Fecal Inoculum Research Articles

Evaluating the potential prebiotic effects of umbu-cajá (Spondias spp.) fruit processing by-product flour on the human intestinal microbiota

This study evaluated the potential prebiotic properties of umbu-cajá (Spondias spp.) fruit processing by-product flour (UCF) on the human intestinal microbiota by monitoring the relative abundance of specific intestinal bacterial populations, microbial metabolic behavior, and antioxidant activity during 48 h of in vitro colonic fermentation using fecal inoculum donated from healthy adult individuals. UCF had a high insoluble fiber content and flavonols, especially quercetin-3-O-glucoside, as the most prevalent phenolic compounds, besides showing antioxidant activity. Medium with UCF increased the relative abundance of Lactobacillus spp./Enterococcus spp., Bifidobacterium spp., and Clostridium histolyticum during the colonic fermentation and decreased the relative abundance of Bacteroides spp./Prevotella spp. and Eubacterium rectale/Clostridium coccoides. Medium with UCF maintained a higher relative abundance of Ruminococcus albus/R. flavefaciens than negative control during the colonic fermentation. UCF stimulated intense metabolic activity of the intestinal microbiota during the colonic fermentation, evidenced by decreased pH values, sugar consumption, and production of several metabolites linked to host health promotion, concomitant with high antioxidant activity. The results characterize UCF as a candidate for prebiotic use and a novel value-added circular ingredient to functionalize foods and nutraceuticals.

292 In vitro fermentation characteristics of dietary fibers using fecal inocula from dogs treated with metronidazole

292 In vitro fermentation characteristics of dietary fibers using fecal inocula from dogs treated with metronidazole

Metatranscriptomic analysis indicates prebiotic effect of isomalto/malto-polysaccharides on human colonic microbiota in-vitro

Isomalto/malto-polysaccharides (IMMPs) are a novel type of soluble dietary fibres with a prebiotic potential promoting growth of beneficial microbes in the gut. However, the mode of action of IMMPs remains unknown. Previous studies on IMMPs showed an increase in total bacteria, especially lactobacilli, and higher production of short chain fatty acids (SCFA) when IMMPs were fed to rats or used during in vitro fermentation. Here we used metatranscriptomics to investigate how IMMPs with different amounts of α − (1 → 6) glycosidic linkages affected microbial function during incubation with human fecal inoculum. We showed that active microbial community dynamics during fermentation varied depending on the type of IMMP used and that the observed changes were reflected in the community gene expression profiles. Based on metatranscriptome analysis, members of Bacteroides, Lactobacillus and Bifidobacterium were the predominant degraders of IMMPs, and the increased gene expression in these bacteria correlated with high amounts of α − (1 → 6) glycosidic linkages. We also noted an increase in relative abundance of these bacteria and an activation of pathways involved in SCFA synthesis. Our findings could provide a baseline for more targeted approaches in designing prebiotics for specific bacteria and to achieve more controlled modulation of microbial activity towards desired health outcomes.

Non-invasive VOCs detection to monitor the gut microbiota metabolism in-vitro

This work implemented a non-invasive volatile organic compounds (VOCs) monitoring approach to study how food components are metabolised by the gut microbiota in-vitro. The fermentability of a model food matrix rich in dietary fibre (oat bran), and a pure prebiotic (inulin), added to a minimal gut medium was compared by looking at global changes in the volatilome. The substrates were incubated with a stabilised human faecal inoculum over a 24-h period, and VOCs were monitored without interfering with biological processes. The fermentation was performed in nitrogen-filled vials, with controlled temperature, and tracked by automated headspace-solid-phase microextraction coupled with gas chromatography–mass spectrometry. To understand the molecular patterns over time, we applied a multivariate longitudinal statistical framework: repeated measurements—ANOVA simultaneous component analysis. The methodology was able to discriminate the studied groups by looking at VOCs temporal profiles. The volatilome showed a time-dependency that was more distinct after 12 h. Short to medium-chain fatty acids showed increased peak intensities, mainly for oat bran and for inulin, but with different kinetics. At the same time, alcohols, aldehydes, and esters showed distinct trends with discriminatory power. The proposed approach can be applied to study the intertwined pathways of gut microbiota food components interaction in-vitro.

