The alarming prevalence of inflammatory bowel disease (IBD) in early childhood is associated with imbalances in the microbiome, the immune response, and environmental factors. Some pathogenic Escherichia coli (E. coli) strains have been found in IBD patients, where they may influence disease progression. Therefore, the discovery of new harmful bacterial strains that have the potential to drive the inflammatory response is of great importance. In this study, we compared the immunomodulatory properties of two E. coli strains of serotype O6: the probiotic E. coli Nissle 1917 and the uropathogenic E. coli O6:K13:H1. Using the epithelial Caco-2 cell line, we investigated the different abilities of the strains to adhere to and invade epithelial cells. We confirmed the potential of E. coli Nissle 1917 to modulate the Th1 immune response in a specific manner in an in vitro setting by stimulating mouse bone marrow-derived dendritic cells (BM-DCs). In gnotobiotic in vivo experiments, we demonstrated that neonatal colonization with E. coli Nissle 1917 achieves a stable high concentration in the intestine and protects mice from the progressive effect of E. coli O6:K13:H1 in developing ulcerative colitis in an experimental model. In contrast, a single-dose treatment with E. coli Nissle 1917 is ineffective in achieving such high concentrations and does not protect against DSS-induced ulcerative colitis in mice neonatally colonized with pathobiont E. coli O6:K13:H1. Despite the stable coexistence of both E. coli strains in the intestinal environment of the mice, we demonstrated a beneficial competitive interaction between the early colonizing E. coli Nissle 1917 and the late-arriving strain O6:K13:H1, suggesting its anti-inflammatory potential for the host. This study highlights the importance of the sequence of bacterial colonization, which influences the development of the immune response in the host gut and potentially impacts future quality of life.

• Keywords
• DSS-experimental colitis, Escherichia coli, immune modulation, mouse model, priority effect,
• Publication type
• Journal Article MeSH

The classical definition of probiotics states that bacteria must be alive to be beneficial for human organism. However, recent reports show that inactivated bacteria or their effector molecules can also possess such properties. In this study, we investigated the physical and immunomodulatory properties of four Bifidobacterium strains in the heat-treated (HT) and untreated (UN) forms. We showed that temperature treatment of bacteria changes their size and charge, which affects their interaction with epithelial and immune cells. Based on the in vitro assays, we observed that all tested strains reduced the level of OVA-induced IL-4, IL-5, and IL-13 in the spleen culture of OVA-sensitized mice. We selected Bifidobacterium longum ssp. longum CCM 7952 (Bl 7952) for further analysis. In vivo experiments confirmed that untreated Bl 7952 exhibited allergy-reducing properties when administered intranasally to OVA-sensitized mice, which manifested in significant suppression of airway inflammation. Untreated Bl 7952 decreased local and systemic levels of Th2 related cytokines, OVA-specific IgE antibodies and simultaneously inhibited airway eosinophilia. In contrast, heat-treated Bl 7952 was only able to reduce IL-4 levels in the lungs and eosinophils in bronchoalveolar lavage, but increased neutrophil and macrophage numbers. We demonstrated that the viability status of Bl 7952 is a prerequisite for the beneficial effects of bacteria, and that heat treatment reduces but does not completely abolish these properties. Further research on bacterial effector molecules to elucidate the beneficial effects of probiotics in the prevention of allergic diseases is warranted.

• Keywords
• Bifidobacterium, Ovalbumin sensitization, allergy, heat inactivation, intranasal administration, probiotic,
• MeSH
• Hypersensitivity * MeSH
• Bifidobacterium * MeSH
• Disease Models, Animal MeSH
• Mice, Inbred BALB C MeSH
• Mice MeSH
• Probiotics * MeSH
• Cell Survival * MeSH
• Inflammation * MeSH
• Animals MeSH
• Check Tag
• Mice MeSH
• Female MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

