Bacterial Bile Salt Hydrolase Research Articles

Zhi-Kang-Yin formula attenuates high-fat diet-induced metabolic disorders through modulating gut microbiota-bile acids axis in mice

BackgroundMetabolic disorders have become one of the global medical problems. Due to the complexity of its pathogenesis, there is still no effective treatment. Bile acids (BAs) and gut microbiota (GM) have been proved to be closely related to host metabolism, which could be important targets for metabolic disorders. Zhi-Kang-Yin (ZKY) is a traditional Chinese medicine (TCM) formula developed by the research team according to theory of TCM and has been shown to improve metabolism in clinic. However, the underlying mechanisms are unclear.Aim of the studyThis study aimed to investigate the potential mechanisms of the beneficial effect of ZKY on metabolism.MethodsHigh-fat diet (HFD)-fed mice were treated with and without ZKY. The glucose and lipid metabolism-related indexes were measured. BA profile, GM composition and hepatic transcriptome were then investigated to analyze the changes of BAs, GM, and hepatic gene expression. Moreover, the relationship between GM and BAs was identified with functional gene quantification and ex vivo fermentation experiment.ResultsZKY reduced weight gain and lipid levels in both liver and serum, attenuated hepatic steatosis and improved glucose tolerance in HFD-fed mice. BA profile detection showed that ZKY changed the composition of BAs and increased the proportion of unconjugated BAs and non-12-OH BAs. Hepatic transcriptomic analysis revealed fatty acid metabolism and BA biosynthesis related pathways were regulated. In addition, ZKY significantly changed the structure of GM and upregulated the gene copy number of bacterial bile salt hydrolase. Meanwhile, ZKY directly promoted the growth of Bifidobacterium, which is a well-known bile salt hydrolase-producing genus. The ex vivo co-culture experiment with gut microbiota and BAs demonstrated that the changes of BAs profile in ZKY group were mediated by ZKY-shifted GM, which led to increased expression of genes associated with fatty acid degradation in the liver.ConclusionOur study indicated that the effect of ZKY on improving metabolism is associated with the modulation of GM-BAs axis, especially, by upregulating the abundance of bile salt hydrolase-expression bacteria and increasing the levels of unconjugated BAs. This study indicates that GM-BAs axis might be an important pathway for improving metabolic disorders by ZKY.

Chronic sucralose consumption inhibits farnesoid X receptor signaling and perturbs lipid and cholesterol homeostasis in the mouse livers, potentially by altering gut microbiota functions

Sucralose has raised concerns regarding its safety and recent studies have demonstrated that sucralose consumption can disrupt the normal gut microbiome and alter metabolic profiles in mice. However, the extent to which this perturbation affects the functional interaction between the microbiota and the host, as well as its potential impact on host health, remains largely unexplored. Here, we aimed to investigate whether chronic sucralose consumption, at levels within the Acceptable Daily Intake (ADI), could disturb key gut microbial functions and lead to adverse health effects in mice. Following six-month sucralose consumption, several bacterial genera associated with bile acid metabolism were decreased, including Lactobacillus and Ruminococcus. Consequently, the richness of secondary bile acid biosynthetic pathway and bacterial bile salt hydrolase gene were decreased in the sucralose-treated gut microbiome. Compared to controls, sucralose-consuming mice exhibited significantly lower ratios of free bile acids and taurine-conjugated bile acids in their livers. Additionally, several farnesoid X receptor (FXR) agonists were decreased in sucralose-treated mice. This reduction in hepatic FXR activation was associated with altered expression of down-stream genes, in the liver. Moreover, the expression of key lipogenic genes was up-regulated in the livers of sucralose-treated mice. Changes in hepatic lipid profiles were also observed, characterized by lower ceramide levels, a decreased PC/PE ratio, and a mildly increase in lipid accumulation. Additionally, sucralose-consumed mice exhibited higher hepatic cholesterol level compared to control mice, with up-regulation of cholesterol efflux genes and down-regulation of genes associated with reverse cholesterol transport. In conclusion, chronic sucralose consumption disrupts FXR signaling activation and perturbs hepatic lipid and cholesterol homeostasis, potentially by diminishing the bile acid metabolic capacity of the gut microbiome. These findings shed light on the complex interplay between sucralose, the gut microbiota, and host metabolism, raising important questions about the safety of its long-term consumption.

