scholarly journals Pleiotropic Regulation of Virulence Genes in Streptococcus mutans by the Conserved Small Protein SprV

2017 ◽  
Vol 199 (8) ◽  
Author(s):  
Manoharan Shankar ◽  
Mohammad S. Hossain ◽  
Indranil Biswas
Keyword(s):  
Dental Caries ◽  
Streptococcus Mutans ◽  
Virulence Factors ◽  
Biofilm Formation ◽  
Normal Growth ◽  
Hypothetical Protein ◽  
Bacteriocin Production ◽  
Transcript Levels ◽  
Content Type ◽  
Tooth Surfaces

ABSTRACT Streptococcus mutans, an oral pathogen associated with dental caries, colonizes tooth surfaces as polymicrobial biofilms known as dental plaque. S. mutans expresses several virulence factors that allow the organism to tolerate environmental fluctuations and compete with other microorganisms. We recently identified a small hypothetical protein (90 amino acids) essential for the normal growth of the bacterium. Inactivation of the gene, SMU.2137, encoding this protein caused a significant growth defect and loss of various virulence-associated functions. An S. mutans strain lacking this gene was more sensitive to acid, temperature, osmotic, oxidative, and DNA damage-inducing stresses. In addition, we observed an altered protein profile and defects in biofilm formation, bacteriocin production, and natural competence development, possibly due to the fitness defect associated with SMU.2137 deletion. Transcriptome sequencing revealed that nearly 20% of the S. mutans genes were differentially expressed upon SMU.2137 deletion, thereby suggesting a pleiotropic effect. Therefore, we have renamed this hitherto uncharacterized gene as sprV (streptococcal pleiotropic regulator of virulence). The transcript levels of several relevant genes in the sprV mutant corroborated the phenotypes observed upon sprV deletion. Owing to its highly conserved nature, inactivation of the sprV ortholog in Streptococcus gordonii also resulted in poor growth and defective UV tolerance and competence development as in the case of S. mutans. Our experiments suggest that SprV is functionally distinct from its homologs identified by structure and sequence homology. Nonetheless, our current work is aimed at understanding the importance of SprV in the S. mutans biology. IMPORTANCE Streptococcus mutans employs several virulence factors and stress resistance mechanisms to colonize tooth surfaces and cause dental caries. Bacterial pathogenesis is generally controlled by regulators of fitness that are critical for successful disease establishment. Sometimes these regulators, which are potential targets for antimicrobials, are lost in the genomic context due to the lack of annotated homologs. This work outlines the regulatory impact of a small, highly conserved hypothetical protein, SprV, encoded by S. mutans. We show that SprV affects the transcript levels of various virulence factors required for normal growth, biofilm formation, stress tolerance, genetic competence, and bacteriocin production.

scholarly journals Disruption of L-Rhamnose Biosynthesis Results in Severe Growth Defects in Streptococcus mutans

2019 ◽  
Vol 202 (6) ◽  
Author(s):  
Andrew P. Bischer ◽  
Christopher J. Kovacs ◽  
Roberta C. Faustoferri ◽  
Robert G. Quivey
Keyword(s):  
Cell Wall ◽  
Dental Caries ◽  
Oral Cavity ◽  
Streptococcus Mutans ◽  
Biofilm Formation ◽  
Etiologic Agent ◽  
Growth Defect ◽  
Cell Wall Polysaccharide ◽  
Content Type ◽  
Severe Growth

