Effect of arginine on the growth and biofilm formation of oral bacteria

BackgroundAlkali production via arginine deiminase system (ADS) of oral bacteria plays a significant role in oral ecology, pH homeostasis and inhibition of dental caries. ADS activity in dental plaque varies greatly between individuals, which may profoundly affect their susceptibility to caries.ObjectiveTo investigate the effect of arginine on the growth and biofilm formation of oral bacteria.Methods and resultsPolymicrobial dental biofilms derived from saliva were formed in a high-throughput active attachment biofilm model and l-arginine (Arg) was shown to reduce the colony forming units (CFU) counts of such biofilms grown for various periods or biofilms derived from saliva of subjects with different caries status. Arg hardly disturbed bacterial growth of Streptococcus mutans, Streptococcus sobrinus, Streptococcus sanguinis and Streptococcus gordonii in BHI medium, but only inhibited biofilm formation of S. mutans. Scanning electron microscope (SEM) showed S. mutans biofilms harboured fewer cells grown with Arg than that without Arg, even in the initial 2 h and 8 h phase. Confocal laser scanning microscope (CLSM) images of poly-microbial dental and S. mutans biofilms revealed the biofilms grown with Arg had lower exopolysaccharide (EPS)/bacteria ratios than those without Arg (P = 0.004, 0.002, respectively). Arg could significantly reduce the production of water-insoluble EPS in S. mutans biofilms (P < 0.001); however, quantitative real-time PCR (qRT-PCR) did not show significantly influence in gene expression of gtfB, gtfC or gtfD (P = 0.32, 0.06, 0.44 respectively).ConclusionsArg could reduce the biomass of poly-microbial dental biofilms and S. mutans biofilms, which may be due to the impact of Arg on water-insoluble EPS. Considering the contribution to pH homeostasis in dental biofilms, Arg may serve as an important agent keeping oral biofilms healthy thus prevent dental caries.     

Novel pit and fissure sealant containing nano-CaF2 and dimethylaminohexadecyl methacrylate with double benefits of fluoride release and antibacterial function

ObjectivePit and fissure sealants with antibacterial and remineralization properties have broad application prospects in caries prevention. The objectives of this study were to: (1) develop a novel pit and fissure sealant containing CaF₂ nanoparticles (nCaF₂) and dimethylaminohexadecyl methacrylate (DMAHDM); and (2) investigate the effects of nCaF₂ and DMAHDM on biofilm response and fluoride (F) ion release for the first time.MethodsHelioseal F was used as a control. Bioactive sealants were formulated with DMAHDM and nCaF₂. Flow properties, enamel shear bond strength, hardness and F ion releases were measured. Streptococcus mutans (S. mutans) biofilms were grown on sealants. Biofilm metabolic activity, lactic acid production, colony-forming units (CFU), and pH of biofilm culture medium were measured.ResultsAdding 5% DMAHDM and 20% nCaF₂ did not reduce the paste flow and enamel bond strength, compared to control (p < 0.05). Hardness of sealants with 20% nCaF₂ and DMAHDM was higher than control (p < 0.05). The F ion release from 20% nCaF₂ was much higher than that of commercial control (p < 0.05). The sealant with DMAHDM reduced the S. mutans biofilm CFU by 4 logs. The pH in biofilm medium of the new bioactive sealant was much higher (pH 6.8) than that of commercial sealant (pH 4.66) (p < 0.05).SignificanceThe new bioactive pit and fissure sealant with nCaF₂ and DMAHDM achieved high fluoride release and strong antibacterial performance. This novel fluoride-releasing and antibacterial sealant is promising to inhibit caries and promote the remineralizaton of enamel and dentin.     