Modification of bile acids metabolism with multi-strain probiotic in patients with diarrhea predominant irritable bowel syndrome: a randomized study

one of the irritable bowel syndrome with diarrhea (IBS-D) mechanisms of development isbile acids (BA) malabsorption (BAM). The promising new therapeutic approach for BAM – probiotic
 bacteria producing bile salt hydrolase (BSH). The aim of the study was to compare the effect of multi-strain probiotic and cholestyramine combination with cholestyramine monotherapy on modifying the
 parameters of BA metabolism in IBS-D patients. Materials and methods. The trial was conducted asa randomized, open, parallel study and included 108 patients with IBS-D divided into 2 groups: casegroup (n=57) that received combination of probiotic (L. rhamnosus, L. plantarum, S. thermophilus, L.acidophilus, B. bifidum, B. longum, B. infantis, S. boulardii) with cholestyramine, and control group(n=51) that received cholestyramine monotherapy during 12 weeks. The total relative activity (RA)of gut bacterial BSH, serum BA (sBA) and fecal BA (fBA) were assessed in all patients. Results: total,
 primary and secondary sBA and fBA, proportions of primary and secondary fBA were significantly dif-ferent after 12 weeks within each group comparing to baseline, p<0.05. In case group the proportions of
 primary and secondary sBA, RA of gut bacterial BSH were significantly different after 12 weeks compar-ing to baseline, p<0.05. All the parameters of BA metabolism except absolute levels of secondary sBA
 were significantly different after 12 weeks between the control and case groups, p<0.05. Conclusions:
 combination of cholestyramine and multi-strain probiotic led to more pronounced alterations of BA me-tabolism and increasing of gut bacterial BSH-activity comparing to cholestyramine monotherapy. It may
 explain possible mechanism of action of probiotics in patients with IBS-D and assumed BAM.

Fluorogenic substrates and pre‐column derivatization for monitoring the activity of bile salt hydrolase from Clostridium perfringens

The bile acid pool has a profound impact on human health and disease. The intestinal microbiota initiates the metabolism of conjugated bile acids through a critical first step catalyzed by bacterial bile salt hydrolase (BSH) and provides unique contributions to the diversity of bile acids. There has been great interest in surveying BSH activity. We compared two substrates with either 2-(7-amino-4-methyl-coumarinyl)acetic acid or 7-amino-4-methyl-coumarin as fluorescent reporters of BSH activity. The BSH-catalyzed conversion of the natural substrate taurocholic acid was followed through an HPLC-based assay by applying 7-nitrobenzo[c][1,2,5]oxadiazole as scavenger for taurine, released in the enzymatic reaction. Hence, a new opportunity to monitor the activity of bile salt hydrolases was introduced.

High-grain diet feeding alters ileal microbiota and disrupts bile acid metabolism in lactating dairy cows.

Bile acids (BAs) play an important role in the regulation of lipid metabolic homeostasis, but little is known about their metabolism in dairy cows fed a high-grain (HG) diet. In the present study, we examined the bacterial community, BA profile, and the FXR/FGF19 signaling pathway in the ileum and liver to investigate the gut microbe-BA metabolism interactions response to HG diet and the changes in the subsequent enterohepatic circulation of dairy cows. The results showed that the ileal bacterial community was altered, with an increase of Paraclostridium, Anaerobutyricum, Shuttleworthia, and Stomatobaculum in the relative abundance in the HG group. Moreover, real-time polymerase chain reaction (PCR) showed that the abundance of total bacteria and bacterial bile-salt hydrolase (BSH) genes was increased in the ileal digesta in the HG group. Meanwhile, HG feeding also decreased the total BA content in the digesta of jejunum and ileum and in feces. HG feeding altered the BA profile in the ileal digesta by increasing unconjugated BAs and decreasing conjugated BAs. In addition, the intestinal FXR/FGF19 signaling pathway was activated. The expression of CYP7A1 (cholesterol 7α-hydroxylase) was depressed, which inhibited BAs synthesis in the liver of cows fed HG. Overall, HG feeding altered the ileal bacterial community and BA profile, and activated FXR/FGF19 signaling pathway, resulting in a decrease of BA level in the ileal digesta via the inhibition of hepatic BA synthesis. The findings provided novel insights into understanding the relationship between gut microbiota and the homeostasis of BAs in dairy cows fed a HG diet.