A steady rise in the number of poly-sensitized patients has increased the demand for effective prophylactic strategies against multi-sensitivities. Probiotic bacteria have been successfully used in clinics and experimental models to prevent allergic mono-sensitization. In the present study, we have investigated whether probiotic bacteria could prevent poly-sensitization by imprinting on the immune system early in life. We used two recombinant variants of probiotic Escherichia coli Nissle 1917 (EcN): i) EcN expressing birch and grass pollen, poly-allergen chimera construct (EcN-Chim), and ii) an "empty" EcN without allergen expression (EcN-Ctrl). Conventional mice (CV) were treated with either EcN-Chim or EcN-Ctrl in the last week of the gestation and lactation period. Gnotobiotic mice received one oral dose of either EcN-Chim or EcN-Ctrl before mating. The offspring from both models underwent systemic allergic poly-sensitization and intranasal challenge with recombinant birch and grass pollen allergens (rBet v 1, rPhl p 1, and rPhl p 5). In the CV setting, the colonization of offspring via treatment of mothers reduced allergic airway inflammation (AAI) in offspring compared to poly-sensitized controls. Similarly, in a gnotobiotic model, AAI was reduced in EcN-Chim and EcN-Ctrl mono-colonized offspring. However, allergy prevention was more pronounced in the EcN-Ctrl mono-colonized offspring as compared to EcN-Chim. Mono-colonization with EcN-Ctrl was associated with a shift toward mixed Th1/Treg immune responses, increased expression of TLR2 and TLR4 in the lung, and maintained levels of zonulin-1 in lung epithelial cells as compared to GF poly-sensitized and EcN-Chim mono-colonized mice. This study is the first one to establish the model of allergic poly-sensitization in gnotobiotic mice. Using two different settings, gnotobiotic and conventional mice, we demonstrated that an early life intervention with the EcN without expressing an allergen is a powerful strategy to prevent poly-sensitization later in life.

• Keywords
• BALB/c, Escherichia coli Nissle 1917, allergic poly-sensitization, germ-free, hygiene hypothesis, mouse model, mucosal tolerance,
• MeSH
• Allergens immunology   MeSH
• Hypersensitivity immunology   MeSH
• Antigens, Plant immunology   MeSH
• Betula immunology   MeSH
• Epithelial Cells immunology   MeSH
• Escherichia coli immunology   MeSH
• Germ-Free Life immunology   MeSH
• Homeostasis immunology   MeSH
• Immune System immunology   MeSH
• Poaceae immunology   MeSH
• Mice, Inbred BALB C MeSH
• Mice MeSH
• Probiotics administration & dosage   MeSH
• Pollen immunology   MeSH
• Animals MeSH
• Check Tag
• Mice MeSH
• Female MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Allergens MeSH
• Antigens, Plant MeSH

Background: Mucosal mast cells (MC) are key players in IgE-mediated food allergy (FA). The evidence on the interaction between gut microbiota, MC and susceptibility to FA is contradictory. Objective: We tested the hypothesis that commensal bacteria are essential for MC migration to the gut and their maturation impacting the susceptibility to FA. Methods: The development and severity of FA symptoms was studied in sensitized germ-free (GF), conventional (CV), and mice mono-colonized with L. plantarum WCFS1 or co-housed with CV mice. MC were phenotypically and functionally characterized. Results: Systemic sensitization and oral challenge of GF mice with ovalbumin led to increased levels of specific IgE in serum compared to CV mice. Remarkably, despite the high levels of sensitization, GF mice did not develop diarrhea or anaphylactic hypothermia, common symptoms of FA. In the gut, GF mice expressed low levels of the MC tissue-homing markers CXCL1 and CXCL2, and harbored fewer MC which exhibited lower levels of MC protease-1 after challenge. Additionally, MC in GF mice were less mature as confirmed by flow-cytometry and their functionality was impaired as shown by reduced edema formation after injection of degranulation-provoking compound 48/80. Co-housing of GF mice with CV mice fully restored their susceptibility to develop FA. However, this did not occur when mice were mono-colonized with L. plantarum. Conclusion: Our results demonstrate that microbiota-induced maturation and gut-homing of MC is a critical step for the development of symptoms of experimental FA. This new mechanistic insight into microbiota-MC-FA axis can be exploited in the prevention and treatment of FA in humans.

• Keywords
• commensal bacteria, food allergy, germ-free, mast cells, mouse models,
• MeSH
• Biomarkers MeSH
• Cell Differentiation genetics   immunology   MeSH
• Cytokines metabolism   MeSH
• Cell Degranulation genetics   immunology   MeSH
• Germ-Free Life MeSH
• Mast Cells immunology   metabolism   MeSH
• Metagenome MeSH
• Metagenomics methods   MeSH
• Microbiota * immunology   MeSH
• Disease Models, Animal MeSH
• Mice MeSH
• Disease Susceptibility MeSH
• Food Hypersensitivity etiology   metabolism   pathology   MeSH
• RNA, Ribosomal, 16S MeSH
• Gastrointestinal Microbiome MeSH
• Animals MeSH
• Check Tag
• Mice MeSH
• Female MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Biomarkers MeSH
• Cytokines MeSH
• RNA, Ribosomal, 16S MeSH

Here, we compared the abilities of polysaccharides L900/2 and L900/3, which were previously isolated from Lactobacillus rhamnosus LOCK 0900, to modulate the immune response to bystander antigens in a mouse model of ovalbumin (OVA) sensitization. In vivo, both polysaccharides reduced the levels of OVA-specific IgE, IgE-dependent basophil degranulation and IgG2a antibodies, but had no effect on the levels of OVA-specific IgA or IgG1. Interestingly, both polysaccharides triggered recall cellular responses with distinct properties. L900/3 significantly suppressed the OVA-induced upregulations of IL-4, IL-5, IL-10 and IL-13 in re-stimulated spleen cells and mesenteric lymph nodes. Our findings support and expand on our previous in vitro studies by demonstrating that polymer L900/3 might modulate the Th1/Th2 balance and could be a promising candidate molecule for preventing allergic sensitization.

