Restoration of gut dysbiosis through Clostridium butyricum and magnesium possibly balance blood glucose levels: an experimental study.

Diabetes mellitus (DM) is a chronic metabolic disorder characterized by an elevated level of blood glucose due to the absence of insulin secretion, ineffectiveness, or lack of uptake of secreted insulin in the body. The improperly diagnosed and poorly managed DM can cause severe damage to organs in the body like the nerves, eyes, heart, and kidneys. This study was aimed at investigating the effect of Clostridium butyricum (probiotic) with magnesium supplementation to evaluate the effect on gut microbial dysbiosis and blood glucose levels. In the laboratory, 6-8 weeks old 24 male albino rats weighing 200-250 g were given free access to water and food. Diabetes was induced using streptozotocin (60 mg/kg) in overnight fasted rats. Diabetic rats were randomly divided into four groups (n = 6, 6 replicates in each group). Metformin (100 mg/kg/day) with a standard basal diet was provided to control group (G0), Clostridium butyricum (1.5 × 105 CFU/day) with standard basal diet was provided to treatment group (G1), magnesium (500 mg/kg/day) was provided to group (G2). Clostridium butyricum (1.5 × 105 CFU/day) and magnesium (300 mg/kg/day) in combination with a standard basal diet was provided to group (G3). Blood Glucose, Magnesium blood test and microbial assay were done. Random blood glucose levels were monitored twice a week for 21 days and were represented as mean of each week. The results conclude that Clostridium butyricum (1.5 × 105 CFU) is very effective in balancing random blood glucose levels from 206.6 ± 67.7 to 85.1 ± 3.8 (p = 0.006) compared to other groups (p > 0.005). The results of stool analysis showed that Clostridium butyricum as probiotic restores microbial dysbiosis as evident by the 105 CFU Clostridium butyricum load in G1, which was higher than G0, G2 and G3 which were 103 and 104 CFU respectively. The findings of this study conclude that Clostridium butyricum supplementation improved blood glucose levels and intestinal bacterial load in type II diabetes mellitus.

Novel mechanism of Clostridium butyricum alleviated coprophagy prevention-induced intestinal inflammation in rabbit.

As bacteria synthesize nutrients primarily in the cecum, coprophagy is indispensable for supplying rabbits with essential nutrients. Recent research has demonstrated its pivotal role in maintaining intestinal microbiota homeostasis and immune regulation in rabbits, although the specific mechanism remains unknown. Here, we used coprophagy prevention (CP) to investigate the effects of coprophagy on the cecum homeostasis and microbiota in New Zealand white rabbits. Furthermore, whether supplementation of Clostridium butyricum (C. butyricum) may alleviate the cecum inflammation and apoptosis caused by CP was also explored. Four groups were randomly assigned: control (Con), sham-coprophagy prevention (SCP), coprophagy prevention (CP), and CP and C. butyricum addition (CPCB). Compared to Con and SCP, CP augmented cecum inflammation and apoptosis, as well as bacterial adhesion to the cecal epithelial mucosa, while decreasing the expression of tight junction proteins (ZO-1, occluding, and claudin-1). The relative abundance of short-chain fatty acids (SCFAs)-producing bacteria was significantly decreased in the CP group. Inversely, there was an increase in the Firmicutes/Bacteroidetes ratio and the relative abundance of Christensenellaceae_R-7_group. Additionally, CP increased the levels of Flagellin, IFN-γ, TNF-a, and IL-1ß in cecum contents and promoted the expression of TLR5/MyD88/NF-κB pathway in cecum tissues. However, the CPCB group showed significant improvements in all parameters compared to the CP group. Dietary C. butyricum supplementation significantly increased the production of SCFAs, particularly butyric acid, triggering anti-inflammatory, tissue repairing, and barrier-protective responses. Notably, CPCB effectively mitigated CP-induced apoptosis and inflammation. In summary, CP disrupts the cecum epithelial barrier and induces inflammation in New Zealand white rabbits, but these effects can be alleviated by C. butyricum supplementation. This process appears to be largely associated with the TLR5/MyD88/NF-κB signaling pathway.

The biotherapeutic Clostridium butyricum MIYAIRI 588 strain potentiates enterotropism of Rorγt+Treg and PD-1 blockade efficacy.