Caffeic acid phenethyl ester suppresses intestinal FXR signaling and ameliorates nonalcoholic fatty liver disease by inhibiting bacterial bile salt hydrolase activity.

Propolis is commonly used in traditional Chinese medicine. Studies have demonstrated the therapeutic effects of propolis extracts and its major bioactive compound caffeic acid phenethyl ester (CAPE) on obesity and diabetes. Herein, CAPE was found to have pharmacological activity against nonalcoholic fatty liver disease (NAFLD) in diet-induced obese mice. CAPE, previously reported as an inhibitor of bacterial bile salt hydrolase (BSH), inhibited BSH enzymatic activity in the gut microbiota when administered to mice. Upon BSH inhibition by CAPE, levels of tauro-β-muricholic acid were increased in theintestine and selectively suppressed intestinal farnesoid X receptor (FXR) signaling. This resulted in lowering of the ceramides in theintestine that resulted from increased diet-induced obesity. Elevated intestinal ceramides are transported to the liver where they promoted fat production. Lowering FXR signaling was also accompanied by increased GLP-1 secretion. In support of this pathway, the therapeutic effects of CAPE on NAFLD were absent in intestinal FXR-deficient mice, and supplementation of mice with C16-ceramide significantly exacerbated hepatic steatosis. Treatment of mice with an antibiotic cocktail to deplete BSH-producing bacteria also abrogated the therapeutic activity of CAPE against NAFLD. These findings demonstrate that CAPE ameliorates obesity-related steatosis at least partly through the gut microbiota-bile acid-FXR pathway via inhibiting bacterial BSH activity and suggests that propolis enriched with CAPE might serve as a promising therapeutic agent for the treatment of NAFLD.

Defective Humoral Immunity Disrupts Bile Acid Homeostasis which Promotes Inflammatory Disease of the Small Bowel

Abstract Mucosal antibodies maintain gut homeostasis by promoting spatial segregation between host tissues and luminal microbes. Whether and how mucosal antibody responses influence gut health through modulation of microbiota composition is unclear. Here, we use a CD19−/− mouse model of antibody-deficiency to demonstrate that a relationship exists between dysbiosis, defects in bile acid homeostasis, and gluten-sensitive enteropathy of the small intestine. The gluten-sensitive small intestine enteropathy that develops in CD19−/− mice is associated with alterations to luminal bile acid composition in the SI, marked by significant reductions in the abundance of conjugated bile acids. Manipulation of bile acid availability, adoptive transfer of functional B cells, and ablation of bacterial bile salt hydrolase activity all influence the severity of small intestine enteropathy in CD19−/− mice. Collectively, results from our experiments support a model whereby mucosal humoral immune responses limit inflammatory disease of the small bowel by regulating bacterial BA metabolism. J.L.K. was supported by a NIAID K22 Award (AI123481), a Jeffrey Modell Network Specific Diseases Research Award, a University of South Carolina ASPIRE-I Award, a pilot project award through the University of South Carolina Center for Alternative Medicine COBRE program (P20GM103641; awarded to Drs. Mitzi and Prakash Nagarkatti), an R21 (AI142409), and 1R01AI155887.

Defective humoral immunity disrupts bile acid homeostasis which promotes inflammatory disease of the small bowel

Mucosal antibodies maintain gut homeostasis by promoting spatial segregation between host tissues and luminal microbes. Whether and how mucosal antibody responses influence gut health through modulation of microbiota composition is unclear. Here, we use a CD19−/− mouse model of antibody-deficiency to demonstrate that a relationship exists between dysbiosis, defects in bile acid homeostasis, and gluten-sensitive enteropathy of the small intestine. The gluten-sensitive small intestine enteropathy that develops in CD19−/− mice is associated with alterations to luminal bile acid composition in the SI, marked by significant reductions in the abundance of conjugated bile acids. Manipulation of bile acid availability, adoptive transfer of functional B cells, and ablation of bacterial bile salt hydrolase activity all influence the severity of small intestine enteropathy in CD19−/− mice. Collectively, results from our experiments support a model whereby mucosal humoral immune responses limit inflammatory disease of the small bowel by regulating bacterial BA metabolism.