AG1®, a Novel Synbiotic, Maintains Gut Barrier Function following Inflammatory Challenge in a Caco-2/THP1-Blue™ Co-Culture Model.

Nutritional interventions to reduce gastrointestinal (GI) permeability are of significant interest to physically active adults and those experiencing chronic health conditions. This in vitro study was designed to assess the impact of AG1, a novel synbiotic, on GI permeability following an inflammatory challenge. Interventions [AG1 (vitamins/minerals, pre-/probiotics, and phytonutrients) and control (control medium)] were fed separately into a human GI tract model (stomach, small intestine, and colon). In the colonic phase, the GI contents were combined with fecal inocula from three healthy human donors. GI permeability was evaluated with transepithelial electrical resistance (TEER) in a Caco-2 (apical)/THP1-Blue™ (basolateral) co-culture model. The apical side received sodium butyrate (positive control) or Caco-2 complete medium (negative control) during baseline testing. In the 24 h experiment, the apical side received colonic simulation isolates from the GI model, and the basolateral side was treated with Caco-2 complete medium, then 6 h treatment with lipopolysaccharide. TEER was assessed at 0 h and 24 h, and inflammatory markers were measured at 30 h in triplicate. Paired samples t-tests were used to evaluate endpoint mean difference (MD) for AG1 vs. control. TEER was higher for AG1 (mean ± SD: 99.89 ± 1.32%) vs. control (mean ± SD: 92.87 ± 1.22%) following activated THP1-induced damage [MD: 7.0% (p < 0.05)]. AG1 maintained TEER similar to the level of the negative control [-0.1% (p = 0.02)]. No differences in inflammatory markers were observed. These in vitro data suggest that acute supplementation with AG1 might stimulate protective effects on GI permeability. These changes may be driven by SCFA production due to the pre-/probiotic properties of AG1, but more research is needed.

In vitro digestion and fermentation of animal-derived fermentable substrates using canine and feline faecal inoculum

Abstract There is increasing interest in understanding the fermentative benefits of animal-derived fermentable substrates (ADFS) in pet foods. While previous research has assessed various ADFS using faecal inoculum derived from the cat, there is no published literature available for the dog. Additionally, very little is understood of the fermentation profiles of ADFS, such as skin and bone. Therefore, faeces were collected from a cohort of cats and dogs fed a complete and balanced high-protein diet and a selection of substrates were analysed in this study. Individual ADFS (tendon, bone cake, mechanically deboned meat (MDM), corium and hydrolysed collagen (a positive control)) were digested in vitro, followed by fermentation using either canine or feline faecal inoculum. Concentrations of butyrate, indole and ammonia were determined after 24 h of fermentation. Regardless of whether cat or dog faecal inoculum was used, fermentation of hydrolysed collagen produced the highest (P &lt; 0.01) concentrations of butyrate and ammonia and the lowest concentrations of indole. For the other substrates, there were differences in the fermentation profiles between the canine and feline inocula. During the feline faecal fermentations, in comparison to the other substrates bone cake produced high (P &lt; 0.05) butyrate concentrations, whereas in the dog faecal fermentations, MDM resulted in high (P &lt; 0.05) butyrate concentrations. In conclusion, ADFS from different alternative co-products lead to different fermentation products, providing valuable information which may be considered in canine and feline dietary formulations.

Enrichment and characterization of human-associated mucin-degrading microbial consortia by sequential passage.

Mucin is a glycoprotein secreted throughout the mammalian gastrointestinal tract that can support endogenous microorganisms in the absence of complex polysaccharides. While several mucin-degrading bacteria have been identified, the interindividual differences in microbial communities capable of metabolizing this complex polymer are not well described. To determine whether community assembly on mucin is deterministic across individuals or whether taxonomically distinct but functionally similar mucin-degrading communities are selected across fecal inocula, we used a 10-day in vitro sequential batch culture fermentation from three human donors with mucin as the sole carbon source. For each donor, 16S rRNA gene amplicon sequencing was used to characterize microbial community succession, and the short-chain fatty acid profile was determined from the final community. All three communities reached a steady-state by day 7 in which the community composition stabilized. Taxonomic comparisons amongst communities revealed that one of the final communities had Desulfovibrio, another had Akkermansia, and all three shared other members, such as Bacteroides. Metabolic output differences were most notable for one of the donor's communities, with significantly less production of acetate and propionate than the other two communities. These findings demonstrate the feasibility of developing stable mucin-degrading communities with shared and unique taxa. Furthermore, the mechanisms and efficiencies of mucin degradation across individuals are important for understanding how this community-level process impacts human health.