ABSTRACT The rhamnose-glucose cell wall polysaccharide (RGP) of Streptococcus mutans plays a significant role in cell division, virulence, and stress protection. Prior studies examined function of the RGP using strains carrying deletions in the machinery involved in RGP assembly. In this study, we explored loss of the substrate for RGP, l-rhamnose, via deletion of rmlD (encoding the protein responsible for the terminal step in l-rhamnose biosynthesis). We demonstrate that loss of rhamnose biosynthesis causes a phenotype similar to strains with disrupted RGP assembly (ΔrgpG and ΔrgpF strains). Deletion of rmlD not only caused a severe growth defect under nonstress growth conditions but also elevated susceptibility of the strain to acid and oxidative stress, common conditions found in the oral cavity. A genetic complement of the ΔrmlD strain completely restored wild-type levels of growth, whereas addition of exogenous rhamnose did not. The loss of rhamnose production also significantly disrupted biofilm formation, an important aspect of S. mutans growth in the oral cavity. Further, we demonstrate that loss of either rmlD or rgpG results in ablation of rhamnose content in the S. mutans cell wall. Taken together, these results highlight the importance of rhamnose production in both the fitness and the ability of S. mutans to overcome environmental stresses. IMPORTANCE Streptococcus mutans is a pathogenic bacterium that is the primary etiologic agent of dental caries, a disease that affects billions yearly. Rhamnose biosynthesis is conserved not only in streptococcal species but in other Gram-positive, as well as Gram-negative, organisms. This study highlights the importance of rhamnose biosynthesis in RGP production for protection of the organism against acid and oxidative stresses, the two major stressors that the organism encounters in the oral cavity. Loss of RGP also severely impacts biofilm formation, the first step in the onset of dental caries. The high conservation of the rhamnose synthesis enzymes, as well as their importance in S. mutans and other organisms, makes them favorable antibiotic targets for the treatment of disease.

Lemon myrtle extract inhibits lactate production by Streptococcus mutans

2021 ◽  
Vol 85 (10) ◽  
pp. 2185-2190
Author(s):  
Yukinori Yabuta ◽  
Yui Sato ◽  
Arisu Miki ◽  
Ryuta Nagata ◽  
Tomohiro Bito ◽  
...  
Keyword(s):  
Lactate Dehydrogenase ◽  
Dental Caries ◽  
Oral Cavity ◽  
Streptococcus Mutans ◽  
Virulence Factors ◽  
Biofilm Formation ◽  
Tooth Enamel ◽  
Lactate Production ◽  
Tooth Decay ◽  
Lactate Dehydrogenase Activity

ABSTRACT Backhousia citriodora (lemon myrtle) extract has been found to inhibit glucansucrase activity, which plays an important role in biofilm formation by Streptococcus mutans. In addition to glucansucrase, various virulence factors in S. mutans are involved in the initiation of caries. Lactate produced by S. mutans demineralizes the tooth enamel. This study investigated whether lemon myrtle extract can inhibit S. mutans lactate production. Lemon myrtle extract reduced the glycolytic pH drop in S. mutans culture and inhibited lactate production by at least 46%. Ellagic acid, quercetin, hesperetin, and myricetin, major polyphenols in lemon myrtle, reduced the glycolytic pH drop and lactate production, but not lactate dehydrogenase activity. Furthermore, these polyphenols reduced the viable S. mutans cell count. Thus, lemon myrtle extracts may inhibit S. mutans-mediated acidification of the oral cavity, thereby preventing dental caries and tooth decay.

scholarly journals A New Small Molecule Specifically Inhibits the Cariogenic Bacterium Streptococcus mutans in Multispecies Biofilms

2011 ◽  
Vol 55 (6) ◽  
pp. 2679-2687 ◽  
Author(s):  
Chang Liu ◽  
Roberta J. Worthington ◽  
Christian Melander ◽  
Hui Wu
Keyword(s):  
Dental Caries ◽  
Small Molecules ◽  
Streptococcus Mutans ◽  
Small Molecule ◽  
Biofilm Formation ◽  
Content Type ◽  
Multispecies Biofilm ◽  
Small Molecule Library ◽  
Cariogenic Bacterium ◽  
Multispecies Biofilms

ABSTRACTStreptococcus mutansis a major cariogenic bacterium. It has adapted to the biofilm lifestyle, which is essential for pathogenesis of dental caries. We aimed to identify small molecules that can inhibit cariogenicS. mutansand to discover lead structures that could give rise to therapeutics for dental caries. In this study, we screened a focused small-molecule library of 506 compounds. Eight small molecules which inhibitedS. mutansat a concentration of 4 μM or less but did not affect cell growth or biofilm formation of commensal bacteria, represented byStreptococcus sanguinisandStreptococcus gordonii, in monospecies biofilms were identified. The active compounds share similar structural properties, which are characterized by a 2-aminoimidazole (2-AI) or 2-aminobenzimidazole (2-ABI) subunit. In multispecies biofilm models, the most active compound also inhibited cell survival and biofilm formation ofS. mutansbut did not affect commensal streptococci. This inhibitor downregulated the expression of six biofilm-associated genes,ftf,pac,relA,comDE,gbpB, andgtfB, in planktonicS. mutanscells, while it downregulated the expression of onlyftf,pac, andrelAin the biofilm cells ofS. mutans. The most potent compound also inhibited production of two key adhesins ofS. mutans, antigen I/II and glucosyltransferase (GTF). However, the compound did not alter the expression of the corresponding genes in bothS. sanguinisandS. gordonii, indicating that it possesses a selective inhibitory activity againstS. mutans.

