ABSTRACT
As part of the development of alternative and environmentally friendly control against phytopathogenic fungi, Burkholderia cepacia could be a useful species notably via the generation of hydrolytic enzymes like chitinases, which can act as a biological control agent. Here, a Burkholderia contaminans S614 strain exhibiting chitinase activity was isolated from a soil in southern Tunisia. Then, response surface methodology (RSM) with a central composite design (CCD) was used to assess the impact of five factors (colloidal chitin, magnesium sulfate, dipotassium phosphate, yeast extract, and ammonium sulfate) on chitinase activity. B. contaminans strain 614 growing in the optimized medium showed up to a 3-fold higher chitinase activity. This enzyme was identified as beta-N-acetylhexosaminidase (90.1 kDa) based on its peptide sequences, which showed high similarity to those of Burkholderia lata strain 383. Furthermore, this chitinase significantly inhibited the growth of two phytopathogenic fungi: Botrytis cinerea M5 and Phoma medicaginis Ph8. Interestingly, a crude enzyme from strain S614 was effective in reducing P. medicaginis damage on detached leaves of Medicago truncatula. Overall, our data provide strong arguments for the agricultural and biotechnological potential of strain S614 in the context of developing biocontrol approaches.
ABSTRACT
BACKGROUND: Obesity is a worldwide health problem and a significant risk factor for diabetes and cardiovascular diseases. Gut microbiota (GM) plays an essential role in obesity, and prebiotics such as polyphenols could be one way to improve microbial dysbiosis-induced obesity. OBJECTIVE: This study was designed to assess the effectiveness of grape seed and skin extract (GSSE), and/or orlistat on obese rats fed with high fat diet by targeting GM modulations. The impact of treatments was also studied in non-obese rats. MATERIAL AND METHODS: Rats were rendered obese or kept with a standard diet for three months. Then they were treated either with GSSE or orlistat or with the combined treatment (GSOR) during three months and then sacrificed. Adipose tissues, blood and faeces were collected and analyzed. RESULTS: In obese rats and to a lesser extent in non-obese rats, treatments decreased the weight of various adipose tissues and the serum levels of cholesterol, LDL, triglycerides, lipase, and CRP and increased HDL and adiponectin. GSOR treatment was even more efficient that orlistat. Obese rats had less GM diversity than non-obese rats and orlistat reduced it even more. However, diversity was restored with GSSE and GSOR treatments. Potential pathogenic Streptococcus alactolyticus/gallolyticus species were greatly increased in obese rats and drastically reduced with the treatments, as wells as other potential pathobionts. CONCLUSIONS: GSSE exerts beneficial effects in obese rats and restores, at least partially, the observed dysbiosis. GSOR induced the highest beneficial effect. Moreover, the various treatments could also enhance physiological and GM modifications in non obese rats.
ABSTRACT
SCOPE: Synthetic emulsifiers have recently been shown to promote metabolic syndrome and considerably alter gut microbiota. Yet, data are lacking regarding the effects of natural emulsifiers, such as plant lecithins rich in essential α-linolenic acid (ALA), on gut and metabolic health. METHODS AND RESULTS: For 5 days, male Swiss mice are fed diets containing similar amounts of ALA and 0, 1, 3, or 10% rapeseed lecithin (RL) or 10% soy lecithin (SL). Following an overnight fast, they are force-fed the same oil mixture and euthanized after 90 minutes. The consumption of lecithin significantly increased fecal levels of the Clostridium leptum group (p = 0.0004), regardless of origin or dose, without altering hepatic or intestinal expression of genes of lipid metabolism. 10%-RL increased ALA abundance in plasma triacylglycerols at 90 minutes, reduced cecal bile acid hydrophobicity, and increased their sulfatation, as demonstrated by the increased hepatic RNA expression of Sult2a1 (p = 0.037) and cecal cholic acid-7 sulfate (CA-7S) concentration (p = 0.05) versus 0%-lecithin. CONCLUSION: After only 5 days, nutritional doses of RL and SL modified gut bacteria in mice, by specifically increasing C. leptum group. RL also increased postprandial ALA abundance and induced beneficial modifications of the bile acid profile. ALA-rich lecithins, especially RL, may then appear as promising natural emulsifiers.
