A Wake-Up Call for the Efficient Use of the Bacterial Resting Cell Process, with Focus on Low Solubility Products.

Micro-organisms are often subjected to stressful conditions. Owing to their capacity to adapt, they try to rapidly cope with the unfavorable conditions by lowering their growth rate, changing their morphology, and developing altered metabolite production and other stress-related metabolism. The stress-related metabolism of the cells which interrupted their growth is often referred to as resting metabolism and can be exploit for specific and high rate production of secondary metabolites. Although the bacterial resting cell process has been described decades ago, we find it worthwhile to bring the process under renewed attention and refer to this type of processes as non-growing metabolically active (NGMA) cell processes. Despite their use may sound counterproductive, NGMA cells can be of interest to increase substrate conversion rates or enable conversion of certain substrates, not accessible to growing cells due to their bacteriostatic nature or requirement of resistance to a multitude of different stress mechanisms. Biomass reuse is an interesting feature to improve the economics of NGMA cell processes. Yet, for lipophilic compounds or compounds with low solubility, biomass separation can be delicate. This review draws the attention on existing examples of NGMA cell processes, summarizing some developmental tools and highlighting drawbacks and opportunities, to answer the research question if NGMA cells can have a distinct added value in industry. Particular elaboration is made on a novel and more broadly applicable strategy to enable biomass reuse for conversions of compounds with low solubility.

Citrus Flavonoid Supplementation Improves Exercise Performance in Trained Athletes.

Previous studies have shown that polyphenol supplementation may be an effective strategy to improve exercise performance, due to their antioxidant character and ability to stimulate NO production. These properties may contribute to exercise performance, yet no conclusive research has been performed in exploring the direct effects of citrus flavonoids on human exercise performance. Therefore, the purpose of this study was to assess whether supplementation of a customized citrus flavonoid (CF) extract for 4 weeks improves cycling time-trial performance in trained male athletes. In a double-blind, randomized, parallel study, 39 healthy, trained males were given a daily dose of either 500 mg of a customized citrus flavonoid extract (CF) or a placebo for 4 weeks. Exercise performance was tested by means of a time-trial test on a cycle ergometer, during which participants had to generate as much power as possible for duration of 10 minutes. Absolute power output significantly increased with 14.9 ± 3.9 W after 4 weeks of CF supplementation, corresponding with a 5.0% increase, compared to 3.8 ± 3.2 W (1.3% increase) in placebo (p < 0.05). In addition, oxygen consumption/power ratio significantly decreased in the CF group compared to placebo (p = 0.001), and a trend was found in the change in peak power output in CF (18.2 ± 23.2 W) versus placebo (-28.4 ± 17.6 W; p = 0.116). The current study is the first convincing report that citrus flavonoid supplementation can improve exercise performance, as shown by a significant increase in power output during the exercise test.

A yeast fermentate improves gastrointestinal discomfort and constipation by modulation of the gut microbiome: results from a randomized double-blind placebo-controlled pilot trial.

