UPLC/Q-TOF-MS-based metabolomics and molecular docking analysis of Bifidobacterium adolescentis exposure to levofloxacin.

Antibiotic-associated diarrhea is a common adverse reaction caused by the widespread use of antibiotics. The decrease in probiotics is one of the reasons why antibiotics cause drug-induced diarrhea. However, few studies have addressed the intrinsic mechanism of antibiotics inhibiting probiotics. To investigate the underlying mechanism of levofloxacin against Bifidobacterium adolescentis, we used a metabolomics mass spectrometry-based approach and molecular docking analysis for a levofloxacin-induced B. adolescentis injury model. The results showed that levofloxacin reduced the survival rate of B. adolescentis and decreased the number of B. adolescentis. The untargeted metabolomics analysis identified 27 potential biomarkers, and many of these metabolites are involved in energy metabolism, amino acid metabolism and the lipid metabolism pathway. Molecular docking showed that levofloxacin can bind with aminoacyl-tRNA synthetase and lactic acid dehydrogenase. This result provides a novel insight into the mechanism of the adverse reactions of levofloxacin.

Ecology- and genome-based identification of the Bifidobacterium adolescentis prototype of the healthy human gut microbiota.

Bifidobacteria are among the first microbial colonizers of the human gut, being frequently associated with human health-promoting activities. In the current study, an in silico methodology based on an ecological and phylogenomic-driven approach allowed the selection of a Bifidobacterium adolescentis prototype strain, i.e., B. adolescentis PRL2023, which best represents the overall genetic content and functional features of the B. adolescentis taxon. Such features were confirmed by in vitro experiments aimed at evaluating the ability of this strain to survive in the gastrointestinal tract of the host and its ability to interact with human intestinal cells and other microbial gut commensals. In this context, co-cultivation of B. adolescentis PRL2023 and several gut commensals revealed various microbe-microbe interactions and indicated co-metabolism of particular plant-derived glycans, such as xylan.IMPORTANCEThe use of appropriate bacterial strains in experimental research becomes imperative in order to investigate bacterial behavior while mimicking the natural environment. In the current study, through in silico and in vitro methodologies, we were able to identify the most representative strain of the Bifidobacterium adolescentis species. The ability of this strain, B. adolescentis PRL2023, to cope with the environmental challenges imposed by the gastrointestinal tract, together with its ability to switch its carbohydrate metabolism to compete with other gut microorganisms, makes it an ideal choice as a B. adolescentis prototype and a member of the healthy microbiota of adults. This strain possesses a genetic blueprint appropriate for its exploitation as a candidate for next-generation probiotics.

Mouse intestinal microbiome modulation by oral administration of a GABA-producing Bifidobacterium adolescentis strain.

IMPORTANCE: The gut microbiome-brain communication signaling has emerged in recent years as a novel target for intervention with the potential to ameliorate some conditions associated with the central nervous system. Hence, probiotics with capacity to produce neurotransmitters, for instance, have come up as appealing alternatives to treat disorders associated with disbalanced neurotransmitters. Herein, we further deep into the effects of administering a gamma-aminobutyric acid (GABA)-producing Bifidobacterium strain, previously demonstrated to contribute to reduce serum glutamate levels, in the gut microbiome composition and metabolic activity in a mouse model. Our results demonstrate that the GABA-producing strain administration results in a specific pattern of gut microbiota modulation, different from the one observed in animals receiving non-GABA-producing strains. This opens new avenues to delineate the specific mechanisms by which IPLA60004 administration contributes to reducing serum glutamate levels and to ascertain whether this effect could exert health benefits in patients of diseases associated with high-glutamate serum concentrations.

Bifidobacterium adolescentis induces Decorin+ macrophages via TLR2 to suppress colorectal carcinogenesis.