In vitro evaluation of composite containing DMAHDM and calcium phosphate nanoparticles on recurrent caries inhibition at bovine enamel-restoration margins

ObjectiveRecurrent caries is a primary reason for restoration failure caused by biofilm acids. The objectives of this study were to: (1) develop a novel multifunctional composite with antibacterial function and calcium (Ca) and phosphate (P) ion release, and (2) investigate the effects on enamel demineralization and hardness at the margins under biofilms.MethodsDimethylaminohexadecyl methacrylate (DMAHDM) and nanoparticles of amorphous calcium phosphate (NACP) were incorporated into composite. Four groups were tested: (1) Commercial control (Heliomolar), (2) Experimental control (0% DMAHDM + 0% NACP), (3) antibacterial group (3% DMAHDM + 0% NACP), (D) antibacterial and remineralizing group (3% DMAHDM + 30% NACP). Mechanical properties and Ca and P ion release were measured. Colony-forming units (CFU), lactic acid and polysaccharide of Streptococcus mutans (S. mutans) biofilms were evaluated. Demineralization of bovine enamel with restorations was induced via S. mutans, and enamel hardness was measured. Data were analyzed via one-way and two-way analyses of variance and Tukey’s multiple comparison tests.ResultsAdding DMAHDM and NACP into composite did not compromise the mechanical properties (P > 0.05). Ca and P ion release of 3% DMAHDM + 30% NACP was increased at cariogenic low pH. Biofilm lactic acid and polysaccharides were greatly decreased via DMAHDM, and CFU was reduced by 4 logs (P < 0.05). Under biofilm acids, enamel hardness at the margins was decreased to about 0.5 GPa for control; it was about 1 GPa for antibacterial group, and 1.3 GPa for antibacterial and remineralizing group (P < 0.05).ConclusionsThe novel 3% DMAHDM + 30% NACP composite had strong antibacterial effects. It substantially reduced enamel demineralization adjacent to restorations under biofilm acid attacks, yielding enamel hardness that was 2-fold greater than that of control composites. The novel multifunctional composite is promising to inhibit recurrent caries.     

Comparative effect of a stannous fluoride toothpaste and a sodium fluoride toothpaste on a multispecies biofilm

ObjectivesThis paper aimed to compare the mode of action of a stannous fluoride-containing toothpaste with a conventional sodium fluoride-containing toothpaste on anti-biofilm properties.MethodsA three-species biofilm model that consists of Streptococcus mutans, Streptococcus sanguinis and Porphyromonas gingivalis was established to compare the anti-biofilm properties of a stannous fluoride-containing toothpaste (CPH), a conventional sodium fluoride-containing toothpaste (CCP) and a negative control (PBS). The 48 h biofilms were subjected to two-minute episodes of treatment with test agents twice a day for 5 consecutive days. Crystal violet staining and XTT assays were used to evaluate the biomass and viability of the treated biofilm. Live/dead staining and bacteria/extracellular polysaccharides (EPS) double-staining were used to visualize the biofilm structure and to quantify microbial/extracellular components of the treated biofilms. Species-specific fluorescent in situ hybridization and quantitative polymerase chain reaction (qPCR) were used to analyze microbial composition of the biofilms after treatment.ResultsThe biomass and viability of the biofilms were significantly reduced after CPH toothpaste treatment. The inhibitory effect was further confirmed by the live/dead staining. The EPS amounts of the three-species biofilm were significantly reduced by CCP and CPH treatments, and CPH toothpaste demonstrated significant inhibition on EPS production. More importantly, CPH toothpaste significantly suppressed S. mutans and P. gingvalis, and enriched S. sanguinis in the three-species biofilm. In all experiments CPH had a significantly greater effect than CCP (p < 0.05) and CCP had a greater effect than PBS (p < 0.05).ConclusionsStannous fluoride-containing toothpaste not only showed better inhibitory effect against oral microbial biofilm, but was also able to modulate microbial composition within multi-species biofilm compared with conventional sodium fluoride-containing toothpaste.     

Correlation of Streptococcus mutans and Streptococcus sanguinis colonization and ex vivo hydrogen peroxide production in carious lesion-free and high caries adults

Objective

This study was conducted to estimate oral colonization by Streptococcus mutans and Streptococcus sanguinis in adults with high and without any caries experience. Furthermore, differences in the amount of hydrogen peroxide (H₂O₂) produced by S. sanguinis isolated from both groups were assessed.

