Bile Salt Hydrolase Activity Research Articles

Assessment of the safety and hepatic lipid-lowering effects of Lactobacillus delbrueckii subsp. lactis CKDB001

Probiotics have been shown to provide health benefits for several metabolic diseases, including obesity, type 2 diabetes, and metabolic dysfunction-associated steatotic liver disease (MASLD), by modulating the gut microbiota. In this study, we evaluated the safety and efficacy of Lactobacillus delbrueckii subsp. lactis CKDB001 as a potential therapeutic candidate for the treatment of MASLD. We evaluated antibiotic resistance, hemolytic, gelatinase, and bile salt hydrolase activities, and the production of biogenic amines and D-lactate using in vitro analyses. We found that L. lactis CKDB001 treatment resulted in significant anti-adipogenic properties in the HepG2 cell line, reducing lipid accumulation and improving lipid profiles through mechanisms involving the upregulation of SIRT1 and PPARα, and downregulation of CD36 and ELOVL6. These results suggest that L. lactis CKDB001 is a safe and effective probiotic for managing MASLD. Further in vivo studies and clinical trials are required to validate these effects and fully elucidate their therapeutic potential and safety profiles.

Lactobacillus johnsonii CCFM1376 improves hypercholesterolemia in mice by regulating the composition of bile acids

Aim: Strains with high bile salt hydrolase (BSH) activity have the potential to regulate cholesterol metabolism. This study aimed to assess the alleviating effect of Lactobacillus johnsonii (L. johnsonii ) CCFM1376, a strain with high BSH activity, on mice with hypercholesterolemia and explore the mechanism of its effect through the modulation of bile acid metabolism. Methods: The BSH activity was measured using the ninhydrin method. C57BL/6J mice were given a high-cholesterol diet to induce hypercholesterolemia with simultaneous gavage of L. johnsonii CCFM1376 for 8 weeks. The biochemical parameters in the serum and liver of hypercholesterolemic mice were measured to assess the alleviating effect of L. johnsonii CCFM1376 on hypercholesterolemia. Bile acid content in the mouse liver, serum, distal ileum contents, and feces was determined using liquid chromatograph mass spectrometer (LC-MS). RNA was extracted from mouse ileum and liver, and the expression levels of relative genes implicated in bile acid metabolism were measured by quantitative real-time PCR (qPCR). Results: Compared to the model group, the group treated with L. johnsonii CCFM1376 exhibited significantly reduced levels of serum total cholesterol (TC) and low-density lipoprotein cholesterol (LDL-C), along with a significant increase in high density lipoproteins cholesterol (HDL-C) level. Moreover, hepatic levels of TC and LDL-C in the CCFM1376 group also decreased significantly. Furthermore, the content and amount of unconjugated bile acids in the hepatic-enteric circulation of the L. johnsonii CCFM1376 group significantly increased, and the total bile acid content in the feces also significantly increased. In the L. johnsonii CCFM1376 group, the relative expression levels of ileal farnesoid X receptor (FXR) and fibroblast growth factor 15 (FGF15) were downregulated, while the relative expression level of CYP7A1 was upregulated. Conclusion: These results indicated L. johnsonii CCFM1376 improves hypercholesterolemia in mice by regulating the composition of bile acids. This provides a reference for probiotic strategy to regulate cholesterol metabolism.

PCR Based Technique for Identification of Bile Salt Hydrolase Positive Lactobacilli

In the burgeoning landscape of probiotic research, the intricate interplay between the gut microbiota and host health has underscored the significance of specific enzymatic activities, notably bile salt hydrolase (BSH) produced by Lactobacilli. Harnessing the potential health benefits associated with BSH activity necessitates a meticulous identification strategy. In response, this study introduces a sophisticated PCR-based technique tailored for the rapid and precise discrimination of lactobacilli strains boasting BSH enzymatic prowess. This PCR-based approach offers a valuable tool for researchers and industry professionals involved in the selection and characterization of probiotic strains with bile salt hydrolase activity. Its speed, accuracy, and adaptability make it a promising method for large-scale screenings and applications in diverse fields, ranging from food science to the development of probiotic supplements aimed at enhancing human health through the modulation of bile acid metabolism. robiotic supplementation, where the modulation of bile acid metabolism holds promise for promoting human health and well-being.

