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.     

Effect of chitosan nanoparticles on the inhibition of Candida spp. biofilm on denture base surface

ObjectivesChitosan nanoparticles (ChNPs) have antifungal effects, however there is a lack of information about the effects of ChNPs against Candida biofilm on denture base surface. This study investigated the ChNPs effect against C. albicans biofilm adhesion and formation, and against Candida spp. biofilm on heat-cured acrylic resin.DesignThe ChNPs were synthetized (3800 μg/mL) and characterized by infra-red spectrophotometry and transmission electron microscopy. The minimum inhibitory/fungicidal concentrations (MIC/MFC) against Candida spp. were determined. The time-kill assay and changes on C. albicans micromorphology were evaluated. The % inhibition of ChNPs on C. albicans biofilm formation and reduction were investigated using 1 min and 8 h exposure. Candida biofilm was developed on resin surfaces and ChNPs were applied every 8 h for 5 days. After, fungal cells were counted (CFU/mL) and the surface roughness (Ra) and vickers microhardness (HV) of resin were analysed. For all experiments, sodium hypochlorite (NaOCl) was used as control. Data were analyzed by ANOVA, Tukey and paired t-tests (α = 0.05).ResultsThe MIC_80% of ChNPs was 30.1 μg/mL. ChNPs at 4 MIC showed complete inhibition in the time-kill assays. Blastoconidia cells were predominant after ChNPs application. The % inhibition ChNPs on C. albicans was proportional to its concentration, regardless of the exposure time. ChNPs decreased the CFU/mL of Candida spp. and showed lower alteration of HV and Ra values of resin surface compared to NaOCl.ConclusionsThe ChNPs inhibited C. albicans biofilm, reduced Candida biofilm on resin and caused small changes in roughness and hardness of acrylic resin surface.     

Fungicidal and inhibitory efficacy of cinnamon and lemongrass essential oils on Candida albicans biofilm established on acrylic resin: An in vitro study

Statement of problemIt is unclear whether cinnamon and lemongrass essential oils can effectively reduce the Candida-biofilm frequently formed on dental devices made from heat-polymerized polymethyl methacrylate (PMMA) resin that contributes to the development of mild oropharyngeal as well as life-threatening candidiasis in patients wearing the devices.PurposeThe purpose of this in vitro study was to determine the efficacy of cinnamon and lemongrass essential oils in eradicating Candida albicans biofilm on heat-polymerized PMMA specimens and to determine whether they retard the formation of fungal biofilm.Material and methodsThe antifungal effect of cinnamon and lemongrass essential oils was determined by using agar disk diffusion and broth microdilution methods to obtain minimum inhibitory concentrations. The mature C albicans biofilm (48 hours) was pre-established on PMMA specimens before being individually treated with various concentrations (½, 1, 2, 4, 8, 16 times minimum inhibitory concentration) of each tested oil for different exposure times (1, 2, 4, 8, and 24 hours). In another experiment, fungal biofilm was established on the PMMA specimens that were primed individually with various concentrations of the tested oils for different times. The 2,3-bis-(2-methoxy-4-nitro-5-sulfophenyl)-2H-tetrazolium-5-carboxanilide (XTT)-reduction assay was used to quantitate biofilm viability in both experiments. Statistical analysis was performed by using the 1-sample Kolmogorov-Smirnov test and 2-way ANOVA followed by the Tukey multiple comparison test (α=.05).ResultsMinimum inhibitory concentration values of cinnamon and lemongrass essential oils against planktonic C albicans were 0.1 μL/mL (0.01% v/v) and 0.4 μL/mL (0.04% v/v). At 8 times the minimum inhibitory concentration, cinnamon oil (0.8 μL/mL or 0.08% v/v) and lemongrass oil (3.2 μL/mL or 0.32% v/v) eradicated the pre-established fungal biofilm by 99.0% in an exposure time of 1 hour. In contrast, high concentrations of 8 and 16 times the minimum inhibitory concentration of cinnamon oil (0.8 μL/mL or 0.08% v/v) and lemongrass oil (6.4 μL/mL or 0.64% v/v) coated on PMMA specimens for 24 hours were only able to inhibit the formation of fungal biofilm by approximately 70.0%.ConclusionsCinnamon and lemongrass essential oils can eliminate pre-established C albicans biofilm and restrain the formation of fungal biofilm on heat-polymerized PMMA specimens. Both effects of the tested essential oils depended on dose and exposure or priming time.     

