Streptococcus Mutans Biofilm Research Articles

Mutanase from Trichoderma harzianum inductively Produced by Mutan: Short-Term Treatment to Degrade Mature Streptococcus mutans Biofilm.

This study aimed to evaluate the disruptive effect of fungal mutanase against cariogenic biofilm after short-term treatment. For that, mature Streptococcus mutans biofilms (n = 9) were exposed to active or inactivated enzymes produced by Trichoderma harzianum for 1 min, two times per day. Biofilms were analyzed by amount of matrix water-insoluble polysaccharides, bacterial viability, acidogenicity, and morphology by scanning electron microscopy (SEM). The group treated with active enzymes (AE) had a significantly lower amount of insoluble polysaccharides (893.30 ± 293.69) when compared to the negative control group (NaCl, 2192.59 ± 361.96), yet no significant difference was found when comparing to the positive control group (CHX, 436.82 ± 151.07). Also, there was no significant effect on bacteria metabolism and viability (P-value < 0.05). Data generated by the quantitative analysis were confirmed through scanning electron microscopy images. Thus, fungal mutanase degraded the biofilm after a short-term treatment without interfering with bacterial viability and metabolism. Such findings offer insight to the development of routine oral care products containing this input.

Evaluation of the antibacterial activity of Enamelast® and Fluor defender® fluoride varnishes against Streptococcus mutans biofilm: an in vitro study in primary teeth

PurposeThe aim of the current work was to compare the antibacterial activity of Enamelast® and Fluor defender® fluoride varnish on biofilm generation by Streptococcus mutans on extracted primary teeth.MethodsThirty-six primary molars were collected and sliced into seventy-two test model disks. All specimens were examined, and the cracked or broken ones were discarded. A total number of specimens (n = 54) were divided into two experimental analyses viz; biofilm formation (n = 27) and microscopic examination (n = 27). Specimens of each analysis were tested under different experimental conditions: a negative control group (n = 9), Fluor defender group (n = 9), and Enamelast group (n = 9). Following treatment, biofilms were generated by adherent Streptococcus mutans on the test model disks on three time intervals: 24 h (n = 3), 48 h (n = 3), and 72 h (n = 3) for each analysis. Then, for biofilm formation analysis, the biofilm was detected spectrophotometrically at 620 nm after being stained by crystal violet. For microscopical analysis, the surfaces of the test model disks were visualized by scanning electron microscopy (SEM), and each image was processed and analyzed using ImageJ software.ResultsAt 48 and 72 h, Enamelast® and Fluor defender®-treated group showed significantly (p < 0.001) slight adhered bacterial cells when compared with the negative control group as revealed by the absorbance and SEM. Compared with the Fluor defender®-treated group, the absorbance of the Enamelast®-treated group showed a significant (p < 0.001) increase by approximately 7- and 16.5-fold at 48 and 72 h, respectively. Similarly, SEM showed that the number of bacterial cells adhered to enamel surfaces in the Fluor defender®-treated group was significantly (p < 0.001) fewer than the Enamelast®-treated group by approximately 36.55% and 20.62% at 48 and 72 h after exposure, respectively.ConclusionWe conclude that the anti-biofilm activity of Fluor defender® against Streptococcus mutans was significantly (p < 0.001) greater than Enamelast® fluoride varnish. The use of Fluor defender® is encouraged as a preventive measure in children with the high risk of developing 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.

Modified Lactoperoxidase System as a Promising Anticaries Agent: In Vitro Studies on Streptococcus mutans Biofilms.