BACKGROUND: The interplay between gut microbiota and tumor microenvironment (TME) in the pathogenesis of colorectal cancer (CRC) is largely unknown. Here, we elucidated the functional role of B. adolescentis and its possible mechanism on the manipulation of Decorin+ macrophages in colorectal cancer. METHODS: The relative abundance of B. adolescentis in tumor or para-tumor tissue of CRC patients was analyzed. The role of B. adolescentis was explored in the CRC animal models. The single cell-RNA sequencing (scRNA-seq) was used to investigate the myeloid cells subsets in TME. The expression level of TLR2/YAP axis and its downstream Decorin in macrophages were tested by Western blot and qRT-PCR. Knockdown of Decorin in Raw264.7 was performed to investigate the effect of Decorin+ macrophages on subcutaneous tumor formation. Multi-immunofluorescence assay examined the number of Decorin+ macrophages on the CRC tissue. RESULTS: We found that the abundance of B. adolescentis was significantly reduced in tumor tissue of CRC patients. Supplementation with B. adolescentis suppressed AOM/DSS-induced tumorigenesis in mice. ScRNA-seq and animal experiment revealed that B. adolescentis increased Decorin+ macrophages. Mechanically, Decorin was activated by TLR2/YAP axis in macrophages. The abundance of B. adolescentis was correlated with the number of Decorin+ macrophages and the expression level of TLR2 in tumor tissue of CRC patients. CONCLUSIONS: These results highlight that B. adolescentis induced Decorin+ macrophages and provide a novel therapeutic target for probiotic-based modulation of immune microenvironment in CRC.

Orally Administered Bifidobacterium adolescentis Diminishes Serum Glutamate Concentration in Mice.

Several studies have described the contribution of glutamate-transforming microbiota to the development of chronic ailments. For instance, the blood concentration of glutamate is higher in some patients with fibromyalgia, chronic fatigue, and pain. Taking advantage of a naturally occurring strain of Bifidobacterium that is able to transform glutamate in γ-aminobutyric caid (GABA), B. adolescentis IPLA60004, we designed a placebo-controlled intervention to test if the presence of this GABA-producing bifidobacteria in mice was able to impact the concentration of glutamate in the blood in comparison with the administration of other strain of the same species lacking the genes of the glutamate decarboxylase (gad) cluster. Animals were fed every day with 8 log CFU of bacteria in a sterilized milk vehicle for 14 days. Samples from feces and blood were collected during this period, and afterwards animals were sacrificed, tissues were taken from different organs, and the levels of different metabolites were analyzed by ultrahigh-performance liquid chromatography coupled to mass spectrometry. The results showed that both bacterial strains orally administered survived in the fecal content, and animals fed B. adolescentis IPLA60004 showed a significant reduction of their glutamate serum concentration, while a nonsignificant decrease was observed for animals fed a reference strain, B. adolescentis LGM10502. The variations observed in GABA were influenced by the gender of the animals, and no significant changes were observed in different tissues of the brain. These results suggest that orally administered GABA-producing probiotics could reduce the glutamate concentration in blood, opening a case for a clinical trial study in chronic disease patients. IMPORTANCE This work presents the results of a trial using mice as a model that were fed with a bacterial strain of the species B. adolescentis, which possesses different active genes capable of degrading glutamate and converting it into GABA. Indeed, the bacterium is able to survive the passage through the gastric tract and, more importantly, the animals reduce over time the concentration of glutamate in their blood. The importance of this result lies in the fact that several chronic ailments, such as fibromyalgia, are characterized by an increase in glutamate. Our results indicate that an oral diet with this probiotic-type bacteria could reduce the concentration of glutamate and, therefore, reduce the symptoms associated with the excess of this neurotransmitter.

Production of GABA in milk fermented by Bifidobacterium adolescentis strains selected on the bases of their technological and gastrointestinal performance.

