Another renaissance for bile acid gastrointestinal microbiology.

The field of bile acid microbiology in the gastrointestinal tract is going through a current rebirth after a peak of activity in the late 1970s and early 1980s. This renewed activity is a result of many factors, including the discovery near the turn of the century that bile acids are potent signalling molecules and technological advances in next-generation sequencing, computation, culturomics, gnotobiology, and metabolomics. We describe the current state of the field with particular emphasis on questions that have remained unanswered for many decades in both bile acid synthesis by the host and metabolism by the gut microbiota. Current knowledge of established enzymatic pathways, including bile salt hydrolase, hydroxysteroid dehydrogenases involved in the oxidation and epimerization of bile acid hydroxy groups, the Hylemon-BjÓ§rkhem pathway of bile acid C7-dehydroxylation, and the formation of secondary allo-bile acids, is described. We cover aspects of bile acid conjugation and esterification as well as evidence for bile acid C3-dehydroxylation and C12-dehydroxylation that are less well understood but potentially critical for our understanding of bile acid metabolism in the human gut. The physiological consequences of bile acid metabolism for human health, important caveats and cautionary notes on experimental design and interpretation of data reflecting bile acid metabolism are also explored.

CHCHD2 mediates glioblastoma cell proliferation, mitochondrial metabolism, hypoxia­induced invasion and therapeutic resistance.

Glioblastoma (GBM) is the most common and malignant primary brain tumor affecting adults and remains incurable. The mitochondrial coiled­coil­helix­coiled­coil­helix domain­containing protein 2 (CHCHD2) has been demonstrated to mediate mitochondrial respiration, nuclear gene expression and cell migration; however, evidence of this in GBM is lacking. In the present study, it was hypothesized that CHCHD2 may play a functional role in U87 GBM cells expressing the constitutively active epidermal growth factor receptor variant III (EGFRvIII). The amplification of the CHCHD2 gene was found to be associated with a decreased patient overall and progression­free survival. The CHCHD2 mRNA levels were increased in high­vs. low­grade glioma, IDH­wt GBMs, and in tumor vs. non­tumor tissue. Additionally, CHCHD2 protein expression was greatest in invasive, EGFRvIII­expressing patient­derived samples. The CRISPR­Cas9­mediated knockout of CHCHD2 in EGFRvIII­expressing U87 cells resulted in an altered mitochondrial respiration and glutathione status, in decreased cell growth and invasion under both normoxic and hypoxic conditions, and in an enhanced sensitivity to cytotoxic agents. CHCHD2 was distributed in both the mitochondria and nuclei of U87 and U87vIII cells, and the U87vIII cells exhibited a greater nuclear expression of CHCHD2 compared to isogenic U87 cells. Incubation under hypoxic conditions, serum starvation and the reductive unfolding of CHCHD2 induced the nuclear accumulation of CHCHD2 in both cell lines. Collectively, the findings of the present study indicate that CHCHD2 mediates a variety of GBM characteristics, and highlights mitonuclear retrograde signaling as a pathway of interest in GBM cell biology.

Diet, Gut Microbiome, and Their End Metabolites Associate With Acute Pancreatitis Risk.

INTRODUCTION: Diet and decreased gut microbiome diversity has been associated with acute pancreatitis (AP) risk. However, differences in dietary intake, gut microbiome, and their impact on microbial end metabolites have not been studied in AP. We aimed to determine differences in (i) dietary intake (ii) gut microbiome diversity and sulfidogenic bacterial abundance, and (iii) serum short-chain fatty acid (SCFA) and hydrogen sulfide (H 2 S) concentrations in AP and control subjects. METHODS: This case-control study recruited 54 AP and 46 control subjects during hospitalization. Clinical and diet data and stool and blood samples were collected. 16S rDNA sequencing was used to determine gut microbiome alpha diversity and composition. Serum SCFA and H 2 S levels were measured. Machine learning (ML) model was used to identify microbial targets associated with AP. RESULTS: AP patients had a decreased intake of vitamin D 3 , whole grains, fish, and beneficial eicosapentaenoic, docosapentaenoic, and docosahexaenoic acids. AP patients also had lower gut microbiome diversity ( P = 0.021) and a higher abundance of sulfidogenic bacteria including Veillonella sp. and Haemophilus sp., which were associated with AP risk. Serum acetate and H 2 S concentrations were significantly higher in the AP group ( P < 0.001 and P = 0.043, respectively). ML model had 96% predictive ability to distinguish AP patients from controls. DISCUSSION: AP patients have decreased beneficial nutrient intake and gut microbiome diversity. An increased abundance of H 2 S-producing genera in the AP and SCFA-producing genera in the control group and predictive ability of ML model to distinguish AP patients indicates that diet, gut microbiota, and their end metabolites play a key role in AP.