Активность кишечной бактериальной гидролазы солей желчных кислот коррелирует с сердечно-сосудистым риском: исследование «случай — контроль»

Актуальность. Уровень сывороточного холестерина частично регулируется благодаря метаболизму желчных кислот в кишечнике, зависящему от активности бактериальной гидролазы солей желчных кислот (ГСЖК). Существующие данные относительно четкого влияния ГСЖК на липидный обмен и сердечно-сосудистый риск недостаточны и противоречивы. Целью исследования было оценить взаимо­связь между относительной активностью (ОА) кишечной бактериальной ГСЖК и уровнями сывороточного холестерина и сердечно-сосудистым риском (ССР). Материалы и методы. Исследование проводилось по дизайну «случай — контроль» и включало 26 относительно здоровых участников (контрольная группа) и 77 пациентов с дислипидемией, без анамнеза тяжелых сердечно-сосудистых событий (основная группа). У участников были определены общая ОА кишечной бактериальной ГСЖК, показатели липидного профиля и уровни ССР по 5 шкалам риска. Результаты. ОА кишечной бактериальной ГСЖК была выше у здоровых участников по сравнению с участниками с дислипидемией (р < 0,001). Была выявлена отрицательная корреляционная связь умеренной силы между ОА ГСЖК и общим холестерином (ОХ) (–0,38) и липопротеинами низкой плотности (ЛПНП) (–0,36) с линейным соотношением, которое определялось уравнением: ЛПНП = –5,33 • ОА ГСЖК + 4,479. Было обнаружено, что с увеличением ОА ГСЖК риск дислипидемии снижается (р < 0,001), ОШ = 1,06 • 10–10 (95% ДИ; 2,5 • 10–15 – 4,5 • 10–6). Выявлена умеренная отрицательная корреляционная взаимосвязь между ОА ГСЖК и ССР, оцененным по шкалам Globorisk (–0,34), Framingham (–0,34), алгоритму ACC/AHA 2013 (–0,32), PROCAM (–0,35) и шкале ВОЗ (–0,34). Выводы. Общая ОА кишечной бактериальной ГСЖК негативно коррелировала с ОХ, ЛПНП, ССР, оцененным по 5 шкалам, и негативно ассоциировалась с риском дислипидемии.

A Gut-Restricted Lithocholic Acid Analog as an Inhibitor of Gut Bacterial Bile Salt Hydrolases.

Bile acids play crucial roles in host physiology by acting both as detergents that aid in digestion and as signaling molecules that bind to host receptors. Gut bacterial bile salt hydrolase (BSH) enzymes perform the gateway reaction leading to the conversion of host-produced primary bile acids into bacterially modified secondary bile acids. Small molecule probes that target BSHs will help elucidate the causal roles of these metabolites in host physiology. We previously reported the development of a covalent BSH inhibitor with low gut permeability. Here, we build on our previous findings and describe the development of a second-generation gut-restricted BSH inhibitor with enhanced potency, reduced off-target effects, and durable in vivo efficacy. Structure-activity relationship (SAR) studies focused on the bile acid core identified a compound, AAA-10, containing a C3-sulfonated lithocholic acid scaffold and an alpha-fluoromethyl ketone warhead as a potent pan-BSH inhibitor. This compound inhibits BSH activity in mouse and human fecal slurry, bacterial cultures, and purified BSH proteins and displays reduced toxicity against mammalian cells compared to first generation compounds. Oral administration of AAA-10 to wild-type mice for 5 days resulted in a decrease in the abundance of the secondary bile acids deoxycholic acid (DCA) and lithocholic acid (LCA) in the mouse GI tract with low systemic exposure of AAA-10, demonstrating that AAA-10 is an effective tool for inhibiting BSH activity and modulating bile acid pool composition in vivo.