Design

Forty adults were divided into: (i) carious lesion-free, without any carious lesion, assessed by the International Caries Detection and Assessment System (ICDAS), or restoration, (CF) and (ii) high caries experience (HC). Saliva samples were collected and seeded on respective agar-plates for enumeration of total streptococci, S. mutans and S. sanguinis (CFU/mL) and compared between groups. Additionally, S. sanguinis colonies obtained from both groups were inoculated on Prussian blue agar for H₂O₂ detection. Production of H₂O₂ was quantified and compared between the two groups. S. sanguinis counts were significantly higher in CF than HC individuals (p < 0.05). Conversely, S. mutans showed significantly higher levels in HC than CF subjects (p < 0.001). S. sanguinis colonies from CF individuals produced significantly larger H₂O₂ halos compared with HC subjects.

Conclusions

S. sanguinis predominates over S. mutans in saliva of adults without caries experience. In those people, S. sanguinis produces more H₂O₂ex vivo.     

Cytotoxicity of novel fluoride solutions and their influence on mineral loss from enamel exposed to a Streptococcus mutans biofilm

ObjectiveThis study evaluated the cytotoxicity, antimicrobial activity and in vitro influence of new fluoridated nanocomplexes on dental demineralization.DesignThe nanocomplexes hydroxypropyl-β-cyclodextrin with 1% titanium tetrafluoride (TiF₄) and γ-cyclodextrin with TiF₄ were compared to a positive control (TiF₄), a blank control (without treatment) and negative controls (hydroxypropyl-β-cyclodextrin, γ-cyclodextrin, deionized water), following 12- and 72-hour complexation periods. The cytotoxicity was assessed using the neutral red dye uptake assay at T1–15 min, T2–30 min and T3–24 h. A minimum bactericidal concentration (MBC) against Streptococcus mutans (ATCC 25175) was performed. Enamel blocks were exposed to an S. mutans biofilm, and the percentage of surface microhardness loss was obtained. Biocompatibility and microhardness data were analysed using ANOVA/Tukey tests (p < 0.05).ResultsAt T1, the cell viability results of the nanocomplexes were similar to that of the blank control. At T2 and T3, the 72 h nanocomplexes demonstrated cell viability results similar to that of the blank, while the 12 h solutions showed results different from that of the blank (p < 0.05). All fluoridated nanocompounds inhibited S. mutans (MBC = 0.25%), while the MBC of TiF₄ alone was 0.13%. All fluoridated compounds presented a percentage of surface microhardness loss lower than that of deionized water (p < 0.05).ConclusionsThe new fluoridated nanocomplexes did not induce critical cytotoxic effects during the experimental periods, whilst they did show bactericidal potential against S. mutans and inhibited enamel mineral loss.     

Antimicrobial and antibiofilm action of Casbane Diterpene from Croton nepetaefolius against oral bacteria

ObjectiveThe antibacterial activity of Casbane Diterpene (CD) was evaluated in vitro against Streptococcus oralis, S. mutans, S. salivarius, S. sobrinus, S. mitis and S. sanguinis. The viability of planktonic cells was analysed by susceptibility tests (MIC and MBC) and antibiofilm action was assayed.MethodsThe minimal inhibitory and bactericidal concentrations (MIC and MBC) of oral Streptococcus were evaluated through microdilution tests. To assay antibiofilm activity, biofilms were generated on 96-wells polystyrene plates under the presence of CD and quantified by a crystal violet technique and colonies forming units counting.ResultsThe CD isolated from Croton nepetaefolius showed antimicrobial effect on planktonic forms and biofilms of oral pathogens, with MIC values of 62.5 μg/mL against Streptococcus oralis and values between 125 and 500 μg/mL against S. mutans, S. salivarius, S. sobrinus, S. mitis and S. sanguinis. CD showed an inhibitory effect on S. mutans biofilm formation at 250 μg/mL, and a decrease on viable cell of 94.28% compared to the normal biofilm growth.ConclusionsThe compound CD can be considered as a promising molecule for the treatment against oral pathogens responsible for dental biofilm.     