The Hylemon-Björkhem pathway of bile acid 7-dehydroxylation: history, biochemistry, and microbiology.

Bile acids are detergents derived from cholesterol that function to solubilize dietary lipids, remove cholesterol from the body, and act as nutrient signaling molecules in numerous tissues with functions in the liver and gut being the best understood. Studies in the early 20th century established the structures of bile acids, and by mid-century, the application of gnotobiology to bile acids allowed differentiation of host-derived "primary" bile acids from "secondary" bile acids generated by host-associated microbiota. In 1960, radiolabeling studies in rodent models led to determination of the stereochemistry of the bile acid 7-dehydration reaction. A two-step mechanism was proposed, which we have termed the Samuelsson-Bergström model, to explain the formation of deoxycholic acid. Subsequent studies with humans, rodents, and cell extracts of Clostridium scindens VPI 12708 led to the realization that bile acid 7-dehydroxylation is a result of a multi-step, bifurcating pathway that we have named the Hylemon-Björkhem pathway. Due to the importance of hydrophobic secondary bile acids and the increasing measurement of microbial bai genes encoding the enzymes that produce them in stool metagenome studies, it is important to understand their origin.

Design of the Building Research in CRC prevention (BRIDGE-CRC) trial: a 6-month, parallel group Mediterranean diet and weight loss randomized controlled lifestyle intervention targeting the bile acid-gut microbiome axis to reduce colorectal cancer risk among African American/Black adults with obesity.

BACKGROUND: Among all racial/ethnic groups, people who identify as African American/Blacks have the second highest colorectal cancer (CRC) incidence in the USA. This disparity may exist because African American/Blacks, compared to other racial/ethnic groups, have a higher prevalence of risk factors for CRC, including obesity, low fiber consumption, and higher intakes of fat and animal protein. One unexplored, underlying mechanism of this relationship is the bile acid-gut microbiome axis. High saturated fat, low fiber diets, and obesity lead to increases in tumor promoting secondary bile acids. Diets high in fiber, such as a Mediterranean diet, and intentional weight loss may reduce CRC risk by modulating the bile acid-gut microbiome axis. The purpose of this study is to test the impact of a Mediterranean diet alone, weight loss alone, or both, compared to typical diet controls on the bile acid-gut microbiome axis and CRC risk factors among African American/Blacks with obesity. Because weight loss or a Mediterranean diet alone can reduce CRC risk, we hypothesize that weight loss plus a Mediterranean diet will reduce CRC risk the most. METHODS: This randomized controlled lifestyle intervention will randomize 192 African American/Blacks with obesity, aged 45-75 years to one of four arms: Mediterranean diet, weight loss, weight loss plus Mediterranean diet, or typical diet controls, for 6 months (48 per arm). Data will be collected at baseline, mid-study, and study end. Primary outcomes include total circulating and fecal bile acids, taurine-conjugated bile acids, and deoxycholic acid. Secondary outcomes include body weight, body composition, dietary change, physical activity, metabolic risk, circulating cytokines, gut microbial community structure and composition, fecal short-chain fatty acids, and expression levels of genes from exfoliated intestinal cells linked to carcinogenesis. DISCUSSION: This study will be the first randomized controlled trial to examine the effects of a Mediterranean diet, weight loss, or both on bile acid metabolism, the gut microbiome, and intestinal epithelial genes associated with carcinogenesis. This approach to CRC risk reduction may be especially important among African American/Blacks given their higher risk factor profile and increased CRC incidence. TRIAL REGISTRATION: ClinicalTrials.gov NCT04753359 . Registered on 15 February 2021.

