Effect of Lacticaseibacillus paracasei K56 with galactooligosaccharide synbiotics on obese individuals: an in vitro fermentation model.

BACKGROUND: The use of synbiotics is emerging as a promising intervention strategy for regulating the gut microbiota and for preventing or reducing obesity, in comparison with the use of probiotics or prebiotics alone. A previous in vivo study revealed that Lacticaseibacillus paracasei K56 (L. paracasei K56) could alleviate obesity induced in high-fat-diet mice; however, the effect of the synbiotic combination of L. paracasei K56 and prebiotics in obese individuals has not been explored fully. RESULTS: The effect of prebiotics on the proliferation of L. paracasei K56 was determined by spectrophotometry. The results showed that polydextrose (PG), xylooligosaccharide (XOS), and galactooligosaccharide (GOS) had a greater potential to be used as substrates for L. paracasei K56 than three other prebiotics (melitose, stachyose, and mannan-oligosaccharide). An in vitro fermentation model based on the feces of ten obese female volunteers was then established. The results revealed that K56_GOS showed a significant increase in GOS degradation rate and short-chain fatty acid (SCFA) content, and a decrease in gas levels, compared with PG, XOS, GOS, K56_PG, and K56_XOS. Changes in these microbial biomarkers, including a significant increase in Bacteroidota, Bifidobacterium, Lactobacillus, Faecalibacterium, and Blautia and a decrease in the Firmicutes/Bacteroidota ratio and Escherichia-Shigella in the K56_GOS group, were associated with increased SCFA content and decreased gas levels. CONCLUSION: This study demonstrates the effect of the synbiotic combination of L. paracasei K56 and GOS on obese individuals and indicates its potential therapeutic role in obesity treatment. © 2024 Society of Chemical Industry.

Efficacy of Bifidobacterium animalis subsp. lactis BL-99 in the treatment of functional dyspepsia: a randomized placebo-controlled clinical trial.

Current treatment for functional dyspepsia (FD) has limited and unsustainable efficacy. Probiotics have the sustainable potential to alleviate FD. This randomized controlled clinical trial (Chinese Clinical Trial Registry, ChiCTR2000041430) assigned 200 FD patients to receive placebo, positive-drug (rabeprazole), or Bifidobacterium animalis subsp. lactis BL-99 (BL-99; low, high doses) for 8-week. The primary outcome was the clinical response rate (CRR) of FD score after 8-week treatment. The secondary outcomes were CRR of FD score at other periods, and PDS, EPS, serum indicators, fecal microbiota and metabolites. The CRR in FD score for the BL-99_high group [45 (90.0%)] was significantly higher than that for placebo [29 (58.0%), p = 0.001], BL-99_low [37 (74.0%), p = 0.044] and positive_control [35 (70.0%), p = 0.017] groups after 8-week treatment. This effect was sustained until 2-week after treatment but disappeared 8-week after treatment. Further metagenomic and metabolomics revealed that BL-99 promoted the accumulation of SCFA-producing microbiota and the increase of SCFA levels in stool and serum, which may account for the increase of serum gastrin level. This study supports the potential use of BL-99 for the treatment of FD.

Dynamic metagenome-scale metabolic modeling of a yogurt bacterial community.

Genome-scale metabolic models and flux balance analysis (FBA) have been extensively used for modeling and designing bacterial fermentation. However, FBA-based metabolic models that accurately simulate the dynamics of coculture are still rare, especially for lactic acid bacteria used in yogurt fermentation. To investigate metabolic interactions in yogurt starter culture of Streptococcus thermophilus and Lactobacillus delbrueckii subsp. bulgaricus, this study built a dynamic metagenome-scale metabolic model which integrated constrained proteome allocation. The accuracy of the model was evaluated by comparing predicted bacterial growth, consumption of lactose and production of lactic acid with reference experimental data. The model was then used to predict the impact of different initial bacterial inoculation ratios on acidification. The dynamic simulation demonstrated the mutual dependence of S. thermophilus and L. d. bulgaricus during the yogurt fermentation process. As the first dynamic metabolic model of the yogurt bacterial community, it provided a foundation for the computer-aided process design and control of the production of fermented dairy products.