The lactoperoxidase (LPO) system shows promise in the prevention of dental caries, a common chronic disease. This system has antimicrobial properties and is part of the non-specific antimicrobial immune system. Understanding the efficacy of the LPO system in the fight against biofilms could provide information on alternative strategies for the prevention and treatment of caries. In this study, the enzymatic system was modified using four different (pseudo)halide substrates (thiocyanate, thiocyanate-iodide mixture, selenocyanate, and iodide). The study evaluated the metabolic effects of applying such modifications to Streptococcus mutans; in particular: (1) biofilm formation, (2) synthesis of insoluble polysaccharides, (3) lactate synthesis, (4) glucose and sucrose consumption, (5) intracellular NAD+ and NADH concentrations, and (6) transmembrane glucose transport efficiency (PTS activity). The results showed that the LPO-iodide system had the strongest inhibitory effect on biofilm growth and lactate synthesis (complete inhibition). This was associated with an increase in the NAD+/NADH ratio and an inhibition of glucose PTS activity. The LPO-selenocyanate system showed a moderate inhibitory effect on biofilm biomass growth and lactate synthesis. The other systems showed relatively small inhibition of lactate synthesis and glucose PTS but no effect on the growth of biofilm biomass. This study provides a basis for further research on the use of alternative substrates with the LPO system, particularly the LPO-iodide system, in the prevention and control of biofilm-related diseases.

A Transcriptional Analysis Showing the Effects of GH12 Combined with Fluoride for Suppressing the Acidogenicity of Streptococcus mutans Biofilms.

The acidogenicity of Streptococcus mutans is important for caries development. The antimicrobial peptide GH12 can affect the integrity of cellular membranes and the virulence factors of S. mutans. Combining GH12 and NaF (GF) efficiently controlled the development of caries, but its mechanisms remained unrevealed. This research intended to verify the effects of GF on the acidogenicity of S. mutans biofilms and to reveal the mechanisms. Lactic acid production assays and pH monitoring assays were conducted to investigate the regulatory effects of the GF treatment on the acidogenicity of S. mutans biofilms. RNA sequencing and bioinformatics analyses were conducted to screen the transcriptional profile affected by the GF treatment. The results demonstrated the GF group had significantly less lactic acid and maintained the broth's pH values above 5.0 for longer times. Thereafter, GO/KEGG enrichment analyses and RT-qPCR validation revealed that the GF treatment mainly restrained the expression of genes related to the carbohydrates' internalization and metabolism. Compared with NaF, the GF treatment further downregulated the carbohydrates transportation genes. Moreover, compared with GH12, the GF treatment affected the membrane's integrity more significantly. Generally, GF treatment could arrest the acidogenicity of S. mutans biofilms, mainly through suppressing carbohydrates transportation and inhibiting overall metabolism.

Antimicrobial activity of probiotic Streptococcus salivarius LAB813 on in vitro cariogenic biofilms

ObjectiveTo investigate the antimicrobial activity of a novel commensal strain of Streptococcus salivarius, LAB813, against Streptococcus mutans biofilms. MethodsThe inhibitory activity of LAB813 towards S. mutans was tested using mono-, dual-, and multi-species cariogenic biofilms formed on three types of orthodontic appliances (metal, ceramic, aligner). The activity of the commercially available probiotic, BLIS M18™ was used as control. ResultsLAB813 significantly inhibited S. mutans biofilms with cell killing approximating 99% for all materials. LAB813 showed effectiveness at inhibiting S. mutans in more complex multi-species biofilms with cell killing approximating 90% for all three materials. When comparing the killing kinetics of the probiotics, LAB813 had a faster rate of killing biofilms than M18. Experiments conducted with cell-free culture supernatant confirmed the presence of an inhibitory substance of proteinaceous nature. The addition of xylitol, a common sugar substitute used for human consumption, potentiated the inhibitory effects of LAB813 against S. mutans embedded in a more complex fungal-bacterial biofilm. ConclusionsLAB813 possesses strong antimicrobial activity, potent anti-biofilm properties, and enhanced antimicrobial activity in the presence of xylitol. The identification and characterization of strain LAB813 exhibiting antimicrobial activity towards S. mutans hold exciting promise for this novel strain to be developed as an oral probiotic for use in the prevention of dental caries.