Effect of Diets Varying in Iron and Saturated Fat on the Gut Microbiota and Intestinal Inflammation: A Crossover Feeding Study among Older Females with Obesity.

Obesity is considered an independent risk factor for colorectal cancer (CRC). Altered nutrient metabolism, particularly changes to digestion and intestinal absorption, may play an important role in the development of CRC. Iron can promote the formation of tissue-damaging and immune-modulating reactive oxygen species. We conducted a crossover, controlled feeding study to examine the effect of three, 3-week diets varying in iron and saturated fat content on the colonic milieu and systemic markers among older females with obesity. Anthropometrics, fasting venous blood and stool were collected before and after each diet. There was a minimum 3-week washout period between diets. Eighteen participants consumed the three diets (72% Black; mean age 60.4 years; mean body mass index 35.7 kg/m2). Results showed no effect of the diets on intestinal inflammation (fecal calprotectin) or circulating iron, inflammation, and metabolic markers. Pairwise comparisons revealed less community diversity between samples (beta diversity, calculated from 16S rRNA amplicon sequences) among participants when consuming a diet low in iron and high in saturated fat vs. when consuming a diet high in iron and saturated fat. More studies are needed to investigate if dietary iron represents a salient target for CRC prevention among individuals with obesity.

Formation of secondary allo-bile acids by novel enzymes from gut Firmicutes.

The gut microbiome of vertebrates is capable of numerous biotransformations of bile acids, which are responsible for intestinal lipid digestion and function as key nutrient-signaling molecules. The human liver produces bile acids from cholesterol predominantly in the A/B-cis orientation in which the sterol rings are "kinked", as well as small quantities of A/B-trans oriented "flat" stereoisomers known as "primary allo-bile acids". While the complex multi-step bile acid 7α-dehydroxylation pathway has been well-studied for conversion of "kinked" primary bile acids such as cholic acid (CA) and chenodeoxycholic acid (CDCA) to deoxycholic acid (DCA) and lithocholic acid (LCA), respectively, the enzymatic basis for the formation of "flat" stereoisomers allo-deoxycholic acid (allo-DCA) and allo-lithocholic acid (allo-LCA) by Firmicutes has remained unsolved for three decades. Here, we present a novel mechanism by which Firmicutes generate the "flat" bile acids allo-DCA and allo-LCA. The BaiA1 was shown to catalyze the final reduction from 3-oxo-allo-DCA to allo-DCA and 3-oxo-allo-LCA to allo-LCA. Phylogenetic and metagenomic analyses of human stool samples indicate that BaiP and BaiJ are encoded only in Firmicutes and differ from membrane-associated bile acid 5α-reductases recently reported in Bacteroidetes that indirectly generate allo-LCA from 3-oxo-Δ4-LCA. We further map the distribution of baiP and baiJ among Firmicutes in human metagenomes, demonstrating an increased abundance of the two genes in colorectal cancer (CRC) patients relative to healthy individuals.

Diversity and distribution of sulfur metabolic genes in the human gut microbiome and their association with colorectal cancer.