Subject(s)
Bile Acids and Salts/analysis , Brassica napus , Gastrointestinal Microbiome/drug effects , Glycine max , Lecithins/administration & dosage , Lipid Metabolism/drug effects , Animals , Bile Acids and Salts/metabolism , Lipids/blood , Male , Mice , Postprandial Period/physiology , alpha-Linolenic Acid/administration & dosageABSTRACT
Given the growing evidence that gut dysfunction, including changes in gut microbiota composition, plays a critical role in the development of inflammation and metabolic diseases, the identification of novel probiotic bacteria with immunometabolic properties has recently attracted more attention. Herein, bacterial strains were first isolated from dairy products and human feces and then screened in vitro for their immunomodulatory activity. Five selected strains were further analyzed in vivo, using a mouse model of diet-induced obesity. C57BL/6 mice were fed a high-fat high-sucrose diet, in combination with 1 of 3 Lactobacillus strains (Lb38, L. plantarum; L79, L. paracasei/casei; Lb102, L. rhamnosus) or Bifidobacterium strains (Bf26, Bf141, 2 different strains of B. animalis ssp. lactis species) administered for 8 wk at 109 colony-forming units/d. Whereas 3 strains showed only modest (Lb38, Bf26) or no (L79) effects, Lb102 and Bf141 reduced diet-induced obesity, visceral fat accretion, and inflammation, concomitant with improvement of glucose tolerance and insulin sensitivity. Further analysis revealed that Lb102 and Bf141 enhanced intestinal integrity markers in association with selective changes in gut microbiota composition. We have thus identified 2 new potential probiotic bacterial strains with immunometabolic properties to alleviate obesity development and associated metabolic disturbances.-Le Barz, M., Daniel, N., Varin, T. V., Naimi, S., Demers-Mathieu, V., Pilon, G., Audy, J., Laurin, E., Roy, D., Urdaci, M. C., St-Gelais, D., Fliss, I, Marette, A. In vivo screening of multiple bacterial strains identifies Lactobacillus rhamnosus Lb102 and Bifidobacterium animalis ssp. lactis Bf141 as probiotics that improve metabolic disorders in a mouse model of obesity.
Subject(s)
Bifidobacterium animalis/physiology , Lacticaseibacillus rhamnosus/physiology , Obesity/diet therapy , Obesity/microbiology , Probiotics/therapeutic use , Adipose Tissue/metabolism , Animals , Chemokines/metabolism , Cytokines/metabolism , Disease Models, Animal , Fatty Acids, Nonesterified/metabolism , Gastrointestinal Microbiome/physiology , Inflammation/metabolism , Male , Mice , Mice, Inbred C57BL , Obesity/metabolism , RNA, Ribosomal, 16S/geneticsABSTRACT
Preventive actions of probiotics as antidiarrheal agents are well documented, but their mechanisms are poorly understood. Two selected probiotics, Bacillus subtilis CU1 and Lactobacillus plantarum CNCM I-4547, were tested in mouse experimental models of diarrhea and the possible mechanisms of action were investigated. Diarrhea was induced in mice by oral castor oil administration or by i.v. injection of lipopolysaccharide (LPS) of Salmonella enteritis. The antidiarrheal drug loperamide was used as control. Fecal water excretion was quantified for 2 h and paracellular permeability and electrical parameters of the colon were assessed in Ussing chambers. The expression of colonic exchangers or channels and of Toll-like receptor 4 (TLR4) was assessed by immunohistochemistry. Prophylactic treatment with B. subtilis CU1 or with L. plantarum CNCM I-4547 reduced LPS-induced diarrhea. The reduction of water excretion was in the same range as those induced by loperamide. In the castor oil model, this effect was only observed with B. subtilis CU1. The two probiotic treatments abolished the increase in paracellular permeability induced by LPS, but not by castor oil. However, only L. plantarum CNCM I-4547 treatment decreased the colonic expression of TLR-4. After B. subtilis CU1, colonic expression of cystic fibrosis transmembrane conductance regulator (CFTR) was reduced and that of Na+/H+ exchanger 3 (NHE3) increased. B. subtilis CU1 may increase the capacity of the colon to absorb excess of water in diarrheic conditions by acting on CFTR and NHE3 expression. The two probiotics strains showed an impact on diarrhea through limitation of water excretion that may involve paracellular permeability or electrolyte transport for L. plantarum CNCM I-4547 and B. subtilis CU1 respectively.
