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1.
Environ Microbiol ; 25(8): 1484-1504, 2023 08.
Article in English | MEDLINE | ID: mdl-36912501

ABSTRACT

Dietary fibre is a major energy source for the human gut microbiota, but it is unclear to what extent the fibre source and complexity affect microbial growth and metabolite production. Cell wall material and pectin were extracted from five different dicotyledon plant sources, apples, beet leaves, beetroots, carrots and kale, and compositional analysis revealed differences in the monosaccharide composition. Human faecal batch incubations were conducted with 14 different substrates, including the plant extracts, wheat bran and commercially available carbohydrates. Microbial activity was determined for up to 72 h by measuring gas and fermentation acid production, total bacteria (by qPCR) and microbial community composition by 16S rRNA amplicon sequencing. The more complex substrates gave rise to more microbiota variation compared with the pectins. The comparison of different plant organs showed that the leaves (beet leaf and kale) and roots (carrot and beetroot) did not give rise to similar bacterial communities. Rather, the compositional features of the plants, such as high arabinan levels in beet and high galactan levels in carrot, appear to be major predictors of bacterial enrichment on the substrates. Thus, in-depth knowledge on dietary fibre composition should aid the design of diets focused on optimizing the microbiota.


Subject(s)
Gastrointestinal Microbiome , Microbiota , Humans , RNA, Ribosomal, 16S/genetics , RNA, Ribosomal, 16S/metabolism , Dietary Fiber/metabolism , Bacteria , Feces/microbiology , Fermentation , Pectins/metabolism
2.
Br J Nutr ; 130(9): 1521-1536, 2023 11 14.
Article in English | MEDLINE | ID: mdl-36847278

ABSTRACT

Only 6 to 8 % of the UK adults meet the daily recommendation for dietary fibre. Fava bean processing lead to vast amounts of high-fibre by-products such as hulls. Bean hull fortified bread was formulated to increase and diversify dietary fibre while reducing waste. This study assessed the bean hull: suitability as a source of dietary fibre; the systemic and microbial metabolism of its components and postprandial events following bean hull bread rolls. Nine healthy participants (53·9 ± 16·7 years) were recruited for a randomised controlled crossover study attending two 3 days intervention sessions, involving the consumption of two bread rolls per day (control or bean hull rolls). Blood and faecal samples were collected before and after each session and analysed for systemic and microbial metabolites of bread roll components using targeted LC-MS/MS and GC analysis. Satiety, gut hormones, glucose, insulin and gastric emptying biomarkers were also measured. Two bean hull rolls provided over 85 % of the daily recommendation for dietary fibre; but despite being a rich source of plant metabolites (P = 0·04 v. control bread), these had poor systemic bioavailability. Consumption of bean hull rolls for 3 days significantly increased plasma concentration of indole-3-propionic acid (P = 0·009) and decreased faecal concentration of putrescine (P = 0·035) and deoxycholic acid (P = 0·046). However, it had no effect on postprandial plasma gut hormones, bacterial composition and faecal short chain fatty acids amount. Therefore, bean hulls require further processing to improve their bioactives systemic availability and fibre fermentation.


Subject(s)
Fabaceae , Gastrointestinal Hormones , Adult , Humans , Healthy Volunteers , Putrescine , Bread/analysis , Chromatography, Liquid , Cross-Over Studies , Tandem Mass Spectrometry , Dietary Fiber/analysis , Fabaceae/metabolism , Deoxycholic Acid , Blood Glucose/analysis
3.
Am J Respir Crit Care Med ; 205(6): 641-650, 2022 03 15.
Article in English | MEDLINE | ID: mdl-34919021

ABSTRACT

Rationale: In murine models, microbial exposures induce protection from experimental allergic asthma through innate immunity. Objectives: Our aim was to assess the association of early life innate immunity with the development of asthma in children at risk. Methods: In the PASTURE farm birth cohort, innate T-helper cell type 2 (Th2), Th1, and Th17 cytokine expression at age 1 year was measured after stimulation of peripheral blood mononuclear cells with LPS in n = 445 children. Children at risk of asthma were defined based on single-nucleotide polymorphisms at the 17q21 asthma gene locus. Specifically, we used the SNP rs7216389 in the GSDMB gene. Wheeze in the first year of life was assessed by weekly diaries and asthma by questionnaire at age 6 years. Measurements and Main Results: Not all cytokines were detectable in all children after LPS stimulation. When classifying detectability of cytokines by latent class analysis, carrying the 17q21 risk allele rs7216389 was associated with risk of wheeze only in the class with the lowest level of LPS-induced activation: odds ratio (OR), 1.89; 95% confidence interval [CI], 1.13-3.16; P = 0.015. In contrast, in children with high cytokine activation after LPS stimulation, no association of the 17q21 risk allele with wheeze (OR, 0.63; 95% CI, 0.29-1.40; P = 0.258, P = 0.034 for interaction) or school-age asthma was observed. In these children, consumption of unprocessed cow's milk was associated with higher cytokine activation (OR, 3.37; 95% CI, 1.56-7.30; P = 0.002), which was in part mediated by the gut microbiome. Conclusions: These findings suggest that within the 17q21 genotype, asthma risk can be mitigated by activated immune responses after innate stimulation, which is partly mediated by a gut-immune axis.