BACKGROUND: Constipation and symptoms of gastrointestinal discomfort such as bloating are common among otherwise healthy individuals, but with significant impact on quality of life. Despite the recognized contribution of the gut microbiome to this pathology, little is known about which group(s) of microorganism(s) are playing a role. A previous study performed in vitro suggests that EpiCor® fermentate has prebiotic-like properties, being able to favorably modulate the composition of the gut microbiome. Therefore, the aim of this study was to investigate the effects of EpiCor fermentate in a population with symptoms of gastrointestinal discomfort and reduced bowel movements and to evaluate its effect at the level of the gut microbiome. METHODS: This pilot study was performed according to a randomized, double-blind, placebo-controlled parallel design. Eighty subjects with symptoms of gastrointestinal discomfort and constipation were allocated to one of two trial arms (placebo or EpiCor fermentate). Randomization was done in a stratified manner according to symptom severity, resulting in two subgroups of patients: severe and moderate. Daily records of gastrointestinal symptoms were assessed on a 5-point scale, and also stool frequency and consistency were documented during a 2-week run-in and a 6-week intervention phases. Averages over two-week intervals were calculated. Constipation-associated quality of life and general perceived stress were assessed at baseline and after 3 and 6 weeks of intervention. Fecal samples were also collected at these same time points. RESULTS: EpiCor fermentate led to a significant improvement of symptoms such as bloating/distension (p = 0.033 and p = 0.024 after 2 and 4 weeks of intervention, respectively), feeling of fullness (p = 0.004 and p = 0.023 after 2 and 4 weeks of intervention, respectively) and general daily scores (p = 0.046 after 2 weeks of intervention) in the moderate subgroup. A significant improvement in stool consistency was observed for the total population (p = 0.023 after 2 weeks of intervention) as well as for the severe subgroup (p = 0.046 after 2 weeks of intervention), and a nearly significant increase in stool frequency was detected for the total cohort (p = 0.083 and p = 0.090 after 2 and 4 weeks of intervention, respectively). These effects were accompanied by an improvement in constipation-associated quality of life and general perceived stress, particularly in the moderate subgroup. Members of the families Bacteroidaceae and Prevotellaceae, two groups of bacteria that have been previously reported to be deficient in constipated patients, were found to increase with EpiCor fermentate in the severe subgroup. In the moderate subgroup, a significant increase in Akkermansia muciniphila was observed. CONCLUSIONS: Despite the relatively low dose administered (500 mg/day), particularly when comparing to the high recommended doses for prebiotic fibers, EpiCor fermentate was able to modulate the composition of the gut microbiome, resulting in improvement of constipation-associated symptoms. Conversely, the reported increase in bowel movements may have altered the gut microbial community by increasing those groups of bacteria that are better adapted to a faster gastrointestinal transit time. TRIAL REGISTRATION: NCT03051399 at ClinicalTrials.gov. Retrospectively registered. Registration date: 13 February 2017.

Arabinoxylan in wheat is more responsible than cellulose for promoting intestinal barrier function in weaned male piglets.

BACKGROUND: The effect of dietary fiber on intestinal function primarily has been ascribed to its interaction with intestinal bacteria in the hindgut, whereas changes in intestinal bacteria in the host have been considered to depend on fiber composition. OBJECTIVES: The objectives of this study were to determine the contribution of the major fiber components to the health-promoting effects of wheat bran on intestinal mucosal barrier function and to elucidate the involvement of microbiota changes in weaned piglets. METHODS: Thirty freshly weaned male piglets were assigned to 5 dietary treatment groups (n = 6) according to litter and weight. The piglets consumed synthetic diets ad libitum for 30 d, including a basal control diet (CON) without fiber components, a wheat bran diet (WB) as reference diet (10% wheat bran), and 3 other diets containing amounts of fiber components equivalent to those in the WB, i.e., an arabinoxylan diet (AX), a cellulose diet (CEL), and a combined arabinoxylan and cellulose diet (CB). RESULTS: The groups consuming diets containing arabinoxylans (i.e., the WB, AX, and CB groups) had increased intestinal secretory immunoglobulin A concentrations, goblet cell number and cecal short-chain fatty acid concentrations, and reduced branched-chain fatty acid concentrations and pH values compared with the CON group. In the WB group, the stimulated secretion of Cl(-) was suppressed (60.8% and 47.5% change in short-circuit current caused by theophylline and carbachol, respectively) in the distal small intestine compared with the CON group. The AX and CB groups also had increased intestinal alkaline phosphatase activities and reduced intestinal transcellular permeability (by 77.3% and 67.2%, respectively) compared with the CON group. Meanwhile, in the WB group, cecal Bacteroidetes and Enterobacteriaceae populations were lower, and the growth of Lactobacillus was higher in the AX and CB groups than in the CON group, whereas no positive effect on intestinal barrier function was observed in the CEL group. CONCLUSION: Arabinoxylan in wheat bran, and not cellulose, is mainly responsible for improving various functional components of the intestinal barrier function and the involvement of microbiota changes.

The HMI™ module: a new tool to study the Host-Microbiota Interaction in the human gastrointestinal tract in vitro.