scholarly journals Multiple Streptococcus mutans Genes Are Involved in Biofilm Formation

2002 ◽  
Vol 68 (12) ◽  
pp. 6283-6291 ◽  
Author(s):  
Akihiro Yoshida ◽  
Howard K. Kuramitsu
Keyword(s):  
Streptococcus Mutans ◽  
Dental Plaque ◽  
Biofilm Formation ◽  
Regulatory Gene ◽  
Hypothetical Protein ◽  
Abiotic Surfaces ◽  
Insertional Inactivation ◽  
Tooth Surfaces ◽  
Virulence Properties

ABSTRACT Streptococcus mutans has been strongly implicated as the principal etiological agent in dental caries. One of the important virulence properties of these organisms is their ability to form biofilms known as dental plaque on tooth surfaces. Since the roles of sucrose and glucosyltransferases in S. mutans biofilm formation have been well documented, we focused our attention on sucrose-independent factors. We have initially identified several mutants that appear to be defective in biofilm formation on abiotic surfaces by an insertional inactivation mutagenesis strategy applied to S. mutans. A total of 27 biofilm-defective mutants were isolated and analyzed in this study. From these mutants, three genes were identified. One of the mutants was defective in the Bacillus subtilis lytR homologue. Another of the biofilm-defective mutants isolated was a yulF homologue, which encodes a hypothetical protein of B. subtilis whose function in biofilm formation is unknown. The vast majority of the mutants were defective in the comB gene required for competence. We therefore have constructed and examined comACDE null mutants. These mutants were also found to be attenuated in biofilm formation. Biofilm formation by several other regulatory gene mutants were also characterized using an in vitro biofilm-forming assay. These results suggest that competence genes as well as the sgp and dgk genes may play important roles in S. mutans biofilm formation.

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 A Novel Gene Involved in the Survival of Streptococcus mutans under Stress Conditions

2013 ◽  
Vol 80 (1) ◽  
pp. 97-103 ◽  
Author(s):  
Dan Li ◽  
Yukie Shibata ◽  
Toru Takeshita ◽  
Yoshihisa Yamashita
Keyword(s):  
Streptococcus Mutans ◽  
Streptococcus Pyogenes ◽  
Biofilm Formation ◽  
Acid Tolerance ◽  
Stress Conditions ◽  
Insertion Mutagenesis ◽  
Trna Modification ◽  
Content Type ◽  
Virulence Proteins ◽  
Random Insertion

ABSTRACTAStreptococcus mutansmutant defective in aciduricity was constructed by random-insertion mutagenesis. Sequence analysis of the mutant revealed a mutation ingidA, which is known to be involved in tRNA modification inStreptococcus pyogenes. Complementation ofgidAbyS. pyogenesgidArecovered the acid tolerance ofS. mutans. Although thegidA-inactivatedS. pyogenesmutant exhibited significantly reduced expression of multiple extracellular virulence proteins, theS. mutansmutant did not. On the other hand, thegidAmutant ofS. mutansshowed reduced ability to withstand exposure to other stress conditions (high osmotic pressure, high temperature, and bacitracin stress) besides an acidic environment. In addition, loss of GidA decreased the capacity for glucose-dependent biofilm formation by over 50%. This study revealed thatgidAplays critical roles in the survival ofS. mutansunder stress conditions, including lower pH.

scholarly journals Oxidative Stressors Modify the Response of Streptococcus mutans to Its Competence Signal Peptides

2017 ◽  
Vol 83 (22) ◽  
Author(s):  
Matthew De Furio ◽  
Sang Joon Ahn ◽  
Robert A. Burne ◽  
Stephen J. Hagen
Keyword(s):  
Oxidative Stress ◽  
Reactive Oxygen Species ◽  
Dental Caries ◽  
Signaling Pathway ◽  
Streptococcus Mutans ◽  
Reactive Oxygen ◽  
Oxygen Species ◽  
Oral Biofilm ◽  
Genetic Competence ◽  
Content Type