The study on the effect of amino acid porphyrin conjugate-mediated antimicrobial photodynamic therapy on Streptococcus mutans biofilm

Antimicrobial Photodynamic Therapy (aPDT) based on the action of visible light and photosensitizers has emerged as a promising microbial reduction and alternative to antibiotics resistance to cariogenic pathogens. The present research aims to evaluate the antimicrobial effect of aPDT mediated by a new photosensitizer (amino acid porphyrin conjugate 4i) on Streptococcus mutans (S. mutans) biofilm. Qualitative morphologic characteristics of S. mutans biofilms are shown by scanning electron microscopy (SEM). The colony plate counting method is used to measure the dark toxicity and the phototoxicity of different concentrations of 4i-aPDT to S. mutans biofilms. MTT assay is conducted to investigate the effect of 4i mediated aPDT on the metabolic activity of S. mutans biofilm. Changes in structure morphology, bacterial density and extracellular matrix of S. mutans biofilm are observed by SEM. The distribution of living and dead bacteria in biofilm is detected using Confocal laser microscopy (CLSM). The results indicate that single laser irradiation has no antibacterial effect on S. mutans biofilms. With the increase of 4i concentration or the prolongation of laser irradiation time, the antibacterial effect of 4i-mediated aPDT on S. mutans biofilm is more statistically significant compared to the control. When the concentration of 62.5 µmol/L 4i is continuously illuminated for 10 min, the logarithm of the colonies in the biofilm shows a reduction of 3.4 log10. MTT assay detected absorbance values of biofilm by 4i-mediated aPDT are the lowest, indicating a significant decrease in biofilm metabolic activity. SEM analysis shows that 4i mediated aPDT reduced the quantity and density of S. mutans. A dense red fluorescence image of the 4i-aPDT treated biofilm is observed under CLSM, indicating that the dead bacteria are widely distributed.

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.

Novel Dental Low-Shrinkage-Stress Composite with Antibacterial Dimethylaminododecyl Methacrylate Monomer.

Current dental resins exhibit polymerization shrinkage causing microleakage, which has the potential to cause recurrent caries. Our objectives were to create and characterize low-shrinkage-stress (LSS) composites with dimethylaminododecyl methacrylate (DMADDM) as an antibacterial agent to combat recurrent caries. Triethylene glycol divinylbenzyl ether and urethane dimethacrylate were used to reduce shrinkage stress. DMADDM was incorporated at different mass fractions (0%, 1.5%, 3%, and 5%). Flexural strength, elastic modulus, degree of conversion, polymerization stress, and antimicrobial activity were assessed. The composite with 5% DMADDM demonstrated higher flexural strength than the commercial group (p < 0.05). The addition of DMADDM in BisGMA-TEGDMA resin and LSS resin achieved clinically acceptable degrees of conversion. However, LSS composites exhibited much lower polymerization shrinkage stress than BisGMA-TEGDMA composite groups (p < 0.05). The addition of 3% and 5% DMADDM showed a 6-log reduction in Streptococcus mutans (S. mutans) biofilm CFUs compared to commercial control (p < 0.001). Biofilm biomass and lactic acid were also substantially decreased via DMADDM (p < 0.05). The novel LSS dental composite containing 3% DMADDM demonstrated potent antibacterial action against S. mutans biofilms and much lower polymerization shrinkage-stress, while maintaining excellent mechanical characteristics. The new composite is promising for dental applications to prevent secondary caries and increase restoration longevity.

Effectiveness of Xylitol Toothpaste on the Growth and Biofilm Formation of Streptococcus mutans ATCC 25175

