Bacterial factors of cariogenicity (literature review).

OBJECTIVE: Introduction: According to the 2003 WHO report, dental caries affects from 60% to 90% of school-age children and adults that makes it one of the most common diseases worldwide. The aim was to systematize data about the modern conception of plaque formation and role of microorganisms in its development. PATIENTS AND METHODS: Materials and methods: Biblosemantic, 50 medical literature sources were systematically reviewed as the material for the research. RESULTS: Review: According to Miller's theory, oral microorganisms can decompose dietary carbohydrates into acids, which in turn dissolve the calcium phosphates found in the enamel, causing it demineralisation Along with Streptococcus mutans, nowadays some other bacteria as Streptococcus sorbinus, Lactobacillus spp. and Actinomyces spp. have been well studied as caries contributors. However, the disease is related to plaque-mediated, because a much larger number of normal oral microflora representatives are involved in creating favourable preconditions for its development. There are a lot of original research papers about a role of bacteria in caries decay but compositions and characters of oral microflora are changing nowadays. Therefore, authors show the main cariogenic bacteria and their factors of pathogenicity which create special conditions for caries lesions. Modern concepts of dental plaque formation and pathogenesis of plaque-assosiative diseases are presented according to the new actual dental research. A lot of attention is paid to the biochemical properties of cariogenic bacteria and chemical process in biofilm. Role of acid and alkali production by oral bacteria in caries decay are shown in this article. Moreover, mechanisms of bacterial acid-fast and acid-tolerance are presented. CONCLUSION: Conclusions: Analysis of literature demonstrates a lot of bacterial pathogenicity factors which play key role in caries development.

A defined-multispecies microbial model for studying enamel caries development.

The aims of this study were to describe and validate an in vitro multispecies microbial biofilm model for caries development by evaluating the effects of varying medium concentration of sucrose (0.5 and 1.0%) and fluoride (0.4, 0.8 and 1.0 ppm F) in study 1, and calcium (1.0 and 2.0 mM Ca) in study 2. Defined-multispecies biofilms, formed by Lactobacillus casei, Streptococcus mutans, S. salivarius and S. sanguinis, were grown on the surface of salivary-pellicle-coated enamel slabs, with known baseline surface hardness; growth medium was changed daily. Counts of viable cells on biofilms and the percentage of surface microhardness change (%SMC), lesion depth (LD) and integrated mineral loss (IML) on enamel slabs were assessed after 4 days of biofilm formation under the tested conditions. Counts of viable cells on biofilms were significantly affected by sucrose, fluoride and calcium concentrations (p < 0.05). There was a decrease in %SMC in response to increased fluoride and calcium concentrations (p < 0.001). Lower IML (p < 0.001) and LD (p < 0.05) were found in the presence of 0.8 and 1.0 ppm F. A negative correlation was found between the response variables (%SMC, LD and IML) and fluoride and calcium concentrations. The results suggest that the microbial caries model developed was able to show distinct levels of caries inhibition in response to fluoride and calcium concentrations, corroborating clinical observations. An effect of sucrose concentration on caries development was found only in the presence of the lowest fluoride concentration.

pH of tooth surface in healthy adolescents at rest and after a glucose rinse: effect of 72 hours of plaque accumulation.

AIM: This study was performed to evaluate the effect of a glucose rinse and of plaque accumulation on pH of tooth surface in healthy adolescents with a device used in gastroenterology and never tested in the oral cavity. MATERIALS AND METHODS: Values of pH were monitored in 12 adolescents using a portable device (pH-day 2® Menfis, bioMedica S.r.l., Bologna, Italy) with a disposable antimonium electrode kept in contact with the interproximal surface of the upper molars for 40 minutes respectively before and after a one-minute rinse with 10 ml of a 10% glucose solution. The same procedure was repeated in the same subjects after 72 hours of plaque accumulation. RESULTS: The device tested resulted difficult to use on the tooth surface because of the size of the active part of the probe. The glucose rinse caused a statistically significant decrease of the mean pH, restrained in basal conditions (d = -0.16, p <0.05), clinically relevant after plaque accumulation (d = -1.24, p <0.05). Time in minutes of pH < 6 grew considerably only in case of combination of plaque accumulation and glucose rinse (d = 20.90, p <0.05). A Stephan's curve of drop and recover of the pH values was not recorded. CONCLUSIONS: The continuous recording of pH of natural plaque present on the tooth could represent an alternative to other techniques found in literature. More studies are necessary to verify the suitability of this new device for the monitoring of pH in the oral cavity.