Efficacy of red propolis hydro-alcoholic extract in controlling Streptococcus mutans biofilm build-up and dental enamel demineralization

ObjectiveThe efficacy of a red propolis hydro-alcoholic extract (RP) in controlling Streptococcus mutans biofilm colonization was evaluated. The effect of RP on dental demineralization was also investigated.MethodsChemical composition was determined by High Performance Liquid Chromatography (HPLC). Minimum Inhibitory and Bactericidal Concentration (MIC and MBC, respectively) were investigated against Streptococcus mutans (ATCC 25175). The cytotoxic potential of 3% RP in oral fibroblasts was observed after 1 and 3 min. Bovine dental enamel blocks (N = 24) were used for S. mutans biofilm formation (48 h), simulating ‘feast or famine’ episodes. Blocks/biofilms were exposed 2×/day, for 3 days, to a cariogenic challenge with sucrose 10% (5 min) and treated (1 min) with: 0.85% saline solution (negative control), 0.12% Chlorhexidine (CHX, positive control for biofilm colonization), 0.05% Sodium Fluoride (NaF, positive control to avoid demineralization) and 3% RP. Biofilms were assessed for viability (CFU/mL), and to observe the concentration of soluble and insoluble extracellular polysaccharides (SEPS and IEPS). Dental demineralization was assessed by the percentage of surface hardness loss (%SHL) and through polarized light microscopy (PLM).ResultsThe RP presented 4.0 pH and ºBrix = 4.8. The p-coumaric acid (17.2 μg/mL) and luteolin (15.23 μg/mL) were the largest contents of phenolic acids and flavonoids, respectively. MIC and MBC of RP were 293 μg/mL and 1172 μg/mL, respectively. The 3% RP showed 43% of viably cells after 1 min. Lower number (p < 0.05) of viable bacteria (CFU/mL) was observed after CHX (1.8 × 10⁵) followed by RP (1.8 × 10⁷) treatments. The lowest concentration (μg/CFU) of SEPS (12.6) and IEPS (25.9) was observed in CHX (p < 0.05) followed by RP (17.1 and 54.3), and both differed from the negative control (34.4 and 63.9) (p < 0.05). Considering the %SHL, all groups differed statistically (p < 0.05) from the negative control (46.6%); but NaF (13.9%), CHX (20.1%) and RP (20.7%) did not differ among them (p > 0.05). After all treatments, suggestive areas of caries lesions were observed by PLM, which were lower for CHX and NaF.ConclusionThe 3% RP reduced S. mutans colonization, decreased concentration of extracellular polysaccharides and reduced dental enamel demineralization.     

Stoichiometric models of sucrose and glucose fermentation by oral streptococci: Implications for free acid formation and enamel demineralization

ObjectivesThe objective of this study is to develop stoichiometric models of sugar fermentation and cell biosynthesis for model cariogenic Streptococcus mutans and non-cariogenic Streptococcus sanguinis to better understand and predict metabolic product formation.MethodsStreptococcus mutans (strain UA159) and Streptococcus sanguinis (strain DSS-10) were grown separately in bioreactors fed brain heart infusion broth supplemented with either sucrose or glucose at 37 °C. Cell mass concentration and fermentation products were measured at different hydraulic residence times (HRT) to determine cell growth yield.ResultsSucrose growth yields were 0.080 ± 0.0078 g cell/g and 0.18 ± 0.031 g cell/g for S. sanguinis and S. mutans, respectively. For glucose, this reversed, with S. sanguinis having a yield of 0.10 ± 0.0080 g cell/g and S. mutans 0.053 ± 0.0064 g cell/g. Stoichiometric equations to predict free acid concentrations were developed for each test case. Results demonstrate that S. sanguinis produces more free acid at a given pH than S. mutans due to lesser cell yield and production of more acetic acid. Greater amounts of free acid were produced at the shortest HRT of 2.5 hr compared to longer HRTs for both microorganisms and substrates.SignificanceThe finding that the non-cariogenic S. sanguinis produces greater amounts of free acids than S. mutans strongly suggests that bacterial physiology and environmental factors affecting substrate/metabolite mass transfer play a much greater role in tooth or enamel/dentin demineralization than acidogenesis. These findings enhance the understanding of fermentation production by oral streptococci and provide useful data for comparing studies under different environmental conditions.     

