Draft genome sequence and probiotic functional property analysis of Lactobacillus gasseri LM1065 for food industry applications.

Probiotics are defined as live organisms in the host that contribute to health benefits. Lactobacillus gasseri LM1065, isolated from human breast milk, was investigated for its probiotic properties based on its genome. Draft genome map and de novo assembly were performed using the PacBio RS II system and hierarchical genome assembly process (HGAP). Probiotic properties were determined by the resistance to gastric conditions, adherence ability, enzyme production, safety assessment and mobile genetic elements. The fungistatic effect and inhibition of hyphae transition were studied using the cell-free supernatant (CFS). L. gasseri LM1065 showed high gastric pepsin tolerance and mild tolerance to bile salts. Auto-aggregation and hydrophobicity were measured to be 61.21% and 61.55%, respectively. The adherence to the human intestinal epithelial cells was measured to be 2.02%. Antibiotic-resistance genes and putative virulence genes were not predicted in the genomic analysis, and antibiotic susceptibility was satisfied by the criteria of the European Food Safety Authority. CFS showed a fungistatic effect and suppressed the tricarboxylic acid cycle in Candida albicans (29.02%). CFS also inhibited the transition to true hyphae and damaged the blastoconidia. This study demonstrates the essential properties of this novel probiotic, L. gasseri LM1065, and potential to inhibit vaginal C. albicans infection.

Effects of Lactobacillus gasseri CP2305 on Mild Menopausal Symptoms in Middle-Aged Women.

Lactobacillus gasseri CP2305 (CP2305) is a paraprobiotic that exhibits beneficial effects on the intestinal function and microbiota, and increases resistance to psychological stress. The stress response mechanism mainly involves the hypothalamic-pituitary-adrenal axis, which is influenced by the gut-brain axis. Furthermore, the gut-brain axis also communicates bidirectionally with the intestinal microbiota. Additionally, the hypothalamic-pituitary-adrenal and hypothalamic-pituitary-gonadal axes share a common route that affects both mental and health aspects in women. This double-blind, placebo-controlled, parallel-group clinical trial aimed to analyze the influence of the intake of CP2305 on mild symptoms associated with menopause. Eighty women aged 40-60 years ingested CP2305 or placebo tablets for six consecutive menstrual cycles. Assessment was based on the observation of climacteric symptoms with two validated questionnaires-the Simplified Menopausal Index (SMI) and the Greene Climacteric Scale (GCS). The results showed that CP2305 provided significant relief in the SMI total score, SMI vasomotor score, SMI psychological score, GCS total score, GCS somatic score, and GCS vasomotor score compared to the placebo. The percentage of women with symptom relief for the SMI total score was 75.0%, with 30 of 40 women in the CP2305 group, and 55.0%, with 22 of 40 women in the placebo group (p = 0.0594). These findings provide new insights into the function of paraprobiotic CP2305 in relieving mild climacteric symptoms in women.

A Synbiotic Combination of Lactobacillus gasseri 505 and Cudrania tricuspidata Leaf Extract Prevents Stress-Induced Testicular Dysfunction in Mice.

This study investigated the effects of a synbiotic combination (Syn) of Lactobacillus gasseri 505 (505) and Cudrania tricuspidata leaf extract (CT) on the hypothalamic-pituitary-gonadal axis in mice under chronic stress. Unpredictable chronic mild stress (UCMS) significantly increased the serum levels of corticosterone, however, treatment with Syn suppressed UCMS-induced increases. Histopathological analysis of the testes showed that these organs experienced some damage during UCMS, but this was repaired following treatment with Syn. Similarly, the transcription levels of gonadotropin-releasing hormone (GnRH), GnRH receptor, and gonadotropins, moreover, testicular development (i.e., Adam5, Adam29, and Spam1) - and steroidogenesis (i.e., Lhr, Egfr, and StAR) -related genes were significantly downregulated by UCMS. These UCMS-induced changes were inhibited by the administration of Syn, which was confirmed by the results of in situ hybridization analysis. These results suggest that the administration of Syn could attenuate the testicular dysfunctions induced by UCMS.

