RESUMO
INTRODUCTION: Streptococcus mutans, known to be a pathogen of dental caries as well as bacteremia and infective endocarditis, is classified into four serotypes, c, e, f and k, based on the structures of serotype-specific polysaccharides. Serotype k was recently designated using blood isolates from Japanese subjects and such strains are considered to be virulent in the bloodstream. The purpose of the present study was to analyse the serotype distribution of strains isolated from Thai subjects and determine whether serotype k strains were present. METHODS: A total of 250 S. mutans strains were isolated from 50 Thai subjects, and serotypes of all strains were determined. Then, molecular and biological analyses were carried out for serotype k strains. RESULTS: Immunodiffusion and polymerase chain reaction analyses showed that serotype c was the most prevalent (70%), followed by serotypes e (22.8%), f (4.4%) and k (2.8%), which indicated that serotype k S. mutans strains occurred in Thai individuals at a similar rate to that previously reported for Japanese and Finnish populations. Molecular analyses of the seven serotype k strains showed extremely low expression of rgpE, which is related to glucose side-chain formation in serotype-specific rhamnose-glucose polymers, similar to previous reports for those other populations. In addition, analysis of the biological properties of the seven serotype k strains demonstrated low levels of sucrose-dependent adhesion, cellular hydrophobicity, dextran-binding activity and phagocytosis susceptibility by human polymorphonuclear leukocytes, which are characteristics similar to those of serotype k strains previously isolated in Japan. CONCLUSION: Our results indicate the possibility of a worldwide prevalence of serotype k strains with properties in common with those of previously reported strains.
Assuntos
Sorotipagem , Streptococcus mutans/classificação , Adolescente , Adulto , Aderência Bacteriana/fisiologia , Proteínas de Bactérias/análise , Dextranos/metabolismo , Feminino , Glucosiltransferases/análise , Humanos , Interações Hidrofóbicas e Hidrofílicas , Imunodifusão , Japão , Masculino , Pessoa de Meia-Idade , Neutrófilos/fisiologia , Fagocitose/fisiologia , Reação em Cadeia da Polimerase , Polissacarídeos Bacterianos/análise , Streptococcus mutans/patogenicidade , Sacarose/metabolismo , Tailândia , Virulência , Adulto JovemRESUMO
It is proposed that the genesis of extracellular membrane vesicles in Gram-negative bacteria is a result of cell wall turnover. Peptidoglycan turnover would cause a turgor on the outer membrane, causing the outer membrane to bulge and finally bleb. Mechanical motion would then shear the blebs into the culture medium.
Assuntos
Bactérias Gram-Negativas/ultraestrutura , Porphyromonas gingivalis/ultraestrutura , Proteínas da Membrana Bacteriana Externa/análise , Parede Celular/química , Parede Celular/metabolismo , Eletroforese em Gel de Poliacrilamida , Bactérias Gram-Negativas/química , Bactérias Gram-Negativas/fisiologia , Humanos , Ácidos Murâmicos/análise , Peptídeos/análise , Porphyromonas gingivalis/química , Porphyromonas gingivalis/fisiologia , Proteínas/análiseRESUMO
OBJECTIVE: Streptococcus mutans, known to be a major pathogen of dental caries, is also considered to cause infective endocarditis. Its 120-kDa Cnm protein binds to type I collagen, which may be a potential virulence factor. In this study, we characterized S. mutans clinical strains focusing on the cnm gene encoding Cnm. DESIGN: A total of 528 S. mutans strains isolated from Japanese, Finnish, and Thai subjects were investigated. Using molecular techniques, the distribution frequency of cnm-positive strains and location of the inserted cnm were analyzed. Furthermore, isogenic mutant strains were constructed by inactivation of the cnm gene, then their biological properties of collagen-binding and glucan-binding were evaluated. Southern hybridization of the genes encoding glucan-binding proteins was also performed. RESULTS: The distribution frequency of cnm-positive strains from Thai subjects was 12%, similar to that previously reported for Japanese and Finnish subjects. Furthermore, the location of insertion of cnm was the same in all cnm-positive clinical isolates. As for the cnm-inactivated mutant strains constructed from 28 clinical isolates, their collagen-binding activity was negligible. In addition, glucan-binding activity in the cnm-positive clinical isolates was significantly reduced and corresponded to a lack of gbpA encoding glucan-binding protein A. CONCLUSIONS: Our results indicate that strains with cnm genes, the most crucial factor for the collagen-binding property of S. mutans, are detectable at similar frequencies over several different geographic locations. In addition, the common properties of these strains are a high level of collagen-binding activity and tendency for a low level of glucan-binding activity.
