scholarly journals Amyloid Aggregation of Streptococcus mutans Cnm Influences Its Collagen-Binding Activity

2021 ◽  
Vol 87 (21) ◽  
Author(s):  
Nicholas M. di Cologna ◽  
Sandip Samaddar ◽  
Carolina A. Valle ◽  
Jonathan Vargas ◽  
Alejandro Aviles-Reyes ◽  
...  
Keyword(s):  
Streptococcus Mutans ◽  
Heart Valves ◽  
Binding Activity ◽  
The Body ◽  
Amyloid Aggregation ◽  
Collagen Binding ◽  
Content Type ◽  
Dental Biofilm ◽  
Laminin Binding ◽  
Binding Glycoprotein

Streptococcus mutans is a keystone pathogen that promotes caries by acidifying the dental biofilm milieu. The collagen- and laminin-binding glycoprotein Cnm is a virulence factor of S. mutans . Expression of Cnm by S. mutans is hypothesized to contribute to niche expansion, allowing colonization of multiple sites in the body, including collagen-rich surfaces such as dentin and heart valves.

scholarly journals Trk2 Potassium Transport System in Streptococcus mutans and Its Role in Potassium Homeostasis, Biofilm Formation, and Stress Tolerance

2016 ◽  
Vol 198 (7) ◽  
pp. 1087-1100 ◽  
Author(s):  
Gursonika Binepal ◽  
Kamal Gill ◽  
Paula Crowley ◽  
Martha Cordova ◽  
L. Jeannine Brady ◽  
...  
Keyword(s):  
Stress Tolerance ◽  
Streptococcus Mutans ◽  
Transport System ◽  
Biofilm Formation ◽  
Cellular Response ◽  
Pathogenic Bacteria ◽  
Transport Systems ◽  
Content Type ◽  
Growth And Survival ◽  
Dental Biofilm

ABSTRACTPotassium (K+) is the most abundant cation in the fluids of dental biofilm. The biochemical and biophysical functions of K+and a variety of K+transport systems have been studied for most pathogenic bacteria but not for oral pathogens. In this study, we establish the modes of K+acquisition inStreptococcus mutansand the importance of K+homeostasis for its virulence attributes. TheS. mutansgenome harbors four putative K+transport systems that included two Trk-like transporters (designated Trk1 and Trk2), one glutamate/K+cotransporter (GlnQHMP), and a channel-like K+transport system (Kch). Mutants lacking Trk2 had significantly impaired growth, acidogenicity, aciduricity, and biofilm formation. [K+] less than 5 mM eliminated biofilm formation inS. mutans. The functionality of the Trk2 system was confirmed by complementing anEscherichia coliTK2420 mutant strain, which resulted in significant K+accumulation, improved growth, and survival under stress. Taken together, these results suggest that Trk2 is the main facet of the K+-dependent cellular response ofS. mutansto environment stresses.IMPORTANCEBiofilm formation and stress tolerance are important virulence properties of caries-causingStreptococcus mutans. To limit these properties of this bacterium, it is imperative to understand its survival mechanisms. Potassium is the most abundant cation in dental plaque, the natural environment ofS. mutans. K+is known to function in stress tolerance, and bacteria have specialized mechanisms for its uptake. However, there are no reports to identify or characterize specific K+transporters inS. mutans. We identified the most important system for K+homeostasis and its role in the biofilm formation, stress tolerance, and growth. We also show the requirement of environmental K+for the activity of biofilm-forming enzymes, which explains why such high levels of K+would favor biofilm formation.

scholarly journals Competition and Caries on Enamel of a Dual-Species Biofilm Model with Streptococcus mutans and Streptococcus sanguinis

