Intestinal Microbial Ecosystem Research Articles

Prebiotic properties of Ganoderma lucidum polysaccharides with special enrichment of Bacteroides ovatus and B. uniformis in vitro

Polysaccharide is the major biologically active constituent in functional mushroom Ganoderma lucidum. This study was aim to investigate the utilizability of G. lucidum polysaccharide by human fecal microbiota and its prebiotic potential based on the Simulator of Human Intestinal Microbial Ecosystem (SHIME). GLP was obtained from fruiting bodies of G. lucidum with average molecular weight of 133.1 KDa. Short-term colonic fermentation of GLP revealed the continuously decreased contents of total carbohydrates and reducing sugar as well as pH. Gas chromatography analysis indicated that GLP promoted the production of acetic, propionic and butyric acids. The 16S rDNA amplicon sequencing analysis showed that GLP improved the microbial diversity and ratio of Bacteroides to Firmicutes., and sharply enriched Bacteroides ovatus and B. uniformis. Therefore, GLP can be utilized by human fecal microbiota and then modify microbial community and metabolic function, and may serve as novel prebiotics to bring health benefits for human.

Tryptophan Supplementation Increases the Production of Microbial-Derived AhR Agonists in an In Vitro Simulator of Intestinal Microbial Ecosystem.

The aryl hydrocarbon receptor (AhR) plays an important role in intestinal homeostasis, and some microbial metabolites of tryptophan are known AhR agonists. In this study, we assessed the impact of tryptophan supplementation on the formation of tryptophan metabolites, AhR activation, and microbiota composition in the simulator of the human intestinal microbial ecosystem (SHIME). AhR activation, microbial composition, and tryptophan metabolites were compared during high tryptophan supplementation (4 g/L tryptophan), control, and wash-out periods. During tryptophan supplementation, the concentration of several tryptophan metabolites was increased compared to the control and wash-out period, but AhR activation by fermenter supernatant was significantly decreased. This was due to the higher levels of tryptophan, which was found to be an antagonist of AhR signaling. Tryptophan supplementation induced most microbial changes in the transverse colon including increased relative abundance of lactobacillus. We conclude that tryptophan supplementation leads to increased formation of AhR agonists in the colon.

Protective effect of flavonoids extract of Hippophae rhamnoides L. on alcoholic fatty liver disease through regulating intestinal flora and inhibiting TAK1/p38MAPK/p65NF-κB pathway

Ethnopharmacological relevanceThe therapeutic properties of Hippophae rhamnoides L. were already known in ancient Greece as well as in Tibetan and Mongolian medicine. Modern studies have indicated that Hippophae rhamnoides L. fermentation liquid protected against alcoholic fatty liver disease (AFLD). However, the underlying mechanism of Hippophae rhamnoides L. flavonoids extract (HLF) treating AFLD remains elusive. Aim of the studyThis study aimed to investigate the hepatoprotective effect of HLF in mice with AFLD and the interaction between AFLD and gut microbiota. Materials and methodsChemical constituents of HLF were analyzed by Liquid Chromatography-Ion Trap-ESI-Mass Spectrometry. The Hepatoprotective effect of HLF was evaluated in mice with AFLD induced by alcohol (six groups, n = 10) daily at doses of 0.1, 0.2, and 0.4 g/kg for 30 consecutive days. At the end of experiment, mice were sacrificed and the liver, serum and feces were harvested for analysis. The liver histological changes were observed by H&E staining and oil red O staining. Moreover, the alterations of fecal microflora were detected by 16S rRNA gene sequencing. The inflammatory related genes were determined by qRT-PCR and western blotting respectively. ResultsThe results showed that the oral administration of HLF remarkably alleviated hepatic lipid accumulation by decreasing the levels of ALT, AST, TG and TC. The levels of TNF-α, TGF-β, and IL-6 were also reduced after treatment with HLF. Meanwhile, the protein and mRNA expression of NF-kB p65, MAPK p38 and TAK-1 in the liver of mice with AFLD were all reduced by HLF compared with model group. Furthermore, the 16S rRNA gene sequencing analysis demonstrated that HLF treatment can help restore the imbalance of intestinal microbial ecosystem and reverse the changes in Fimicutes/Bacterodietes, Clostridiales, Lachnospiraceae, S24-7, and Prevotella in mice with AFLD. ConclusionHLF can effectively ameliorate liver injury in mice with AFLD, and regulate the composition of gut microbiota. Its regulatory mechanism may be related to TAK1/p38MAPK/p65NF-κB pathway. This study may provide novel insights into the mechanism of HLF on AFLD and a basis for promising clinical usage.