Symbiotic probiotic communities with multiple targets successfully combat obesity in high-fat-diet-fed mice

ABSTRACT Probiotics hold great potential for treating metabolic diseases such as obesity. Given the complex and multifactorial nature of these diseases, research on probiotic combination with multiple targets has become popular. Here, we choose four obesity-related targets to perform high-throughput screening, including pancreatic lipase activity, bile salt hydrolase activity, glucagon-like peptide-1 secretion and adipocyte differentiation. Then, we obtained 649 multi-strain combinations with the requirement that each must cover all these targets in principle. After in vitro co-culture and in vivo co-colonization experiments, only four (<0.7%) combinations were selected as symbiotic probiotic communities (SPCs). Next, genome-scale metabolic model analysis revealed that these SPCs showed lower metabolic resource overlap and higher metabolic interaction potential involving amino acid metabolism (Ammonium, L−Lysine, etc.) and energy metabolism (Phosphate, etc.). Further animal experiments demonstrated that all SPCs exhibited a good safety profile and excellent effects in improving obesity and associated glucose metabolism disruptions and depression-like behaviors in high-fat-diet-fed mice. This anti-obesity improvement was achieved through reduced cholesterol level, fat accumulation and inhibited adipocyte differentiation. Taken together, our study provides a new perspective for designing multi-strain combinations, which may facilitate greater therapeutic effect on obesity and other complex diseases in the future.

Fetal programming by the parental microbiome of offspring behavior, and DNA methylation and gene expression within the hippocampus.

The microorganisms colonizing the gastrointestinal tract of animals, collectively referred to as the gut microbiome, affect numerous host behaviors dependent on the central nervous system (CNS). Studies comparing germ-free mice to normally colonized mice have demonstrated influences of the microbiome on anxiety-related behaviors, voluntary activity, and gene expression in the CNS. Additionally, there is epidemiologic evidence supporting an intergenerational influence of the maternal microbiome on neurodevelopment of offspring and behavior later in life. There is limited experimental evidence however directly linking the maternal microbiome to long-term neurodevelopmental outcomes, or knowledge regarding mechanisms responsible for such effects. Here we show that that the maternal microbiome has a dominant influence on several offspring phenotypes including anxiety-related behavior, voluntary activity, and body weight. Adverse outcomes in offspring were associated with features of the maternal microbiome including bile salt hydrolase activity gene expression ( bsh ), abundance of certain bile acids, and hepatic expression of Slc10a1 . In cross-foster experiments, offspring resembled their birth dam phenotypically, despite faithful colonization in the postnatal period with the surrogate dam microbiome. Genome-wide methylation analysis of hippocampal DNA identified microbiome- associated differences in methylation of 196 loci in total, 176 of which show conserved profiles between mother and offspring. Further, single-cell transcriptional analysis revealed accompanying differences in expression of several differentially methylated genes within certain hippocampal cell clusters, and vascular expression of genes associated with bile acid transport. Inferred cell-to-cell communication in the hippocampus based on coordinated ligand-receptor expression revealed differences in expression of neuropeptides associated with satiety. Collectively, these data provide proof-of-principle that the maternal gut microbiome has a dominant influence on the neurodevelopment underlying certain offspring behaviors and activities, and selectively affects genome methylation and gene expression in the offspring CNS in conjunction with that neurodevelopment.

