RESUMO
Abstract Objective: To assess the antibacterial activity of Psidium guajava fractions and their effects on adhesion of a multispecies biofilm consisting of Streptococcus gordonii, Fusobacterium nucleatum, and Porphyromonas gingivalis in vitro. Material and Methods: Guava leaves were obtained from the mountains of northern Peru, where they grow wild and free of pesticides. The antimicrobial activity of 25 mg/mL petroleum ether, 25 mg/mL dichloromethane and 25 mg/mL methanol fractions of P. guajava was evaluated by measuring inhibition halos, as well as the effect on the adhesion of multispecies biofilms at 4, 7 and 10 days of growth by measuring the optical density. In addition, antimicrobial susceptibility was compared using the Kruskal-Wallis test and its multiple comparison tests, and differences in mean biofilm adhesion between each fraction were assessed by repeated measures analysis and the Tukey multiple comparison test. Results: The rank-based Kruskal-Wallis test highlighted differences in the effects of the fractions on the zone of inhibition for each oral bacterium, including S. gordonii (p=0.000), F. nucleatum (p=0.000), and P. gingivalis (p=0.000), the Tukey test showed that the group treated with 0.12% chlorhexidine exhibited the least amount of adhesion, followed by the group treated with the 1.56 mg/mL methanol fraction. Conclusion: The methanol fraction of P. guajava had an antibacterial effect on S. gordonii and P. gingivalis, and the 1.56 mg/mL methanol fraction decreased biofilm adhesion.
Assuntos
Periodontite/microbiologia , Biofilmes , Psidium/química , Streptococcus gordonii/patogenicidade , Antibacterianos/farmacologia , Infecções Estreptocócicas , Técnicas In Vitro , Análise de VariânciaRESUMO
The treatment of periodontitis has numerous positive effects on established chronic health conditions, including cardiovascular disease and diabetes. However, ethical considerations do limit the establishment of human trials to investigate whether periodontitis promotes the early stages of chronic conditions. Therefore, the aim of this study was to investigate whether periodontitis induces endothelial dysfunction in hyperlipidemic apolipoprotein E gene-deficient (ApoE-/-) mice. Forty-five 8-week-old ApoE-/- mice were challenged by oral lavage with Porphyromonas gingivalis and Streptococcus gordonii for 4 weeks. A subgroup of animals (n = 15-17/group) was placed in a metabolic chamber immediately before euthanasia at 4 weeks to measure VO2/CO2 concentrations and voluntary locomotion. In infected and control animals alveolar bone levels were measured by x-ray imaging and endothelial function was determined by measuring endothelial-dependent vasorelaxation of aortic rings. The mRNA expression levels of serum amyloid A and tumor necrosis factor were determined in liver tissues by qRT PCR and protein concentrations in serum by ELISA. Caecal contents were analysed by sequencing to determine changes to the gut microbiota to investigate linkages between microbiome and systemic changes. The results showed that oral lavage of P. gingivalis and S. gordonii for 4 weeks, initiated periodontitis in ApoE-/- mice, similar to the human situation. The oral inflammation was accompanied by a significant increase in mRNA expression of pro-inflammatory mediators serum amyloid A1 and tumor necrosis factor in the liver. Mice with periodontitis also exhibited impaired endothelial-dependent vasorelaxation responses to acetylcholine. This systemic response was connected to increased energy expenditure, locomotion and respiratory quotient. No differences were detected in caecal microbiota between the infected and control animals. Overall, this is the first report that provide evidence that periodontitis induces endothelial dysfunction in mice. Other systemic responses observed in response to the local reaction need further investigation. The study suggests that early prevention of periodontitis may help limit the early stages of endothelial dysfunction that is linked to atherogenesis in humans.