The Modulation of Chaihu Shugan Formula on Microbiota Composition in the Simulator of the Human Intestinal Microbial Ecosystem Technology Platform and its Influence on Gut Barrier and Intestinal Immunity in Caco-2/THP1-Blue™ Cell Co-Culture Model.

The traditional Chinese medicine (TCM)–Chaihu Shugan Formula (CSF), consisting of several Chinese botanical drugs like Bupleurum, is derived from the ancient Chinese pharmacopeia. It has been used for more than thousands of years in various suboptimal health statuses and diseases induced by chronic stress based on empirical therapy. Recent studies confirm the role of CSF in the development of many diseases, including depression, stress-induced hepatic injury and tumors. However, little has been known about the mechanisms behind the health effects of CSF. Here, we investigate the influence of CSF on the modulation of the simulated colonic microbiota of five healthy donors, gut barrier integrity, and intestinal immunity by combining the simulator of the human intestinal microbial ecosystem (SHIME®) technology platform with co-culture of intestinal and immune cells. This approach revealed that CSF stimulated the production of SCFA (acetate, propionate and butyrate) across donors while significantly lowering the production of branched SCFA (bSCFA). In terms of community composition, CSF stimulated a broad spectrum of health-related Bifidobacterium species, which are potent acetate and lactate producers. At the same time, it lowered the abundance of opportunistic pathogenic Escherichia coli. Later, we explore the effect of colonic fermentation of CSF on the gut barrier and intestinal immunity in the Caco-2/THP1-blue™ cell co-culture model. Based on the study using SHIME technology platform, CSF showed protective effects on inflammation-induced intestinal epithelial barrier disruption in all donors. Also, the treatment of CSF showed pronounced anti-inflammatory properties by strongly inducing anti-inflammatory cytokines IL-6 and IL-10 and reducing pro-inflammatory cytokine TNF-α. These findings demonstrate a significant modulatory effect of CSF on intestinal gut microbiota. CSF-microbial fermentation products improved the gut barrier and controlled intestinal inflammation.

Effects of Dietary Poly-β-Hydroxybutyrate Supplementation on the Growth, Non-specific Immunity, and Intestinal Microbiota of the Sea Cucumber Apostichopus japonicus

