Streptococcus Mutans Biofilm Formation Research Articles

Evaluation of Streptococcus mutans biofilm formation and acidogenicity of infant milk formulas for treating cow milk allergy: An in vitro study.

When infants cannot consume breast milk, the most commonly available alternative milk formula is cow milk-based. Due to a rise in the prevalence of cow milk protein allergy (CMPA) among children, this study aimed to assess the biofilm formation and acidogenicity of cow milk-based formulas as well as milk formulas suggested for children with CMPA. Cow milk-based formulas with 0%, 10%, or 18% sucrose added, partially hydrolyzed formula (pHF), extensively hydrolyzed formula (eHF), amino acid-based formula (AAF), and soy-based formulas with 0%, or 11% sucrose added were evaluated. Streptococcus mutans was used as a representative microorganism associated with caries. The acidogenicity after 24-h incubation was assessed by the pH of the formed biofilm and lactic acid formation. Biofilm formation was quantified using crystal violet staining. Additionally, the biofilm characteristics were determined using confocal laser scanning microscopy (CLSM). Comparisons were made among formulas without added sucrose to observe protein-based differences. Furthermore, formulas with different sucrose percentages were compared to explore the impact of sucrose content. When comparing the formulas without added sucrose, the biofilm formation in the cow milk-based formula and pHF were significantly greater than the soy-based formula, eHF, and AAF. In the presence of S. mutans, all formulas reduced the biofilm pH below the critical enamel pH. The cow milk-based formula and AAF showed a significantly lower biofilm pH than the pHF, soy-based, and eHF groups, while the lactic acid production was markedly higher in the cow milk-based formula, pHF and AAF, compared with the eHF and soy-based formula. Adding sucrose into the cow milk-based and soy-based formulas substantially increased biofilm mass. The biofilm pH of the cow milk-based formulas, with or without sucrose, was significantly lower than that of the soy-based formulas. The CLSM indicated distinct biofilm characteristics among the different protein-based formulas, with sucrose supplementation promoting S. mutans aggregation in cow milk-based formula biofilm and increased density and intact biofilm in the soy-based formula. All assessed milk formulas had caries-inducing factors, including those without supplemental sucrose. Among them, the eHF demonstrated the least caries-inducing factors, attributed to its minimal biofilm formation and the highest biofilm pH.

Comparative evaluation of anti-biofilm and anti- adherence potential of plant extracts against Streptococcus mutans: A therapeutic approach for oral health

Dental caries predominantly attributed to the cariogenic nature of Streptococcus mutans, continue to pose a substantial global challenge to oral health. In response to this challenge, this study aimed to evaluate the effectiveness of leaf extracts (LEs) and essential oils (EOs) derived from different medicinal plants in inhibiting the growth of Streptococcus mutans biofilm. In vitro and in silico approaches were employed to identify active compounds and assess their inhibitory effects on S. mutans. Minimum Inhibitory Concentration (MIC) and Minimum Bactericidal Concentration (MBC) were measured to determine the anti-biofilm and anti-adherence activity against S. mutans. Biofilm viability (CFU/mL) and extracellular polymeric substance (EPS) concentration were quantified. GC-MS analysis was utilized to identify active compounds in the most effective plant extracts exhibiting anti-S. mutans activity. A high-throughput screening focused on the interaction between these compounds and the target enzyme SortaseA (SrtA) using molecular docking was performed. Results indicated that Cymbopogon citratus displayed the highest efficacy in reducing S. mutans biofilm formation and adhesion activity, achieving 90 % inhibition at an MIC value of 12 μg/mL. Among the 12 bioactive compounds identified, trans-Carvyl acetate exhibited the lowest binding energy with SrtA (-6.0 Kcal/mole). Trans-Carvyl acetate also displayed favorable pharmacokinetic properties. This study provides novel insights into the anti-S. mutans properties of C. citratus and suggests its potential as a therapeutic approach for oral health. Further research is needed to explore the combined effect of plant extracts for enhanced protection against dental caries.

Antibiofilm Properties and Demineralization Suppression in Early Enamel Lesions Using Dental Coating Materials.

