Roseburia Intestinalis Research Articles

Roseburia intestinalis inhibits interleukin‑17 excretion and promotes regulatory T cells differentiation in colitis.

Roseburia intestinalis (R. intestinalis) is one of the dominant intestinal bacterial microbiota and is decreased in patients with inflammatory bowel disease (IBD). It helps protect colonic mucosa against the development of inflammation and subsequent IBD, however its underlying mechanisms are unclear. The aim of the present study was to evaluate the anti-inflammatory properties of R. intestinalis in vitro and in an animal model of IBD. The effects of R. intestinalis on disease activity index (DAI) scores, intestinal pathology, the expression of interleukin (IL)-17 and the frequency of CD4+CD25+Foxp3+ regulatory T cells (Treg) were evaluated in vivo in a model of 2,4,6-trinitrobenzenesulfonic acid solution (TNBS)-induced colitis. Compared with the control group, TNBS-treated mice had significantly higher secretion of IL-17, higher DAI scores, a lower ratio of Treg, reduced colon lengths and higher histological scores for colon inflammation. The administration of R. intestinalis significantly downregulated the expression of IL-17, increased the ratio of Treg and ameliorated the high DAI scores and the pathological signs of inflammation in the colon compared with mice treated with TNBS alone. Gene expression profiling was also used to detect the expression of IL-17 in human IBD and healthy control specimens. To extend these findings to an in vitro model of inflammation the human colon epithelial cell line NCM460 was stimulated with lipopolysaccharide (LPS) to induce inflammation and co-cultured with R. intestinalis and changes in IL-17 expression were evaluated. R. intestinalis inhibited the LPS-induced secretion of IL-17 by NCM460 cells. In conclusion, these results demonstrate that R. intestinalis inhibits IL-17 secretion and promotes Treg differentiation in colitis, suggesting that R. intestinalis could be of potential use in the treatment of IBD.

Enteric Delivery of Regenerating Family Member 3 alpha Alters the Intestinal Microbiota and Controls Inflammation in Mice With Colitis

Paneth cell dysfunction causes deficiencies in intestinal C-type lectins and antimicrobial peptides, which leads to dysbiosis of the intestinal microbiota, alters the mucosal barrier, and promotes development of inflammatory bowel diseases. We investigated whether transgenic (TG) expression of the human regenerating family member 3 alpha gene (REG3A) alters the fecal microbiota and affects development of colitis in mice. We performed studies with C57BL/6 mice that express human regenerating family member 3 alpha (hREG3A) in hepatocytes, via the albumin gene promoter. In these mice, hREG3A travels via the bile to the intestinal lumen. Some mice were given dextran sodium sulfate (DSS) to induce colitis. Feces were collected from mice and the composition of the microbiota was analyzed by 16S ribosomal RNA sequencing. The fecal microbiome was also analyzed from mice that express only 1 copy of human REG3A transgene but were fed feces from control mice (not expressing hREG3A) as newborns. Mice expressing hREG3A were monitored for DSS-induced colitis after cohousing or feeding feces from control mice. Colitis was induced in another set of control and hREG3A-TG mice by administration of trinitrobenzene sulfonic acid; some mice were given intrarectal injections of the hREG3A protein. Colon tissues were collected from mice and analyzed by histology and immunohistochemistry to detect mucin 2, as well as by 16S ribosomal RNA fluorescence in situ hybridization, transcriptional analyses, and quantitative polymerase chain reaction. We measured levels of reactive oxygen species (ROS) in bacterial cultures and fecal microbiota using 2',7'-dichlorofluorescein diacetate and flow cytometry. The fecal microbiota of mice that express hREG3A had a significant shift in composition, compared with control mice, with enrichment of Clostridiales (Ruminococcaceae, Lachnospiraceae) and depletion of Bacteroidetes (Prevotellaceae); the TG mice developed less-severe colitis following administration of DSS than control mice, associated with preserved gut barrier integrity and reduced bacterial translocation, epithelial inflammation, and oxidative damage. A similar shift in the composition of the fecal microbiota occurred after a few months in TG mice heterozygous for REG3A that harbored a wild-type maternal microbiota at birth; these mice developed less-severe forms of colitis following DSS administration. Cohoused and germ-free mice fed feces from REG3A-TG mice and given DSS developed less-severe forms of colitis and had reduced lipopolysaccharide activation of the toll-like receptor 4 and increased survival times compared with mice not fed feces from REG3A-TG mice. REG3A TG mice developed only mild colonic inflammation after exposure to 2,4,6-trinitrobenzene sulfonic acid, compared with control mice. Control mice given intrarectal hREG3A and exposed to 2,4,6-trinitrobenzene sulfonic acid showed less colon damage and inflammation than mice not given intrarectal hREG3A. Fecal samples from REG3A-TG mice had lower levels of ROS than feces from control mice during DSS administration. Addition of hREG3A to bacterial cultures reduced levels of ROS and increased survival of oxygen-sensitive commensal bacteria (Faecalibacterium prausnitzii and Roseburia intestinalis). Mice with hepatocytes that express hREG3A, which travels to the intestinal lumen, are less sensitive to colitis than control mice. We found hREG3A to alter the colonic microbiota by decreasing levels of ROS. Fecal microbiota from REG3A-TG mice protect non-TG mice from induction of colitis. These findings indicate a role for reduction of oxidative stress in preserving the gut microbiota and its ability to prevent inflammation.