Defective Humoral Immunity Disrupts Bile Acid Homeostasis Which Promotes Inflammatory Disease of the Small Bowel

Abstract Mucosal antibodies maintain gut homeostasis by promoting spatial segregation between host tissues and luminal microbes. Whether and how mucosal antibody responses influence gut health through modulation of microbiota composition is unclear. One critical metabolic function carried out exclusively by bacteria in the gut is bio-transformation of host bile acids (BAs), and dysregulation of BAs metabolism has been linked to numerous inflammatory and metabolic diseases in humans. Here, we use a CD19−/− mouse model of antibody-deficiency to demonstrate that a relationship exists between dysbiosis, defects in BA homeostasis, and enteropathy of the small intestine (SI). SI enteropathy that develops in CD19−/− mice is associated with alterations to the luminal BA pool in the SI, marked by significant reductions in the abundance of conjugated BAs. Manipulation of BA availability, adoptive transfer of functional B cells, and ablation of bacterial bile salt hydrolase (bsh) activity all influence the severity of SI enteropathy in CD19−/− mice. Collectively, results from our experiments support a model whereby mucosal humoral immune responses limit inflammatory disease of the small bowel by regulating bacterial BA metabolism.

Lactobacillus bile salt hydrolase substrate specificity governs bacterial fitness and host colonization

Primary bile acids (BAs) are a collection of host-synthesized metabolites that shape physiology and metabolism. BAs transit the gastrointestinal tract and are subjected to a variety of chemical transformations encoded by indigenous bacteria. The resulting microbiota-derived BA pool is a mediator of host-microbiota interactions. Bacterial bile salt hydrolases (BSHs) cleave the conjugated glycine or taurine from BAs, an essential upstream step for the production of deconjugated and secondary BAs. Probiotic lactobacilli harbor a considerable number and diversity of BSHs; however, their contribution to Lactobacillus fitness and colonization remains poorly understood. Here, we define and compare the functions of multiple BSHs encoded by Lactobacillus acidophilus and Lactobacillus gasseri Our genetic and biochemical characterization of lactobacilli BSHs lend to a model of Lactobacillus adaptation to the gut. These findings deviate from previous notions that BSHs generally promote colonization and detoxify bile. Rather, we show that BSH enzymatic preferences and the intrinsic chemical features of various BAs determine the toxicity of these molecules during Lactobacillus growth. BSHs were able to alter the Lactobacillus transcriptome in a BA-dependent manner. Finally, BSHs were able to dictate differences in bacterial competition in vitro and in vivo, defining their impact on BSH-encoding bacteria within the greater gastrointestinal tract ecosystem. This work emphasizes the importance of considering the enzymatic preferences of BSHs alongside the conjugated/deconjugated BA-bacterial interaction. These results deepen our understanding of the BA-microbiome axis and provide a framework to engineer lactobacilli with improved bile resistance and use probiotics as BA-altering therapeutics.

MODIFICATION OF GUT BACTERIAL BILE SALT HYDROLASE ACTIVITY AND CARDIOVASCULAR RISK: A RANDOMIZED STUDY

Introduction: secretion of bacterial bile salt hydrolase (BSH) is one of the main mechanisms by which gut microbiota play role in cholesterol metabolism. There are limiting and controversial data regarding the clear effect of gut BSH activity correction on modification of serum cholesterol and cardiovascular risk (CVR). Aim of investigation was to evaluate the relationship between modification of the gut bacterial BSH relative activity (RA) by probiotic L. plantarum and serum cholesterol with CVR levels. Methods: the study was conducted as open, comparative, randomized, parallel and included 26 almost healthy participants (healthy control group) and 77 patients with dyslipidemia and without anamnesis of major cardiovascular events, that were divided in two groups: main treatment group (n=41) received combination therapy (capsules with Lactobacillus plantarum in the amount of 2*109 CFU one time a day and tablets simvastatin 20 mg one time a day) and control treatment group (n=36) received monotherapy (simvastatin 20 mg one time a day) during 12 weeks. Before and after 12 weeks of treatment the assessment of total RA of gut BSH, lipid profile and CVR level according to 5 risk scores were performed. Results: at baseline the RA of BSH was higher in healthy adults comparing to participants with dyslipidemia (p<0,001); after 12 weeks of treatment there wasn`t difference between healthy control and only main treatment groups (р=0,45). It was found that with increasing of RA of gut bacterial BSH, the risk of failure of treatment efficacy endpoints achievement (³20% reduction of values) decreased regarding: total cholesterol (TC) (p=0,0306), OR=0,00133 (95% CI; 3,28*10-6-0,538); low-density lipoproteins (LDL) (p<0,001), OR=5,65*10-14(95% CI; 6,38*10-20-5*10-8); CVR level according to Framingham score (р=0,0035), OR=4,09*10-5(95% CI; 4,66*10-8-0,0359); CVR level according to 2013 ACC/AHA algorithm (р=0,0135), OR=3,8*10-4(95% CI; 7,34*10-7-0,197); CVR level according to PROCAM score (p=0,00125), OR=8,38*10-6 (95%; CI; 6,93*10-9-0,0101). Conclusions: additional supplementation with BSH-producing bacteria L. plantarum was more effective in increasing of BSH activity compared to simvastatin monotherapy. Increasing of BSH RA by L. plantarum was associated with higher chances to achieve treatment efficacy goals regarding reduction of TC, LDL and CVR levels according to Framingham, 2013 ACC/AHA algorithm and PROCAM scores.