Immune checkpoint inhibitors (ICI) have been positioned as a standard of care for patients with advanced non-small-cell lung carcinomas (NSCLC). A pilot clinical trial has reflected optimistic association between supplementation with Clostridium butyricum MIYAIRI 588 (CBM588) and ICI efficacy in NSCLC. However, it remains to be established whether this biotherapeutic strain may be sufficient to heighten the immunogenicity of the tumor draining lymph nodes to overcome resistance to ICI. Herein, we report that supplementation with CBM588 led to an improved responsiveness to antibody targeting programmed cell death protein 1 (aPD-1). This was statistically associated with a significant decrease in α-diversity of gut microbiota from CBM588-treated mice upon PD-1 blockade. At the level of the tumor-draining lymph node, such combination of treatment significantly lowered the frequency of microbiota-modulated subset of regulatory T cells that express Retinoic Orphan Receptor gamma t (Rorγt+ Treg). Specifically, this strongly immunosuppressive was negatively correlated with the abundance of bacteria that belong to the family of Ruminococcaceae. Accordingly, the colonic expression of both indoleamine 2,3-Dioxygenase 1 (IDO-1) and interleukin-10 (IL-10) were heightened in mice with greater PD-1 blockade efficacy. The CBM588-induced ability to secrete Interleukin-10 of lamina propria mononuclear cells was heightened in tumor bearers when compared with cancer-free mice. Conversely, blockade of interleukin-10 signaling preferentially enhanced the capacity of CD8+ T cells to secrete Interferon gamma when being cocultured with CBM588-primed lamina propria mononuclear cells of tumor-bearing mice. Our results demonstrate that CBM588-centered intervention can adequately improve intestinal homeostasis and efficiently overcome resistance to PD-1 blockade in mice.

Combination of thalidomide and Clostridium butyricum relieves chemotherapy-induced nausea and vomiting via gut microbiota and vagus nerve activity modulation.

Nausea and vomiting (CINV) are distressful and widespread side effects of chemotherapy, and additional efficient regimens to alleviate CINV are urgently needed. In the present study, colorectal cancer (CRC) mice model induced by Azoxymethane (AOM)/Dextran Sodium Sulfate (DSS) was employed to evaluate the cancer suppression and CINV amelioration effect of the combination of thalidomide (THD) and Clostridium butyricum. Our results suggested that the combination of THD and C. butyricum abundantly enhanced the anticancer effect of cisplatin via activating the caspase-3 apoptosis pathway, and also ameliorated CINV via inhibiting the neurotransmitter (e.g., 5-HT and tachykinin 1) and its receptor (e.g., 5-HT3R and NK-1R) in brain and colon. Additionally, the combination of THD and C. butyricum reversed the gut dysbacteriosis in CRC mice by increasing the abundance of Clostridium, Lactobacillus, Bifidobacterium, and Ruminococcus at the genus level, and also led to increased expression of occludin and Trek1 in the colon, while decreased expression of TLR4, MyD88, NF-κB, and HDAC1, as well as the mRNA level of IL-6, IL-1ß, and TNF-α. In all, these results suggest that the combination of THD and C. butyricum had good efficacy in enhancing cancer treatments and ameliorating CINV, which thus provides a more effective strategy for the treatment of CRC.

Supplemental Clostridium butyricum modulates skeletal muscle development and meat quality by shaping the gut microbiota of lambs.

This study evaluated the contributions of Clostridium butyricum on skeletal muscle development, gastrointestinal flora and meat quality of lambs. Eighteen Dorper (♂) × Small Tailed Han sheep (♀) crossed ewe lambs of similar weight (27.43 ± 1.94 kg; age, 88 ± 5 days) were divided into two dietary treatments. The control group was fed the basal diet (C group), and the probiotic group was supplemented with C. butyricum on the basis of the C group (2.5 × 108 cfu/g, 5 g/day/lamb; P group) for 90 d. The results showed that dietary C. butyricum elevated growth performance, muscle mass, muscle fiber diameter and cross-sectional area, and decreased the shear force value of meat (P < 0.05). Moreover, C. butyricum supplementation accelerated protein synthesis by regulating the gene expression of IGF-1/Akt/mTOR pathway. We identified 54 differentially expressed proteins that regulated skeletal muscle development through different mechanisms by quantitative proteomics. These proteins were associated with ubiquitin-protease, apoptosis, muscle structure, energy metabolism, heat shock, and oxidative stress. The metagenomics sequencing results showed that Petrimonas at the genus level and Prevotella brevis at the species level in the rumen, while Lachnoclostridium, Alloprevotella and Prevotella at the genus level in the feces, were significantly enriched in the P group. Also, butyric acid and valeric acid levels were elevated in both rumen and feces of the P group. Overall, our results support the idea that C. butyricum could change gastrointestinal flora, and affect skeletal muscle development and meat quality of lambs by modulating gut-muscle axis.