Ceftriaxone causes dysbiosis and changes intestinal structure in adjuvant obesity treatment.

BACKGROUND: Obesity is still a worldwide public health problem, requiring the development of adjuvant therapies to combat it. In this context, modulation of the intestinal microbiota seems prominent, given that the composition of the intestinal microbiota contributes to the outcome of this disease. The aim of this work is to investigate the treatment with an antimicrobial and/or a potential probiotic against overweight. METHODS: Male C57BL/6J mice were subjected to a 12-week overweight induction protocol. After that, 4-week treatment was started, with mice divided into four groups: control, treated with distilled water; potential probiotic, with Lactobacillus gasseri LG-G12; antimicrobial, with ceftriaxone; and antimicrobial + potential probiotic with ceftriaxone in the first 2 weeks and L. gasseri LG-G12 in the subsequent weeks. RESULTS: The treatment with ceftriaxone in isolated form or in combination with the potential probiotic provided a reduction in body fat. However, such effect is supposed to be a consequence of the negative action of ceftriaxone on the intestinal microbiota composition, and this intestinal dysbiosis may have contributed to the destruction of the intestinal villi structure, which led to a reduction in the absorptive surface. Also, the effects of L. gasseri LG-G12 apparently have been masked by the consumption of the high-fat diet. CONCLUSIONS: The results indicate that the use of a ceftriaxone in the adjuvant treatment of overweight is not recommended due to the potential risk of developing inflammatory bowel disease.

Insight into phenotypic and genotypic differences between vaginal Lactobacillus crispatus BC5 and Lactobacillus gasseri BC12 to unravel nutritional and stress factors influencing their metabolic activity.

The vaginal microbiota, normally characterized by lactobacilli presence, is crucial for vaginal health. Members belonging to L. crispatus and L. gasseri species exert crucial protective functions against pathogens, although a total comprehension of factors that influence their dominance in healthy women is still lacking. Here we investigated the complete genome sequence and comprehensive phenotypic profile of L. crispatus strain BC5 and L. gasseri strain BC12, two vaginal strains featured by anti-bacterial and anti-viral activities. Phenotype microarray (PM) results revealed an improved capacity of BC5 to utilize different carbon sources as compared to BC12, although some specific carbon sources that can be associated to the human diet were only metabolized by BC12, i.e. uridine, amygdalin, tagatose. Additionally, the two strains were mostly distinct in the capacity to utilize the nitrogen sources under analysis. On the other hand, BC12 showed tolerance/resistance towards twice the number of stressors (i.e. antibiotics, toxic metals etc.) with respect to BC5. The divergent phenotypes observed in PM were supported by the identification in either BC5 or BC12 of specific genetic determinants that were found to be part of the core genome of each species. The PM results in combination with comparative genome data provide insights into the possible environmental factors and genetic traits supporting the predominance of either L. crispatus BC5 or L. gasseri BC12 in the vaginal niche, giving also indications for metabolic predictions at the species level.

Heat inactivation partially preserved barrier and immunomodulatory effects of Lactobacillus gasseri LA806 in an in vitro model of bovine mastitis.

Probiotics could help combat infections and reduce antibiotic use. As use of live bacteria is limited in some cases by safety or regulatory concerns, the potential of inactivated bacteria is worth investigating. We evaluated the potential of live and heat-inactivated Lactobacillus gasseri LA806 to counteract Staphylococcus aureus and Escherichia coli infection cycles in an in vitro model of bovine mastitis. We assessed the ability of live and inactivated LA806 to impair pathogen colonisation of bovine mammary epithelial cells (bMECs) and to modulate cytokine expression by pathogen-stimulated bMECs. Live LA806 induced a five-fold decrease in S. aureus adhesion and internalisation (while not affecting E. coli colonisation) and decreased pro-inflammatory cytokine expression by S. aureus-stimulated bMECs (without interfering with the immune response to E. coli). The ability of inactivated LA806 ability to diminish S. aureus colonisation was two-fold lower than that of the live strain, but its anti-inflammatory properties were barely impacted. Even though LA806 effects were impaired after inactivation, both live and inactivated LA806 have barrier and immunomodulatory properties that could be useful to counteract S. aureus colonisation in the bovine mammary gland. As S. aureus is involved in various types of infection, LA806 potential would worth exploring in other contexts.