The present study evaluated the effects of dietary supplementation with poly-β-hydroxybutyrate (PHB) on the growth performance, non-specific immunity, immune-related gene expression in the Nuclear factor-κB (NF-κB) signaling pathway, and intestinal microbiota of the sea cucumber Apostichopus japonicus. During a 63-day feeding trial, the sea cucumber was fed with basal diet (control, C treatment) and treatment diets supplemented at different doses of PHB, i.e., 0.5% (P treatment), 1% (OP treatment), and 3% PHB (TP treatment) (w/w) followed by a stimulation test of inactivated Vibrio splendidus for 7 days. All PHB treatments significantly promoted the specific growth rate and weight gain rate of the sea cucumber (P < 0.05). The 1% PHB significantly enhanced phagocytic, respiratory burst, superoxide dismutase, alkaline phosphatase, acid phosphatase, catalase, and lysozyme activities in the coelomocytes of A. japonicus (P < 0.05). However, no elevated activities of catalase and lysozyme were observed in the P treatment (P < 0.05), and only increased catalase activity appeared in the TP treatment (P < 0.05). Quantitative real-time PCR revealed significantly higher expression levels of Aj-p105, Aj-p50, and Aj-rel in the mid-intestine tissue of the sea cucumber in the PHB treatments (P < 0.05), and the relative expression level Aj-lys gene was significantly higher in the OP treatment (P < 0.05) than that in the control. After injection of inactivated V. splendidus, the relative expression level of four immune-related genes in the OP treatment was significantly up-regulated at 24 h (P < 0.05). The richness of intestinal microbiota in PHB treatments significantly increased, while diversity in TP treatment significantly decreased (P < 0.05). The relative abundances of Rhodobacteraceae in the PHB treatments were significantly higher than that in the control (P < 0.05). Network analysis revealed that 0.5 and 1% PHB supplement enhanced the stability of the intestinal microbial ecosystem. Functional prediction revealed that the PHB diet significantly increased some potential functions of intestinal microbiota, involving amino acid, lipid, and nucleotide metabolisms. In summary, dietary supplementation with a suitable dose of PHB had shown multiple beneficial effects on A. japonicus. Considering collectively the above, the optimum dose of 1% PHB addition to the diet of A. japonicus is recommended.

Health Effect of N-Nitroso Diethylamine in Treated Water on Gut Microbiota Using a Simulated Human Intestinal Microbiota System

Chlorination disinfection byproducts (CDBPs) can exert adverse human health effects. Many toxicology-based studies confirmed the health hazards of CDBPs, but little research has been done on gut microbiome. We explored the effect of CDBPs on intestinal microbiota in the Simulator of the Human Intestinal Microbial Ecosystem (SHIME). The results showed that CDBPs slightly inhibited the production of short-chain fatty acids, and the abundance of Actinobacteria decreased in the transverse colon and descending colon. The abundance of Proteobacteria increased in the ascending colon and descending colon, while it decreased in the transverse colon. The abundance of Firmicutes decreased in both the ascending colon and descending colon. In particular, the abundance of Lachnospiraceae members, Bilophila, Oscillospira, Parabacteroides, Desulfovibrio, and Roseburia increased in the ascending colon, while the abundance of Sutterella, Bacteroides, Escherichia, Phascolarctobacterium, Clostridium, Citrobacter, and Klebsiella increased in the descending colon. The Shannon index differed significantly in both the ascending colon and descending colon before and after exposure. Overall, we demonstrate the feasibility of applying the SHIME model to studying the effects of intestinal toxicity on health of chlorinated by-products. The findings of this study improve our understanding of the health impact of CDBPs on the intestinal microbiota and better control of CDBPs in treated water is recommended.

Anti-obesity effects of Chenpi: an artificial gastrointestinal system study.

SummaryThe gut microbiota plays a significant role in human health; however, the complex relationship between gut microbial communities and host health is still to be thoroughly studied and understood. Microbes in the distal gut contribute to host health through the biosynthesis of vitamins and essential amino acids and the generation of important metabolic by‐products from dietary components that are left undigested by the small intestine. Aged citrus peel (Chenpi) is used in traditional Chinese medicine to lower cholesterol, promote weight loss and treat various gastrointestinal symptoms. This study investigated how the microbial community changes during treatment with Chenpi using the Simulator of the Human Intestinal Microbial Ecosystem (SHIME). Two preparations of Chenpi extract were tested: Chenpi suspended in oil only and Chenpi in a viscoelastic emulsion. Short‐chain fatty acids (SCFAs) were measured during treatment to monitor changes in the microbial community of the colon presenting a decrease in production for acetic, propionic and butyric acid (ANOVA (P < 0.001) during the 15 days of treatment. 16S rRNA sequencing of microbial samples showed a clear difference between the two treatments at the different sampling times (ANOSIM P < 0.003; ADOSIM P < 0.002 [R 2 = 69%]). Beta diversity analysis by PcoA showed differences between the two Chenpi formulations for treatment day 6. These differences were no longer detectable as soon as the Chenpi treatment was stopped, showing a reversible effect of Chenpi on the human microbiome. 16S rRNA sequencing of microbial samples from the descending colon showed an increase in Firmicutes for the treatment with the viscoelastic emulsion. At the genus level, Roseburia, Blautia, Subdoligranulum and Eubacterium increased in numbers during the viscoelastic emulsion treatment. This study sheds light on the anti‐obesity effect of a polymethoxyflavone (PMFs)‐enriched Chenpi extract and creates a foundation for the identification of ‘obesity‐prevention’ biomarkers in the gut microbiota.