ABSTRACT
Prostaglandin E2 (PGE2 ) plays a critical role in intestinal mucosal tolerance and barrier integrity. Cyclooxygenase-2 (COX-2)-dependent PGE2 production involves mobilisation of arachidonic acid. Lactobacillus rhamnosus GG (LbGG) is one of the most widely used probiotics reported to colonise the colonic mucosa. LbGG contributes to the protection of the small intestine against radiation injury through the repositioning of mucosal COX-2 expressing cells. However, it is unknown if LbGG modulates PGE2 production in the colonic mucosa under homeostasis and the major cellular elements involved in these processes. Colonic epithelial and CD90+ mesenchymal stromal cells, also known as (myo) fibroblasts (CMFs), are abundant innate immune cells in normal colonic mucosa able to produce PGE2 . Herein, we tested the hypothesis that under colonic mucosal homeostasis, LbGG modulates the eicosanoid pathway resulting in increased PGE2 production in both epithelial and stromal cells. Among the five tested human colonic epithelial cell lines, only exposure of Caco-2 to LbGG for 24 hr led to the mobilisation of arachidonic acid with concomitant increase in the components within the leukotriene and COX-2-dependent PGE2 pathways. By contrast, CMFs isolated from the normal human colonic mucosa responded to LbGG with increased expression of COX-2 and PGE2 in the prostaglandin pathway, but not 5-LO in the leukotriene pathway. Oral gavage of C57BL/6 mice for 5 days with LbGG (5 × 108 Colony-Forming Unit (CFU)/dose) increased COX-2 expression in the colonic mucosa. The majority of cells upregulating COX-2 protein expression were located in the colonic lamina propria and colocalised with α-SMA+ cells corresponding to the CMF phenotype. This process was myeloid differentiation factor-88-dependent, because silencing of myeloid differentiation factor-88 expression in CMFs abrogated LbGG-induced upregulation of COX-2 in culture and in vivo. Taken together, our data suggest that LbGG increases release of COX-2-mediated PGE2 , contributing to the maintenance of mucosal homeostasis in the colon and CMFs are among the major contributors to this process.
Subject(s)
Cyclooxygenase 2/metabolism , Dinoprostone/metabolism , Lacticaseibacillus rhamnosus , Myeloid Differentiation Factor 88/metabolism , Probiotics/pharmacology , Administration, Oral , Animals , Arachidonate 5-Lipoxygenase/metabolism , Arachidonic Acid/metabolism , Caco-2 Cells , Colon/cytology , Colon/microbiology , Homeostasis , Humans , Mice, Inbred C57BL , Mice, Transgenic , Myeloid Differentiation Factor 88/genetics , Myofibroblasts/metabolism , Myofibroblasts/microbiology , Probiotics/administration & dosageABSTRACT
Enterococcus species, principally Enterococcus faecium are used as probiotics since a long time with preference in animal applications but safety considerations were updated and also new uses as probiotics can be envisaged. Fifteen Enterococcus strains isolated from different foods were identified and analyzed for virulence factors and antibiotic resistance. Three Enterococcus durans strains were selected to study their immunomodulatory properties on PBMC and Caco2 cells. Two strains presented a profile toward a mild inflammatory Th1 response considering TNF-α/IL-10 and IL-1ß/IL-10 cytokines ratios. The third strain EP1, presented an anti-inflammatory potential and was selected for in vivo studies. In mice, the strain was well tolerated and did not cause any adverse effects. EP1 administration increased the amount of IgA+ cells in mesenteric lymph node (MLN) after 7 days of administration. In fecal samples, the IgA content increased gradually and significantly from day 7 to day 21 in treated group. Additionally, IL-17, IL-6, IL-1ß, IFN-γ, and CXCL1 gene expression significantly decreased on day 21 in Peyer's patches and IL-17 decreased in MLN. Mice treated with the probiotic showed significant lower mRNA levels of pro-inflammatory cytokines and mucins in the ileum at day 7 while their expression was normalized at day 21. Colonic expression of il-1ß, il6, and mucins remain diminished at day 21. Ileum and colon explants from treated mice stimulated in vitro with LPS showed a significant reduction in IL-6 and an increase in IL-10 secretion suggesting an in vivo protective effect of the probiotic treatment against a proinflammatory stimulus. Interestingly, analysis of feces microbiota demonstrated that EP1 administration increase the amount of Faecalibacterium prausnitzii, a butyrate-producing bacteria, which is known for its anti-inflammatory effects. In conclusion, we demonstrated that EP1 strain is a strong sIgA inducer and possess mucosal anti-inflammatory properties. This strain also modulates gut microbiota increasing Faecalibacterium prausnitzii, a functionally important bacterium. Thus, E. durans EP1 is not only a good candidate to increases F. prausnitzii in some cases of dysbiosis but can also be interesting in gut inflammatory disorders therapy.