Evaluation of ion release and the recharge ability of glass-ionomer cement containing BioUnion filler using an in vitro saliva-drop setting assembly

ObjectiveA glass-ionomer cement (GIC) containing BioUnion filler has been reported to release Zn²⁺ under acidic conditions and to inhibit oral bacteria on its surface. However, previous results are based on in vitro experiments under static conditions. This study aimed to assemble an in vitro saliva-drop setting to simulate in vivo conditions of the oral cavity and to investigate the ion releasing and recharging properties of the GIC containing BioUnion filler.MethodsThe effective concentrations of Zn²⁺ and F^? against Streptococcus mutans and saliva-derived multi-species biofilms were determined. Artificial saliva was dropped on the GIC containing BioUnion filler using the in vitro saliva-drop setting assembly and was periodically replaced with acetic acid. Ion release/recharge properties were investigated by measuring the release concentrations of Zn²⁺ and F^?.ResultsThe concentration of Zn²⁺ released from the BioUnion filler-containing GIC during seven days with repeated exposure to acid could be maintained at the level to inhibit S. mutans and saliva-derived multi-species biofilm formation. Moreover, the BioUnion filler-containing GIC could be recharged with Zn²⁺ and F^? by the application of a tooth gel containing Zn²⁺ and F^?. The release concentration of Zn²⁺ after recharging was significantly higher than the effective concentration of Zn²⁺ to hinder S. mutans and saliva-derived multi-species biofilm formation on material surfaces.SignificanceThe GIC containing BioUnion filler was shown to have the potential to inhibit biofilm formation in the oral cavity. In addition, recharging Zn²⁺ and F^? would further enhance the effect of the GIC containing BioUnion filler.     

In silico analysis of the competition between Streptococcus sanguinis and Streptococcus mutans in the dental biofilm

During dental caries, the dental biofilm modifies the composition of the hundreds of involved bacterial species. Changing environmental conditions influence competition. A pertinent model to exemplify the complex interplay of the microorganisms in the human dental biofilm is the competition between Streptococcus sanguinis and Streptococcus mutans. It has been reported that children and adults harbor greater numbers of S. sanguinis in the oral cavity, associated with caries‐free teeth. Conversely, S. mutans is predominant in individuals with a high number of carious lesions. Competition between both microorganisms stems from the production of H₂O₂ by S. sanguinis and mutacins, a type of bacteriocins, by S. mutans. There is limited evidence on how S. sanguinis survives its own H₂O₂ levels, or if it has other mechanisms that might aid in the competition against S. mutans, nonetheless. We performed a genomic and metabolic pathway comparison, coupled with a comprehensive literature review, to better understand the competition between these two species. Results indicated that S. sanguinis can outcompete S. mutans by the production of an enzyme capable of metabolizing H₂O₂. S. mutans, however, lacks the enzyme and is susceptible to the peroxide from S. sanguinis. In addition, S. sanguinis can generate energy through gluconeogenesis and seems to have evolved different communication mechanisms, indicating that novel proteins may be responsible for intra‐species communication.     

Antibacterial and physical properties of EGCG-containing glass ionomer cements

Objectives

To evaluate the effect of the addition of epigallocatechin-3-gallate (EGCG) on the antibacterial and physical properties of glass ionomer cement (GIC).

Methods

A conventional GIC, Fuji IX, was used as a control. EGCG was incorporated into GIC at 0.1% (w/w) and used as the experimental group. Chlorhexidine (CHX) was added into GIC at 1% (w/w) as a positive control. The anti-biofilm effect of the materials was assessed by a colorimetric technique (MTT assay) and scanning electron microscopy (SEM). The leaching antibacterial activity of the materials on Streptococcus mutans was evaluated by an agar-diffusion test. The flexural strength of the materials was evaluated using a universal testing machine and the surface microhardness was measured using a microhardness tester. The fluoride-releasing property of the materials was tested by ion chromatography.

Results

The optical density (OD) values of the GIC-EGCG group were significantly decreased at 4 h compared with the GIC group, but only a slightly decreased tendency was observed at 24 h (P > 0.05). No inhibition zones were detected in the GIC group during the study period. Significant differences were found between each group (P < 0.05). Compared with the control group, there was a significant increase in the flexural strength and surface microhardness for the GIC-EGCG group (P < 0.05). The fluoride ion release was not influenced by EGCG-incorporation (P > 0.05).

Conclusions

These findings suggested that GIC-containing 0.1% (w/w) EGCG is a promising restorative material with improved mechanical properties and a tendency towards preferable antibacterial properties.