Ameliorating the efficacy of galacto-oligosaccharides on ovalumin-induced allergic dermatitis symptoms in Balb/c mice by regulating Th2 immune response and the ecosystem of gut microbiota.

In this study, we evaluated the effect of oral administration of galacto-oligosaccharide (GOS), famous biological molecules that are comprised of galactose and lactose, on ovalbumin (OVA)-induced allergic dermatitis. OVA-induced mice were divided into the OVA-administered group (OVA-C), promethazine hydrochloride-administered group (PC), and 100 and 200 mg kg-1 GOS-administered groups (GL and GH, respectively). GOS administration significantly improved epidermal thickness and decreased CD4+ cell numbers. The concentrations of IgE, OVA-specific IgE, and inflammatory cytokines (IL-4, IL-5, and INF-γ) in GH group mice were significantly lower than those in OVA-C group mice. Firmicutes and Bacteroidetes were identified as the major phyla in the intestinal microbiota in mice, and the relative abundance of Deferribacteres was significantly lower in the GH group than in the OVA-C group. Deferribacteraceae and Mucispirillum species were significantly lower in the GH group than in the OVA-C group. The relative abundance of Muribaculum species was significantly lower, but those of Lachnospira and Lactococcus species were significantly higher in the GH group than in the OVA-C group. Our results suggest that the alleviation effect of GOS on allergic dermatitis induced by OVA sensitization was achieved by regulating hypersensitive immune responses by improving the intestinal microbial ecosystem.

A novel functional herbal tea containing probiotic Bacillus coagulans GanedenBC30: An in vitro study using the Simulator of the Human Intestinal Microbial Ecosystem (SHIME)

• Herbal tea as carrier of the probiotic Bacillus coagulans GanedenBC 30. • Spores of B. coagulans GanedenBC 30 markedly survive to infusion treatments. • Colonization and persistence in gut ecosystems investigated using the SHIME. • B. coagulans GanedenBC 30 quickly colonizes mucosal compartments and persists. • Viable cells in colon reactors are largely above the probiotic threshold. This study aimed at investigating the potential of a novel functional herbal tea to act as the carrier for the probiotic Bacillus coagulans GanedenBC 30 . Spores markedly survived to infusion treatments and the simulation of the gastrointestinal transit only slightly affected the survival, which was 94.8 ± 2.8%. An in-depth investigation using the Simulator of the Human Intestinal Microbial Ecosystem (SHIME) which incorporated mucin-covered microcosms was performed. Q-PCR with targeted primers demonstrated that the probiotic quickly colonized mucosal compartments and still persisted after 4 days of wash out. As estimated by plate count combined with 16S rRNA sequencing, B. coagulans GanedenBC 30 showed almost the same behavior in luminal compartments both during herbal tea intake and wash out. By summing the luminal and mucosal values and referring to whole volume of colon bioreactors, the content of viable cells of B. coagulans GanedenBC 30 was largely above the probiotic threshold.