Assuntos
Apolipoproteínas E/genética , Infecções por Bacteroidaceae/diagnóstico por imagem , Hiperlipidemias/genética , Periodontite/microbiologia , Placa Aterosclerótica/diagnóstico por imagem , Animais , Bactérias/classificação , Bactérias/genética , Bactérias/isolamento & purificação , Modelos Animais de Doenças , Metabolismo Energético , Microbioma Gastrointestinal , Técnicas de Inativação de Genes , Hiperlipidemias/microbiologia , Masculino , Camundongos , Periodontite/diagnóstico por imagem , Periodontite/genética , Filogenia , Placa Aterosclerótica/microbiologia , Porphyromonas gingivalis/patogenicidade , Análise de Sequência de RNA , Proteína Amiloide A Sérica/genética , Proteína Amiloide A Sérica/metabolismo , Streptococcus gordonii/patogenicidade , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/metabolismo , Raios XRESUMO
Streptococcus gordonii and Streptococcus sanguinis belong to the Mitis group streptococci, which mostly are commensals in the human oral cavity. Though they are oral commensals, they can escape their niche and cause infective endocarditis, a severe infection with high mortality. Several virulence factors important for the development of infective endocarditis have been described in these two species. However, the background for how the commensal bacteria, in some cases, become pathogenic is still not known. To gain a greater understanding of the mechanisms of the pathogenic potential, we performed a comparative analysis of 38 blood culture strains, S. sanguinis (n = 20) and S. gordonii (n = 18) from patients with verified infective endocarditis, along with 21 publicly available oral isolates from healthy individuals, S. sanguinis (n = 12) and S. gordonii (n = 9). Using whole genome sequencing data of the 59 streptococci genomes, functional profiles were constructed, using protein domain predictions based on the translated genes. These functional profiles were used for clustering, phylogenetics and machine learning. A clear separation could be made between the two species. No clear differences between oral isolates and clinical infective endocarditis isolates were found in any of the 675 translated core-genes. Additionally, random forest-based machine learning and clustering of the pan-genome data as well as amino acid variations in the core-genome could not separate the clinical and oral isolates. A total of 151 different virulence genes was identified in the 59 genomes. Among these homologs of genes important for adhesion and evasion of the immune system were found in all of the strains. Based on the functional profiles and virulence gene content of the genomes, we believe that all analysed strains had the ability to become pathogenic.
Assuntos
Endocardite Bacteriana/microbiologia , Endocardite/microbiologia , Genoma Bacteriano , Infecções Estreptocócicas/microbiologia , Streptococcus gordonii/genética , Streptococcus sanguis/genética , Fatores de Virulência/genética , Endocardite/patologia , Endocardite Bacteriana/patologia , Endocárdio/microbiologia , Endocárdio/patologia , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Aprendizado de Máquina , Boca/microbiologia , Boca/patologia , Filogenia , Infecções Estreptocócicas/patologia , Streptococcus gordonii/classificação , Streptococcus gordonii/isolamento & purificação , Streptococcus gordonii/patogenicidade , Streptococcus sanguis/classificação , Streptococcus sanguis/isolamento & purificação , Streptococcus sanguis/patogenicidade , Simbiose/fisiologia , Virulência , Fatores de Virulência/classificação , Fatores de Virulência/metabolismoRESUMO
Streptococcus gordonii and Streptococcus sanguinis are primary colonizers of the tooth surface. Although generally non-pathogenic in the oral environment, they are a frequent cause of infective endocarditis. Both streptococcal species express a serine-rich repeat surface adhesin that mediates attachment to sialylated glycans on mucin-like glycoproteins, but the specific sialoglycan structures recognized can vary from strain to strain. Previous studies have shown that sialoglycan binding is clearly important for aortic valve infections caused by some S. gordonii, but this process did not contribute to the virulence of a strain of S. sanguinis. However, these streptococci can bind to different subsets of sialoglycan structures. Here we generated isogenic strains of S. gordonii that differ only in the type and range of sialoglycan structures to which they adhere and examined whether this rendered them more or less virulent in a rat model of endocarditis. The findings indicate that the recognition of specific sialoglycans can either enhance or diminish pathogenicity. Binding to sialyllactosamine reduces the initial colonization of mechanically-damaged aortic valves, whereas binding to the closely-related trisaccharide sialyl T-antigen promotes higher bacterial densities in valve tissue 72 hours later. A surprising finding was that the initial attachment of streptococci to aortic valves was inversely proportional to the affinity of each strain for platelets, suggesting that binding to platelets circulating in the blood may divert bacteria away from the endocardial surface. Importantly, we found that human and rat platelet GPIbα (the major receptor for S. gordonii and S. sanguinis on platelets) display similar O-glycan structures, comprised mainly of a di-sialylated core 2 hexasaccharide, although the rat GPIbα has a more heterogenous composition of modified sialic acids. The combined results suggest that streptococcal interaction with a minor O-glycan on GPIbα may be more important than the over-all affinity for GPIbα for pathogenic effects.
Assuntos
Endocardite Bacteriana/imunologia , Glicoproteínas/imunologia , Ácidos Siálicos/imunologia , Infecções Estreptocócicas/imunologia , Streptococcus gordonii/imunologia , Streptococcus sanguis/imunologia , Animais , Modelos Animais de Doenças , Endocardite Bacteriana/patologia , Feminino , Humanos , Masculino , Ratos , Ratos Sprague-Dawley , Índice de Gravidade de Doença , Infecções Estreptocócicas/patologia , Streptococcus gordonii/patogenicidade , Streptococcus sanguis/patogenicidadeRESUMO
Pylephlebitis is a condition with thrombophlebitis of the portal mesenteric venous system. Herein, we report a patient suggesting odontogenic bacteremia as a risk factor of pylephlebitis. He was diagnosed as superior mesenteric vein thrombophlebitis, and blood cultures grew Gemella sanguinis and Streptococcus gordonii.