Circulating and Tissue-Resident CD4+ T Cells With Reactivity to Intestinal Microbiota Are Abundant in Healthy Individuals and Function Is Altered During Inflammation

Background & AimsInteractions between commensal microbes and the immune system are tightly regulated and maintain intestinal homeostasis, but little is known about these interactions in humans. We investigated responses of human CD4+ T cells to the intestinal microbiota. We measured the abundance of T cells in circulation and intestinal tissues that respond to intestinal microbes and determined their clonal diversity. We also assessed their functional phenotypes and effects on intestinal resident cell populations, and studied alterations in microbe-reactive T cells in patients with chronic intestinal inflammation.MethodsWe collected samples of peripheral blood mononuclear cells and intestinal tissues from healthy individuals (controls, n = 13−30) and patients with inflammatory bowel diseases (n = 119; 59 with ulcerative colitis and 60 with Crohn’s disease). We used 2 independent assays (CD154 detection and carboxy-fluorescein succinimidyl ester dilution assays) and 9 intestinal bacterial species (Escherichia coli, Lactobacillus acidophilus, Bifidobacterium animalis subsp lactis, Faecalibacterium prausnitzii, Bacteroides vulgatus, Roseburia intestinalis, Ruminococcus obeum, Salmonella typhimurium, and Clostridium difficile) to quantify, expand, and characterize microbe-reactive CD4+ T cells. We sequenced T-cell receptor Vβ genes in expanded microbe-reactive T-cell lines to determine their clonal diversity. We examined the effects of microbe-reactive CD4+ T cells on intestinal stromal and epithelial cell lines. Cytokines, chemokines, and gene expression patterns were measured by flow cytometry and quantitative polymerase chain reaction.ResultsCirculating and gut-resident CD4+ T cells from controls responded to bacteria at frequencies of 40−4000 per million for each bacterial species tested. Microbiota-reactive CD4+ T cells were mainly of a memory phenotype, present in peripheral blood mononuclear cells and intestinal tissue, and had a diverse T-cell receptor Vβ repertoire. These cells were functionally heterogeneous, produced barrier-protective cytokines, and stimulated intestinal stromal and epithelial cells via interleukin 17A, interferon gamma, and tumor necrosis factor. In patients with inflammatory bowel diseases, microbiota-reactive CD4+ T cells were reduced in the blood compared with intestine; T-cell responses that we detected had an increased frequency of interleukin 17A production compared with responses of T cells from blood or intestinal tissues of controls.ConclusionsIn an analysis of peripheral blood mononuclear cells and intestinal tissues from patients with inflammatory bowel diseases vs controls, we found that reactivity to intestinal bacteria is a normal property of the human CD4+ T-cell repertoire, and does not necessarily indicate disrupted interactions between immune cells and the commensal microbiota. T-cell responses to commensals might support intestinal homeostasis, by producing barrier-protective cytokines and providing a large pool of T cells that react to pathogens.

Abstract 238: Evidence of early colorectal cancer risk and prevention pathways in the fecal microbiome of colonoscopy patients: associations with diet and circulating adipocytokines