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.     

Inhibition of microbial adhesion to plastic surface and human buccal epithelial cells by Rhodomyrtus tomentosa leaf extract

Objective

The adherence of oral pathogenic microorganisms to host tissues is the initial step for successful process of oral diseases. This study aimed to determine the effect of the Rhodomyrtus tomentosa leaf extract and rhodomyrtone, an antibacterial compound from R. tomentosa leaf, on adhesion of some oral pathogens to polystyrene plastic surface and human buccal epithelial cells.

Methods

The minimum inhibitory concentration (MIC) was evaluated using broth microdilution method. The microbial adhesion to the plastic surface and buccal cells was determined using microtiter plate method and microscopy technique.

Results

The ethanol extract of leaf demonstrated antibacterial activity against oral microorganisms including Staphylococcus aureus ATCC 25923, Streptococcus mutans (clinical isolate), and Candida albicans ATCC 90028 with the MIC values of 31.25, 15.62, and 1000 μg/ml, respectively. Rhodomyrtone displayed activity with the MIC values of 0.78 and 0.39 μg/ml against S. aureus ATCC 25923 and S. mutans, respectively. The MIC value of the compound against C. albicans ATCC 90028 was more than 100 μg/ml which was the highest test concentration. All pathogenic microorganisms treated with the extract and rhodomyrtone at their subinhibitory concentrations resulted in a decrease in their adherence ability to both plastic surface and buccal cells.

Conclusion

It is suggested that R. tomentosa extract and rhodomyrtone may be useful in therapy or as prophylaxis in infections involving oral pathogens.     

Genotypic distribution of Candida albicans in dental biofilm of Chinese children associated with severe early childhood caries

AimCandida albicans causes a wide range of infections in the oral mucosa, especially candidiasis. A strong association has been found between C. albicans and dental caries. In this study, we investigated the presence and genotypic distribution of C. albicans in the dental biofilm of Chinese children with severe early childhood caries (S-ECC).DesignDental biofilm samples were collected from 41 Chinese children (21 children with S-ECC and 20 children without caries) aged 3–6 years. Samples collected were cultured in CHROMagar Candida (CA) medium. Cellular DNA of typical C. albicans isolates in the CA medium was isolated, and PCR using primers reported to span a transposable intron region in the 25S rRNA gene was performed to determine genotypic subgroups.ResultsC. albicans was detected in 57.1% of individuals with S-ECC. Additionally, three genotypic subgroups of C. albicans (genotypes A, B, and C) were found in children with S-ECC, and genotype A was dominant.ConclusionsC. albicans is an important component of dental biofilm associated with S-ECC, and C. albicans genotypic subgroup A is the dominant strain in the dental biofilm of children with S-ECC.     

Oral spray containing plant-derived compounds is effective against common oral pathogens