This study aimed to investigate the effects of dental coating materials on Streptococcus mutans biofilm formation. The test materials were PRG Barrier Coat (PRG), BioCoat Ca (BioC), and FluorDental Jelly (FluorJ). Bovine enamel specimens were demineralized to mimic early enamel lesions. The biofilm was developed on a specimen treated with one of the materials by using a modified Robbins device flow-cell system. Scanning electron and fluorescence confocal laser scanning microscopy, viable and total cell counts, and gene expression assessments of the antibiofilm were performed. Ion incorporation was analyzed using a wavelength-dispersive X-ray spectroscopy electron probe microanalyzer. All materials allowed biofilm formation but reduced its volume. FluorJ was the only material that inhibited biofilm accumulation and had a bactericidal effect, revealing 0.66 log CFU in viable cells and 1.23 log copy reduction in total cells compared with the untreated group after 24 h of incubation. The ions released from PRG varied depending on the element. BioC contributed to enamel remineralization by supplying calcium ions while blocking the acid produced from the biofilm. In summary, the dental coating materials physically prevented acid attacks from the biofilm while providing ions to the enamel to improve its mechanical properties.

Inhibition of Streptococcus mutans growth and biofilm formation through protein acetylation.

Numerous cellular processes are regulated in response to the metabolic state of the cell, and one such regulatory mechanism involves lysine acetylation. Lysine acetylation has been proven to play an important role in the virulence of Streptococcus mutans, a major cariogenic bacterial species. S. mutans' glucosyltransferases (Gtfs) are responsible for synthesizing extracellular polysaccharides (EPS) and contributing to biofilm formation. One of the most common nonsteroidal anti-inflammatory drugs is acetylsalicylic acid (ASA), which can acetylate proteins through a nonenzymatic transacetylation reaction. Herein, we investigated the inhibitory effects of ASA on S. mutans. ASA treatment was observed to impede the growth of S. mutans, leading to a reduction in the production of water-insoluble EPS and the formation of biofilm. Moreover, ASA decreased the enzyme activity of Gtfs while increasing the protein acetylation level. The in vivo anticaries efficacy of ASA has further been proved using the rat caries model. In conclusion, ASA as an acetylation agent attenuated the cariogenic virulence of S. mutans, suggesting the potential value of protein acetylation on antimicrobial and anti-biofilm applications to S. mutans.

[Role and related mechanisms of LiaSR two-component system in acid tolerance and biofilm formation of Streptococcus mutans].

Objective: To investigate the role and related mechanisms of the LiaSR two-component system in acid tolerance and biofilm formation abilities of Streptococcus mutans (Sm) 593. Methods: The growth curves of various Sm strains in pH=5.5 brian heart infusion (BHI) medium were analyzed. And colony forming unit (CFU) was also performed to evaluate the acid tolerance of Sm. Laurdan probe, H+-K+adenosine triphosphate (ATP)ase activity analysis kit, proton permeability assay and real-time fluorescence quantitative PCR (RT-qPCR) were conducted to detect the acid tolerant mechanisms of LiaSR two-component system in Sm. Crystal violet staining, CFU, SYTOX probe and anthrone-sulfuric method were used to analyze the properties and structures of the Sm biofilms. RT-qPCR was conducted to detect the expression levels of underlying regulated genes. Results: The growth of mutants in acidic BHI were inhibited (P<0.05). The acid tolerance of mutants significantly decreased compared to the wild-type strain (P<0.05). In mutants, the activity of H+-ATPase (917.06±59.53 and 469.53±47.65) were elevated by 7.22-folds and 3.70-folds compared to the wild-type strain (127.00±50.71) (P<0.001, P<0.001) and the encoded gene atpD (3.39±0.21 and 1.94±0.17) were also elevated by 3.39-folds and 1.94-folds compared to the wild-type strain (1.00±0.15) (P<0.001, P=0.001). The Laurdan generalized polarization of mutants (0.18±0.04 and 0.18±0.05) increased significantly compared to the wild-type strain (0.08±0.05) (P=0.006, P=0.003) and the expression levels of fabM gene were decreased in mutants (0.52±0.11 and 0.57±0.05) by 1/2 (P=0.014, P=0.022). In liaR deletion mutant, the reduced terminal pH (4.76±0.01) can also be observed (P<0.001). The total amount of the biofilms of three Sm didn't show significant differences (P>0.05). But the number of viable bacteria of mutants' biofilms were decreased [Sm 593: (12.00±2.80)×107 CFU/ml; Sm ΔliaS: (2.95±1.13)×107 CFU/ml; Sm ΔliaR: (7.25±1.60)×107 CFU/ml] (P=0.001, P=0.024). The extracellular DNA were increased by 18.00-folds and 6.50-folds in mutants' biofilms (128.73±15.65 and 46.38±5.52) compared to the wild-type strain (7.16±3.62) (P<0.001, P=0.003). Water-soluble exopolysaccharides could be found up-regulated in liaS deletion mutant [(138.73±10.12) μg/ml] (P=0.003) along with the expression level of gtfC gene (1.65±0.39) (P=0.014). The expression level of gtfD were elevated by 47.43-folds and 16.90-folds in mutants (P<0.001, P=0.010). Conclusions: The LiaSR two-component system can promote the expression of fabM gene and increase the fluidity of Sm which contributes to acid tolerance. The LiaR can also decrease the proton permeability and restrict the entrance of H+. The LiaSR two-component system can negatively regulate the production of the extracellular matrix in Sm biofilm.