MODIFICATION OF GUT BACTERIAL BILE SALT HYDROLASE ACTIVITY AND CARDIOVASCULAR RISK: A RANDOMIZED STUDY

Introduction: secretion of bacterial bile salt hydrolase (BSH) is one of the main mechanisms by which gut microbiota play role in cholesterol metabolism. There are limiting and controversial data regarding the clear effect of gut BSH activity correction on modification of serum cholesterol and cardiovascular risk (CVR). Aim of investigation was to evaluate the relationship between modification of the gut bacterial BSH relative activity (RA) by probiotic L. plantarum and serum cholesterol with CVR levels. Methods: the study was conducted as open, comparative, randomized, parallel and included 26 almost healthy participants (healthy control group) and 77 patients with dyslipidemia and without anamnesis of major cardiovascular events, that were divided in two groups: main treatment group (n=41) received combination therapy (capsules with Lactobacillus plantarum in the amount of 2*109 CFU one time a day and tablets simvastatin 20 mg one time a day) and control treatment group (n=36) received monotherapy (simvastatin 20 mg one time a day) during 12 weeks. Before and after 12 weeks of treatment the assessment of total RA of gut BSH, lipid profile and CVR level according to 5 risk scores were performed. Results: at baseline the RA of BSH was higher in healthy adults comparing to participants with dyslipidemia (p<0,001); after 12 weeks of treatment there wasn`t difference between healthy control and only main treatment groups (р=0,45). It was found that with increasing of RA of gut bacterial BSH, the risk of failure of treatment efficacy endpoints achievement (≥20% reduction of values) decreased regarding: total cholesterol (TC) (p=0,0306), OR=0,00133 (95% CI; 3,28*10-6-0,538); low-density lipoproteins (LDL) (p<0,001), OR=5,65*10-14(95% CI; 6,38*10-20-5*10-8); CVR level according to Framingham score (р=0,0035), OR=4,09*10-5(95% CI; 4,66*10-8-0,0359); CVR level according to 2013 ACC/AHA algorithm (р=0,0135), OR=3,8*10-4(95% CI; 7,34*10-7-0,197); CVR level according to PROCAM score (p=0,00125), OR=8,38*10-6 (95%; CI; 6,93*10-9-0,0101). Conclusions: additional supplementation with BSH-producing bacteria L. plantarum was more effective in increasing of BSH activity compared to simvastatin monotherapy. Increasing of BSH RA by L. plantarum was associated with higher chances to achieve treatment efficacy goals regarding reduction of TC, LDL and CVR levels according to Framingham, 2013 ACC/AHA algorithm and PROCAM scores.

Development of a covalent inhibitor of gut bacterial bile salt hydrolases.