BACKGROUND: Recent scientific developments have shed more light on the importance of the host-microbe interaction, particularly in the gut. However, the mechanistic study of the host-microbe interplay is complicated by the intrinsic limitations in reaching the different areas of the gastrointestinal tract (GIT) in vivo. In this paper, we present the technical validation of a new device--the Host-Microbiota Interaction (HMI) module--and the evidence that it can be used in combination with a gut dynamic simulator to evaluate the effect of a specific treatment at the level of the luminal microbial community and of the host surface colonization and signaling. RESULTS: The HMI module recreates conditions that are physiologically relevant for the GIT: i) a mucosal area to which bacteria can adhere under relevant shear stress (3 dynes cm(-2)); ii) the bilateral transport of low molecular weight metabolites (4 to 150 kDa) with permeation coefficients ranging from 2.4 × 10(-6) to 7.1 × 10(-9) cm sec(-1); and iii) microaerophilic conditions at the bottom of the growing biofilm (PmO2 = 2.5 × 10(-4) cm sec(-1)). In a long-term study, the host's cells in the HMI module were still viable after a 48-hour exposure to a complex microbial community. The dominant mucus-associated microbiota differed from the luminal one and its composition was influenced by the treatment with a dried product derived from yeast fermentation. The latter--with known anti-inflammatory properties--induced a decrease of pro-inflammatory IL-8 production between 24 and 48 h. CONCLUSIONS: The study of the in vivo functionality of adhering bacterial communities in the human GIT and of the localized effect on the host is frequently hindered by the complexity of reaching particular areas of the GIT. The HMI module offers the possibility of co-culturing a gut representative microbial community with enterocyte-like cells up to 48 h and may therefore contribute to the mechanistic understanding of host-microbiome interactions.

Effects of wheat bran extract containing arabinoxylan oligosaccharides on gastrointestinal parameters in healthy preadolescent children.

OBJECTIVES: We assessed whether wheat bran extract (WBE) containing arabinoxylan-oligosaccharides (AXOS) elicited a prebiotic effect and modulated gastrointestinal (GI) parameters in healthy preadolescent children upon consumption in a beverage. METHODS: This double-blind randomized placebo-controlled crossover trial evaluated the effects of consuming WBE at 0 (control) or 5.0 g/day for 3 weeks in 29 healthy children (8-12 years). Fecal levels of microbiota, short-chain fatty acids, branched-chain fatty acids, ammonia, moisture, and fecal pH were assessed at the end of each treatment and at the end of a 1-week run-in (RI) period. In addition, the subjects completed questionnaires scoring distress severity of 3 surveyed GI symptoms. Finally, subjects recorded defecation frequency and stool consistency. RESULTS: Nominal fecal bifidobacteria levels tended to increase after 5 g/day WBE consumption (P = 0.069), whereas bifidobacteria expressed as percentage of total fecal microbiota was significantly higher upon 5 g/day WBE intake (P = 0.002). Additionally, 5 g/day WBE intake induced a significant decrease in fecal content of isobutyric acid and isovaleric acid (P < 0.01), markers of protein fermentation. WBE intake did not cause a change in distress severity of the 3 surveyed GI symptoms (flatulence, abdominal pain/cramps, and urge to vomit) (P > 0.1). CONCLUSIONS: WBE is well tolerated at doses up to 5 g/day in healthy preadolescent children. In addition, the intake of 5 g/day exerts beneficial effects on gut parameters, in particular an increase in fecal bifidobacteria levels relative to total fecal microbiota, and reduction of colonic protein fermentation.

Metabolic fate of polyphenols in the human superorganism.

Dietary polyphenols are components of many foods such as tea, fruit, and vegetables and are associated with several beneficial health effects although, so far, largely based on epidemiological studies. The intact forms of complex dietary polyphenols have limited bioavailability, with low circulating levels in plasma. A major part of the polyphenols persists in the colon, where the resident microbiota produce metabolites that can undergo further metabolism upon entering systemic circulation. Unraveling the complex metabolic fate of polyphenols in this human superorganism requires joint deployment of in vitro and humanized mouse models and human intervention trials. Within these systems, the variation in diversity and functionality of the colonic microbiota can increasingly be captured by rapidly developing microbiomics and metabolomics technologies. Furthermore, metabolomics is coming to grips with the large biological variation superimposed on relatively subtle effects of dietary interventions. In particular when metabolomics is deployed in conjunction with a longitudinal study design, quantitative nutrikinetic signatures can be obtained. These signatures can be used to define nutritional phenotypes with different kinetic characteristics for the bioconversion capacity for polyphenols. Bottom-up as well as top-down approaches need to be pursued to link gut microbial diversity to functionality in nutritional phenotypes and, ultimately, to bioactivity of polyphenols. This approach will pave the way for personalization of nutrition based on gut microbial functionality of individuals or populations.

