Fabíola Galbiatti de Carvalho

Analysis by confocal laser scanning microscopy of the MDPB bactericidal effect on S. mutans biofilm CLSM analysis of MDPB bactericidal effect on biofilm

Since bacteria remain in the dentin following caries removal, restorative materials with antibacterial properties are desirable to help maintaining the residual microorganisms inactive. The adhesive system Clearfil Protect Bond (PB) contains the antibacterial monomer 12-methacryloyloxydodecylpyridinium bromide (MDPB) in its primer, which has shown antimicrobial activity. However, its bactericidal effect against biofilm on the dentin has been little investigated. Objective The aim of this study was to analyze by confocal laser scanning microscopy (CLSM) and viable bacteria counting (CFU) the MDPB bactericidal effect against S. mutans biofilm on the dentin surface. Material and Methods Bovine dentin surfaces were obtained and subjected to S. mutans biofilm formation in BHI broth supplemented with 1% (w/v) sucrose for 18 h. Samples were divided into three groups, according to the primer application (n=3): Clearfil Protect Bond (PB), Clearfil SE Bond, which does not contain MDPB, (SE) and saline (control group). After the biofilm formation, Live/Dead stain was applied directly to the surface of each sample. Next, 10 µL of each primer were applied on the samples during 590 s for the real-time CLSM analysis. The experiment was conducted in triplicate. The primers and saline were also applied on the other dentin samples during 20, 90, 300 and 590 s (n=9 for each group and period evaluated) and the CFU were assessed by colonies counting. Results The results of the CLSM showed that with the SE application, although non-viable bacteria were detected at 20 s, there was no increase in their count during 590 s. In contrast, after the PB application there was a gradual increase of non-viable bacteria over 590 s. Conclusions The quantitative analysis demonstrated a significant decrease of S. mutans CFU at 90 s PB exposure and only after 300 s of SE application. Protect Bond showed an earlier antibacterial effect than SE Bond.

Antifungal activity, mode of action and anti-biofilm effects of Laurus nobilis Linnaeus essential oil against Candida spp.

OBJECTIVE The present study demonstrated the antifungal potential of the chemically characterized essential oil (EO) of Laurus nobilis L. (bay laurel) against Candida spp. biofilm adhesion and formation, and further established its mode of action on C. albicans. METHODS L. nobilis EO was obtained and tested for its minimum inhibitory and fungicidal concentrations (MIC/MFC) against Candida spp., as well as for interaction with cell wall biosynthesis and membrane ionic permeability. Then we evaluated its effects on the adhesion, formation, and reduction of 48hC. albicans biofilms. The EO phytochemical profile was determined by gas chromatography coupled to mass spectrometry (GC/MS). RESULTS The MIC and MFC values of the EO ranged from (250 to 500) μg/mL. The MIC values increased in the presence of sorbitol (osmotic protector) and ergosterol, which indicates that the EO may affect cell wall biosynthesis and membrane ionic permeability, respectively. At 2 MIC the EO disrupted initial adhesion of C. albicans biofilms (p<0.05) and affected biofilm formation with no difference compared to nystatin (p>0.05). When applied for 1min, every 8h, for 24h and 48h, the EO reduced the amount of C. albicans mature biofilm with no difference in relation to nystatin (p>0.05). The phytochemical analysis identified isoeugenol as the major compound (53.49%) in the sample. CONCLUSIONS L. nobilis EO has antifungal activity probably due to monoterpenes and sesquiterpenes in its composition. This EO may affect cell wall biosynthesis and membrane permeability, and showed deleterious effects against C. albicans biofilms.

Protective effect of calcium nanophosphate and CPP-ACP agents on enamel erosion

The aim of this study was to assess the effect of different remineralizing agents on enamel microhardness (KHN) and surface topography after an erosive challenge. Forty-eight human enamel specimens (4 × 4 mm) were randomly assigned to 4 groups: control (no treatment), fluoride varnish, calcium nanophosphate paste and casein phosphopeptide-amorphous calcium phosphate paste (CPP-ACP). Both pastes were applied for 5 minutes, and fluoride varnish, for 24 h. Four daily erosive cycles of 5 minutes of immersion in a cola drink and 2 h in artificial saliva were conducted for 5 days. KHN readings were performed at baseline and after 5 days. The percentage of enamel hardness change (%KHN) was obtained after erosion. The surface topography was evaluated by atomic force microscopy (AFM). The data were tested using ANOVA, Tukeys and paired-T tests (p < 0.05). After an erosive challenge, there was no statistically significant difference between the control (96.8 ± 11.4 KHN / 72.4 ± 3.0%KHN) and the varnish (91.7 ± 14.1 KHN / 73.4 ± 5.5%KHN) groups. The nanophosphate group showed lower enamel hardness loss (187.2 ± 27.9 / 49.0 ± 7.9%KHN), compared with the CPP-ACP group (141.8 ± 16.5 / 60.6 ± 4.0%KHN), and both were statistically different from the varnish and the control groups. AFM images showed a rough surface for the control and the varnish groups, a non-homogeneous layer with globular irregularities for CPP-ACP, and a thick homogeneous layer for the nanophosphate group. None of the agents provided protection against the development of erosion; however, nanophosphate paste was able to reduce enamel surface softening after the erosive challenge.