BACKGROUND: Recent evidence implicates microbial sulfidogenesis as a potential trigger of colorectal cancer (CRC), highlighting the need for comprehensive knowledge of sulfur metabolism within the human gut. Microbial sulfidogenesis produces genotoxic hydrogen sulfide (H2S) in the human colon using inorganic (sulfate) and organic (taurine/cysteine/methionine) substrates; however, the majority of studies have focused on sulfate reduction using dissimilatory sulfite reductases (Dsr). RESULTS: Here, we show that genes for microbial sulfur metabolism are more abundant and diverse than previously observed and are statistically associated with CRC. Using ~ 17,000 bacterial genomes from publicly available stool metagenomes, we studied the diversity of sulfur metabolic genes in 667 participants across different health statuses: healthy, adenoma, and carcinoma. Sulfidogenic genes were harbored by 142 bacterial genera and both organic and inorganic sulfidogenic genes were associated with carcinoma. Significantly, the anaerobic sulfite reductase (asr) genes were twice as abundant as dsr, demonstrating that Asr is likely a more important contributor to sulfate reduction in the human gut than Dsr. We identified twelve potential pathways for reductive taurine metabolism and discovered novel genera harboring these pathways. Finally, the prevalence of metabolic genes for organic sulfur indicates that these understudied substrates may be the most abundant source of microbially derived H2S. CONCLUSIONS: Our findings significantly expand knowledge of microbial sulfur metabolism in the human gut. We show that genes for microbial sulfur metabolism in the human gut are more prevalent than previously known, irrespective of health status (i.e., in both healthy and diseased states). Our results significantly increase the diversity of pathways and bacteria that are associated with microbial sulfur metabolism in the human gut. Overall, our results have implications for understanding the role of the human gut microbiome and its potential contributions to the pathogenesis of CRC. Video abstract.

Bile Acids, Gut Microbes, and the Neighborhood Food Environment-a Potential Driver of Colorectal Cancer Health Disparities.

Bile acids (BAs) facilitate nutrient digestion and absorption and act as signaling molecules in a number of metabolic and inflammatory pathways. Expansion of the BA pool and increased exposure to microbial BA metabolites has been associated with increased colorectal cancer (CRC) risk. It is well established that diet influences systemic BA concentrations and microbial BA metabolism. Therefore, consumption of nutrients that reduce colonic exposure to BAs and microbial BA metabolites may be an effective method for reducing CRC risk, particularly in populations disproportionately burdened by CRC. Individuals who identify as Black/African American (AA/B) have the highest CRC incidence and death in the United States and are more likely to live in a food environment with an inequitable access to BA mitigating nutrients. Thus, this review discusses the current evidence supporting diet as a contributor to CRC disparities through BA-mediated mechanisms and relationships between these mechanisms and barriers to maintaining a low-risk diet.

Treatment With the CSF1R Antagonist GW2580, Sensitizes Microglia to Reactive Oxygen Species.

Microglia activation and proliferation are hallmarks of many neurodegenerative disorders and may contribute to disease pathogenesis. Neurons actively regulate microglia survival and function, in part by secreting the microglia mitogen interleukin (IL)-34. Both IL-34 and colony stimulating factor (CSF)-1 bind colony stimulating factor receptor (CSFR)1 expressed on microglia. Systemic treatment with central nervous system (CNS) penetrant, CSFR1 antagonists, results in microglia death in a dose dependent matter, while others, such as GW2580, suppress activation during disease states without altering viability. However, it is not known how treatment with non-penetrant CSF1R antagonists, such as GW2580, affect the normal physiology of microglia. To determine how GW2580 affects microglia function, C57BL/6J mice were orally gavaged with vehicle or GW2580 (80mg/kg/d) for 8 days. Body weights and burrowing behavior were measured throughout the experiment. The effects of GW2580 on circulating leukocyte populations, brain microglia morphology, and the transcriptome of magnetically isolated adult brain microglia were determined. Body weights, burrowing behavior, and circulating leukocytes were not affected by treatment. Analysis of Iba-1 stained brain microglia indicated that GW2580 treatment altered morphology, but not cell number. Analysis of RNA-sequencing data indicated that genes related to reactive oxygen species (ROS) regulation and survival were suppressed by treatment. Treatment of primary microglia cultures with GW2580 resulted in a dose-dependent reduction in viability only when the cells were concurrently treated with LPS, an inducer of ROS. Pre-treatment with the ROS inhibitor, YCG063, blocked treatment induced reductions in viability. Finally, GW2580 sensitized microglia to hydrogen peroxide induced cell death. Together, these data suggest that partial CSF1R antagonism may render microglia more susceptible to reactive oxygen and nitrogen species.

Comparison between handheld ultrasound and regional and whole-body dual energy x-ray absorptiometry (DXA) for body fat assessment.