15 Effects of post-MSC DDGS inclusion in gestation diets compared with other fiber sources on in vitro gas and volatile fatty acid production

Abstract In vitro fermentation has been used to characterize hindgut fermentation of fibrous ingredients fed to grower pigs. However, there is little to no work investigating hindgut fermentation of fibrous feedtuffs in gestating sows, who have greater fermentative capacity compared with grower pigs. The objective of this study was to characterize hindgut gas production and volatile fatty acids (VFA) following in vitro fermentation using fecal inoculum collected from the sows when fed either post-protein separation dried-distiller’s grains with solubles (post-MSC DDGS), soyhulls, or sugar beet pulp. Pregnant sows (n = 22; parity 0 to 1; body weigh = 190 ± 10 kg) were allocated to 1 of 6 dietary treatments in a crossover design with two collection periods (period 1 from d 40-59 of gestation and period 2 from d 60-79 of gestation) to attain at least 7 observations per treatment. Two of the six diets were practical corn-soybean meal-based gestation diets with either soyhulls (17.5% inclusion) or post-MSC DDGS (15% inclusion). The remaining four diets included a corn-basal diet and the basal diet plus 30% inclusion of post-MSC DDGS, soyhulls, or sugar beet pulp. Sows were fed experimental diets (2.2 kg/d in a single feeding) in two 20 d collection periods. For in vitro fermentation, fresh fecal samples were collected on d 16 of each collection period, prepared as inoculum, and subjected to the gas production measurement technique through in vitro fermentation. Supernatant residue from the in vitro fermentation was subjected to VFA analysis by gas chromatography. Data were analyzed with ANOVA and fixed to a mixed linear model using treatment as the main effect and collection period as a random variable. Total gas production of the practical diet containing post-MSC DDGS diet was 132 ml/g less (P = 0.03) than the practical soyhull diet. While the total concentration of total VFA did not differ between practical diets, the concentration of isovalerate tended to be greater for the post-MSC DDGS diet compared with the soyhull diet (P = 0.09). Total gas production was 140 and 375 mg/L less for the post-MSC DDGS than soyhulls and sugar beet pulp, respectively (P &amp;lt; 0.01). The concentrations of butyrate and isovalerate (relative to total VFA) were approximately 7 and 2%, respectively, greater for the post-MSC DDGS than in the soyhulls or sugar beet pulp fermentation residues (P &amp;lt; 0.05). Concentrations of other VFA relative to total VFA did not differ between fiber sources. In conclusion, post-MSC DDGS can contribute to sustainable swine production when fed to gestating by lowering total gas production and being a source of volatile fatty acids.

A systematic study of sheep faeces to be used as an alternative inoculum source in the Hohenheim gas test