Assuntos
Bacteriemia/complicações , Bacteriemia/microbiologia , Gemella/patogenicidade , Veias Mesentéricas/patologia , Streptococcus gordonii/patogenicidade , Tromboflebite/complicações , Antibacterianos/uso terapêutico , Implantes Dentários/efeitos adversos , Humanos , Masculino , Veias Mesentéricas/diagnóstico por imagem , Pessoa de Meia-Idade , Boca/microbiologia , Veia Porta , Fatores de Risco , Tromboflebite/diagnóstico por imagem , Tromboflebite/patologia , Extração Dentária/efeitos adversosRESUMO
Streptococcus gordonii is commonly found in the periapical endodontic lesions of patients with apical periodontitis, a condition characterized by inflammation and periapical bone loss. Since bone metabolism is controlled by osteoclastic bone resorption and osteoblastic bone formation, we investigated the effects of S. gordonii on the differentiation and function of osteoclasts and osteoblasts. For the determination of bone resorption activity in vivo, collagen sheets soaked with heat-killed S. gordonii were implanted on mouse calvaria, and the calvarial bones were scanned by micro-computed tomography. Mouse bone marrow-derived macrophages (BMMs) were stimulated with M-CSF and RANKL for 2 days and then differentiated into osteoclasts in the presence or absence of heat-killed S. gordonii. Tartrate-resistant acid phosphatase staining was performed to determine osteoclast differentiation. Primary osteoblast precursors were differentiated into osteoblasts with ascorbic acid and ß-glycerophosphate in the presence or absence of heat-killed S. gordonii. Alkaline phosphatase staining and alizarin red S staining were conducted to determine osteoblast differentiation. Western blotting was performed to examine the expression of transcription factors including c-Fos, NFATc1, and Runx2. Heat-killed S. gordonii induced bone destruction in a mouse calvarial implantation model. The differentiation of RANKL-primed BMMs into osteoclasts was enhanced in the presence of heat-killed S. gordonii. Heat-killed S. gordonii increased the expression of c-Fos and NFATc1, which are essential transcription factors for osteoclast differentiation. On the other hand, heat-killed S. gordonii inhibited osteoblast differentiation and reduced the expression of Runx2, an essential transcription factor for osteoblast differentiation. S. gordonii exerts bone resorptive activity by increasing osteoclast differentiation and reducing osteoblast differentiation, which may be involved in periapical bone resorption.
Assuntos
Reabsorção Óssea/microbiologia , Diferenciação Celular , Osteoblastos , Osteoclastos , Osteogênese , Streptococcus gordonii/patogenicidade , Fosfatase Alcalina , Animais , Ácido Ascórbico/metabolismo , Reabsorção Óssea/diagnóstico por imagem , Subunidade alfa 1 de Fator de Ligação ao Core/metabolismo , Citocinas , Modelos Animais de Doenças , Glicerofosfatos/metabolismo , Fator Estimulador de Colônias de Macrófagos/metabolismo , Macrófagos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Fatores de Transcrição NFATC/metabolismo , Periodontite Periapical , Proteínas Proto-Oncogênicas c-fos/metabolismo , Ligante RANK/metabolismo , Fatores de Transcrição , Regulação para Cima , Microtomografia por Raio-XRESUMO
Periodontal disease consists of chronic gingival inflammation characterized by both degradation of the periodontal connective tissue and alveolar bone loss. Drug therapy is used as an auxiliary treatment method in severe chronic periodontitis, aggressive periodontitis, and periodontitis-associated systemic disease. Nal-P-113, a modified antimicrobial peptide, specifically replaces the histidine residues of P-113 with the bulky amino acid ß-naphthylalanine, and our previous studies have verified that this novel peptide is not toxic to the human body within a certain concentration range. The objective of the present study was to evaluate the effect of Nal-P-113 on periodontal pathogens and periodontal status in clinical studies. In a split-mouth clinical trial, the pocket depth and bleeding index values tended to decrease in the experimental group compared with those in the control group. SEM results verified that Nal-P-113 restrained the maturation of plaque. Based on real-time polymerase chain reaction, the levels of Fusobacterium nucleatum, Streptococcus gordonii, Treponema denticola, and Porphyromonas gingivalis in subgingival plaque were decreased when the subjects were given Nal-P-113. Bacterial growth curve analysis and a biofilm susceptibility assay verified that Nal-P-113 at a concentration of 20 µg/mL restrained the growth of S. gordonii, F. nucleatum, and P. gingivalis and biofilm formation. Therefore, Nal-P-113 effectively reduces periodontal pathogens and ameliorates periodontal status.