OBJECTIVES: To determine the extent of agreement between a handheld ultrasound (US) attached to an android tablet and the reference method dual energy x-ray absorptiometry (DXA) for the measurement of adiposity. METHODS: A whole-body DXA scan and abdominal adipose tissue thickness measurements using a handheld US were obtained from 104 adults (63 females, 41 males). Body fat percent (BF%), total fat mass (kg), and trunk fat mass (kg) were obtained from DXA. Subcutaneous adipose tissue (SAT), superficial subcutaneous adipose tissue (SSAT), and deep subcutaneous adipose tissue (DSAT) thickness were obtained from US. Sex-specific total fat mass, trunk fat mass, and BF% estimates by US were compared with DXA. Spearman's correlations and Bland-Altman plots were used to assess agreement between the methods. RESULTS: US SAT correlated strongly with total fat mass for both females (rs = 0.74) and males (rs = 0.87) as did trunk fat mass (females, rs = 0.81; males, rs = 0.83); as did SSAT and DSAT (females: rs = 0.65 and rs = 0.66; males: rs = 0.63 and rs = 0.85, respectively, all p-values < 0.0001). Bland-Altman plots demonstrated strong agreement for total and trunk fat mass for both males and females. For BF%, acceptable limits of agreement were observed for males but not for females, substantial proportional bias as indicated by a negative slope was noted for BF% using SAT (r = -0.298, p = 0.0177). CONCLUSION: The handheld US and technique to analyze abdominal adipose tissue thickness showed strong agreement with DXA results and generated highly comparable estimates for total and trunk fat mass for both sexes.

Support policies that foster a healthy food environment and incentivize healthy food purchases to mitigate cancer inequities.

The COVID-19 pandemic has highlighted the inequitable access to resources, leading to a disproportionate burden of disease in vulnerable communities in the USA. However, these inequities in health outcomes are not limited to COVID-19. Approximately 18% of cancers are related to dietary behaviors and excess body weight. Underserved communities, such as minority racial/ethnic groups living in neighborhoods of low socioeconomic status, experience barriers to healthy eating including lack of access to high-quality healthy foods and higher availability of unhealthy foods and beverages in local retail food outlets. Strikingly, these same populations are more likely to die from cancers related to dietary intake and obesity like colorectal, liver, and pancreatic cancers. To reduce cancer inequities, policy makers can act by supporting programs that incentivize healthy food purchases and improve the local food environment in underserved communities.

Completion of the gut microbial epi-bile acid pathway.

Bile acids are detergent molecules that solubilize dietary lipids and lipid-soluble vitamins. Humans synthesize bile acids with α-orientation hydroxyl groups which can be biotransformed by gut microbiota to toxic, hydrophobic bile acids, such as deoxycholic acid (DCA). Gut microbiota can also convert hydroxyl groups from the α-orientation through an oxo-intermediate to the ß-orientation, resulting in more hydrophilic, less toxic bile acids. This interconversion is catalyzed by regio- (C-3 vs. C-7) and stereospecific (α vs. ß) hydroxysteroid dehydrogenases (HSDHs). So far, genes encoding the urso- (7α-HSDH & 7ß-HSDH) and iso- (3α-HSDH & 3ß-HSDH) bile acid pathways have been described. Recently, multiple human gut clostridia were reported to encode 12α-HSDH, which interconverts DCA and 12-oxolithocholic acid (12-oxoLCA). 12ß-HSDH completes the epi-bile acid pathway by converting 12-oxoLCA to the 12ß-bile acid denoted epiDCA; however, a gene(s) encoding this enzyme has yet to be identified. We confirmed 12ß-HSDH activity in cultures of Clostridium paraputrificum ATCC 25780. From six candidate C. paraputrificum ATCC 25780 oxidoreductase genes, we discovered the first gene (DR024_RS09610) encoding bile acid 12ß-HSDH. Phylogenetic analysis revealed unforeseen diversity for 12ß-HSDH, leading to validation of two additional bile acid 12ß-HSDHs through a synthetic biology approach. By comparison to a previous phylogenetic analysis of 12α-HSDH, we identified the first potential C-12 epimerizing strains: Collinsella tanakaei YIT 12063 and Collinsella stercoris DSM 13279. A Hidden Markov Model search against human gut metagenomes located putative 12ß-HSDH genes in about 30% of subjects within the cohorts analyzed, indicating this gene is relevant in the human gut microbiome.