There is an increasing interest in producing foods enriched in gamma-aminobutyric acid (GABA), due to their purported health promoting attributes. GABA is the main inhibitor neurotransmitter of the central nervous system, and several microbial species are capable to produce it through decarboxylation of glutamate. Among them, several lactic acid bacteria species have been previously investigated as an appealing alternative to produce GABA enriched foods via microbial fermentation. In this work we report for the first time an investigation into the possibility of utilizing high GABA-producing Bifidobacterium adolescentis strains as a mean to produce fermented probiotic milks naturally enriched in GABA. To this end, in silico and in vitro analyses were conducted in a collection of GABA-producing B. adolescentis strains, with the main goal to scrutinize their metabolic and safety traits, including antibiotic resistance patterns, as well as their technological robustness and performance to survive a simulated gastrointestinal passage. One of the strains, IPLA60004, exhibited better survival to lyophilization and cold storage (for up to 4 weeks at 4 °C), as well as survival to gastrointestinal passage, as compared to the other strains under investigation. Besides, the elaboration of milk drinks fermented with this strain, yielded products with the highest GABA concentration and viable bifidobacterial cell counts, achieving conversion rates of the precursor, monosodium glutamate (GMS), up to 70 %. To our knowledge, this is the first report on the elaboration of GABA enriched milks through fermentation with B. adolescentis.

Insights into ß-manno-oligosaccharide uptake and metabolism in Bifidobacterium adolescentis DSMZ 20083 from whole-genome microarray analysis.

Metabolism of non-digestible dietary glycans directly influences the structure and composition of human gut microbiota and, in turn, the host health. ß-Mannans form an integral component of the modern diet as naturally occurring dietary fibre or additives in processed foods. In the present study, in vitro fermentation and TLC studies were used to determine the ability of adult-associated Bifidobacterium adolescentis DSMZ 20083 to utilise ß-manno-oligosaccharides from guar gum, locust bean gum, konjac root, and copra meal generated using GH26 endo-ß-mannanase (ManB-1601). Further, to gain insights into the underlying molecular mechanism, a whole-genome microarray analysis, RT-qPCR, and molecular docking studies were employed to reconstruct the copra meal ß-manno-oligosaccharides (CM-ß-MOS) utilisation pathway in B. adolescentis DSMZ 20083. B. adolescentis DSMZ 20083 grew appreciably (O.D600 nm up to 0.8) on all tested ß-manno-oligosaccharides but maximally on CM-ß-MOS. CM-ß-MOS having DP2-3 were found to deplete from the fermentation media. Whole-genome transcriptome analysis, RT-qPCR, and molecular docking studies suggested that in B. adolescentis DSMZ 20083, ABC & MFS transporters are possibly involved in the uptake of DP ≥ 2 and DP ≥ 3 linear CM-ß-MOS, respectively, while GH1 ß-glucosidase, and GH32 ß-fructofuranosidase possibly cleave linear CM-ß-MOS into monosaccharides. Sugar absorption and utilisation pathways; Bifid shunt, ABC transport system, pyruvate metabolism, glycolysis/gluconeogenesis, pentose, and glucouronate inter-conversions were also found up-regulated following the growth on CM-ß-MOS. This is the first study reporting on possible molecular determinants used by B. adolescentis DSMZ 20083 to utilise ß-manno-oligosaccharides. Our studies can prove resourceful to food and nutraceutical industries, aiming at precision microbiome modulation using ß-manno-oligosaccharides.

Bifidobacterium adolescentis Is Effective in Relieving Type 2 Diabetes and May Be Related to Its Dominant Core Genome and Gut Microbiota Modulation Capacity.

The prevalence of diabetes mellitus is increasing globally. Probiotics have been shown to be an effective intervention for diabetes. This study focused on the relieving effects and possible mechanisms of 16 strains of two dominant Bifidobacterium species (B. bifidum and B. adolescentis, which exist in the human gut at different life stages) on type 2 diabetes (T2D). The results indicated that more B. adolescentis strains appeared to be superior in alleviating T2D symptoms than B. bifidum strains. This effect was closely related to the ability of B. adolescentis to restore the homeostasis of the gut microbiota, increase the abundance of short-chain fatty acid-producing flora, and alleviate inflammation in mice with T2D. In addition, compared with B. bifidum, B. adolescentis had a higher number of core genes, and these genes were more evolutionarily stable, including unique environmental tolerance, carbon and nitrogen utilization genes, and a blood sugar regulation gene, glgP. This may be one of the reasons why B. adolescentis is more likely to colonize in the adult gut and show a superior ability to relieve T2D. This study provides insights into future studies aimed at investigating probiotics for the treatment of metabolic diseases.