ABSTRACT
Bacillus subtilis CU1 is a recently described probiotic strain with beneficial effects on immune health in elderly subjects. The following work describes a series of studies supporting the safety of the strain for use as an ingredient in food and supplement preparations. Using a combination of 16S rDNA and gyrB nucleotide analyses, the species was identified as a member of the Bacillus subtilis complex (B. subtilis subsp. spizizenii). Further characterization of the organism at the strain level was achieved using random amplified polymorphic DNA polymerase chain reaction (RAPD PCR) and pulsed field gel electrophoresis (PFGE) analyses. B. subtilis CU1 did not demonstrate antibiotic resistance greater than existing regulatory cutoffs against clinically important antibiotics, did not induce hemolysis or produce surfactant factors, and was absent of toxigenic activity in vitro. Use of B. subtilis CU1 as a probiotic has recently been evaluated in a 16-week randomized, double-blind, placebo-controlled, parallel-arm study, in which 2 × 109 spores per day of B. subtilis CU1 were administered for a total 40 days to healthy elderly subjects (4 consumption periods of 10 days separated by 18-day washouts). This work describes safety related endpoints not previously reported. B. subtilis CU1 was safe and well-tolerated in the clinical subjects without undesirable physiological effects on markers of liver and kidney function, complete blood counts, hemodynamic parameters, and vital signs.
Subject(s)
Bacillus subtilis/physiology , Food Safety , Probiotics/toxicity , Aged , Anti-Bacterial Agents/pharmacology , Bacillus subtilis/drug effects , Bacillus subtilis/genetics , Bacillus subtilis/pathogenicity , Consumer Product Safety , DNA Fingerprinting , DNA Gyrase/genetics , DNA, Bacterial/genetics , Double-Blind Method , Drug Resistance, Bacterial , Electrophoresis, Gel, Pulsed-Field , Female , Hemolysis , Humans , Male , Microbial Sensitivity Tests , Middle Aged , Polymerase Chain Reaction , RNA, Ribosomal, 16S/genetics , Ribotyping , Risk Assessment , Time FactorsABSTRACT
Although the use of probiotics based on Bacillus strains to fight off intestinal pathogens and antibiotic-associated diarrhea is widespread, the mechanisms involved in producing their beneficial effects remain unclear. Here, we studied the ability of compounds secreted by the probiotic Bacillus clausii strain O/C to counteract the cytotoxic effects induced by toxins of two pathogens, Clostridium difficile and Bacillus cereus, by evaluating eukaryotic cell viability and expression of selected genes. Coincubation of C. difficile and B. cereus toxic culture supernatants with the B. clausii supernatant completely prevented the damage induced by toxins in Vero and Caco-2 cells. The hemolytic effect of B. cereus was also avoided by the probiotic supernatant. Moreover, in these cells, the expression of rhoB, encoding a Rho GTPase target for C. difficile toxins, was normalized when C. difficile supernatant was pretreated using the B. clausii supernatant. All of the beneficial effects observed with the probiotic were abolished by the serine protease inhibitor phenylmethylsulfonyl fluoride (PMSF). Suspecting the involvement of a secreted protease in this protective effect, a protease was purified from the B. clausii supernatant and identified as a serine protease (M-protease; GenBank accession number Q99405). Experiments on Vero cells demonstrated the antitoxic activity of the purified protease against pathogen supernatants. This is the first report showing the capacity of a protease secreted by probiotic bacteria to inhibit the cytotoxic effects of toxinogenic C. difficile and B. cereus strains. This extracellular compound could be responsible, at least in part, for the protective effects observed for this human probiotic in antibiotic-associated diarrhea.