ObjectivesPlant-derived compounds are a good source of therapeutic agents and inhibitors of inflammatory process. Dental caries, periodontal diseases and candidiasis are common oral infections caused by virulent biofilms. The objectives of this study were to develop oral spray containing plant-derived compounds; α-mangostin (α-MG) and/or lawsone methyl ether (2-methoxy-1,4-naphthoquinone) (LME) and determine its antimicrobial, anti-biofilm, and anti-inflammatory activities.DesignOral spray formulations were prepared containing α-MG (5 mg/ml) and/or LME (250 μg/ml). Antimicrobial activity against Candida albicans, Streptococcus mutans, and Porphyromonas gingivalis and anti-biofilm formation activities were determined as well as cytotoxicity and anti-inflammatory effects.ResultsThe oral spray demonstrated antimicrobial activity against all three of the oral pathogens tested with stronger effects on C. albicans and S. mutans than P. gingivalis. The formulation containing α-MG (2.5 mg/ml) and LME (125 ug/ml) reduced growth of the microorganisms about 1–2 Log CFU/ml at 1–3 h and the killing effects were complete at 24 h. Based on biofilm assay, the oral spray containing both α-MG and LME showed greater inhibitory effects than those with α-MG or LME. In addition, the oral spray containing both α-MG and LME demonstrated more inhibition of nitric oxide production than α-MG alone. All the formulations were safe and demonstrated greater anti-inflammatory activity at lower concentration (<6.25 μg/ml) than at a higher concentration.ConclusionOral spray containing α-MG and/or LME is effective against common oral pathogens without significant cytotoxicity. Thus, it has the potential to prevent the infections and may serve as adjunctive treatment to conventional therapy.     

The in vivo anti-Candida albicans activity of flavonoids

BackgroundEmerging drug-resistant strains of Candida albicans have led to the recurrence of fungal disease, rendering conventional drug therapies ineffective. Although in vitro studies on flavonoids as novel antifungal products have shown promising results, there is currently limited information regarding their in vivo effects. The aim of this review is to evaluate in vivo studies on the antifungal activity of flavonoids against C. albicans, as novel therapeutic agents. In this regard, we conducted broad searches of PubMed, Web of Science, and Embase covering the years 2009–2020.HighlightFlavonoids represent new natural therapeutic compounds to treat oral candidiasis. Among subclasses of flavonoids, flavonols and chalcones appear to have the most significant antifungal activities. Oral administration of Canthin-6-one, a flavonol, has the potential to damage fungal cell membrane while having minimal toxic effects on mice. Similarly, topical oral application of lichochalcone-A, a chalcone, reduces oral candidiasis in mice. There appears to be structural similarities in the hydroxyl residues among compounds within the same subclass of flavonoids, which may contribute to antibiofilm activity. Oral topical application of flavonoids shows low toxicity and has clinical relevance as potential novel antifungal treatments.ConclusionFlavonoids are a group of natural products exhibiting antifungal activity. The subclasses flavonols and chalcones appear to have the most significant in vivo antifungal activity against C. albicans infections in mouse models. Specifically, quercetin (flavonol) has been applied via vaginal gavage in a murine vulvovaginal candidiasis model, whereas lichochalcone-A (chalcone) has shown topical oral application in C. albicans-inoculated mice. Both compounds show efficacy in fungal elimination via biofilm inhibition for their respective subclasses. The translational significance of these in vivo studies should be examined in clinical trials of selected potent compounds for the treatment of oral candidiasis. Further studies are necessary to elucidate the specific mechanisms of flavonoids as antifungal agents.     

Study of virulence factor of Candida species in oral lesions and its association with potentially malignant and malignant lesions

ObjectiveThe aim of this study was to explore the association between malignant and premalignant lesions and the virulence factor profile of Candida spp. recovered from different oral lesions.DesignCandida spp. isolated from malignant lesions (squamous cell carcinoma, OC, n = 25), atypical lichen planus (AL, n = 11), chronic candidiasis (CC, n = 25), and asymptomatic carriers (WI, n = 15, control strains.) Isolates were identified in chromogenic medium, colony morphology and biochemical tests. The lipolytic and proteinase activity was determined on supplemented agar with olive oil and BSA, respectively. The biofilm formation with XTT reduction assay and cellular surface hydrophobicity (CSH) by water-hydrocarbon method were performed.ResultsAll isolates recovered from oral lesions produced the four virulence factors studied with significantly higher levels than in WI isolates. Interestingly, lipolytic activity was absent in WI isolates. The proteolytic activity was similar in AL and OC isolates. OC isolates showed significantly higher CSH values than other clinical isolates. Non-albicans species showed higher biofilm formation than C.albicans (P = 0.03.) There were no significant differences in virulence factors among species. A strong positive correlation was found between proteinase and lipase activity (r = 0.90, P < 0.0001), and between hydrophobicity and biofilm (R = 0.81, P < 0.0001.)ConclusionsOur results indicate that OC Candida isolates exhibited a significant higher attributes of virulence than other lesions fungus isolates, providing evidence about the association between Candida pathogenicity and lesions severity.     