Spirulina platensis biomass enhances the proliferation rate of Lactobacillus acidophilus 5 (La-5) and combined with La-5 impact the gut microbiota of medium-age healthy individuals through an in vitro gut microbiome model

This study first evaluated the stimulatory effect of S. platensis biomass on the growth of L. acidophilus and the metabolic activity during fermentation (37 °C, 72 h) in a culture medium. The results demonstrated a higher impact of S. platensis biomass than fructooligosaccharide (FOS), an established prebiotic. Higher L. acidophilus proliferation rates and metabolic activity were observed (lower pH values and higher concentrations of acetic, lactic, and propionic acids) in the presence of S. platensis. Then, we evaluated the effects of the S. platensis biomass (1.5 g, twice a day, 5 days) in association with L. acidophilus (106 CFU/g) on the gut microbiota composition of medium-age healthy individuals through the Simulator of Human Intestinal Microbial Ecosystem (SHIME®) and measurement of metabolites. L. acidophilus (La5) and L. acidophilus + S. platensis (Spi-La5) could positively modulate the intestinal microbiota. The administration of La5 resulted in increases in Bacteroides, Megasphaera, Lactobacillus, and Parabacteroides genus abundance, with a consequent decrease in ammonium ions. The administration of Spi-La5 increased the abundance of the genus Erysipelatoclostridium, Roseburia, Enterococcus, Bifidobacterium, Coriobacteriaceae UCG-003, Enterobacter, and Paraclostridium. The results demonstrate that the intestinal microbiota was differently modified by administrating La5 and Spi-La5 and indicate the latter as an alternative for microbiota positive modulation in healthy individuals.

The bioavailability of soy isoflavones in vitro and their effects on gut microbiota in the simulator of the human intestinal microbial ecosystem

Since soy isoflavones have many potentially beneficial properties, they have received increasing attention. However, there are few studies on their bioavailability and effects on gut microbiota. In this experiment, the simulator of the human intestinal microbial ecosystem (SHIME) was used to study the bioavailability of soy isoflavones and their effects on gut microbiota. The effects of soy isoflavones on the growth and antibacterial properties of probiotics were also studied in vitro. The results shows that the digestion of soy isoflavones mainly occurs in the colon, and the antioxidant capacity changes with the progress of digestion. Soy isoflavone can regulate gut microbiota, inhibit the growth of harmful bacteria, and affect the growth of obesity-related bacteria. Moreover, soy isoflavones can promote the growth of probiotics and significantly improve their antibacterial ability in vitro. Overall, soy isoflavones have the potential to be a functional food by improving gut microbiota.

Modulation of human gut microbiota composition and metabolites by arabinogalactan and Bifidobacterium longum subsp. longum BB536 in the Simulator of the Human Intestinal Microbial Ecosystem (SHIME®)

• Arabinogalactan (AG) gave higher amounts of SCFA, A. muciniphila and F. prausnitzii than starch . • The SCFA changes induced by the AG/ B. longum combination continued after B. longum supplementation. • The AG/ B. longum combination increased AG degradation, butyrate and F. prausnitzii . • The effect of the AG/ B. longum combination was dependent on the colon region. The Mucosal-Simulator of the Human Intestinal Microbial Ecosystem (SHIME) was used to differentiate the gut microbial fermentation of arabinogalactan (AG) and starch before (T1), during (T2), and after (T3) one-week Bifidobacterium longum subsp. longum BB536 supplementation. Adding 10 g/L of either AG or starch to the basal medium (T1) promoted short-chain fatty acid (SCFA) production in three successive SHIME colon vessels. AG fermentation (T1) resulted in higher total SCFAs ( P < 0.05) and luminal live Faecalibacterium prausnitzii ( P < 0.0001) in the transverse colon than starch fermentation (T1). Compared to AG fermentation alone (T1), B. longum supplementation (T2) significantly enhanced butyrate production ( P < 0.05) and the abundance of luminal live F. prausnitzii ( P < 0.0001) in the transverse colon. The results indicate that combining a potential prebiotic and a probiotic strain increases SCFA production and particular gut commensals which could have a beneficial effect on gut health.

Studies and Application of Sialylated Milk Components on Regulating Neonatal Gut Microbiota and Health.