Subject(s)
Asthma , Chromosomes, Human, Pair 17 , Lipopolysaccharides , Alleles , Animals , Asthma/genetics , Cattle , Cytokines/genetics , Female , Humans , Immunity, Innate , Leukocytes, Mononuclear , Mice , Respiratory Sounds/genetics
4.
Clin Nutr ; 40(8): 5009-5019, 2021 08.
Article in English | MEDLINE | ID: mdl-34364241

ABSTRACT

BACKGROUND & AIMS: Siblings of people with Crohn's disease (CD) share aspects of the disease phenotype (raised faecal calprotectin, altered microbiota), which are markers of risk for their own development of CD. The aim was to determine whether supplementation with prebiotic oligofructose/inulin induces a prebiotic response and impacts the risk phenotype in CD patients and siblings. METHODS: Patients with inactive CD (n = 19, CD activity index <150) and 12 of their unaffected siblings (with calprotectin >50 µg/g) ingested oligofructose/inulin (15 g/day) for three weeks. Faecal microbiota (qPCR), intestinal permeability (lactulose-rhamnose test), blood T cells (flow-cytometry) and calprotectin (ELISA) were measured at baseline and follow-up. RESULTS: Following oligofructose/inulin, calprotectin did not significantly change in patients (baseline mean 537 SD 535 µg/g; follow-up mean 974 SD 1318 µg/g, p = 0.08) or siblings (baseline mean 73 SD 90 µg/g: follow up mean 58 SD 72 µg/g, p = 0.62). Faecal Bifidobacteria and Bifidobacterium longum increased in patients and siblings; Bifidobacterium adolescentis and Roseburia spp. increased only in siblings. Compared with patients, siblings had a greater magnitude change in Bifidobacteria (+14.6% vs +0.4%, p = 0.028), B. adolescentis (+1.1% vs 0.0% p = 0.006) and Roseburia spp. (+1.5% vs -0.1% p = 0.004). Intestinal permeability decreased significantly in patients after oligofructose/inulin to a level that was similar to siblings. Blood T cell abundance reduced in siblings but not patients following oligofructose/inulin. CONCLUSIONS: Oligofructose/inulin supplementation did not significantly impact calprotectin, but the prebiotic effect was more marked in healthy siblings compared with patients with inactive CD and was associated with alterations in other CD risk markers. Future research should focus on dietary intervention, including with prebiotics, in the primary prevention of CD.


Subject(s)
Crohn Disease/microbiology , Crohn Disease/prevention & control , Fructans/administration & dosage , Prebiotics/administration & dosage , Siblings , Adolescent , Adult , Feces/chemistry , Feces/microbiology , Female , Flow Cytometry , Healthy Volunteers , Humans , Intestines/microbiology , Inulin/administration & dosage , Leukocyte L1 Antigen Complex/analysis , Male , Oligosaccharides/administration & dosage , Permeability , Phenotype , Pilot Projects , T-Lymphocytes/microbiology , Young Adult
5.
Nat Med ; 26(11): 1766-1775, 2020 11.
Article in English | MEDLINE | ID: mdl-33139948

ABSTRACT

Growing up on a farm is associated with an asthma-protective effect, but the mechanisms underlying this effect are largely unknown. In the Protection against Allergy: Study in Rural Environments (PASTURE) birth cohort, we modeled maturation using 16S rRNA sequence data of the human gut microbiome in infants from 2 to 12 months of age. The estimated microbiome age (EMA) in 12-month-old infants was associated with previous farm exposure (ß = 0.27 (0.12-0.43), P = 0.001, n = 618) and reduced risk of asthma at school age (odds ratio (OR) = 0.72 (0.56-0.93), P = 0.011). EMA mediated the protective farm effect by 19%. In a nested case-control sample (n = 138), we found inverse associations of asthma with the measured level of fecal butyrate (OR = 0.28 (0.09-0.91), P = 0.034), bacterial taxa that predict butyrate production (OR = 0.38 (0.17-0.84), P = 0.017) and the relative abundance of the gene encoding butyryl-coenzyme A (CoA):acetate-CoA-transferase, a major enzyme in butyrate metabolism (OR = 0.43 (0.19-0.97), P = 0.042). The gut microbiome may contribute to asthma protection through metabolites, supporting the concept of a gut-lung axis in humans.