[Effect of a novel pH-responsive tertiary amine monomer dodecylmethylaminoethyl methacrylate modified resin adhesive on biofilm formation of Streptococcus mutans and Lactobacillus casei in vitro].

Objective: To explore the application prospect of a new pH-responsive tertiary amine monomer dodecylmethylaminoethyl methacrylate (DMAEM) modified resin adhesive (DMAEM@RA) in the prevention and treatment of secondary caries. Methods: Five percents DMAEM was added to the resin adhesive to synthesize DMAEM@RA for modifying. Streptococcus mutans (Sm) and Lactobacillus casei (Lc) biofilms were cultured on resin adhesive and DMAEM@RA, respectively. The culture systems were set up at pH=7.4, 6.0, 5.5, and 5.0. The antimicrobial activity of DMAEM@RA was evaluated by quantitative PCR. The effects of DMAEM@RA on biofilm thickness, bacterial amount, and extracellular polysaccharides were studied by scanning electron microscope (SEM) and extracellular polysaccharide staining. Real-time fluorescence quantitative PCR was used to study the effect of DMAEM@RA on the expression levels of cariogenic genes in Sm. Results: DMAEM@RA could significantly reduce the amount of Sm and Lc under acidic conditions, especially Lc. At pH=5.0, the logarithm value of co-cultured Sm bacteria [lg (CFU/ml)] in DMAEM@RA group (7.58±0.01) was significantly lower than that in control group (7.87±0.03) (t=14.32, P<0.001), and the logarithm value of Lc bacteria [lg (CFU/ml)] (7.29±0.04) was also significantly lower than that in control group (7.93±0.15) (t=6.93, P=0.002). SEM observed that the bacteria decreased and the cell fragments appeared in DMAEM@RA group. In addition, DMAEM@RA significantly reduced the biomass of extracellular polysaccharides in the dual-species biofilm under acidic conditions. At pH=5.0, the biomass of extracellular polysaccharides in DMAEM@RA group [(25.13±3.14) mm3/mm2] was significantly lower than that in the control group [(42.66±7.46) mm3/mm2] (t=3.75, P=0.020). DMAEM@RA could significantly up-regulate the expressions of gtfB and gtfC genes in Sm under acidic conditions. At pH=5.0, gtfB and gtfC genes were significantly up-regulated by (14.64± 0.44) times and (2.99±0.20) times, respectively (t=-42.74, P<0.001; t=-13.55, P<0.001). Conclusions: The DMAEM@RA has a good antibacterial effect under acidic conditions, demonstrating that it has a good potential to prevent the occurrence and development of secondary caries.