Bile salt hydrolase (BSH) enzymes are widely expressed by human gut bacteria and catalyze the gateway reaction leading to secondary bile acid formation. Bile acids regulate key metabolic and immune processes by binding to host receptors. There is an unmet need for a potent tool to inhibit BSHs across all gut bacteria in order to study the effects of bile acids on host physiology. Here, we report the development of a covalent pan-inhibitor of gut bacterial BSH. From a rationally designed candidate library, we identified a lead compound bearing an alpha-fluoromethyl ketone warhead that modifies BSH at the catalytic cysteine residue. Strikingly, this inhibitor abolished BSH activity in conventional mouse feces. Mice gavaged with a single dose of this compound displayed decreased BSH activity and decreased deconjugated bile acid levels in feces. Our studies demonstrate the potential of a covalent BSH inhibitor to modulate bile acid composition in vivo.

Antibacterial activities of a polyphenolic-rich extract prepared from American cranberry (Vaccinium macrocarpon) fruit pomace against Listeria spp.

Cranberry (Vaccinium macrocarpon) fruits are known for their high polyphenolics content making them a rich source of antioxidants. These polyphenolics have been reported to promote human health and are gaining attention for their antimicrobial activities against foodborne pathogens. We investigated the antimicrobial activity of an ethanolic extract (#FC111-1) prepared from cranberry pomace against Listeria spp. Many polyphenolics were identified in this extract which could be responsible for growth-inhibitory effects against 12 Listeria strains including L. monocytogenes. The minimum inhibitory concentration (MIC) of #FC111-1 determined in cation adjusted Muller Hinton broth (CAMHB) was approximately 2 mg/mL for 11 (91.7%) of these strains. The inclusion of 2–8 mg/mL (1–4 × MIC) of #FC111-1 decreased (>3 log10) viable bacterial cells of all Listeria strains in CAMHB over a 24 h period, while dose-dependently reducing bacterial salt tolerance, bacterial bile-salt hydrolase activity, bacterial biofilm formation capacity, and increasing cell-membrane permeability. The #FC111-1 extract (0.4 and 0.8% concentrations) had no effect on L. monocytogenes survival in a cooked chicken-breast meat model, highlighting the influence of protein-rich matrices on antibacterial activity and the need to consider the role of food composition when using extracts or polyphenolics from cranberry fruits to improve food-safety.

2578. Narrow-Spectrum Antibiotic Treatment of Clostridium difficile Infection Improves Preservation of Intestinal Metabolic Profile

BackgroundCommensal gut bacteria are thought to protect against C. difficile infection (CDI) by producing metabolites that inhibit C. difficile germination and growth. Alternatively, the protective effect could reflect nutrient competition or other mechanisms of chemical communication that also involve the host. CDI treatment using a broad-spectrum antibiotic such as vancomycin (VAN) dramatically depletes commensal bacteria. This dysbiosis can persist for several weeks after end-of-therapy (EOT), and is associated with increased recurrence risk. In this study, we investigate the hypothesis that treating CDI with a more selective antibiotic reduces collateral damage to the intestinal microbiota, preserving or restoring a CDI-inhibitory metabolic profile.MethodsStool samples were collected from CDI patients treated with either a narrow- (RDZ) or broad-spectrum antibiotic (VAN) at days 1, 10 (EOT), 25, and 40. Global metabolite profiles were measured by untargeted LC-MS.ResultsUntargeted metabolite analysis showed broad differences in the metabolic activity of intestinal microbiota of RDZ- and VAN-treated subjects (Figure 1). At EOT, 28% of LC-MS features detected in both RDZ and VAN samples were differentially present (FDR corrected P-value <0.05). Over 80% of the differentially present features were elevated in the RDZ group, indicating diminished capacity of microbiota from VAN subjects to generate diverse metabolic products. Pathway analysis found significant differences in purine, taurine, tyrosine, and bile acid metabolites. The VAN group showed a 5-fold decrease in free taurine, a major conjugation substrate of primary bile acids released by bacterial bile salt hydrolases. VAN treatment also decreased fermentation products of aromatic amino acids and amino sugars derived from mucin degradation. Oligosaccharides were the major metabolite class elevated in VAN subjects.ConclusionOur data suggest that RDZ treatment correlates with enhanced preservation of bacteria-derived ligands regulating intestinal immune function and substrates of bacterial metabolism. These metabolic profile differences between a narrow- and broad-spectrum antibiotic may contribute to their varying efficacy in preventing CDI recurrence. Disclosures All authors: No reported disclosures.