Lactobacillus paracasei ET-22 and derived postbiotics reduce halitosis and modulate oral microbiome dysregulation - a randomized, double-blind placebo-controlled clinical trial.

Oral microbial dysbiosis is the primary etiologic factor for halitosis and may be the critical preventive target for halitosis. This study included randomized controlled trials (RCTs) assessing the effects of Lactobacillus paracasei ET-22 live and heat-killed bacteria on halitosis and the related oral microbiome. 68 halitosis subjects were divided into placebo, ET-22 live (ET-22.L) and ET-22 heat-killed (ET-22.HK) groups. Subjects took different lozenges three times a day for 4 weeks and underwent saliva collection and assessment of breath volatile sulfur compound (VSC) levels at the beginning and end of the intervention. Salivary volatile organic compounds were measured using HS-SPME-GC/MS, and the microbiome profile was determined by 16S rRNA gene amplicon sequencing. A positive decrease in breath volatile sulfur compound (VSC) levels was observed in the means of both ET-22.L and ET-22.HK groups after 4 weeks of intervention, being more marked in the ET-22.L group (p = 0.0148). Moreover, ET-22.L and ET-22.HK intervention remarkably changed the composition of total salivary volatile organic compounds (VOCs) and aroma-active VOCs. Key undesirable VOCs, such as indole, pyridine, nonanoic acid, benzothiazole, and valeric acid, were significantly reduced. Meanwhile, ET-22.L or ET-22.HK also altered the taxonomic composition of the salivary microbiome. The halitosis pathogens Rothia and Streptococcus were significantly reduced in the ET-22.HK group and the pathogenic Solobacterium and Peptostreptococcus were significantly inhibited in the ET-22.L group. Collectively, our study suggests that both ET-22.L and ET-22.HK can significantly inhibit the production of undesirable odor compounds in subjects with halitosis, which may be related to the changes of the oral microbiome.

Effect of different doses of Lacticaseibacillus paracasei K56 on body fat and metabolic parameters in adult individuals with obesity: a pilot study.

BACKGROUND: Studies have shown that probiotics have an effect on reducing body fat on a strain-specific and dose-response bases. The purpose of this study was to evaluate the effect of a novel probiotic strain Lacticaseibacillus paracasei K56 on body fat and metabolic biomarkers in adult individuals with obesity. METHODS: 74 adult subjects with obesity (body mass index ≥ 30 kg/m2, or percent body fat > 25% for men, percent body fat > 30% for women) were randomized into 5 groups and supplemented with different doses of K56 (groups VL_K56, L_K56, H_K56, and VH_K56: K56 capsules, 2 × 107 CFU/day, 2 × 109 CFU/day, 2 × 1010 CFU/day, 2 × 1011 CFU/day, respectively) or placebo (group Pla: placebo capsule) for 60 days. Subjects were advised to maintain their original dietary intake and physical activity. Anthropometric measurements, body composition assessment, and metabolic parameters were measured at baseline and after 60 days of intervention. RESULTS: The results showed that the L_K56 group had significant decreases in percent body fat (p = 0.004), visceral fat area (p = 0.0007), total body fat mass (p = 0.018), trunk body fat mass (p = 0.003), waist circumference (p = 0.003), glycosylated hemoglobin(p = 0.002) at the end of the study compared with baseline. There were non-significant reductions in Body weight and BMI in the L_K56, H_K56, VL_K56 groups, whereas increases were observed in the placebo and VH_K56 groups compared with baseline values. In addition, K56 supplementation modulated gut microbiota characteristics and diversity indices in the L-K56 group. However, mean changes in body fat mass, visceral fat area, weight, body mass index, waist circumference and hip circumference were not significantly different between groups. CONCLUSIONS: The results suggest that supplementation with different doses of Lacticaseibacillus paracasei K56 has certain effect on reducing body fat and glycosylated hemoglobin, especially at a dose of 109 CFU/day. TRIAL REGISTRATION: clinicaltrials.gov Identifier: NCT04980599.

Postbiotics Derived from L. paracasei ET-22 Inhibit the Formation of S. mutans Biofilms and Bioactive Substances: An Analysis.