Probiotics and Immunostimulant modulate intestinal flora diversity in Reeves' Turtle (Mauremys reevesii) and effects of Clostridium butyricum on its spleen transcriptome.

In this study, we investigated the effects of Clostridium butyricum (group A), Bacillus subtilis (group B), and the immune enhancer algal ß-1,3 glucan (group C) on the intestinal flora of Reeves' turtle Mauremys reevesii and the effects of C. butyricum on the transcriptome of M. reevesii splenic immune tissues. Reeve' turtles were assigned to four groups, each containing three replicates from 18 samples. Juvenile turtles with an initial weight of 106.35 ± 0.03 g were fed a basic diet containing no probiotics (group D), or a basic diet containing C. butyricum TF20201120, B.subtilis, or algal ß-1,3 glucan supplement, respectively. After the turtles had been fed for 60, 90, and 120 d of the experimental period, high-throughput sequencing of the 16S rRNA gene revealed no significant difference in alpha diversity among the four groups at 60 days of feeding (P > 0.05), and at 90 days, the alpha diversity in group A was significantly different (P < 0.05), with an increase of 26.62% in the Shannon index and a decrease of 83.33% in the Simpson index; at 120 d, the alpha diversity (Shannon index) showed a decreasing trend in order for groups A, B, and C, At the phylum level, the abundance of Bacteroidetes, Proteobacteria, and Fusobacteria in group A increased significantly with increasing feeding time (P < 0.05), At the genus level, the abundance of Ruminococcaceae and Anaerotruncus in group A increased significantly compared with that in the other three groups (P < 0.05). Transcriptome analysis showed that 384 genes were differentially expressed in the spleen of M. reevesii, 195 genes were upregulated and 189 genes were downregulated, and C. butyricum TF201120 regulated the hematopoietic cell lineage signaling pathway in the spleen of M. reevesii (P < 0.05). The regulation of several identified immune-related genes was confirmed by qPCR. These results showed that C. butyricum, B. subtilis, and the immune enhancer algal ß-1,3 glucan can improve the intestinal flora of M. reevesii, with C. butyricum TF20201120 being the most effective and significantly enhancing the immunity of M. reevesii.

Clostridium butyricum alleviates LPS-induced acute immune stress in goats by regulating bacterial communities and blood metabolites.

The mitigation and prevention of acute immune stress are essential for livestock production. Clostridium butyricum (C. butyricum) has shown positive effects in stabilizing intestinal microbiota disorders, improving immune function and inhibiting disease development, but its effects on ruminants are unclear. Therefore, the current trial hypothesized that C. butyricum could improve goats' immune function and antioxidant capacity by regulating bacterial communities and blood metabolism and effectively alleviating the acute immune stress induced by Lipopolysaccharides (LPS). Sixteen healthy goats were fed C. butyricum for 70 days, and the goats were challenged with LPS on day 71. Blood and feces were collected at 0 h and 6 h after the challenge to evaluate the effects of C. butyricum on their intestinal microbiota, immune function, antioxidant function, and plasma metabolites. The results showed that C. butyricum had no significant effect on plasma biochemical parameters at the beginning of the LPS challenge. However, supplementation with C. butyricum increased plasma levels of IgA, IgG, T-SOD, and T-AOC (P < 0.05), but TNF-α, IL-6, and MDA were decreased (P < 0.05). In contrast, IL-10 showed an increasing trend (P < 0.10). Rectal microbiota analysis showed that C. butyricum significantly increased the relative abundance of Epsilonbacteraeota at the phylum level of goats; at the genus level, the relative abundances of Campylobacter and Anaerorhabdus]_furcosa_group were also significantly increased (P < 0.05). Christensenellaceae_R-7_group as the dominant microbiota also showed a significant increase in their abundance values, while Clostridium and Lachnospiraceae_UCG-001 were significantly lower (P < 0.05). When the LPS challenge continued up to 6 h, dietary supplementation with C. butyricum still resulted in significantly higher plasma concentrations of IgA, IL-10, and T-SOD in goats than in the control group, reducing TNF-α levels (P < 0.05). In addition, plasma levels of T-CHOL and LDL were significantly reduced, and the expression of d-proline was significantly upregulated according to metabolomic analysis (P < 0.05). In conclusion, dietary supplementation with C. butyricum helped optimize the expression of bacterial communities and plasma metabolites to enhance the ability of goats to alleviate acute immune stress.

The protective effects of dietary Clostridium butyricum supplementation on hepatic ischemia reperfusion injury in rats.

