Saliva Decreases Sucrose-Induced Cariogenicity in an Experimental Biological Caries Model.

OBJECTIVE: Whether a minimum quantity of saliva inhibit the caries process remains uncertain. This study aimed to investigate the impact of saliva dilutions on an in vitro caries model using Streptococcus mutans (S. mutans) biofilms. METHODS: S. mutans biofilms were cultivated on enamel and root dentin slabs, in culture media containing different proportions of saliva (v/v): 0%, 5%, 10%, 25%, 50%, 75%, and 100% saliva, and exposed to a 10% sucrose solution (5 min, 3x/day), with appropriate controls. After 5 (enamel) and 4 (dentin) days, demineralization, biomass, viable bacteria, and polysaccharide formation were analyzed. The acidogenicity of the spent media was monitored overtime. Each assay was performed in triplicate across two independent experiments (n = 6). RESULTS: In both enamel and dentin, an inverse relationship was observed between acidogenicity, demineralization, and the proportion of saliva. Even small quantities of saliva incorporated into the media led to a noticeable reduction in enamel and dentin demineralization. Saliva presence resulted in significant reductions in biomass, viable S. mutans cells, and polysaccharides, with the effects being concentration-dependent for both tissues. CONCLUSIONS: High quantities of saliva can almost completely inhibit sucrose-induced cariogenicity, while even small amounts exhibit a dose-dependent caries-protective effect.

Extended biofilm formation time by Streptococcus sanguinis modifies its non-cariogenic behavior, in vitro.

Although the commensal Streptococcus sanguinis [ S. sanguinis] is isolated from caries-free people, it can ferment carbohydrates producing acids. We aimed to characterize S. sanguinis cariogenic potential as a function of different enamel biofilm formation periods, in vitro. Saliva-coated enamel slabs were inoculated with S. sanguinis to form initial biofilms for 8, 12 or 16 h in presence of sucrose and followed by a period in medium with glucose for 16, 12 or 8 h, respectively, until completion of 24 h. To simulate cariogenic challenges, S. sanguinis biofilms were exposed to 10% sucrose for 5 minutes, 3x/day for 5 days. Biofilm biomass, viable cells, total proteins, intracellular and extracellular polysaccharides production, acidogenicity and enamel demineralization were determined. Biofilms of Streptococcus mutans [ S. mutans ] served as caries-positive control. Biofilms of S. sanguinis forming on enamel for 12 and 16 h showed higher demineralization than those formed during 8 h, but lower than S. mutans biofilms, regardless of the initial biofilm formation time. No differences were detected in the biofilm properties among the different biofilm formation times tested for S. sanguinis . Increased enamel initial biofilm formation time by S. sanguinis appears to induce a cariogenic potential, but lower than S. mutans .

Extended biofilm formation time by Streptococcus sanguinis modifies its non-cariogenic behavior, in vitro

Abstract Although the commensal Streptococcus sanguinis [ S. sanguinis] is isolated from caries-free people, it can ferment carbohydrates producing acids. We aimed to characterize S. sanguinis cariogenic potential as a function of different enamel biofilm formation periods, in vitro. Saliva-coated enamel slabs were inoculated with S. sanguinis to form initial biofilms for 8, 12 or 16 h in presence of sucrose and followed by a period in medium with glucose for 16, 12 or 8 h, respectively, until completion of 24 h. To simulate cariogenic challenges, S. sanguinis biofilms were exposed to 10% sucrose for 5 minutes, 3x/day for 5 days. Biofilm biomass, viable cells, total proteins, intracellular and extracellular polysaccharides production, acidogenicity and enamel demineralization were determined. Biofilms of Streptococcus mutans [ S. mutans ] served as caries-positive control. Biofilms of S. sanguinis forming on enamel for 12 and 16 h showed higher demineralization than those formed during 8 h, but lower than S. mutans biofilms, regardless of the initial biofilm formation time. No differences were detected in the biofilm properties among the different biofilm formation times tested for S. sanguinis . Increased enamel initial biofilm formation time by S. sanguinis appears to induce a cariogenic potential, but lower than S. mutans .