Abstract Accumulating evidence suggests that the gut microbiome’s role in early colorectal cancer etiology extends beyond the pro-carcinogenic activities of specific pathogens and is largely influenced by the wider microbial community of commensal bacteria. To identify early microbiome-related pathways and potential dietary intervention targets, we conducted an epidemiologic study among cancer-free colonoscopy patients at known and varied risk of colorectal neoplasia. Sporadic patients undergoing colonoscopy screening provided consent and fasting blood. Polyps, if found, were removed at colonoscopy and ~1 month later (prior studies show the effect of the colonoscopy prep dissipates within this time period), eligible patients provided a stool sample via mail kit (n=47). Patients completed comprehensive dietary assessments and clinicopathologic factors, including screening history, were abstracted from electronic medical records. We characterized the fecal microbiome via whole genome shotgun sequencing (Illumina HiSeq) and circulating blood adipocytokines via Luminex multiplex assays (Millipore). Majority of the patients were female and recently diagnosed with precancerous polyps, primarily tubular and sessile serrated adenomas. Polyp patients were more likely to be obese and to not consume alcohol, but otherwise similar by age, diet composition, and other risk factors to patients with normal colonoscopy findings. We observed no effects of time interval between colonoscopy and fecal sample collection, or of recent polyp diagnosis/removal, on microbial alpha or beta diversity. Correlation networks between species revealed that Faecalibacterium prausnitzii inversely correlated with E. coli, the most abundant species in our sample, and co-occurred with other short chain fatty acid-metabolizing bacteria, including Anaerostipes hadrus and Roseburia hominis, known butyrate-producers. Among recurrent patients (n=14), Bacteroides fragilis co-occurred with Bilophila wadsworthia (r=0.6; P=0.02). Higher intake of fiber and/or overall diet quality, as defined by the Healthy Eating Index, was associated with several bacteria largely linked to butyrate production (e.g., Bifidobacterium animalis, F. prausnitzii, Roseburia intestinalis, Coprococcus eutactus, Eubacterium eligens). These same dietary factors were inversely correlated with Blautia hydrogenotrophica, an acetogen, and other bacteria implicated in inflammation and colorectal cancer (e.g., B. fragilis). Biologically plausible associations between microbial (functional gene content) pathways important to cancer risk (e.g., one-carbon, lysine, carbohydrate and fatty acid metabolism) with dietary factors and circulating adipocytokines involved in immunity, inflammation, and glucose metabolism support the functionality of these diet-microbe relationships. Citation Format: Carrie R. Daniel, Kristi L. Hoffman, G S. Raju, Samir M. Hanash, Diane S. Hutchinson, Nadim J. Ajami, Richard G. Fowler, Gladys J. Browman, Akhil Sood, Paul Scheet, Xifeng Wu, Joseph F. Petrosino. Evidence of early colorectal cancer risk and prevention pathways in the fecal microbiome of colonoscopy patients: associations with diet and circulating adipocytokines [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 238. doi:10.1158/1538-7445.AM2017-238

Effect of dietary supplementation with Lactobacillus acidophilus D2/CSL (CECT 4529) on caecum microbioma and productive performance in broiler chickens.

This study examines the effects of the dietary supplementation with Lactobacillus acidophilus D2/CSL (CECT 4529) (LA) on productive performances, incidence of foot pad dermatitis and caecum microbioma in broiler chickens. A total of 1,100 one-day old male Ross 308 chicks were divided into 2 groups of 16 replicates with 25 birds each and reared from 1–41 d. One group was fed a basal diet (CON) and the other group the same diet supplemented with LA. Caecum contents were collected from 4 selected birds at day one and 5 selected birds at the end of the rearing period. Then, they were submitted to DNA extraction and whole DNA shotgun metagenomic sequencing. Overall, the LA supplementation produced a significant beneficial effect on body weight gain between 15–28 d and improved feed conversion rate in the overall period. On the contrary, litter moisture, pH and incidence of the foot pad lesions were not affected by LA. Birds treated with LA showed a lower occurrence of pasty vent at both 14 and 28 d. At the end of the rearing period, Lachanospiraceae were significantly higher in LA birds in comparison to CON (17.07 vs 14.39%; P = 0.036). Moreover, Ruminococcus obeum, Clostridium clostridioforme, Roseburia intestinalis, Lachnospiraceae bacterium 14-2T and Coprococcus eutactus were significantly higher in LA birds in comparison to CON. The relative abundance of Lactobacillus acidophilus was comparable between LA and CON groups. However, a positive effect was observed in relation to the metabolic functions in the treated group, with particular reference to the higher abundance of β-glucosidase. In conclusion, the LA supplementation improved broiler productive performances and metabolic functions promoting animal health.

Autoantibody cross-reactivity with a microbial protein from a prevalent human gut commensal in antiphospholipid syndrome

Abstract Given the vast antigenic source of the gut microbiota, we hypothesized that human gut commensal bacteria induce and sustain autoreactivity via cross-reactivity. As a paradigm, we studied the antiphospholipid syndrome (APS), which is a serious autoimmune clotting disorder of unknown etiology but with a well-defined major autoantigen, b2-glycoprotein I (b2GPI). Infectious triggers are implicated in transient autoantibody production, but the persistent stimuli for anti-b2GPI antibodies remain unknown. To this end, we characterized b2GPI-specific monoclonal autoantibody reactivity to an in silico identified candidate commensal, Roseburia intestinalis (R.int), that is abundant in human stool from APS patients as detected by a species-specific PCR. Using an APS-derived monoclonal antibody, P1-117, that binds to the major B cell epitope in domain I of b2GPI (RGGMR), we tested for cross-reactivity to anaerobically cultured R.int and its candidate cross-reactive R.int protein (WP_006857340.1). P1-117 bound significantly to R.int lysates whereas a control antibody specific for another epitope within b2GPI did not. Further, P1-117 displayed significant binding to the recombinantly expressed R.int cross-reactive candidate protein. BALB/cJ mice immunized with whole heat-killed R.int displayed significant cross-reactive splenocytes and low titers of cross-reactive IgG autoantibodies. In conclusion, we present here in vitro and in vivo studies supporting cross-reactivity of pathogenic b2GPI-specific autoantibody with a common gut commensal. This study provides a concept for persistently colonizing gut commensals as chronic cross-reactive triggers in genetically susceptible autoimmune hosts such as APS patients.