Effects of a Potential Probiotic Strain Lactobacillus gasseri ATCC 33323 on Helicobacter pylori-Induced Inflammatory Response and Gene Expression in Coinfected Gastric Epithelial Cells.

In the present study, we aimed to investigate the modulatory effects of a potential probiotic bacterium Lactobacillus gasseri ATCC 33323 on Helicobacter pylori-induced inflammatory response and gene expression in human gastric adenocarcinoma (AGS) cell line. The gastric epithelial cells were coinfected with a collection of H. pylori clinical strains alone or in combination with L. gasseri at a multiplicity of infection (MOI) of 1:100 for each bacterium, and incubated for different time points of 3, 6, and 12 h. IL-8 secretion from coinfected AGS cells after incubation at each time point was measured by an enzyme-linked immunosorbent assay (ELISA). The mRNA expression of IL-8, Bcl-2, ß-catenin, integrin α5, and integrin ß1 genes was determined by quantitative RT-PCR amplification of total RNA extracted from coinfected epithelial cells. L. gasseri significantly (P < 0.05 and P < 0.01) decreased the production of IL-8 in AGS cells coinfected with H. pylori strains at 6 h post-infection. We also detected that L. gasseri significantly (P < 0.05) down-regulated the gene expression level of IL-8 in H. pylori-stimulated AGS cells after 6 and 12 h of coinfection. Similarly, L. gasseri caused a significant decrease (P < 0.05) in mRNA expression of Bcl-2, ß-catenin, integrin α5, and integrin ß1 genes in AGS cells at 3 and 6 h after infection with H. pylori strains as compared with non-infected control cells. In conclusion, our results demonstrated that L. gasseri ameliorates H. pylori-induced inflammation and could be developed as a supplementation to the current treatment regimens administrated against H. pylori infection.

A Probiotic Lactobacillus gasseri Alleviates Escherichia coli-Induced Cognitive Impairment and Depression in Mice by Regulating IL-1ß Expression and Gut Microbiota.

Excessive expression of interleukin (IL)-1ß in the brain causes depression and cognitive dysfunction. Herein, we investigated the effect of Lactobacillus gasseri NK109, which suppressed IL-1ß expression in activated macrophages, on Escherichia coli K1-induced cognitive impairment and depression in mice. Germ-free and specific pathogen-free mice with neuropsychiatric disorders were prepared by oral gavage of K1. NK109 alleviated K1-induced cognition-impaired and depressive behaviors, decreased the expression of IL-1ß and populations of NF-κB+/Iba1+ and IL-1R+ cells, and increased the K1-suppressed population of BDNF+/NeuN+ cells in the hippocampus. However, its effects were partially attenuated by celiac vagotomy. NK109 treatment mitigated K1-induced colitis and gut dysbiosis. Tyndallized NK109, even if lysed, alleviated cognitive impairment and depression. In conclusion, NK109 alleviated neuropsychiatric disorders and colitis by modulating IL-1ß expression, gut microbiota, and vagus nerve-mediated gut-brain signaling.

Lactobacillus gasseri JM1 with potential probiotic characteristics alleviates inflammatory response by activating the PI3K/Akt signaling pathway in vitro.