Development of a Topical Amphotericin B and Bursera graveolens Essential Oil-Loaded Gel for the Treatment of Dermal Candidiasis.

The higher molecular weight and low solubility of amphotericin B (AmB) hinders its topical administration. The aim of this study was to incorporate Bursera graveolens essential oil into an AmB topical gel (AmB + BGEO gel) in order to promote the diffusion of the drug through the skin in the treatment of cutaneous candidiasis. AmB + BGEO gel formulation was determined using a factorial experiment. Physical and chemical parameters, stability, in vitro release profile and ex vivo permeation in human skin were evaluated. In vitro antimicrobial activity was studied using strains of C. albicans, C. glabrata and C. parapsilosis. The tolerability was evaluated using in vitro and in vivo models. AmB + BGEO gel presented appropriate characteristics for topical administration, including pH of 5.85, pseudoplastic behavior, optimal extensibility, as well as high stability and acceptable tolerability. In vitro release studies showed that the formulation releases the drug following a Boltzmann sigmoidal model. Finally, AmB + BGEO gel exhibited higher amount of drug retained inside the skin and lower Minimum Inhibitory Concentration than a formulation sans essential oil. Therefore, these results suggest that the incorporation of B. graveolens essential oil in the formulation could be used as strategy to promote a local effect in the treatment of cutaneous candidiasis.

Competition and Caries on Enamel of a Dual-Species Biofilm Model with Streptococcus mutans and Streptococcus sanguinis.

Imbalances within the dental biofilm trigger dental caries, currently considered a dysbiosis and the most prevalent noncommunicable disease. There is still a gap in knowledge about the dynamics of enamel colonization by bacteria from the dental biofilm in caries. The aim, therefore, was to test whether the sequence of enamel colonization by a typically commensal and a cariogenic species modifies biofilm's cariogenicity. Dual-species biofilms of Streptococcus mutans and Streptococcus sanguinis on saliva-coated enamel slabs were inoculated in different sequences: S. mutans followed by S. sanguinis (Sm-Ss), S. sanguinis followed by S. mutans (Ss-Sm), S. mutans and S. sanguinis inoculated at the same time (Sm=Ss), and the single-species controls S. mutans followed by S. mutans (Sm-Sm) and S. sanguinis followed by S. sanguinis (Ss-Ss). Biofilms were exposed to 10% sucrose 3 times per day for 5 days, and the slabs/biofilms were retrieved to assess demineralization, viable cells, biomass, proteins, polysaccharides, and H2O2 production. Compared with Sm-Sm, primary inoculation with S. sanguinis reduced demineralization (P < 0.05). Both Ss-Sm and Sm=Ss sequences showed reduction in biomass, protein, and polysaccharide content (P < 0.05). The highest S. sanguinis viable count and H2O2 production level and the lowest acidogenicity were observed when S. sanguinis colonized enamel before S. mutans (P < 0.05). Initial enamel adherence with commensal biofilms seems to induce more intense competition against more typically cariogenic species, reducing cariogenicity.IMPORTANCE The concept of caries as an ecological disease implies the understanding of the intricate relationships among the populating microorganisms. Under frequent sugar exposure, some bacteria from the dental biofilm develop pathogenic traits that lead to imbalances (dysbiosis). Depending on which microorganism colonizes the dental surface first, different competition strategies may be developed. Studying the interactions in the entire dental biofilm is not an easy task. In this study, therefore, we modeled the interplay among these microorganisms using a caries-inducing species (S. mutans) and a health-associated species (S. sanguinis). Initial enamel adherence with S. sanguinis seems to induce more intense competition against typically caries-inducing species. Besides continuous exposure with sugars, early colonization of the enamel by highly cariogenic species like S. mutans appears to be needed to develop caries lesions as well. Promoting early colonization by health-associated bacteria such as S. sanguinis could help to maintain oral health, delaying dysbiosis.

Streptococcus mutans and Streptococcus sanguinis Expression of Competition-Related Genes, Under Sucrose.