Assuntos
Antibacterianos/administração & dosagem , Inflamação/tratamento farmacológico , Peptídeos/administração & dosagem , Doenças Periodontais/tratamento farmacológico , Adulto , Antibacterianos/química , Biofilmes/efeitos dos fármacos , Feminino , Fusobacterium nucleatum/efeitos dos fármacos , Fusobacterium nucleatum/patogenicidade , Gengiva/efeitos dos fármacos , Gengiva/microbiologia , Humanos , Inflamação/microbiologia , Inflamação/patologia , Masculino , Pessoa de Meia-Idade , Peptídeos/química , Doenças Periodontais/microbiologia , Bolsa Periodontal/tratamento farmacológico , Bolsa Periodontal/microbiologia , Porphyromonas gingivalis/efeitos dos fármacos , Porphyromonas gingivalis/patogenicidade , Streptococcus gordonii/efeitos dos fármacos , Streptococcus gordonii/patogenicidade , Treponema denticola/efeitos dos fármacos , Treponema denticola/patogenicidade , beta-Alanina/análogos & derivados , beta-Alanina/químicaRESUMO
BACKGROUND: Streptococcus gordonii is an infrequent cause of infective endocarditis (IE); associated spondylodiskitis has not yet been described in the literature. PURPOSE: We describe 2 patients who presented with new-onset, severe back pain; blood cultures revealed S. gordonii bacteremia, which led to the diagnosis of spondylodiskitis and IE. We review our 2-decade experience with S. gordonii bacteremia to describe the clinical and epidemiological characteristics of these patients. RESULTS: In our hospital over the last 20 years (1998-2017), a total of 15 patients with S. gordonii bacteremia were diagnosed, including 11 men and 4 women, and the mean age was 65 ± 22 (range 23-95). The most common diagnosis was IE (9 patients), spondylodiskitis (the presented 2 patients, who in addition were diagnosed with endocarditis), necrotizing fasciitis (1), sternitis (1), septic arthritis (1) and pneumonia (1). The 11 patients with IE were treated with penicillin ± gentamicin, or ceftriaxone for 6 weeks, 5 required valve surgery and 10/11 (91%) attained complete cure. The 2 patients with diskitis required 2-3 months of intravenous antibiotics to achieve complete cure. CONCLUSION: Spondylodiskitis was the presenting symptom of 2/11 (18%) patients with S. gordonii endocarditis. Spondylodiskitis should probably be looked for in patients diagnosed with S. gordonii endocarditis and back pain as duration of antibiotic treatment to achieve complete cure may be considerably longer.
Assuntos
Discite/etiologia , Discite/microbiologia , Endocardite Bacteriana/etiologia , Endocardite Bacteriana/microbiologia , Infecções Estreptocócicas/complicações , Streptococcus gordonii/patogenicidade , Adulto , Idoso , Idoso de 80 Anos ou mais , Antibacterianos/uso terapêutico , Artrite Infecciosa , Bacteriemia/tratamento farmacológico , Discite/tratamento farmacológico , Discite/epidemiologia , Endocardite Bacteriana/tratamento farmacológico , Endocardite Bacteriana/epidemiologia , Fasciite Necrosante , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Pneumonia , Infecções Estreptocócicas/tratamento farmacológico , Infecções Estreptocócicas/epidemiologia , Streptococcus gordonii/efeitos dos fármacos , Adulto JovemRESUMO
Many human infections are polymicrobial in origin, and interactions among community inhabitants shape colonization patterns and pathogenic potential 1 . Periodontitis, which is the sixth most prevalent infectious disease worldwide 2 , ensues from the action of dysbiotic polymicrobial communities 3 . The keystone pathogen Porphyromonas gingivalis and the accessory pathogen Streptococcus gordonii interact to form communities in vitro and exhibit increased fitness in vivo 3,4 . The mechanistic basis of this polymicrobial synergy, however, has not been fully elucidated. Here we show that streptococcal 4-aminobenzoate/para-amino benzoic acid (pABA) is required for maximal accumulation of P. gingivalis in dual-species communities. Metabolomic and proteomic data showed that exogenous pABA is used for folate biosynthesis, and leads to decreased stress and elevated expression of fimbrial adhesins. Moreover, pABA increased the colonization and survival of P. gingivalis in a murine oral infection model. However, pABA also caused a reduction in virulence in vivo and suppressed extracellular polysaccharide production by P. gingivalis. Collectively, these data reveal a multidimensional aspect to P. gingivalis-S. gordonii interactions and establish pABA as a critical cue produced by a partner species that enhances the fitness of P. gingivalis while diminishing its virulence.