Inhibitory effects of Bacillus velezensis ID-A01 supernatant against Streptococcus mutans

BackgroundDental caries is a chronic oral disease caused by microbial infections, which result in erosion of the dental enamel and cause irreversible damage. Therefore, proper disease management techniques and the creation of an environment that prevents intraoral growth and biofilm formation of Streptococcus mutans in the early stages, are crucial to prevent the potential progression of dental plaque to disease. Here, we aimed to investigate antimicrobial and antibiofilm effects of the Bacillus velezensis ID-A01 supernatant (ID23029) against S. mutans, and its inhibitory effects on acidogenesis.ResultsA killing kinetics assay showed a peak lethality percentage of 94.5% after 6 h of exposure to ID23029. In sucrose-exposed conditions, ID23029 inhibited lactic acid formation, preventing the pH from falling below the threshold for enamel demineralization, and inhibited up to 96.6% of biofilm formation. This effect was maintained in the presence of lysozyme. Furthermore, ID23029 retained up to 92% lethality, even at an intraoral concentration at which lysozyme is ineffective against S. mutans.ConclusionsThis study demonstrates the potential of the B. velezensis ID-A01 supernatant for the prevention and treatment of dental caries. Its eventual use in dental practice is encouraged, although further studies are required to confirm its beneficial effects.

Recombinant Prevotella melaninogenica α-1,3 glucanase and Capnocytophaga ochracea α-1,6 glucanase as enzymatic tools for in vitro degradation of S. mutans biofilms.

Dental biofilms represent a serious oral health problem playing a key role in the development of caries and other oral diseases. In the present work, we cloned and expressed in E. coli two glucanases, Prevotella melaninogenica mutanase (PmGH87) and Capnocytophaga ochracea dextranase (CoGH66), and characterized them biochemically and biophysically. Their three-dimensional structures were elucidated and discussed. Furthermore, we tested the capacity of the enzymes to hydrolyze mutan and dextran to prevent formation of Streptococcus mutans biofilms, as well as to degrade pre- formed biofilms in low and abundant sugar conditions. The percentage of residual biofilm was calculated for each treatment group in relation to the control, as well as the degree of synergism. Our results suggest that both PmGH87 and CoGH66 are capable of inhibiting biofilm formation grown under limited or abundant sucrose conditions. Degradation of pre-formed biofilms experiments reveal a time-dependent effect for the treatment with each enzyme alone. In addition, a synergistic and dose-dependent effects of the combined enzymatic treatment with the enzymes were observed. For instance, the highest biomass degradation was 95.5% after 30min treatment for the biofilm grown in low sucrose concentration, and 93.8% after 2h treatment for the biofilm grown in sugar abundant condition. Strong synergistic effects were observed, with calculated degree of synergism of 5.54 and 3.18, respectively and their structural basis was discussed. Jointly, these data can pave the ground for the development of biomedical applications of the enzymes for controlling growth and promoting degradation of established oral biofilms.

Biomimetic Approach to Counter Streptococcus mutans Biofilm: An In Vitro Study on Seashells.

Aim This study aimed to investigate the anti-adherent property of the seashell surface and periostracum to prevent the formation of Streptococcus mutans biofilm. Materials and methods The seashells were initially collected from the natural urban beach, and an antibiofilm assay of the shells with and without periostracum was performed against Streptococcus mutans. Furthermore, the seashells were analyzed with a stylus profilometer (Mitutoyo Surftest SJ-301, Mitutoyo America Corporation, Illinois, USA), atomic force microscope (AFM; Nanosurf Easyscan 2, Nanosurf Inc., USA), contact angle assessment, Fourier-transform infrared (FTIR) spectroscopy analysis, and scanning electron microscopy (SEM; JEOL USA, Inc., FE-SEM IT800, Massachusetts, USA) analysis. The ability of seashells to prevent the attachment of Streptococcus mutans and form a biofilm with and without periostracum was studied by crystal violet assay. Results The results revealed that shells without periostracum promoted higher biofilm formation when compared to those having intact periostracum (by 15%, p<0.001). Shell 1 showed the highest biofilm formation, whereas shell 3 showed the least biofilm formation due to the differences in their surface morphologies. The remaining shells (4, 2, 6, and 5) showed interspersed biofilm formation. Conclusion In summary, our study was able to correlate the topologies of the shell surface with the biofilm formed by Streptococcus mutans with the wetting behavior of those shell surfaces and their roughness. More hydrophobic surfaces (with intact periostracum) were observed to lead to less attachment (correlation coefficient=-0.67). This study can pave the way for designing such biomimetic surfaces to prevent bacterial attachment.