Lactobacillus gasseri JM1, a novel strain isolated from infant feces, exhibited common probiotic properties such as high acid tolerance, bile salt tolerance, and adhesion to epithelial Caco-2 cells, suggesting its ability to survive in the gastrointestinal tract and confer potential probiotic action on the host. In the current study, we aimed to evaluate the immunomodulatory activity of L. gasseri JM1 and explore the underlying signaling pathways in vitro. The results showed that pretreatment with L. gasseri JM1 alleviated lipopolysaccharide-induced inflammatory response, as evidenced by downregulation of genes encoding proinflammatory cytokines [IL1B, IL6, IL8, and tumor necrosis factor-α (TNFA)] and upregulation of genes encoding anti-inflammatory cytokines [IL4, IL10, transforming growth factor-ß3 (TGFB3), and IFNG]. A high level of gene expression was noted for toll-like receptor 2 and NOD-like receptor 2. Meanwhile, transcriptomic sequencing obtained 84 differentially expressed genes. Kyoto Encyclopedia of Genes and Genomes analysis revealed the phosphatidylinositol 3-kinase (PI3K)/protein kinase B (Akt) signaling pathway was activated by L. gasseri JM1 in Caco-2 cells. Inhibitor of PI3K/Akt played various roles in the release of cytokines, indicating that the pathway was involved in protecting the host against lipopolysaccharide stress. Moreover, whole-genome sequencing revealed fundamental genetic properties of L. gasseri JM1 and provided clues for probiotic characteristics. In summary, the strain could exert immunomodulatory effects via the toll-like receptor 2 and NOD2-mediated PI3K/Akt signaling pathway and be regarded as a potential probiotic.

Unveiling the presence of epigenetic mark by Lactobacillus supplementation in high-fat diet-induced metabolic disorder in Sprague-Dawley rats.

Gut dysbiosis, particularly bacteria from Firmicutes and Bacteroidetes phyla, plays a fundamental role in the progression of metabolic disorders. Probiotics have shown to restore the gut microbiota composition in metabolic disorders with subsequent beneficial effects. Recent studies have reported that several species of Lactobacillus as probiotic supplementation improve insulin sensitivity and glucose metabolism. Nonetheless, whether Lactobacillus could influence the epigenetic modifications that underlie insulin-resistant conditions is still unexplored. Therefore, the current study examined the therapeutic effects and underlying epigenetic mechanisms of three different species of Lactobacillus in the high-fat diet (HFD)-induced insulin-resistant rats. Three different species of Lactobacillus; Lactobacillus casei, Lactobacillus gasseri, and Lactobacillus rhamnosus were individually supplemented orally (109 CFU/mL) to insulin-resistant SD rats for 12 weeks. Lactobacillus supplementation led to a significant reduction in the hyperglycemia, hyperinsulinemia, and hyperlipidemia associated with HFD-induced insulin resistance. Histopathological examination also indicated the protective effects of Lactobacillus supplementation against the hepatic and intestinal damage caused by the high-fat diet. Lactobacillus supplementation also down-regulated the expression of FOXO1, a major transcription factor of insulin signaling. In addition, at the epigenetic level, Lactobacillus supplementation predominantly prevented methylation and demethylation of H3K79me2 and H3K27me3, respectively. Chromatin Immunoprecipitation (ChIP) coupled with quantitative PCR (ChIP-qPCR) assay revealed the presence of cross-talk between these two histone modifications at the promoter region of FOXO1. Taken together, this is the first report to observe that the effects of Lactobacillus supplementation involve alteration in FOXO1 expression via cross-talking between H3K79me2 and H3K27me3 histone modifications.

Anti-Inflammatory Effect of a Peptide Derived from the Synbiotics, Fermented Cudrania tricuspidata with Lactobacillus gasseri, on Inflammatory Bowel Disease.