Subject(s)
Bacillus cereus/pathogenicity , Bacillus clausii/metabolism , Bacterial Toxins/toxicity , Clostridioides difficile/pathogenicity , Probiotics/pharmacology , Subtilisins/metabolism , Animals , Caco-2 Cells , Cell Line, Tumor , Cell Survival/drug effects , Chlorocebus aethiops , Culture Media, Conditioned/pharmacology , Humans , Intestinal Mucosa/microbiology , Intestinal Mucosa/pathology , Phenylmethylsulfonyl Fluoride/pharmacology , Protease Inhibitors/pharmacology , Subtilisins/antagonists & inhibitors , Vero Cells , rhoB GTP-Binding Protein/metabolismABSTRACT
BACKGROUND: Bacillus probiotics health benefits have been until now quite poorly studied in the elderly population. This study aimed to assess the effects of Bacillus subtilis CU1 consumption on immune stimulation and resistance to common infectious disease (CID) episodes in healthy free-living seniors. RESULTS: One hundred subjects aged 60-74 were included in this randomized, double-blind, placebo-controlled, parallel-arms study. Subjects consumed either the placebo or the probiotic (2.10(9) B. subtilis CU1 spores daily) by short periodical courses of 10 days intermittently, alternating 18-day course of break. This scheme was repeated 4 times during the study. Symptoms of gastrointestinal and upper/lower respiratory tract infections were recorded daily by the subjects throughout the study (4 months). Blood, saliva and stool samples were collected in a predefined subset of the first forty-four subjects enrolled in the study. B. subtilis CU1 supplementation did not statistically significantly decrease the mean number of days of reported CID symptoms over the 4-month of study (probiotic group: 5.1 (7.0) d, placebo group: 6.6 (7.3) d, P = 0.2015). However, in the subset of forty-four randomized subjects providing biological samples, we showed that consumption of B. subtilis CU1 significantly increased fecal and salivary secretory IgA concentrations compared to the placebo. A post-hoc analysis on this subset showed a decreased frequency of respiratory infections in the probiotc group compared to the placebo group. CONCLUSION: Taken together, our study provides evidence that B. subtilis CU1 supplementation during the winter period may be a safe effective way to stimulate immune responses in elderly subjects.
ABSTRACT
[This corrects the article on p. 291 in vol. 39, PMID: 26301190.].
ABSTRACT
Over the past decade, growing evidence has established the gut microbiota as one of the most important determinants of metabolic disorders such as obesity and type 2 diabetes. Indeed, obesogenic diet can drastically alter bacterial populations (i.e., dysbiosis) leading to activation of pro-inflammatory mechanisms and metabolic endotoxemia, therefore promoting insulin resistance and cardiometabolic disorders. To counteract these deleterious effects, probiotic strains have been developed with the aim of reshaping the microbiome to improve gut health. In this review, we focus on benefits of widely used probiotics describing their potential mechanisms of action, especially their ability to decrease metabolic endotoxemia by restoring the disrupted intestinal mucosal barrier. We also discuss the perspective of using new bacterial strains such as butyrate-producing bacteria and the mucolytic Akkermansia muciniphila, as well as the use of prebiotics to enhance the functionality of probiotics. Finally, this review introduces the notion of genetically engineered bacterial strains specifically developed to deliver anti-inflammatory molecules to the gut.
ABSTRACT
Among nine chitinase-producing strains isolated from Tunisian soil, one isolate called S213 exhibited a potent chitinolytic activity. S213 strain was identified as Bacillus licheniformis by API 50CH system and sequence analysis of its partial 16S ribosomal DNA. Chitinolytic activity was induced either by colloidal chitin or fungal cell walls, and the highest chitinase activity reached at the late stationary phase exhibiting optimal temperature and pH of 50-60 °C and pH 6.0, respectively. SDS-PAGE analysis of the secreted colloidal chitin-induced proteins showed a major protein of about 65 kDa. This protein was identified as chitinase on the basis of its peptide sequences which displayed high homology with chitinase sequence of B. licheniformis ATCC 14580. Moreover, chitinolytic activity containing supernatant inhibited the growth of several phytopathogenic fungi including Phoma medicaginis. Interestingly, S213 strain reduced efficiently the damping-off disease caused by P. medicaginis in Medicago truncatula and should be envisaged in enzyme-based biopesticides against phytopathogen application.