Fluoride Release/Uptake from Different Orthodontic Adhesives: A 30-Month Longitudinal Study

The aim of this study was to test the null hypothesis that there is no difference in fluoride release between resin-modified glass ionomer cements (RMGICs) and composites in the long term. The materials were divided into 5 groups: a nonfluoride-releasing composite - Group TXT (Transbond XT), a fluoride-releasing composite - Group QC (Quick-Cure), and three RMGICs - Groups FOLC, FOB and MC (Fuji Ortho LC, Fuji Ortho Band, and Multi-Cure). Fluoride release was measured at time intervals of 1 h, 1, 7, 14, 21 and 29 days, followed by further evaluations performed at 6, 12, 18, 24 and 30 months using selective ion electrodes connected to an ionic analyzer. Fluoride releasing and re-releasing experiments were analyzed using the Kruskal-Wallis test and Mann-Whitney test with the Bonferroni correction. The amount of fluoride released by FOB was larger in comparison with the other adhesives (p=0.01). In the long-term, FOLC and MC had a similar performance (p>0.05). The composites presented a low fluoride release, but fluoride ion uptake and re-release capacity of QC was statistically significant (p<0.05) during the experiment. In conclusion, the null hypothesis was rejected, the RMGIC Fuji Ortho Band and the composite Quick-Cure presented greater fluoride release and re-release capacity when recharged.

Mechanical and Biological Properties of Acrylic Resins Manipulated and Polished by Different Methods

This study evaluated the influence of the manipulation technique and polishing method on the flexural strength and cytotoxicity of acrylic resins. Two manipulation techniques and three polishing methods were used in the fabrication of acrylic plates that were divided into 6 groups (n=10). Groups MM, MC and MW: mass technique with mechanical polishing, chemical polishing and without polishing, respectively; and Groups SM, SC and SW: Saturation technique with mechanical polishing, chemical polishing and without polishing, respectively). Flexural strength was tested in a universal testing machine and the cytotoxicity assay used cell cultures (L-929) for periods of 24 h to 168 h. Flexural strength and cytotoxicity data were assessed using two-way and three-way ANOVA, respectively (α=0.05), followed by post hoc Bonferroni test for multiple comparisons. The effect of combinations of manipulation techniques and polishing methods on flexural strength showed significant differences only between Group SC and Groups MW, MM and MC (p<0.01). Cell viability ranged from 51% (3.9%) to 87,6% (3.2) in the 24-h time interval, and from 87.8% (5.0) to 95.7% (3.1%) in the 168-h time interval. With the increase of cell viability, from the third day (72 h), there was no significant difference among the groups, except between MM and SC (p<0.01) at 72 h. In conclusion, the manipulation technique and polishing method had more influence on the cytotoxicity than on flexural strength.

In vivo biocompatibility versus degree of conversion of resin-reinforced cements in different time periods

This study focused on test the null hypothesis that there is no difference between the degree of conversion and biocompatibility of different resin reinforced glass ionomer cements (RRGICs). Forty‐eight male Wistar rats were used, distributed into four groups (n = 12), as follows: Group C (Control, polyethylene), Group FOB (Fuji Ortho Band), Group UBL (Ultra band Lok), and Group MCG (Multicure Glass), in subcutaneous tissue. The events of edema, necrosis, granulation tissue, multinuclear giant cells, young fibroblasts, and collagen formation were analyzed at 7, 15, and 30 days. The degree of conversion was evaluated by the Fourier method. Biocompatibility and degree of conversion were assessed using the Kruskal–Wallis and Dunn tests, and ANOVA and Tukeys test, respectively (P < 0.05). It was observed that, there was significant difference between Groups FOB and UBL for the presence of young fibroblasts at 15 days (P = 0.034) and between the Control and MCG Groups for the presence of multinucleated giant cells at 30 days (P = 0.009). Monomer conversion increased progressively until day 30, with significant difference between Group FOB and Groups UBL and MCG (P = 0.013) at 15 days. The null hypothesis was partially accepted, Fuji Ortho Band showed a less monomer conversion and a smaller number of young fibroblasts in the time of 15 days. Microsc. Res. Tech. 77:335–340, 2014.

