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Studies in type 2 diabetes (T2D) rodent models suggest the potential for individual microbial taxa in the human gut to improve or worsen metabolic disease via specific metabolites (signals) in hepatic or general circulation. We previously reported significant improvement in postprandial glucose control in T2D subjects following ingestion of a 5-strain novel probiotic formulation (INT) in a placebo (PBO) controlled setting. (DOI:10.1136/bmjdrc-2020-001319) Here we report measurements of fecal and fasting circulating plasma metabolites, implicated as mediators of microbiome effects, collected before and after the 12-week intervention. Consistent with the hypothesized benefits of increased colonic production of butyrate, plasma butyrate increased in INT but not PBO (within group, concentration increase = 0.15 µM, p = 0.007; between group, p = 0.11), but not acetate nor propionate, paralleling changes in subject stool and production by in vitro culture. We detected changes consistent with enhanced fatty acid oxidation in INT relative to placebo, especially a decrease in medium chain fatty acids, their carnitinylates, and a decrease in bilirubin degradation byproducts. We also detected increases in certain bile acids, especially ursodeoxycholic acid (UDCA). In vitro monoculture supernatant showed that the formulation’s C. butyricum strain specifically and efficiently synthesizes UDCA during butyrogenic growth in rich media amended with the primary bile acid chenodeoxycholic acid. Untargeted data also uncovered previously undocumented use of sulfonylurea (SFU), accounting for 6 out of 7 subjects with A1c non-response in the INT group. To our knowledge, this is the first description of an increase in circulating butyrate and UDCA due solely to a bacterial probiotic intervention in humans with T2D, and may help explain the observed improvement in postprandial glucose control. Disclosure P. Mcmurdie: Employee; Self; Pendulum Therapeutics. M. K. Stoeva: Employee; Self; Pendulum Therapeutics. M. Nemchek: Employee; Self; Pendulum Therapeutics. S. Tyagi: None. J. Gines: Employee; Self; Pendulum Therapeutics. J. Eid: Stock/Shareholder; Self; Pendulum Therapeutics. O. G. Kolterman: Board Member; Self; GlySens Incorporated, Consultant; Self; ADOCIA, Cirius Therapeutics, Nanoprecision Medical, Viacyte, Inc., Employee; Self; Pendulum Therapeutics.

Clostridium butyricum is a butyrate-producing human gut symbiont that has been safely used as a probiotic for decades. C. butyricum strains have been investigated for potential protective or ameliorative effects in a wide range of human diseases, including gut-acquired infection, intestinal injury, irritable bowel syndrome, inflammatory bowel disease, neurodegenerative disease, metabolic disease, and colorectal cancer. In this review we summarize the studies on C. butyricum supplementation with special attention to proposed mechanisms for the associated health benefits and the supporting experimental evidence. These mechanisms center on molecular signals (especially butyrate) as well as immunological signals in the digestive system that cascade well beyond the gut to the liver, adipose tissue, brain, and more. The safety of probiotic C. butyricum strains appears well-established. We identify areas where additional human randomized controlled trials would provide valuable further data related to the strains’ utility as an intervention.

IntroductionA growing body of evidence suggests that specific, naturally occurring gut bacteria are under-represented in the intestinal tracts of subjects with type 2 diabetes (T2D) and that their functions, like gut barrier stability and butyrate production, are important to glucose and insulin homeostasis. The objective of this study was to test the hypothesis that enteral exposure to microbes with these proposed functions can safely improve clinical measures of glycemic control and thereby play a role in the overall dietary management of diabetes.Research design and methodsWe evaluated whether a probiotic comprised of these anaerobic bacteria would enhance dietary management by (1) manufacturing two novel probiotic formulations containing three (WBF-010) or five (WBF-011) distinct strains in a Current Good Manufacturing Practice (cGMP) facility, (2) establishing consistent live-cell concentrations, (3) confirming safety at target concentrations dispensed in both animal and human studies and (4) conducting a 12-week parallel, double-blind, placebo-controlled, proof-of-concept study in which subjects previously diagnosed with T2D (n=76) were randomly assigned to a two times a day regimen of placebo, WBF-010 or WBF-011.ResultsNo safety or tolerability issues were observed. Compared with the placebo group, subjects administered WBF-011 (which contains inulin, Akkermansia muciniphila, Clostridium beijerinckii, Clostridium butyricum, Bifidobacterium infantis and Anaerobutyricum hallii) significantly improved in the primary outcome, glucose total area under the curve (AUC): −36.1 mg/dL/180 min, p=0.0500 and also improved in secondary outcomes, glycated hemoglobin (A1c): −0.6, glucose incremental-AUC: −28.6 mg/dL/180 min.ConclusionsTo our knowledge, this is the first randomized controlled trial to administer four of the five strains to human subjects with T2D. This proof-of-concept study (clinical trial number NCT03893422) shows that the intervention was safe and well tolerated and that supplementation with WBF-011 improves postprandial glucose control. The limited sample size and intersubject variability justifies future studies designed to confirm and expand on these observations.