Implication of fermentable carbohydrates targeting the gut microbiota on conjugated linoleic acid production in high-fat-fed mice.

In vitro experiments have shown that isolated human gut bacteria are able to metabolise PUFA into conjugated PUFA like conjugated linoleic acids (CLA). The hypothesis of the present paper was that high-fat (HF) diet feeding and supplementation with fermentable carbohydrates that have prebiotic properties modulate the in vivo production of CLA by the mouse gut microbiota. Mice were treated for 4 weeks as follows: control (CT) groups were fed a standard diet; HF groups were fed a HF diet rich in linoleic acid (18 : 2n-6); the third groups were fed with the HF diet supplemented with either inulin-type fructans (HF-ITF) or arabinoxylans (HF-Ax). HF diet feeding increased rumenic acid (cis-9,trans-11-18 : 2 CLA) content both in the caecal and liver tissues compared with the CT groups. ITF supplementation had no major effect compared with the HF diet whereas Ax supplementation increased further rumenic acid (cis-9,trans-11-18 : 2 CLA) in the caecal tissue. These differences between both prebiotics may be linked to the high fat-binding capacity of Ax that provides more substrates for bacterial metabolism and to differential modulation of the gut microbiota (specific increase in Roseburia spp. in HF-Ax v. HF). In conclusion, these experiments supply the proof of concept that the mouse gut microbiota produces CLA in vivo, with consequences on the level of CLA in the caecal and liver tissues. We postulate that the CLA-producing bacteria could be a mediator to consider in the metabolic effects of both HF diet feeding and prebiotic supplementation.

Barcoded pyrosequencing analysis of the microbial community in a simulator of the human gastrointestinal tract showed a colon region-specific microbiota modulation for two plant-derived polysaccharide blends.

The combination of a Simulator of the Human Intestinal Microbial Ecosystem with ad hoc molecular techniques (i.e. pyrosequencing, denaturing gradient gel electrophoresis and quantitative PCR) allowed an evaluation of the extent to which two plant polysaccharide supplements could modify a complex gut microbial community. The presence of Aloe vera gel powder and algae extract in product B as compared to the standard blend (product A) improved its fermentation along the entire simulated colon. The potential extended effect of product B in the simulated distal colon, as compared to product A, was confirmed by: (i) the separate clustering of the samples before and after the treatment in the phylogenetic-based dendrogram and OTU-based PCoA plot only for product B; (ii) a higher richness estimator (+33 vs. -36 % of product A); and (iii) a higher dynamic parameter (21 vs. 13 %). These data show that the combination of well designed in vitro simulators with barcoded pyrosequencing is a powerful tool for characterizing changes occurring in the gut microbiota following a treatment. However, for the quantification of low-abundance species-of interest because of their relationship to potential positive health effects (i.e. bifidobacteria or lactobacilli)-conventional molecular ecological approaches, such as PCR-DGGE and qPCR, still remain a very useful complementary tool.

An exploratory study into the putative prebiotic activity of fructans isolated from Agave angustifolia and the associated anticancer activity.

Linear inulin-type fructan (ITF) prebiotics have a putative role in the prevention of colorectal cancer, whereas relatively little is known about branched fructans. This study aims to investigate the fermentation properties and potential prebiotic activity of branched fructans derived from Agave angustifolia Haw, using the Simulator of Human Intestinal Microbial Ecosystem (SHIME) model. The proximal, transverse and distal vessels were used to investigate fructan fermentation throughout the colon and to assess the alterations of the microbial composition and fermentation metabolites (short chain fatty acids and ammonia). The influence on bioactivity of the fermentation supernatant was assessed by MTT, Comet and transepithelial electrical resistance (TER), respectively. Addition of Agave fructan to the SHIME model significantly increased (P < 0.05), bifidobacteria populations (proximal and transverse), SCFA concentrations (proximal, transverse and distal) and decreased ammonia concentrations in the distal vessel. Furthermore, the fermentation supernatant significantly (P < 0.05) increased the TER of a Caco-2 cell monolayer (%) and decreased fluorescein-based paracellular flux, suggesting enhanced barrier function and reduced epithelial barrier permeability (proximal and distal vessel). While cytotoxicity and genotoxicity remained unaltered in response to the presence of Agave fructans. To conclude, branched Agave fructans show indications of prebiotic activity, particularly in relation to colon health by exerting a positive influence on gut barrier function, an important aspect of colon carcinogenesis.