Inhibition of Quorum Sensing Controlled Virulence Factors and Biofilm Formation of Streptococcus mutans Isolated From Orthodontic Subjects by 4-Hydroxycinnamic Acid.

Introduction Dental plaque biofilms are a collection of microorganisms that are adhered to the tooth enamel surface. Inhibition of plaque biofilms is required to prevent dental caries and periodontitis and currently, there are many chemical and herbal products in use for inhibition of biofilms but with limited success. Materials and methods Dental plaque collection was done from subjects undergoing orthodontic therapy followed by isolation of Streptococcus mutans. Isolated S. mutans were subjected to disk diffusion assay with 4-HCA (baseline 10mg/mL) for the zone of inhibition and broth micro-dilution to evaluate the minimum inhibitory concentration (MIC) and sub-MIC. Crystal violet staining was done for biofilm inhibition assay. Results Growth of S. mutans was inhibited by 4-HCA at concentrations as low as 0.31 mg/mL. 4-HCA (40μL) inhibited the bacterial growth and a clear zone (15 mm) was observed. 4-Hydroxycinnamic acids treated culture showed progressive reduction in the biofilm production at the concentration of 0.01 mg/mL. The 4-HCA concentration as low as 4 mg and 2 mg has remarkably inhibited biofilm formation of 49.3% and 34.3%, respectively. Conclusion The anti-quorum sensing and anti-biofilm activity of 4-Hydroxycinnamic acid against S. mutans isolated from subjects undergoing orthodontic treatment showed a remarkable result.

The inhibitory effect of berberine chloride hydrate on Streptococcus mutans biofilm formation at different pH values.

Streptococcus mutans (S. mutans) is one of the major cariogenic bacteria of dental caries owing to its ability to adhere to tooth surfaces and biofilm formation. Berberine chloride hydrate (BH), a quaternary ammonium salt alkaloid, has diverse pharmacological efforts against microorganisms. However, the effect of BH on S. mutans biofilm has not been reported. Considering that berberine is a quaternary ammonium salt alkaloid, which needs to adapt to a large variation in pH values and the acid resistance of S. mutans, we employed three groups including pH 5 (acidic), pH 8 (alkaline), and unprocessed group (neutral) to examine the antibiofilm activities of BH against S. mutans during different pH values. In this study, we found BH effectively suppresses S. mutans biofilm formation as well as its cariogenic virulence including acid production and EPS synthesis significantly, and the inhibitory effort was reduced under acidic condition whereas elevated under alkaline condition. In addition, we preliminarily explored the influence of pH values on the structural stability and biosafety of BHas well as the underlying mechanism of inhibition of S. mutans biofilm formation with BH. Our study showed BH could maintain a good structural stability and low toxicity to erythrocytes at different pH values. And BH could downregulate the expression of srtA, spaP, and gbpC, which play critical roles in the adhesion process, promoting bacterial colonization and biofilm formation. Furthermore, comX and ldh expression levels were downregulated in BH-treated group, which might explain its inhibitory effect on acid production.IMPORTANCEDental caries is a common chronic detrimental disease, which could cause a series of oral problem including oral pain, difficulties in eating, and so on. Recently, many natural products have been considered as fundamental sources of therapeutic drugs to prevent caries. Berberine as a plant extract showed good antibiofilm abilities against microorganism. Our study focuses on its antibiofilm abilities against S. mutans, which was defined as major cariogenic bacterium and explored the role of pH values and possible underlying mechanisms in the inhibitory effect of BH on S. mutans biofilm formation. This study demonstrated a promising prospect for BH as an adjuvant drug in the prevention and management of dental caries.

Antimicrobial peptide temporin derivatives inhibit biofilm formation and virulence factor expression of Streptococcus mutans.