Background: Several toothpaste ingredients have also been proven to prevent dental caries. An alternative material like xylitol can reduce biofilms and caries’ formation. Objective: This study aims to test the antibacterial xylitol toothpaste against Streptococcus mutans biofilm. Materials and Methods: The type of this research was true experimental by using two kinds of toothpaste with xylitol active ingredient (samples 1 and 2) and toothpaste with fluoride (sample 3). The toothpaste was diluted using the serial dilution method, and then the minimum inhibitory concentration (MIC), minimum bactericide concentration (MBC), minimum biofilm eradication concentration (MBEC), and minimum biofilm inhibition concentration (MBIC) were examined. The data were analyzed with ANOVA and Pearson correlation with a significance value of p0.005. Results: The MIC values of samples 1, 2, and 3 were 0.39%, 0.78%, and 0.39%, respectively. The MBC of samples 1, 2, and 3 were 12.5%, 25%, and 6.25%, respectively. The results of MBIC samples 1, 2, and 3 were 1.56%, 1.56%, and 0.78%, respectively. The results of MBEC samples 1, 2, and 3 were 6.25%, 6.25%, and 1.56%, respectively. There was a difference in the decrease of the mass of Streptococcus mutans ATCC 25175 biofilm after administration using xylitol and fluoride toothpaste, with a p-value of 0.0295. Additionally, there is a correlation between the time of administration of xylitol toothpaste with the changes in the mass of the biofilm Streptococcus mutans ATCC 25175. Conclusion: Xylitol toothpaste can inhibit the growth and eradication of Streptococcus mutans biofilm ATCC 25175.

Preparation of Nanocomposites for Antibacterial Orthodontic Invisible Appliance Based on Piezoelectric Catalysis.

Compared to fixed orthodontic appliances with brackets, thermoplastic invisible orthodontic aligners offer several advantages, such as high aesthetic performance, good comfort, and convenient oral health maintenance, and are widely used in orthodontic fields. However, prolonged use of thermoplastic invisible aligners may lead to demineralization and even caries in most patients' teeth, as they enclose the tooth surface for an extended period. To address this issue, we have created PETG composites that contain piezoelectric barium titanate nanoparticles (BaTiO3NPs) to obtain antibacterial properties. First, we prepared piezoelectric composites by incorporating varying amounts of BaTiO3NPs into PETG matrix material. The composites were then characterized using techniques such as SEM, XRD, and Raman spectroscopy, which confirmed the successful synthesis of the composites. We cultivated biofilms of Streptococcus mutans (S. mutans) on the surface of the nanocomposites under both polarized and unpolarized conditions. We then activated piezoelectric charges by subjecting the nanocomposites to 10 Hz cyclic mechanical vibration. The interactions between the biofilms and materials were evaluated by measuring the biofilm biomass. The addition of piezoelectric nanoparticles had a noticeable antibacterial effect on both the unpolarized and polarized conditions. Under polarized conditions, nanocomposites demonstrated a greater antibacterial effect than under unpolarized conditions. Additionally, as the concentration of BaTiO3NPs increased, the antibacterial rate also increased, with the surface antibacterial rate reaching 67.39% (30 wt% BaTiO3NPs). These findings have the potential for application in wearable, invisible appliances to improve clinical services and reduce the need for cleaning methods.

Antibiofilm and mechanical properties of silver nanowire-modified glass ionomer cement

BackgroundGlass ionomer cement (GIC) is a commonly used restorative material in dentistry, but GIC does not have significant antibiofilm effects and its mechanical strength is limited. ObjectiveTo investigate the antibiofilm and mechanical properties of a newly developed silver nanowire (AgNW) modified GIC. MethodsGICs were modified with different nanosilver formulations including 0.05%wt AgNW, 0.5%wt AgNW, 0.05%wt silver nanoparticle (AgNP) and 0.5%wt AgNP. Biofilms of Streptococcus mutans were cultured for 72 h on GIC specimens. Scanning electron microscopy (SEM) was conducted to observe the accumulation and morphology of Streptococcus mutans on the material surfaces. Another set of biofilm/specimens was treated with SYTO-9 and the biofilm development was evaluated by quantifying the visible stain with an inverted fluorescence microscope. The compressive strength of the specimens was assessed according to ISO 9917–1:2007. The colour of GICs was compared using the VITA Easyshade system. ResultsSEM images showed that fewer biofilms were accumulated on nanosilver-modified GICs. The Streptococcus mutans quantity was significantly lower in all nanosilver-modified groups compared to conventional GIC (p<0.0001). GIC modified with 0.5% AgNP showed slightly lower compressive strength than the negative control (58.3 ± 12.9 MPa vs 78.3 ± 13.8 MPa); but there is no significant difference between all study groups. The colour change between AgNW-modified GIC and conventional GIC was significantly lower than that between AgNP-modified GIC and conventional GIC. ConclusionAgNW-modified GIC showed superior antibiofilm effect and comparable compressive strength to conventional GIC. In addition, the complement of AgNW would not influence the colour stability of GIC as much as AgNP. Clinical SignificanceThis study developed a novel AgNW-modified GIC material. It showed good antibiofilm and mechanical properties and would not influence the colour stability of GIC. The AgNW-modified GIC has a large potential for clinical use and biomedical application.