ABSTRACT In vitro studies of gas production (GP) using rumen fluid obtained from rumen-cannulated animals are common in feed evaluation for ruminants. The objective was to compare sheep faeces with rumen fluid from dairy cows as inocula in the Hohenheim gas test (HGT) using a large number of different-quality feeds. It then was evaluated whether GP obtained by using rumen inoculum (RI-GP) can be estimated from GP using faecal inoculum (FI-GP). Ninety feeds for ruminants including roughages (n = 27), total mixed rations (n = 6), commercial compound feeds for dairy cows (n = 18), cereal grains (n = 10), energy-rich by-products (n = 5), legume grains (n = 6), oilseed meals (n = 6), and other protein-rich feeds (n = 12) were incubated with either rumen inoculum (RI) or faecal inoculum (FI), and GP was recorded at 2, 4, 8, 12, 24, 48, and 72 h. Nonlinear equations were fitted to the GP data to determine GP kinetic parameters. Correlations between RI-GP and FI-GP were calculated for different time points. Linear regressions were calculated to estimate RI-GP from FI-GP using the entire data set or subsets of roughages and concentrates. GP kinetic parameters were lower for all feed categories when incubated with FI instead of RI. On average of all feed samples, the potential GP was 9 mL/200 mg dry matter (DM) lower and the rate of GP was 3.1%/h lower with FI than RI. The estimation of kinetic data for FI included a lag phase of 1.51 h on average, whereas no lag phase was estimated for RI. Estimated parameters indicated an overall lower fermentation activity of FI than RI. However, there was a very similar progression of GP curves of RI-GP and FI-GP. RI-GP after 24 h was significantly correlated with FI-GP in the time period of 24 to 60 h (r = 0.973 – 0.982, including all feed samples). Linear regression analysis showed that RI-GP after 24 h can be estimated from FI-GP after 24 h (Slope = 1.02, R2 = 0.97) and 48 h (Slope = 1.1, R2 = 0.97) including the complete data set. Splitting the data set into roughages and concentrate feeds did not lead to distinctly higher estimation accuracy. In conclusion, sheep faeces can be used as an alternative inoculum for studying a wide range of different-quality feedstuffs. The standard gas test may be modified without the need for rumen-cannulated animals.

Complementary feeding in infants with cystic fibrosis: In vitro nutrient digestibility and impact on colonic microbiota

The aim was assessing macronutrient digestibility of common purees in the context of children with cystic fibrosis (CF) starting complementary feeding. A static in vitro digestion model was set up reproducing the conditions of infants with CF and healthy controls. Egg, chicken, infant formula and infant cereal were subjected to digestion individually and in combination with other ingredients. The resulting digesta was subjected to colonic fermentation with the faecal inoculum of an exclusively breast-fed infant with CF. Results showed overall low proteolysis and lipolysis, with positive interaction between egg and chicken with potato, vegetables, and olive oil. Contrarily, the combination of infant cereals and formula had a negative interaction. Notably, the dose of pancreatic enzymes had no effect on lipolysis. Purees rather than in individual foods impacted colonic microbiota, by reducing Bifidobacterium while increasing Veillonella. This study could guide future dietary recommendations for complementary feeding in infants with CF.

Cascade Microbial Metabolism of Ferulic Acid In Vitro Fermented by the Human Fecal Inoculum.

Ferulic acid (FA), predominantly existing in most cereals, can modulate the gut microbiome, but the influences of its metabolites on the microbial population and FA-transforming microorganisms are still unclear. In this study, FA and its potential phenolic metabolites were fermented in vitro for 24 h with the human fecal inoculum. A comparable short chain fatty acid (SCFA) production trend was observed in the presence and absence of substrates, suggesting limited contribution of FA mechanism to SCFA formation. Dihydroferulic acid, 3-(3,4-dihydroxyphenyl)propionic acid, and 3-(3-hydroxyphenyl)propionic acid were ascertained to be successive metabolites of FA, by tracking the intermediate variation. FA remarkably promoted the absolute abundances of total bacteria, while different metabolites affected bacterial growth of selective genera. Specific genera were identified as quantitatively correlating to the content of FA and its metabolites. Ultimately, FA-mediated gut microbiota modulation involves both the action of metabolizing microbes and the regulation effects of metabolites on bacterial growth.

Meta-analysis of the Microbial Diversity Cultured in Bioreactors Simulating the Gut Microbiome

Understanding the intricate ecological interactions within the gut microbiome and unravelling its impact on human health is a challenging task. Bioreactors are valuable tools that have contributed to our understanding of gut microbial ecology. However, there is a lack of studies describing and comparing the microbial diversity cultivated in these models. This knowledge is crucial for refining current models to reflect the gastrointestinal microbiome accurately. In this study, we analysed the microbial diversity of 1512 samples from 18 studies available in public repositories that employed cultures performed in batches and various bioreactor models to cultivate faecal microbiota. Community structure comparison between samples using t-distributed stochastic neighbour embedding and the Hellinger distance revealed a high variation between projects. The main driver of these differences was the inter-individual variation between the donor faecal inocula. Moreover, there was no overlap in the structure of the microbial communities between studies using the same bioreactor platform. In addition, α-diversity analysis using Hill numbers showed that highly complex bioreactors did not exhibit higher diversities than simpler designs. However, analyses of five projects in which the samples from the faecal inoculum were also provided revealed an amplicon sequence variants enrichment in bioreactors compared to the inoculum. Finally, a comparative analysis of the taxonomy of the families detected in the projects and the GMRepo database revealed bacterial families exclusively found in the bioreactor models. These findings highlight the potential of bioreactors to enrich low-abundance microorganisms from faecal samples, contributing to uncovering the gut microbial “dark matter”.