Assuntos
Infecções por Bacteroidaceae/microbiologia , Coinfecção/microbiologia , Interações Microbianas , Porphyromonas gingivalis/metabolismo , Porphyromonas gingivalis/patogenicidade , Infecções Estreptocócicas/microbiologia , Streptococcus gordonii/metabolismo , Ácido 4-Aminobenzoico/metabolismo , Ácido 4-Aminobenzoico/farmacologia , Adesinas Bacterianas/metabolismo , Animais , Aderência Bacteriana , Biofilmes , Coinfecção/metabolismo , Modelos Animais de Doenças , Disbiose , Feminino , Humanos , Metabolômica , Camundongos , Camundongos Endogâmicos BALB C , Periodontite/microbiologia , Porphyromonas gingivalis/efeitos dos fármacos , Porphyromonas gingivalis/crescimento & desenvolvimento , Proteômica , Streptococcus gordonii/efeitos dos fármacos , Streptococcus gordonii/genética , Streptococcus gordonii/patogenicidade , Virulência , para-Aminobenzoatos/metabolismo , para-Aminobenzoatos/farmacologiaRESUMO
Periodontitis are bacterial inflammatory diseases, where the bacterial biofilms present on the tooth-supporting tissues switch from a healthy state towards a pathogenic state. Among bacterial species involved in the disease, Porphyromonas gingivalis has been shown to induce dysbiosis, and to induce virulence of otherwise healthy bacteria like Streptococcus gordonii. During biofilm development, primary colonizers such as S. gordonii first attach to the surface and allow the subsequent adhesion of periodontal pathogens such as P. gingivalis. Interactions between those two bacteria have been extensively studied during the adhesion step of the biofilm. The aim of the study was to understand interactions of both species during the growing phase of the biofilm, for which little knowledge is available, using a mathematical model. This two-species biofilm model was based on a substrate-dependent growth, implemented with damage parameters, and validated thanks to data obtained on experimental biofilms. Three different hypothesis of interactions were proposed and assayed using this model: independence, competition between both bacteria species, or induction of toxicity by one species for the other species. Adequacy between experimental and simulated biofilms were found with the last hypothetic mathematical model. This new mathematical model of two species bacteria biofilms, dependent on different substrates for growing, can be applied to any bacteria species, environmental conditions, or steps of biofilm development. It will be of great interest for exploring bacterial interactions in biofilm conditions.
Assuntos
Biofilmes , Modelos Biológicos , Boca/microbiologia , Porphyromonas gingivalis/fisiologia , Streptococcus gordonii/fisiologia , Humanos , Microscopia Eletrônica de Varredura , Porphyromonas gingivalis/crescimento & desenvolvimento , Porphyromonas gingivalis/patogenicidade , Streptococcus gordonii/crescimento & desenvolvimento , Streptococcus gordonii/patogenicidadeRESUMO
A range of Streptococcus bacteria are able to interact with blood platelets to form a thrombus (clot). Streptococcus gordonii is ubiquitous within the human oral cavity and amongst the common pathogens isolated from subjects with infective endocarditis. Two cell surface proteins, Hsa and Platelet adherence protein A (PadA), in S. gordonii mediate adherence and activation of platelets. In this study, we demonstrate that PadA binds activated platelets and that an NGR (Asparagine-Glycine-Arginine) motif within a 657 amino acid residue N-terminal fragment of PadA is responsible for this, together with two other integrin-like recognition motifs RGT and AGD. PadA also acts in concert with Hsa to mediate binding of S. gordonii to cellular fibronectin and vitronectin, and to promote formation of biofilms. Evidence is presented that PadA and Hsa are each reliant on the other's active presentation on the bacterial cell surface, suggesting cooperativity in functions impacting both colonization and pathogenesis.
Assuntos
Adesinas Bacterianas/metabolismo , Proteínas da Membrana Bacteriana Externa/metabolismo , Proteínas de Transporte/metabolismo , Matriz Extracelular/metabolismo , Interações Hospedeiro-Patógeno , Ativação Plaquetária , Streptococcus gordonii/patogenicidade , Fatores de Virulência/metabolismo , Aderência Bacteriana , Biofilmes/crescimento & desenvolvimento , Hemaglutininas Virais , Humanos , Proteínas de Membrana/metabolismo , Streptococcus gordonii/crescimento & desenvolvimento , Streptococcus gordonii/fisiologiaRESUMO
Periodontitis is a highly prevalent disease caused in part by an aberrant host response to the oral multi-species biofilm. A balance between the oral bacteria and host immunity is essential for oral health. Imbalances in the oral microbiome lead to an uncontrolled host inflammatory response and subsequent periodontal disease (i.e. gingivitis and periodontitis). TREM-1 is a signaling receptor present on myeloid cells capable of acting synergistically with other pattern recognition receptors leading to amplification of inflammatory responses. The aim of this study was to investigate the activation of the TREM-1 pathway in the human monocyte-like cell line THP-1 exposed to both oral pathogens and commensals. The relative expression of the genes encoding TREM-1 and its adapter protein DAP12 were determined by quantitative real-time polymerase chain reaction. The surface expression of TREM-1 was determined by flow cytometry. Soluble TREM-1 and cytokines were measured by enzyme-linked immunosorbent assay. The results demonstrate that both commensal and pathogenic oral bacteria activate the TREM-1 pathway, resulting in a proinflammatory TREM-1 activity-dependent increase in proinflammatory cytokine production.
