Estudio in vivo del efecto sobre la microbiota oral de la aplicación de un barniz de flúor tras la reducción del esmalte interproximal con fines ortodóncicos

Abstract

Interproximal enamel reduction (IPR) is a technique commonly applied in orthodontics in order to obtain more space to align teeth and maintain long-term alignment. It has been shown that the IPR procedure results in an enamel surface that is more susceptible to demineralization phenomena, therefore, regardless of the IPR method used, it is important to protect that surface. Fixed orthodontic appliances such as brackets create new undercut areas favorable to plaque accumulation, which causes a change in the oral microbiota. Microorganisms are analyzed and classified with 16S RNA gene sequencing techniques. Within the context in which oral diseases are generated by dysbiosis, our study aims to determine if the application of an orthodontic appliance to a patient destabilizes the system and generates dysbiosis, for this reason we will characterize the oral microbiota of patients wearing fixed multi-brackets appliances. On the other hand, most of the published studies on the oral microbiota do not take into account that there may be different microorganisms depending on the surface of the tooth, so we find it interesting to be able to study the microbiota that exists on the different surfaces of the enamel (vestibular -V-, palatal -P- and interproximal -IP-) in individuals. To the best of our knowledge, there are no studies in the literature that evaluate in vivo changes in the oral microbiota in patients with multi-bracket fixed appliances who undergo IPR and fluoride varnish is applied as a preventive measure, which motivated us to to carry out this esearch whose general objective was to determine if the microbiota of the interproximal surface of the enamel in patients wearing multibracket fixed appliances is affected by the IPR procedure and the application of a fluoride varnish. To this end, 20 patients who were candidates for wearing fixed orthodontic appliances were selected from whom plaque samples were taken from the V, P and IP surfaces of the upper right (experimental side) and left (control side) first premolars. The patients were divided into two groups (n=10): group 1 (undergoing IPR) and group 2 (undergoing IPR with fluoride varnish application). The sampling was carried out in a period of 3 months with four different times, namely: T0: Oral hygiene instructions were given to patients and fluoride-free toothpaste was given. Plaque samples were taken. They had previously been told that they should not brush from the night before the sample was taken. T1: 1 month after T0, plaque samples were taken prior to the cementation of the appliance in the patients. T2: 1 month after the cementation of the appliance, plaque samples were taken. During this time, the IPR procedure was performed on both groups and after it, Bifluorid® 12 was applied to group 2. The IPR was performed between the first upper right premolar (1.4) and the second upper right premolar (1.5) with separation strips (Horico®, Berlin, Germany) and diamond bur (Komet®). Bifluorid® 12 fluoride varnish was applied after the IPR procedure in group 2 patients on the interproximal surface of 1.4 and 1.5 following the manufacturer's instructions: with the dental surfaces dry, it was applied using a brush (Pele Tim®). distributing it on the surfaces leaving a thin layer. T3: 2 months after the cementation of the appliance. The last samples were taken and the patient was told that he could continue using his usual toothpaste. All patients underwent two lactate measurements, one at baseline and another after 10 minutes. These measurements were taken from the same sample. A pH measurement was also performed. The DNA extraction from our samples was carried out in one of the laboratories of FISABIO, the Foundation for the Promotion of Health and Biomedical Research of the Valencian Community, with the MagNA PURE LC DNA Isolation kit II robot (Roche®). The readings obtained after massive sequencing of the hypervariable region of the 16S ribosomal gene were analyzed using the Dada2 program. The entire comparison and statistical process was performed using R (programming language and an environment for statistical and graphical analysis). The p value was adjusted using the false discovery rate (FDR) system (p<0.05). The significance of the differences between the proportions of each taxon, between the pH and lactate values between the groups, was calculated using the Wilcoxon test (p<0.05). In our study, differences were also observed between bacteria a month after cementing the brackets, producing an increase in bacteria associated with gingivitis and periodontitis. Bacterial diversity also increases in IP and P, but not in the V surface. The fact that there is less diversity in the V surface may be related to hygiene. Regarding the IPR, no significant differences were observed after performing this procedure, it seems that the microbiota stabilizes over time, although it is interesting to observe that all the species that are more abundant before the IPR are strictly anaerobic while those that are after of IPR can live in the presence of oxygen. The species that increased most significantly after the application of a fluoride varnish was Haemophilus parainfluenzae, associated with health. On the other hand, we found genres related to oral diseases that decreased significantly after the application of fluoride. By analyzing changes in pH, we find that pH tends to improve. After performing IPR, the pH tends to worsen in group 1 (IPR), but not in group 2 (IPR+F-), therefore, a buffer effect is created when fluoride is administered that maintains the pH. It is an expected and promising result, although the differences are not significant.