Subject(s)
Asthma/epidemiology , Butyrates/metabolism , Coenzyme A-Transferases/genetics , Gastrointestinal Microbiome/genetics , Adolescent , Asthma/genetics , Asthma/microbiology , Asthma/pathology , Bacteria/classification , Bacteria/genetics , Bacteria/metabolism , Butyrates/isolation & purification , Child , Feces/chemistry , Female , Humans , Infant , Lung/metabolism , Lung/pathology , Male , RNA, Ribosomal, 16S/genetics
6.
Toxins (Basel) ; 12(10)2020 10 13.
Article in English | MEDLINE | ID: mdl-33066173

ABSTRACT

Mycotoxins are important food contaminants that commonly co-occur with modified mycotoxins such as mycotoxin-glucosides in contaminated cereal grains. These masked mycotoxins are less toxic, but their breakdown and release of unconjugated mycotoxins has been shown by mixed gut microbiota of humans and animals. The role of different bacteria in hydrolysing mycotoxin-glucosides is unknown, and this study therefore investigated fourteen strains of human gut bacteria for their ability to break down masked mycotoxins. Individual bacterial strains were incubated anaerobically with masked mycotoxins (deoxynivalenol-3-ß-glucoside, DON-Glc; nivalenol-3-ß-glucoside, NIV-Glc; HT-2-ß-glucoside, HT-2-Glc; diacetoxyscirpenol-α-glucoside, DAS-Glc), or unconjugated mycotoxins (DON, NIV, HT-2, T-2, and DAS) for up to 48 h. Bacterial growth, hydrolysis of mycotoxin-glucosides and further metabolism of mycotoxins were assessed. We found no impact of any mycotoxin on bacterial growth. We have demonstrated that Butyrivibrio fibrisolvens, Roseburia intestinalis and Eubacterium rectale hydrolyse DON-Glc, HT-2 Glc, and NIV-Glc efficiently and have confirmed this activity in Bifidobacterium adolescentis and Lactiplantibacillus plantarum (DON-Glc only). Prevotella copri and B. fibrisolvens efficiently de-acetylated T-2 and DAS, but none of the bacteria were capable of de-epoxydation or hydrolysis of α-glucosides. In summary we have identified key bacteria involved in hydrolysing mycotoxin-glucosides and de-acetylating type A trichothecenes in the human gut.


Subject(s)
Bacteria/enzymology , Bacterial Proteins/metabolism , Gastrointestinal Microbiome , Intestines/microbiology , Mycotoxins/metabolism , Acetylation , Bacteria/classification , Bacteria/growth & development , Food Microbiology , Glucosides/metabolism , Humans , Hydrolysis , Mycotoxins/adverse effects , Risk Assessment , Substrate Specificity , Time Factors , Trichothecenes/metabolism
7.
mSystems ; 5(5)2020 Sep 08.
Article in English | MEDLINE | ID: mdl-32900872

ABSTRACT

Lactate can be produced by many gut bacteria, but in adults its accumulation in the colon is often an indicator of microbiota perturbation. Using continuous culture anaerobic fermentor systems, we found that lactate concentrations remained low in communities of human colonic bacteria maintained at pH 6.5, even when dl-lactate was infused at 10 or 20 mM. In contrast, lower pH (5.5) led to periodic lactate accumulation following lactate infusion in three fecal microbial communities examined. Lactate accumulation was concomitant with greatly reduced butyrate and propionate production and major shifts in microbiota composition, with Bacteroidetes and anaerobic Firmicutes being replaced by Actinobacteria, lactobacilli, and Proteobacteria Pure-culture experiments confirmed that Bacteroides and Firmicutes isolates were susceptible to growth inhibition by relevant concentrations of lactate and acetate, whereas the lactate-producer Bifidobacterium adolescentis was resistant. To investigate system behavior further, we used a mathematical model (microPop) based on 10 microbial functional groups. By incorporating differential growth inhibition, our model reproduced the chaotic behavior of the system, including the potential for lactate infusion both to promote and to rescue the perturbed system. The modeling revealed that system behavior is critically dependent on the proportion of the community able to convert lactate into butyrate or propionate. Communities with low numbers of lactate-utilizing bacteria are inherently less stable and more prone to lactate-induced perturbations. These findings can help us to understand the consequences of interindividual microbiota variation for dietary responses and microbiota changes associated with disease states.IMPORTANCE Lactate is formed by many species of colonic bacteria, and can accumulate to high levels in the colons of inflammatory bowel disease subjects. Conversely, in healthy colons lactate is metabolized by lactate-utilizing species to the short-chain fatty acids butyrate and propionate, which are beneficial for the host. Here, we investigated the impact of continuous lactate infusions (up to 20 mM) at two pH values (6.5 and 5.5) on human colonic microbiota responsiveness and metabolic outputs. At pH 5.5 in particular, lactate tended to accumulate in tandem with decreases in butyrate and propionate and with corresponding changes in microbial composition. Moreover, microbial communities with low numbers of lactate-utilizing bacteria were inherently less stable and therefore more prone to lactate-induced perturbations. These investigations provide clear evidence of the important role these lactate utilizers may play in health maintenance. These should therefore be considered as potential new therapeutic probiotics to combat microbiota perturbations.