Histological analysis of biocompatibility of ionomer cements with an acid-base reaction

The purpose of this study was to evaluate the inflammatory and cure events of acid-based reactions using glass ionomer cement used for cementation of crowns, bridges, onlays and orthodontic bands implanted in subcutaneous tissue, at different time intervals. A total of 48 male Wistar rats were used, distributed into 4 groups (n = 12), as follows: Group C (control, polyethylene), Group ME (Meron), Group KC (Ketac Cem) and Group PR (Precedent). The animals were sacrificed after time intervals of 7, 15 and 30 days, and their tissues were analyzed under an optical microscope for such events as inflammatory infiltrate, edema, necrosis, granulation tissue, multinucleated giant cells, young fibroblasts and collagen. The results was assessed using Kruskal-Wallis and Dunns tests (p < 0.05). In the initial period, intense inflammatory infiltrate was observed for all the materials with no significant difference among them (p = 0.104). Groups PR and KC showed significant difference in relation to Group C, at 7 days (p = 0.025) and 15 days (p = 0.006). Edema and giant cells were more expressive in Group ME, differing significantly from Groups C (p = 0.023) and KC (p = 0.039), respectively, at 7 days. Group ME showed a statistically significant difference in relation to Groups PR and KC for the presence of young fibroblasts (p = 0.009) and for collagen (p = 0.002), at 7 days. Within the limits of this in vivo study, Precedent and Ketac Cem glass ionomer cements showed better tissue healing with a greater number of fibroblasts and collagen, as compared to Meron.

Thickness and nanomechanical properties of protective layer formed by TiF4 varnish on enamel after erosion

The layer formed by fluoride compounds on tooth surface is important to protect the underlying enamel from erosion. However, there is no investigation into the properties of protective layer formed by NaF and TiF4 varnishes on eroded enamel. This study aimed to evaluate the thickness, topography, nanohardness, and elastic modulus of the protective layer formed by NaF and TiF4 varnishes on enamel after erosion using nanoindentation and atomic force microscopy (AFM). Human enamel specimens were sorted into control, NaF, and TiF4 varnish groups (n = 10). The initial nanohardness and elastic modulus values were obtained and varnishes were applied to the enamel and submitted to erosive challenge (10 cycles: 5 s cola drink/5 s artificial saliva). Thereafter, nanohardness and elastic modulus were measured. Both topography and thickness were evaluated by AFM. The data were subjected to ANOVA, Tukeys test and Students t test (α = 0.05). After erosion, TiF4 showed a thicker protective layer compared to the NaF group and nanohardness and elastic modulus values were significantly lower than those of the control group. It was not possible to measure nanohardness and elastic modulus in the NaF group due to the thin protective layer formed. AFM showed globular deposits, which completely covered the eroded surface in the TiF4 group. After erosive challenge, the protective layer formed by TiF4 varnish showed significant properties and it was thicker than the layer formed by NaF varnish.

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

OBJECTIVES Chitosan 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. DESIGN The 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). RESULTS The MIC80% 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. CONCLUSIONS The ChNPs inhibited C. albicans biofilm, reduced Candida biofilm on resin and caused small changes in roughness and hardness of acrylic resin surface.

Efeito do flúor e do xilitol na atividade antimicrobiana de dentifrícios infantis

The aim of this study was to evaluate the antimicrobial effect toothpastes for infants containing fluoride and xylitol presented against Streptococcus mutans, through an agar diffusion test. Six toothpastes were categorized according to the presence/concentration of fluoride and xylitol: 1-Condor Bambinos; 2-Bitufo Cocorico; 3-Colgate Smiles; 4-Bitufo Penelope; 5-Tandy and 6-Aquafresh Kids. After the solidification of an agar medium in the dishes, 200 µL of the inoculum (S. mutans UA 159) were dispensed on the medium and five holes were made in it for the application of the dentifrices, totalizing 10 wells for each one. The plates were incubated at 37oC for 48h. The inhibition zones were analyzed through the ANOVA one-way and Tukey (α=0.05) tests. There was a significant difference between fluoridated dentifrices, as the Aquafresh showed the biggest inhibition halos against S. mutans. The concentration of fluoride influenced the antimicrobial activity of child toothpastes; xylitol, however, had no influence.