• MeSH
• Allergens immunology   MeSH
• Polysaccharides, Bacterial administration & dosage   isolation & purification   MeSH
• Basophils immunology   MeSH
• Cytokines biosynthesis   MeSH
• Cell Degranulation MeSH
• Immunization * MeSH
• Immunoglobulin E blood   MeSH
• Immunoglobulin G blood   MeSH
• Immunologic Factors administration & dosage   isolation & purification   MeSH
• Lacticaseibacillus rhamnosus chemistry   MeSH
• Mice MeSH
• Serpins immunology   MeSH
• Animals MeSH
• Check Tag
• Mice MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Allergens MeSH
• Polysaccharides, Bacterial MeSH
• Cytokines MeSH
• Immunoglobulin E MeSH
• Immunoglobulin G MeSH
• Immunologic Factors MeSH
• SERPIN-B5 MeSH Browser
• Serpins MeSH

Germ-free animals have been used to define the vital role of commensal bacteria on the maturation of the host immune system. However, the role of bacterial residues in diet in this setting is poorly understood. Here we investigated the effect of bacterial contamination in sterile diet on the level of allergic sensitization in germ-free mice. Sterile grain-based diets ST1 and R03 were tested for the level of bacterial contamination. ST1 contained higher amount of bacterial DNA, approximately ten times more endotoxin, and induced higher, TLR4-dependent, cytokine production in dendritic cells compared to R03. In a germ-free mouse model of sensitization to the major birch pollen allergen Bet v 1, feeding on ST1 for at least two generations was associated with decreased production of allergen-specific IgE and IgG1 antibodies in sera in comparison to R03. Furthermore, reduced levels of allergen-specific and ConA-induced cytokines IL-4, IL-5 and IL-13 accompanied by increased levels of IFN-γ were detected in splenocytes cultures of these mice. Our results show that contamination of experimental diet with bacterial residues, such as endotoxin, significantly affects the development of allergic sensitization in germ-free mice. Therefore, careful selection of sterile food is critical for the outcomes of germ-free or gnotobiotic experimental models of immune-deviated diseases.

• MeSH
• Hypersensitivity immunology   pathology   MeSH
• Antigens, Plant immunology   MeSH
• Cell Differentiation drug effects   MeSH
• Breeding MeSH
• Cytokines biosynthesis   MeSH
• Dendritic Cells drug effects   MeSH
• Diet * MeSH
• DNA, Bacterial analysis   MeSH
• Endotoxins toxicity   MeSH
• Epitopes immunology   MeSH
• Germ-Free Life MeSH
• HEK293 Cells MeSH
• Immunization * MeSH
• Immunoglobulin A immunology   MeSH
• Immunoglobulin G immunology   MeSH
• DNA Contamination MeSH
• Cells, Cultured MeSH
• Humans MeSH
• Ligands MeSH
• Mitogens pharmacology   MeSH
• Mice, Inbred BALB C MeSH
• Spleen pathology   MeSH
• Toll-Like Receptor 4 metabolism   MeSH
• Digestive System immunology   MeSH
• Animals MeSH
• Check Tag
• Humans MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Antigens, Plant MeSH
• Bet v 1 allergen, Betula MeSH Browser
• Cytokines MeSH
• DNA, Bacterial MeSH
• Endotoxins MeSH
• Epitopes MeSH
• Immunoglobulin A MeSH
• Immunoglobulin G MeSH
• Ligands MeSH
• Mitogens MeSH
• Toll-Like Receptor 4 MeSH

Increasing numbers of clinical trials and animal experiments have shown that probiotic bacteria are promising tools for allergy prevention. Here, we analyzed the immunomodulatory properties of three selected lactobacillus strains and the impact of their mixture on allergic sensitization to Bet v 1 using a gnotobiotic mouse model. We showed that Lactobacillus (L.) rhamnosus LOCK0900, L. rhamnosus LOCK0908 and L. casei LOCK0919 are recognized via Toll-like receptor 2 (TLR2) and nucleotide-binding oligomerization domain-containing protein 2 (NOD2) receptors and stimulate bone marrow-derived dendritic cells to produce cytokines in species- and strain-dependent manners. Colonization of germ-free (GF) mice with a mixture of all three strains (Lmix) improved the intestinal barrier by strengthening the apical junctional complexes of enterocytes and restoring the structures of microfilaments extending into the terminal web. Mice colonized with Lmix and sensitized to the Bet v 1 allergen showed significantly lower levels of allergen-specific IgE, IgG1 and IgG2a and an elevated total IgA level in the sera and intestinal lavages as well as an increased transforming growth factor (TGF)-β level compared with the sensitized GF mice. Splenocytes and mesenteric lymph node cells from the Lmix-colonized mice showed the significant upregulation of TGF-β after in vitro stimulation with Bet v 1. Our results show that Lmix colonization improved the gut epithelial barrier and reduced allergic sensitization to Bet v 1. Furthermore, these findings were accompanied by the increased production of circulating and secretory IgA and the regulatory cytokine TGF-β. Thus, this mixture of three lactobacillus strains shows potential for use in the prevention of increased gut permeability and the onset of allergies in humans.