The objective of this study was to evaluate the effects of peptides derived from synbiotics on improving inflammatory bowel disease (IBD). Five-week-old male C57BL/6 mice were administered with dextran sulfate sodium (DSS) via drinking water for seven days to induce IBD (IBD group). The mice in the IBD group were orally administered with PBS (IBD-PBS-positive control), Lactobacillus gasseri 505 (IBD-Pro), fermented powder of CT extract with L. gasseri 505 (IBD-Syn), ß-casein: LSQSKVLPVPQKAVPYPQRDMP (IBD-Pep 1), or α s2-casein: VYQHQKAMKPWIQPKTKVIPYVRYL (IBD-Pep 2) (both peptides are present in the synbiotics) for four more days while inducing IBD. To confirm IBD induction, the weights of the animals and the disease activity index (DAI) scores were evaluated once every two days. Following treatment of probiotics, synbiotics, or peptides for 11 days, the mice were sacrificed. The length of the small and large intestines was measured. The expression of the proinflammatory cytokines IL-1ß, IL-6, TNF-α, and COX-2 in the large intestine was measured. Large intestine tissue was fixed in 10% formalin and stained with hematoxylin and eosin for histopathological analysis. The body weights decreased and DAI scores increased in the IBD group, but the DAI scores were lower in the IBD-Pep 2 group than those in the IBD group treated with PBS, Pro, Syn, or Pep 1. The lengths of the small and large intestines were shorter in the IBD group than in the group without IBD, and the expression levels of the proinflammatory cytokines were lower (p < 0.05) in the IBD-Pep 2 group than those in the IBD-PBS-positive control group. In addition, histopathological analysis showed that IBD was ameliorated in the Pep 2-treated group. These results indicate that Pep 2 derived from α s2-casein was effective in alleviating IBD-associated inflammation. Thus, we showed that these peptides can alleviate inflammation in IBD.

Cancer-protective effect of a synbiotic combination between Lactobacillus gasseri 505 and a Cudrania tricuspidata leaf extract on colitis-associated colorectal cancer.

Previously, a synbiotic combination of probiotic Lactobacillus gasseri 505 (LG) and a new prebiotic, Cudrania tricuspidata leaf extract (CT) in fermented milk, designated FCT, showed an in vitro immunomodulatory effect and antioxidant activity. Although synbiotic combination might have cancer-protective effects, these activities have not been fully validated in vivo. Ten-week treatment of LG, CT, or FCT to azoxymethane (AOM)/dextran sodium sulfate (DSS)-induced colitis-associated colorectal cancer (CAC) mouse model reduced both the incidence of colonic tumors and damage to the colonic mucosa effectively, suggesting a cancer-protective effect. To understand these, biomarkers associated with inflammation, colon barrier, apoptosis, and cancer cell proliferation were monitored in AOM/DSS group versus LG/CT/FCT groups. A synbiotic combination (FCT) down-regulated pro-inflammatory cytokines (TNF-α, IFN-γ, IL-1ß, and IL-6) and inflammation-associated enzymes (iNOS and COX-2), and up-regulated anti-inflammatory cytokines (IL-4 and IL-10). In addition, colon barrier experiment revealed that biomarkers of mucus layer (MUC-2 and TFF3) and tight junction (occludin and ZO-1) were up-regulated. Subsequent apoptosis experiment showed that pro-apoptotic factors (p53, p21, and Bax) were up-regulated and anti-apoptotic factors (Bcl-2 and Bcl-xL) were down-regulated. Furthermore, comparative metagenome analysis of gut microbiota revealed that Staphylococcus decreased but Lactobacillus, Bifidobacterium, and Akkermansia increased, supporting their protective effects, accompanied by increased short-chain fatty acids (SCFAs). Taken together, the FCT administration showed cancer-protective effects by reducing the risk of colitis-associated colon cancer via regulation of inflammation, carcinogenesis, and compositional change of gut microbiota. Consequently, the synbiotic combination (FCT) could be a novel potential health-protective natural agent against CAC.

Synergistic effect of Lactobacillus gasseri and Cudrania tricuspidata on the modulation of body weight and gut microbiota structure in diet-induced obese mice.