Assuntos
Bactérias/imunologia , Bactérias/patogenicidade , Monócitos/microbiologia , Doenças Periodontais/imunologia , Receptor Gatilho 1 Expresso em Células Mieloides/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Células Cultivadas , Citocinas/genética , Citometria de Fluxo , Humanos , Imunidade Inata , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Doenças Periodontais/microbiologia , Porphyromonas gingivalis/imunologia , Porphyromonas gingivalis/patogenicidade , Reação em Cadeia da Polimerase em Tempo Real , Transdução de Sinais , Streptococcus gordonii/imunologia , Streptococcus gordonii/patogenicidade , Simbiose , Células THP-1 , Receptor Gatilho 1 Expresso em Células Mieloides/genéticaRESUMO
Streptococcus gordonii is a commensal inhabitant of human oral biofilms. Previously, we identified an enzyme called SdbA that played an important role in biofilm formation by S. gordonii. SdbA is thiol-disulfide oxidoreductase that catalyzes disulfide bonds in secreted proteins. Surprisingly, inactivation of SdbA results in enhanced biofilm formation. In this study we investigated the basis for biofilm formation by the ΔsdbA mutant. The results revealed that biofilm formation was mediated by the interaction between the CiaRH and ComDE two-component signalling systems. Although it did not affect biofilm formation by the S. gordonii parent strain, CiaRH was upregulated in the ΔsdbA mutant and it was essential for the enhanced biofilm phenotype. The biofilm phenotype was reversed by inactivation of CiaRH or by the addition of competence stimulating peptide, the production of which is blocked by CiaRH activity. Competition assays showed that the enhanced biofilm phenotype also corresponded to increased oral colonization in mice. Thus, the interaction between SdbA, CiaRH and ComDE affects biofilm formation both in vitro and in vivo.
Assuntos
Proteínas de Bactérias/genética , Histidina Quinase/genética , Proteínas de Membrana/genética , Proteínas Quinases/genética , Infecções Estreptocócicas/microbiologia , Streptococcus gordonii/genética , Animais , Proteínas de Bactérias/metabolismo , Biofilmes/crescimento & desenvolvimento , Regulação Bacteriana da Expressão Gênica , Histidina Quinase/metabolismo , Humanos , Proteínas de Membrana/metabolismo , Camundongos , Mutação , Óperon/genética , Fenótipo , Proteínas Quinases/metabolismo , Transdução de Sinais , Infecções Estreptocócicas/genética , Streptococcus gordonii/patogenicidadeRESUMO
Streptococcus gordonii SspA and SspB proteins, members of the antigen I/II (AgI/II) family of Streptococcus adhesins, mediate adherence to cysteine-rich scavenger glycoprotein gp340 and cells of other oral microbial species. In this article we investigated further the mechanism of coaggregation between S. gordonii DL1 and Actinomyces oris T14V. Previous mutational analysis of S. gordonii suggested that SspB was necessary for coaggregation with A. oris T14V. We have confirmed this by showing that Lactococcus lactis surrogate host cells expressing SspB coaggregated with A. oris T14V and PK606 cells, while L. lactis cells expressing SspA did not. Coaggregation occurred independently of expression of A. oris type 1 (FimP) or type 2 (FimA) fimbriae. Polysaccharide was prepared from cells of A. oris T14V and found to contain 1,4-, 4,6- and 3,4-linked glucose, 1,4-linked mannose, and 2,4-linked galactose residues. When immobilized onto plastic wells this polysaccharide supported binding of L. lactis expressing SspB, but not binding of L. lactis expressing other AgI/II family proteins. Purified recombinant NAVP region of SspB, comprising amino acid (aa) residues 41-847, bound A. oris polysaccharide but the C-domain (932-1470 aa residues) did not. A site-directed deletion of 29 aa residues (Δ691-718) close to the predicted binding cleft within the SspB V-region ablated binding of the NAVP region to polysaccharide. These results infer that the V-region head of SspB recognizes an actinomyces polysaccharide ligand, so further characterizing a lectin-like coaggregation mechanism occurring between two important primary colonizers.