Fecal Bacteria Act as Novel Biomarkers for Noninvasive Diagnosis of Colorectal Cancer

Purpose: Gut microbiota have been implicated in the development of colorectal cancer. We evaluated the utility of fecal bacterial marker candidates identified by our metagenome sequencing analysis for colorectal cancer diagnosis.Experimental Design: Subjects (total 439; 203 colorectal cancer and 236 healthy subjects) from two independent Asian cohorts were included. Probe-based duplex quantitative PCR (qPCR) assays were established for the quantification of bacterial marker candidates.Results: Candidates identified by metagenome sequencing, including Fusobacterium nucleatum (Fn), Bacteroides clarus (Bc), Roseburia intestinalis (Ri), Clostridium hathewayi (Ch), and one undefined species (labeled as m7), were examined in fecal samples of 203 colorectal cancer patients and 236 healthy controls by duplex-qPCR. Strong positive correlations were demonstrated between the quantification of each candidate by our qPCR assays and metagenomics approach (r = 0.801-0.934, all P < 0.0001). Fn was significantly more abundant in colorectal cancer than controls (P < 0.0001), with AUROC of 0.868 (P < 0.0001). At the best cut-off value maximizing sum of sensitivity and specificity, Fn discriminated colorectal cancer from controls with a sensitivity of 77.7%, and specificity of 79.5% in cohort I. A simple linear combination of four bacteria (Fn + Ch + m7-Bc) showed an improved diagnostic ability compared with Fn alone (AUROC = 0.886, P < 0.0001) in cohort I. These findings were further confirmed in an independent cohort II. In particular, improved diagnostic performances of Fn alone (sensitivity 92.8%, specificity 79.8%) and four bacteria (sensitivity 92.8%, specificity 81.5%) were achieved in combination with fecal immunochemical testing for the detection of colorectal cancer.Conclusions: Stool-based colorectal cancer-associated bacteria can serve as novel noninvasive diagnostic biomarkers for colorectal cancer. Clin Cancer Res; 23(8); 2061-70. ©2016 AACR.

Low abundance of colonic butyrate-producing bacteria in HIV infection is associated with microbial translocation and immune activation.

Gut microbial translocation is a major driving force behind chronic immune activation during HIV-1 infection. HIV-1-related intestinal dysbiosis, including increases in mucosa-associated pathobionts, may influence microbial translocation and contribute to mucosal and systemic inflammation. Thus, it is critical to understand the mechanisms by which gut microbes and their metabolic products, such as butyrate, influence immune cell function during HIV-1 infection. A cross-sectional study was performed to compare the relative abundance of butyrate-producing bacterial (BPB) species in colonic biopsies and stool of untreated, chronic HIV-1-infected (n = 18) and HIV-1-uninfected (n = 14) study participants. The effect of exogenously added butyrate on gut T-cell activation and HIV-1 infection was evaluated using an ex-vivo human intestinal cell culture model. Species were identified in 16S ribosomal RNA sequence datasets. Ex-vivo isolated lamina propria mononuclear cells were infected with C-C chemokine receptor type 5-tropic HIV-1Bal, cultured with enteric gram-negative bacteria and a range of butyrate doses, and lamina propria T-cell activation and HIV-1 infection levels measured. Relative abundance of total BPB and specifically of Roseburia intestinalis, were lower in colonic mucosa of HIV-1-infected versus HIV-1-uninfected individuals. In HIV-1-infected study participants, R. intestinalis relative abundance inversely correlated with systemic indicators of microbial translocation, immune activation, and vascular inflammation. Exogenous butyrate suppressed enteric gram-negative bacteria-driven lamina propria T-cell activation and HIV-1 infection levels in vitro. Reductions in mucosal butyrate from diminished colonic BPB may exacerbate pathobiont-driven gut T-cell activation and HIV replication, thereby contributing to HIV-associated mucosal pathogenesis.

Roseburia spp.: a marker of health?