Saliva Decreases Sucrose-Induced Cariogenicity in an Experimental Biological Caries Model.

Whether a minimum quantity of saliva inhibit the caries process remains uncertain. This study aimed to investigate the impact of saliva dilutions on an in vitro caries model using Streptococcus mutans (S. mutans) biofilms. S. mutans biofilms were cultivated on enamel and root dentin slabs, in culture media containing different proportions of saliva (v/v): 0%, 5%, 10%, 25%, 50%, 75%, and 100% saliva, and exposed to a 10% sucrose solution (5 min, 3x/day), with appropriate controls. After 5 (enamel) and 4 (dentin) days, demineralization, biomass, viable bacteria, and polysaccharide formation were analyzed. The acidogenicity of the spent media was monitored overtime. Each assay was performed in triplicate across two independent experiments (n = 6). In both enamel and dentin, an inverse relationship was observed between acidogenicity, demineralization, and the proportion of saliva. Even small quantities of saliva incorporated into the media led to a noticeable reduction in enamel and dentin demineralization. Saliva presence resulted in significant reductions in biomass, viable S. mutans cells, and polysaccharides, with the effects being concentration-dependent for both tissues. High quantities of saliva can almost completely inhibit sucrose-induced cariogenicity, while even small amounts exhibit a dose-dependent caries-protective effect.

Surface properties and Streptococcus mutans biofilm adhesion of ion-releasing resin-based composite materials

ObjectiveTo evaluate the adhesion of Streptococcus mutans (S. mutans) and related surface properties of ion-releasing resin-based composite (RBC) restorative materials. MethodsTwo ion-releasing RBCs, Activa (ACT) and Cention-N (CN), were compared to a conventional RBC (Z350) and a resin-modified glass ionomer cement (Fuji-II-LC). Ten disk-shaped specimens were fabricated for each material (n = 40). After standardized surface polishing procedure, the surface properties of the specimens were evaluated using surface roughness measurements by a profilometer and hydrophobicity using water contact angle measurements. To assess bacterial adhesion, the number of S. mutans bacteria was calculated from colony-forming units (CFU). Confocal laser scanning microscope analysis was done for qualitative & quantitative assessment. The data were analyzed using One-way ANOVA followed by Tukey's post-hoc test to compare the mean values of surface roughness, water contact angle and CFU values. To compare the mean dead cell percentage Kruskal-Wallis rank test and Conover test were used. A p-value of ≤ 0.05 was used to report the statistical significance. ResultsZ350 and ACT had the smoothest surfaces, followed by CN, and the roughest surface was seen in FUJI-II-LC. The lowest water contact angles were seen in CN, and Z350, and the highest were in ACT. S. mutans counts were the highest in ACT and the lowest in Z350 and CN. CN and Fuji-II-LC registered the highest percentage of dead bacterial cells, while the lowest were in ACT. SignificanceSurface properties did not significantly influence bacterial adhesion. More S. mutans bacteria accumulated on ACT than on the nanofilled composite and on CN. CN had antibacterial effects against Streptococcus mutans biofilms.