PURPOSE: This study investigated the effects of oral administration of Clostridium butyricum (C. butyricum) on inflammation, oxidative stress, and gut flora in rats with hepatic ischemia reperfusion injury (HIRI). METHODS: The rats from C. butyricum group were given C. butyricum for 5 days. Then, hepatic ischemia for 30 min and reperfusion for 6 h were performed in all the rats. After the animals were sacrificed, alanine transaminase (ALT), aspartate aminotransferase (AST), lipopolysaccharide (LPS) in serum, short-chain fatty acids (SCFAs), and gut microbiota composition in feces, and malondialdehyde (MDA), glutathione (GSH), tumor necrosis factor-α (TNF-α), interleukin 6 (IL-6), Toll-like receptor 4 (TLR4), nuclear factor-kappa Bp65 (NF-κBp65) and histological analysis in the liver were performed. RESULTS: The rats given C. butyricum showed decreased ALT, AST, LPS, and MDA; improved GSH and histological damage; changes in SCFAs; declined TNF-α, IL-6, TLR4, and pNF-κBp65/NF-κBp65; and changes in the gut microbial composition, which decreased the Firmicutes/Bacteroidetes ratio and increased the relative abundance (RA) of probiotics. CONCLUSIONS: C. butyricum supplementation protected against HIRI by regulating gut microbial composition, which contributed to the decreased LPS and attenuation of inflammation and oxidative stress. These indicate C. butyricum may be a potent clinical preoperative dietary supplement for HIRI.

Protease or Clostridium butyricum addition to a low-protein diet improves broiler growth performance.

Low-protein (LP) feeds are used in the poultry industry to combat the increasing consumption of protein resources and reduce environmental pollution caused by excessive nitrogen excretion. Dietary supplementation of protease or Clostridium butyricum increases the growth performance of broilers; however, it is unclear whether they counteract the negative effects of LP diets. The effects of protease and C. butyricum on growth performance, intestinal morphology, anti-oxidant capacity, anti-inflammatory response, and microbial community of broilers have not been studied extensively. Here, 450 healthy 1-day-old Cobb500 broilers were allocated to five groups, according to different diets: basal diet (Control); LP diet (LP; 2% less crude protein than the control); LP diet + 200 g/t HuPro protease (LPH); LP diet + 1.0 × 109 CFU/t C. butyricum (LPC); and basal diet + 200 g/t oxytetracycline (Antibiotic). Supplementing both C. butyricum and protease improved the growth performance of broilers. The supplementation of HuPro protease under low-protein conditions could achieve a breeding effect similar to that of the positive control (Antibiotic). Supplementing C. butyricum could maintain intestinal barrier function, alleviate the inflammatory response, and increase ileal and cecal short-chain fatty acid concentrations. Both C. butyricum and protease altered the bacterial diversity in the cecum, increased Bacteroidetes abundance, and resulted in higher abundance of Rikenellaceae RC9 gut spp. and lower abundance of Alistipes spp. in broilers. This study demonstrates the positive effects of proteases and C. butyricum on broilers and serves as a reference for the selection of appropriate supplementation for broilers in the poultry industry. KEY POINTS: • Low-protein diet had a negative effect on growth performance of broilers. • Protease significantly reduced feed conversion rate. • Clostridium butyricum had positive effects on broilers.

Protective Effect of Clostridium butyricum on Escherichia coli-Induced Endometritis in Mice via Ameliorating Endometrial Barrier and Inhibiting Inflammatory Response.

Endometritis is a common reproductive disease occurs both in human and animals. Clostridium butyricum is a Gram-positive anaerobic bacterium that can ferment various carbohydrates into butyric acid. In this study, we investigated the effects of C. butyricum on Escherichia coli-induced endometritis and clarified the underlying mechanism. We first verified the protective effect of C. butyricum in vivo by establishing a mouse model of E. coli-induced endometritis. It was determined that C. butyricum pretreatment significantly reversed E. coli-induced uterine histopathological changes. Meanwhile, C. butyricum pretreatment significantly decreased the production of pro-inflammatory mediators and the levels of myeloperoxidase (MPO) and malondialdehyde (MDA). We found that C. butyricum could inhibit TLR4-mediated phosphorylation of NF-κB and the activity of histone deacetylase (HDAC). Furthermore, C. butyricum significantly increased the expression of the tight junction proteins (TJPs) ZO-1, claudin-3, and occludin. Additionally, treatment with C. butyricum culture supernatant dramatically suppressed the degree of inflammation in the uterus, and inactivated C. butyricum did not exert a protective effect. We subsequently investigated butyrate levels in both the uterus and blood and observed a marked augment in the C. butyricum treatment group. Collectively, our data suggest that C. butyricum maintains epithelial barrier function and suppresses inflammatory response during E. coli-induced endometritis and that the protective effect of C. butyricum may be related to the production of butyrate. IMPORTANCE Endometritis is a common reproductive disease both in human and animals. It impairs female fertility by disrupting endometrial function. Antibiotics are widely used to treat endometritis in clinical practice, but the misuse of antibiotics often leads to antibiotic resistance. Therefore, there is an urgent need for new therapeutic agents to treat bacterial endometritis and overcome bacterial resistance. In this study, we found that C. butyricum could protect from E. coli-induced endometritis.