The genus Roseburia consists of obligate Gram-positive anaerobic bacteria that are slightly curved, rod-shaped and motile by means of multiple subterminal flagella. It includes five species: Roseburia intestinalis, R. hominis, R. inulinivorans, R. faecis and R. cecicola. Gut Roseburia spp. metabolize dietary components that stimulate their proliferation and metabolic activities. They are part of commensal bacteria producing short-chain fatty acids, especially butyrate, affecting colonic motility, immunity maintenance and anti-inflammatory properties. Modification in Roseburia spp. representation may affect various metabolic pathways and is associated with several diseases (including irritable bowel syndrome, obesity, Type 2 diabetes, nervous system conditions and allergies). Roseburia spp. could also serve as biomarkers for symptomatic pathologies (e.g., gallstone formation) or as probiotics for restoration of beneficial flora.

Crystal structures of a group II intron maturase reveal a missing link in spliceosome evolution.

Group II introns are self-splicing ribozymes that are essential in many organisms, and they are hypothesized to share a common evolutionary ancestor with the spliceosome. While structural similarity of RNA components supports this connection, it is of interest to determine whether associated protein factors also share an evolutionary heritage. Here we present the crystal structures of reverse transcriptase (RT) domains from two group II intron encoded proteins (maturases) from Roseburia intestinalis and Eubacterium rectale, obtained at 1.2 Å and 2.1 Å respectively. Their architecture is more similar to the spliceosomal Prp8 RT-like domain than to any other RTs, and they share substantial similarity with flaviviral RNA polymerases. The RT domain itself is sufficient for binding intron RNA with high affinity and specificity, and it is contained within an active RT enzyme. These studies provide a foundation for understanding structure-function relationships within group II intron–maturase complexes.

Determination of potential metabolic pathways of human intestinal bacteria by modeling growth kinetics from cross-feeding dynamics

Microbial cross-feeding is essential for a healthy commensal bacteria community in the human gut. Here we present mathematical models that account for the various types of cross-feeding by human commensal intestinal bacteria. The model bacteria include a mixed but unknown microbial community (fecal slurry), Eubacterium hallii, Roseburia intestinalis, Roseburia inulinivorans, and Anaserostipes caccae. These mathematical models demonstrate that a carbon balance approach together with chemical kinetic analysis and parameters estimated from model fitting can be used to determine which of several potential metabolic pathways are employed by cross-feeding bacterial communities to produce their metabolites. The approach can be used to estimate growth kinetics either if the population of bacteria is known, or if the population is mixed and unknown. Based on chemical kinetic analysis, an alternative view of the metabolic pathway of E. hallii is proposed. The modeling suggested that the production of butyrate by E. hallii from lactate and acetate was a second rather than a third-order reaction. Furthermore, the process by which both R. inulinivorans and R. intestinalis degraded carbohydrates and acetate was a second order reaction, and the consumption ratio was found to be approximately 1mM FE oligofructose to 1mM acetate for both Roseburia strains. As well as estimating metabolic parameters, the approach has also suggested candidate metabolic pathways for those systems that could be tested experimentally.

Comparison study of gut microbiota in case of Parkinson’s disease and other neurological disorders

Currently the role of microbiota in diseases pathogenesis, its therapeutic and diagnostic potential are of the utmost interest for scientists and medical doctors. Parkinson’s disease is neurodegenerative disorder for which microbiota’s dysbiosis was previously shown. The main goal of the study is to compare the colon microbiota composition in case of Parkinson’s disease and other neurological pathologies, including idiopathic familial dystonia, essential tremor, multiple sclerosis, multiple system atrophy in order to determine the intestinal flora landscape specific to Parkinson’s disease. Material and methods. One hundred twenty-six patients, 93 with Parkinson’s disease and 33 with other neurological pathology were examined. For all patients, physical examination and fecal samples collection were performed. Microbiota taxonomic composition was analyzed by sequencing of bacterial 16S rRNA genes followed by bioinformatic and statistical analysis. As a result of the study, significant differences between groups in microbiota composition were found. Gut microbiota of patients with Parkinson’s disease was characterized by increase of Desulfovibrio piger, Lactobacillus mucosae, Yokenella regensburgei, Alistipes indistinctus, Oscillospira capillosus, Clostridium bolteae, Soleaferrea massiliensis, Butyricimonas virosa, Dorea massiliensis, Victivallis vadensis abundances. Patients with other neurological diseases had increased levels of bacteria belonging to Blautia, Intestinibacter, Coprococcus genera and Anoxystipes fissicatena, Fusobacterium periodonticum, Gemmiger formicilis, Papillibacter cinnamivorans, Roseburia faecis, Lachnoclostridium indolis, Clostridium populeti, Clostridium tertium, Roseburia intestinalis, Eubacterium desmolans, Eubacterium cylindroides, Clostridium clariflavum, Eubacterium eligens, Coprococcus eutactus, Intestinibacter bartlettii species in their gut microbiota. Consequently, gut microbiota in case of Parkinson’s disease was different from the microbiota of patients with other neurological diseases, including neuroinflammatory and neurodegenerative disorders, in terms of taxonomic diversity and composition.

