Bile salt hydrolase catalyses formation of amine-conjugated bile acids.

Bacteria in the gastrointestinal tract produce amino acid bile acid amidates that can affect host-mediated metabolic processes1-6; however, the bacterial gene(s) responsible for their production remain unknown. Herein, we report that bile salt hydrolase (BSH) possesses dual functions in bile acid metabolism. Specifically, we identified a previously unknown role for BSH as an amine N-acyltransferase that conjugates amines to bile acids, thus forming bacterial bile acid amidates (BBAAs). To characterize this amine N-acyltransferase BSH activity, we used pharmacological inhibition of BSH, heterologous expression of bsh and mutants in Escherichia coli and bsh knockout and complementation in Bacteroides fragilis to demonstrate that BSH generates BBAAs. We further show in a human infant cohort that BBAA production is positively correlated with the colonization of bsh-expressing bacteria. Lastly, we report that in cell culture models, BBAAs activate host ligand-activated transcription factors including the pregnane X receptor and the aryl hydrocarbon receptor. These findings enhance our understanding of how gut bacteria, through the promiscuous actions of BSH, have a significant role in regulating the bile acid metabolic network.

Preservation of conjugated primary bile acids by oxygenation of the small intestinal microbiota in vitro.

Bile acids play a critical role in the emulsification of dietary lipids, a critical step in the primary function of the small intestine, which is the digestion and absorption of food. Primary bile acids delivered into the small intestine are conjugated to enhance functionality, in part, by increasing aqueous solubility and preventing passive diffusion of bile acids out of the gut lumen. Bile acid function can be disrupted by the gut microbiota via the deconjugation of primary bile acids by bile salt hydrolases (BSHs), leading to their conversion into secondary bile acids through the expression of bacterial bile acid-inducible genes, a process often observed in malabsorption due to small intestinal bacterial overgrowth. By modeling the small intestinal microbiota in vitro using human small intestinal ileostomy effluent as the inocula, we show here that the infusion of physiologically relevant levels of oxygen, normally found in the proximal small intestine, reduced deconjugation of primary bile acids, in part, through the expansion of bacterial taxa known to have a low abundance of BSHs. Further recapitulating the small intestinal bile acid composition of the small intestine, limited conversion of primary into secondary bile acids was observed. Remarkably, these effects were preserved among four separate communities, each inoculated with a different small intestinal microbiota, despite a high degree of taxonomic variability under both anoxic and aerobic conditions. In total, these results provide evidence for a previously unrecognized role that the oxygenated environment of the small intestine plays in the maintenance of normal digestive physiology. IMPORTANCE: Conjugated primary bile acids are produced by the liver and exist at high concentrations in the proximal small intestine, where they are critical for proper digestion. Deconjugation of these bile acids with subsequent transformation via dehydroxylation into secondary bile acids is regulated by the colonic gut microbiota and reduces their digestive function. Using an in vitro platform modeling the small intestinal microbiota, we analyzed the ability of this community to transform primary bile acids and studied the effect of physiological levels of oxygen normally found in the proximal small intestine (5%) on this metabolic process. We found that oxygenation of the small intestinal microbiota inhibited the deconjugation of primary bile acids in vitro. These findings suggest that luminal oxygen levels normally found in the small intestine may maintain the optimal role of bile acids in the digestive process by regulating bile acid conversion by the gut microbiota.

Surgery for Crohn's Disease Is Associated With a Dysbiotic Microbiome and Metabolome: Results From Two Prospective Cohorts.