Amelioration of ammonia-induced intestinal oxidative stress by dietary Clostridium butyricum in giant freshwater prawn (Macrobrachium rosenbergii).

The use of Clostridium butyricum in crustacean aquaculture for anti-abiotic stress is yet unknown. Feeds were formulated containing 0, 125, 250, 500, and 1000 mg/kg Clostridium butyricum (2 × 107 CFU/g), respectively. The giant freshwater prawns (Macrobrachium rosenbergii) were fed for 8 weeks in triplicate. The results showed that C. butyricum-supplemented groups improved growth performance significantly with the optimum level at 610 mg/kg. Ammonia stress reduced hemolymph glucose, total protein, total cholesterol, and triglyceride concentrations while dietary C. butyricum significantly increased hemolymph glucose and total protein levels after the ammonia challenge. Ammonia stress increased inducible nitric oxide synthase (iNOS) and nitric oxide (NO) levels, and the treatments supplemented with C. butyricum had considerably enhanced levels of iNOS and NO after stress. Treatment with C. butyricum increased the level of superoxide dismutase (SOD), and decreased the level of malondialdehyde (MDA) and superoxide anion, with the 125 mg/kg treated groups having the extreme value. Furthermore, C. butyricum-treated groups reduced the expression of HSPs after ammonia stress while the ammonia stress induced the expression of HSP60, HSP70, and HSP90. Dietary C. butyricum elevated the expression of peroxiredoxin-5 and toll in response to ammonia stress. The results indicate that dietary supplementation with 125-500 mg/kg of C. butyricum (2 × 107 CFU/g) improved biochemical and antioxidant features as well as intestinal immunity of M. rosenbergii under ammonia challenge by activating the toll signal pathway.

Role of Clostridium butyricum, Bacillus subtilis, and algae-sourced ß-1,3 glucan on health in grass turtle.

This study investigated the effects of two probiotics namely Clostridium butyricum and Bacillus subtilis, and one prebiotic known as algae-sourced ß-1,3 glucan, on the overall performances of grass turtles (Chinemys reevesii) juveniles. Growth performance, immune responses, enzymatic antioxidant activities, intestinal histomorphology, and disease resistance against the challenge with Aeromonas veronii were assessed. Two hundred and sixteen (216) juvenile turtles with an average initial weight of 106.35 ± 0.03 g were divided into four groups, each containing three replicates with 18 turtles per each replicate, which were fed a basic diet (control group, GD) and a basal diet supplemented with C. butyricum 1.0 × 108 CFU per kg (GA group), or with B. subtilis 1.0 × 108 CFU per kg (GB group) and with algal-sourced ß-1,3-glucan 50 mg per kg (GC group), respectively. After the turtles had been fed for 60 d, 90 d, and 120 d of the experimental period, the growth performance and survival rate (SR), intestinal digestive enzyme, hepatic and intestinal antioxidant capacity, serum biochemical indexes, and immune performance were measured. The results showed that the weight gain rate and SR were significantly enhanced (P < 0.05) after fed probiotics and algae-sourced ß-1,3-glucan in all test times;The pepsin, amylase, acid phosphatase, total antioxidant capacity, triglyceride, alkaline phosphatase, urea nitrogen, cholesterol, total protein, IgA, IgG, IgM at 120 d were significantly enhanced (P<0.05) after fed C. butyricum. The intestinal villi heights, widths, and the thickness of the muscle layer were significantly higher in groups GA, GB, and GC than those reared within the GD control group (P < 0.05). After injecting the challenge by A. veronii the survival rate of grass turtles in the GA group (75%) was significantly higher than the other three groups (P<0.05), while there was no significant difference between the GB and GC groups compared with the control GD group, respectively (P>0.05). Overall, these results indicated that dietary supplementation with probiotics or algae-sourced ß-1,3 glucan, exhibited positive effects on C. reevesii. In particular, C. butyricum, showed the greatest improvements relating to growth, immune response, antioxidant activity, intestinal health, and disease resistance.