8.
mBio ; 11(4)2020 07 14.
Article in English | MEDLINE | ID: mdl-32665271

ABSTRACT

We investigated the requirement of 15 human butyrate-producing gut bacterial strains for eight B vitamins and the proteinogenic amino acids by a combination of genome sequence analysis and in vitro growth experiments. The Ruminococcaceae species Faecalibacterium prausnitzii and Subdoligranulum variabile were auxotrophic for most of the vitamins and the amino acid tryptophan. Within the Lachnospiraceae, most species were prototrophic for all amino acids and several vitamins, but biotin auxotrophy was widespread. In addition, most of the strains belonging to Eubacterium rectale and Roseburia spp., but few of the other Lachnospiraceae strains, were auxotrophic for thiamine and folate. Synthetic coculture experiments of five thiamine or folate auxotrophic strains with different prototrophic bacteria in the absence and presence of different vitamin concentrations were carried out. This demonstrated that cross-feeding between bacteria does take place and revealed differences in cross-feeding efficiency between prototrophic strains. Vitamin-independent growth stimulation in coculture compared to monococulture was also observed, in particular for F. prausnitzii A2-165, suggesting that it benefits from the provision of other growth factors from community members. The presence of multiple vitamin auxotrophies in the most abundant butyrate-producing Firmicutes species found in the healthy human colon indicates that these bacteria depend upon vitamins supplied from the diet or via cross-feeding from other members of the microbial community.IMPORTANCE Microbes in the intestinal tract have a strong influence on human health. Their fermentation of dietary nondigestible carbohydrates leads to the formation of health-promoting short-chain fatty acids, including butyrate, which is the main fuel for the colonic wall and has anticarcinogenic and anti-inflammatory properties. A good understanding of the growth requirements of butyrate-producing bacteria is important for the development of efficient strategies to promote these microbes in the gut, especially in cases where their abundance is altered. The demonstration of the inability of several dominant butyrate producers to grow in the absence of certain vitamins confirms the results of previous in silico analyses. Furthermore, establishing that strains prototrophic for thiamine or folate (butyrate producers and non-butyrate producers) were able to stimulate growth and affect the composition of auxotrophic synthetic communities suggests that the provision of prototrophic bacteria that are efficient cross feeders may stimulate butyrate-producing bacteria under certain in vivo conditions.


Subject(s)
Bacteria/genetics , Butyrates/metabolism , Fermentation , Microbiota , Vitamins/biosynthesis , Bacteria/metabolism , Clostridiales/genetics , Clostridiales/physiology , Colon/microbiology , Faecalibacterium prausnitzii/genetics , Faecalibacterium prausnitzii/physiology , Humans , Ruminococcus/genetics , Ruminococcus/physiology
9.
Environ Microbiol ; 22(6): 2150-2164, 2020 06.
Article in English | MEDLINE | ID: mdl-32141148

ABSTRACT

A clone encoding carboxymethyl cellulase activity was isolated during functional screening of a human gut metagenomic library using Lactococcus lactis MG1363 as heterologous host. The insert carried a glycoside hydrolase family 9 (GH9) catalytic domain with sequence similarity to a gene from Coprococcus eutactus ART55/1. Genome surveys indicated a limited distribution of GH9 domains among dominant human colonic anaerobes. Genomes of C. eutactus-related strains harboured two GH9-encoding and four GH5-encoding genes, but the strains did not appear to degrade cellulose. Instead, they grew well on ß-glucans and one of the strains also grew on galactomannan, galactan, glucomannan and starch. Coprococcus comes and Coprococcus catus strains did not harbour GH9 genes and were not able to grow on ß-glucans. Gene expression and proteomic analysis of C. eutactus ART55/1 grown on cellobiose, ß-glucan and lichenan revealed similar changes in expression in comparison to glucose. On ß-glucan and lichenan only, one of the four GH5 genes was strongly upregulated. Growth on glucomannan led to a transcriptional response of many genes, in particular a strong upregulation of glycoside hydrolases involved in mannan degradation. Thus, ß-glucans are a major growth substrate for species related to C. eutactus, with glucomannan and galactans alternative substrates for some strains.


Subject(s)
Clostridiales/growth & development , Gastrointestinal Microbiome , beta-Glucans , Bacterial Proteins/genetics , Clostridiales/genetics , Gene Expression , Glucans/pharmacology , Glycoside Hydrolases/genetics , Humans , Proteomics
10.
Sci Rep ; 8(1): 15566, 2018 10 22.
Article in English | MEDLINE | ID: mdl-30349136

ABSTRACT

Dietary fibers (DF) can prevent obesity in rodents fed a high-fat diet (HFD). Their mode of action is not fully elucidated, but the gut microbiota have been implicated. This study aimed to identify the effects of seven dietary fibers (barley beta-glucan, apple pectin, inulin, inulin acetate ester, inulin propionate ester, inulin butyrate ester or a combination of inulin propionate ester and inulin butyrate ester) effective in preventing diet-induced obesity and links to differences in cecal bacteria and host gene expression. Mice (n = 12) were fed either a low-fat diet (LFD), HFD or a HFD supplemented with the DFs, barley beta-glucan, apple pectin, inulin, inulin acetate ester, inulin propionate ester, inulin butyrate ester or a combination of inulin propionate ester and inulin butyrate ester for 8 weeks. Cecal bacteria were determined by Illumina MiSeq sequencing of 16S rRNA gene amplicons. Host responses, body composition, metabolic markers and gene transcription (cecum and liver) were assessed post intervention. HFD mice showed increased adiposity, while all of the DFs prevented weight gain. DF specific differences in cecal bacteria were observed. Results indicate that diverse DFs prevent weight gain on a HFD, despite giving rise to different cecal bacteria profiles. Conversely, common host responses to dietary fiber observed are predicted to be important in improving barrier function and genome stability in the gut, maintaining energy homeostasis and reducing HFD induced inflammatory responses in the liver.