Subject(s)
Antifungal Agents/isolation & purification , Ascomycota/drug effects , Chitin/metabolism , Chitinases/isolation & purification , Mitosporic Fungi/enzymology , Antifungal Agents/metabolism , Antifungal Agents/pharmacology , Ascomycota/pathogenicity , Bacillus , Cell Wall/metabolism , Chitin/chemistry , Chitinases/metabolism , Chitinases/pharmacology , DNA, Bacterial/genetics , DNA, Bacterial/isolation & purification , RNA, Ribosomal, 16S/genetics , Soil Microbiology , TunisiaABSTRACT
The vaginal microbiota of healthy, fertile women is dominated by lactobacilli. As a defence mechanism, these bacteria produce H2O2 to discourage colonization of the vagina by undesirable micro-organisms. In particular, Lactobacillus jensenii CECT 4306 is a strong producer of H2O2 and has been found to protect itself from the bactericidal effects of this compound through the activity of extracellular peroxidases. However, this peroxidase activity is dependent on the presence of Fe(3+), which is found in elevated concentrations in the vaginal mucosa as a consequence of the menstrual discharge. The aim of the present work was to evaluate whether Fe(3+) is able to modulate other potential probiotic properties of strain 4306. We found that Fe(3+) enhances the adhesion of L. jensenii CECT 4306 to mucin and to HT-29 and HT-29 MTX cells, and, in addition, improves the anti-inflammatory profile, as judged by an increase in the ratio of IL-10/IL-12p70 that were secreted by macrophages. A comparison of total, secreted and surface proteins produced in the presence and absence of Fe(3+) revealed significant differences in the concentration of the moonlighting protein glyceraldehyde-3-phosphate dehydrogenase (GAPDH). In conclusion, Fe(3+) seems to improve the probiotic characteristics of L. jensenii CECT 4306, and future research of the interactions of this strain with its vaginal environment may reveal further information about different aspects of its probiotic potential.
Subject(s)
Iron/metabolism , Lactobacillus/metabolism , Probiotics , Vagina/microbiology , Bacterial Adhesion , Bacterial Proteins , Cell Line , Female , Ferric Compounds/metabolism , Humans , Immunomodulation , Microbiota , Mucous Membrane/immunology , Mucous Membrane/microbiology , Proteome , Proteomics , Vagina/immunologyABSTRACT
Probiotics represent a potential strategy to influence the host's immune system thereby modulating immune response. Lipoteichoic Acid (LTA) is a major immune-stimulating component of Gram-positive cell envelopes. This amphiphilic polymer, anchored in the cytoplasmic membrane by means of its glycolipid component, typically consists of a poly (glycerol-phosphate) chain with D-alanine and/or glycosyl substitutions. LTA is known to stimulate macrophages in vitro, leading to secretion of inflammatory mediators such as Nitric Oxide (NO). This study investigates the structure-activity relationship of purified LTA from three probiotic Bacillus strains (Bacillus cereus CH, Bacillus subtilis CU1 and Bacillus clausii O/C). LTAs were extracted from bacterial cultures and purified. Chemical modification by means of hydrolysis at pH 8.5 was performed to remove D-alanine. The molecular structure of native and modified LTAs was determined by (1)H NMR and GC-MS, and their inflammatory potential investigated by measuring NO production by RAW 264.7 macrophages. Structural analysis revealed several differences between the newly characterized LTAs, mainly relating to their D-alanylation rates and poly (glycerol-phosphate) chain length. We observed induction of NO production by LTAs from B. subtilis and B. clausii, whereas weaker NO production was observed with B. cereus. LTA dealanylation abrogated NO production independently of the glycolipid component, suggesting that immunomodulatory potential depends on D-alanine substitutions. D-alanine may control the spatial configuration of LTAs and their recognition by cell receptors. Knowledge of molecular mechanisms behind the immunomodulatory abilities of probiotics is essential to optimize their use.
Subject(s)
Alanine/analysis , Alanine/immunology , Bacillus/chemistry , Lipopolysaccharides/analysis , Lipopolysaccharides/immunology , Probiotics/chemistry , Teichoic Acids/analysis , Teichoic Acids/immunology , Animals , Bacillus/immunology , Cell Line , Gas Chromatography-Mass Spectrometry , Hydrolysis , Immunologic Factors/analysis , Immunologic Factors/chemistry , Immunologic Factors/immunology , Lipopolysaccharides/chemistry , Macrophages/drug effects , Magnetic Resonance Spectroscopy , Mice , Molecular Structure , Nitric Oxide/metabolism , Structure-Activity Relationship , Teichoic Acids/chemistryABSTRACT
Inflammatory bowel disease (IBD) is an autoimmune disease characterized by a chronic inflammation of the gastrointestinal tract mucosa and is related to an abnormal immune response to commensal bacteria. Our aim of the present work has been to explore the levels of antibodies (IgG and IgA) raised against extracellular proteins produced by LAB and its association with IBD. We analyzed, by Western-blot and ELISA, the presence of serum antibodies (IgA and IgG) developed against extracellular protein fractions produced by different food bacteria from the genera Bifidobacterium and Lactobacillus. We used a sera collection consisting of healthy individuals (HC, n = 50), Crohn's disease patients (CD, n = 37), and ulcerative colitis patients (UC, n = 15). Levels of IgA antibodies developed against a cell-wall hydrolase from Lactobacillus casei subsp. rhamnosus GG (CWH) were significantly higher in the IBD group (P < 0.002; n = 52). The specificity of our measurements was confirmed by measuring IgA antibodies developed against the CWH peptide 365-VNTSNQTAAVSAS-377. IBD patients appeared to have different immune response to food bacteria. This paper sets the basis for developing systems for early detection of IBD, based on the association of high levels of antibodies developed against extracellular proteins from food and probiotic bacteria.