Clinical significance

Modification of the glass ionomer cements with EGCG to improve the antibacterial and physical properties showed some encouraging results. This suggested that the modification of GIC with EGCG might be an effective strategy to be used in the dental clinic. However, this was only an in vitro study and clinical trials would need to verify true outcomes.     

Hydroalcoholic extracts of Myracrodruon urundeuva All. and Qualea grandiflora Mart. leaves on Streptococcus mutans biofilm and tooth demineralization

ObjectivesThis study evaluated the effect of the hydroalcoholic extracts of Myracrodruon urundeuva All. and Qualea grandiflora Mart. leaves (alone or combined) on the viability of Streptococcus mutans biofilm and on the prevention of enamel demineralization.MethodsStrain of S. mutans (ATCC 21175) was reactivated in BHI broth. Minimum inhibitory concentration, minimum bactericidal concentration, minimum inhibition biofilm concentration and minimum eradication biofilm concentration were determined in order to choose the concentrations to be tested under biofilm model. S. mutans biofilm (5 × 10⁵ CFU/ml) was produced on bovine enamel, using McBain saliva under 0.2% sucrose exposure, for 3 days. The biofilm was daily treated with the extracts for 1 min. The biofilm viability was tested by fluorescence and the enamel demineralization was measured using TMR.ResultsMyracrodruon urundeuva All. (Isolated or combined) at the concentrationsc ≥0.625 mg/ml was able to reduce bacteria viability, while Qualea Grandflora Mart. alone had antimicrobial effect at 5 mg/ml only (p < 0.05). On the other hand, none of the extracts were able to reduce enamel demineralization.ConclusionsThe hydroalcoholic extracts of Myracrodruon urundeuva All. and Qualea grandiflora Mart. leaves (isolated or combined) have antimicrobial action; however, they do not prevent enamel caries under S. mutans biofilm model.     

Dual-functional adhesive containing amorphous calcium phosphate nanoparticles and dimethylaminohexadecyl methacrylate promoted enamel remineralization in a biofilm-challenged environment

ObjectiveThe cariogenic biofilm on enamel, restoration, and bonding interface is closely related to dental caries and composite restoration failure. Enamel remineralization at adhesive interface is conducive to protecting bonding interface and inhibiting secondary caries. This study intended to assess the remineralization efficiency of adhesive with dimethylaminohexadecyl methacrylate (DMAHDM) and nanoparticles of amorphous calcium phosphate (NACP) on initial caries lesion of biofilm-coated enamel.MethodsArtificial initial carious lesion was created via 72-hour immersion in demineralization solution and cariogenic biofilm was formed after 24-hour culture of Streptococcus mutans (S. mutans). Specimens were then divided into 4 groups: enamel control, enamel treated with NACP, DMAHDM and NACP+DMAHDM respectively. Samples next underwent 7-day cycling, 4 h in BHIS (brain heart infusion broth containing 1 % sucrose) and 20 h in AS (artificial saliva) per day. The pH of BHIS was tested daily. So did the concentration of calcium and phosphate in BHIS and AS. Live/dead staining, colony-forming unit (CFU) count, and lactic acid production of biofilms were measured 7 days later. The enamel remineralization efficiency was evaluated by microhardness testing and transverse microradiography (TMR) quantitatively.ResultsEnamel of NACP+DMAHDM group demonstrated excellent remineralization effectiveness. And the NACP+DMAHDM adhesive released a great number of Ca²⁺ and PO₄^3- ions, increased pH to 5.81 via acid neutralization, decreased production of lactic acid, and reduced CFU count of S. mutans (P < 0.05).SignificanceThe NACP+DMAHDM adhesive would be applicable to preventing secondary caries, strengthening enamel-adhesive interface, and extending the lifespan of composite restoration.     