High-fat diet (HFD)-induced obesity has been associated with alteration of gut microbiota alongside body weight gain. In this study, the synbiotic effect of Lactobacillus gasseri 505 (LG) and Cudrania tricuspidata (CT) in HFD-induced mice was revealed. After feeding mice with high-fat diet for 10 weeks, combination of LG and CT (LG_CT) exhibited the greatest reduction in the final body weight (11.9%). Moreover, microbial diversity significantly increased, and Principal Coordinate Analysis (PCoA) revealed that the LG_CT group showed closer cluster to NORM. At phylum level, the Firmicutes/Bacteroidetes (F/B) ratio increased in HFD, and the abundance of Bacteroidetes was restored by LG and CT. At genus level, notable changes in Alistipes, Desulfovibrio, Bilophila, and Acetatifactor were observed. Helicobacter elevated to 16.2% in HFD and diminished dramatically to less than 0.01% in LG and/or CT. At species level, L. gasseri increased after the administration of LG (0.54%) and LG_CT (1.14%), suggesting that LG may grow and colonize in the gut and CT can function as a prebiotic. Finally, functional analysis revealed certain metabolic factors correlated with body weight and gut microbiota. This study serves as a potential basis for the application of L. gasseri 505 and C. tricuspidata in the prevention and treatment of diet-induced obesity.Key Points • Combination of L. gasseri (LG) and C. tricuspidata (CT) reduced body weight gain.• Microbial diversity significantly increased in LG_CT treatment.• Abundance of microorganisms involved with leanness increased in LG, CT, and LG_CT.• Body weight is associated with some metabolic functions of gut microbiota.

The effects of viable and non-viable Lactobacillus gasseri CP2305 cells on colonic ion transport and corticotropin releasing factor-induced diarrhea.

The effect of non-viable lactic acid bacteria on gastrointestinal physiology and dysfunction remains still unclear. Previous clinical trials have reported that Lactobacillus gasseri CP2305 (CP2305) exerts stress-relieving and anti-flatulent effects regardless of cell viability. In this study, we investigated the effect of viable and non-viable CP2305 cells on electrical field stimulation (EFS)-evoked increases in short-circuit current (Isc) using the Ussing chamber technique. In mucosal-submucosal preparations of rats, both viable and non-viable CP2305 cells significantly and acutely inhibited the EFS-evoked increases in Isc in the middle and distal colon and rectum but not in proximal colon. The inhibition of EFS-evoked Isc differed from strain to strain. Peripheral injection of corticotropin releasing factor (CRF) is known to mimic diarrhea symptoms in rats. Therefore, we examined the chronic effects of CP2305 cells on CRF-induced diarrhea in the rat model. Treatment with viable and non-viable CP2305 cells significantly improved CRF-induced diarrhea in the rat model. However, the treatment did not affect the fecal pellet output. These findings suggest that CP2305 has an important role in gastrointestinal physiology and dysfunction.

Lactobacillus gasseri Suppresses the Production of Proinflammatory Cytokines in Helicobacter pylori-Infected Macrophages by Inhibiting the Expression of ADAM17.

The ability of Helicobacter pylori to evade the host immune system allows the bacterium to colonize the host for a lifetime. Long-term infection with H. pylori causes chronic inflammation, which is the major risk factor for the development of gastric ulcers and gastric cancer. Lactobacilli are part of the human microbiota and have been studied as an adjunct treatment in H. pylori eradication therapy. However, the molecular mechanisms by which lactobacilli act against H. pylori infection have not been fully characterized. In this study, we investigated the anti-inflammatory effects of Lactobacillus strains upon coincubation of host macrophages with H. pylori. We found that Lactobacillus gasseri Kx110A1 (L. gas), a strain isolated from a human stomach, but not other tested Lactobacillus species, blocked the production of the proinflammatory cytokines TNF and IL-6 in H. pylori-infected macrophages. Interestingly, L. gas also inhibited the release of these cytokines in LPS or LTA stimulated macrophages, demonstrating a general anti-inflammatory property. The inhibition of these cytokines did not occur through the polarization of macrophages from the M1 (proinflammatory) to M2 (anti-inflammatory) phenotype or through the altered viability of H. pylori or host cells. Instead, we show that L. gas suppressed the release of TNF and IL-6 by reducing the expression of ADAM17 (also known as TNF-alpha-converting enzyme, TACE) on host cells. Our findings reveal a novel mechanism by which L. gas prevents the production of the proinflammatory cytokines TNF and IL-6 in host macrophages.

Application of Lactobacillus gasseri 63 AM supernatant to Pseudomonas aeruginosa-infected wounds prevents sepsis in murine models of thermal injury and dorsal excision.