2020 ◽  
Vol 86 (21) ◽  
Author(s):  
Natalia Díaz-Garrido ◽  
Carla P. Lozano ◽  
Jens Kreth ◽  
Rodrigo A. Giacaman
Keyword(s):  
Streptococcus Mutans ◽  
Single Species ◽  
Viable Count ◽  
Noncommunicable Disease ◽  
Continuous Exposure ◽  
Content Type ◽  
Streptococcus Sanguinis ◽  
Dental Biofilm ◽  
Intense Competition ◽  
Dental Surface

ABSTRACT Imbalances within the dental biofilm trigger dental caries, currently considered a dysbiosis and the most prevalent noncommunicable disease. There is still a gap in knowledge about the dynamics of enamel colonization by bacteria from the dental biofilm in caries. The aim, therefore, was to test whether the sequence of enamel colonization by a typically commensal and a cariogenic species modifies biofilm’s cariogenicity. Dual-species biofilms of Streptococcus mutans and Streptococcus sanguinis on saliva-coated enamel slabs were inoculated in different sequences: S. mutans followed by S. sanguinis (Sm-Ss), S. sanguinis followed by S. mutans (Ss-Sm), S. mutans and S. sanguinis inoculated at the same time (Sm=Ss), and the single-species controls S. mutans followed by S. mutans (Sm-Sm) and S. sanguinis followed by S. sanguinis (Ss-Ss). Biofilms were exposed to 10% sucrose 3 times per day for 5 days, and the slabs/biofilms were retrieved to assess demineralization, viable cells, biomass, proteins, polysaccharides, and H2O2 production. Compared with Sm-Sm, primary inoculation with S. sanguinis reduced demineralization (P < 0.05). Both Ss-Sm and Sm=Ss sequences showed reduction in biomass, protein, and polysaccharide content (P < 0.05). The highest S. sanguinis viable count and H2O2 production level and the lowest acidogenicity were observed when S. sanguinis colonized enamel before S. mutans (P < 0.05). Initial enamel adherence with commensal biofilms seems to induce more intense competition against more typically cariogenic species, reducing cariogenicity. IMPORTANCE The concept of caries as an ecological disease implies the understanding of the intricate relationships among the populating microorganisms. Under frequent sugar exposure, some bacteria from the dental biofilm develop pathogenic traits that lead to imbalances (dysbiosis). Depending on which microorganism colonizes the dental surface first, different competition strategies may be developed. Studying the interactions in the entire dental biofilm is not an easy task. In this study, therefore, we modeled the interplay among these microorganisms using a caries-inducing species (S. mutans) and a health-associated species (S. sanguinis). Initial enamel adherence with S. sanguinis seems to induce more intense competition against typically caries-inducing species. Besides continuous exposure with sugars, early colonization of the enamel by highly cariogenic species like S. mutans appears to be needed to develop caries lesions as well. Promoting early colonization by health-associated bacteria such as S. sanguinis could help to maintain oral health, delaying dysbiosis.

scholarly journals Contribution of the Interaction of Streptococcus mutans SerotypekStrains with Fibrinogen to the Pathogenicity of Infective Endocarditis

2014 ◽  
Vol 82 (12) ◽  
pp. 5223-5234 ◽  
Author(s):  
Ryota Nomura ◽  
Masatoshi Otsugu ◽  
Shuhei Naka ◽  
Noboru Teramoto ◽  
Ayuchi Kojima ◽  
...  
Keyword(s):  
Infective Endocarditis ◽  
Streptococcus Mutans ◽  
Heart Valve ◽  
Heart Valves ◽  
Binding Assays ◽  
Bacterial Dna ◽  
Content Type ◽  
Fibrinogen Binding ◽  
Level Of Aggregation