The complex structure of bile salt hydrolase from Lactobacillus salivarius reveals the structural basis of substrate specificity

The gut bacterial bile salt hydrolase (BSH) plays a critical role in host lipid metabolism and energy harvest. Therefore, BSH is a promising microbiome target to develop new therapies to regulate obesity in humans and novel non-antibiotic growth promoters for food animals. We previously reported the 1.90 Å apo crystal structure of BSH from Lactobacillus salivarius (lsBSH). In this study, we soaked the lsBSH crystal with glycocholic acid (GCA), a substrate, and obtained a 2.10 Å structure containing complex of lsBSH bound to GCA and cholic acid (CA), a product. The substrate/product sits in the water-exposed cavity molded by Loops 2 and 3. While the glycine moiety of GCA is exposed into a highly polar pocket, the sterane core of GCA is stabilized by aromatic and hydrophobic interactions. Comparison of product binding with BSH from Clostridium perfringenes reveals a distinct orientation of the sterane core in the binding site. The stability of the substrate-lsBSH complex and the putative catalytic mechanism were explored with molecular dynamics simulations. Site-directed mutagenesis of lsBSH demonstrated that Cys2 and Asn171 are critical for enzymatic activity, while Tyr24, Phe65 and Gln257 contribute to the substrate specificity. Together, this study provides structural insights into BSH-substrate interaction, the mechanism of catalysis and substrate specificity, which facilitate rational design of BSH inhibitors.

Taxonomic profiling and populational patterns of bacterial bile salt hydrolase (BSH) genes based on worldwide human gut microbiome

BackgroundBile salt hydrolase plays an important role in bile acid-mediated signaling pathways, which regulate lipid absorption, glucose metabolism, and energy homeostasis. Several reports suggest that changes in the composition of bile acids are found in many diseases caused by dysbacteriosis.ResultsHere, we present the taxonomic identification of bile salt hydrolase (BSH) in human microbiota and elucidate the abundance and activity differences of various bacterial BSH among 11 different populations from six continents. For the first time, we revealed that bile salt hydrolase protein sequences (BSHs) are distributed in 591 intestinal bacterial strains within 117 genera in human microbiota, and 27.52% of these bacterial strains containing BSH paralogs. Significant variations are observed in BSH distribution patterns among different populations. Based on phylogenetic analysis, we reclassified these BSHs into eight phylotypes and investigated the abundance patterns of these phylotypes among different populations. From the inspection of enzyme activity among different BSH phylotypes, BSH-T3 showed the highest enzyme activity and is only found in Lactobaclillus. The phylotypes of BSH-T5 and BSH-T6 mainly from Bacteroides with high percentage of paralogs exhibit different enzyme activity and deconjugation activity. Furthermore, we found that there were significant differences between healthy individuals and patients with atherosclerosis and diabetes in some phylotypes of BSHs though the correlations were pleiotropic.ConclusionThis study revealed the taxonomic and abundance profiling of BSH in human gut microbiome and provided a phylogenetic-based system to assess BSHs activity by classifying the target sequence into specific phylotype. Furthermore, the present work disclosed the variation patterns of BSHs among different populations of geographical regions and health/disease cohorts, which is essential to understand the role of BSH in the development and progression of related diseases.

A selective gut bacterial bile salt hydrolase alters host metabolism.

The human gut microbiota impacts host metabolism and has been implicated in the pathophysiology of obesity and metabolic syndromes. However, defining the roles of specific microbial activities and metabolites on host phenotypes has proven challenging due to the complexity of the microbiome-host ecosystem. Here, we identify strains from the abundant gut bacterial phylum Bacteroidetes that display selective bile salt hydrolase (BSH) activity. Using isogenic strains of wild-type and BSH-deleted Bacteroides thetaiotaomicron, we selectively modulated the levels of the bile acid tauro-β-muricholic acid in monocolonized gnotobiotic mice. B. thetaiotaomicron BSH mutant-colonized mice displayed altered metabolism, including reduced weight gain and respiratory exchange ratios, as well as transcriptional changes in metabolic, circadian rhythm, and immune pathways in the gut and liver. Our results demonstrate that metabolites generated by a single microbial gene and enzymatic activity can profoundly alter host metabolism and gene expression at local and organism-level scales.