Introduction. Severely burned patients are susceptible to bacterial infection within their burn wounds, which frequently leads to sepsis, multiple organ failure and death. The opportunistic pathogen Pseudomonas aeruginosa, an organism inherently resistant to multiple antibiotics, is a common cause of sepsis in these patients.Aim. Development of a topical treatment unrelated to conventional antibiotics is essential for prevention of P. aeruginosa infection and sepsis, leading to a role for the direct application of probiotics or their by-products.Methodology. We examined the effectiveness of 20× concentrated supernatant from Lactobacillus gasseri strain 63 AM (LgCS) grown in de Man, Rogosa and Sharpe broth in inhibiting P. aeruginosa biofilms in vitro, as well as in reducing wound bioburden and P. aeruginosa sepsis in vivo.Results. LgCS inhibited the growth of P. aeruginosa strain PAO1, prevented its biofilm development and eliminated partially developed PAO1 biofilms. In the murine model of thermal injury, a single injection of LgCS following injury and PAO1 infection reduced mortality to 0 % and prevented systemic spread (sepsis). Furthermore, a second injection of LgCS 24 h after the first eliminated PAO1 from the wound. In the murine dorsal excision infection model, either LgCS or ceftazidime treatment of the PAO1-infected wound significantly reduced the mortality rate among infected mice, while combining LgCS with ceftazidime eliminated mortality.Conclusion. These results suggest the potential of LgCS in preventing sepsis from P. aeruginosa infection in severely burned and other immunocompromised patients.

Putative Adhesion Factors in Vaginal Lactobacillus gasseri DSM 14869: Functional Characterization.

Lactobacilli play an important role in the maintenance of a healthy vaginal microbiota, and some select species are widely used as probiotics. Vaginal isolates of Lactobacillus gasseri DSM 14869 and Lactobacillus rhamnosus DSM 14870 were previously selected to develop the probiotic EcoVag capsules and showed therapeutic effects in women with bacterial vaginosis (BV). However, the molecular mechanisms involved in their probiotic activity are largely unknown. In this study, we identified three cell surface molecules in L. gasseri DSM 14869 that promote adhesion to vaginal epithelial cells (VEC) by constructing dedicated knockout mutants, including exopolysaccharides (EPSs), a protein containing MucBP-like domains (N506_1778), and a putative novel adhesin (N506_1709) with rib/alpha-like domain repeats. EPS knockout mutants revealed 20-fold and 14-fold increases in adhesion to Caco-2 and HeLa cells, respectively, compared with wild type, while the adhesion to VEC was reduced 30% by the mutation, suggesting that EPSs might mediate tissue tropism for vaginal cells. A significant decrease in adhesion to Caco-2 cells, HeLa cells, and VEC was observed in the N506_1778 knockout mutant. The N506_1709 mutant showed no significant difference for the adhesion to Caco-2 and HeLa cells compared with wild type (WT); in contrast, the adhesion to VEC revealed a significant decrease (42%), suggesting that N506_1709 might mediate specific binding to stratified squamous epithelial cells, and this putative novel adhesin was annotated as Lactobacillus vaginal epithelium adhesin (LVEA). Thus, we have discovered an important role for EPSs and a novel adhesin, LVEA, in the adhesive capacity of a vaginal probiotic Lactobacillus strain.IMPORTANCE Lactobacilli are known to contribute to the maintenance of a healthy vaginal microbiota and some are selected as probiotics for the prevention or treatment of urogenital diseases, such as bacterial vaginosis. However, the molecular mechanisms for these health-promoting effects are not fully understood. Here, we functionally identified three cell surface factors of a Lactobacillus gasseri strain potentially involved in its adhesion to vaginal epithelial cells, including exopolysaccharides (EPSs) and two sortase-dependent proteins (N506_1778 and N506_1709). We could demonstrate the tissue-specific adhesion of EPSs to vaginal cells and that N506_1709 might be a novel adhesin specifically mediating bacterial binding to stratified squamous epithelial cells. The results provide important new information on the molecular mechanisms of vaginal Lactobacillus spp. adhesion.