ABSTRACTStreptococcus mutans, a pathogen responsible for dental caries, is occasionally isolated from the blood of patients with bacteremia and infective endocarditis (IE). Our previous study demonstrated that serotypek-specific bacterial DNA is frequently detected inS. mutans-positive heart valve specimens extirpated from IE patients. However, the reason for this frequent detection remains unknown. In the present study, we analyzed the virulence of IE fromS. mutansstrains, focusing on the characterization of serotypekstrains, most of which are positive for the 120-kDa cell surface collagen-binding protein Cbm and negative for the 190-kDa protein antigen (PA) known as SpaP, P1, antigen I/II, and other designations. Fibrinogen-binding assays were performed with 85 clinical strains classified by Cbm and PA expression levels. The Cbm+/PA−group strains had significantly higher fibrinogen-binding rates than the other groups. Analysis of platelet aggregation revealed that SA31, a Cbm+/PA−strain, induced an increased level of aggregation in the presence of fibrinogen, while negligible aggregation was induced by the Cbm-defective isogenic mutant SA31CBD. A rat IE model with an artificial impairment of the aortic valve created using a catheter showed that extirpated heart valves in the SA31 group displayed a prominent vegetation mass not seen in those in the SA31CBD group. These findings could explain why Cbm+/PA−strains are highly virulent and are related to the development of IE, and the findings could also explain the frequent detection of serotypekDNA inS. mutans-positive heart valve clinical specimens.

scholarly journals A Nonfimbrial Adhesin ofAggregatibacter actinomycetemcomitansMediates Biofilm Biogenesis

2018 ◽  
Vol 87 (1) ◽  
Author(s):  
David R. Danforth ◽  
Gaoyan Tang-Siegel ◽  
Teresa Ruiz ◽  
Keith P. Mintz
Keyword(s):  
Biofilm Formation ◽  
Protein A ◽  
Three Dimensional ◽  
Binding Activity ◽  
Periodontal Pathogen ◽  
Collagen Binding ◽  
Content Type ◽  
Mutant Strains ◽  
Different Strains ◽  
Biofilm Development

ABSTRACTPeriodontitis is an inflammatory disease caused by polymicrobial biofilms. The periodontal pathogenAggregatibacter actinomycetemcomitansdisplays two proteinaceous surface structures, the fimbriae and the nonfimbrial extracellular matrix binding protein A (EmaA), as observed by electron microscopy. Fimbriae participate in biofilm biogenesis and the EmaA adhesins mediate collagen binding. However, in the absence of fimbriae,A. actinomycetemcomitansstill retains the potential to form robust biofilms, suggesting that other surface macromolecules participate in biofilm development. Here, isogenic mutant strains lacking EmaA structures, but still expressing fimbriae, were observed to have reduced biofilm potential. In strains lacking both EmaA and fimbriae, biofilm mass was reduced by 80%. EmaA enhanced biofilm formation in different strains, independent of the fimbriation state or serotype. Confocal microscopy revealed differences in cell density within microcolonies between the EmaA positive and mutant strains. EmaA-mediated biofilm formation was found to be independent of the glycosylation state and the precise three-dimensional conformation of the protein, and thus this function is uncorrelated with collagen binding activity. The data suggest that EmaA is a multifunctional adhesin that utilizes different mechanisms to enhance bacterial binding to collagen and to enhance biofilm formation, both of which are important forA. actinomycetemcomitanscolonization and subsequent infection.

scholarly journals O-Polysaccharide Glycosylation Is Required for Stability and Function of the Collagen Adhesin EmaA of Aggregatibacter actinomycetemcomitans

2012 ◽  
Vol 80 (8) ◽  
pp. 2868-2877 ◽  
Author(s):  
Gaoyan Tang ◽  
Teresa Ruiz ◽  
Keith P. Mintz
Keyword(s):  
Cellular Localization ◽  
Proteolytic Enzymes ◽  
Aggregatibacter Actinomycetemcomitans ◽  
Binding Activity ◽  
Mrna Levels ◽  
Collagen Binding ◽  
Content Type ◽  
Protein Levels ◽  
Mutant Strains ◽  
And Function