Clostridium butyricum, a butyrate-producing potential probiotic, alleviates experimental colitis through epidermal growth factor receptor activation.

Inflammatory bowel disease (IBD) is an idiopathic inflammatory bowel disease. Modulation of gut microbiota with dietary and nutritional targets is a feasible strategy for the prevention and treatment of IBD. In this study, we focused on Clostridium butyricum Prazmowski (CB), a butyrate-producing potential probiotic. We found that CB feeding decreased the disease activity index, colon inflammation/injury score and cell apoptosis in an experimental colitis mouse model, as well as elevated the level of SCFAs in cecal feces. CB could also balance the inflammatory cytokines, protect tight junctions, and increase the number of goblet cells and MUC2 production in mice, accompanied by EGFR signaling activation triggered by heparin-binding epidermal growth factor (HB-EGF) and amphiregulin (AREG). From the perspective of mechanism, the CB supernatant (CBS) stimulated EGFR activation in colon epithelial cell lines in concentration-dependent and time-dependent manners. CBS reduced the damage of tight junctions induced by H2O2, and inhibition of EGFR could suppress the protective effect of CBS. In conclusion, CB could protect the gut barrier and alleviate experimental colitis through the transactivation of EGFR signaling in intestinal epithelial cells induced by ligands (HB-EGF and AREG). This study identified the potential efficacy of CB as a preventive strategy for IBD and showed the broad prospect of CB as a food supplement.

Clostridium butyricum and Its Derived Extracellular Vesicles Modulate Gut Homeostasis and Ameliorate Acute Experimental Colitis.

Microbiological treatments are expected to have a role in the future management of inflammatory bowel disease (IBD). Clostridium butyricum (C. butyricum) is a probiotic microorganism that exhibits beneficial effects on various disease conditions. Although many studies have revealed that C. butyricum provides protective effects in mice with colitis, the way C. butyricum establishes beneficial results in the host remains unclear. In this study, we investigated the mechanisms by which C. butyricum modifies the gut microbiota, produces bacterial metabolites that may be involved, and, specifically, how microbial extracellular vesicles (EVs) positively influence IBD, using a dextran sulfate sodium (DSS)-induced colitis murine model in mice. First, we showed that C. butyricum provides a protective effect against colitis, as evidenced by the prevention of body weight loss, a reduction in the disease activity index (DAI) score, a shortened colon length, decreased histology score, and an improved gut barrier function, accompanied by reduced levels of pathogenic bacteria, including Escherichia/Shigella, and an increased relative abundance of butyrate-producing Clostridium sensu stricto-1 and Butyricicoccus. Second, we also confirmed that the gut microbiota and metabolites produced by C. butyricum played key roles in the attenuation of DSS-induced experimental colitis, as supported by the profound alleviation of colitis effects following fecal transplantation or fecal filtrate insertion supplied from C. butyricum-treated mice. Finally, C. butyricum-derived EVs protected the gut barrier function, improved gut microbiota homeostasis in ulcerative colitis, and contributed to overall colitis alleviation. IMPORTANCE This study indicated that C. butyricum provided a prevention effect against colitis mice, which involved protection of the intestinal barrier and positively regulating gut microbiota. Furthermore, we confirmed that the gut microbiota and metabolites that were induced by C. butyricum also contributed to the attenuation of DSS-induced colitis. Importantly, C. butyricum-derived EVs showed an effective impact in alleviating colitis.

Co-interventions with Clostridium butyricum and soluble dietary fiber targeting the gut microbiota improve MAFLD via the Acly/Nrf2/NF-κB signaling pathway.

Purpose: The pathogenesis of metabolic associated fatty liver disease (MAFLD) is complex. Lipid metabolic disorder, chronic inflammation, and oxidative stress are the core events for MAFLD. Dietary intervention is an important treatment strategy for preventing the onset and progression of MAFLD. Clostridium butyricum (CB) and soluble dietary fiber (SDF) are often considered beneficial for health. We explored how two microbiota-targeted interventions (SDF and CB) influence the hepatic immune system, oxidative stress, and lipid metabolism in MAFLD mice. Methods: To explore the role of SDF and CB in MAFLD, we generated MAFLD mouse models by feeding C57BL/6 mice with a high-fat diet (HFD). After 8 weeks of intervention, we measured immune cell function, lipid metabolism, and oxidative stress levels in the livers of mice. Results: Single intervention with SDF or CB was not effective in improving MAFLD; however, co-interventions with SDF and CB increased microbiota diversity and decreased inflammation, oxidative stress, and lipid synthesis. Moreover, we determined that co-intervention with SDF and CB mediated fatty acid oxidation by activating the Acly/Nrf2/NF-κB signaling pathway. Most importantly, co-intervention exerted anti-inflammatory effects by inhibiting the differentiation of macrophages into pro-inflammatory M1 macrophages. Conclusion: This study show that co-intervention with SDF and CB can improve MAFLD, and co-intervention with  SDF and CB are suggested to be potential gut microbiota modulators and therapeutic substances for MAFLD.