Effects of blue-light LED toothbrush on reducing dental plaque and gingival inflammation in orthodontic patients with fixed appliances: a crossover randomized controlled trial

BackgroundPatients with fixed orthodontic appliances have higher plaque accumulation and gingival inflammation. Our aim was to compare the effectiveness of a light emitting diode (LED) toothbrush with a manual toothbrush in reducing dental plaque and gingival inflammation in orthodontic patients with fixed appliances, and to investigate the effect of the LED toothbrush on Streptococcus mutans (S. mutans) biofilm in vitro.MethodsTwenty-four orthodontic patients were recruited and randomly assigned into 2 groups: (1) started with manual and (2) started with LED toothbrushes. After a 28-day usage and 28-day wash-out period, the patients switched to the other intervention. The plaque and gingival indices were determined at baseline and 28 days after each intervention. The patients’ compliance and satisfaction scores were collected using questionnaires. For the in vitro experiments, S. mutans biofilm was divided into 5 groups (n = 6) with 15-, 30-, 60-, or 120-sec LED exposure, and without LED exposure as a control group.ResultsThere was no significant difference in the gingival index between the manual and LED toothbrush groups. The manual toothbrush was significantly more effective in reducing the plaque index in the proximal area on the bracket side (P = 0.031). However, no significant difference was found between the two groups in other areas around the brackets or on the non-bracket side. After LED exposure in vitro, the percentages of bacterial viability after LED exposure for 15–120 s were significantly lower compared with the control (P = 0.006).ConclusionClinically, the LED toothbrush was not more effective in reducing dental plaque or gingival inflammation than the manual toothbrush in orthodontic patients with fixed appliances. However, the blue light from the LED toothbrush significantly reduced the number of S. mutans in biofilm when it was exposed to the light for at least 15 s in vitro.Clinical Trial RegistrationThai Clinical Trials Registry (TCTR20210510004). Registered 10/05/2021.

Functional liposome loaded curcumin for the treatment of Streptococcus mutans biofilm.

Introduction: Plaque biofilms, mainly formed by Streptococcus mutans (S. mutans), play an important role in the occurrence and development of dental caries. Antibiotic treatment is the traditional way to control plaque. However, problems such as poor drug penetration and antibiotic resistance have encouraged the search for alternative strategies. In this paper, we hope to avoid antibiotic resistance through the antibacterial effect of curcumin, a natural plant extract with photodynamic effects, on S. mutans. However, the clinical application of curcumin is limited due to its low water solubility, poor stability, high metabolic rate, fast clearance rate, and limited bioavailability. In recent years, liposomes have become a widely used drug carrier due to their numerous advantages, such as high drug loading efficiency, high stability in the biological environment, controlled release, biocompatibility, non-toxic, and biodegradability. So, we constructed a curcumin-loaded liposome (Cur@LP) to avoid the defect of curcumin. Methods: Cur@LP functioned with NHS can adhere to the surface of the S. mutans biofilm by condensation reaction. Liposome (LP) and Cur@LP was characterized by transmission electron microscopy (TEM) and dynamic light scattering (DLS). The cytotoxicity of Cur@LP was evaluated by CCK-8 assay and LDH assay. The adhesion of Cur@LP to S. mutans biofilm was observed by confocal laser scanning microscope (CLSM). The antibiofilm efficiency of Cur@LP were evaluated by crystal violet staining, CLSM, and scanning electron microscope (SEM). Results: The mean diameter of LP and Cur@LP were 206.67 ± 8.38nm and 312 ± 18.78nm respectively. The ζ-potential of LP and Cur@LP were ∼-19.3mV and ∼-20.8mV respectively. The encapsulation efficiency of Cur@LP was (42.61 ± 2.19) %, and curcumin was rapidly released up to ±21% at 2h. Cur@LP has negligible cytotoxicity, and can effectively adhered to the S. mutans biofilm and inhibited its growth. Discussion: Curcumin has been widely studied in many fields such as cancer, which can be attributed to its antioxidant and anti-inflammatory effects. At present, there are few studies on the delivery of curcumin to S. mutans biofilm. In this study, we verified the adhesion and antibiofilm of Cur@LP to S. mutans biofilm. This biofilm removal strategy has the potential to be translated into the clinic.

Evaluation of antibacterial properties of fluoride-containing mouth rinses differing in their acidic compound using a Streptococcus mutans biofilm.