ABSTRACTAggregatibacter actinomycetemcomitansis hypothesized to colonize through the interaction with collagen and establish a reservoir for further dissemination. The trimeric adhesin EmaA ofA. actinomycetemcomitansbinds to collagen and is modified with sugars mediated by an O-antigen polysaccharide ligase (WaaL) that is associated with lipopolysaccharide (LPS) biosynthesis (G. Tang and K. Mintz, J. Bacteriol.192:1395–1404, 2010). This investigation characterized the function and cellular localization of EmaA glycosylation. The interruption of LPS biogenesis by using genetic and pharmacological methods changed the amount and biophysical properties of EmaA molecules in the outer membrane. InrmlCandwaaLmutant strains, the membrane-associated EmaA was reduced by 50% compared with the wild-type strain, without changes in mRNA levels. The membrane-associated EmaA protein levels were recovered by complementation with the corresponding O-polysaccharide (O-PS) biosynthetic genes. In contrast, another trimeric autotransporter, epithelial adhesin ApiA, was not affected in the same mutant background. The inhibition of undecaprenyl pyrophosphate recycling by bacitracin resulted in a similar decrease in the membrane-associated EmaA protein. This effect was reversed by removal of the compound. A significant decrease in collagen binding activity was observed in strains expressing the nonglycosylated form of EmaA. Furthermore, the electrophoretic mobility shifts of the EmaA monomers found in the O-PS mutant strains were associated only with the membrane-associated protein and not with the cytoplasmic pre-EmaA protein, suggesting that this modification does not occur in the cytoplasm. The glycan modification of EmaA appears to be required for collagen binding activity and protection of the protein against degradation by proteolytic enzymes.

scholarly journals Contribution of Streptococcus mutans Strains with Collagen-Binding Proteins in the Presence of Serum to the Pathogenesis of Infective Endocarditis

2017 ◽  
Vol 85 (12) ◽  
Author(s):  
Masatoshi Otsugu ◽  
Ryota Nomura ◽  
Saaya Matayoshi ◽  
Noboru Teramoto ◽  
Kazuhiko Nakano
Keyword(s):  
Infective Endocarditis ◽  
Streptococcus Mutans ◽  
Pathogenic Bacteria ◽  
Matrix Protein ◽  
Binding Proteins ◽  
Ex Vivo ◽  
Extracellular Matrix Protein ◽  
Dna Encoding ◽  
Collagen Binding ◽  
Content Type

ABSTRACT Streptococcus mutans, a major pathogen of dental caries, is considered one of the causative agents of infective endocarditis (IE). Recently, bacterial DNA encoding 120-kDa cell surface collagen-binding proteins (CBPs) has frequently been detected from S. mutans-positive IE patients. In addition, some of the CBP-positive S. mutans strains lacked a 190-kDa protein antigen (PA), whose absence strengthened the adhesion to and invasion of endothelial cells. The interaction between pathogenic bacteria and serum or plasma is considered an important virulence factor in developing systemic diseases; thus, we decided to analyze the pathogenesis of IE induced by S. mutans strains with different patterns of CBP and PA expression by focusing on the interaction with serum or plasma. CBP-positive (CBP+)/PA-negative (PA−) strains showed prominent aggregation in the presence of human serum or plasma, which was significantly greater than that with CBP+/PA-positive (PA+) and CBP-negative (CBP−)/PA+ strains. Aggregation of CBP+/PA− strains was also observed in the presence of a high concentration of type IV collagen, a major extracellular matrix protein in serum. In addition, aggregation of CBP+/PA− strains was drastically reduced when serum complement was inactivated. Furthermore, an ex vivo adherence model and an in vivo rat model of IE showed that extirpated heart valves infected with CBP+/PA− strains displayed prominent bacterial mass formation, which was not observed following infection with CBP+/PA+ and CBP−/PA+ strains. These results suggest that CBP+/PA− S. mutans strains utilize serum to contribute to their pathogenicity in IE.

scholarly journals The Collagen Binding Protein Cnm Contributes to Oral Colonization and Cariogenicity of Streptococcus mutans OMZ175