Fermentation dynamics of bile salt hydrolase production in Heyndrickxia coagulans ATCC 7050 and Lactiplantibacillus plantarum ATCC 10012: Addressing ninhydrin assay limitations with a novel HPTLC–MS method

Bile salt hydrolase (BSH), a pivotal enzyme in cholesterol management, holds significant promise in both human and animal subjects. This study investigated the effect of fermentation dynamics in Heyndrickxia coagulans ATCC 7050 and Lactiplantibacillus plantarum ATCC 10012 to enhance BSH production. Cultivation of cultures in MRS and M17 media revealed that MRS medium enhanced BSH production by 235.98 % in H. coagulans ATCC 7050 and 147.37 % in L. plantarum ATCC 10012, compared to M 17 medium. Additionally, varying oxygen concentration levels indicated that H. coagulans ATCC 7050 exhibited its minimum doubling time of 79.8 ± 0.64 min in anaerobic conditions, whereas L.plantarum ATCC 10012 demonstrated its minimum doubling time of 85.5 ± 1.2 min under microaerophilic conditions. However, their highest BSH activity was observed during the stationary phase under anaerobic conditions, yielding 17.14 ± 0.78 U/mL by H. coagulans ATCC 7050 and 19.04 ± 0.81 U/mL by L.plantarum ATCC 10012. Furthermore, it was observed that both organisms did not retain BSH within their cells. BSH activity was assessed using ninhydrin assay that detected free taurine liberated from sodium taurocholate. However, ninhydrin can yield false–positive results owing to its interaction with other free amino acids. To subjugate this limitation, the study introduced a novel and sensitive HPTLC–MS method capable of accurately detecting taurine. By comprehending fermentation dynamics and selecting appropriate conditions, BSH production increased 2.1–fold in both organisms. These findings illuminate critical insights, offering a pathway for novel strategies to enhance the BSH–producing capabilities of these LAB strains.

Characterization of indigenous lactobacilli from dairy fermented foods of Haryana as potential probiotics utilizing multiple attribute decision-making approach

The interest in region-specific ethnic fermented foods and their functional microbiota is rising. The demands for functional foods are continuously rising, so research is going on to develop nutritious food with many beneficial attributes and low safety concerns. The present study was designed to isolate and characterize lactobacilli probiotic candidates from locally resourced fermented foods (dahi, lassi, and raabadi) to make ready-to-eat fermented functional products later. Cultures were isolated from 82 fermented food samples collected from different villages. The initial experiments of gram staining, catalase test, and carbohydrate fermentation were assessed for the morphology, purity, and primary characterization on the genus level, which was verified through molecular characterization using PCR. Seven lactobacilli strains (no. MS001-MS007) were then assessed for safety, probiotic candidacy, phytase degradation, and biofilm forming abilities. All seven bacterial cultures showed no hemolytic activity and antibiotic sensitivity against more than 14 antibiotics out of 20. All seven lactobacilli isolates were able to tolerate pH 3.0, 0.3% bile 0.5% pancreatin, lysozyme (100 mg/L to 300 mg/L) and also shown possessed phytase degradation ability. All the cultures showed antioxidative potential and biofilm formation ability. Culture MS007 showed considerably higher bile salt hydrolase activity among all the isolates, whereas MS005 possessed excellent phytate degradation ability among others. Bacterial strains were identified using 16S rRNA gene sequencing. Moreover, the order of preference of isolates was calculated using the multidimensional Technique for Order of Preference by Similarity to Ideal Solution (TOPSIS) based on probiotic and other functional properties. The most promising attributes showing cultures were recognised as Limosilactobacillus fermentum MS005 and Lactiplantibacillus plantarum MS007, which could be further used for functional food product development.Graphical

Influence of gut microbiome composition on effect variations of anti-cholesterol treatment among individual mice

This study investigated if the variation in the effect of anti-cholesterol (AC) treatment on individual mice are related to gut microbiome composition. The bile salt hydrolase (BSH) activity of 23 commercial fermented milk products was examined to select a fermented milk product for AC treatment. Mice were fed to different diets for 6 weeks: high-fat (660 % of total calories from fat; D1), high-dietary fibre (20 % cellulose; D2), and low-fat (17.2 % of total calories from fat; D3) diets to change their gut microbiomes. Subsequently, faecal microbiome was transplanted (FMT) into mice treated with high cholesterol diet contained 2 % cholesterol, followed by AC or non-AC (sterile tap water, STW) treatments. Control groups with normal (NC) and high-cholesterol diets (PC) were prepared for both AC and STW treatment. All experimental groups were subjected to serum and liver cholesterol, cholesterol metabolism-related (CMR) gene expression, and intestinal microbiome analyses. D3-FMT mice showed the most significant enhancements in cholesterol ratio and decreased hepatic cholesterol levels with AC treatment. Moreover, upregulation of the Cyp7a1 gene expression was observed in this group. Furthermore, the intestinal microbiome analysis indicated higher abundances of BSH-producing Eubacterium, Bifidobacterium, and Parabacteroides in the D3-FMT + AC group compare to others, potentially contributing to increased bile acid synthesis.