Globally, dental caries is one of the most common non-communicable diseases for patients of all ages; Streptococcus mutans (S. mutans) is its principal pathogen. Lactobacillus paracasei (L. paracasei) shows excellent anti-pathogens and immune-regulation functions in the host. The aim of this study is to evaluate the effects of L. paracasei ET-22 on the formation of S. mutans biofilms. The living bacteria, heat-killed bacteria, and secretions of L. paracasei ET-22 were prepared using the same number of bacteria. In vitro, they were added into artificial-saliva medium, and used to coculture with the S. mutans. Results showed that the living bacteria and secretions of L. paracasei ET-22 inhibited biofilm-growth, the synthesis of water-soluble polysaccharide and water-insoluble polysaccharide, and virulence-gene-expression levels related to the formation of S. mutans biofilms. Surprisingly, the heat-killed L. paracasei ET-22, which is a postbiotic, also showed a similar regulation function. Non-targeted metabonomics technology was used to identify multiple potential active-substances in the postbiotics of L. paracasei ET-22 that inhibit the formation of S. mutans biofilms, including phenyllactic acid, zidovudine monophosphate, and citrulline. In conclusion, live bacteria and its postbiotics of L. paracasei ET-22 all have inhibitory effects on the formation of S. mutans biofilm. The postbiotics of L. paracasei ET-22 may be a promising biological anticariogenic-agent.

Bifidobacterium animalis subsp. lactis BL-99 ameliorates colitis-related lung injury in mice by modulating short-chain fatty acid production and inflammatory monocytes/macrophages.

Pulmonary inflammation as one of the extraintestinal manifestations of ulcerative colitis (UC) has attracted extensive attention, and its pathogenesis is closely related to gut dysbiosis. Bifidobacterium animalis subsp. lactis BL-99 (BL-99) can alleviate osteoporosis caused by UC, but less research has been done on other extraintestinal manifestations (EIM) caused by UC. This study aimed to explore the role and potential mechanisms of BL-99 on DSS-induced pulmonary complications in colitis mice. The results showed that BL-99 decreased weight loss, disease activity index score, colonic pathology score, and the production of pro-inflammatory cytokines (e.g., TNF-α, IL-1ß, and IL-6) in colitis mice. BL-99 also alleviated DSS-induced lung pathological damage by suppressing the infiltration of pro-inflammatory cytokines, inflammatory monocytes, and macrophages. Furthermore, 16S rRNA gene sequencing showed lower abundances of several potentially pathogenic bacteria (e.g., Burkholderia, Shigella, and Clostridium perfringens) and enrichment in specific beneficial bacteria (e.g., Adlercreutzia and Bifidobacterium animalis) in colitis mice with BL-99 treatment. Targeted metabolomics suggested that BL-99 intervention promoted the production of intestinal acetate and butyrate. Finally, we observed that the pulmonary expression of primary acetate and butyrate receptors, including FFAR2, FFAR3, and, GPR109a, was up-regulated in BL-99-treated mice, which negatively correlated with inflammatory monocytes and macrophages. Altogether, these results suggest that BL-99 might be utilized as a probiotic intervention to prevent the incidence of colitis-related lung injury owing to its ability to shape the intestinal microbiota and suppress inflammation.

Lacticaseibacillus paracasei K56 Attenuates High-Fat Diet-Induced Obesity by Modulating the Gut Microbiota in Mice.

