Direct and indirect effects of different dentifrices on the initial bacterial colonization of enamel in situ overnight.

OBJECTIVE: The aim of this study was to investigate the direct and indirect influence of fluoridated toothpastes and fluoride-free toothpaste with hydroxyapatite (HAP) as active ingredient on initial bacterial colonization on enamel in situ. METHODS: For this clinical-experimental pilot study, eight subjects were instructed to brush their teeth with three different toothpastes (Elmex® : 1400 ppm AmF, Meridol® : 1400 ppm AmF +SnF2, Karex® : HAP), using each for two consecutive days. As a control, brushing without toothpaste was performed. To evaluate bacterial colonization, subject wore splints with buccally placed bovine enamel platelets overnight. Two modes were tested. In a first pass (regimen A), the splints were inserted after toothbrushing to examine the indirect effects of the dentifrices. In order to investigate the direct effects, the specimens were brushed in situ in a second pass (regimen B). Biofilm formation was visualized and quantified using fluorescence microscopy (DAPI and BacLight) and transmission electron microscopy (TEM). RESULTS: For brushing regimen A (indirect effect of dentifrices), no statistical differences were detected between any of the tested dentifrices or the control. Likewise, no statistically significant differences were recorded for brushing regimen B (direct effect of dentifrices). Furthermore, no differences between the different brushing techniques were determined with regard to the ultrastructure of the overnight biofilm. CONCLUSION: Within the limitations of the present pilot study, it can be concluded that in patients with good oral hygiene, dentifrices and their chemical composition have no statistically significant effect on the initial bacterial colonization of enamel platelets in situ, irrespectively of the mode of application.

Hydroxyapatite-Based Solution as Adjunct Treatment for Biofilm Management: An In Situ Study.

Synthetic hydroxyapatite-based solution is a bioinspired material that may present anti-adhesive properties, restraining the dental biofilm formation without causing adverse effects. This in situ study aims to evaluate the effects of three different hydroxyapatite (HAP) watery solutions as a mouthwash against biofilm adhesion on different dental material surfaces under oral conditions. Hence, four volunteers carried maxillary splints containing enamel, titanium, ceramics, and polymethyl-methacrylate resin (PMMA) samples. Three HAP watery solutions (5%) were prepared with HAP particles presenting different shapes and sizes (HAP I, HAP II, HAP III). During 24 h, the volunteers rinsed two times with one of the following selected tested solution: HAP I, HAP II, HAP III, water, or chlorhexidine 0.2% (CHX). The first rinse was performed 3 min after pellicle formation; the second rinse occurred after a 12 h interval. The surface analysis was performed by scanning electron microscopy (SEM), fluorescence microscopy (FM), and transmission electron microscopy (TEM). Statistical and microscopic analysis showed that most samples treated with any HAP solution revealed reduced biofilm coverage presenting comparable results to CHX treated samples, however without altering the microorganisms' viability. In conclusion, the results of this investigation showed that a pure hydroxyapatite-based mouthrinse could be a promising bioinspired adjunct solution for biofilm management.

Modification of in situ Biofilm Formation on Titanium by a Hydroxyapatite Nanoparticle-Based Solution.

Oral biofilms play an essential role on peri-implant disease development. Synthetic hydroxyapatite nanoparticles (nHAP) are a bioinspired material that has structural and functional similarities to dental enamel apatite and may provide preventive properties against biofilm formation. This study aimed to investigate the effects of an experimental nHAP solution on biofilm formation on polished and non-polished titanium under oral conditions. Five volunteers carried maxillary splints with non-polished and polished titanium and followed a 48 h rinsing protocol with the proposed nHAP solution, and with chlorhexidine 0.2% (CHX) and water, as controls. Samples were analyzed by fluorescence microscopy (FM), scanning electron microscopy (SEM) and transmission electron microscopy (TEM). FM showed a significant reduction of biofilms on polished samples treated with nHAP (p = 0.0485) compared with water, without differences between nHAP and CHX (p > 0.9999). Analyzing biofilm viability, polished samples rinsed with nHAP showed significantly fewer dead bacteria than CHX (p = 0.0079), but there was no significant difference in viability between polished samples rinsed with water and nHAP (p = 0.9268). A significantly higher biofilm coverage was observed on the non-polished surfaces compared to the polished surfaces when nHAP was applied (p = 0.0317). This difference between polished and non-polished surfaces was not significant when water (p = 0.1587) or CHX (p = 0.3413) rinsing were applied. SEM and TEM analysis supported the FM findings, that polished samples rinsed with nHAP presented fewer biofilm coverage compared to samples rinsed with water. In conclusion, the nHAP solution reduced the biofilm formation on polished Ti surfaces without altering bacterial viability, providing a novel approach for the management of biofilm formation on biomaterials.