Streptococcus mutans synthesizes 3 glucosyltransferases (Gtfs) associated with cariogenic biofilms, while commensal Streptococcus sanguinis produces only one; gtfP and hydrogen peroxide (H2O2) by SpxB. The aim was to test the hypothesis that under a sucrose-induced cariogenic challenge, the expression of competition-related genes is differentially regulated depending on whether S. sanguinis or S. mutans primarily colonize enamel. Dual-species biofilms of S. sanguinis and S. mutans were formed under different colonization sequences on enamel slabs and exposed to 10% sucrose for 5 min, 3×/day for 5 days. Biofilms were analyzed for the transcriptional response of competition-related genes encoding gtfB, gtfC, and gtfD for S. mutans and gtfP and spxB for S. sanguinis. In addition, acidogenicity (pH) and viable cells in each of the conditions were determined. For all the genes, a downregulation was observed during simultaneous colonization by both bacterial species. In contrast, gtfB was upregulated when S. sanguinis was the first colonizer (p < 0.05). Both gtfC and gtfD were upregulated during sequential inoculation with S. sanguinis as the first colonizer. An eleven-fold upregulation of gtfP was observed in biofilms with S. mutans as initial colonizer (p < 0.05), with a moderate increase in spxB expression. The lowest pH values and viable cells of S. sanguinis were observed when S. mutans first colonized the enamel slabs, compared to the other conditions (p < 0.05). Demanding sucrose-challenged oral environment requires increased expression of virulence traits to effectively compete and thrive in the dental biofilm, especially when the competitor has already colonized the ecological niche.

Cariogenicity induced by commercial carbonated beverages in an experimental biofilm-caries model.

OBJECTIVES: Frequent consumption of sugars-containing carbonated beverages has been associated with caries, but the consequences on the dental biofilm remain unclear. The aim was to evaluate the effect of commercial carbonated beverages and their sugar-free version on enamel and dentine demineralization and on the cariogenic properties of Streptococcus mutans biofilms. MATERIALS AND METHODS: Biofilms of S. mutans UA159 were grown on enamel and dentin slabs and exposed 3 times/day for 5 min, to a commercial cola or orange-flavored carbonated beverage or to their sugar-free version. Biofilms/slabs were recovered to assess biomass, viable microorganisms, protein content and polysaccharides. Demineralization was estimated by the variation of Knoop surface microhardness. RESULTS: Exposures to the biofilm with sugars-containing carbonated beverages resulted in similar biomass, viable microorganisms, proteins, and polysaccharides than sucrose (P < 0.05). The sugar-free cola and orange-flavored drink showed lower effect on the biofilm, as compared with sucrose or their sugared version (P < 0.05). All of the products tested, included the sugar-free, showed higher demineralization than the negative control (P < 0.05). CONCLUSIONS: Sugars-containing carbonated beverages enhance cariogenic activity of S. mutans biofilms, comparable with sucrose. Sugar-free carbonated beverages also have a high demineralizing potential, without affecting biofilm properties.

Creación de un Currículo de Competencias en Cariología para Estudiantes de Pregrado de Odontología en Chile / Development of a Competency-based Curriculum in Cariology for Undergraduate Dental Students in Chile

RESUMEN: Objetivo: Creación de un currículo de competencias mínimas en Cariología, para la formación de los Cirujano-Dentistas egresados de las escuelas de Odontología de Chile. Metodologías: A partir de una reunión de académicos de las Universidades de Talca y de Chile (año 2011), se elaboró una propuesta de currículo inicial, basado en los dominios propuestos por la Unión Europea (Schulte AG y cols). Durante el año 2016, dicha propuesta fue analizada mediante diálogos digitales y grupos de trabajo, con la participación del 96% de las Escuelas de Odontología existentes en el país, que concluyeron en un documento intermedio. Este documento fue analizado, discutido y perfeccionado durante el Taller para el Desarrollo de un Currículo de Competencias Mínimas en Cariología para las Escuelas de Odontología Chilenas (22/Mayo/2017, Talca, organizado por la Universidad de Talca y la Universidad de Chile) con la asistencia de representantes del 96% de las escuelas dentales chilenas, Ministerio de Salud de Chile, Colegio de Cirujano-Dentistas de Chile y con la asesoría de los profesores de Cariología Dres. Margherita Fontana y Carlos González-Cabezas (Universidad de Michigan, Ann Arbor, EEUU). Cada grupo de trabajo revisó el documento y envió nuevos comentarios, los que fueron incorporados en el documento final por una comisión asesora. Resultados: El documento del Currículo en Cariología se organizó en 5 Dominios: 1. Conocimientos base; 2. Determinación de Riesgo, diagnóstico de caries y detección de lesiones de caries; 3. Toma de decisiones y manejo preventivo no operatorio; 4. Toma de decisiones y manejo operatorio y 5. Cariología basada en la evidencia, en la práctica clínica y de salud pública. Se consensuaron las definiciones operacionales, las competencias principales y las sub-competencias para cada uno de los dominios. Las sub-competencias fueron clasificadas en tres niveles: A: Ser competente en; B: Tener conocimientos sobre y C: Estar familiarizado con. El documento final fue enviado a todos los participantes del taller para su aprobación y difusión en cada una de las instituciones involucradas. Conclusiones: Se logró, por medio de consenso, la construcción del Currículo de Competencias mínimas en Cariología para estudiantes de pregrado de Odontología en las universidades chilenas.