This study investigated the effects of Lacticaseibacillus paracasei K56 (L. paracasei K56) on body weight, body composition, and glycolipid metabolism in mice with high-fat diet-induced obesity and explored the underlying mechanisms. Male C57BL/6J mice were fed a high-fat diet for 8 weeks to induce obesity; then, the obese mice were gavaged with or without L. paracasei K56 for 10 weeks. The body weight, body composition, fat mass, blood lipid, blood glucose, and hormones of the mice were evaluated. Moreover, the fatty acid synthesis (FAS) and peroxisome proliferator-activated receptor γ (PPAR-γ) expressions in the liver were detected via Western blotting. 16S rRNA gene sequencing was adopted to determine the gut microbiota alterations. The high-fat diet successfully induced obesity, as indicated by the abnormal increase in body weight, visceral fat, fat mass, blood lipids, fasting blood glucose, and insulin-resistance. Moreover, the FAS expression in the liver was significantly increased, whereas the PPAR-γ expression was significantly decreased. The relative abundance of Proteobacteria, Actinobacteria and Patescibacteria was also significantly increased, and that of Verrucomicrobia was significantly decreased. However, these indicators of mice supplemented with L. paracasei K56 were significantly opposite to those of obese mice. The Ruminococcuaceae_UCG-013, Akkermansia, Prevotellaceae_UCG-001, Muribaculum, and Lachnospiraceae_NK4A136 groups were significantly negatively correlated with body weight, blood lipids, and blood glucose-related indicators, whereas Coriobacteriaceae_UCG-002, Enterorhabdus, Raoultibacter, Acinetobacter, Romboutsia, Leuconostoc, and Erysipelatoclostridium were significantly positively correlated with these indicators. L. paracasei K56 might be a promising probiotic strain that could effectively slow down the body weight gain, reduce fat accumulation, alleviate insulin-resistance, and restore pancreatic ß-cell function in obese mice by regulating the gut microbiota.

In vitro Study of Bifidobacterium lactis BL-99 With Fructooligosaccharide Synbiotics Effected on the Intestinal Microbiota.

Probiotics and prebiotics relieve constipation by altering the composition of the intestinal microbiota. However, their synergistic mechanism of action remains unclear. Herein, an in vitro fermentation model was constructed to examine the synergistic effects of Bifidobacterium lactis BL-99 and fructooligosaccharide (FOS) on the regulation of intestinal microbiota from a population with constipation. The utilization of FOS was promoted by BL-99, and the increase rate being 22.33%. Relative to the BL-99 and the FOS groups, the BL-99_FOS group showed a highly significant increase in acetic acid content (P < 0.01) and a marked decrease in CO2 and H2S contents (P < 0.01) in the fermentation broth. In addition, the BL-99_FOS combination significantly changed the structure of the intestinal microbiota, enhanced the relative abundances of beneficial bacteria that relieved constipation, including Bifidobacterium, Fecalibacterium, Lactobacillus, Subdoligranulum, and Blautia, and decreased those of the harmful bacteria, including Bilophila and Escherichia-Shigella. These findings suggested that BL-99 and FOS synergistically regulated the composition and structure of the intestinal microbiota from the population with constipation and increased acetic acid and decreased CO2 and H2S levels, thereby providing a theoretical basis for the application of synbiotics.

Bifidobacterium infantis strain YLGB-1496 possesses excellent antioxidant and skin barrier-enhancing efficacy in vitro.

Atopic dermatitis (AD) is a recurring allergic skin disease that has a high incidence. Orally applied Bifidobacteria ameliorate signs of irritated skin and enhance the skin barrier. The present study investigated the safety and efficacy of a topically used cell-free culture supernatant (CFS) from a Bifidobacterium infantis strain using in vitro evaluation methods. The results showed that CFS had strong free radical scavenging activity on DPPH, ABTS, ·OH and O2 -radicals. CFS treatment fundamentally reduced the intracellular reactive oxygen species (ROS) and malondialdehyde (MDA) contents and improved the activities of antioxidant enzymes (CAT, SOD and GSH-Px) in H2 O2 -treated HaCaT cells. Notably, the upregulation of skin physical barrier gene (FLG, LOR, IVL, AQP3 and TGM1) expression and skin antimicrobial peptide gene (CAMP, hBD-2 and hBD-3) expression by CFS might contribute to skin barrier resistance. CFS was non-irritating to the skin and eyes. CFS from the Bifidobacterium infantis strain had strong antioxidant properties on the skin and strengthened skin barrier function, and it was safe for topical use.

Bifidobacterium lactis BL-99 protects mice with osteoporosis caused by colitis via gut inflammation and gut microbiota regulation.