Breast milk is rich in sialic acids (SA), which are commonly combined with milk oligosaccharides and glycoconjugates. As a functional nutrient component, SA-containing milk components have received increasing attention in recent years. Sialylated human milk oligosaccharides (HMOs) have been demonstrated to promote the growth and metabolism of beneficial gut microbiota in infants, bringing positive outcomes to intestinal health and immune function. They also exhibit antiviral and bacteriostatic activities in the intestinal mucosa of new-borns, thereby inhibiting the adhesion of pathogens to host cells. These properties play a pivotal role in regulating the intestinal microbial ecosystem and preventing the occurrence of neonatal inflammatory diseases. In addition, some recent studies also support the promoting effects of sialylated HMOs on neonatal bone and brain development. In addition to HMOs, sialylated glycoproteins and glycolipids are abundant in milk, and are also critical to neonatal health. This article reviews the current research progress in the regulation of sialylated milk oligosaccharides and glycoconjugates on neonatal gut microbiota and health.

Differential fermentation of raw and processed high-amylose and waxy maize starches in the Simulator of the Human Intestinal Microbial Ecosystem (SHIME®)

This study aimed to evaluate the colonic fermentation profiles of both high-amylose and waxy maize starches in raw, cooked, and cooked-cooled forms, as measured by changes in short-chain fatty acid production and microbiota composition, using the Mucosal-Simulator of the Human Intestinal Microbial Ecosystem (SHIME). Adding starch (6 g/L) to basal feed promoted butyrate production in SHIME proximal and distal colon vessels. The cooked high-amylose starch induced the highest butyrate production in both colon vessels, while the cooked waxy starch induced the highest butyrate production in the distal but not the proximal colon vessel. All starch preparations increased Bifidobacteriaceae abundance in the proximal colon vessel (2- to 6-fold) and the Firmicutes/Bacteroidetes ratio. The cooked high-amylose starch increased Akkermansiaceae abundance in the distal (2-fold) but not the proximal colon vessel. The results indicate the cooking and cooling effect on starch fermentation outcome in successive colon regions depends on the starch amylose content.

In-Vitro Characterization of Growth Inhibition against the Gut Pathogen of Potentially Probiotic Lactic Acid Bacteria Strains Isolated from Fermented Products.

Lactic acid bacteria (LAB) are probiotic candidates that may restore the balance of microbiota populations in intestinal microbial ecosystems by controlling pathogens and thereby promoting host health. The goal of this study was to isolate potential probiotic LAB strains and characterize their antimicrobial abilities against pathogens in intestinal microbiota. Among 54 LAB strains isolated from fermented products, five LAB strains (NSMJ15, NSMJ16, NSMJ23, NSMJ42, and NFFJ04) were selected as potential probiotic candidates based on in vitro assays of acid and bile salt tolerance, cell surface hydrophobicity, adhesion to the intestinal epithelium, and antagonistic activity. Phylogenetic analysis based on 16S rRNA genes showed that they have high similarities of 99.58–100% to Lacticaseibacillus paracasei strains NSMJ15 and NFFJ04, Lentilactobacillus parabuchneri NSMJ16, Levilactobacillus brevis NSMJ23, and Schleiferilactobacillus harbinensis NSMJ42. To characterize their antimicrobial abilities against pathogens in intestinal microbiota, the impact of cell-free supernatant (CFS) treatment in 10% (v/v) fecal suspensions prepared using pooled cattle feces was investigated using in vitro batch cultures. Bacterial community analysis using rRNA amplicon sequencing for control and CFS-treated fecal samples at 8 and 16 h incubation showed the compositional change after CFS treatment for all five LAB strains. The changed compositions were similar among them, but there were few variable increases or decreases in some bacterial groups. Interestingly, as major genera that could exhibit pathogenicity and antibiotic resistance, the members of Bacillus, Escherichia, Leclercia, Morganella, and Vagococcus were decreased at 16 h in all CFS-treated samples. Species-level classification suggested that the five LAB strains are antagonistic to gut pathogens. This study showed the probiotic potential of the five selected LAB strains; in particular, their antimicrobial properties against pathogens present in the intestinal microbiota. These strains would therefore seem to play an important role in modulating the intestinal microbiome of the host.