Antiadherent activity of Schinus terebinthifolius and Croton urucurana extracts on in vitro biofilm formation of Candida albicans and Streptococcus mutans

Objective

To evaluate the antiadherent property of crude, methanol and acetate methanol extract fractions from Schinus terebinthifolius and Croton urucurana in hydroalcoholic (HA) and dimethylsulfoxide (DMSO) solvents on in vitro biofilms formed by Streptococcus mutans and Candida albicans strains.

Design

The minimal concentration of adherence (MICA) was determined to evaluate the antiadherent potential of extracts on the in vitro biofilm formation. The extracts of plants were subjected to thin layer chromatography (TLC) in order to detect what class of compounds was responsible for the antiadherent activity. Data were estimated by analysis of variance (ANOVA) complemented by Tukey test level of significance set at 5%.

Results

Both plants demonstrated inhibition of S. mutans and C. albicans on in vitro biofilm formation. The biofilms of C. albicans were more efficiently inhibited by the S. terebinthifolius fraction of acetate–methanol and methanol in hydroalcoholic solvents (p < 0.05). The S. mutans biofilms adherence was best inhibited by the S. terebinthifolius crude extract and its methanolic fraction, both in hydroalcoholic solvent (p < 0.05). TLC of crude extracts and fractions of S. terebinthifolius detected the presence of several active compounds, including phenolic compounds, anthraquinones, terpenoids, and alkaloids. C. urucurana extracts confirmed activity for both microorganisms (p < 0.05). However, higher concentrations were needed to achieve antiadherent activity, mainly to inhibit in vitro biofilm formation of C. albicans.

Conclusion

The antiadherent potential of both plants on in vitro biofilms formed by C. albicans and S. mutans were confirmed, suggesting the importance of studies about these extracts for therapeutic prevention of oral diseases associated with oral biofilms.     

Susceptibility of Candida albicans and Candida dubliniensis to erythrosine- and LED-mediated photodynamic therapy

The effect of erythrosine- and LED-mediated photodynamic therapy (PDT) on planktonic cultures and biofilms of Candida albicans and Candida dubliniensis was evaluated. Planktonic cultures of standardized suspensions (10⁶ cells/mL) of C. albicans and C. dubliniensis were treated with erythrosine concentrations of 0.39–200 μM and LEDs in a 96-well microtiter plate. Biofilms formed by C. albicans and C. dubliniensis in the bottom of a 96-well microtiter plate were treated with 400 μM erythrosine and LEDs. After PDT, the biofilms were analysed by scanning electron microscopy (SEM). The antimicrobial effect of PDT against planktonic cultures and biofilms was verified by counting colony-forming units (CFU/mL), and the data were submitted to analysis of variance and the Tukey test (P < 0.05). C. albicans and C. dubliniensis were not detectable after PDT of planktonic cultures with erythrosine concentrations of 3.12 μM or higher. The CFU/mL values obtained from biofilms were reduced 0.74 log₁₀ for C. albicans and 0.21 log₁₀ for C. dubliniensis. SEM revealed a decrease in the quantity of yeasts and hyphae in the biofilm after PDT. In conclusion, C. albicans and C. dubliniensis were susceptible to erythrosine- and LED-mediated PDT, but the biofilms of both Candida species were more resistant than their planktonic counterparts.     