ABSTRACTThe dental caries pathogenStreptococcus mutansis continually exposed to several types of stress in the oral biofilm environment. Oxidative stress generated by reactive oxygen species has a major impact on the establishment, persistence, and virulence ofS. mutans. Here, we combined fluorescent reporter-promoter fusions with single-cell imaging to study the effects of reactive oxygen species on activation of genetic competence inS. mutans. Exposure to paraquat, which generates superoxide anion, produced a qualitatively different effect on activation of expression of the gene for the master competence regulator, ComX, than did treatment with hydrogen peroxide (H2O2), which can yield hydroxyl radical. Paraquat suppressed peptide-mediated induction ofcomXin a progressive and cumulative fashion, whereas the response to H2O2displayed a strong threshold behavior. Low concentrations of H2O2had little effect on induction ofcomXor the bacteriocin genecipB, but expression of these genes declined sharply if extracellular H2O2exceeded a threshold concentration. These effects were not due to decreased reporter gene fluorescence. Two different threshold concentrations were observed in the response to H2O2, depending on the gene promoter that was analyzed and the pathway by which the competence regulon was stimulated. The results show that paraquat and H2O2affect theS. mutanscompetence signaling pathway differently, and that some portions of the competence signaling pathway are more sensitive to oxidative stress than others.IMPORTANCEStreptococcus mutansinhabits the oral biofilm, where it plays an important role in the development of dental caries. Environmental stresses such as oxidative stress influence the growth ofS. mutansand its important virulence-associated behaviors, such as genetic competence.S. mutanscompetence development is a complex behavior that involves two different signaling peptides and can exhibit cell-to-cell heterogeneity. Although oxidative stress is known to influenceS. mutanscompetence, it is not understood how oxidative stress interacts with the peptide signaling or affects heterogeneity. In this study, we used fluorescent reporters to probe the effect of reactive oxygen species on competence signaling at the single-cell level. Our data show that different reactive oxygen species have different effects onS. mutanscompetence, and that some portions of the signaling pathway are more acutely sensitive to oxidative stress than others.

scholarly journals Essential Roles of thesppRAFructose-Phosphate Phosphohydrolase Operon in Carbohydrate Metabolism and Virulence Expression byStreptococcus mutans

2018 ◽  
Vol 201 (2) ◽  
Author(s):  
Lin Zeng ◽  
Robert A. Burne
Keyword(s):  
Dental Caries ◽  
Organic Acids ◽  
Streptococcus Mutans ◽  
Biochemical Characterization ◽  
Constitutive Expression ◽  
Extracellular Dna ◽  
Tooth Surface ◽  
Oral Diseases ◽  
Sugar Phosphate ◽  
Content Type

ABSTRACTThe dental caries pathogenStreptococcus mutanscan ferment a variety of sugars to produce organic acids. Exposure ofS. mutansto certain nonmetabolizable carbohydrates, such as xylitol, impairs growth and can cause cell death. Recently, the presence of a sugar-phosphate stress inS. mutanswas demonstrated using a mutant lacking 1-phosphofructokinase (FruK) that accumulates fructose-1-phosphate (F-1-P). Here, we studied an operon inS. mutans,sppRA, which was highly expressed in thefruKmutant. Biochemical characterization of a recombinant SppA protein indicated that it possessed hexose-phosphate phosphohydrolase activity, with preferences for F-1-P and, to a lesser degree, fructose-6-phosphate (F-6-P). SppA activity was stimulated by Mg2+and Mn2+but inhibited by NaF. SppR, a DeoR family regulator, repressed the expression of thesppRAoperon to minimum levels in the absence of the fructose-derived metabolite F-1-P and likely also F-6-P. The accumulation of F-1-P, as a result of growth on fructose, not only inducedsppAexpression, but it significantly altered biofilm maturation through increased cell lysis and enhanced extracellular DNA release. Constitutive expression ofsppA, via a plasmid or by deletingsppR, greatly alleviated fructose-induced stress in afruKmutant, enhanced resistance to xylitol, and reversed the effects of fructose on biofilm formation. Finally, by identifying three additional putative phosphatases that are capable of promoting sugar-phosphate tolerance, we show thatS. mutansis capable of mounting a sugar-phosphate stress response by modulating the levels of certain glycolytic intermediates, functions that are interconnected with the ability of the organism to manifest key virulence behaviors.IMPORTANCEStreptococcus mutansis a major etiologic agent for dental caries, primarily due to its ability to form biofilms on the tooth surface and to convert carbohydrates into organic acids. We have discovered a two-gene operon inS. mutansthat regulates fructose metabolism by controlling the levels of fructose-1-phosphate, a potential signaling compound that affects bacterial behaviors. With fructose becoming increasingly common and abundant in the human diet, we reveal the ways that fructose may alter bacterial development, stress tolerance, and microbial ecology in the oral cavity to promote oral diseases.