BACKGROUND & AIMS: Crohn's disease is associated with alterations in the gut microbiome and metabolome described as dysbiosis. We characterized the microbial and metabolic consequences of ileal resection, the most common Crohn's disease surgery. METHODS: Patients with and without intestinal resection were identified from the Diet to Induce Remission in Crohn's Disease and Study of a Prospective Adult Research Cohort with Inflammatory Bowel Disease studies. Stool samples were analyzed with shotgun metagenomics sequencing. Fecal butyrate was measured with 1H nuclear magnetic resonance spectroscopy. Fecal bile acids and plasma 7α-hydroxy-4-cholesten-3-one (C4) was measured with mass spectrometry. RESULTS: Intestinal resection was associated with reduced alpha diversity and altered beta diversity with increased Proteobacteria and reduced Bacteroidetes and Firmicutes. Surgery was associated with higher representation of genes in the KEGG pathway for ABC transporters and reduction in genes related to bacterial metabolism. Surgery was associated with reduced concentration of the But gene but this did not translate to reduced fecal butyrate concentration. Surgery was associated with decreased abundance of bai operon genes, with increased plasma C4 concentration, increased primary bile acids and reduced secondary bile acids, including isoLCA. Additionally, Egerthella lenta, Adlercreutzia equalofaciens, and Gordonibacter pamelaeae were lower in abundance among patients with prior surgery in both cohorts. CONCLUSIONS: In 2 different populations, prior surgery in Crohn's disease is associated with altered fecal microbiome. Patients who had undergone ileal resection had reduction in the potentially beneficial bacteria E lenta and related actinobacteria and secondary bile acids, including isoLCA, suggesting that these could be biomarkers of patients at higher risk for disease progression.

Disruption of intestinal oxygen balance in acute colitis alters the gut microbiome.

The juxtaposition of well-oxygenated intestinal colonic tissue with an anerobic luminal environment supports a fundamentally important relationship that is altered in the setting of intestinal injury, a process likely to be relevant to diseases such as inflammatory bowel disease. Herein, using two-color phosphorometry to non-invasively quantify both intestinal tissue and luminal oxygenation in real time, we show that intestinal injury induced by DSS colitis reduces intestinal tissue oxygenation in a spatially defined manner and increases the flux of oxygen from the tissue into the gut lumen. By characterizing the composition of the microbiome in both DSS colitis-affected gut and in a bioreactor containing a stable human fecal community exposed to microaerobic conditions, we provide evidence that the increased flux of oxygen into the gut lumen augments glycan degrading bacterial taxa rich in glycoside hydrolases which are known to inhabit gut mucosal surface. Continued disruption of the intestinal mucus barrier through such a mechanism may play a role in the perpetuation of the intestinal inflammatory process.

Dietary carbohydrates regulate intestinal colonization and dissemination of Klebsiella pneumoniae.

Bacterial translocation from the gut microbiota is a source of sepsis in susceptible patients. Previous work suggests that overgrowth of gut pathobionts, including Klebsiella pneumoniae, increases the risk of disseminated infection. Our data from a human dietary intervention study found that, in the absence of fiber, K. pneumoniae bloomed during microbiota recovery from antibiotic treatment. We thus hypothesized that dietary nutrients directly support or suppress colonization of this gut pathobiont in the microbiota. Consistent with our study in humans, complex carbohydrates in dietary fiber suppressed the colonization of K. pneumoniae and allowed for recovery of competing commensals in mouse models. In contrast, through ex vivo and in vivo modeling, we identified simple carbohydrates as a limiting resource for K. pneumoniae in the gut. As proof of principle, supplementation with lactulose, a nonabsorbed simple carbohydrate and an FDA-approved therapy, increased colonization of K. pneumoniae. Disruption of the intestinal epithelium led to dissemination of K. pneumoniae into the bloodstream and liver, which was prevented by dietary fiber. Our results show that dietary simple and complex carbohydrates were critical not only in the regulation of pathobiont colonization but also disseminated infection, suggesting that targeted dietary interventions may offer a preventative strategy in high-risk patients.

Challenges in IBD Research 2024: Environmental Triggers.