Cell factory for γ-aminobutyric acid (GABA) production using Bifidobacterium adolescentis.

BACKGROUND: Bifidobacteria are gram-positive, probiotic, and generally regarded as safe bacteria. Techniques such as transformation, gene knockout, and heterologous gene expression have been established for Bifidobacterium, indicating that this bacterium can be used as a cell factory platform. However, there are limited previous reports in this field, likely because of factors such as the highly anaerobic nature of this bacterium. Bifidobacterium adolescentis is among the most oxygen-sensitive Bifidobacterium species. It shows strain-specific gamma-aminobutyric acid (GABA) production. GABA is a potent bioactive compound with numerous physiological and psychological functions. In this study, we investigated whether B. adolesentis could be used for mass production of GABA. RESULTS: The B. adolescentis 4-2 strain isolated from a healthy adult human produced approximately 14 mM GABA. It carried gadB and gadC, which encode glutamate decarboxylase and glutamate GABA antiporter, respectively. We constructed pKKT427::Pori-gadBC and pKKT427::Pgap-gadBC plasmids carrying gadBC driven by the original gadB (ori) and gap promoters, respectively. Recombinants of Bifidobacterium were then constructed. Two recombinants with high production abilities, monitored by two different promoters, were investigated. GABA production was improved by adjusting the fermentation parameters, including the substrate concentration, initial culture pH, and co-factor supplementation, using response surface methodology. The optimum initial cultivation pH varied when the promoter region was changed. The ori promoter was induced under acidic conditions (pH 5.2:4.4), whereas the constitutive gap promoter showed enhanced GABA production at pH 6.0. Fed-batch fermentation was used to validate the optimum fermentation parameters, in which approximately 415 mM GABA was produced. The conversion ratio of glutamate to GABA was 92-100%. CONCLUSION: We report high GABA production in recombinant B. adolescentis. This study provides a foundation for using Bifidobacterium as a cell factory platform for industrial production of GABA.

Understanding the Xylooligosaccharides Utilization Mechanism of Lactobacillus brevis and Bifidobacterium adolescentis: Proteins Involved and Their Conformational Stabilities for Effectual Binding.

Xylooligosaccharides having various degrees of polymerization such as xylobiose, xylotriose, and xylotetraose positively affect human health by interacting with gut proteins. The present study aimed to identify proteins present in gut microflora, such as xylosidase, xylulokinase, etc., with the help of retrieved whole-genome annotations and find out the mechanistic interactions of those with the above substrates. The 3D structures of proteins, namely Endo-1,4-beta-xylanase B (XynB) from Lactobacillus brevis and beta-D-xylosidase (Xyl3) from Bifidobacterium adolescentis, were computationally predicted and validated with the help of various bioinformatics tools. Molecular docking studies identified the effectual binding of these proteins to the xylooligosaccharides, and the stabilities of the best-docked complexes were analyzed by molecular dynamic simulation. The present study demonstrated that XynB and Xyl3 showed better effectual binding toward Xylobiose with the binding energies of - 5.96 kcal/mol and - 4.2 kcal/mol, respectively. The interactions were stabilized by several hydrogen bonding having desolvation energy (- 6.59 and - 7.91). The conformational stabilities of the docked complexes were observed in the four selected complexes of XynB-xylotriose, XynB-xylotetraose, Xyl3-xylobiose, and Xyn3-xylotriose by MD simulations. This study showed that the interactions of these four complexes are stable, which means they have complex metabolic activities among each other. Extending these studies of understanding, the interaction between specific probiotics enzymes and their ligands can explore the detailed design of synbiotics in the future.