The effect of in vitro simulated colonic pH gradients on microbial activity and metabolite production using common prebiotics as substrates

BackgroundThe interplay between gut microbiota (GM) and the metabolization of dietary components leading to the production of short-chain fatty acids (SCFAs) is affected by a range of factors including colonic pH and carbohydrate source. However, there is still only limited knowledge on how the GM activity and metabolite production in the gastrointestinal tract could be influenced by pH and the pH gradient increases along the colon.ResultsHere we investigate the effect of pH gradients corresponding to levels typically found in the colon on GM composition and metabolite production using substrates inulin, lactose, galactooligosaccharides (GOS), and fructooligosaccharide (FOS) in an in vitro colon setup. We investigated 3 different pH regimes (low, 5.2 increasing to 6.4; medium, 5.6 increasing to 6.8 and high, 6.0 increasing to 7.2) for each fecal inoculum and found that colonic pH gradients significantly influenced in vitro simulated GM structure, but the influence of fecal donor and substrate was more pronounced. Low pH regimes strongly influenced GM with the decreased relative abundance of Bacteroides spp. and increased Bifidobacterium spp. Higher in vitro simulated colonic pH promoted the production of SCFAs in a donor- and substrate-dependent manner. The butyrate producer Butyricimonas was enriched at higher pH conditions, where also butyrate production was increased for inulin. The relative abundance of Phascolarctobacterium, Bacteroides, and Rikenellaceae also increased at higher colonic pH, which was accompanied by increased production of propionate with GOS and FOS as substrates.ConclusionsTogether, our results show that colonic substrates such as dietary fibres influence GM composition and metabolite production, not only by being selectively utilized by specific microbes, but also because of their SCFA production, which in turn also influences colonic pH and overall GM composition and activity. Our work provides details about the effect of the gradients of rising pH from the proximal to distal colon on fermenting dietary substrates in vitro and highlights the importance of considering pH in GM research.

Combined dilute alkali and milling process enhances the functionality and gut microbiota fermentability of insoluble corn fiber

In this study, we developed a process combining dilute alkali (NaOH or NaHCO3) and physical (disk milling and/or ball milling) treatments to improve the functionality and fermentability of corn fiber. The results showed that combining chemical with physical processes greatly improved the functionality and fermentability of corn fiber. Corn fiber treated with NaOH followed by disk milling (NaOH-DM-CF) had the highest water retention (19.5 g/g), water swelling (38.8 mL/g), and oil holding (15.5 g/g) capacities. Moreover, NaOH-DM-CF produced the largest amount (42.9 mM) of short-chain fatty acid (SCFA) during the 24-hr in vitro fermentation using porcine fecal inoculum. In addition, in vitro fermentation of NaOH-DM-CF led to a targeted microbial shifting to Prevotella (genus level), aligning with a higher fraction of propionic acid. The outstanding functionality and fermentability of NaOH-DM-CF were attributed to its thin and loose structure, decreased ester linkages and acetyl groups, and enriched structural carbohydrate exposure.

In vitro fermentation characteristics of different types of fiber-rich ingredients by pig fecal inoculum.