Bile Salt Hydrolase Activity of Cholesterol-Lowering Probiotic Lactic Acid Bacteria Isolated from Indigenous Fermented Foods

Bile Salt Hydrolase Activity of Cholesterol-Lowering Probiotic Lactic Acid Bacteria Isolated from Indigenous Fermented Foods

Interplay between Bile Acids and Intestinal Microbiota: Regulatory Mechanisms and Therapeutic Potential for Infections.

Bile acids (BAs) play a crucial role in the human body's defense against infections caused by bacteria, fungi, and viruses. BAs counteract infections not only through interactions with intestinal bacteria exhibiting bile salt hydrolase (BSH) activity but they also directly combat infections. Building upon our research group's previous discoveries highlighting the role of BAs in combating infections, we have initiated an in-depth investigation into the interactions between BAs and intestinal microbiota. Leveraging the existing literature, we offer a comprehensive analysis of the relationships between BAs and 16 key microbiota. This investigation encompasses bacteria (e.g., Clostridioides difficile (C. difficile), Staphylococcus aureus (S. aureus), Escherichia coli, Enterococcus, Pseudomonas aeruginosa, Mycobacterium tuberculosis (M. tuberculosis), Bacteroides, Clostridium scindens (C. scindens), Streptococcus thermophilus, Clostridium butyricum (C. butyricum), and lactic acid bacteria), fungi (e.g., Candida albicans (C. albicans) and Saccharomyces boulardii), and viruses (e.g., coronavirus SARS-CoV-2, influenza virus, and norovirus). Our research found that Bacteroides, C. scindens, Streptococcus thermophilus, Saccharomyces boulardii, C. butyricum, and lactic acid bacteria can regulate the metabolism and function of BSHs and 7α-dehydroxylase. BSHs and 7α-dehydroxylase play crucial roles in the conversion of primary bile acid (PBA) to secondary bile acid (SBA). It is important to note that PBAs generally promote infections, while SBAs often exhibit distinct anti-infection roles. In the antimicrobial action of BAs, SBAs demonstrate antagonistic properties against a wide range of microbiota, with the exception of norovirus. Given the intricate interplay between BAs and intestinal microbiota, and their regulatory effects on infections, we assert that BAs hold significant potential as a novel approach for preventing and treating microbial infections.

Gut microbiota, dietary taurine, and fiber shift taurine homeostasis in adipose tissue of calorie-restricted mice to impact fat loss

Previously, we demonstrated that caloric restriction (CR) stimulates the synthesis, conjugation, secretion, and deconjugation of taurine and bile acids in the intestine, as well as their reuptake. Given taurine's potent antiobesogenic properties, this study aimed to assess whether the CR-induced shift in taurine homeostasis contributes to adipose tissue loss. To verify that, male C57Bl/6 mice were subjected to 20% CR or ad libitum feeding, with variations in cage bedding and gut microbiota conditions. Additional groups received taurine supplementation or were fed a low-taurine diet (LTD). The results showed that in CR animals, taurine derived from the intestine was preferentially trafficked to epididymal white adipose tissue (eWAT) over other tested organs. Besides increased levels of taurine transporter TauT, gene expression of Cysteine dioxygenase (Cdo) involved in taurine synthesis was upregulated in CR eWAT. Taurine concentration in adipocytes was inversely correlated with fat pad weight of CR mice. Different types of cage bedding did not impact eWAT taurine levels; however, the lack of bedding and consumption of a diet high in soluble fiber did. Depleting gut microbiota with antibiotics or inhibiting bile salt hydrolase (BSH) activity reduced WAT taurine concentration in CR mice. Taurine supplementation increased taurine levels in WAT and brown adipose tissue (BAT), promoting fat loss in CR animals. LTD consumption blunted WAT loss in CR animals, with negligible impact on BAT. This study provides multiple insights into taurine's role in CR-triggered fat loss and describes a novel communication path between the liver, gut, microbiota, and WAT, with taurine acting as a messenger.