• MeSH
• Hypersensitivity immunology   prevention & control   MeSH
• Antigens, Plant toxicity   MeSH
• Germ-Free Life immunology   MeSH
• HEK293 Cells MeSH
• Immunoglobulin A immunology   MeSH
• Immunoglobulin G immunology   MeSH
• Lacticaseibacillus rhamnosus immunology   MeSH
• Lacticaseibacillus casei immunology   MeSH
• Humans MeSH
• Mice, Inbred BALB C MeSH
• Mice MeSH
• Nod2 Signaling Adaptor Protein immunology   MeSH
• Intestinal Mucosa immunology   MeSH
• Toll-Like Receptor 2 immunology   MeSH
• Animals MeSH
• Check Tag
• Humans MeSH
• Mice MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Antigens, Plant MeSH
• Bet v 1 allergen, Betula MeSH Browser
• Immunoglobulin A MeSH
• Immunoglobulin G MeSH
• NOD2 protein, human MeSH Browser
• Nod2 protein, mouse MeSH Browser
• Nod2 Signaling Adaptor Protein MeSH
• TLR2 protein, human MeSH Browser
• Tlr2 protein, mouse MeSH Browser
• Toll-Like Receptor 2 MeSH

BACKGROUND: Reduced microbial diversity has been associated with inflammatory bowel disease (IBD) and probiotic bacteria have been proposed for its prevention and/or treatment. Nevertheless, comparative studies of strains of the same subspecies for specific health benefits are scarce. Here we compared two Bifidobacterium longum ssp. longum strains for their capacity to prevent experimental colitis. METHODS: Immunomodulatory properties of nine probiotic bifidobacteria were assessed by stimulation of murine splenocytes. The immune responses to B. longum ssp. longum CCM 7952 (Bl 7952) and CCDM 372 (Bl 372) were further characterized by stimulation of bone marrow-derived dendritic cell, HEK293/TLR2 or HEK293/NOD2 cells. A mouse model of dextran sulphate sodium (DSS)-induced colitis was used to compare their beneficial effects in vivo. RESULTS: The nine bifidobacteria exhibited strain-specific abilities to induce cytokine production. Bl 372 induced higher levels of both pro- and anti-inflammatory cytokines in spleen and dendritic cell cultures compared to Bl 7952. Both strains engaged TLR2 and contain ligands for NOD2. In a mouse model of DSS-induced colitis, Bl 7952, but not Bl 372, reduced clinical symptoms and preserved expression of tight junction proteins. Importantly, Bl 7952 improved intestinal barrier function as demonstrated by reduced FITC-dextran levels in serum. CONCLUSIONS: We have shown that Bl 7952, but not Bl 372, protected mice from the development of experimental colitis. Our data suggest that although some immunomodulatory properties might be widespread among the genus Bifidobacterium, others may be rare and characteristic only for a specific strain. Therefore, careful selection might be crucial in providing beneficial outcome in clinical trials with probiotics in IBD.

• MeSH
• Bifidobacterium classification   physiology   MeSH
• Dendritic Cells microbiology   pathology   MeSH
• HEK293 Cells MeSH
• Immunoenzyme Techniques MeSH
• Colitis chemically induced   prevention & control   MeSH
• Humans MeSH
• Disease Models, Animal * MeSH
• Mice, Inbred BALB C MeSH
• Mice MeSH
• Probiotics pharmacology   MeSH
• Nod2 Signaling Adaptor Protein genetics   metabolism   MeSH
• Dextran Sulfate toxicity   MeSH
• Intestines microbiology   physiopathology   MeSH
• Toll-Like Receptor 2 genetics   metabolism   MeSH
• Animals MeSH
• Check Tag
• Humans MeSH
• Mice MeSH
• Female MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Nod2 Signaling Adaptor Protein MeSH
• Dextran Sulfate MeSH
• Tlr2 protein, mouse MeSH Browser
• Toll-Like Receptor 2 MeSH