This in vitro study assessed the antibacterial effect on Streptococcus mutans biofilms of mouth rinses with 700 ppm F- (derived from NaF) that differed only in their acid compounds (malic (A), citric (B), tartaric (C), fumaric (D), hydrochloric (E), phosphoric (F), and lactic (G) acid) used to adjust pH. S. mutans (ATCC 25175) was grown for 22 h at 37°C, harvested, resuspended in simulated body fluid and biofilm formation followed for 24 h at 37°C. Thereafter, biofilms were treated with experimental rinses for 30 s, and placed in TAM48 isothermal microcalorimeter at 37°C for 72 h. Applying Gompertz growth model parameters lag time and growth rate were determined from heatflow curves; additionally, reduction of active biofilms was calculated. Moreover, samples were live/dead stained and analyzed by confocal scanning microscopy. All mouth rinses were showing statistically significant lag time and reduction of active biofilm (p<0.05, A 19.1+/-2.3h and 58.5+/-7.7%, B 15.5+/-1.1h and 41.9+/-5.3%, C 17.6+/-1.9h and 53.1+/-7.5%, D 18.4+/-2.4h and 55.8+/-8.8%, E 20.2+/-3.3h and 61.5+/-10.0%, F 20.2+/-3.0h and 61.6+/-9.3%, and G 18.3+/-2.5h and 55.3+/-8.9%). Interestingly, there were no differences found between the treated groups (p>0.05, A 0.064+/-0.004 1/h, B 0.063+/-0.005 1/h, C 0.065+/-0.004 1/h, D 0.067+/-0.004 1/h, E 0.066+/-0.006 1/h, F 0.067+/-0.004 1/h, G 0.066+/-0.006 1/h) for the maximum growth rate. Vitality staining supported these findings.. The present investigation demonstrates that the type of acid compounds used to produce the rinses did not show any negative effect on the antimicrobial properties of the tested products as all of them exhibited a similar efficacy against S. mutans biofilms.

Enhancement of hypericin nanoparticle-mediated sonoinduced disruption of biofilm and persister cells of Streptococcus mutans by dermcidin-derived peptide DCD-1L.

Streptococcus mutans is considered a major significant contributor to dental caries and its effective removal is difficult due to the formation of biofilm. Therefore, the development of adjuvant therapeutic strategies with anti-biofilm properties is a promising approach. In the present study, we examined the effect of dermcidin-derived peptide DCD-1L on the antibacterial activity of hypericin nanoparticle (HypNP)-mediated antimicrobial sonodynamic therapy (aSDT) against persister cells growing- and biofilm cultures of S. mutans. Following synthesis and confirmation of HypNP, the fractional inhibitory concentration (FIC) index of HypNP and DCD-1L was determined by checkerboard assay. Cellular uptake of HypNP-DCD-1L and generation of endogenous reactive oxygen species (ROS) were assessed and followed by the determination of antimicrobial sonoactivity of HypNP-DCD-1L against persister cells growing- and biofilm cultures of S. mutans. The water-insoluble extracellular polysaccharide (EPS) and expression of the gtfD, comDE, and smuT genes were then evaluated in persister cells growing- and biofilm cultures of S. mutans. There was a synergistic activity in the combination of HypNP and DCD-1L against S. mutans with an FIC index value of 0.37. The HypNP-DCD-1L-mediated aSDT also displayed the highest cellular uptake and endogenous ROS generation by bacterial cells. When biofilm and persister cells of S. mutans were treated with HypNP-DCD-1L and subsequently exposed to ultrasound waves, 5.1 log and 3.8 log reductions, respectively, in bacterial numbers were observed (P<0.05). According to the data, EPS in both persister cells growing- and biofilm cultures of S. mutans were significantly decreased after exposure to the HypNP-DCD-1L-mediated aSDT (P<0.05). In addition, the quantitative real-time PCR data illustrated the high level of similarities in very low-expression profiles of the gtfD before and after all treated groups for persister cells. While, following HypNP-DCD-1L-mediated aSDT treatment, the expression levels of gtfD, comDE, and smuT were significantly lower in treated persister cells growing- and biofilm cultures of S. mutans in comparison with control groups (P<0.05). Combined, the results of this study indicate that ultrasound waves-activated HypNP-DCD-1L can sonoinactivate S. mutans biofilms and persister cells, as well as reduce effectively pathogenicity potency of S. mutans. Hence, HypNP-DCD-1L-mediated aSDT may be proposed as a promising adjunctive therapeutic approach for dental caries.