Changes in the Bile Acid Pool and Timing of Female Puberty: Potential Novel Role of Hypothalamic TGR5.

The regulation of pubertal timing and reproductive axis maturation is influenced by a myriad of physiologic and environmental inputs yet remains incompletely understood. To contrast differences in bile acid isoform profiles across defined stages of reproductive maturity in humans and a rat model of puberty and to characterize the role of bile acid signaling via hypothalamic expression of bile acid receptor populations in the rodent model. Secondary analysis and pilot studies of clinical cohorts, rodent models, ex vivo analyses of rodent hypothalamic tissues. Bile acid concentrations is the main outcome measure. Lower circulatory conjugated:deconjugated bile acid concentrations and higher total secondary bile acids were observed in postmenarcheal vs pre-/early pubertal adolescents, with similar shifts observed in infantile (postnatal day [PN]14) vs early juvenile (PN21) rats alongside increased tgr5 receptor mRNA expression within the mediobasal hypothalamus of female rats. 16S rRNA gene sequencing of the rodent gut microbiome across postnatal life revealed changes in the gut microbial composition predicted to have bile salt hydrolase activity, which was observed in parallel with the increased deconjugated and increased concentrations of secondary bile acids. We show that TGR5-stimulated GnRH release from hypothalamic explants is mediated through kisspeptin receptors and that early overexpression of human-TGR5 within the arcuate nucleus accelerates pubertal onset in female rats. Bile acid isoform shifts along stages of reproductive maturation are conserved across rodents and humans, with preclinical models providing mechanistic insight for the neuroendocrine-hepatic-gut microbiome axis as a potential moderator of pubertal timing in females.

Safety evaluation of Bifidobacterium animalis subsp. lactis BLa80 under in vitro, and in vivo conditions

Bifidobacterium animalis subsp. lactis BLa80 is a new probiotic strain with extensive applications in food products both domestically and internationally. Given the rising consumption of this probiotic, its safety assessment is increasingly crucial in the food industry. This study evaluates the safety of strain BLa80 using a combination of in vitro and in vivo assays along with genomic analysis. Methods included exposing the strain to artificial gastric and intestinal fluids, as well as a medium containing bile salts, to stimulate human digestive conditions. The strain showed high tolerance to gastric fluid at pH of 2.5 and to 0.3 % bile salts. It maintained a 99.92 % survival rate in intestinal fluid. Additional tests assessed hemolytic activity, antibiotic susceptibility (revealing sensitivity to 7 antibiotics), and biogenic amine production using HPLC-ELSD, confirming the absence of histamine, and other harmful amines. Bile salt hydrolase activity was demonstrated qualitatively, and metabolic byproducts were quantitatively analyzed using a D-/l-lactic acid assay kit, showing that BLa80 produces 1.48 mg/mL of l-lactic acid and no harmful d-lactic acid. Genomic analysis confirmed the absence of virulence or pathogenicity genes, and a 90-day oral toxicity study in rats confirmed no toxic effects at various doses. Overall, these findings support the safety classification of the strain BLa80.