Subject(s)
Dietary Fiber/therapeutic use , Gastrointestinal Microbiome , Obesity/diet therapy , Animals , Cecum/metabolism , Cecum/microbiology , Dietary Fiber/administration & dosage , Liver/metabolism , Male , Mice , Mice, Inbred C57BL , Obesity/microbiology
11.
ISME J ; 12(2): 610-622, 2018 02.
Article in English | MEDLINE | ID: mdl-29192904

ABSTRACT

The diet provides carbohydrates that are non-digestible in the upper gut and are major carbon and energy sources for the microbial community in the lower intestine, supporting a complex metabolic network. Fermentation produces the short-chain fatty acids (SCFAs) acetate, propionate and butyrate, which have health-promoting effects for the human host. Here we investigated microbial community changes and SCFA production during in vitro batch incubations of 15 different non-digestible carbohydrates, at two initial pH values with faecal microbiota from three different human donors. To investigate temporal stability and reproducibility, a further experiment was performed 1 year later with four of the carbohydrates. The lower pH (5.5) led to higher butyrate and the higher pH (6.5) to more propionate production. The strongest propionigenic effect was found with rhamnose, followed by galactomannans, whereas fructans and several α- and ß-glucans led to higher butyrate production. 16S ribosomal RNA gene-based quantitative PCR analysis of 22 different microbial groups together with 454 sequencing revealed significant stimulation of specific bacteria in response to particular carbohydrates. Some changes were ascribed to metabolite cross-feeding, for example, utilisation by Eubacterium hallii of 1,2-propanediol produced from fermentation of rhamnose by Blautia spp. Despite marked inter-individual differences in microbiota composition, SCFA production was surprisingly reproducible for different carbohydrates, indicating a level of functional redundancy. Interestingly, butyrate formation was influenced not only by the overall % butyrate-producing bacteria in the community but also by the initial pH, consistent with a pH-dependent shift in the stoichiometry of butyrate production.


Subject(s)
Bacteria/metabolism , Dietary Carbohydrates/metabolism , Fatty Acids, Volatile/metabolism , Microbiota , Bacteria/genetics , Bacteria/isolation & purification , Butyrates/metabolism , Eubacterium/metabolism , Feces/microbiology , Fermentation , Galactose/analogs & derivatives , Humans , Mannans/metabolism , Propionates/metabolism , Reproducibility of Results , Rhamnose/metabolism
12.
Appl Environ Microbiol ; 84(2)2018 01 15.
Article in English | MEDLINE | ID: mdl-29101203

ABSTRACT

Mycotoxin contamination of cereal grains causes well-recognized toxicities in animals and humans, but the fate of plant-bound masked mycotoxins in the gut is less well understood. Masked mycotoxins have been found to be stable under conditions prevailing in the small intestine but are rapidly hydrolyzed by fecal microbiota. This study aims to assess the hydrolysis of the masked mycotoxin deoxynivalenol-3-glucoside (DON3Glc) by the microbiota of different regions of the porcine intestinal tract. Intestinal digesta samples were collected from the jejunum, ileum, cecum, colon, and feces of 5 pigs and immediately frozen under anaerobic conditions. Sample slurries were prepared in M2 culture medium, spiked with DON3Glc or free deoxynivalenol (DON; 2 nmol/ml), and incubated anaerobically for up to 72 h. Mycotoxin concentrations were determined using liquid chromatography-tandem mass spectrometry, and the microbiota composition was determined using a quantitative PCR methodology. The jejunal microbiota hydrolyzed DON3Glc very slowly, while samples from the ileum, cecum, colon, and feces rapidly and efficiently hydrolyzed DON3Glc. No further metabolism of DON was observed in any sample. The microbial load and microbiota composition in the ileum were significantly different from those in the distal intestinal regions, whereas those in the cecum, colon and feces did not differ.IMPORTANCE Results from this study clearly demonstrate that the masked mycotoxin DON3Glc is hydrolyzed efficiently in the distal small intestine and large intestine of pigs. Once DON is released, toxicity and absorption in the distal intestinal tract likely occur in vivo This study further supports the need to include masked metabolites in mycotoxin risk assessments and regulatory actions for feed and food.