Relationship between gap size and dentine secondary caries formation assessed in a microcosm biofilm model.

Secondary caries can develop at the tooth-restoration interface, depending on the presence of a gap and its size, but this process could be inhibited by fluoride. The aim of this study was to assess the relationship between gap size and dentine secondary caries adjacent to composite resin (CR) or glass ionomer (GI) restorations, using a microcosm biofilm model in a constant depth film fermentor (CDFF). Dentine discs restored with CR (Z250) or GI (Vitremer) with gap sizes of 0, 50, 100, 180 or 250 microm were mounted on the CDFF. Microcosm biofilms were formed on the restored discs and daily subjected to 8 pulses of 10% sucrose solution. On the 18th day, dentine mineral loss and lesion depth around the restorations were determined by transverse microradiography. The effect of gap size was overall not statistically significant either with regard to mineral loss (p = 0.449) or lesion depth (p = 0.328), but greater mineral loss and lesion depth were found adjacent to CR than to GI (p < 0.001). However, Spearman correlation showed that mineral loss and lesion depth increased with gap size for CR (p < 0.001) but not for GI (p > 0.05). The findings support the conclusion that fluoride released from GI inhibits dentine demineralization adjacent to restorations, irrespective of gap width.

Chlorhexidine efficacy in preventing lesion formation in enamel and dentine: an in situ study.

BACKGROUND: Clinical studies on the caries-preventive properties of chlorhexidine mouthrinses are limited and the results are inconclusive. AIM: The aim of this study was to elucidate the contribution of a 0.2% chlorhexidine mouthrinse to the protection of enamel and dentine against demineralization. METHODS: In this randomized two-treatment, two-leg study 14 individuals wearing partial prostheses were enrolled. Sound enamel and dentine specimens were placed in situ for 4 weeks. Twice daily, a mouthrinse was performed with either chlorhexidine or saline (control) depending on the experimental group the participant was allocated to. After the experimental period, plaque samples were collected from the surface of the specimens and from natural tooth surfaces to assess the organic acid production upon a sucrose challenge. The specimens were analyzed for mineral loss by transversal microradiography. RESULTS: This study could not demonstrate a significantly better protection of enamel and dentine against demineralization by the chlorhexidine treatment compared to saline. No differences in acid production of plaque samples from the chlorhexidine-treated and control groups were observed. This result was also found for plaque samples originating from the natural tooth surfaces. CONCLUSION: Mouth rinsing with 0.2% chlorhexidine did not prevent demineralization of dentine and enamel in situ.

Salivary flow patterns and the health of hard and soft oral tissues.

BACKGROUND: This nonsystematic review summarizes the effects of saliva on some of the diseases affecting the hard and soft oral tissues. RESULTS: Saliva enters the mouth at several locations, and the different secretions are not well-mixed. Saliva in the mouth forms a thin film, the velocity of which varies greatly at different sites. This variation appears to account for the site specificity of smooth-surface caries and supragingival calculus deposition. Saliva protects against dental caries, erosion, attrition, abrasion, candidiasis and the abrasive mucosal lesions seen commonly in patients with hyposalivation. These effects are the result of saliva's being a source of the acquired enamel pellicle; promoting the clearance of sugar and acid from the mouth; being supersaturated with respect to tooth mineral; containing buffers, urea for plaque base formation, and antibacterial and antifungal factors; and lubricating the oral mucosa, making it less susceptible to abrasive lesions. CLINICAL IMPLICATIONS: For optimal oral health, people should keep food and liquids in the mouth as briefly as possible. The most important time for toothbrushing is just before bedtime, because salivary flow is negligible during sleep and the protective effects of saliva are lost. Chewing sugar-free gum or sucking on sugar-free candies stimulates salivary flow, which benefits hard and soft oral tissues in many ways.