Enzymatic analysis of truncation mutants of a type II pullulanase from Bifidobacterium adolescentis P2P3, a resistant starch-degrading gut bacterium.

A putative type II pullulanase gene, pulP, was identified in Bifidobacterium adolescentis P2P3. PulP possesses an α-amylase domain at the N-terminus and a pullulanase type I domain at the C-terminus, as well as three carbohydrate-binding modules (one CBM25 and two CBM41s) between them. The native PulP and four truncated mutant recombinant proteins (PulPΔCΔP, PulPΔP, PulPΔAΔC, and PulPΔA), in which each of the two catalytic domains and/or the CBMs were deleted, were produced in Escherichia coli and their specific properties were characterized. The removal of either catalytic domain abolished the corresponding catalytic activity of the wild-type enzyme. Deletion of the C-terminal domain resulted in a drastic decrease in the optimal temperature and thermostability, indicating that the pullulanase domain might be related to the temperature dependency of the enzyme. In addition, the elimination of the CBMs in the mutant proteins led to a loss of binding affinity toward raw substrates as well as the loss of their hydrolysis activities compared to the wild-type enzyme. HPAEC and TLC analyses proved that PulP and its mutants could hydrolyze α-glucans into maltotriose as their main product. These results suggest that PulP may play an important role in α-glucan metabolism in B. adolescentis P2P3.

A gap-filling algorithm for prediction of metabolic interactions in microbial communities.

The study of microbial communities and their interactions has attracted the interest of the scientific community, because of their potential for applications in biotechnology, ecology and medicine. The complexity of interspecies interactions, which are key for the macroscopic behavior of microbial communities, cannot be studied easily experimentally. For this reason, the modeling of microbial communities has begun to leverage the knowledge of established constraint-based methods, which have long been used for studying and analyzing the microbial metabolism of individual species based on genome-scale metabolic reconstructions of microorganisms. A main problem of genome-scale metabolic reconstructions is that they usually contain metabolic gaps due to genome misannotations and unknown enzyme functions. This problem is traditionally solved by using gap-filling algorithms that add biochemical reactions from external databases to the metabolic reconstruction, in order to restore model growth. However, gap-filling algorithms could evolve by taking into account metabolic interactions among species that coexist in microbial communities. In this work, a gap-filling method that resolves metabolic gaps at the community level was developed. The efficacy of the algorithm was tested by analyzing its ability to resolve metabolic gaps on a synthetic community of auxotrophic Escherichia coli strains. Subsequently, the algorithm was applied to resolve metabolic gaps and predict metabolic interactions in a community of Bifidobacterium adolescentis and Faecalibacterium prausnitzii, two species present in the human gut microbiota, and in an experimentally studied community of Dehalobacter and Bacteroidales species of the ACT-3 community. The community gap-filling method can facilitate the improvement of metabolic models and the identification of metabolic interactions that are difficult to identify experimentally in microbial communities.

Enhanced Storage of Anaerobic Bacteria through Polymeric Encapsulation.

Live microbes such as lactobacilli have long been used as probiotic supplements and, more recently, have been explored as live biotherapeutic products with the potential to treat a range of conditions. Among these microbes is a category of anaerobes that possess therapeutic potential while exhibiting unique oxygen sensitivity and thus requiring careful considerations in the formulation and storage processes. Existing microbial formulation development has focused on facultative anaerobes with natural oxygen tolerance; a few strategies have been reported for anaerobes with demonstrated oxygen intolerance, warranting novel approaches toward addressing the challenges for these oxygen-sensitive anaerobes. Here, we develop a polymeric encapsulation system for the formulation and storage of Bifidobacterium adolescentis (B. adolescentis), a model anaerobe that loses viability in aerobic incubation at 37 °C within 1 day. We discover that this strain remains viable under aerobic conditions for 14 days at 4 °C, enabling formulation development such as solution casting and air drying in an aerobic environment. Next, through a systematic selection of polymer encapsulants and excipients, we show that encapsulation with poly(vinyl alcohol) (PVA) acts as an oxygen barrier and facilitates long-term storage of B. adolescentis, which is partially attributed to reduced generation of reactive oxygen species. Lastly, PVA-based formulations can produce oral capsule-loaded films and edible gummy bears, demonstrating its compatibility with both pharmaceutical and food dosage forms.