Subject(s)
Gastrointestinal Microbiome/drug effects , Gastrointestinal Microbiome/physiology , Glucosides/pharmacology , Intestines/microbiology , Mycotoxins/pharmacology , Trichothecenes/metabolism , Trichothecenes/pharmacology , Anaerobiosis , Animals , Batch Cell Culture Techniques , Edible Grain/chemistry , Feces/chemistry , Feces/microbiology , Food Contamination , Gastrointestinal Microbiome/genetics , Humans , Hydrolysis , Intestines/anatomy & histology , Jejunum/microbiology , Jejunum/physiology , Mycotoxins/analysis , Mycotoxins/metabolism , Mycotoxins/toxicity , Polymerase Chain Reaction , Swine , Trichothecenes/analysis
13.
J Agric Food Chem ; 66(2): 485-497, 2018 Jan 17.
Article in English | MEDLINE | ID: mdl-29236499

ABSTRACT

Human colonic bacteria have an important impact on the biotransformation of flavonoid glycosides and their conversion can result in the formation of bioactive compounds. However, information about the microbial conversion of complex glycosylated flavonoids and the impact on the gut microbiota are still limited. In this study, in vitro fermentations with selected flavonoid O- and C-glycosides and three different fecal samples were performed. As a result, all flavonoid glycosides were metabolized via their aglycones yielding smaller substances. Main metabolites were 3-(4-hydroxyphenyl)propionic acid, 3-phenylpropionic acid, and phenylacetic acid. Differences in the metabolite formation due to different time courses between the donors were determined. Therefore, from all fermentations, the ones with a specific donor were always slower resulting in a lower number of metabolites compared to the others. For example, tiliroside was totally degraded from 0 h (105 ± 13.2 µM) within the first 24 h, while in the fermentations with fecal samples from other donors, tiliroside (107 ± 52.7 µM at 0 h) was not detected after 7 h anymore. In general, fermentation rates of C-glycosides were slower compared to the fermentation rates of O-glycosides. The O-glycoside tiliroside was degraded within 4 h while the gut microbiota converted the C-glycoside vitexin within 13 h. However, significant changes (p < 0.05) in the microbiota composition and short chain fatty acid levels as products of carbohydrate fermentation were not detected between incubations with different phenolic compounds. Therefore, microbiota diversity was not affected and a significant prebiotic effect of phenolic compounds cannot be assigned to flavonoid glycosides in food-relevant concentrations.


Subject(s)
Apigenin/metabolism , Feces/chemistry , Gastrointestinal Microbiome , Kaempferols/metabolism , Phenols/metabolism , Apigenin/chemistry , Bacteria/genetics , Bacteria/isolation & purification , Bacteria/metabolism , Feces/microbiology , Humans , Intestinal Mucosa/metabolism , Intestines/microbiology , Kaempferols/chemistry , Molecular Structure , Phenols/chemistry
14.
Food Res Int ; 100(Pt 3): 375-384, 2017 10.
Article in English | MEDLINE | ID: mdl-28964360

ABSTRACT

The almost forgotten crop amaranth has gained renewed interest in recent years due to its immense nutritive potential. Health beneficial effects of certain plants are often attributed to secondary plant metabolites such as phenolic compounds. As these compounds undergo significant metabolism after consumption and are in most cases not absorbed very well, it is important to gain knowledge about absorption, biotransformation, and further metabolism in the human body. Whilst being hardly found in other edible plants, caffeoylisocitric acid represents the most abundant low molecular weight phenolic compound in many leafy amaranth species. Given that this may be a potentially bioactive compound, gastrointestinal microbial degradation of this substance was investigated in the present study by performing in vitro fermentation tests using three different fecal samples as inocula. The (phenolic) metabolites were analyzed using high-performance liquid chromatography/electrospray ionization tandem mass spectrometry (HPLC-ESI-MS/MS). Furthermore, quantitative polymerase chain reaction (qPCR) analyses were carried out to study the influence on the microbiome and its composition. The in vitro fermentations led to different metabolite profiles depending on the specific donor. For example, the metabolite 3-(4-hydroxyphenyl)propionic acid was observed in one fermentation as the main metabolite, whereas 3-(3-hydroxyphenyl)propionic acid was identified in the other fermentations as important. A significant change in selected microorganisms of the gut microbiota however was not detected. In conclusion, caffeoylisocitric acid from amaranth, which is a source of several esterified phenolic acids in addition to chlorogenic acid, can be metabolized by the human gut microbiota, but the metabolites produced vary between individuals.


Subject(s)
Amaranthus/metabolism , Caffeic Acids/metabolism , Chlorogenic Acid/metabolism , Gastrointestinal Microbiome/physiology , Isocitrates/metabolism , Chromatography, High Pressure Liquid , Feces/microbiology , Humans , In Vitro Techniques , Polymerase Chain Reaction , Reference Values , Spectrometry, Mass, Electrospray Ionization
15.
Gastroenterology ; 153(4): 936-947, 2017 10.
Article in English | MEDLINE | ID: mdl-28625832