Unveiling the Impact of Lactic Acid Bacteria on Blood Lipid Regulation for Cardiovascular Health

Lactic acid bacteria (LAB) are a group of microorganisms which are beneficial and well-characterized with respect to the flavor and texture of food products via fermentation. The accumulated literature has suggested that dietary intake of fermented foods rich in LAB is related to different health-promoting benefits; however, in recent years, emerging evidence suggests a contribution of LAB to blood lipid regulation and cardiovascular health via certain mechanisms. Different potential mechanisms for the lipid regulatory effects of LAB may include the interaction of hydroxymethylglutaryl-CoA (HMG-CoA) reductase and bile salt hydrolase activity and bile salt metabolism; gut microbiome modulation; and regulation of mRNA expression of genes related to fat metabolism in animal models and human studies. This review comprehensively aims to answer whether/how LAB influence blood lipids in both animal models and human studies while also uncovering the underlying mechanisms linking LAB to lipid metabolism.

Ketogenic diet-induced bile acids protect against obesity through reduced calorie absorption.

The low-carbohydrate ketogenic diet (KD) has long been practiced for weight loss, but the underlying mechanisms remain elusive. Gut microbiota and metabolites have been suggested to mediate the metabolic changes caused by KD consumption, although the particular gut microbes or metabolites involved are unclear. Here, we show that KD consumption enhances serum levels of taurodeoxycholic acid (TDCA) and tauroursodeoxycholic acid (TUDCA) in mice to decrease body weight and fasting glucose levels. Mechanistically, KD feeding decreases the abundance of a bile salt hydrolase (BSH)-coding gut bacterium, Lactobacillus murinus ASF361. The reduction of L. murinus ASF361 or inhibition of BSH activity increases the circulating levels of TDCA and TUDCA, thereby reducing energy absorption by inhibiting intestinal carbonic anhydrase 1 expression, which leads to weight loss. TDCA and TUDCA treatments have been found to protect against obesity and its complications in multiple mouse models. Additionally, the associations among the abovementioned bile acids, microbial BSH and metabolic traits were consistently observed both in an observational study of healthy human participants (n = 416) and in a low-carbohydrate KD interventional study of participants who were either overweight or with obesity (n = 25). In summary, we uncover a unique host-gut microbiota metabolic interaction mechanism for KD consumption to decrease body weight and fasting glucose levels. Our findings support TDCA and TUDCA as two promising drug candidates for obesity and its complications in addition to a KD.

A gut microbiota-independent mechanism shapes the bile acid pool in mice with MASH

Background & AimsAn imbalance between primary and secondary bile acids contributes to the development of metabolic dysfunction-associated steatohepatitis (MASH). The precise mechanisms underlying changes in the bile acid pool in MASH remain to be identified. As gut bacteria convert primary bile acids to secondary bile acids, we investigated the contribution of the gut microbiota and its metabolizing activities to bile acid alterations in MASH. MethodsTo disentangle the influence of MASH from environmental and dietary factors, high fat diet fed foz/foz mice were compared to their high fat diet fed wildtype littermates. We developed functional assays (stable isotope labeling and in vitro experiments) to extend the analyses beyond a mere study of gut microbiota composition (16S rRNA gene sequencing). Key findings were confirmed in C57BL/6J mice were fed a western and high fructose diet, as an independent mouse model of MASH. ResultsWhile mice with MASH exhibited lower levels of secondary 7α-dehydroxylated bile acids (3.5-fold lower, p=0.0008), the gut microbial composition was similar in mice with and without MASH. Similar gut microbial bile salt hydrolase and 7α-dehydroxylating activities could not explain the low levels of secondary 7α-dehydroxylated bile acids. Furthermore, the 7α-dehydroxylating activity was unaffected by Clostridium scindens administration in mice with a non-standardized gut microbiota.By exploring alternative mechanisms, we identified an increased bile acid 7α-rehydroxylation mediated by liver CYP2A12 and CYP2A22 enzymes (4.0-fold higher, p<0.0001), that reduces secondary 7α-dehydroxylated bile acid levels in MASH. ConclusionsThis study reveals a gut microbiota-independent mechanism that alters the level of secondary bile acids and contributes to the development of MASH in mice. Impact and implicationsWhile changes in bile acid levels are implicated in the development of metabolic dysfunction-associated steatohepatitis (MASH), the precise mechanisms underpinning these alterations remain elusive. In this study, we investigated the mechanisms responsible for the changes in bile acid levels in mouse models of MASH. Our results support that neither the composition nor the metabolic activity of the gut microbiota can account for the alterations in the bile acid pool. Instead, we identified hepatic 7α-rehydroxylation of secondary bile acids as a gut microbiota-independent factor contributing to the reduced levels of secondary bile acids in mice with MASH. Further investigation is warranted to understand bile acid metabolism and its physiological implications in clinical MASH. Nonetheless, our findings hold promise for exploring novel therapeutic interventions for MASH.