Studies show that BP+MR are decreased inT2D; the impact of administering specific strains has not been reported. A 12-week, double blind, placebo-controlled study of 2 MFs prepared GMP using only GRAS constituents (MF-1: BP; MF-2: BP + MR) in T2D (drug naive + metformin ± SFU, n=76), showed improved GC. See Table for demographics and key results. The primary endpoint was change from baseline (∆) in total glucose AUC0-180 min during a Boost Plus™ (720 cal) MTT. At 12 weeks, MF-2 yielded significant ∆s in both total (T) and incremental (I) glucose AUC0-180 min, (p=0.05 and 0.007 respectively). MF-1 improved only I glucose AUC0-180 min (p=0.05). Neither MF-1 or MF-2 impacted fasting glucose, T or I insulin AUC0-180 min, weight, BMI, HOMA-IR, lipids, or c-reactive protein. No safety or tolerability concerns were observed. Pre and post fecal microbial analyses are pending. Post hoc analyses revealed 2 interesting results. 1) SFU subjects across both study groups showed minimal response while subjects not using SFUs had greater ∆s in both T glucose AUC0-180 min (-24.4%) and placebo-corrected ∆ A1c (-0.7%). 2) MF-2 showed significant reductions in ALT consistent with improved liver health. In conclusion, 1) Interventions directed at increasing BP+MR may be of benefit in T2D. 2) MF-2 appears to be a safe and effective non-pharmacologic intervention for T2D which merits further investigation. Disclosure O.G. Kolterman: Advisory Panel; Spouse/Partner; Intarcia Therapeutics, Inc. Advisory Panel; Self; Zafgen, Inc. Board Member; Self; American Diabetes Association, GlySens Incorporated, Viacyte, Inc. Consultant; Self; ADOCIA, Cirius Therapeutics Inc, DiaVacs Inc, JDRF, Nanoprecisionmedical, NuSirt Biopharma, Inc., Renova Therapeutics, Sensulin, LLC. Employee; Self; Whole Biome Inc. Stock/Shareholder; Self; Corcept Therapeutics, GI Therapeutics, Xeris Pharmaceuticals, Inc. Other Relationship; Spouse/Partner; American Diabetes Association. F. Perraudeau: Employee; Self; Whole Biome. M.S. Iyer: Employee; Self; Whole Biome Inc. J.S. Eid: Board Member; Self; Whole Biome Inc. J. Bullard: Employee; Self; Whole Biome Inc. Stock/Shareholder; Self; Whole Biome Inc. M.F. Schicklberger: Employee; Self; Whole Biome Inc. A. Cheng: None. N. Justice: Employee; Self; Whole Biome. P. McMurdie: Employee; Self; Whole Biome, Inc. B. Stoneburner: None. J. Gines: Employee; Self; Whole Biome Inc. S. Tyagi: None. C. Cutcliffe: Employee; Self; Whole Biome Inc.

ObjectiveColorectal cancer (CRC) is the second leading cause of cancer-associated mortality in the USA. The faecal microbiome may provide non-invasive biomarkers of CRC and indicate transition in the adenoma–carcinoma sequence. Re-analysing raw sequence and metadata from several studies uniformly, we sought to identify a composite and generalisable microbial marker for CRC.DesignRaw 16S rRNA gene sequence data sets from nine studies were processed with two pipelines, (1) QIIME closed reference (QIIME-CR) or (2) a strain-specific method herein termed SS-UP (Strain Select, UPARSE bioinformatics pipeline). A total of 509 samples (79 colorectal adenoma, 195 CRC and 235 controls) were analysed. Differential abundance, meta-analysis random effects regression and machine learning analyses were carried out to determine the consistency and diagnostic capabilities of potential microbial biomarkers.ResultsDefinitive taxa, including Parvimonas micra ATCC 33270, Streptococcus anginosus and yet-to-be-cultured members of Proteobacteria, were frequently and significantly increased in stools from patients with CRC compared with controls across studies and had high discriminatory capacity in diagnostic classification. Microbiome-based CRC versus control classification produced an area under receiver operator characteristic (AUROC) curve of 76.6% in QIIME-CR and 80.3% in SS-UP. Combining clinical and microbiome markers gave a diagnostic AUROC of 83.3% for QIIME-CR and 91.3% for SS-UP.ConclusionsDespite technological differences across studies and methods, key microbial markers emerged as important in classifying CRC cases and such could be used in a universal diagnostic for the disease. The choice of bioinformatics pipeline influenced accuracy of classification. Strain-resolved microbial markers might prove crucial in providing a microbial diagnostic for CRC.

Growing interest in the gut microbiome has revealed links between gut microbiome composition and many diseases including not only infections but also obesity, diabetes and inflammatory bowel disease. The microbiome composition can undergo changes as a result of interactions between gut microbiome and exogenous agents. However, the impact of commonly used pharmaceuticals on the gut microbiome has not been well characterized. In this study, we characterized the impact of 24 common pharmaceuticals on gut microbiome compositional shift in mice and in a human fecal culture system. Mice were dosed with each optimized pharmaceutical compound daily for five consecutive days. Fecal samples were collected on day -1, 0, 4 and 5 of the administration. For the human fecal culture system, human stool samples were collected and immediately stored at -80 degree C. After resuspension of the sample in reduced medium, fecal cultures were incubated with individual compounds for 24 hours in an anaerobic chamber. DNA was extracted, then the V4 region of 16S rRNA gene was amplified by PCR and sequenced using the Illumina MiSeq. A negative binomial Wald test was used to detect microbial taxa difference. Principal component analysis exhibits greatest shift in mice microbiome by antimicrobial drugs such as Azithromycin, Amoxicillin and Metronidazole as well as Type 2 diabetes drugs such as Acarbose and Metformin between day 0 and day 5. Permutational multivariate analysis of variance using Euclidean distance exhibited greatest significant microbiome shift by Acarbose, Amoxicillin and Metformin (p 0.1). These results suggest different impact of administration of antimicrobial agents (e.g. protein synthesis inhibitor) and modulation of glucose availability to the intestinal microbiome. Similar microbiome shifts were also observed in the anaerobic human fecal culture system. Understanding the impact of pharmaceuticals to the gut microbiome could ultimately be useful in improving predictions of the therapeutic intervention outcomes on variety of individuals.