ABSTRACT

BACKGROUND & AIMS: Dietary restriction of fermentable carbohydrates (a low FODMAP diet) has been reported to reduce symptoms in some patients with irritable bowel syndrome (IBS). We performed a randomized, placebo-controlled study to determine its effects on symptoms and the fecal microbiota in patients with IBS. METHODS: We performed a 2×2 factorial trial of 104 patients with IBS (18-65 years old), based on the Rome III criteria, at 2 hospitals in the United Kingdom. Patients were randomly assigned (blinded) to groups given counselling to follow a sham diet or diet low in FODMAPs for 4 weeks, along with a placebo or multistrain probiotic formulation, resulting in 4 groups (27 receiving sham diet/placebo, 26 receiving sham diet/probiotic, 24 receiving low FODMAP diet /placebo, and 27 receiving low FODMAP diet/probiotic). The sham diet restricted a similar number of staple and non-staple foods as the low FODMAP diet; the diets had similar degrees of difficulty to follow. Dietary counselling was given to patients in all groups and data on foods eaten and compliance were collected. The incidence and severity of 15 gastrointestinal symptoms and overall symptoms were measured daily for 7 days before the study period; along with stool frequency and consistency. At baseline, global and individual symptoms were measured, along with generic and disease-specific health-related quality of life, using standard scoring systems. All data were collected again at 4 weeks, and patients answered questions about adequate symptom relief. Fecal samples were collected at baseline and after 4 weeks and analyzed by quantitative PCR and 16S rRNA sequencing. The co-primary endpoints were adequate relief of symptoms and stool Bifidobacterium species abundance at 4 weeks. RESULTS: There was no significant interaction between the interventions in adequate relief of symptoms (P = .52) or Bifidobacterium species (P = .68). In the intention-to-treat analysis, a higher proportion of patients in the low FODMAP diet had adequate symptom relief (57%) than in the sham diet group (38%), although the difference was not statistically significant (P = .051). In the per-protocol analysis, a significantly higher proportion of patients on the low FODMAP diet had adequate symptom relief (61%) than in the sham diet group (39%) (P = .042). Total mean IBS-Severity Scoring System score was significantly lower for patients on the low FODMAP diet (173 ± 95) than the sham diet (224 ± 89) (P = .001), but not different between those given probiotic (207 ± 98) or placebo (192 ± 93) (P = .721) Abundance of Bifidobacterium species was lower in fecal samples from patients on the low FODMAP diet (8.8 rRNA genes/g) than patients on the sham diet (9.2 rRNA genes/g) (P = .008), but higher in patients given probiotic (9.1 rRNA genes/g) than patients given placebo (8.8 rRNA genes/g) (P = .019). There was no effect of the low FODMAP diet on microbiota diversity in fecal samples. CONCLUSIONS: In a placebo-controlled study of patients with IBS, a low FODMAP diet associates with adequate symptom relief and significantly reduced symptom scores compared with placebo. It is not clear whether changes resulted from collective FODMAP restriction or removal of a single component, such as lactose. Co-administration of the multistrain probiotic increased numbers of Bifidobacterium species, compared with placebo, and might be given to restore these bacteria to patients on a low FODMAP diet. Trial registration no: ISRCTN02275221.


Subject(s)
Bifidobacterium/growth & development , Diet, Carbohydrate-Restricted , Dietary Carbohydrates/adverse effects , Gastrointestinal Microbiome , Intestines/microbiology , Irritable Bowel Syndrome/diet therapy , Probiotics/therapeutic use , Adult , Bifidobacterium/classification , Bifidobacterium/genetics , Combined Modality Therapy , Dietary Carbohydrates/metabolism , Feces/microbiology , Female , Fermentation , Humans , Irritable Bowel Syndrome/diagnosis , Irritable Bowel Syndrome/microbiology , London , Male , Middle Aged , Quality of Life , Remission Induction , Ribotyping , Severity of Illness Index , Surveys and Questionnaires , Time Factors , Treatment Outcome , Young Adult
16.
Microbiome ; 4(1): 61, 2016 11 25.
Article in English | MEDLINE | ID: mdl-27884202

ABSTRACT

BACKGROUND: Sequencing-based analysis has become a well-established approach to deciphering the composition of the gut microbiota. However, due to the complexity of accessing sufficient material from colonoscopic biopsy samples, most studies have focused on faecal microbiota analysis, even though it is recognised that differences exist between the microbial composition of colonic biopsies and faecal samples. We determined the suitability of colonic lavage samples to see if it had comparable microbial diversity composition to colonic biopsies as they are without the limitations associated with sample size. We collected paired colonic biopsies and lavage samples from subjects who were attending for colorectal cancer screening colonoscopy. RESULTS: Next-generation sequencing and qPCR validation were performed with multiple bioinformatics analyses to determine the composition and predict function of the microbiota. Colonic lavage samples contained significantly higher numbers of operational taxonomic units (OTUs) compared to corresponding biopsy samples, however, diversity and evenness between lavage and biopsy samples were similar. The differences seen were driven by the presence of 12 OTUs which were in higher relative abundance in biopsies and were either not present or in low relative abundance in lavage samples, whilst a further 3 OTUs were present in higher amounts in the lavage samples compared to biopsy samples. However, predicted functional community profiling based on 16S ribosomal ribonucleic acid (rRNA) data indicated minimal differences between sample types. CONCLUSIONS: We propose that colonic lavage samples provide a relatively accurate representation of biopsy microbiota composition and should be considered where biopsy size is an issue.