Effects of a wheat bran extract containing arabinoxylan oligosaccharides on gastrointestinal health parameters in healthy adult human volunteers: a double-blind, randomised, placebo-controlled, cross-over trial.

Wheat bran extract (WBE) is a food-grade soluble fibre preparation that is highly enriched in arabinoxylan oligosaccharides. In this placebo-controlled cross-over human intervention trial, tolerance and effects on colonic protein and carbohydrate fermentation were studied. After a 1-week run-in period, sixty-three healthy adult volunteers consumed 3, 10 and 0 g WBE/d for 3 weeks in a random order, with 2 weeks' washout between each treatment period. Fasting blood samples were collected at the end of the run-in period and at the end of each treatment period for analysis of haematological and clinical chemistry parameters. Additionally, subjects collected a stool sample for analysis of microbiota, SCFA and pH. A urine sample, collected over 48 h, was used for analysis of p-cresol and phenol content. Finally, the subjects completed questionnaires scoring occurrence frequency and distress severity of eighteen gastrointestinal symptoms. Urinary p-cresol excretion was significantly decreased after WBE consumption at 10 g/d. Faecal bifidobacteria levels were significantly increased after daily intake of 10 g WBE. Additionally, WBE intake at 10 g/d increased faecal SCFA concentrations and lowered faecal pH, indicating increased colonic fermentation of WBE into desired metabolites. At 10 g/d, WBE caused a mild increase in flatulence occurrence frequency and distress severity and a tendency for a mild decrease in constipation occurrence frequency. In conclusion, WBE is well tolerated at doses up to 10 g/d in healthy adults volunteers. Intake of 10 g WBE/d exerts beneficial effects on gut health parameters.

A Nine-Strain Bacterial Consortium Improves Portal Hypertension and Insulin Signaling and Delays NAFLD Progression In Vivo.

The gut microbiome has a recognized role in Non-alcoholic fatty liver disease (NAFLD) and associated comorbidities such as Type-2 diabetes and obesity. Stool transplantation has been shown to improve disease by restoring endothelial function and insulin signaling. However, more patient-friendly treatments are required. The present study aimed to test the effect of a defined bacterial consortium of nine gut commensal strains in two in vivo rodent models of Non-alcoholic steatohepatitis (NASH): a rat model of NASH and portal hypertension (PHT), and the Stelic animal (mouse) model (STAM™). In both studies the consortium was administered orally q.d. after disease induction. In the NASH rats, the consortium was administered for 2 weeks and compared to stool transplant. In the STAM™ study administration was performed for 4 weeks, and the effects compared to vehicle or Telmisartan at the stage of NASH/early fibrosis. A second group of animals was followed for another 3 weeks to assess later-stage fibrosis. In the NASH rats, an improvement in PHT and endothelial function was observed. Gut microbial compositional changes also revealed that the consortium achieved a more defined and richer replacement of the gut microbiome than stool transplantation. Moreover, liver transcriptomics suggested a beneficial modulation of pro-fibrogenic pathways. An improvement in liver fibrosis was then confirmed in the STAM™ study. In this study, the bacterial consortium improved the NAFLD activity score, consistent with a decrease in steatosis and ballooning. Serum cytokeratin-18 levels were also reduced. Therefore, administration of a specific bacterial consortium of defined composition can ameliorate NASH, PHT, and fibrosis, and delay disease progression.

Bacterial monocultures, propionate, butyrate and H2O2 modulate the expression, secretion and structure of the fasting-induced adipose factor in gut epithelial cell lines.