Patients diagnosed with inflammatory bowel disease or related conditions also frequently suffer from osteoporosis as a consequence of changes in the intestinal microenvironment and consequent dysbiosis. We hypothesized that anti-inflammatory probiotic treatment would be sufficient to alleviate intestinal inflammation and thereby prevent the development of osteoporosis. To that end, the ability of Bifidobacterium lactis BL-99 administration to protect against bone loss in an experimental model of dextran sodium sulfate-induced ulcerative colitis (UC) was analyzed, and the underlying molecular mechanisms were interrogated in detail. The results of these analyses revealed that BL-99 administration suppressed colitis-associated weight loss (P < 0.05), disease activity index scores, and the production of proinflammatory cytokines (TNF-α, IL-1ß, IL-6, and IL-17) (P < 0.05). Colon tissue pathological sections similarly revealed BL-99-mediated reductions in tissue injury severity. Micro-computed tomography (Micro-CT) analyses further exhibited significant improvements in percent bone volume (BV/TV) as well as trabecular number and thickness in BL-99-treated animals (P < 0.05). Such probiotic supplementation also resulted in pronounced changes in the composition of the gut microbiota. Moreover, BL-99 intervention markedly increased the expression of intestinal barrier-related proteins (Claudin-1, MUC2, ZO-1, and Occludin). Together, these results suggest that BL-99 can be utilized as a beneficial probiotic preparation to prevent the incidence of osteoporosis in UC patients owing to its ability to shape the intestinal microflora and to suppress inflammatory cytokine production.

Safety Evaluation of Bifidobacterium lactis BL-99 and Lacticaseibacillus paracasei K56 and ET-22 in vitro and in vivo.

Probiotics have been reported to play a major role in maintaining the balance of microbiota in host. Consumption of food with probiotics has increased with consumer concerns regarding healthy diets and wellness. Correspondingly, safety evaluation of probiotics for human consumption has become increasingly important in food industry. Herein, we aimed to test the safety of Bifidobacterium lactis BL-99 and Lacticaseibacillus paracasei K56 and ET-22 strains in vitro and in vivo. In results, these strains were found to be negative for mucin degradation and platelet aggregation test. Additionally, the three strains were susceptible to eight antibiotics. In accordance with bacterial reversion mutation (Ames) assay, the tested strains had no genetic mutagenicity. Finally, it was confirmed that there were no dose-dependent mortality and toxicity throughout multidose oral toxicity tests in rats. Our findings demonstrated that B. lactis BL-99 and L. paracasei K56 and ET-22 can achieve the generally recognized as safe (GRAS) status as probiotics in the future.

Design and evaluation of novel primers for the detection of genes encoding diverse enzymes of methylotrophy and autotrophy.

The phylogenetic significance of the diversity of key enzymes of methylotrophic and autotrophic metabolism is discussed. Primers for these key enzymes were designed using gene sequences encoding methanol dehydrogenase (mxaF; using subsets from database sequences for 22 Bacteria), hydroxypyruvate reductase (hpr; 36 sequences), methylamine dehydrogenase (mauA; 12 sequences), methanesulfonate monooxygenase (msmA; four sequences), and the ccbL and cbbM genes of ribulose bisphosphate carboxylase (26 and 23 sequences). These were effective in amplifying the correct gene products for the target genes in reference organisms and in test organisms not previously shown to contain the genes, as well as in some methylotrophic Proteobacteria isolated from the human mouth. The availability of the new primers increases the probability of detecting diverse examples of the genes encoding these key enzymes both in natural populations and in isolated bacterial strains.

Facultative methylotrophs from the human oral cavity and methylotrophy in strains of Gordonia, Leifsonia, and Microbacterium.

We show that bacteria with methylotrophic potential are ubiquitous in the human mouth microbiota. Numerous strains of Actinobacteria (Brevibacterium, Gordonia, Leifsonia, Microbacterium, Micrococcus, Rhodococcus) and Proteobacteria (Achromobacter, Klebsiella, Methylobacterium, Pseudomonas, Ralstonia) were isolated, and one strain of each of the eleven genera was studied in detail. These strains expressed enzymes associated with methylotrophic metabolism (methanol, methylamine, and formate dehydrogenases), and the assimilation of one-carbon compounds by the serine pathway (hydroxypyruvate reductase). Methylotrophic growth of the strains was enhanced by the addition of glass beads to cultures, suggesting that they may naturally occur in biofilms in the mouth. This is the first report of Gordonia, Leifsonia, and Rhodococcus being present in the mouth and of the unequivocal demonstration for the first time of the methylotrophic potential of strains of Gordonia, Leifsonia, and Microbacterium.