scholarly journals Understanding the Basis of METH Mouth Using a Rodent Model of Methamphetamine Injection, Sugar Consumption, and Streptococcus mutans Infection

mBio ◽  
2021 ◽  
Vol 12 (2) ◽  
Author(s):  
Hiu Ham Lee ◽  
Preethi Sudhakara ◽  
Shreena Desai ◽  
Kildare Miranda ◽  
Luis R. Martinez
Keyword(s):  
Streptococcus Mutans ◽  
Biofilm Formation ◽  
Human Saliva ◽  
Tooth Decay ◽  
Content Type ◽  
Oral Rinse ◽  
Tooth Grinding ◽  
Oral Tissue ◽  
Public Health Strategies ◽  
The Impact

ABSTRACT “METH mouth” is a common consequence of chronic methamphetamine (METH) use, resulting in tooth decay and painful oral tissue inflammation that can progress to complete tooth loss. METH reduces the amount of saliva in the mouth, promoting bacterial growth, tooth decay, and oral tissue damage. This oral condition is worsened by METH users’ compulsive behavior, including high rates of consumption of sugary drinks, recurrent tooth grinding, and a lack of frequent oral hygiene. Streptococcus mutans is a Gram-positive bacterium found in the oral cavity and associated with caries in humans. Hence, we developed a murine model of METH administration, sugar intake, and S. mutans infection to mimic METH mouth in humans and to investigate the impact of this drug on tooth colonization. We demonstrated that the combination of METH and sucrose stimulates S. mutans tooth adhesion, growth, and biofilm formation in vivo. METH and sucrose increased the expression of S. mutans glycosyltransferases and lactic acid production. Moreover, METH contributes to the low environmental pH and S. mutans sucrose metabolism, providing a plausible mechanism for bacterium-mediated tooth decay. Daily oral rinse treatment with chlorhexidine significantly reduces tooth colonization in METH- and sucrose-treated mice. Furthermore, human saliva inhibits S. mutans colonization and biofilm formation after exposure to either sucrose or the combination of METH and sucrose. These findings suggest that METH might increase the risk of microbial dental disease in users, information that may help in the development of effective public health strategies to deal with this scourge in our society. IMPORTANCE “METH mouth” is characterized by severe tooth decay and gum disease, which often causes teeth to break or fall out. METH users are also prone to colonization by cariogenic bacteria such as Streptococcus mutans. In addition, this oral condition is aggravated by METH users’ compulsive behavior, including the consumption of beverages with high sugar content, recurrent tooth grinding, and a lack of frequent oral hygiene. We investigated the effects of METH and sugar consumption on S. mutans biofilm formation and tooth colonization. Using a murine model of METH administration, sucrose ingestion, and oral infection, we found that the combination of METH and sucrose increases S. mutans adhesion and biofilm formation on the teeth of C57BL/6 mice. However, daily chlorhexidine-based oral rinse treatment reduces S. mutans tooth colonization. Similarly, METH has been associated with dry mouth or hyposalivation in users. Hence, we assessed the impact of human saliva on biofilm formation and demonstrated that surface preconditioning with saliva substantially reduces S. mutans biofilm formation. Our results are significant because to our knowledge, this is the first basic science study focused on elucidating the fundamentals of METH mouth using a rodent model of prolonged drug injection and S. mutans oral infection. Our findings may have important translational implications for the development of treatments for the management of METH mouth and more effective preventive public health strategies that can be applied to provide effective dental care for METH users in prisons, drug treatment centers, and health clinics.

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