Reclassification of Eubacterium rectale (Hauduroy et al. 1937) Prévot 1938 in a new genus Agathobacter gen. nov. as Agathobacter rectalis comb. nov., and description of Agathobacter ruminis sp. nov., isolated from the rumen contents of sheep and cows.

Three strains of a butyrate-producing bacterium were isolated from the rumen contents of grazing sheep and cows. The strains were anaerobic, with Gram-positive cell walls, straight-to-slightly-curved, rod-shaped, non-spore-forming and single flagellate. C14 : 1, C14 : 0, C16 : 0 and C16 : 1 were the predominant fatty acids. The cell-wall peptidoglycan type was A1γ. The DNA G+C content varied from 41.4 to 42.2 mol%. 16S rRNA gene sequence similarities between the isolates and Eubacterium rectale, Roseburia hominis and Roseburia intestinalis were found to be 96, 95 and 95 %, respectively. The phylogenetic tree showed that the strains constituted a different taxon, separate from other taxa with validly published names and forming a cluster with strains of Eubacterium rectale. On the basis of phenotypic, chemotaxonomic and phylogenetic results (16S RNA, dnaK, groEL, atpA genes), the isolates are considered to represent a novel species of a new genus of the family Lachnospiraceae, for which the name Agathobacter ruminis gen. nov., sp. nov. is proposed (type strain JK623T = DSM 29029T = LMG 28559T). We also propose the transfer of Eubacterium rectale to the new genus as Agathobacter rectalis gen. nov., comb nov. This new genus represents saccharoclastic, chemo-organotrophic and obligatory anaerobic, non-spore-forming rods with Gram-positive membrane. The main fermentation products on peptone yeast glucose (PYG) medium were butyrate, acetate, hydrogen and lactate. The type species of the genus is Agathobacter rectalis gen. nov., comb nov. (Prévot, 1938) with type strain ATCC 33656T ( = JCM 17463T).

Iron Modulates Butyrate Production by a Child Gut Microbiota In Vitro.

ABSTRACTThe aim of this study was to investigate the effect of iron (Fe) availability on butyrate production in the complex bacterial ecosystem of the human gut. Hence, different Fe availabilities were mimicked in an in vitro colonic fermentation model (the polyfermenter intestinal model called PolyFermS) inoculated with immobilized gut microbiota from a child and in batch cultures of the butyrate producer Roseburia intestinalis. Shifts in the microbial community (16S rRNA sequencing and quantitative PCR), metabolic activity (high-performance liquid chromatography), and expression of genes involved in butyrate production were assessed. In the PolyFermS, moderate Fe deficiency resulted in a 1.4-fold increase in butyrate production and a 5-fold increase in butyryl-coenzyme A (CoA):acetate CoA-transferase gene expression, while very strong Fe deficiency significantly decreased butyrate concentrations and butyrate-producing bacteria compared with the results under normal Fe conditions. Batch cultures of R. intestinalis grown in a low-Fe environment preferentially produced lactate and had reduced butyrate and hydrogen production, in parallel with upregulation of the lactate dehydrogenase gene and downregulation of the pyruvate:ferredoxin-oxidoreductase gene. In contrast, under high-Fe conditions, R. intestinalis cultures showed enhanced butyrate and hydrogen production, along with increased expression of the corresponding genes, compared with the results under normal-Fe conditions. Our data reveal the strong regulatory effect of Fe on gut microbiota butyrate producers and on the concentrations of butyrate, which contributes to the maintenance of host gut health.

Microorganisms linked to inflammatory bowel disease-associated dysbiosis differentially impact host physiology in gnotobiotic mice.

Studying host-microbiota interactions are fundamental to understanding the mechanisms involved in intestinal homeostasis and inflammation. In this work, we analyzed these interactions in mice that were mono-associated with six microorganisms that are representative of inflammatory bowel disease (IBD)-associated dysbiosis: the bacteria Bacteroides thetaiotaomicron, adhesive-invasive Escherichia coli (AIEC), Ruminococcus gnavus and Roseburia intestinalis; a yeast used as a probiotic drug, Saccharomyces boulardii CNCM I-745; and another yeast, Candida albicans. Extensive ex vivo analyses including colon transcriptomics, histology, immune response, bile acid metabolism and short-chain fatty acid production were studied. We showed that B. thetaiotaomicron had the highest impact on the immune system because it was almost able to recapitulate the effects of the entire conventional microbiota and notably induced Treg pathways. Furthermore, these analyses uncovered the effects of E. coli AIEC LF82 on indoleamine 2,3-dioxygenase expression and of S. boulardii CNCM I-745 on angiogenesis. These results were confirmed in vitro in human cell lines. Finally, our results suggested that R. gnavus has major effects on metabolism, and notably on tryptophan metabolism. This work therefore reveals that microorganisms with a potential role in intestinal homeostasis and inflammation have specific impacts on the host, and it suggests several tracks to follow to understand intestinal homeostasis and IBD pathogenesis better, providing new insights to identify novel therapeutic targets.