The role of sucrose in cariogenic dental biofilm formation--new insight.

Dental caries is a biofilm-dependent oral disease, and fermentable dietary carbohydrates are the key environmental factors involved in its initiation and development. However, among the carbohydrates, sucrose is considered the most cariogenic, because, in addition to being fermented by oral bacteria, it is a substrate for the synthesis of extracellular (EPS) and intracellular (IPS) polysaccharides. Therefore, while the low pH environment triggers the shift of the resident plaque microflora to a more cariogenic one, EPS promote changes in the composition of the biofilms' matrix. Furthermore, it has recently been shown that the biofilm formed in the presence of sucrose presents low concentrations of Ca, P(i), and F, which are critical ions involved in de- and remineralization of enamel and dentin in the oral environment. Thus, the aim of this review is to explore the broad role of sucrose in the cariogenicity of biofilms, and to present a new insight into its influence on the pathogenesis of dental caries.

Glucansucrases: mechanism of action and structure-function relationships.

Glucansucrases are produced principally by Leuconostoc mesenteroides and oral Streptococcus species, but also by the lactic acid bacteria (Lactococci, Lactobacilli). They catalyse the synthesis of high molecular weight D-glucose polymers, named glucans, from sucrose. In the presence of efficient acceptors, they catalyse the synthesis of low molecular weight oligosaccharides. Glucosidic bond synthesis occurs without the mediation of nucleotide activated sugars and cofactors are not necessary. Glucansucrases have an industrial value because of the production of dextrans and oligosaccharides and a biological importance by their key role in the cariogenic process. They were identified more than 50 years ago. The first glucansucrase encoding gene was cloned more than 10 years ago. But the mechanism of their action remains incompletely understood. However, in order to synthesise oligosaccharides of biological interest or to develop vaccines against dental caries, elucidation of the factors determining the regiospecificity and the regioselectivity of glucansucrases is necessary. The cloning of glucansucrase encoding genes in addition to structure-function relationship studies have allowed the identification of important amino acid residues and have shown that glucansucrases are composed of two functional domains: a core region (ca. 1000 amino acids) involved in sucrose binding and splitting and a C-terminal domain (ca. 500 amino acids) composed of a series of tandem repeats involved in glucan binding. Enzymology studies have enabled different models for their action mechanism to be proposed. The use of secondary structure prediction has led to a clearer knowledge of structure-function relationships of glucansucrases. However, mainly due to the large size of these enzymes, data on the three-dimensional structure of glucansucrases (given by crystallography and modelling) remain necessary to clearly identify those features which determine function.

pH modulation and salivary sugar clearance of different chocolates in children: A randomized clinical trial.

BACKGROUND: Sugars that occur naturally in foods and those added in processed foods may act as the source for fermentable carbohydrates and may initiate caries process. Among all the foods consumed by children, chocolates form an important constituent. A wide variety of chocolates are available in the Indian market and very few studies have compared their acidogenicity and salivary sugar clearance. OBJECTIVES: To compare the acidogenicity and salivary sugar clearance of 6 different commercially available chocolates in the Indian market. MATERIALS AND METHODS: Thirty subjects aged 10-15 years were selected randomly from one of the available public schools in Nellore city. Six commercially available chocolates in the Indian market were divided into three groups, unfilled (dark and milk chocolate), filled (wafer and fruit and nuts chocolate), and candy (hard milk and mango-flavored candy) groups. Plaque pH values and salivary sugar clearance rates are assessed at baseline, 5, 10, 15, 20, and 30 min after consumption. All the data obtained were statistically evaluated using independent sample t-test and one-way ANOVA for multiple group comparisons. RESULTS: Mango-flavored candy had maximum fall in plaque pH and least fall in plaque pH was recorded with milk chocolate. Fruit and nuts chocolate had a maximum clearance of salivary sugar and least fall in the salivary sugar clearance was recorded with dark chocolate. When the plaque pH and salivary sugar clearance of all the chocolates were assessed, it was seen that the values were statistically significant at all the time intervals (P < 0.05). CONCLUSION: Dark chocolate had a high fall in pH and milk chocolate had low salivary sugar clearance which signifies that unfilled chocolates are more cariogenic than other chocolates. Even though mango-flavored candy had maximum fall in plaque pH, its salivary sugar clearance was high.