Lactobacillus rhamnosus GG ameliorates triptolide-induced liver injury through modulation of the bile acid-FXR axis

Triptolide (TP) is the principal bioactive compound of Tripterygium wilfordii with significant anti-tumor, anti-inflammatory and immunosuppressive activities. However, its severe hepatotoxicity greatly limits its clinical use. The underlying mechanism of TP-induced liver damage is still poorly understood. Here, we estimate the role of the gut microbiota in TP hepatotoxicity and investigate the bile acid metabolism mechanisms involved. The results of the antibiotic cocktail (ABX) and fecal microbiota transplantation (FMT) experiment demonstrate the involvement of intestinal flora in TP hepatotoxicity. Moreover, TP treatment significantly perturbed gut microbial composition and reduced the relative abundances of Lactobacillus rhamnosus GG (LGG). Supplementation with LGG reversed TP-induced hepatotoxicity by increasing bile salt hydrolase (BSH) activity and reducing the increased conjugated bile acids (BA). LGG supplementation upregulates hepatic FXR expression and inhibits NLRP3 inflammasome activation in TP-treated mice. In summary, this study found that gut microbiota is involved in TP hepatotoxicity. LGG supplementation protects mice against TP-induced liver damage. The underlying mechanism was associated with the gut microbiota-BA-FXR axis. Therefore, LGG holds the potential to prevent and treat TP hepatotoxicity in the clinic.

Monitoring probiotic properties and safety evaluation of antilisterial Enterococcus faecium strains with cholesterol-lowering potential from raw Cow's milk

Enterococci are an important member of lactic acid bacteria due to their presence in fermented foods, tolerance to low pH and bile salts, bile salt hydrolase (BSH) activity, reducing cholesterol, and producing bacteriocin. However, probiotic and safety issues of enterococci are still debated. It is important to investigate probiotic properties and safety evaluation of these genus because they are intentionally or unintentionally present in foods. In this study, probiotic properties (tolerance to bile salts and simulated gastric juice, antimicrobial and BSH activity, cholesterol lowering potential, auto/co-aggregation, and cell surface hydrophobicity) and safety evaluation (antibiotic resistance, virulence factor, haemolysis and DNase activity, Galleria mellonella infection model, gelatinase activity, and biofilm formation) of Enterococcus faecium (E. faecium) BH04, BH12, BH84, and BH99 isolated raw cow's milk were determined. All strains had high tolerance to bile salts (47.70%–103.84%) and simulated gastric juice (76.80%–82.05%), and antimicrobial activity on L. monocytogenes ATCC 7644 by protein-like substances such as bacteriocin. The strains had strong BSH property and reduced cholesterol by 69.47%–95.00%. The strains were resistant only to gentamicin, kanamycin, and streptomycin and did not have haemolysis and DNase activity. Only gelE and esp genes were detected in some strains. According to the results, it is thought that the E. faecium strains could be possible probiotic candidates. Moreover, this study is the first report on Enterococcus faecium strains isolated from raw cow's milk, which tolerate to bile salt and simulated gastric juice, have BSH potential and antimicrobial activity by protein-like substances, and reduce cholesterol level.

Recombinant Bile Salt Hydrolase Enhances the Inhibition Efficiency of Taurodeoxycholic Acid against Clostridium perfringens Virulence.