Berberine alters gut microbial function through modulation of bile acids.

BACKGROUND: Berberine (BBR) is a plant-based nutraceutical that has been used for millennia to treat diarrheal infections and in contemporary medicine to improve patient lipid profiles. Reduction in lipids, particularly cholesterol, is achieved partly through up-regulation of bile acid synthesis and excretion into the gastrointestinal tract (GI). The efficacy of BBR is also thought to be dependent on structural and functional alterations of the gut microbiome. However, knowledge of the effects of BBR on gut microbiome communities is currently lacking. Distinguishing indirect effects of BBR on bacteria through altered bile acid profiles is particularly important in understanding how dietary nutraceuticals alter the microbiome. RESULTS: Germfree mice were colonized with a defined minimal gut bacterial consortium capable of functional bile acid metabolism (Bacteroides vulgatus, Bacteroides uniformis, Parabacteroides distasonis, Bilophila wadsworthia, Clostridium hylemonae, Clostridium hiranonis, Blautia producta; B4PC2). Multi-omics (bile acid metabolomics, 16S rDNA sequencing, cecal metatranscriptomics) were performed in order to provide a simple in vivo model from which to identify network-based correlations between bile acids and bacterial transcripts in the presence and absence of dietary BBR. Significant alterations in network topology and connectivity in function were observed, despite similarity in gut microbial alpha diversity (P = 0.30) and beta-diversity (P = 0.123) between control and BBR treatment. BBR increased cecal bile acid concentrations, (P < 0.05), most notably deoxycholic acid (DCA) (P < 0.001). Overall, analysis of transcriptomes and correlation networks indicates both bacterial species-specific responses to BBR, as well as functional commonalities among species, such as up-regulation of Na+/H+ antiporter, cell wall synthesis/repair, carbohydrate metabolism and amino acid metabolism. Bile acid concentrations in the GI tract increased significantly during BBR treatment and developed extensive correlation networks with expressed genes in the B4PC2 community. CONCLUSIONS: This work has important implications for interpreting the effects of BBR on structure and function of the complex gut microbiome, which may lead to targeted pharmaceutical interventions aimed to achieve the positive physiological effects previously observed with BBR supplementation.

Crosstalk between microglia and patient-derived glioblastoma cells inhibit invasion in a three-dimensional gelatin hydrogel model.

BACKGROUND: Glioblastoma is the most common and deadly form of primary brain cancer, accounting for more than 13,000 new diagnoses annually in the USA alone. Microglia are the innate immune cells within the central nervous system, acting as a front-line defense against injuries and inflammation via a process that involves transformation from a quiescent to an activated phenotype. Crosstalk between GBM cells and microglia represents an important axis to consider in the development of tissue engineering platforms to examine pathophysiological processes underlying GBM progression and therapy. METHODS: This work used a brain-mimetic hydrogel system to study patient-derived glioblastoma specimens and their interactions with microglia. Here, glioblastoma cells were either cultured alone in 3D hydrogels or in co-culture with microglia in a manner that allowed secretome-based signaling but prevented direct GBM-microglia contact. Patterns of GBM cell invasion were quantified using a three-dimensional spheroid assay. Secretome and transcriptome (via RNAseq) were used to profile the consequences of GBM-microglia interactions. RESULTS: Microglia displayed an activated phenotype as a result of GBM crosstalk. Three-dimensional migration patterns of patient-derived glioblastoma cells showed invasion was significantly decreased in response to microglia paracrine signaling. Potential molecular mechanisms underlying with this phenotype were identified from bioinformatic analysis of secretome and RNAseq data. CONCLUSION: The data demonstrate a tissue engineered hydrogel platform can be used to investigate crosstalk between immune cells of the tumor microenvironment related to GBM progression. Such multi-dimensional models may provide valuable insight to inform therapeutic innovations to improve GBM treatment.

Effects of taurocholic acid metabolism by gut bacteria: A controlled feeding trial in adult African American subjects at elevated risk for colorectal cancer.