PWE-361 Colonic microbiome-metabonome network interactions in african americans and native africans: a prospective 2-week food exchange study

Introduction Higher rates of colon cancer are seen in African Americans (65:100,000) compared to rural South Africans ( Method This was a prospective, 2-week food exchange study. 20 middle aged healthy Native Africans (NA) and 20 African Americans (AA) (age 50–65, BMI 18.5–35 kg/m 2 ) were studied on their usual diet and after dietary modification to either a high meat and fat, low fibre western diet (protein 27%, fat 52%, carbohydrate 21%, total fibre 12 g/d) or a high fibre African diet under supervision for 2 weeks. Colonoscopy was performed for mucosal biomarkers of cancer risk from the colon (Ki67). Urinary and faecal samples were analysed by 600MHz 1HNMR (Bruker) and interpreted in Matlab v14a. Microbiome analysis was performed by the HITChip microarray. Metabolic reaction networks of metabolites found differentially expressed between different dietary comparisons, were created using the MetaboNetworks software. Results Orthoganol Partial Least Square Discriminant Analysis regression coefficient plots of AA (faecal: Q2Y = 0.66, R2X = 37.2%, urinary Q2Y = 0.51, R2X 20.8%) and and NA (faecal: Q2Y = 0.37, R2X = 30.7%, urinary Q2Y = 0.83, R2X 27.8%) samples demonstrated distinct metabolic phenotypes for each dietary intervention. Increased diversity of metabolites was seen in faecal water of NAs consuming their usual diet. Faecal metabolites demonstrated a higher number of significant correlations with faecal microbiota compared with urinary metabolites. Eubacterium rectale , Lachnospira pectinoschiza , Roseburia intestinalis and Lactobacillus bovis correlated strongly with butyrate, acetate and formate. The faceal metabolic network pathway analysis demonstrated reciprocal changes after dietary interventions reflecting reductions in short chain fatty acids and increases in glycosylated proteins and choline metabolism in NAs, and increased amino acid metabolism in AAs. Reciprocal changes in urinary metabolites derived from microbial metabolism of green vegetables, e.g. N -acetyl- S -methyl- L -cysteine sulfoxide (p 2 = 0.64), were observed, suggesting increased phytochemical consumption may influence mucosal inflammation. Conclusion A high fat low fibre western diet is associated with a specific, modifiable metabolic phenotype. This is significantly influenced by the gut microbiome, which in turn may have an important role in the cause of colon cancer. Disclosure of interest J. Li: None Declared, J. Kinross Grant/ Research Support from: Imperial BRC, Conflict with: Director of Getwell media, J. Posma: None Declared, L. Lahti: None Declared, V. Naidoo: None Declared, J. Ou: None Declared, K. Newton: None Declared, R. Gaskins: None Declared, E. Zoetendal: None Declared, S. O9Keefe: None Declared, J. Nicholson: None Declared.

A common gut commensal as a cross-reactive candidate in antiphospholipid syndrome (HUM3P.250)

Abstract Antiphospholipid syndrome (APS) is an autoimmune clotting disease characterized by autoantibodies interfering with coagulation. Given the vast antigenic potential of the gut microbiota, we hypothesized that adaptive immune responses to gut commensals may sustain chronic autoreactivity via cross-reactivity. We identified in in silico searches the common, gram-positive gut commensal Roseburia intestinalis as a prime candidate for molecular mimicry, containing peptide sequences highly homologous to both T and B cell immunodominant epitopes of the major autoantigen in APS, the plasma protein b2-glycoprotein I (b2GPI). PBMCs from APS patients proliferated significantly more to R. intestinalis protein extracts compared to phylogenetically closely related Eubacterium rectale, which lacks homologous peptides. We therefore cloned b2GPI-specific CD4 memory T cells and tested cross-reactivity with peptides in vitro. Preliminary data supports that IL-2 but not IL-4, IL-10 or IL-17 secretion can be elicited from an autoantigen-specific Th1 clone in response to a R. intestinalis peptide. Furthermore, Western blotting using patient plasma samples revealed specific bands for R. intestinalis including a potential cross-reactive protein at 24 kDa. Pre-adsorption using recombinant b2GPI prior to Western blotting showed a decrease in this candidate band. These data support a working model whereby autoreactive T and B cell responses are sustained by cross-reactivity with a common gut commensal.