The acidogenic potential of reference foods and snacks at interproximal sites in the human dentition.

Telemetry was used to evaluate changes in plaque pH at interproximal sites in the dentition of five human subjects following the ingestion of a variety of reference foods and snacks. Short-term telemetry (30 min) revealed that most of the substrates yielded both pH minima and total responses similar to that obtained with a 10% sucrose rinse. Aged Cheddar cheese and skim milk were much less acidogenic than were the other foods.

Comparison of methods for monitoring changes in the pH of human dental plaque.

Changes in human dental plaque pH can be used to obtain estimates of the acidogenic potential of ingested foods. The presence of acid in plaque is influenced by a large number of host, microbial, and substrate factors. Several useful methods have been developed for monitoring changes in plaque pH. Plaque sampling involves repeated removal of small samples of plaque from a number of teeth at intervals after food ingestion, dispersion of the sample, and in vitro measurement of pH. Touch electrode methods utilize glass or antimony microelectrodes, which are placed onto plaque in situ where direct readings can be obtained. Telemetry methods involve placement of glass microelectrodes or ion-sensitive field effect transistors within the dentition. Plaque is allowed to accumulate, and pH changes can subsequently be transmitted with radio or wire. Each of the methods has clear advantages and limitations. The methods have been simultaneously compared in human volunteers using solutions of fermentable carbohydrate. Inter-method differences in response were observed depending upon the site of measurement. Data obtained from caries-prone surfaces via telemetry showed lower pH minima and retarded returns to resting pH levels. The technology is available for controlled comparative plaque pH studies, with the method of choice depending upon the goals of the investigation. It is essential that the results be compared to data obtained with other models designed to evaluate the cariogenic potential of foods.

Remineralization of caries lesions extending into dentin.

Remineralization is one aspect of the overall process of tooth decay. However, it is primarily studied in shallow lesions. The aim of this study was to explore whether caries lesions in enamel and extending into the dentin can be remineralized. A single-section model was developed for the longitudinal and non-destructive monitoring of changes in enamel and dentin. Lesions at least 200 microm into dentin were formed in undersaturated acetate buffers. Next, the lesions were divided into groups (three treatment and one control) and remineralized. The treatments were: weekly immersion in 1,000 ppm fluoride, single treatment with methanehydroxybisphosphonate, and a constant level of 1 ppm fluoride. De- and remineralization was assessed by transverse microradiography. Remineralization was observed in enamel, but also in dentin, indicating that, deep into dentin, the pores become supersaturated to apatite formation. Treatments affected remineralization only in the outer part of enamel. Both findings are explained by a relatively fast diffusion of mineral ions, with precipitation being rate-limiting. The results suggest that dentin remineralization, underneath enamel, can be achieved and could possibly be used in clinical treatment strategies.

Factors affecting the resting pH of in vitro human microcosm dental plaque and Streptococcus mutans biofilms.