Colorectal cancer (CRC) is the third leading cause of cancer and second leading cause of cancer death in the United States. Recent evidence has linked a high fat and animal protein diet and microbial metabolism of host bile acids as environmental risk factors for CRC development. We hypothesize that the primary bile salt taurocholic acid (TCA) is a key, diet-controlled metabolite whose use by bacteria yields a carcinogen and tumor-promoter, respectively. The work is motivated by our published data indicating hydrogen sulfide (H2S) and secondary bile acid production by colonic bacteria, serve as environmental insults contributing to CRC risk. The central aim of this study is to test whether a diet high in animal protein and saturated fat increases abundance of bacteria that generate H2S and pro-inflammatory secondary bile acids in African Americans (AAs) at high risk for CRC. Our prospective, randomized, crossover feeding trial will examine two microbial mechanisms by which an animal-based diet may support the growth of TCA metabolizing bacteria. Each subject will receive two diets in a crossover design- an animal-based diet, rich in taurine and saturated fat, and a plant-based diet, low in taurine and saturated fat. A mediation model will be used to determine the extent to which diet (independent variable) and mucosal markers of CRC risk and DNA damage (dependent variables) are explained by colonic bacteria and their functions (mediator variables). This research will generate novel information targeted to develop effective dietary interventions that may reduce the unequal CRC burden in AAs.

The 'in vivo lifestyle' of bile acid 7α-dehydroxylating bacteria: comparative genomics, metatranscriptomic, and bile acid metabolomics analysis of a defined microbial community in gnotobiotic mice.

The formation of secondary bile acids by gut microbes is a current topic of considerable biomedical interest. However, a detailed understanding of the biology of anaerobic bacteria in the genus Clostridium that are capable of generating secondary bile acids is lacking. We therefore sought to determine the transcriptional responses of two prominent secondary bile acid producing bacteria, Clostridium hylemonae and Clostridium hiranonis to bile salts (in vitro) and the cecal environment of gnotobiotic mice. The genomes of C. hylemonae DSM 15053 and C. hiranonis DSM 13275 were closed, and found to encode 3,647 genes (3,584 protein-coding) and 2,363 predicted genes (of which 2,239 are protein-coding), respectively, and 1,035 orthologs were shared between C. hylemonae and C. hiranonis. RNA-Seq analysis was performed in growth medium alone, and in the presence of cholic acid (CA) and deoxycholic acid (DCA). Growth with CA resulted in differential expression (>0.58 log2FC; FDR < 0.05) of 197 genes in C. hiranonis and 118 genes in C. hylemonae. The bile acid-inducible operons (bai) from each organism were highly upregulated in the presence of CA but not DCA. We then colonized germ-free mice with human gut bacterial isolates capable of metabolizing taurine-conjugated bile acids. This consortium included bile salt hydrolase-expressing Bacteroides uniformis ATCC 8492, Bacteroides vulgatus ATCC 8482, Parabacteroides distasonis DSM 20701, as well as taurine-respiring Bilophila wadsworthia DSM 11045, and deoxycholic/lithocholic acid generating Clostridium hylemonae DSM 15053 and Clostridium hiranonis DSM 13275. Butyrate and iso-bile acid-forming Blautia producta ATCC 27340 was also included. The Bacteroidetes made up 84.71% of 16S rDNA cecal reads, B. wadsworthia, constituted 14.7%, and the clostridia made up <.75% of 16S rDNA cecal reads. Bile acid metabolomics of the cecum, serum, and liver indicate that the synthetic community were capable of functional bile salt deconjugation, oxidation/isomerization, and 7α-dehydroxylation of bile acids. Cecal metatranscriptome analysis revealed expression of genes involved in metabolism of taurine-conjugated bile acids. The in vivo transcriptomes of C. hylemonae and C. hiranonis suggest fermentation of simple sugars and utilization of amino acids glycine and proline as electron acceptors. Genes predicted to be involved in trimethylamine (TMA) formation were also expressed.

A prospective cohort analysis of gut microbial co-metabolism in Alaska Native and rural African people at high and low risk of colorectal cancer.