Temporin-GHa obtained from the frog Hylarana guentheri showed bactericidal efficacy against Streptococcus mutans. To enhance its antibacterial activity, the derived peptides GHaR and GHa11R were designed, and their antibacterial performance, antibiofilm efficacy and potential in the inhibition of dental caries were evaluated. Bacterial survival assay, fluorescent staining assay and transmission electron microscopy observation were applied to explore how the peptides inhibited and killed S. mutans. The antibiofilm efficacy was assayed by examining exopolysaccharide (EPS) and lactic acid production, bacterial adhesion and cell surface hydrophobicity. The gene expression level of virulence factors of S. mutans was detected by qRT-PCR. Finally, the impact of the peptides on the caries induced ability of S. mutans was measured using a rat caries model. It has been shown that the peptides inhibited biofilm rapid accumulation by weakening the initial adhesion of S. mutans and reducing the production of EPS. Meanwhile, they also decreased bacterial acidogenicity and aciduricity, and ultimately prevented caries development in vivo. GHaR and GHa11R might be promising candidates for controlling S. mutans infections.

Virulence factors analysis and determination of the suitable chemical agent to inhibit Streptococcus mutans growth and biofilm formation

Streptococcus mutans is one of the most important causes of tooth decay and oral diseases, and it has received wide attention from researchers worldwide. The present work aimed to investigate, evaluation and comparisons effect of the antimicrobial and anti-biofilm property of some commercial compounds (chlorhexidine, hyaluronic acid, sodium fluoride and silver nanoparticles) against oral bacteria pathogens of S. mutans. In this work, S. mutans ATCC 25175 was used and its virulence factors were analyzed using Seed Viewer and The Comprehensive Antibiotic Resistance Database (CARD). The antibacterial susceptibility testing (AST) confirms effects of chlorhexidine mouthwash on S. mutans ATCC 25175 beggar than all components. The minimum inhibitory concentration (MIC) minimum bactericidal concentration (MBC) and Minimum biofilm inhibition concentration (MBIC) of the commercial compounds were determined. The statistical analysis was performed using One-Way ANOVA (Tukey test). The VITEK2system was used to confirm the biochemical features of S. mutans ATCC 25175 and susceptibility testing was done to compare the results with Seed Viewer and CARD analysis. The findings showed that The MICs, MBCs, and MBIC means of chlorhexidine against S. mutans ATCC 25175 was 0.0079, 0.0038, and 0.0039 μg/mL respectively, where its biological activity was significantly (P < 0.05) higher compared to other commercial compounds included in this study. The S. mutans strain used in this study showed sensitivity to all tested antibiotics and the Seed Viewer and CARD reported that it has several virulence factors such as adhesion genes (hsp33, Cpa, PrtF, PrtF2, MsmRL, Nra, SOF, LepAL, Spy0135, and RofA) and antibiotic-resistant genes (vanY, vanT, and mdeA). This study concluded that chlorhexidine was the best in inhibiting S. mutans growth as well as preventing biofilm formation, and the study recommends further investigation about antibiotic resistance genes and susceptibility profile.

Lysine lactylation regulates metabolic pathways and biofilm formation in Streptococcus mutans.

In eukaryotes, lactate produced during glycolysis is involved in regulating multiple metabolic processes through lysine lactylation (Kla). To explore the potential link between metabolism and Kla in prokaryotes, we investigated the distribution of Kla in the cariogenic bacterium Streptococcus mutans during planktonic growth in low-sugar conditions and in biofilm-promoting, high-sugar conditions. We identified 1869 Kla sites in 469 proteins under these two conditions, with the biofilm growth state showing a greater number of lactylated sites and proteins. Although high sugar increased Kla globally, it reduced lactylation of RNA polymerase subunit α (RpoA) at Lys173. Lactylation at this residue inhibited the synthesis of extracellular polysaccharides, a major constituent of the cariogenic biofilm. The Gcn5-related N-acetyltransferase (GNAT) superfamily enzyme GNAT13 exhibited lysine lactyltransferase activity in cells and lactylated Lys173 in RpoA in vitro. Either GNAT13 overexpression or lactylation of Lys173 in RpoA inhibited biofilm formation. These results provide an overview of the distribution and potential functions of Kla and improve our understanding of the role of lactate in the metabolic regulation of prokaryotes.