Clostridium butyricum Alleviates Enterotoxigenic Escherichia coli K88-Induced Oxidative Damage Through Regulating the p62-Keap1-Nrf2 Signaling Pathway and Remodeling the Cecal Microbial Community.

Clostridium butyricum (CB) can enhance antioxidant capacity and alleviate oxidative damage, but the molecular mechanism by which this occurs remains unclear. This study used enterotoxigenic Escherichia coli (ETEC) K88 as a pathogenic model, and the p62-Keap1-Nrf2 signaling pathway and intestinal microbiota as the starting point to explore the mechanism through which CB alleviates oxidative damage. After pretreatment with CB for 15 d, mice were challenged with ETEC K88 for 24 h. The results suggest that CB pretreatment can dramatically reduce crypt depth (CD) and significantly increase villus height (VH) and VH/CD in the jejunum of ETEC K88-infected mice and relieve morphological lesions of the liver and jejunum. Additionally, compared with ETEC-infected group, pretreatment with 4.4×106 CFU/mL CB can significantly reduce malondialdehyde (MDA) level and dramatically increase superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) levels in the serum. This pretreatment can also greatly increase the mRNA expression levels of tight junction proteins and genes related to the p62-Keap1-Nrf2 signaling pathway in the liver and jejunum in ETEC K88-infected mice. Meanwhile, 16S rDNA amplicon sequencing revealed that Clostridium disporicum was significantly enriched after ETEC K88 challenge relative to the control group, while Lactobacillus was significantly enriched after 4.4×106 CFU/mL CB treatment. Furthermore, 4.4×106 CFU/mL CB pretreatment increased the short-chain fatty acid (SCFA) contents in the cecum of ETEC K88-infected mice. Moreover, we found that Lachnoclostridium, Roseburia, Lactobacillus, Terrisporobacter, Akkermansia, and Bacteroides are closely related to SCFA contents and oxidative indicators. Taken together, 4.4×106 CFU/mL CB pretreatment can alleviate ETEC K88-induced oxidative damage through activating the p62-Keap1-Nrf2 signaling pathway and remodeling the cecal microbiota community in mice.

Clostridium butyricum enhances colonization resistance against Clostridioides difficile by metabolic and immune modulation.

Clostridioides difficile infection (CDI) represents the leading cause of nosocomial diarrhea worldwide and is associated with gut dysbiosis and intestinal damage. Clostridium butyricum MIYAIRI 588 (CBM 588) contributes significantly to reduce epithelial damage. However, the impacts of CBM 588 on antibacterial therapy for CDI are not clear. Here we show that CBM 588 enhanced the antibacterial activity of fidaxomicin against C. difficile and negatively modulated gut succinate levels to prevent C. difficile proliferation and downregulate tumor necrosis factor-α (TNF-α) producing macrophages in the colon lumina propria (cLP), resulting in a significant decrease in colon epithelial damage. Additionally, CBM 588 upregulated T cell-dependent pathogen specific immunoglobulin A (IgA) via interleukin (IL)-17A producing CD4+ cells and plasma B cells in the cLP, and Th17 cells in the cLP enhanced the gut epithelial barrier function. IL-17A and succinic acid modulations with CBM 588 enhance gut colonization resistance to C. difficile and protect the colon tissue from CDI.

Combination of supercritical CO2 and high-power ultrasound for the inactivation of fungal and bacterial spores in lipid emulsions.