The antimicrobial and antibiofilm properties of allicin against Candida albicans and Staphylococcus aureus – A therapeutic potential for denture stomatitis

The objective of this study is to determine the therapeutic efficacy of allicin against Candida albicans (C. albicans) and Staphylococcus aureus (S. aureus), the common etiological agents for denture stomatitis (DS). The minimum inhibitory concentration (MICs), minimum bactericidal concentrations (MBCs) and minimum fungicidal concentration (MFCs) of allicin were determined by the broth microdilution method followed by checkerboard microdilution method for a synergistic interaction between allicin + nystatin and allicin + CHX. The potential of allicin to eradicate C. albicans and S. aureus biofilms was assessed by treating biofilm formed on self- polymerized acrylic resin with allicin at a sub-MIC concentration for 5 min. The commercial denture cleanser (brand X) was used as a positive control. A Kruskal-Wallis test followed by the post-hoc Mann-Whitney U test was applied (SPSS 20.0), and the level of significance was set at P < 0.05. Allicin exhibited antimicrobial activity against C. albicans (MIC:8 µg/ml and MFC:16 µg/ml) and S. aureus (MIC:8 µg/ml and MBC:8 µg/ml). A synergistic interaction was observed between allicin + nystatin and allicin + CHX (FICI ≤ 0.5). Allicin exhibited significant biofilm eradication against C. albicans and S. aureus biofilms with percentages of 50.0% and 52.6%, respectively. The results of this study suggest a possible application of allicin in treating C. albicans and S. aureus infection in DS.     

Effect of Long-term Exposure to Peptides on Mono- and Multispecies Biofilms in Dentinal Tubules

IntroductionThe aim of this study was to evaluate the antibiofilm effectiveness of 2% chlorhexidine (CHX) and peptides 1018 and DJK-5 used either alone or in a mixture (peptide and 2% CHX) against Enterococcus faecalis and multispecies biofilms in dentin canals after short-term and long-term exposure.MethodsOne hundred eighty dentin blocks were prepared and filled with E. faecalis or multispecies bacteria by centrifugation. Three-week-old biofilms in dentin were subjected to 2% CHX, DJK-5 (10 μg/mL), 1018 (10 μg/mL), DJK-5 + 2% CHX, or 1018 + 2% CHX for short-term (1 or 3 minutes), short-term exposure after 24 hours, and long-term exposure (24 hours of exposure). The antibacterial efficacy was determined by live/dead bacterial viability staining and confocal laser scanning microscopy.ResultsPeptide DJK-5 with or without CHX was the most effective agent against all the biofilms (P < .05), killing 77% of biofilm bacteria in 1 minute. No significant difference in bacterial killing was detected between the first 3 minutes of exposure (>81%) and after 24 hours of exposure (83%) to DJK-5 or DJK-5 + CHX. Chlorhexidine and peptide 1018 had a weaker antibiofilm effect than DJK-5, and their effect was time dependent (P < .05) with a maximum killing of 60% after 24 hours of exposure.ConclusionsPeptide DJK-5 alone and together with CHX had a rapid antibacterial effect against dentin infection. An additional antibacterial effect by CHX and peptide 1018 was achieved after a 24-hour long-term exposure.     

Antibacterial and mechanical properties of arginine-containing glass ionomer cements