BACKGROUND: Alaska Native (AN) people have the world's highest recorded incidence of sporadic colorectal cancer (CRC) (∼91:100,000), whereas rural African (RA) people have the lowest risk (<5:100,000). Previous data supported the hypothesis that diet affected CRC risk through its effects on the colonic microbiota that produce tumor-suppressive or -promoting metabolites. OBJECTIVES: We investigated whether differences in these metabolites may contribute to the high risk of CRC in AN people. METHODS: A cross-sectional observational study assessed dietary intake from 32 AN and 21 RA healthy middle-aged volunteers before screening colonoscopy. Analysis of fecal microbiota composition by 16S ribosomal RNA gene sequencing and fecal/urinary metabolites by 1H-NMR spectroscopy was complemented with targeted quantification of fecal SCFAs, bile acids, and functional microbial genes. RESULTS: Adenomatous polyps were detected in 16 of 32 AN participants, but not found in RA participants. The AN diet contained higher proportions of fat and animal protein and less fiber. AN fecal microbiota showed a compositional predominance of Blautia and Lachnoclostridium, higher microbial capacity for bile acid conversion, and low abundance of some species involved in saccharolytic fermentation (e.g., Prevotellaceae, Ruminococcaceae), but no significant lack of butyrogenic bacteria. Significantly lower concentrations of tumor-suppressive butyrate (22.5 ± 3.1 compared with 47.2 ± 7.3 SEM µmol/g) coincided with significantly higher concentrations of tumor-promoting deoxycholic acid (26.7 ± 4.2 compared with 11 ± 1.9 µmol/g) in AN fecal samples. AN participants had lower quantities of fecal/urinary metabolites than RA participants and metabolite profiles correlated with the abundance of distinct microbial genera in feces. The main microbial and metabolic CRC-associated markers were not significantly altered in AN participants with adenomatous polyps. CONCLUSIONS: The low-fiber, high-fat diet of AN people and exposure to carcinogens derived from diet or environment are associated with a tumor-promoting colonic milieu as reflected by the high rates of adenomatous polyps in AN participants.

Plasticity in the Human Gut Microbiome Defies Evolutionary Constraints.

The gut microbiome of primates, including humans, is reported to closely follow host evolutionary history, with gut microbiome composition being specific to the genetic background of its primate host. However, the comparative models used to date have mainly included a limited set of closely related primates. To further understand the forces that shape the primate gut microbiome, with reference to human populations, we expanded the comparative analysis of variation among gut microbiome compositions and their primate hosts, including 9 different primate species and 4 human groups characterized by a diverse set of subsistence patterns (n = 448 samples). The results show that the taxonomic composition of the human gut microbiome, at the genus level, exhibits increased compositional plasticity. Specifically, we show unexpected similarities between African Old World monkeys that rely on eclectic foraging and human populations engaging in nonindustrial subsistence patterns; these similarities transcend host phylogenetic constraints. Thus, instead of following evolutionary trends that would make their microbiomes more similar to that of conspecifics or more phylogenetically similar apes, gut microbiome composition in humans from nonindustrial populations resembles that of generalist cercopithecine monkeys. We also document that wild cercopithecine monkeys with eclectic diets and humans following nonindustrial subsistence patterns harbor high gut microbiome diversity that is not only higher than that seen in humans engaging in industrialized lifestyles but also higher compared to wild primates that typically consume fiber-rich diets.IMPORTANCE The results of this study indicate a discordance between gut microbiome composition and evolutionary history in primates, calling into question previous notions about host genetic control of the primate gut microbiome. Microbiome similarities between humans consuming nonindustrialized diets and monkeys characterized by subsisting on eclectic, omnivorous diets also raise questions about the ecological and nutritional drivers shaping the human gut microbiome. Moreover, a more detailed understanding of the factors associated with gut microbiome plasticity in primates offers a framework to understand why humans following industrialized lifestyles have deviated from states thought to reflect human evolutionary history. The results also provide perspectives for developing therapeutic dietary manipulations that can reset configurations of the gut microbiome to potentially improve human health.