Subject(s)
Colitis, Ulcerative/immunology , Crohn Disease/immunology , Inflammatory Bowel Diseases/immunology , Lacticaseibacillus casei/immunology , Probiotics , Adult , Antibodies/blood , Antibodies/immunology , Bacterial Proteins/immunology , Bacterial Proteins/isolation & purification , Bifidobacterium/immunology , Bifidobacterium/pathogenicity , Colitis, Ulcerative/blood , Colitis, Ulcerative/microbiology , Crohn Disease/blood , Crohn Disease/microbiology , Female , Food Microbiology , Humans , Immunity, Cellular , Immunoglobulin A/blood , Immunoglobulin G/blood , Inflammatory Bowel Diseases/blood , Inflammatory Bowel Diseases/pathology , Lactobacillus/immunology , Lactobacillus/pathogenicity , Lacticaseibacillus casei/pathogenicity , Male , Middle AgedABSTRACT
Lactobacilli are generally regarded as safe; however, certain strains have been associated with cases of infection. Our workgroup has already assessed many functional properties of Lactobacillus kefiri, but parameters regarding safety must be studied before calling them probiotics. In this work, safety aspects and antimicrobial activity of L. kefiri strains were studied. None of the L. kefiri strains tested caused α- or ß-hemolysis. All the strains were susceptible to tetracycline, clindamycin, streptomycin, ampicillin, erythromycin, kanamycin, and gentamicin; meanwhile, two strains were resistant to chloramphenicol. On the other hand, all L. kefiri strains were able to inhibit both Gram(+) and Gram(-) pathogens. Regarding the in vitro results, L. kefiri CIDCA 8348 was selected to perform in vivo studies. Mice treated daily with an oral dose of 10(8) CFU during 21 days showed no signs of pain, lethargy, dehydration, or diarrhea, and the histological studies were consistent with those findings. Moreover, no differences in proinflammatory cytokines secretion were observed between treated and control mice. No translocation of microorganisms to blood, spleen, or liver was observed. Regarding these findings, L. kefiri CIDCA 8348 is a microorganism isolated from a dairy product with a great potential as probiotic for human or animal use.
Subject(s)
Cultured Milk Products/adverse effects , Lactobacillus/metabolism , Probiotics/administration & dosage , Animals , Cultured Milk Products/microbiology , Gram-Negative Bacteria/drug effects , Gram-Positive Bacteria/drug effects , Humans , Lactobacillus/pathogenicity , Mice , Microbial Sensitivity Tests , Probiotics/adverse effectsABSTRACT
We investigated the mucus-binding properties of aggregating and non-aggregating potentially probiotic strains of kefir-isolated Lactobacillus kefiri, using different substrates. All the strains were able to adhere to commercial gastric mucin (MUCIN) and extracted mucus from small intestine (SIM) and colon (CM). The extraction of surface proteins from bacteria using LiCl or NaOH significantly reduced the adhesion of three selected strains (CIDCA 8348, CIDCA 83115 and JCM 5818); although a significant proportion (up to 50%) of S-layer proteins were not completely eliminated after treatments. The surface (S-layer) protein extracts from all the strains of Lb. kefiri were capable of binding to MUCIN, SIM or CM, and no differences were observed among them. The addition of their own surface protein extract increased adhesion of CIDCA 8348 and 83115 to MUCIN and SIM, meanwhile no changes in adhesion were observed for JCM 5818. None of the seven sugars tested had the ability to inhibit the adhesion of whole bacteria to the three mucus extracts. Noteworthy, the degree of bacterial adhesion reached in the presence of their own surface protein (S-layer) extract decreased to basal levels in the presence of some sugars, suggesting an interaction between the added sugar and the surface proteins. In conclusion, the ability of these food-isolated bacteria to adhere to gastrointestinal mucus becomes an essential issue regarding the biotechnological potentiality of Lb. kefiri for the food industry.