Previous research showed that an intestinal microbial community represses the fasting-induced adipose factor (FIAF) in the gut epithelium, thereby increasing fat storage in the host. This study was designed to investigate the overall effect of different bacterial species and metabolites on FIAF in intestinal (Caco-2, HT-29 and HCT-116) and hepatic (HepG2) cancer cell lines. First, we showed that FIAF was present in different isoforms, and secreted as N-glycosylated proteins, exclusively at the basal side of the cell monolayer. Second, co-incubation of cell lines with bacterial monocultures and metabolites altered both FIAF production and isoform appearance. Propionate and/or butyrate treatment increased FIAF expression and cleavage in all tested cell lines. In contrast, different bacteria induced cell line-specific FIAF modulation. Clostridium perfringens induced FIAF isoform changes in Caco-2 cells. Enterococcus faecalis and Bacteroides thetaiotaomicron treatment resulted in cell line-specific FIAF increases, whereas Escherichia coli significantly decreased FIAF expression in HCT-116 cells. Treatment with H(2) O(2) and peroxide-producing E. faecalis strains induced FIAF isoform changes in Caco-2 cells. Since bacteria and bacterial metabolites alter both FIAF production and isoform appearance, further investigation may reveal an important role for bacteria in FIAF-regulated physiological processes, such as cell differentiation and fat metabolism.

Arabinoxylans and inulin differentially modulate the mucosal and luminal gut microbiota and mucin-degradation in humanized rats.

The endogenous gut microbiota affects the host in many ways. Prebiotics should favour beneficial intestinal microbes and thus improve host health. In this study, we investigated how a novel class of potential prebiotic long-chain arabinoxylans (LC-AX) and the well-established prebiotic inulin (IN) modulate the gut microbiota of humanized rats. Six weeks after axenic rats were inoculated with a human faecal microbiota, their colonic microbiota was similar to this inoculum (∼ 70%), whereas their caecal microbiota was enriched with Verrucomicrobia and Firmicutes concomitant with lower abundance of Bacteroidetes. Moreover, different Bifidobacterium species colonized the lumen (B. adolescentis) and mucus (B. longum and B. bifidum). Both LC-AX and IN increased SCFA levels and induced a shift from acetate towards health-promoting propionate and butyrate respectively. By applying a high-resolution phylogenetic micro-array (HITChip) at the site of fermentation (caecum), IN and LC-AX were shown to stimulate bacterial groups with known butyrate-producers (Roseburia intestinalis, Eubacterium rectale, Anaerostipes caccae) and bifidobacteria (B. longum) respectively. Prebiotic administration also resulted in lower caecal abundances of the mucin-degrading Akkermansia muciniphila and potentially more mucin production by the host. Both factors might explain the increased caecal mucin levels for LC-AX (threefold) and IN (sixfold). These mucins were degraded along the colon, resulting in high faecal abundances of Akkermansia muciniphila for LC-AX and especially IN-treated rats. Finally, the microbial changes caused an adaptation period for the host with less weight gain, after which the host fine-tuned the interaction with this altered microbiota. Our results demonstrate that next to IN, LC-AX are promising prebiotic compounds by stimulating production of health-promoting metabolites by specific microbes in the proximal regions. Further, prebiotic supplementation shifted mucin degradation to distal regions, where mucin-degraders may produce beneficial metabolites (e.g. propionate by Akkermansia muciniphila), so that prebiotics may potentially improve gut health along the entire length of the intestine.

Adjuvant Effect of Orally Applied Preparations Containing Non-Digestible Polysaccharides on Influenza Vaccination in Healthy Seniors: A Double-Blind, Randomised, Controlled Pilot Trial.