Cross‐reactivity of Gut Commensals and Autoantigen in Antiphospholipid Syndrome

Antiphospholipid syndrome (APS) is an autoimmune clotting disorder of unknown etiology targeting mainly the autoantigen ß2‐glycoprotein I (β2GPI). Given the vast antigenic potential of the gut microbiota and its general influence on autoimmune disease models, we hypothesized that a homeostatic adaptive immune response to gut commensals can trigger and sustain autoreactive T and B cells via cross‐reactivity with b2GPI. Protein BLAST and Clustal Omega queries identified the common human gut commensal Roseburia intestinalis as containing peptide sequences with high homology to the immunodominant ß2GPI T and B cell epitopes. We present here initial studies examining T and B cell reactivity to gut commensals in human APS patients. Patient PBMCs proliferated significantly more to R. intestinalis protein extracts compared to phylogenetically related Eubacterium rectale, which lacks homologous sequences. We successfully cloned ß2GPI‐specific memory T cells using a T cell library assay to determine next if these clones cross‐react with candidate commensals. Western blot experiments using patient plasma samples showed reactivity with several commensals, notably also with R. intestinalis including one protein at 24 kDa that may be cross‐reactive based on in silico experiments. We next adsorbed plasma from APS and control subjects with recombinant ß2GPI prior to Western blotting observing a decrease in the R. intestinalis 24 kDa band. These data support a working model where lymphocytes specific to commensals cross‐react with human autoantigen and vice versa. This process could sustain APS pathogenesis in vivo.

Diets high in resistant starch and arabinoxylan modulate digestion processes and SCFA pool size in the large intestine and faecal microbial composition in pigs

The effects of a high level of dietary fibre (DF) either as arabinoxylan (AX) or resistant starch (RS) on digestion processes, SCFA concentration and pool size in various intestinal segments and on the microbial composition in the faeces were studied in a model experiment with pigs. A total of thirty female pigs (body weight 63.1 (sem 4.4) kg) were fed a low-DF, high-fat Western-style control diet (WSD), an AX-rich diet (AXD) or a RS-rich diet (RSD) for 3 weeks. Diet significantly affected the digestibility of DM, protein, fat, NSP and NSP components, and the arabinose:xylose ratio, as well as the disappearance of NSP and AX in the large intestine. RS was mainly digested in the caecum. AX was digested at a slower rate than RS. The digesta from AXD-fed pigs passed from the ileum to the distal colon more than twice as fast as those from WSD-fed pigs, with those from RSD-fed pigs being intermediate (P< 0.001). AXD feeding resulted in a higher number of Faecalibacterium prausnitzii, Roseburia intestinalis, Blautia coccoides-Eubacterium rectale, Bifidobacterium spp. and Lactobacillus spp. in the faeces sampled at week 3 of the experimental period (P< 0.05). In the caecum, proximal and mid colon, AXD feeding resulted in a 3- to 5-fold higher pool size of butyrate compared with WSD feeding, with the RSD being intermediate (P <0.001). In conclusion, the RSD and AXD differently affected digestion processes compared with the WSD, and the AXD most efficiently shifted the microbial composition towards butyrogenic species in the faeces and increased the large-intestinal butyrate pool size.

Preweaning modulation of intestinal microbiota by oligosaccharides or amoxicillin can contribute to programming of adult microbiota in rats

ObjectiveIncreasing evidence suggests that early nutrition has programming effects on adult health. Identifying mechanisms underlying nutritional programming would aid in the design of new disease prevention strategies. The intestinal microbiota could be a key player in this programming because it affects host metabolic homeostasis, postnatal gut colonization is sensitive to early nutrition, and initial microbial set-up is thought to shape microbiota composition for life. The aim of this study was to determine whether early manipulation of intestinal microbiota actually programs adult microbiota in rats. MethodsSuckling rats pups were supplemented with fructo-oligosaccharides, galacto-oligosaccharides/long-chain fructan mix (GOS/lcF, 9/1), acidic oligosaccharides, amoxicillin, or vehicle from the fifth to the fourteenth day of life, and weaned to standard chow at day 21. Ceco-colonic microbiota was characterized at 14 and 131 d by real-time polymerase chain reaction analysis. ResultsAt day 14, all treatments affected microbiota. Amoxicillin had the most significant effect. All oligosaccharides decreased Firmicutes levels, whereas only fructo-oligosaccharides and GOS/lcF increased bifidobacteria. At day 131, most of these effects had faded away but a significant, albeit minor, adult microbiota programming was observed for rats that received GOS/lcF mix before weaning, regarding Roseburia intestinalis cluster, one subdivision of the Erysipelotrichaceae family as well as butyrate kinase gene. ConclusionsAs revealed by a targeted quantitative polymerase chain reaction approach, programming of adult intestinal microbiota seems to vary according to the nature of the preweaning microbiotal modulator. This suggests that intestinal microbiota may, only under specific circumstances, serve as a relay of neonatal nutrition and thus potentially contribute to nutritional programming of host physiology.