Antibacterial and mechanical properties of propolis added to glass ionomer cement

Objective:To investigate whether adding ethanolic extracts of propolis (EEP) might influence the antibacterial and mechanical (shear-peel band strength [SPBS]) properties of conventional glass ionomer cement (GIC) used in orthodontic band cementation.Materials and Methods:The cement was divided into four groups: one using the original composition and three with 10%, 25%, and 50% EEP added to the liquid and then manipulated. An antimicrobial assay, broth-dilution method was used to determine the antibacterial capacity of the GIC containing EEP. Eighty teeth were used for the mechanical assay, and an Instron testing machine was used to evaluate the SPBS. Kolmogorov-Smirnov and Kruskal-Wallis tests were used for statistical analyses.Results:GIC with the addition of 25% and 50% EEP activated inhibition of Streptococcus mutans (ATCC 25175) growth, but this effect did not occur in the group to which 10% EEP was added or in the control GIC group. There was no significant difference between the groups in terms of SPBS (P > .05).Conclusions:The addition of EEP may increase antibacterial properties without negatively modifying the mechanical properties of conventional GIC.     

New perspective to improve dentin–adhesive interface stability by using dimethyl sulfoxide wet-bonding and epigallocatechin-3-gallate

ObjectivesTo determine whether dentin–adhesive interface stability would be improved by dimethyl sulfoxide (DMSO) wet-bonding and epigallocatechin-3-gallate (EGCG).MethodsEtched dentin surfaces from sound third molars were randomly assigned to five groups according to different pretreatments: group 1, water wet-bonding (WWB); group 2, 50% (v/v) DMSO wet-bonding (DWB); groups 3–5, 0.01, 0.1, and 1 wt% EGCG-incorporated 50% (v/v) DMSO wet-bonding (0.01%, 0.1%, and 1%EGCG/DWB). Singlebond universal adhesive was applied to the pretreated dentin surfaces, and composite buildups were constructed. Microtensile bond strength (μTBS) and interfacial nanoleakage were respectively examined after 24 h water storage or 1-month collagenase ageing. In situ zymography andStreptococcus mutans (S. mutans) biofilm formation were also investigated.ResultsAfter collagenase ageing, μTBS of groups 4 (0.1%EGCG/DWB) and 5 (1%EGCG/DWB) did not decrease (p > 0.05) and was higher than that of the other three groups (p < 0.05). Nanoleakage expression of groups 4 and 5 was less than that of the other three groups (p < 0.05), regardless of collagenase ageing. Metalloproteinase activities within the hybrid layer in groups 4 and 5 were suppressed. Furthermore, pretreatment with 1%EGCG/DWB (group 5) efficiently inhibited S. mutans biofilm formation along the dentin–adhesive interface.SignificanceThis study suggested that the synergistic action of DMSO wet-bonding and EGCG can effectively improve dentin–adhesive interface stability. This strategy provides clinicians with promising benefits to achieve desirable dentin bonding performance and to prevent secondary caries, thereby extending the longevity of adhesive restorations.     

Chemical composition of Pistacia vera L. oleoresin and its antibacterial,anti-virulence and anti-biofilm activities against oral streptococci,including Streptococcus mutans

ObjectiveThe aim of this study was to characterize the chemical composition of oleoresin of Pistacia vera L. and to determine its antimicrobial and anti-virulence activity versus selected oral streptococci.DesignA gaschromatografic analysis of the oleoresin was performed. The antimicrobial and anti-virulence activity of the oleoresin and its fractions was evaluated by the Minimum Inhibitory Concentration (MIC) and/or Minimum Bactericidal Concentration (MBC), biofilm production and haemolytic activity inhibition experiments.ResultsThe oleoresin MBCs were ≥1024 μg/mL for all tested strains; the neutral and acidic fraction MBCs ranged from 128 to 2048 μg/mL. Essential oil’s MBCs (from 256 to 2048 μg/mL) were almost identical to MICs, suggesting a bactericidal effect. P. vera oleoresin at sub-lethal concentrations significantly reduced biofilm production by Streptococcus mutans (up to 49.4%) and by Streptococcus sanguinis (up to 71.2%). In addition, the acidic fraction showed a specific anti-biofilm activity against S. mutans (up to 41.3% reduction). A significant dose-dependent reduction in the haemolytic activity of S. mutans (up to 65.9%) and of S. anginosus (up to 78.3%) was observed after growth in the presence of oleoresin at sub-lethal concentrations. The acidic fraction reduced haemolytic activity (up to 54.3% at 64 μg/mL) of S. mutans only.ConclusionsGiven the anti-virulence activity of the P. vera oleoresin and its acidic fraction against S. mutans, our findings suggest their potential use in oral hygiene. These data represent the first step in the exploitation of P. vera L. oleoresin.     