Subject(s)
Bacterial Adhesion , Gastric Mucosa/microbiology , Intestinal Mucosa/microbiology , Lactobacillus/physiology , Mucus/microbiology , Probiotics , Animals , Bacterial Adhesion/drug effects , Colon , Cultured Milk Products/microbiology , Hexoses/pharmacology , Intestine, Small , Membrane Proteins/pharmacology , SwineABSTRACT
SCOPE: The human/microbiota cross-talk is partially mediated by bacteria-derived peptides like Serine-Threonine peptide (STp), which is resistant to gut proteolysis, is found in the human healthy colon and induces regulatory properties on gut dendritic cells (DCs); here we characterized human gut DC in ulcerative colitis (UC) patients and studied the effect of STp on their properties. METHODS AND RESULTS: Human colonic DC from healthy controls and UC patients were isolated, conditioned for 24 h +/- STp and characterized by flow cytometry, immunohistochemistry, and electron microscopy. Expression of immature DC markers DC-SIGN and ILT3, and Toll-like receptors were increased on gut UC-DC. Langerin (involved in phagocytosis), lymph node homing marker CCR7, and activation markers CD40/CD80/CD86 were decreased in UC. Gut DC had restricted stimulatory capacity for T-cells in UC. Conditioning of DC with STp in vitro reduced Toll-like receptor expression, increased CD40 and CD80 expression, and restored their stimulatory capacity. CONCLUSION: Colonic DCs display an abnormal immature phenotype in UC, which was partially restored following STp treatment. Bacteria-derived metabolites, like STp, seem to have a role in gut homeostasis that is missing in UC so they might lead a new era of probiotic products setting the basis for nondrug dietary therapy in inflammatory bowel disease.
Subject(s)
Colitis, Ulcerative/therapy , Dendritic Cells/metabolism , Lactobacillus plantarum/metabolism , Peptides/pharmacology , Serine/pharmacology , Threonine/pharmacology , Adult , Antigens, CD/genetics , Antigens, CD/metabolism , B7-1 Antigen/genetics , B7-1 Antigen/metabolism , CD40 Antigens/genetics , CD40 Antigens/metabolism , Case-Control Studies , Cell Adhesion Molecules/genetics , Cell Adhesion Molecules/metabolism , Cell Proliferation/drug effects , Colitis, Ulcerative/microbiology , Dendritic Cells/drug effects , Female , Gastrointestinal Tract/drug effects , Gastrointestinal Tract/metabolism , Gastrointestinal Tract/microbiology , Healthy Volunteers , Humans , Immunohistochemistry , Inflammatory Bowel Diseases/pathology , Inflammatory Bowel Diseases/therapy , Lectins, C-Type/genetics , Lectins, C-Type/metabolism , Male , Mannose-Binding Lectins/genetics , Mannose-Binding Lectins/metabolism , Membrane Glycoproteins , Middle Aged , Probiotics/administration & dosage , Receptors, Cell Surface/genetics , Receptors, Cell Surface/metabolism , Receptors, Immunologic , T-Lymphocytes/metabolism , Toll-Like Receptors/genetics , Toll-Like Receptors/metabolismABSTRACT
An antagonist L194 strain against Phoma medicaginis pathogenic fungi was isolated from Tunisian soil (vicinity of Tunis) and identified as Bacillus subtilis based on biochemical characteristics and partial 16S rDNA sequence. When cells were grown in a minimal medium for 24 h, spore culture supernatant exhibited 2-fold higher antifungal activity than vegetative cells. MALDI-TOF mass spectrometry analysis showed that L194 spores produced mainly iturins, surfactins and fengycins with long-chain fatty acids, and other not yet identified compounds. Both vegetative cells and spores of L194 efficiently reduced germination of P. medicaginis conidia. As revealed by atomic force microscopy, L194 spores modified conidia morphology from a regular to a deflated shape. Data suggest that lipopeptides interacted with the cytoplasmic membrane, causing pore formation. In vivo, L194 spores were highly protective against P. medicaginis by reducing disease symptoms and alleviating growth disturbance of Medicago truncatula seedlings. As a whole, the lipopeptide-producing L194 strain may be successfully used in biocontrol of plant diseases induced by phytopathogenic fungi such as P. medicaginis.