Senior individuals can suffer from immunosenescence and novel strategies to bolster the immune response could contribute to healthy ageing. In this double-blind, randomised, controlled pilot trial, we investigated the ability of non-digestible polysaccharide (NPS) preparations to enhance the immune response in a human vaccination model. In total, 239 subjects (aged 50-79 years) were randomised to consume one of five different NPS (yeast ß-glucan (YBG), shiitake ß-glucan (SBG), oat ß-glucan (OBG), arabinoxylan (AX), bacterial exopolysaccharide (EPS)) or control (CTRL) product daily for five weeks. After two weeks of intervention, subjects were vaccinated with seasonal influenza vaccine. The post-vaccination increases in haemagglutination inhibition antibody titres and seroprotection rate against the influenza strains were non-significantly enhanced in the NPS intervention groups compared to CTRL. Specifically, a trend towards a higher mean log2 fold increase was observed in the AX group (uncorrected p = 0.074) combined with a trend for an increased seroprotection rate, AX group (48.7%) compared to CTRL (25.6%) (uncorrected p = 0.057), for the influenza A H1N1 strain. Subjects consuming AX also had a reduced incidence of common colds compared to CTRL (1 vs. 8; p = 0.029 in Fisher exact test). No adverse effects of NPS consumption were reported. The findings of this pilot study warrant further research to study AX as an oral adjuvant to support vaccine efficacy.

Microbial community development in a dynamic gut model is reproducible, colon region specific, and selective for Bacteroidetes and Clostridium cluster IX.

Dynamic, multicompartment in vitro gastrointestinal simulators are often used to monitor gut microbial dynamics and activity. These reactors need to harbor a microbial community that is stable upon inoculation, colon region specific, and relevant to in vivo conditions. Together with the reproducibility of the colonization process, these criteria are often overlooked when the modulatory properties from different treatments are compared. We therefore investigated the microbial colonization process in two identical simulators of the human intestinal microbial ecosystem (SHIME), simultaneously inoculated with the same human fecal microbiota with a high-resolution phylogenetic microarray: the human intestinal tract chip (HITChip). Following inoculation of the in vitro colon compartments, microbial community composition reached steady state after 2 weeks, whereas 3 weeks were required to reach functional stability. This dynamic colonization process was reproducible in both SHIME units and resulted in highly diverse microbial communities which were colon region specific, with the proximal regions harboring saccharolytic microbes (e.g., Bacteroides spp. and Eubacterium spp.) and the distal regions harboring mucin-degrading microbes (e.g., Akkermansia spp.). Importantly, the shift from an in vivo to an in vitro environment resulted in an increased Bacteroidetes/Firmicutes ratio, whereas Clostridium cluster IX (propionate producers) was enriched compared to clusters IV and XIVa (butyrate producers). This was supported by proportionally higher in vitro propionate concentrations. In conclusion, high-resolution analysis of in vitro-cultured gut microbiota offers new insight on the microbial colonization process and indicates the importance of digestive parameters that may be crucial in the development of new in vitro models.

Exploration of isoxanthohumol bioconversion from spent hops into 8-prenylnaringenin using resting cells of Eubacterium limosum.

Hops is an almost unique source of the potent phytoestrogen 8-prenylnaringenin (8-PN). As hops contain only low levels of 8-PN, synthesis may be more attractive than extraction. A strain of the Gram-positive Eubacterium limosum was isolated previously for 8-PN production from more abundant precursor isoxanthohumol (IX) from hops. In this study, spent hops, an industrial side stream from the beer industry, was identified as interesting source of IX. Yet, hop-derived compounds are well-known antibacterial agents and the traces of a large variety of different compounds in spent hops interfered with growth and IX conversion. Critical factors to finally enable bacterial 8-PN production from spent hops, using a food and feed grade medium, were evaluated in this research. The use of bacterial resting cells and complex medium at a pH of 7.8-8 best fulfilled the requirements for 8-PN production and generated a solid basis for development of an economic process.

Development and validation of the Simulator of the Canine Intestinal Microbial Ecosystem (SCIME)1.