Assuntos
Actinomyces/fisiologia , Adesinas Bacterianas/metabolismo , Polissacarídeos Bacterianos/metabolismo , Streptococcus gordonii/fisiologia , Adesinas Bacterianas/química , Adesinas Bacterianas/genética , Sequência de Aminoácidos , Aderência Bacteriana , Humanos , Lectinas , Interações Microbianas , Polissacarídeos Bacterianos/química , Polissacarídeos Bacterianos/isolamento & purificação , Streptococcus gordonii/genética , Streptococcus gordonii/patogenicidadeRESUMO
Damaged cardiac valves attract blood-borne bacteria, and infective endocarditis is often caused by viridans group streptococci. While such bacteria use multiple adhesins to maintain their normal oral commensal state, recognition of platelet sialoglycans provides an intermediary for binding to damaged valvular endocardium. We use a customized sialoglycan microarray to explore the varied binding properties of phylogenetically related serine-rich repeat adhesins, the GspB, Hsa, and SrpA homologs from Streptococcus gordonii and Streptococcus sanguinis species, which belong to a highly conserved family of glycoproteins that contribute to virulence for a broad range of Gram-positive pathogens. Binding profiles of recombinant soluble homologs containing novel sialic acid-recognizing Siglec-like domains correlate well with binding of corresponding whole bacteria to arrays. These bacteria show multiple modes of glycan, protein, or divalent cation-dependent binding to synthetic glycoconjugates and isolated glycoproteins in vitro. However, endogenous asialoglycan-recognizing clearance receptors are known to ensure that only fully sialylated glycans dominate in the endovascular system, wherein we find these particular streptococci become primarily dependent on their Siglec-like adhesins for glycan-mediated recognition events. Remarkably, despite an excess of alternate sialoglycan ligands in cellular and soluble blood components, these adhesins selectively target intact bacteria to sialylated ligands on platelets, within human whole blood. These preferred interactions are inhibited by corresponding recombinant soluble adhesins, which also preferentially recognize platelets. Our data indicate that circulating platelets may act as inadvertent Trojan horse carriers of oral streptococci to the site of damaged endocardium, and provide an explanation why it is that among innumerable microbes that gain occasional access to the bloodstream, certain viridans group streptococci have a selective advantage in colonizing damaged cardiac valves and cause infective endocarditis.
Assuntos
Adesinas Bacterianas/metabolismo , Plaquetas/metabolismo , Endocardite Bacteriana/sangue , Streptococcus gordonii/metabolismo , Streptococcus sanguis/metabolismo , Fatores de Virulência/metabolismo , Feminino , Humanos , Masculino , Análise Serial de Proteínas , Lectinas Semelhantes a Imunoglobulina de Ligação ao Ácido Siálico/metabolismo , Streptococcus gordonii/patogenicidade , Streptococcus sanguis/patogenicidadeRESUMO
Oral streptococci adhere to tooth-immobilized glycoprotein 340 (GP340) via the surface protein antigen I/II (AgI/II) and its homologs as the first step in pathogenesis. Studying this interaction using recombinant proteins, we observed that calcium increases the conformational stability of the scavenger-rich cysteine repeat (SRCRs) domains of GP340. Our results also show that AgI/II adheres specifically with nanomolar affinity to the calcium-induced SRCR conformation in an immobilized state and not in solution. This interaction is significantly dependent on the O-linked carbohydrates present on the SRCRs. This study also establishes that a single SRCR domain of GP340 contains the two surfaces to which the apical and C-terminal regions of AgI/II noncompetitively adhere. Compared with the single SRCR domain, the three tandem SRCR domains displayed a collective/cooperative increase in their bacterial adherence and aggregation. The previously described SRCRP2 peptide that was shown to aggregate several oral streptococci displayed limited aggregation and also nonspecific adherence compared to SRCR domains. Finally, we show distinct species-specific adherence/aggregation between Streptococcus mutans AgI/II and Streptococcus gordonii SspB in their interaction with the SRCRs. This study concludes that identification of the metal ion and carbohydrate adherence motifs on both SRCRs and AgI/II homologs could lead to the development of anti-adhesive inhibitors that could deter the adherence of pathogenic oral streptococci and thereby prevent the onset of infections.
Assuntos
Receptores de Superfície Celular/química , Receptores de Superfície Celular/metabolismo , Antígenos de Bactérias/metabolismo , Aderência Bacteriana/fisiologia , Cálcio/metabolismo , Proteínas de Ligação ao Cálcio , Proteínas de Ligação a DNA , Glicosilação , Humanos , Modelos Moleculares , Boca/microbiologia , Ligação Proteica , Conformação Proteica , Domínios e Motivos de Interação entre Proteínas , Estabilidade Proteica , Proteínas Recombinantes/química , Proteínas Recombinantes/metabolismo , Streptococcus gordonii/metabolismo , Streptococcus gordonii/patogenicidade , Streptococcus mutans/metabolismo , Streptococcus mutans/patogenicidade , Proteínas Supressoras de TumorRESUMO
Whole cells of wild-type strains of Streptococcus gordonii and Streptococcus mutans induced Toll-like receptor 2 (TLR2)-mediated nuclear factor-κB (NF-κB) activation, whereas those of lipoprotein (LP)-deficient strains did not. All strains upregulated the proliferation of TLR2(+/+) splenocytes more strongly than TLR2(-/-) splenocytes. However, significant differences were not observed between the cytokine-inducing activities of wild-type and LP-deficient strains toward TLR2(+/+) and TLR2(-/-) splenocytes. Muramyl dipeptide as well as whole cells not only induced nucleotide-binding oligomerization domain 2 (NOD2)-mediated activation of NF-κB but also enhanced the proliferation of TLR2(-/-) as well as TLR2(+/+) splenocytes. Wild-type strains of these streptococci were more resistant to clearance from blood and organs (liver and spleen) in TLR2(+/+) but not TLR2(-/-) mice and induced production of larger amounts of blood TNF-α than the LP-deficient strains. Wild-type strains of both species adhered to human vascular endothelial cells more strongly than did the LP-deficient strains. Thus, this study suggested that LP plays an important role in the recognition of these streptococci by the host in vivo as well as in vitro and that these streptococci possess some components recognized by NOD2 and/or TLR2 that are involved in the mitogenic activity toward splenocytes.