Salvia dumetorum essential oil: GC-MS analysis, antibacterial activity and effect on the formation of Streptococcus mutans biofilms

Salvia dumetorum essential oil (SDEO) was obtained using a Clevenger apparatus by hydrodistillation approach. The chemical composition of the essential oil was determined by GC-MS analysis. In this study SDEO was screened for its antibacterial activity against Staphylococcus aureus, Bacillus subtilis, Escherichia coli, and Candida albicans and the amount of biofilm formed by Streptococcus mutans bacteria. For the first time the chemical composition of SDEO growing in Central Kazakhstan was established. The results obtained from GC-MS showed the predominance of sesquiterpenoids (54.15%). The antibacterial study results showed that SDEO exhibited strong antibacterial activity against B. subtilis and S. aureus in vitro and also demonstrated an inhibitory effect on S. mutans biofilm formation on 1% sucrose medium. During the study, no antibacterial activity was detected against E. coli and C. albicans. These results demonstrated that SDEO can be used in the development of new antibacterial and anti-caries therapeutic dental products.

Application of Three Compounds Extracted from Cynodon dactylon against Streptococcus mutans Biofilm Formation to Prevent Oral Diseases.

Streptococcus mutans bacteria form a biofilm called plaque that causes oral diseases, including tooth decay. Therefore, inhibition of biofilm formation is essential to maintaining good oral health. The health and nutritional benefits of Cynodon dactylon are well documented, but very little is known about its use to treat against oral diseases. The aim of this study was to detect the adhesion strength of the S. mutans bacterial biofilm in 100 cases in the Rajshahi region and evaluate the inhibitory activity of different compound extracts of C. dactylon on the S. mutans bacterial biofilm by determining the composition of isolated compounds using phytochemical analysis. Nuclear magnetic resonance (NMR) spectroscopy confirmed that three specific compounds from C. dactylon were discovered in this study: 3,7,11,15 tetramethyl hexadec-2-4dien 1-o1, compound 3,7,11,15 tetramethylhexadec-2-en-1-o1 from phytol derivatives, and stigmasterol. Results indicated that the compound of 3,7,11,15-tetramethyl-hexadec-2-en-1-ol exhibited higher antibiofilm activities on S. mutans than those of the other compound extracts. A lower level of minimum inhibitory concentration was exposed by 3, 7, 11,15 tetramethyl hexadeca-2-en-1-o1 (T2) on S. mutans at 12.5 mL. In this case, the compound of 3,7,11,15 tetramethyl hexadec 2en-1-o1 was used, and patients showed a mean value and standard error reduced from 3.42 ± 0.21 to 0.33 ± 0.06 nm. The maximum inhibition was (80.10%) in the case of patient no. 17, with a value of p < 0.05 found for S. mutans to which 12.5 μL/mL ethyl acetate extract was applied. From these findings, it may be concluded that C. dactylon extracts can be incorporated into various oral preparations to prevent tooth decay.

Prevention of bacterial biofilm formation on orthodontic brackets by non-crosslinked chitosan coating

During orthodontic treatment, the patients are susceptible to dental caries as a result of the bacterial adhesion and biofilm formation around the orthodontic brackets. Prevention of the caries-related biofilm formation is of significance for maintaining both aesthetics and health of the teeth. Herein, the brackets were functionalized with antibacterial activity via coating a layer of non-crosslinked chitosan (CS). We firstly demonstrated the ability of free CS scaffolds (not coated on brackets) to inhibit the formation of Streptococcus mutans biofilms (inhibition rate 94.3 % for CS-0.3 mg) and to eradicate the mature biofilms (biofilm loss rate 99.8 % for CS-1.2 mg). Further, the inhibition of S. mutans biofilm formation on brackets by CS coating was investigated for the first time. As a result, the CS-coated brackets (Br-CS) kept the great biofilm inhibition capacity of free CS scaffolds. In detail, the Br-CS, prepared by immersing brackets in CS solutions (containing 1.0, 2.5, 5.0 and 10 mg/mL CS) and freeze-drying, showed the biofilm inhibition rate of 48.5 %, 88.6 %, 96.4 % and 99.6 %, respectively. In conclusion, coating orthodontic brackets with the non-crosslinked CS is a potential approach for inhibiting biofilm formation and protecting patients from dental caries.

Impacts of a DUF2207 Family Protein on Streptococcus mutans Stress Tolerance Responses and Biofilm Formation.