Dietary fibers with varying physicochemical properties have different fermentation characteristics, which may differently impact host health. The present study aimed to determine the fermentation characteristics including gas production kinetics, short-chain fatty acids (SCFAs) production and microbial composition of different fibrous ingredients using in vitro fermentation by fecal microbiota. Sugar beet pule (SBP), wheat bran (WB), dried corn distillers grains with solubles (DDGS), rice bran (RB) and alfalfa meal (AM) were selected to fermentation in vitro for 36 h. The results showed that SBP had the greatest gas production. SBP had the highest in vitro dry matter fermentability (IVDMF) and production of acetate, propionate and total SCFAs, followed by WB, which were all greater than DDGS, AM and RB. The alpha-diversity was higher in the DDGS, AM and RB groups than in the WB and SBP groups. Differences in microbial community composition were observed among groups. The relative abundance of Treponema was highest in WB group. RB group showed lower Prevotella abundance than other groups but had higher Succinivibrio abundance. Interestingly, the Lactobacillus reached the highest abundances in the DDGS group. Correlation analysis indicated that the relative abundance of Treponema and Prevotella was positively associated with the gas production, IVDMF and SCFAs, whereas norank_f_Muribaculaceae, Rikenellaceae_RC9_gut_group, Lysinibacillus and Succinivibrio were the opposite. Collectively, WB and SBP were fermented rapidly by fecal microbiota compared to DDGS, AM and RB. Different fiber sources have different fiber compositions and fermentation properties that affect the microbial compositins and SCFAs production. © 2024 Society of Chemical Industry.

M-Batches to Simulate Luminal and Mucosal Human Gut Microbial Ecosystems: A Case Study of the Effects of Coffee and Green Tea.

Gastrointestinal simulations in vitro have only limited approaches to analyze the microbial communities inhabiting the mucosal compartment. Understanding and differentiating gut microbial ecosystems is crucial for a more comprehensive and accurate representation of the gut microbiome and its interactions with the host. Herein is suggested, in a short-term and static set-up (named "M-batches"), the analysis of mucosal and luminal populations of inhabitants of the human colon. After varying several parameters, such as the fermentation volume and the fecal inoculum (single or pool), only minor differences in microbial composition and metabolic production were identified. However, the pool created with feces from five donors and cultivated in a smaller volume (300 mL) seemed to provide a more stable luminal ecosystem. The study of commercially available coffee and green tea in the M-batches suggested some positive effects of these worldwide known beverages, including the increase in butyrate-producing bacteria and lactobacilli populations. We hope that this novel strategy can contribute to future advances in the study of intestinal ecosystems and host-microbe relationships and help elucidate roles of the microbiome in health and disease.

In vitro screening of the impact of dietary prebiotic components, probiotic strains, and their symbiotic combinations on colonic microbiota in children with cystic fibrosis.

Children with Cystic Fibrosis (CF) are more likely to have intestinal dysbiosis due to recurrent antibiotic therapy and the conventional hypercaloric diet administered to them. This study aimed at evaluating the effect of isolated prebiotic components and probiotic strains, and their combinations as potential synbiotics, on the intestinal microbiota of CF children. A static in vitro colonic fermentation model was used by colonizing vials with faecal inoculum, a culture medium, and the substrates to be tested. Post treatment, aliquots were taken to determine ammonium, lactate, and short-chain fatty acids production and to profile the microbiota composition by 16s rRNA sequencing. At genus level, Escherichia-Shigella decreased (15.8%) with the treatment pectin + L. rhamnosus, followed by the beta-glucan + L. salivarius (15.5%). Inversely, the most increase in Bacteroides (44%) was obtained by the treatment with Pectin + L. reuteri. Lactate and acetic acid production was significantly increased with prebiotics and their combinations with L. rhamnosus and L. salivarius. In conclusion, the use of beta-glucan and pectin in combination with probiotic strains from the Lactobacillaceae family suggest potential to modulate dysbiosis and metabolic activity on CF colonic microbiota, encouraging further studies in animal studies or clinical settings to confirm the findings in vivo.

Modulation of the gut microbiota and short-chain fatty acid production by gac fruit juice and its fermentation in in vitro colonic fermentation.