Catalase-mimetic gold nanoparticles inhibit the antagonistic action of Lactobacillus gasseri toward foodborne enteric pathogens in associative cultures.

Gold nanoparticles (AuNPs) have been previously shown to induce gut dysbiosis during colitis in mice, but the underlying mechanism is not clear yet. Here, we evaluated the effects of AuNPs (5 nm diameter, coated with tannic acid, polyvinylpyrrolidone or citrate) on H2O2 accumulation and pathogen antagonization by an intestinal strain of Lactobacillus gasseri under aerobic cultural conditions. AuNPs (0.65 µg/mL) reduced over 50% of H2O2 accumulation by L. gasseri, and significantly inhibited the antagonistic action of L. gasseri on growth of four foodborne enteric pathogens, i.e. Salmonella enterica serovar Typhimurium, Escherichia coli, Listeria monocytogenes, and Staphylococcus aureus in associative cultures.

The membrane phospholipid cardiolipin plays a pivotal role in bile acid adaptation by Lactobacillus gasseri JCM1131T.

Bile acids exhibit strong antimicrobial activity as natural detergents, and are involved in lipid digestion and absorption. We investigated the mechanism of bile acid adaptation in Lactobacillus gasseri JCM1131T. Exposure to sublethal concentrations of cholic acid (CA), a major bile acid in humans, resulted in development of resistance to otherwise-lethal concentrations of CA by this intestinal lactic acid bacterium. As this adaptation was accompanied by decreased cell-membrane damage, we analyzed the membrane lipid composition of L. gasseri. Although there was no difference in the proportions of glycolipids (~70%) and phospholipids (~20%), adaptation resulted in an increased abundance of long-sugar-chain glycolipids and a 100% increase in cardiolipin (CL) content (to ~50% of phospholipids) at the expense of phosphatidylglycerol (PG). In model vesicles, the resistance of PG vesicles to solubilization by CA increased with increasing CL/PG ratio. Deletion of the two putative CL synthase genes, the products of which are responsible for CL synthesis from PG, decreased the CL content of the mutants, but did not affect their ability to adapt to CA. Exposure to CA restored the CL content of the two single-deletion mutants, likely due to the activities of the remaining CL synthase. In contrast, the CL content of the double-deletion mutant was not restored, and the lipid composition was modified such that PG predominated (~45% of total lipids) at the expense of glycolipids. Therefore, CL plays important roles in bile acid resistance and maintenance of the membrane lipid composition in L. gasseri.

Long-Term Administration of Conjugated Estrogen and Bazedoxifene Decreased Murine Fecal ß-Glucuronidase Activity Without Impacting Overall Microbiome Community.

Conjugated estrogens (CE) and Bazedoxifene (BZA) combination is used to alleviate menopause-associated symptoms in women. CE+BZA undergo first-pass-metabolism in the liver and deconjugation by gut microbiome via ß-glucuronidase (GUS) enzyme inside the distal gut. To date, the impact of long-term exposure to CE+BZA on the gut microbiome or GUS activity has not been examined. Our study using an ovariectomized mouse model showed that CE+BZA administration did not affect the overall cecal or fecal microbiome community except that it decreased the abundance of Akkermansia, which was identified as a fecal biomarker correlated with weight gain. The fecal GUS activity was reduced significantly and was positively correlated with the abundance of Lactobacillaceae in the fecal microbiome. We further confirmed in Escherichia coli K12 and Lactobacillus gasseri ADH that Tamoxifen-, 4-hydroxy-Tamoxifen- and Estradiol-Glucuronides competed for GUS activity. Our study for the first time demonstrated that long-term estrogen supplementation directly modulated gut microbial GUS activity. Our findings implicate that long-term estrogen supplementation impacts composition of gut microbiota and microbial activity, which affects estrogen metabolism in the gut. Thus, it is possible to manipulate such activity to improve the efficacy and safety of long-term administered estrogens for postmenopausal women or breast cancer patients.