Clostridium perfringens is the main pathogen of chicken necrotic enteritis (NE) causing huge economic losses in the poultry industry. Although dietary secondary bile acid deoxycholic acid (DCA) reduced chicken NE, the accumulation of conjugated tauro-DCA (TDCA) raised concerns regarding DCA efficacy. In this study, we aimed to deconjugate TDCA by bile salt hydrolase (BSH) to increase DCA efficacy against the NE pathogen C. perfringens. Assays were conducted to evaluate the inhibition of C. perfringens growth, hydrogen sulfide (H2S) production, and virulence gene expression by TDCA and DCA. BSH activity and sequence alignment were conducted to select the bsh gene for cloning. The bsh gene from Bifidobacterium longum was PCR-amplified and cloned into plasmids pET-28a (pET-BSH) and pDR111 (pDR-BSH) for expressing the BSH protein in E. coli BL21 and Bacillus subtilis 168 (B-sub-BSH), respectively. His-tag-purified BSH from BL21 cells was evaluated by SDS-PAGE, Coomassie blue staining, and a Western blot (WB) assays. Secretory BSH from B. subtilis was analyzed by a Dot-Blot. B-sub-BSH was evaluated for the inhibition of C. perfringens growth. C. perfringens growth reached 7.8 log10 CFU/mL after 24 h culture. C. perfringens growth was at 8 vs. 7.4, 7.8 vs. 2.6 and 6 vs. 0 log10 CFU/mL in 0.2, 0.5, and 1 mM TDCA vs. DCA, respectively. Compared to TDCA, DCA reduced C. perfringens H2S production and the virulence gene expression of asrA1, netB, colA, and virT. BSH activity was observed in Lactobacillus johnsonii and B. longum under anaerobe but not L. johnsonii under 10% CO2 air. After the sequence alignment of bsh from ten bacteria, bsh from B. longum was selected, cloned into pET-BSH, and sequenced at 951 bp. After pET-BSH was transformed in BL21, BSH expression was assessed around 35 kDa using Coomassie staining and verified for His-tag using WB. After the subcloned bsh and amylase signal peptide sequence was inserted into pDR-BSH, B. subtilis was transformed and named B-sub-BSH. The transformation was evaluated using PCR with B. subtilis around 3 kb and B-sub-BSH around 5 kb. Secretory BSH expressed from B-sub-BSH was determined for His-tag using Dot-Blot. Importantly, C. perfringens growth was reduced greater than 59% log10 CFU/mL in the B-sub-BSH media precultured with 1 vs. 0 mM TDCA. In conclusion, TDCA was less potent than DCA against C. perfringens virulence, and recombinant secretory BSH from B-sub-BSH reduced C. perfringens growth, suggesting a new potential intervention against the pathogen-induced chicken NE.

Antibiotic-Induced Gut Microbial Dysbiosis Reduces the Growth of Weaning Rats via FXR-Mediated Hepatic IGF-2 Inhibition.

The gut microbiota plays a crucial role in postnatal growth, particularly in modulating the development of animals during their growth phase. In this study, we investigated the effects of antibiotic-induced dysbiosis of the gut microbiota on the growth of weaning rats by administering a non-absorbable antibiotic cocktail (ABX) in water for 4 weeks. ABX treatment significantly reduced body weight and feed intake in rats. Concurrently, ABX treatment decreased microbial abundance and diversity in rat ceca, predominantly suppressing microbes associated with bile salt hydrolase (BSH) activity. Furthermore, decreased appetite may be attributed to elevated levels of glucagon-like peptide-1 (GLP-1) in the serum, along with reduced neuropeptide Y (NPY) and increased cocaine and amphetamine-regulated transcript (CART) in the hypothalamus at the mRNA level. Importantly, concentrations of insulin-like growth factor 1 (IGF-1) and insulin-like growth factor 2 (IGF-2) were decreased in the serum and liver of antibiotic-treated rats. These alterations were associated with significant down-regulation of IGF-2 mRNA in the liver and significantly decreased farnesoid X receptor (FXR) protein expression and binding to the IGF-2 promoter. These results indicate that antibiotic-induced gut microbial dysbiosis not only impacts bile acid metabolism but also diminishes rat growth through the FXR-mediated IGF-2 pathway.