Effect of fragaria vesca, hamamelis and tormentil on the initial bacterial colonization in situ.

OBJECTIVE: The presentin situ study aims to examine the influence of the polyphenolic tea drugs fragaria vesca, hamamelis and tormentil on the initial oral bioadhesion. DESIGN: Initial biofilm formation was performed on bovine enamel slabs which were carried intraorally by 12 subjects. After 1 min of intraoral pellicle formation, the subjects rinsed with fragaria vesca, tormentil (0.8 mg/8 mL) and hamamelis (0.2 mg/8 mL) for 10 min. Tap water served as negative control, 0.2 % CHX as positive control. The investigations took place on different days (wash-out: 2 days). Afterwards, fluorescence microscopy has been performed per test solution (n = 5) and per subject (n = 12) to visualize bacterial adhesion and glucan formation (8 h oral exposition) with DAPI, ConA and BacLight. Additionally, TEM was used to visualize the pellicle ultrastructure and expectorate samples. Statistical evaluation was carried out using the Kruskal-Wallis- (p < 0.5), Mann-Whitney U test (p < 0.5) and Bonferroni-Holm-correction (p < 0.1). RESULTS: Rinsing with the polyphenolic tea extracts reduced significantly initial bacterial colonization (DAPI) compared to the negative control. There was no significant difference betweenfragaria vesca, hamamelis and tormentil. All solutions showed a reducing effect on the glucan formation. No significant difference was observed between fragaria vesca and CHX. Considerable alterations of the pellicle's ultrastructure manifested by an increase in thickness and electron density resulted from rinsing with the three polyphenolic aqueous extracts. CONCLUSIONS: Fragaria vesca, hamamelis and tormentil significantly reduce initial bioadhesion and glucan formation in situ and are therefore recommended as adjuvant antibacterial oral therapeutics.

Activity and distribution pattern of enzymes in the in-situ pellicle of children.

OBJECTIVE: This study investigated, for the first time, pellicle enzymes with respect to their activity, distribution and fluorescence pattern in children with different caries experience. DESIGN: In-situ pellicles were collected from 41 children (aged 4-6 years) with different caries status; 17 of them were caries-free (dmf = 0), 12 had dental restorations but no current caries (dmf ≥ 2) and 12 had at least two carious lesions (dmf ≥ 2). Bovine enamel samples were fixed on individual upper jaw braces for pellicle formation. After 30 min of intraoral exposure, the pellicle and saliva samples were analysed for the activities of amylase, lysozyme, peroxidase and glucosyltransferase (GTF). The distribution of these enzymes, including GTF-isoforms B, C and D, and the pellicle ultrastructure were examined by gold-immunolabelling and transmission electron microscopy (TEM). Furthermore, interactions between pellicle enzymes and adherent bacteria were visualised using combined fluorescence and immunofluorescence labelling. RESULTS: There were no significant differences in the pellicle enzyme activities between the study groups. TEM analysis revealed the absence of GTF C and D in the pellicle of caries-active children. Amylase, peroxidase and GTF-isoforms showed a random distribution within the pellicle layer; lysozyme was found in the form of clusters. A similar ultrastructural pattern was observed for all subjects. Fluorescence labelling technique enabled visualisation of all enzymes, except for GTF B. CONCLUSION: Pellicle enzyme activities and ultrastructure are not associated with children's caries status. Further investigation is needed to assess the influence of individual GTF-isoforms on caries susceptibility in children.