BdPUL12 depolymerizes ß-mannan-like glycans into mannooligosaccharides and mannose, which serve as carbon sources for Bacteroides dorei and gut probiotics.

Symbiotic bacteria, including members of the Bacteroides genus, are known to digest dietary fibers in the gastrointestinal tract. The metabolism of complex carbohydrates is restricted to a specified subset of species and is likely orchestrated by polysaccharide utilization loci (PULs) in these microorganisms. ß-Mannans are plant cell wall polysaccharides that are commonly found in human nutrients. Here, we report the structural basis of a PUL cluster, BdPUL12, which controls ß-mannan-like glycan catabolism in Bacteroides dorei. Detailed biochemical characterization and targeted gene disruption studies demonstrated that a key glycoside hydrolase, BdP12GH26, performs the initial attack on galactomannan or glucomannan likely via an endo-acting mode, generating mannooligosaccharides and mannose. Importantly, coculture assays showed that the B. dorei promoted the proliferation of Lactobacillus helveticus and Bifidobacterium adolescentis, likely by sharing mannooligosaccharides and mannose with these gut probiotics. Our findings provide new insights into carbohydrate metabolism in gut-inhabiting bacteria and lay a foundation for novel probiotic development.

Three-level hybrid modeling for systematic optimization of biocatalytic synthesis: α-glucosyl glycerol production by enzymatic trans-glycosylation from sucrose.

Mechanism-based kinetic models are rigorous tools to analyze enzymatic reactions, but their extension to actual conditions of the biocatalytic synthesis can be difficult. Here, we demonstrate (mechanistic-empirical) hybrid modeling for systematic optimization of the sucrose phosphorylase-catalyzed glycosylation of glycerol from sucrose, to synthesize the cosmetic ingredient α-glucosyl glycerol (GG). The empirical model part was developed to capture nonspecific effects of high sucrose concentrations (up to 1.5 M) on microscopic steps of the enzymatic trans-glycosylation mechanism. Based on verified predictions of the enzyme performance under initial rate conditions (Level 1), the hybrid model was expanded by microscopic terms of the reverse reaction to account for the full-time course of GG synthesis (Level 2). Lastly (Level 3), the application of the hybrid model for comprehensive window-of-operation analysis and constrained optimization of the GG production (~250 g/L) was demonstrated. Using two candidate sucrose phosphorylases (from Leuconostoc mesenteroides and Bifidobacterium adolescentis), we reveal the hybrid model as a powerful tool of "process decision making" to guide rational selection of the best-suited enzyme catalyst. Our study exemplifies a closing of the gap between enzyme kinetic models considered for mechanistic research and applicable in technologically relevant reaction conditions; and it highlights the important benefit thus realizable for biocatalytic process development.

Bifidobacterium adolescentis Isolated from Different Hosts Modifies the Intestinal Microbiota and Displays Differential Metabolic and Immunomodulatory Properties in Mice Fed a High-Fat Diet.