ABSTRACT: Objective: Development of a minimum set of competencies in Cariology that every dentist graduated from a Dental School in Chile must have. Methodology: Starting from a meeting of scholars from the Universities of Talca and Chile (year 2011), an initial proposal for a curriculum was developed, based on the domains proposed by the European Cariology Curriculum (Schulte, et al, 2011). During 2016, this proposal was discussed through online dialogues and working groups, with the participation of 95.2% of the Chilean dental schools, which resulted in an intermediate document. This document was analyzed, discussed and refined during the Workshop for the Development of a Curriculum of Minimum Competencies in Cariology for Chilean Dental Schools (May 22, 2017, Talca, organized by the Universities of Talca and Chile) with the attendance of representatives from 95.2% of the Chilean dental schools, the Chilean Ministry of Health, Chilean College od Dentists and with the assistance of the professors of Cariology Margherita Fontana and Carlos González-Cabezas (University of Michigan, Ann Arbor, USA). Each working group revised the document and provided feedback, which was incorporated in the final document by an advisory committee, elected on the day of the workshop, including the authors of the present article. Results: The Cariology Curriculum was organized in 5 Domains: 1. Basic knowledge; 2. Risk assessment, caries diagnosis and caries lesion detection; 3. Decision-making and non-operative preventive treatment; 4. Decision making and operative treatment; and 5. Evidence-based, clinical and public health practice. Operational definitions, main competencies and sub-competencies for each domain were agreed. Sub-competencies were classified into three levels: A: Be competent in; B: Have knowledge about, and C: Be familiar with. The final document was sent to all the participants of the workshop for dissemination in each of the institutions involved. Conclusions: The development of the Competency-based Curriculum in Cariology for undergraduate dental students at Chilean universities was achieved through consensus.

Frequency of sucrose exposure on the cariogenicity of a biofilm-caries model.

OBJECTIVE: Although sucrose is considered the most cariogenic carbohydrate in the human diet, the question of how many exposures are needed to induce damage on the hard dental tissues remains unclear. To approach this question, different frequencies of daily sucrose exposure were tested on a relevant biological caries model. MATERIALS AND METHODS: Biofilms of the Streptococcus mutans were formed on enamel slabs and exposed to cariogenic challenges with 10% sucrose for 5 min at 0, 1, 3, 5, 8, or 10 times per day. After 5 days, biofilms were retrieved to analyze biomass, protein content, viable bacteria, and polysaccharide formation. Enamel demineralization was evaluated by percentage of microhardness loss (percentage surface hardness loss [%SHL]). RESULTS: Biomass, protein content, polysaccharide production, acidogenicity of the biofilm, and %SHL proportionally increased with the number of daily exposures to sucrose (P < 0.05). One daily sucrose exposure was enough to induce 20% more demineralization than the negative unexposed control. Higher frequencies induced greater demineralization and more virulent biofilms, but eight and ten exposures were not different between them in most of the analyzed variables (P > 0.05). CONCLUSIONS: Higher sucrose exposure seems to increase cariogenicity, in a frequency-dependent manner, by the modification of bacterial virulent properties.