Whereas a wide variety of in vitro models have been developed and validated to assess the effect of specific food ingredients on the human gut microbiome, such models have only been developed and applied to a limited extent for companion animals. Since the use of pre- and probiotics to improve gut health is an emerging research topic in the field of companion animals and as dogs are often used as laboratory animals in developing and testing of pharmaceuticals, the current study aimed to establish an adequate canine in vitro model. This consisted of a four-stage reactor composed of a stomach and small intestinal compartment followed by a proximal and distal colon. This semi-continuous gastrointestinal tract model allowed a long-term, region-dependent, and pH-controlled simulation of the colon-associated microbial community of dogs. Upon reaching a functional steady state, the simulated canine microbial community composition proved to be representative of the in vivo situation. Indeed, the predominant bacterial phyla present in the in vitro proximal and distal colon corresponded with the main bacterial phyla detected in the fecal material of the dogs, resulting in an average community composition along the simulated canine gastrointestinal tract of 50.5% Firmicutes, 34.5% Bacteroidetes, 7.4% Fusobacteria, 4.9% Actinobacteria, and 2.7% Proteobacteria. A parallel in vivo-in vitro comparison assessing the effects of fructooligosaccharides (FOS) on the canine microbial community composition showed a consistent stimulation of Lactobacillus concentrations in the in vivo fecal samples as well as in the in vitro canine gut model. Furthermore, the in vitro platform provided additional insights about the prebiotic effect of FOS supplementation of dogs, such as a reduced abundance of Megamonas spp. which are only present in very low abundance in in vivo fecal samples, indicating an interesting application potential of the developed canine in vitro model in research related to gastrointestinal health of dogs.

Cosupplementation of isoflavones, prenylflavonoids, and lignans alters human exposure to phytoestrogen-derived 17beta-estradiol equivalents.

The microbial metabolism of dietary phytoestrogens varies considerably among individuals and influences the final exposure to bioactive compounds. In view of the increasing number of food supplements combining several classes of phytoestrogens, the microbial potential to activate various proestrogens within an individual was evaluated in 3 randomized dietary crossovers. Treatment allocation was based on participants' eligibility (>45% in vitro bioactivation of >or=2 separate proestrogens by fecal cultures; n = 40/100). After a run-in of >or=4 d, participants were given soy-, hop-, and/or flax-based food supplements dosed either separately (SOY: 2.83 mg daidzein aglycone equivalents/supplement, HOP: 1.20 mg isoxanthohumol (IX)/supplement, or FLAX: 2.08 mg secoisolariciresinol (SECO) aglycone equivalents/supplement; reference intervention) or simultaneously (MIX; test intervention) 3 times/d for 5 d, followed by a wash-out period (>or=7 d) and the second intervention. Before and after each (co)supplementation, spot urine and serum were collected. In total, 22 equol, 19 8-prenylnaringenin (8-PN), and 21 enterolactone (ENL) producers completed the SOY+MIX, HOP+MIX, and FLAX+MIX trials, respectively. The microbial bioactivation of daidzein, IX, and SECO, generally decreased upon coincubation in vitro (equol: 4.4%, P = 0.164; 8-PN: 20.5%, P < 0.001; ENL: 44.3%, P < 0.001) and cosupplementation in vivo (equol: 28.3%, P = 0.009; 8-PN: 35.4%, P = 0.107; ENL: 35.9%, P = 0.003). Although the bioavailabilities of total isoflavones, prenylflavonoids, and lignans were not significantly affected upon coadministration, participants were exposed to lower phytoestrogen-derived 17beta-estradiol equivalents. In conclusion, the bioavailability of phytoestrogens, especially when given in mixtures, is subject to high interindividual variation. These findings support the importance of personalized screening when assessing the efficacy of such products and mixtures.

In vitro model to study the modulation of the mucin-adhered bacterial community.

There is an upsurge of interest in gastro-intestinal microbiology to improve the balance between positive and negative commensals. Mucosal bacteria make closer contact with the host than luminal ones and can therefore have a stronger health impact. An in vitro adhesion assay was developed to study the mucin colonization of bacteria from the mixed microbial communities of the Simulator of the Human Intestinal Microbial Ecosystem. Adhesion capacity differed substantially between bacteria and decreased from lactobacilli over fecal coliforms, bifidobacteria, and clostridia to total anaerobes. Lactobacillus rhamnosus GG adhered most selectively. Further, intestinal water lowered adhesion compared to phosphate-buffered saline. By processing the data to an Adhesion-Related Prebiotic Index, it was found that intestinal water stimulated adherence of positive commensals. Arabinoxylans decreased the adhesion capacity matrix independently, whereas inulin had less or no influence. Measurements of bacterial surface tension, surface hydrophobicity, liquid surface tension, and viscosity showed that bacterial adhesion to mucin agar is a matter of both non-specific and specific interactions. The developed methodology can be useful for the characterization of the relevant but barely investigated mucin-associated bacterial community in health and disease (e.g., IBD) as well as for its modulation with functional foods like prebiotics.