Assuntos
Citocinas/metabolismo , Lipoproteínas/imunologia , NF-kappa B/imunologia , Streptococcus gordonii/imunologia , Streptococcus mutans/imunologia , Receptor 2 Toll-Like/imunologia , Animais , Aderência Bacteriana , Sangue/microbiologia , Proliferação de Células , Células Cultivadas , Células Endoteliais/microbiologia , Feminino , Humanos , Leucócitos Mononucleares/imunologia , Lipoproteínas/deficiência , Fígado/microbiologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Boca/microbiologia , Baço/imunologia , Baço/microbiologia , Streptococcus gordonii/isolamento & purificação , Streptococcus gordonii/patogenicidade , Streptococcus mutans/isolamento & purificação , Streptococcus mutans/patogenicidade , Receptor 2 Toll-Like/deficiênciaRESUMO
Oral streptococci comprise a numerically prominent group of oral bacteria that occur primarily on the human tooth surface as members of the biofilm community, commonly referred to as dental plaque. These streptococci are not only causative of dental caries and are primers for colonization of periodontopathic bacteria, but also well known for their ability to colonize damaged heart valves, identified most frequently as primary etiological agents of infective endocarditis. A number of streptococcal cell surface components are known to contribute to colonization of the tooth surface including putative adhesins recognizing host sialic acid (sialic acid-binding adhesins). Interactions mediated by these adhesins include the attachment of these bacteria to saliva-coated hydroxyapatite and their adhesion to erythrocytes, both of which are abolished or reduced by sialidase pretreatment of the corresponding host sialoglycoconjugate receptors. The sialic acid-binding adhesin on Streptococcus gordonii, an early colonizer on the tooth surface, has been molecularly analyzed. The adhesin, Hsa (203-kDa protein), consists of an N-terminal non repetitive region (NR1) including a signal sequence, a relatively short serine-rich region (SR1), a second non repetitive region (NR2), a long serine-rich region (SR2) containing 113 dodecapeptide repeats accounting for 75% of the whole protein, and a C-terminal cell wall anchoring domain. Therefore, it has been suggested that NR2, the putative sialic acid-binding domain of Hsa, is presented on the bacterial surface at the end of a long molecular stalk formed by SR2. The present review deals with the function and pathogenicity of oral streptococcal adhesins.
Assuntos
Adesinas Bacterianas/química , Adesinas Bacterianas/genética , Proteínas de Transporte/química , Proteínas de Transporte/genética , Infecções Estreptocócicas , Streptococcus gordonii/genética , Streptococcus gordonii/patogenicidade , Animais , Placa Dentária/microbiologia , Endocardite/microbiologia , Hemaglutininas Virais , Humanos , Ácido N-Acetilneuramínico/metabolismo , Ligação Proteica , Sinais Direcionadores de Proteínas/fisiologia , Estrutura Terciária de Proteína/fisiologiaRESUMO
Infective endocarditis is a typical biofilm-associated infectious disease frequently caused by commensal streptococci, but the contribution of host factors in biofilm formation is unclear. We found that platelets are essential for in vitro biofilm formation by Streptococcus mutans or Streptococcus gordonii grown in human plasma. The biofilms were composed of bacterial floes embedded with platelet aggregates in layers, and a similar architecture was also detected in situ on the injured valves of a rat model of experimental endocarditis. Similar to planktonic cells, the streptococci in biofilms were also able to induce platelet aggregation, which facilitates multilayer biofilm formation. Entrapping of platelets directly enhances the resistance of streptococcal biofilms to clindamycin. Prophylactic antibiotics or aspirin can reduce but not prevent or abolish biofilm formation on injured heart valves. Therefore, the platelet is a host factor for commensal streptococci in the circulation to consolidate biofilm formation and protect bacteria against antibiotics.