Rutin-gallium complex mediated antimicrobial photodynamic therapy: An in vitro studies against Streptococcus mutans biofilms

BackgroundDiscoloration of teeth following antimicrobial photodynamic therapy (aPDT) is a serious concern. Common photosensitizers are colored, and access to a photosensitizer that does not leave color on the teeth or is the same color as the enamel and dentin is highly demanded. The physicochemical characterization, anti-virulence, and antimicrobial effects of a novel rutin-gallium (III) (Rt-Ga) complex as novel photosensitizer are presented herein. Materials and MethodsPhotophysical properties and cytotoxicity of the Rt-Ga complex were evaluated in comparison with the parental rutin. Intracellular reactive oxygen species (ROS) generation following Rt-Ga complex-mediated aPDT was measured using the fluorescent 2′,7′-dichlorofluorescein diacetate (H2DCF-DA) method. The anti-biofilm effects of Rt-Ga complex-mediated aPDT on Streptococcus mutans were assessed using a colorimetric assay. The virulence‑associated gtfB gene expression was assessed following Rt-Ga complex-mediated aPDT by quantitative real‑time PCR. ResultsThe photophysical properties of the Rt-Ga complex revealed a significant blue-shift in absorption (60 nm shift) and increased extinction coefficient (4100 M −1 cm −1; at λmax = 450 nm). Average (± SEM) DCF fluorescence intensities in an arbitrary unit (A.U.) were 7.1 ± 0.9, 4.1 ± 0.5, and 1.7 ± 0.3 for 10.0 μM of Rt-Ga complex-mediated aPDT, 7.5 μM of Rt-Ga complex-mediated aPDT, and 10.0 μM of Rt-Ga complex alone, respectively. The corresponding DCF fluorescence intensities were 710% (P = 0.001), 410% (P = 0.001), and 170% (P = 0.02) of the untreated S. mutans as the control group (1.0 ± 0.1 A.U.), respectively. The novel Rt-Ga complex-mediated aPDT exhibited no toxicity against primary human gingival fibroblast cells, a dose dependent decrease in S. mutans biofilm cell survival and virulence were observed (91.4% and 11.7-fold, respectively). ConclusionThe Rt-Ga complex-mediated anti-virulence and antimicrobial photodynamic effects were superior to the one caused by rutin alone making the Rt-Ga complex a more promising photosensitizer than the parent material.

The Activity of Calcium Glycerophosphate and Fluoride against Cariogenic Biofilms of Streptococcus mutans and Candida albicans Formed In Vitro.

This study evaluated the effects of calcium glycerophosphate (CaGP), with or without fluoride (F), on dual-species biofilms of Streptococcus mutans and Candida albicans. The biofilms were treated three times with 0.125, 0.25, and 0.5% CaGP solutions, with or without 500 ppm F (NaF). Additionally, 500 and 1100 ppm F-solutions and artificial saliva served as controls. After the final treatment, the microbial viability and biofilm structure, metabolic activity, total biomass production, and the composition of the extracellular matrix composition were analyzed. Regardless of the presence of F, 0.25 and 0.5% CaGP promoted a higher biomass production and metabolic activity increase than the controls (p < 0.05). F-free CaGP solutions reduced bacterial cell population significantly more than the 500 ppm F group or the negative control (p < 0.05). All the groups reduced the proteins, and 0.5% CaGP combined with F led to the highest reduction in the carbohydrate and nucleic acids content of the extracellular matrix (p < 0.05). It can be concluded that CaGP alone affected the number of bacterial cells and, when combined with F, reduced its production of biomass, metabolic activity, and the expression of the extracellular matrix components.