Mucosal immunization with non-living antigens usually requires the use of an adjuvant. The adjuvant activity of Bacillus firmus in the mucosal immunization of mice was described by our laboratory previously. In the present study, subcellular localization of B. firmus activities was followed. After mechanical disintegration, subcellular components of bacterium were fractionated by differential centrifugation and salting out. Bacterial cell walls, cytoplasmic membrane fraction, soluble cytoplasmic proteins, and ribosomal fractions were isolated. Their effect on the mouse immune system was studied. Lymphocyte proliferation and immunoglobulin formation in vitro were stimulated by bacterial cell wall (BCW), cytoplasmic membrane (CMF), and ribosomal fractions. BCW and CMF increased antibody formation after intratracheal immunization of mice with influenza A and B viruses, and increased protection against subsequent infection with influenza virus. The BCW fraction even induced intersubtypic cross-protection: Mice immunized with A/California/7/04 (H3N2) + BCW were resistant to the infection by the highly pathogenic A/PR/8/34 (H1N1) virus.

• MeSH
• Adjuvants, Immunologic administration & dosage   isolation & purification   MeSH
• Bacillus chemistry   MeSH
• Orthomyxoviridae Infections prevention & control   MeSH
• Cells, Cultured MeSH
• Lymphocytes drug effects   MeSH
• Disease Models, Animal MeSH
• Mice MeSH
• Cell Proliferation drug effects   MeSH
• Antibody Formation drug effects   MeSH
• Influenza Vaccines administration & dosage   MeSH
• Influenza A virus immunology   MeSH
• Influenza B virus 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
• Adjuvants, Immunologic MeSH
• Influenza Vaccines MeSH

The effect of nonpathogenic G+ bacterium B. firmus (BF) on stimulation of mouse peritoneal cells in vitro was evaluated by testing nitric-oxide-synthesis induction and cytokine formation. The reactivity was compared of peritoneal cells from two inbred mouse strains, C57B1/6 and BALB/c, which differ in their immunological reactivity. Peritoneal macrophages from C57B1/6 produced more nitric oxide after a 1-d cultivation with inactivated BF than those of BALB/c mice. In both strains, production can be further increased by adding exogenous IFN-gamma to the culture. There were no significant differences between peritoneal cells of these two mouse strains in cytokine production after optimal in vitro stimulation with BF. BF effectively activated peritoneal cells for the production of TNF-alpha, IL-1beta and IL-10, delipidated bacterium (DBF) being more efficient than BF in induction of IL-10 and TNF-alpha. On the other hand, BF had only small effect on IFN-gamma production and no detectable effect on IL-12 production. Macrophage activation by BF/DBF can represent one of the mechanisms responsible for previously described immunomodulatory activity of BF.

• MeSH
• Macrophage Activation * MeSH
• Bacillus immunology   MeSH
• Cell Culture Techniques methods   MeSH
• Cytokines immunology   metabolism   MeSH
• Lipopolysaccharides pharmacology   MeSH
• Mice, Inbred BALB C MeSH
• Mice, Inbred C57BL MeSH
• Mice MeSH
• Organic Chemicals pharmacology   MeSH
• Nitric Oxide metabolism   MeSH
• Peritoneal Cavity cytology   MeSH
• Immunity, Innate MeSH
• Animals MeSH
• Check Tag
• Mice MeSH
• Female MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Comparative Study MeSH
• Names of Substances
• Cytokines MeSH
• Lipopolysaccharides MeSH
• Nocardia delipidated cell mitogen MeSH Browser
• Organic Chemicals MeSH
• Nitric Oxide MeSH

Inactivated Bacillus firmus (BF), G+ nonpathogenic bacterium of the external environment, was coupled to ovalbumin (OVA) and used in immunization experiments as antigen carrier. Balb/c mice were immunized thrice intra-tracheally and intra-nasally with conjugates of OVA and BF. Surprisingly, administration of OVA-BF conjugates inhibited anti-OVA IgG response in both sera and mucosal secretions if compared to an exposure to OVA alone. The suppression of antigen-specific antibody production was accompanied by promotion of TH1 phenotype.