Cardiovascular Health and The Intestinal Microbial Ecosystem: The Impact of Cardiovascular Therapies on The Gut Microbiota.

It has become evident over the past several years that the intestinal microbial ecosystem plays a critical role in the development and prevention of cardiovascular diseases (CVDs) and other metabolic disorders, such as hypertension, obesity, diabetes mellitus, and metabolic syndrome. The intestinal microbiota ecosystem functions as a major virtual endocrine organ that interacts and responds to molecules’ signals within the host. Several meta-organismal pathways are involved in the gut–host interaction, including trimethylamine-N-oxide (TMAO) and short-chain fatty acids (SCFA). Host phenotype and cardiovascular diseases (CVDs) varying from hypertension, insulin resistance, and obesity to more specific inflammatory processes, such as atherosclerosis and hypercoagulability, have shown to be affected by the gut–host interaction. Additionally, several studies that involved animals and humans demonstrated a striking connection between the development of new CVDs and an imbalance in the gut microbiota composition along with the presence of their derived metabolites. Through this review article, we aim to evaluate the role of the normal gut microbiota ecosystem, its association with CVDs, effects of the therapies used to control and manage CVDs in the gut microbiota environment and explore potential therapeutic interventions to amplify disease outcomes in patients with CVDs.

Treatment with a spore-based probiotic containing five strains of Bacillus induced changes in the metabolic activity and community composition of the gut microbiota in a SHIME® model of the human gastrointestinal system

MegaSporeBiotic™ is an oral, spore-based probiotic comprised of five Bacillus spp. (Bacillus indicus HU36, Bacillus subtilis HU58, Bacillus coagulans SC208, Bacillus licheniformis SL307, and Bacillus clausii SC109). The effects of MegaSporeBiotic™ on gut microbiota activity and community composition were evaluated for the first time using an in vitro model of the human gastrointestinal tract, the simulator of the human intestinal microbial ecosystem (SHIME®), under healthy conditions. Following a stabilization period and a control period (2 weeks each), the reactor feed was supplemented with daily MegaSporeBiotic™ for 3 weeks (treatment period). Changes in microbial community activity and composition between the control and treatment periods were evaluated for each colon compartment (ascending [AC], transverse [TC], and descending colon [DC]). Propionate levels increased significantly in the TC (week 2, P = 0.02; week 3, P = 0.0019) and DC (week 2, P = 0.03) with treatment while lactate levels significantly decreased in the TC (week 3, P = 0.03). Ammonium levels were significantly decreased during the final week of treatment (TC, P = 0.02; DC, P = 0.03). Overall, Akkermansia muciniphila, Bifidobacteria spp., and Firmicutes increased with treatment while Lactobacillus spp. and Bacteroidetes decreased. The Firmicutes:Bacteroidetes ratio increased with treatment in the AC compartment. MegaSporeProbiotic™ treatment resulted in changes in metabolism and increased bacterial diversity.

Comparison of protection and release behavior of different capsule polymer combinations based on L. acidophilus survivability and function and caffeine release