The incidence of obesity, which is closely associated with the gut microbiota and chronic inflammation, has rapidly increased in the past 40 years. Therefore, the probiotic-based modification of the intestinal microbiota composition has been developed as a strategy for the treatment of obesity. In this study, we selected four Bifidobacterium adolescentis strains isolated from the feces of newborn and elderly humans to investigate whether supplementation with B. adolescentis of various origins could alleviate obesity in mice. Male C57BL/6J mice fed a high-fat diet (HFD, 60% energy as fat) received one of the following 14-week interventions: (i) B. adolescentis N4_N3, (ii) B. adolescentis Z25, (iii) B. adolescentis 17_3, (iv) B. adolescentis 2016_7_2, and (v) phosphate-buffered saline. The metabolic parameters, thermogenesis, and immunity of all treated mice were measured. Cecal and colonic microbial profiles were determined by 16S rRNA gene sequencing. Intestinal concentrations of short-chain fatty acids (SCFAs) were measured by gas chromatography-mass spectrometry (GC-MS). The B. adolescentis strains isolated from the feces of elderly humans (B. adolescentis Z25, 17_3, and 2016_7_2) decreased the body weight or weight gain of mice, whilst the strain isolated from the newborn (B. adolescentis N4_N3) increased the body weight of mice. The B. adolescentis strains isolated from the elderly also increased serum leptin concentrations and induced the expression of thermogenesis- and lipid metabolism-related genes in brown adipose tissue. All the B. adolescentis strains alleviated inflammations in the spleen and brain and modified the cecal and colonic microbiota. Particularly, all strains reversed the HFD-induced depletion of Bifidobacterium and reduced the development of beta-lactam resistance. In addition, the B. adolescentis strains isolated from the elderly increased the relative abundances of potentially beneficial genera, such as Bacteroides, Parabacteroides, and Faecalibaculum. We speculate that such increased abundance of commensal bacteria may have mediated the alleviation of obesity, as B. adolescentis supplementation decreased the intestinal production of SCFAs, thereby reducing energy delivery to the host mice. Our results revealed that certain strains of B. adolescentis can alleviate obesity and modify the gut microbiota of mice. The tested strains of B. adolescentis showed different effects on lipid metabolism and immunity regulation, with these effects related to whether they had been isolated from the feces of newborn or elderly humans. This indicates that B. adolescentis from different sources may have disparate effects on host health possibly due to the transmission of origin-specific functions to the host.

Bifidobacterium adolescentis Alleviates Liver Steatosis and Steatohepatitis by Increasing Fibroblast Growth Factor 21 Sensitivity.

The gut microbiota is a newly identified contributor to the development of non-alcoholic fatty liver disease (NAFLD). Previous studies of Bifidobacterium adolescentis (B. adolescentis), a species of Bifidobacterium that is common in the human intestinal tract, have demonstrated that it can alleviate liver steatosis and steatohepatitis. Fibroblast growth factor 21 (FGF21) has long been considered as a biomarker of NAFLD, and recent studies have shown the protective effect of FGF21 analogs on NAFLD. We wondered whether B. adolescentis treatment would alleviate NAFLD via the interaction with FGF21. To this end, male C57BL/6J mice on a choline-deficient high-fat diet (CDHFD) were treated with drinking water supplemented with B. adolescentis for 8 weeks, followed by the acute administration of recombinant mouse FGF21 protein (rmFGF21) to conduct the FGF21 response test. Consistent with previous studies, B. adolescentis supplementation reversed the CDHFD-induced liver steatosis and steatohepatitis. This was evaluated on the NAFLD activity score (NAS), reduced liver enzymes, and lipid accumulation. Further studies demonstrated that B. adolescentis supplementation preserved the gut barrier, reduced the gut microbiota-derived lipopolysaccharide (LPS), and inhibited the hepatic TLR4/NF-κB pathway. This was accompanied by the elevated expressions of the receptors of FGF21, fibroblast growth factor receptor 1 (FGFR1) and ß-klotho (KLB), in the liver and the decreased expression of FGF21. The results of FGF21 response test showed that B. adolescentis supplementation alleviated the CDHFD-induced FGF21 resistance. In vivo experiments suggested that LPS could suppress the expression of FGF21 and KLB in a dose-dependent manner. Collectively, this study showed that B. adolescentis supplementation could alleviate NAFLD by increasing FGF21 sensitivity.

Bifidobacterium adolescentis as a key member of the human gut microbiota in the production of GABA.