This study aimed to investigate the effects of gac fruit juice and its probiotic fermentation (FGJ) utilizing Lactobacillus paracasei on the modulation of the gut microbiota and the production of short-chain fatty acids (SCFAs). We conducted a comparison between FGJ, non-fermented gac juice (GJ), and control samples through in vitro digestion and colonic fermentation using the human gut microbiota derived from fecal inoculum. Our findings revealed that both GJ and FGJ led to an increase in the viability of Lactobacilli, with FGJ exhibiting even higher levels compared to the control. The results from the 16S rDNA amplicon sequencing technique showed that both GJ and FGJ exerted positive impact on the gut microbiota by promoting beneficial bacteria, notably Lactobacillus mucosae and Bacteroides vulgatus. Additionally, both GJ and FGJ significantly elevated the levels of SCFAs, particularly acetic, propionic, and n-butyric acids, as well as lactic acid, in comparison to the control. Notably, FGJ exhibited a more pronounced effect on the gut microbiota compared to GJ. This was evident in its ability to enhance species richness, reduce the Firmicutes to Bacteroidetes (F/B) ratio, promote Akkermansia, and inhibit pathogenic Escherichia coli. Moreover, FGJ displayed enhanced production of SCFAs, especially acetic and lactic acids, in contrast to GJ. Our findings suggest that the probiotic fermentation of gac fruit enhances its functional attributes in promoting a balanced gut microbiota. This beverage demonstrates potential as a functional food with potential advantages for sustaining intestinal health.

The use of an in vitro fecal fermentation model to uncover the beneficial role of omega-3 and punicic acid in gut microbiota alterations induced by a Western diet.

The influence of gut microbiota in the onset and development of several metabolic diseases has gained attention over the last few years. Diet plays an essential role in gut microbiota modulation. Western diet (WD), characterized by high-sugar and high-fat consumption, alters gut microbiome composition, diversity index, microbial relative levels, and functional pathways. Despite the promising health effects demonstrated by polyunsaturated fatty acids, their impact on gut microbiota is still overlooked. The effect of Fish oil (omega-3 source) and Pomegranate oil (punicic acid source), and a mixture of both oils in gut microbiota modulation were determined by subjecting the oil samples to in vitro fecal fermentations. Cecal samples from rats from two different dietary groups: a control diet (CD) and a high-fat high-sugar diet (WD), were used as fecal inoculum. 16S amplicon metagenomics sequencing showed that Fish oil + Pomegranate oil from the WD group increased α-diversity. This sample can also increase the relative abundance of the Firmicutes and Bacteroidetes phylum as well as Akkermansia and Blautia, which were affected by the WD consumption. All samples were able to increase butyrate and acetate concentration in the WD group. Moreover, tyrosine concentrations, a precursor for dopamine and norepinephrine, increase in the Fish oil + Pomegranate oil WD sample. GABA, an important neurotransmitter, was also increased in WD samples. These results suggest a potential positive impact of these oils' mixture on gut-brain axis modulation. It was demonstrated, for the first time, the great potential of using a mixture of both Fish and Pomegranate oil to restore the gut microbiota changes associated with WD consumption.

Presence of digestible starch impacts in vitro fermentation of resistant starch.

Starch is an important energy source for humans. Starch escaping digestion in the small intestine will transit to the colon to be fermented by gut microbes. Many gut microbes express α-amylases that can degrade soluble starch, but only a few are able to degrade intrinsic resistant starch (RS), which is insoluble and highly resistant to digestion (≥80% RS). We studied the in vitro fermentability of eight retrograded starches (RS-3 preparations) differing in rapidly digestible starch content (≥70%, 35-50%, ≤15%) by a pooled adult faecal inoculum and found that fermentability depends on the digestible starch fraction. Digestible starch was readily fermented yielding acetate and lactate, whereas resistant starch was fermented much slower generating acetate and butyrate. Primarily Bifidobacterium increased in relative abundance upon digestible starch fermentation, whereas resistant starch fermentation also increased relative abundance of Ruminococcus and Lachnospiraceae. The presence of small fractions of total digestible starch (±25%) within RS-3 preparations influenced the fermentation rate and microbiota composition, after which the resistant starch fraction was hardly fermented. By short-chain fatty acid quantification, we observed that six individual faecal inocula obtained from infants and adults were able to ferment digestible starch, whereas only one adult faecal inoculum was fermenting intrinsic RS-3. This suggests that, in contrast to digestible starch, intrinsic RS-3 is only fermentable when specific microbes are present. Our data illustrates that awareness is required for the presence of digestible starch during in vitro fermentation of resistant starch, since such digestible fraction might influence and overrule the evalution of the prebiotic potential of resistant starches.