Oral administration of active pharmaceutical ingredients, nutraceuticals, enzymes or probiotics requires an appropriate delivery system for optimal bioactivity and absorption. The harsh conditions during the gastrointestinal transit can degrade the administered products, hampering their efficacy. Enteric or delayed-release pharmaceutical formulations may help overcome these issues. In a Simulator of Human Intestinal Microbial Ecosystem model (SHIME) and using caffeine as a marker for release kinetics and L. acidophilus survivability as an indicator for protection, we compared the performance of ten capsule configurations, single or DUOCAP® combinations. The function of L. acidophilus and its impact on the gut microbiota was further tested in three selected capsule types, combinations of DRcaps® capsule in DRcaps® capsule (DR-in-DR) and DRcaps® capsule in Vcaps® capsule (DR-in-VC) and single Vcaps® Plus capsule under colonic conditions. We found that under stomach and small intestine conditions, DR-in-DR and DR-in-VC led to the best performance both under fed and fasted conditions based on the slow caffeine release and the highest L. acidophilus survivability. The Vcaps® Plus capsule however, led to the quickest caffeine and probiotic release. When DR-in-DR, DR-in-VC and single Vcaps® Plus capsules were tested through the whole gastrointestinal tract, including under colonic conditions, caffeine release was found to be slower in capsules containing DRcaps® capsules compared to the single Vcaps® capsules. In addition, colonic survival of L. acidophilus was significantly increased under fasted conditions in DR-in-DR or DR-in-VC formulation compared to Vcaps® Plus capsule. To assess the impact of these formulations on the microbial function, acetate, butyrate and propionate as well as ammonia were measured. L. acidophilus released from DR-in-DR or DR-in-VC induced a significant increase in butyrate and a decrease in ammonia, suggesting a proliferation of butyrate-producing bacteria and reduction in ammonia-producing bacteria. These data suggest that L. acidophilus included in DR-in-DR or DR-in-VC reaching the colon is viable and functional, potentially contributing to changes in colonic microbiota composition and diversity.

Inclusion of small intestinal absorption and simulated mucosal surfaces further improve the Mucosal Simulator of the Canine Intestinal Microbial Ecosystem (M-SCIME™)

While a large set of in vitro models are available to study the effects of specific food ingredients (e.g. pre- and probiotics) on the human gut microbiome, the availability of such models for companion animals is limited. Since improving gut health of such animals is an emerging research field, the Simulator of the Canine Intestinal Microbial Ecosystem (SCIME™) was recently developed and validated with in vivo data. The current study presents a further improvement of this model by using an alternative method for feed preparation, i.e. by administering digestive enzymes to mimic upper gastro-intestinal digestion followed by a dialysis approach to mimic small intestinal absorption. As opposed to the previously implemented method, this resulted in a more optimal simulation of protein digestion and absorption. Further, upon entrance in the colon, increased production of the health-promoting butyrate and lower levels of Lactobacillus spp. and Bifidobacterium spp. were observed, which corresponded better with obtained in vivo data. A second model improvement consisted of the implementation of a mucosal environment to not only simulate luminal but also mucosal microbiota. In consistency with the human model for which this technology was previously validated, it was found that for all canine microbiota mucin beads were enriched with members of the Lachnospiraceae (~ Clostridium cluster XIVa), a family containing multiple well-known butyrate producers. The SCIME™ was thus upgraded to a so-called Mucosal SCIME™ (M-SCIME™). In conclusion, the current study presents improvements of the SCIME™, further increasing the relevance of obtained data with this in vitro model for dogs.

Nutrient load acts as a driver of gut microbiota load, community composition and metabolic functionality in the simulator of the human intestinal microbial ecosystem.

A recently introduced quantitative framework for gut microbiota analysis indicated that microbial load alterations can be linked to various diseases, making it essential to pinpoint its determinants. We identified nutrient load as a main driver of the quantitative microbial community composition and functionality in vitro by stepwise decreasing standardized feed concentrations from 100% to 33%, 20% and 10% in 5-day intervals. While the proportional composition and metabolic profile were mainly determined by the inter-individual variability (35% and 41%), nutrient load accounted for 58%, 23% and 65% of the observed variation in the microbial load, quantitative composition and net daily metabolite production, respectively. After the 10-fold nutrient reduction, the microbial load decreased by 79.72± 9% and 82.96± 1.66% in the proximal and distal colon, respectively, while the net total short-chain fatty acid production dropped by 79.42± 4.42% and 84.58± 2.42%, respectively. The majority of microbial taxa quantitatively decreased, whereas a select group of nutritional specialists, such as Akkermansia muciniphila and Bilophila wadsworthia, and a number of opportunistic pathogens remained unaffected. This shows that nutrient load is an important driver of the human gut microbiome and should be considered in future in vitro and in vivo dietary research.