ObjectiveThe study investigated the effect of incorporating l-arginine (Arg) in a glass ionomer cement (GIC) on its mechanical properties and antibacterial potential.MethodsPre-determined proportions (1%, 2%, and 4% by wt.) of Arg were incorporated in GIC powder; while GIC without Arg served as control. The flexural strength, nanohardness, surface roughness, elemental analysis using SEM-EDX (n = 6) and F/Arg/Ca/Al/Si release in deionized water for 21 days were assessed. The antibacterial potential was evaluated in a multi-species biofilm model with Streptococcus mutans, Streptococcus sanguinis, Streptococcus gordonii, and Lactobacillus acidophilus for 72 h. Real-time qPCR was used to analyse biofilm bacterial concentrations. Propidium monoazide modification of real-time qPCR was performed to quantify viable/dead bacteria. The pH, lactic acid, ADS activity, and H₂O₂ metabolism were measured. Confocal microscopy was used to investigate the biofilm bacterial live/dead cells, density, and thickness.ResultsThere was no difference in flexural strength among the different groups (p > 0.05). No significant difference in nanohardness and surface roughness was observed between 4% Arg + GIC and control (p > 0.05). The 4% Arg + GIC showed significantly higher F/Arg/Al/Si release than the other groups (p < 0.05), reduced total bacterial concentration and growth inhibition of viable S. mutans and S. sanguinis (p < 0.05). Lactic acid formation for 4% Arg + GIC was significantly higher than 1% Arg + GIC (p < 0.05). The spent media pH of 4% Arg + GIC was higher than the other groups (p < 0.05), with proportionately lower ammonia and higher H₂O₂ released (p < 0.05).SignificanceAddition of 4% l-arginine in GIC enhanced its antibacterial activity via a biofilm modulatory effect for microbial homeostasis, with no detrimental effect on its mechanical properties.     

Effectiveness of Antibiotic Medicaments against Biofilm Formation of Enterococcus faecalis and Porphyromonas gingivalis

IntroductionIn this study we compared the antibacterial effect of triple antibiotic paste (TAP), double antibiotic paste (DAP), and calcium hydroxide [Ca(OH)₂] against Enterococcus faecalis and Porphyromonas gingivalis biofilm.MethodsThe minimum inhibitory concentration (MIC), minimum bactericidal concentration (MBC), minimum biofilm inhibitory concentration (MBIC), and biofilm formation were measured by using microtiter plate methods. The 2 bacteria were treated with different dilutions of TAP, DAP, and Ca(OH)₂ solutions. The turbidities of the bacterial cultures in the microtiter plate were measured by optical density at 490 nm by using a spectrophotometer. Data were analyzed by 2-way analysis of variance (α = 0.05).ResultsFor TAP, the MIC and MBIC values were 0.003 mg/mL for E. faecalis and 0.006 mg/mL for P. gingivalis. The MBC values for TAP were 0.3 mg/mL for both bacteria. The MIC and MBIC values for DAP were 0.001 mg/mL for E. faecalis and P. gingivalis. The MBC values for DAP were 0.14 mg/mL for both bacteria. Biofilm formation of the 2 bacteria was significantly decreased with TAP and DAP at all tested dilutions (P < .0001) compared with control groups; however, TAP and DAP biofilm formations were not significantly different from each other. Ca(OH)₂ significantly decreased bacterial biofilm formation compared with the control, but it was significantly less than TAP and DAP (P < .05).ConclusionsBoth TAP and DAP were more effective than Ca(OH)₂ against E. faecalis and P. gingivalis. DAP can be considered an effective and comparable antibacterial substitute for TAP.     

Synergistic activity of lysozyme and antifungal agents against Candida albicans biofilms on denture acrylic surfaces

Denture related oral candidiasis is a recalcitrant fungal infection not easily resolved by topical antifungals. The antimycotic protein lysozyme, in saliva is an important host defense mechanism although its activity against Candida biofilms on denture acrylic has not been evaluated.

Objectives

(i) To establish a clinically relevant denture acrylic assay model to develop standardized Candida albicans biofilms, and (ii) assess the inhibitory effects of lysozyme alone and, the latter combined with antifungals (nystatin, amphotericin B, ketoconazole and 5-fluorocytosine) on sessile Candida cells and, finally (iii) to visualize the accompanying ultrastructural changes.

Design

The rotating-disc biofilm reactor was used to develop standardized 48 h Candida biofilms on acrylic discs in YNB/100 mM glucose medium and the biofilm metabolic activity was monitored using a tetrazolium reduction assay.