Gamma aminobutyric acid (GABA) is the principal inhibitory neurotransmitter playing a key role in anxiety and depression disorders in mammals. Recent studies revealed that members of the gut microbiota are able to produce GABA modulating the gut-brain axis response. Among members of the human gut microbiota, bifidobacteria are well known to establish many metabolic and physiologic interactions with the host. In this study, we performed genome analyses of more than 1,000 bifidobacterial strains publicly available revealing that Bifidobacterium adolescentis taxon might represent a model GABA producer in human gastrointestinal tract. Moreover, the in silico screening of human/animal metagenomic datasets showed an intriguing association/correlation between B. adolescentis load and mental disorders such as depression and anxiety. Interestingly, in vitro screening of 82 B. adolescentis strains allowed identifying two high GABA producers, i.e. B. adolescentis PRL2019 and B. adolescentis HD17T2H, which were employed in an in vivo trial in rats. Feeding Groningen rats with a supplementation of B. adolescentis strains, confirmed the ability of these microorganisms to stimulate the in vivo production of GABA highlighting their potential implication in gut-brain axis interactions.

Protective effects of Bifidobacterium adolescentis on collagen-induced arthritis in rats depend on timing of administration.

Emerging studies have addressed the role of probiotics in inflammation modulation via modifying gut microbiota. Perturbed gut microbiota is recognized as a pivotal trigger in the pathogenesis of rheumatoid arthritis (RA), and manipulating gut microbiota at the early phase may be helpful to alleviate the disease based on the fact that dysbiosis occurred prior to clinical arthritis. The current study compared the effects of preventive and therapeutic treatment with Bifidobacterium adolescentis on collagen induced arthritis (CIA) in rats. Early B. adolescentis administration before CIA modelling performed better than late B. adolescentis treatment in reducing the clinical symptoms, rebalancing the pro- and anti-inflammatory responses and maintaining the fecal concentration of short chain fatty acids (SCFAs), as well as restoring the intestinal dysbiosis. Preventive B. adolescentis treatment restored the gut microbiota to a normal level while late B. adolescentis fed rats showed clearly different gut microbial profiles. In addition, there were slight discrepancies between early- and late- treatment of B. adolescentis in the production of specific auto-antibodies and tight junction proteins. All those results highlighted that early treatment of probiotics in arthritis might be a better timing for alleviating arthritis.

Mucosal-associated invariant T-Cell (MAIT) activation is altered by chlorpyrifos- and glyphosate-treated commensal gut bacteria.

Mucosal-associated invariant T-cells (MAIT) can react to metabolites of the vitamins riboflavin and folate which are produced by the human gut microbiota. Since several studies showed that the pesticide chlorpyrifos (CPF) and glyphosate (GLP) can impair the gut microbiota, the present study was undertaken to investigate the impact of CPF and GLP treatment on the metabolism of gut microbiota and the resulting bacteria-mediated modulation of MAIT cell activity. Here, Bifidobacterium adolescentis (B. adolescentis), Lactobacillus reuteri (L. reuteri), and Escherichia coli (E. coli) were treated with CPF (50-200 µM) or GLP (75-300 mg/L) and then used in MAIT cell stimulation assays as well as in vitamin and proteome analyses. All three bacteria were nonpathogenic and chosen as representatives of a healthy human gut microflora. The results showed that E. coli activated MAIT cells whereas B. adolescentis and L. reuteri inhibited MAIT cell activation. CPF treatment significantly increased E. coli-mediated MAIT cell activation. Treatment of B. adolescentis and L. reuteri with CPF and GLP weakened the inhibition of MAIT cell activation. Riboflavin and folate production by the test bacteria was influenced by CPF treatment, whereas GLP had only minor effects. Proteomic analysis of CPF-treated E. coli revealed changes in the riboflavin and folate biosynthesis pathways. The findings here suggest that the metabolism of the analyzed bacteria could be altered by exposure to CPF and GLP, leading to an increased pro-inflammatory immune response.