Subject(s)
Bacteria/classification , Biopsy/methods , Colon/microbiology , Gastrointestinal Microbiome/genetics , Intestinal Mucosa/microbiology , Therapeutic Irrigation/methods , Bacteria/genetics , Base Sequence , Colonoscopy/methods , Computational Biology/methods , Feces/microbiology , High-Throughput Nucleotide Sequencing , Humans , RNA, Ribosomal, 16S/genetics , Sequence Analysis, DNA
17.
ISME J ; 8(11): 2218-30, 2014 Nov.
Article in English | MEDLINE | ID: mdl-24763370

ABSTRACT

There is growing interest in understanding how diet affects the intestinal microbiota, including its possible associations with systemic diseases such as metabolic syndrome. Here we report a comprehensive and deep microbiota analysis of 14 obese males consuming fully controlled diets supplemented with resistant starch (RS) or non-starch polysaccharides (NSPs) and a weight-loss (WL) diet. We analyzed the composition, diversity and dynamics of the fecal microbiota on each dietary regime by phylogenetic microarray and quantitative PCR (qPCR) analysis. In addition, we analyzed fecal short chain fatty acids (SCFAs) as a proxy of colonic fermentation, and indices of insulin sensitivity from blood samples. The diet explained around 10% of the total variance in microbiota composition, which was substantially less than the inter-individual variance. Yet, each of the study diets induced clear and distinct changes in the microbiota. Multiple Ruminococcaceae phylotypes increased on the RS diet, whereas mostly Lachnospiraceae phylotypes increased on the NSP diet. Bifidobacteria decreased significantly on the WL diet. The RS diet decreased the diversity of the microbiota significantly. The total 16S ribosomal RNA gene signal estimated by qPCR correlated positively with the three major SCFAs, while the amount of propionate specifically correlated with the Bacteroidetes. The dietary responsiveness of the individual's microbiota varied substantially and associated inversely with its diversity, suggesting that individuals can be stratified into responders and non-responders based on the features of their intestinal microbiota.


Subject(s)
Intestines/microbiology , Microbiota , Obesity/diet therapy , Obesity/microbiology , Adult , Aged , Bacteria/classification , Bacteria/genetics , Bacteria/isolation & purification , Bacteria/metabolism , Cross-Over Studies , Diet, Reducing , Fatty Acids, Volatile/analysis , Feces/chemistry , Feces/microbiology , Fermentation , Humans , Male , Metabolic Syndrome/diet therapy , Metabolic Syndrome/microbiology , Middle Aged , Phylogeny
18.
Gut ; 63(10): 1578-86, 2014 Oct.
Article in English | MEDLINE | ID: mdl-24398881

ABSTRACT

OBJECTIVE: Crohn's disease (CD) is associated with intestinal dysbiosis, altered blood T cell populations, elevated faecal calprotectin (FC) and increased intestinal permeability (IP). CD-associated features present in siblings (increased risk of CD) but not in healthy controls, provide insight into early CD pathogenesis. We aimed to (1) Delineate the genetic, immune and microbiological profile of patients with CD, their siblings and controls and (2) Determine which factors discriminate between groups. DESIGN: Faecal microbiology was analysed by quantitative PCR targeting 16S ribosomal RNA, FC by ELISA, blood T cell phenotype by flow cytometry and IP by differential lactulose-rhamnose absorption in 22 patients with inactive CD, 21 of their healthy siblings and 25 controls. Subject's genotype relative risk was determined by Illumina Immuno BeadChip. RESULTS: Strikingly, siblings shared aspects of intestinal dysbiosis with patients with CD (lower concentrations of Faecalibacterium prausnitzii (p=0.048), Clostridia cluster IV (p=0.003) and Roseburia spp. (p=0.09) compared with controls). As in CD, siblings demonstrated a predominance of memory T cells (p=0.002) and elevated naïve CD4 T cell ß7 integrin expression (p=0.01) compared with controls. FC was elevated (>50 µg/g) in 8/21 (38%) siblings compared with 2/25 (8%) controls (p=0.028); whereas IP did not differ between siblings and controls. Discriminant function analysis determined that combinations of these factors significantly discriminated between groups (χ(2)=80.4, df=20, p<0.001). Siblings were separated from controls by immunological and microbiological variables. CONCLUSIONS: Healthy siblings of patients with CD manifest immune and microbiological abnormalities associated with CD distinct from their genotype-related risk and provide an excellent model in which to investigate early CD pathogenesis.


Subject(s)
Crohn Disease/immunology , Crohn Disease/microbiology , Dysbiosis/immunology , Dysbiosis/microbiology , Intestinal Mucosa/microbiology , Microbiota , T-Lymphocytes/immunology , Adolescent , Adult , Enzyme-Linked Immunosorbent Assay , Feces/microbiology , Female , Genotype , Humans , Immunophenotyping , Male , Siblings , United Kingdom , Young Adult
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