For the first time, this study addresses the intensification of supercritical carbon dioxide (SC-CO2) treatments using high-power ultrasound (HPU) for the inactivation of fungal (Aspergillus niger) and bacterial (Clostridium butyricum) spores in oil-in-water emulsions. The inactivation kinetics were analyzed at different pressures (100, 350 and 550 bar) and temperatures (50, 60, 70, 80, 85 °C), depending on the microorganism, and compared to the conventional thermal treatment. The inactivation kinetics were satisfactorily described using the Weibull model. Experimental results showed that SC-CO2 enhanced the inactivation level of both spores when compared to thermal treatments. Bacterial spores (C.butyricum) were found to be more resistant to SC-CO2 + HPU, than fungal (A.niger) ones, as also observed in the thermal and SC-CO2 treatments. The application of HPU intensified the SC-CO2 inactivation of C.butyricum spores, e.g. shortening the total inactivation time from 10 to 3 min at 85 °C. However, HPU did not affect the SC-CO2 inactivation of A.niger spores. The study into the effect of a combined SC-CO2 + HPU treatment has to be necessarily extended to other fungal and bacterial spores, and future studies should elucidate the impact of HPU application on the emulsion's stability.

Effects of Clostridium butyricum on growth performance, metabonomics and intestinal microbial differences of weaned piglets.

BACKGROUND: Weaning stress of piglets causes a huge economic loss to the pig industry. Balance and stability of the intestinal microenvironment is an effective way to reduce the occurance of stress during the weaning process. Clostridium butyricum, as a new microecological preparation, is resistant to high temperature, acid, bile salts and some antibiotics. The aim of present study is to investigate the effects of C. butyricum on the intestinal microbiota and their metabolites in weaned piglets. RESULTS: There was no statistical significance in the growth performance and the incidence of diarrhoea among the weaned piglets treated with C. butyricum during 0-21 days experimental period. Analysis of 16S rRNA gene sequencing results showed that the operational taxonomic units (OTUs), abundance-based coverage estimator (ACE) and Chao index of the CB group were found to be significantly increased compared with the NC group (P < 0.05). Bacteroidetes, Firmicutes and Tenericutes were the predominant bacterial phyla in the weaned piglets. A marked increase in the relative abundance of Megasphaera, Ruminococcaceae_NK4A214_group and Prevotellaceae_UCG-003, along with a decreased relative abundance of Ruminococcaceae_UCG-005 was observed in the CB group, when compared with the NC group (P < 0.05). With the addition of C. butyricum, a total of twenty-two significantly altered metabolites were obtained in the feces of piglets. The integrated pathway analysis by MetaboAnalyst indicated that arginine and proline metabolism; valine, leucine and isoleucine biosynthesis; and phenylalanine metabolism were the main three altered pathways, based on the topology. Furthermore, Spearman's analysis revealed some altered gut microbiota genus such as Oscillospira, Ruminococcaceae_NK4A214_group, Megasphaera, Ruminococcaceae_UCG-005, Prevotella_2, Ruminococcaceae_UCG-002, Rikenellaceae_RC9_gut_group and Prevotellaceae_UCG-003 were associated with the alterations in the fecal metabolites (P < 0.05), indicating that C. butyricum presented a potential protective impact through gut microbiota. The intestinal metabolites changed by C. butyricum mainly involved the variation of citrulline, dicarboxylic acids, branched-chain amino acid and tryptophan metabolic pathways. CONCLUSIONS: Overall, this study strengthens the idea that the dietary C. butyricum treatment can significantly alter the intestinal microbiota and metabolite profiles of the weaned piglets, and C. butyricum can offer potential benefits for the gut health.

Clinical Significance of Probiotics for Children with Idiopathic Nephrotic Syndrome.

We previously reported that a decrease in butyrate-producing bacteria in the gut is a potential cause of regulatory T cell (Treg) abnormalities in children with idiopathic nephrotic syndrome (INS). Therefore, we hypothesized that administration of butyrate-producing bacteria might reduce INS relapse and the need for immunosuppressants in these patients. Twenty patients in remission from INS (median age 5.3 years, 15 boys) were enrolled in the study and assigned to receive either daily oral treatment with a preparation of 3 g Clostridium butyricum or no probiotic treatment. The number of relapses and requirement for immunosuppressive agents were compared between the two groups. In the probiotic treatment group, analyses of the gut microbiota and Treg measurements were also performed. Probiotic-treated patients experienced fewer INS relapses per year compared with non-probiotic-treated patients (p = 0.016). Further, administration of rituximab in the probiotic treatment group was significantly less frequent compared with the non-probiotic-treated group (p = 0.025). In the probiotic treatment group, analyses before and after probiotic treatment revealed the significant increases in the relative abundance of butyrate-producing bacteria (p = 0.017) and blood Treg counts (p = 0.0065). Thus, oral administration of butyrate-producing bacteria during INS remission may reduce the frequency of relapse and the need for immunosuppressive agents.