced a consistent metabolic response depending on the donor microbiota. Irrespective to the DF supplemented, the Bacteroidaceae-Ruminococcaceae dominated microbiota produced more butyrate (up to 96%), while the Prevotellaceae-Ruminococcaceae dominated microbiota produced more propionate (up to 40%). Changes in abundance of specific bacterial taxa upon DF supplementation explained the observed changes in short-chain fatty acid profiles. Our data suggest that the metabolic profile of SCFA profile may be the most suitable and robust read-out to characterize microbiota-modulating effects of a DF and highlights importance to understand the inter-individual response to a prebiotic treatment for mechanistic understanding and human application.

Effects of monobutyrin and tributyrin on liver lipid profile, caecal microbiota composition and SCFA in high-fat diet-fed rats.

Butyric acid has been shown to have suppressive effects on inflammation and diseases related to the intestinal tract. The aim of the present study was to investigate whether supplementation of two glycerol esters, monobutyrin (MB) and tributyrin (TB), would reach the hindgut of rats, thus having an effect on the caecal profile of SCFA, microbiota composition and some risk markers associated with chronic inflammation. For this purpose, rats were fed high-fat diets after adding MB (1 and 5g/kg) and TB (5g/kg) to a diet without any supplementation (high-fat control; HFC). A low-fat (LF) diet was also included. In the liver, total cholesterol concentrations, LDL-cholesterol concentrations, LDL:HDL ratio, and succinic acid concentrations were reduced in rats given the MB and TB (5g/kg) diets, compared with the group fed the HFC diet. These effects were more pronounced in MB than TB groups as also expressed by down-regulation of the gene Cyp8b1. The composition of the caecal microbiota in rats fed MB and TB was separated from the group fed the HFC diet, and also the LF diet, as evidenced by the absence of the phylum TM7 and reduced abundance of the genera Dorea (similar to LF-fed rats) and rc4-4. Notably, the caecal abundance of Mucispirillum was markedly increased in the MB group compared with the HFC group. The results suggest that dietary supplementation of MB and TB can be used to counteract disturbances associated with a HFC diet, by altering the gut microbiota, and decreasing liver lipids and succinic acid concentrations.