• MeSH
• Lymphocyte Activation MeSH
• Antigens administration & dosage   MeSH
• Administration, Intranasal MeSH
• Bacillus immunology   MeSH
• Immunization MeSH
• Immunoglobulin G biosynthesis   blood   MeSH
• Mice, Inbred BALB C MeSH
• Mice MeSH
• Drug Carriers MeSH
• Ovalbumin administration & dosage   immunology   MeSH
• Antibodies, Bacterial biosynthesis   blood   MeSH
• Immunity, Mucosal MeSH
• In Vitro Techniques MeSH
• Th1 Cells immunology   MeSH
• Trachea 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
• Antigens MeSH
• Immunoglobulin G MeSH
• Drug Carriers MeSH
• Ovalbumin MeSH
• Antibodies, Bacterial MeSH

Functions of T cells were determined after intranasal and intratracheal immunization of mice with ovalbumin (Ova) and Bacillus firmus (Bf), a Gram-positive nonpathogenic bacterium of the external environment, or delipidated Bf (dBf) as adjuvants, with the aim to elucidate the mechanism of support of Ova-specific antibody production caused by Bf that had been observed in an identical experiment. Neither Bf nor dBf in a mixture with Ova stimulated Ova-specific T-cell response tested as antigen-specific blast transformation. By contrast, a mild polyclonal stimulation was observed in splenocytes from mice given dBf. In vitro incubation of splenocytes with 100 micrograms (but not 10 micrograms) of Bf or dBf led to a highly significant inhibition of proliferation below the control level in all groups of animals. Supernatants of splenocyte cultures were further tested for cytokine production. IL-10 and IFN-gamma were released after in vitro challenge with dBf and in some cases also with Bf. Analysis of sera demonstrated that administration of Ova + adjuvant brought about an increase in anti-Ova IgG1, IgG2a and IgG2b whereas treatment with Ova alone caused a rise in IgG1 only. The role of Bf or dBf in the enhancement of antigen-specific antibody production could be in influencing macrophages and inducing cytokine milieu composed of IL-10, IFN-gamma and other factors that leads to a bystander stimulation of specifically activated Ova-B cell receptor (Ova-BCR)-bearing cells.

• MeSH
• Adjuvants, Immunologic administration & dosage   pharmacology   MeSH
• Lymphocyte Activation immunology   MeSH
• Administration, Intranasal MeSH
• Bacillus immunology   MeSH
• Cytokines immunology   metabolism   MeSH
• Immunization methods   standards   MeSH
• Immunoglobulin G blood   MeSH
• Mice, Inbred BALB C MeSH
• Mice MeSH
• Ovalbumin administration & dosage   immunology   MeSH
• T-Lymphocytes 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
• Adjuvants, Immunologic MeSH
• Cytokines MeSH
• Immunoglobulin G MeSH
• Ovalbumin MeSH

Bacillus firmus (a Gram-positive nonpathogenic and harmless bacterium), was shown to be a strong polyclonal activator of mouse B lymphocytes as estimated by ELISA testing of Ig concentrations in culture supernatants after incubation of BALB/c mouse splenocytes with inactivated bacillus. Synthesis of all main Ig classes and all IgG subclasses was stimulated in vitro, the considerable effect on IgA formation being the most interesting feature. B cell stimulation was T cell dependent, as was demonstrated by the effect of B. firmus on all Ig isotypes and by comparison of lymphocyte response of nu/nu mice and heterozygous nu/+ mice. The effect of B. firmus on splenocyte proliferation was stimulatory or suppressive depending on the dose of the bacterium. Increased synthesis of IFN-gamma and IL-10 (detected by ELISA in splenocyte culture supernatants) showed probable stimulation of Th1 and Th2 subpopulations. Considering the stimulatory effect on IgA formation and macrophage stimulation, B. firmus seems to be a prospective mucosal adjuvant and/or probiotic.

• MeSH
• Lymphocyte Activation * drug effects   MeSH
• B-Lymphocytes drug effects   immunology   MeSH
• Bacillus chemistry   immunology   MeSH
• Enzyme-Linked Immunosorbent Assay MeSH
• Immunoglobulin G biosynthesis   drug effects   MeSH
• Cells, Cultured MeSH
• Mice, Inbred BALB C MeSH
• Mice MeSH
• Antibodies, Bacterial biosynthesis   MeSH
• T-Lymphocytes 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
• Immunoglobulin G MeSH
• Antibodies, Bacterial MeSH