Novel nanotechnology and near-infrared photodynamic therapy to kill periodontitis-related biofilm pathogens and protect the periodontium

ObjectivePeriodontal tissue destruction and tooth loss are increasingly a worldwide problem as the population ages. Periodontitis is caused by bacterial infection and biofilm plaque buildup. Therefore, the objectives of this study were to: (1) develop a near-infrared light (NIR)-triggered core-shell nanostructure of upconversion nanoparticles and TiO₂ ([email protected]₂), and (2) investigate its inhibitory effects via antibacterial photodynamic therapy (aPDT) against periodontitis-related pathogens.MethodsThe core β-NaYF₄:Yb³⁺,Tm³⁺ were synthesized via thermal decomposition and further modified with the TiO₂ shell via a hydrothermal method. The core-shell structure and the upconversion fluorescence-induced aPDT treatment via 980 nm laser were studied. Three periodontitis-related pathogens Streptococcus sanguinis (S. sanguinis), Porphyromonas gingivalis (P. gingivalis) and Fusobacterium nucleatum (F. nucleatum) were investigated. The killing activity against planktonic bacteria was detected by a time-kill assay. Single species 4-day biofilms on dentin were tested by live/dead staining, colony-forming units (CFU), and metabolic activity.ResultsThe hexagonal shaped [email protected]₂ had an average diameter of 39.7 nm. [email protected]₂ nanoparticles had positively charged (+12.4 mV) surface and were biocompatible and non-cytotoxic. Under the excitation of NIR light (980 nm), the core NaYF₄:Yb³⁺,Tm³⁺ UCNPs could emit intense ultraviolet (UV) light, which further triggered the aPDT function of the shell TiO₂ via energy transfer, thereby realizing the remarkable antibacterial effects against planktons and biofilms of periodontitis-associated pathogens. NIR-triggered [email protected]₂ achieved much greater reduction in biofilms than control (p < 0.05). Biofilm CFU was reduced by 3–4 orders of magnitude via NIR-triggered aPDT, which is significantly greater than that of negative control and commercial aPDT control groups. The killing ef?cacy of [email protected]₂-based aPDT against the three species was ranked to be: S. sanguinis < F. nucleatum = P. gingivalis. Metabolic activities of biofilms were also greatly reduced via NIR-triggered aPDT (p < 0.05).SignificanceUpconversion fluorescence-based aPDT achieved strong inhibiting effects against all three species of periodontitis-related pathogens. This novel nanotechnology demonstrated a high promise to inhibit periodontitis, with exciting potential to combat other oral infectious diseases such as deep endodontic infections.     

Characterization of polymethylmethacrylate microspheres loaded with silver and doxycycline for dental materials applications

ObjectivesThe manufacturing of polymethylmethacrylate(PMMA) microspheres loaded with doxycycline(DOX) and/or silver sulfate(Ag₂SO₄) to be incorporated into glass ionomer cement(GIC).MethodsPMMA microspheres were manufactured with Ag₂SO₄(1–5%) and/or DOX(5–15%). Particle size, encapsulation efficiency and drug release were measured by light microscope, ICP, and HPLC. Microspheres were added to a dental GIC(20%w/w). Drug release and DTS were investigated. Minimum inhibitory concentration and antibacterial effects of PMMA microspheres into GIC materials were tested.ResultsThe median diameter of 50 µm was obtained for microspheres. DOX was encapsulated at an efficiency of 8.3% using a theoretical loading of 15%DOX + 5%Ag₂SO₄. The Ag₂SO₄ encapsulation efficiency was 0.63% using a theoretical loading of 5%AgSO₄. All groups showed burst release within the first day and continued released up to 15 days, with 60–83% of DOX and approximately 30% of silver. For GIC, approximately 15% of DOX and 0.18% of silver were released in a 7-day period. Microbiological results showed an antimicrobial effect against S. mutans when the lead formulation of microspheres was added. The DTS was reduced by the inclusion of microspheres.SignificancePMMA microspheres containing DOX and Ag₂SO₄ offer a sustained antimicrobial activity for dental applications and promising potential for the biomedical field.