Results

The biofilm metabolic activity was similar in 18 identical denture acrylic discs (p < 0.05) thus validating the rotating-disc biofilm model. Very low concentrations of lysozyme (6.25 μg/ml) significantly (p < 0.01) inhibited Candida biofilm formation indicating that lysozyme may likely regulate intra-oral Candida biofilm development. Although 100 μg/ml lysozyme killed 45% of sessile Candida cells, further increasing its concentration (up to 240 μg/ml) had no such effect. Nystatin, amphotericin B, and ketoconazole in association with 100 μg/ml lysozyme exhibited effective synergistic killing of biofilm Candida in comparison to drug-free controls. Scanning electron and confocal scanning laser microscopy analysis confirmed the latter trends.

Conclusion

Our results indicate that agents found in biological fluids such as lysozyme could be a safe adjunct to antifungals in future treatment strategies for recalcitrant candidal infections.     

Evaluation of antibiofilm and mechanical properties of new nanocomposites based on acrylic resins and silver vanadate nanoparticles

ObjectiveThe purpose of this study was evaluate, for the first time, the impact of incorporation of nanostructured silver vanadate (β-AgVO₃) in antibiofilm and mechanical properties of dental acrylic resins (poly(methyl methacrylate), PMMA).DesignThe β-AgVO₃ was synthesized and characterized by X-ray diffraction (XRD), Fourier transform infrared spectroscopy (FTIR), scanning electron microscopy, and microanalysis (SEM/EDS). Resins specimens were prepared with 0–10% wt.% β-AgVO₃ and characterized by SEM, XRD and optical microscopy_. The antibiofim activity of the samples against Candida albicans and Streptococcus mutans was investigated by XTT reduction test, colony-forming units (CFUs), and confocal laser scanning microscopy (CLSM). The flexural strength, hardness, and surface roughness of the samples containing β-AgVO₃ were compared with the pure PMMA matrix.ResultsThe incorporation of 10% β-AgVO₃ significantly reduced the metabolic activity of C. albicans and S. mutans (p < 0.05). There was a reduction in microbial load (CFU/mL) of microorganisms for the different concentrations used (p < 0.05), which was confirmed by confocal microscopy. The addition of β-AgVO₃ did not change the mechanical properties of hardness and surface roughness of the resins (p > 0.05). However, flexural strength decreased with the addition of amounts greater than 1% (p < 0.05).Conclusionsβ-AgVO₃ additions in dental acrylic resin may have an impact on inhibition of biofilm of main microorganisms associated with dental prostheses. However, the viability of clinical use should be evaluated in function of changed promoted in some mechanical properties.     

Correction to: Synergistic effect of thymoquinone and nystatin in the treatment of oral candidiasis; an in vitro study

The effectiveness of antifungal agents may be insufficient against resistant strains in some cases of oral candidiasis. The aim of this study was to evaluate the antifungal effect of thymoquinone against Candida albicans, Candida tropicalis, Candida glabrata and Candida krusei strains and the synergistic antifungal activity of these strains in combination with nystatin. To evaluate in vitro antifungal activity and interactions between thymoquinone and nystatin, substances were tested against Candida albicans ATCC 10,231, C. tropicalis ATCC 750, C.krusei ATCC 6258 and C. glabrata ATCC 2001 standard strains both individually and combinationally via microdilution method. MIC and ΣFIC index value were analysed. The Kruskal Wallis test and Bonferroni test were used for statistical evaluations. Statistical significance was set at p < 0.05. A statistically significant difference was observed between the mean ranks of all Candida species and doses of thymoquinone, nystatin, and the combination thymoquinone-nystatin (p < 0.05). MIC values for thymoquinone were determined as 15 μg/mL for C. albicans, C. tropicalis and C. krusei while it was 30 μg/mL for C. glabrata. Moreover, MIC for nystatin was found as 1.875 μg/mL for C. albicans, C. tropicalis and C. krusei, whereas it was 7.5 μg/mL in C. glabrata. Interaction assays and ΣFIC index value revealed that, TQ and nystatin have a synergistic effect against to all strains. Thymoquinone was found to have antifungal activity on Candida species and synergistic effect when combined with nystatin.