Assuntos
Adesinas Bacterianas/genética , Proteínas de Transporte/genética , Colágeno/metabolismo , Streptococcus mutans/genética , Southern Blotting , Cárie Dentária/microbiologia , Finlândia , Regulação Bacteriana da Expressão Gênica/fisiologia , Glucanos/metabolismo , Humanos , Japão , Reação em Cadeia da Polimerase , Ligação Proteica/genética , Saliva/microbiologia , Streptococcus mutans/isolamento & purificação , Streptococcus mutans/patogenicidade , TailândiaRESUMO
Porphyromonas gingivalis is thought to be one of the major pathogenic organisms of adult periodontitis. Of the several virulence factors associated with the pathology it causes, evidence is now presented suggesting that outer membrane vesicles, which form from blebbing of the outer membrane, may also contribute to the pathogenesis of this bacterium. To evaluate this possibility, outer membrane vesicles were isolated from cultures of P. gingivalis and tested for their ability to promote inflammation and for their effects on the biosynthesis of E-selectin and ICAM-1 adhesion molecules and MHC class II glycoproteins. The results indicate that these vesicles are capable of inducing acute inflammation characterized by the accumulation of a large number of neutrophils in the connective tissue. This cellular response corresponds to the vesicle-mediated biosynthesis and surface membrane expression of E-selectin and ICAM-1 by vascular endothelial cells. In contrast, IFN-gamma-dependent synthesis of MHC class II molecules was found to be inhibited by vesicles. Inhibition of HLA-DR expression occurred regardless of whether vesicles were added at the same time as, 24 h before, or 24 h after IFN-gamma stimulation of endothelial cells, suggesting that the inhibitory effects occur at both the membrane and intracellular level. These findings, taken together, indicate that P. gingivalis membrane vesicles are capable of inducing and regulating cellular responses involved in inflammation and initiation of acquired immunity. Membrane vesicles are composed of muramyl peptides, periplasmic proteins and outer membrane constituents. The combination of these components probably contribute to the immune regulatory functions reported herein.
Assuntos
Membrana Celular/imunologia , Endotélio Vascular/imunologia , Antígenos HLA-DR/biossíntese , Interferon gama/farmacologia , Porphyromonas gingivalis/imunologia , Animais , Tecido Conjuntivo/imunologia , Selectina E/biossíntese , Endotélio Vascular/citologia , Endotélio Vascular/metabolismo , Citometria de Fluxo , Humanos , Inflamação/etiologia , Molécula 1 de Adesão Intercelular/biossíntese , Camundongos , Neutrófilos/fisiologia , Proteínas Recombinantes , Cordão UmbilicalRESUMO
Vaccinia virus complement control protein (VCP) has been shown to possess the ability to inhibit both classical and alternative complement pathway activation. The newly found ability of this protein to bind to heparin has been shown in previous studies to result in uptake by mast cells, possibly promoting tissue persistence. It has also been shown to reduce chemotactic migration of leukocytes by blocking chemokine binding. In addition, this study shows that VCP-through its ability to bind to glycosaminoglycans (heparin-like molecules) on the surface of human endothelial cells-is able to block antibody binding to surface major histocompatibility complex class I molecules. Since heparin binding is critical for many functions of this protein, we have attempted to characterize the molecular basis for this interaction. Segments of this protein, generated by genetic engineering of the DNA encoding VCP into the Pichia pastoris expression system, were used to localize the regions with heparin binding activity. These regions were then analyzed to more specifically define their properties for binding. It was found that the number of putative binding sites (K/R-X-K/R), the overall positive charge, and the percentage of positively charged amino acids within the protein were responsible for this interaction.