Locus SMU.243 in Streptococcus mutans was annotated as a member of the DUF2207 family proteins highly conserved in all bacteria but with unknown function. To investigate its role in S. mutans physiology, a SMU.243-deficient mutant was constructed using allelic exchange mutagenesis, and the impacts of SMU.243 deletion on bacterial growth, stress tolerance response, and biofilm formation were analyzed. Compared to the wild-type UA159, S. mutans lacking SMU.243 displayed a reduced growth rate and a reduced overnight culture density (p < 0.01) when grown at low pH and in the presence of methyl viologen. Relative to the parent strain, the deficient mutant also had a reduced survival rate following incubation in a buffer of pH 2.8 (p < 0.01) and in a buffer containing hydrogen peroxide at 58 mM after 60 min (p < 0.001) and had a reduced capacity in biofilm formation especially in the presence of sucrose (p < 0.01). To study any ensuing functional/phenotypical links between SMU.243 and uppP, which is located immediately downstream of SMU.243 and encodes an undecaprenyl pyrophosphate phosphatase involved in recycling of carrier lipid undecaprenyl phosphate, a uppP deficient mutant was generated using allelic exchange mutagenesis. Unlike the SMU.243 mutant, deletion of uppP affected cell envelope biogenesis and caused major increases in susceptibility to bacitracin. In addition, two variant morphological mutants, one forming rough colonies and the other forming mucoid, smooth colonies, also emerged following the deletion of uppP. The results suggest that the SMU.243-encoded protein of the DUF2207 family in S. mutans plays an important role in stress tolerance response and biofilm formation, but unlike the downstream uppP, does not seem to be involved in cell envelope biogenesis, although the exact roles in S. mutans' physiology awaits further investigation.

Inhibition of biofilm formation and virulence factors of cariogenic oral pathogen Streptococcus mutans by natural flavonoid phloretin

ABSTRACT Objectives To evaluate the effect and mechanism of action of the flavonoid phloretin on the growth and sucrose-dependent biofilm formation of Streptococcus mutans. Methods Minimum inhibitory concentration, viability, and biofilm susceptibility assays were conducted to assess antimicrobial and antibiofilm effect of phloretin. Biofilm composition and structure were analysed with scanning electron microscopy (SEM) and confocal laser scanning microscopy (CLSM). Water-soluble (WSG) and water-insoluble glucan (WIG) were determined using anthrone method. Lactic acid measurements and acid tolerance assay were performed to assess acidogenicity and aciduricity. Reverse transcription quantitative PCR (RT-qPCR) was used to measure the expression of virulence genes essential for surface attachment, biofilm formation, and quorum sensing. Results Phloretin inhibited S. mutans growth and viability in a dose-dependent manner. Furthermore, it reduced gtfB and gtfC gene expression, correlating with the reduction of extracellular polysaccharides (EPS)/bacteria and WIG/WSG ratio. Inhibition of comED and luxS gene expression, involved in stress tolerance, was associated with compromised acidogenicity and aciduricity of S. mutans. Conclusions Phloretin exhibits antibacterial properties against S. mutans, modulates acid production and tolerance, and reduces biofilm formation. Clinical significance Phloretin is a promising natural compound with pronounced inhibitory effect on key virulence factors of the cariogenic pathogen, S. mutans.

Investigation of Components in Roasted Green Tea That Inhibit Streptococcus mutans Biofilm Formation.

Streptococcus mutans form oral biofilms (BFs) and cause dental caries. Roasted green tea (RGT) is prepared by roasting the tea plant, and RGT-specific polyphenols are produced during the roasting process. Catechins, polyphenols in green tea, have BF inhibitory activity against S. mutans; therefore, RGT-specific polyphenols are also expected to have this activity. However, there are few reports on the structural and functional properties of RGT. This study aimed to investigate the inhibitory activity of RGT against S. mutans BF formation and to investigate the active compounds. RGT extract fractionation and BF inhibitory assay were performed. Strong activity was confirmed in the RGT fractions that had medium-high hydrophobicity, were rich in phenolic hydroxyl groups, and lacked catechins. A peak comprising compounds with molecular weights of 918 (mw918) and 1050 (mw1050) was purified from the fraction. Since BF inhibitory activity was confirmed for this peak, these compounds were considered to be part of the active ingredients. The mw918 polyphenol was detected only in RGT and it was thought to be produced during the roasting process. The results of this research will serve as a basis for the future application of RGT as a safe and effective anti-caries agent.