     

Cytotoxicity and estrogenicity in simulated dental wastewater after grinding of resin-based materials

ObjectiveThis study evaluated the cytotoxic and estrogenic effects of dust and eluates released into simulated wastewater after grinding of dental resin-based materials.MethodsFour materials were used: ceram.x® universal, Filtek™ Supreme XTE, Lava™ Ultimate and Core-X™ flow. From each composite material, samples (5 × 2 mm, n = 50) were prepared according to the manufacturers’ instructions. Lava™ Ultimate was used as blocks. All samples were ground to dust with a diamond bur (106 μm) and suspended in distilled water at 60 mg/mL. After storage for 72 h, the suspensions were separated into a soluble (eluate) and a particulate (dust) fraction. Eluates and dusts were evaluated for inhibition of Vibrio fischeri bioluminescence and cytotoxicity on human A549 lung cells (WST-1-Assay). The estrogenic activity was assessed by YES-Assay using Saccharomyces cerevisiae. Additionally, dental monomers (BisGMA, BisEMA, UDMA, TEGDMA, HEMA) and Bisphenol A were investigated.ResultsAll eluates showed inhibition of V. fischeri bioluminescence at concentrations above 1.1 mg/mL (p < 0.05). The activity of the eluates of ceram.x® universal and Filtek™ Supreme XTE was significantly higher than Lava™ Ultimate and Core-X™ flow (p < 0.05). In the WST-1-Assay, all materials induced cytotoxic effects at concentrations of 0.1 mg/mL (p < 0.05), while no significant differences were detected among them. The tested materials revealed no estrogenic activity. All dental monomers and Bisphenol A showed concentration dependent cytotoxic effects (p < 0.05), whereas only Bisphenol A induced an estrogenic effect (p < 0.01).SignificanceDust and eluates of resin-based dental materials released into wastewater exert bactericidal and cytotoxic effects in vitro. However, they reveal no estrogenic effect.     

Interaction of Candida albicans with periodontal ligament fibroblasts limits biofilm formation over elastomer silicone disks

ObjectiveCandida albicans is the most numerous commensal and potentially pathological yeast in the human oral cavity. The purpose herein is to investigate the ability of C. albicans to form a biofilm in the presence of periodontal ligament (PDL) fibroblasts.Material and methodsSilicone elastomer disks (SE) were transferred to wells containing PDL cells. C. albicans suspension was added to each well. The whole mixed culture was then allowed to form a biofilm for 48 h. Biofilms were quantified by tetrazolium-salt-based (2,3-bis(2-methoxy-4-nitro-5-sulfophenyl)-5-[(phenyl amino) carbonyl]- 2H-tetrazolium hydroxide (XTT). Furthermore, biofilm was visualized by confocal scanning laser and scanning electron microscopy. Migration of C. albicans and its ability to form biofilms in presence of PDL cells was determined by using a transwell system. Last, elutes obtained from co-culturing C. albicans and PDL cells were added to SE disks and covered with C. albicans. The culture plate was then incubated to allow biofilm formation. Biofilms formed over SE disks were quantified using XTT.ResultsPDL cells significantly limited the biofilm formation at incubation interval of 48 h. PDL cells induced less biofilm compared to mature and thick hyphae in the absence of PDL cells as seen in confocal scanning laser and scanning electron microscopy. The presence of PDL cells limited the migration and formation of biofilm by C. albicans. Elutes obtained from co-culturing PDL cells with C. albicans for one hour induced significantly less biofilm.ConclusionsThis is the first study to report that PDL cells exhibit antifungal activity. While the exact mechanism of how PDL cells limited biofilm formation is yet unknown, it was clear that competent PDL cells promote resistance to C. albicans biofilm formation.