Environmental factors play an important role in inflammatory bowel diseases (IBD; Crohn's disease, [CD], ulcerative colitis [UC]). As part of the Crohn's & Colitis Challenges 2024 agenda, the Environmental Triggers workgroup summarized the progress made in the field of environmental impact on IBD since the last Challenges cycle in this document. The workgroup identified 4 unmet gaps in this content area pertaining to 4 broad categories: (1) Epidemiology; (2) Exposomics and environmental measurement; (3) Biologic mechanisms; and (4) Interventions and Implementation. Within epidemiology, the biggest unmet gaps were in the study of environmental factors in understudied populations including racial and ethnic minority groups and in populations witnessing rapid rise in disease incidence globally. The workgroup also identified a lack of robust knowledge of how environmental factors may impact difference stages of the disease and for different disease-related end points. Leveraging existing cohorts and targeted new prospective studies were felt to be an important need for the field. The workgroup identified the limitations of traditional questionnaire-based assessment of environmental exposure and placed high priority on the identification of measurable biomarkers that can quantify cross-sectional and longitudinal environmental exposure. This would, in turn, allow for identifying the biologic mechanisms of influence of environmental factors on IBD and understand the heterogeneity in effect of such influences. Finally, the working group emphasized the importance of generating high-quality data on effective environmental modification on an individual and societal level, and the importance of scalable and sustainable methods to deliver such changes.

Environmental factors are important in inflammatory bowel diseases. It is a high priority to identify environmental factors impacting different disease stages and in different populations, develop biomarkers for such exposures, and generate evidence for modifying them to improve outcomes.

The intestinal microbiome of inflammatory bowel disease across the pediatric age range.

Dysbiosis is associated with pediatric and adult-onset inflammatory bowel disease (IBD), but the role of dysbiosis and the microbiome in very early onset IBD (VEO-IBD) has not yet been described. Here, we aimed to demonstrate the impact of age and inflammation on microbial community structure using shotgun metagenomic sequencing in children with VEO-IBD, pediatric-onset IBD, and age-matched pediatric healthy controls (HC) observed longitudinally over the course of 8 weeks. We found disease-related differences in alpha and beta diversity between HC and children with IBD or VEO-IBD. Using a healthy microbial maturity index modeled from HC across the age range to characterize their gut microbiota, we found that children with pediatric-onset IBD and VEO-IBD had lower maturity than their age-matched HC groups, suggesting a disease effect on the microbial community. In addition, patients with pediatric IBD had significantly lower maturity than those with VEO-IBD, who had more heterogeneity at the youngest ages, highlighting differences in these two cohorts that were not captured in standard comparisons of alpha and beta diversity. These results demonstrate that young age and inflammation independently impact microbial community structure. However, the effect is not additive in the youngest patients, likely because of the heterogeneous and dynamic stool microbiome in this population.

Neutrophil-mediated hypoxia drives pathogenic CD8+ T cell responses in cutaneous leishmaniasis.

Cutaneous leishmaniasis caused by Leishmania parasites exhibits a wide range of clinical manifestations. Although parasites influence disease severity, cytolytic CD8+ T cell responses mediate disease. Although these responses originate in the lymph node, we found that expression of the cytolytic effector molecule granzyme B was restricted to lesional CD8+ T cells in Leishmania-infected mice, suggesting that local cues within inflamed skin induced cytolytic function. Expression of Blimp-1 (Prdm1), a transcription factor necessary for cytolytic CD8+ T cell differentiation, was driven by hypoxia within the inflamed skin. Hypoxia was further enhanced by the recruitment of neutrophils that consumed oxygen to produce ROS and ultimately increased the hypoxic state and granzyme B expression in CD8+ T cells. Importantly, lesions from patients with cutaneous leishmaniasis exhibited hypoxia transcription signatures that correlated with the presence of neutrophils. Thus, targeting hypoxia-driven signals that support local differentiation of cytolytic CD8+ T cells may improve the prognosis for patients with cutaneous leishmaniasis, as well as for other inflammatory skin diseases in which cytolytic CD8+ T cells contribute to pathogenesis.