2015 ◽  
Vol 83 (5) ◽  
pp. 2001-2010 ◽  
Author(s):  
James H. Miller ◽  
Alejandro Avilés-Reyes ◽  
Kathy Scott-Anne ◽  
Stacy Gregoire ◽  
Gene E. Watson ◽  
...  
Keyword(s):  
Dental Caries ◽  
Streptococcus Mutans ◽  
Binding Protein ◽  
Oral Epithelium ◽  
Pathogenic Potential ◽  
Collagen Binding ◽  
Native Strain ◽  
Content Type ◽  
Root Tissues

Streptococcus mutansis the etiological agent of dental caries and one of the many bacterial species implicated in infective endocarditis. The expression of the collagen-binding protein Cnm byS. mutanshas been associated with extraoral infections, but its relevance for dental caries has only been theorized to date. Due to the collagenous composition of dentinal and root tissues, we hypothesized that Cnm may facilitate the colonization of these surfaces, thereby enhancing the pathogenic potential ofS. mutansin advancing carious lesions. As shown for extraoral endothelial cell lines, Cnm mediates the invasion of oral keratinocytes and fibroblasts byS. mutans. In this study, we show that in the Cnm+native strain, OMZ175, Cnm mediates stringent adhesion to dentinal and root tissues as well as collagen-coated surfaces and promotes both cariogenicity and carriagein vivo. In vitro,ex vivo, andin vivoexperiments revealed that while Cnm is not universally required forS. mutanscariogenicity, it contributes to (i) the invasion of the oral epithelium, (ii) enhanced binding on collagenous surfaces, (iii) implantation of oral biofilms, and (IV) the severity of caries due to a native Cnm+isolate. Taken together, our findings reveal that Cnm is a colonization factor that contributes to the pathogenicity of certainS. mutansstrains in their native habitat, the oral cavity.

scholarly journals Amyloid aggregation of Streptococcus mutans Cnm influences its collagen-binding activity

2021 ◽  
Author(s):  
Nicholas de Mojana di Cologna ◽  
Sandip Samaddar ◽  
Carolina Valle ◽  
Jonathan A Vargas ◽  
Alejandro Aviles-Reyes ◽  
...  
Keyword(s):  
Quaternary Structure ◽  
Ex Vivo ◽  
In Silico Analysis ◽  
Binding Activity ◽  
Amyloid Aggregation ◽  
Binding Assays ◽  
Collagen Binding ◽  
Microtiter Plates

The glycosylated collagen- and laminin-binding surface adhesin Cnm is present in approximately 20% of S. mutans clinical isolates and is associated with systemic infections and increased caries risk. Other surface-associated collagen-binding proteins of S. mutans such as P1 and WapA have been demonstrated to form an amyloid quaternary structure with functional implications within biofilms. In silico analysis predicted that the b-sheet rich N-terminal collagen-binding domain (CBD) of Cnm has propensity for amyloid aggregation, whereas the threonine-rich C-terminal domain was predicted to be disorganized. In this study, thioflavin-T fluorescence and electron microscopy were used to show that Cnm forms amyloids either in its native glycosylated or recombinant non-glycosylated forms and that the CBD of Cnm is the main amyloidogenic unit of Cnm. We then performed a series of in vitro, ex vivo and in vivo assays to characterize the amylogenic properties of Cnm.  In addition, Congo red birefringence indicated that Cnm is a major amyloidogenic protein of S. mutans biofilms. Competitive binding assays using collagen-coated microtiter plates and dental roots, a substrate rich in collagen, revealed that Cnm monomers inhibit S. mutans binding to collagenous substrates whereas Cnm amyloid aggregates lose this property. Thus, while Cnm contributes to recognition and initial binding of S. mutans to collagen-rich surfaces, Cnm amyloid aggregation appears to represent a mechanism to modulate this activity in mature biofilms.

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