The aim was to examine factors that potentially control the resting pH, defined as the pH unaffected by meals, of microcosm dental plaques and Streptococcus mutans biofilms under standard conditions, and to examine the effect of supplying urea at concentrations found intraorally. Microcosm plaques were cultured from plaque bacteria-enriched saliva in an 'artificial mouth' with a continuous supply of a medium including 0.25% mucin [Basal Medium Mucin, (BMM), 3.6 ml/hr per plaque] and a periodic supply of sucrose. The steady-state resting pH was 6.4 (range +/- 0.1) in BMM containing no urea and supplied at the standard flowrate. This is a robust property of the ecosystem. In one experiment with a replicated (n = 9) set of measurements, the resting pH was approx. pH 6.3, 6.4, 6.7 and 7.3 with 0, 1, 5 and 20 mmol/l urea in the BMM. The magnitude of sucrose- and urea-induced pH responses was unaffected by elevating the resting pH to produce parallel pH curves. The sucrose-induced pH curves were analogous to those classically reported by Stephan that showed an association between caries activity and increasingly acidic plaque pH responses to glucose. Stopping the BMM flow caused a pH rise, indicating continuing net alkali generation from BMM components in the absence of a fluid flow. Step. mutans monoculture biofilms had an acidic resting pH of 5.0 to 5.3, which increased to 6.8 following an adventitious superinfection by Bacillus cereus. It was concluded that the resting pH in plaque results from a delicate balance between alkali and acid generation, which is in turn dependent both on the bacterial composition of the plaque and on the supply of substrates and buffers from, and metabolite clearance into, flowing oral fluid. In vivo the resting pH will vary with site-specific changing saliva flows. Urea continuously supplied at concentrations normal for saliva and gingival crevicular fluid can raise the resting pH of microcosm plaque by an amount tat in vivo would probably be significant in reducing dental caries.

Low-cariogenicity of trehalose as a substrate.

OBJECTIVES: The effects of trehalose on cariogenesis by mutans streptococci were investigated. METHODS: Inhibited effect of trehalose on water-insoluble glucan (WIG) synthesis from sucrose by glucosyltransferase (GTase) of mutans streptococci was assayed. The acid fermentability of trehalose by mutans streptococci was determined by the measurements of pH, and amounts of lactic acid production. Plaque pH was determined by the measurements of collected plaque from volunteers after sugar mouth-rinse. Rat experimental caries was investigated by feeding a sucrose and/or trehalose diet. RESULTS: Trehalose was not utilized as a substrate for GTase. In addition, trehalose inhibited synthesis of WIG by GTase in the presence of sucrose. Trehalose showed weaker and slower acid fermentation than sucrose by mutans streptococci. The levels of lactic acid production from trehalose by Streptococcus mutans and Streptococcus sobrinus were 24.2 and 59.8% of those from sucrose, respectively. The minimum plaque pH after sucrose mouth-rinse was lower than those after trehalose mouth-rinse in all subjects. Plaque pH after trehalose mouth-rinse never reached critical pH. The substitution of trehalose for sucrose in the rat diet significantly reduced caries scores. Furthermore, rats fed diets containing sucrose and trehalose had significantly lower caries scores than those fed a sucrose diet. CONCLUSIONS: These results suggested that trehalose might be not only lowly cariogenic but also anti-cariogenic, and is promising as a sugar substitute.

Estimation of the caries-related risk associated with infant formulas.

PURPOSE: Baby bottle tooth decay (BBTD) affects 6% of children under three years of age and is associated with inappropriate bottle use. The objective of this study was to estimate the caries-related risk associated with 26 infant formulas and whole milk. METHODS: First, the plaque pH of adult volunteers was monitored before and after an oral rinse with infant formula to determine the minimum pH obtained in response to each formula. Second, Streptococcus sobrinus 6715 was cultured in each infant formula, and the increase in the number of colony forming units was measured. Third, each infant formula was incubated with powdered enamel and the solubility of enamel mineral was calculated in the absence of bacteria. Fourth, each formula was mixed with standardized concentrations of acid to determine the buffering capabilities. Finally, enamel windows were created on extracted permanent molars and exfoliated primary incisor crowns that were then colonized with mutans streptococci and incubated with infant formula. Caries was assessed visually and radiographically for 18 weeks. The length of time required for the development of enamel caries, dentinal caries and pulpal involvement was recorded. RESULTS: One-way or two-way ANOVA of these five assays demonstrated that 1. Plaque pH varied in response to oral rinsing with infant formula and most formulas did have the ability to reduce the pH significantly below the pH obtained after rinsing with water 2. Some infant formulas supported significant bacterial growth 3. Enamel mineral was dissolved by incubation with certain infant formula 4. The buffer capacity varied among the infant formulas tested 5. The length of time required for caries to reach dentin or pulp differed for the formulas, with some formulas causing dentinal caries by 3.4 weeks and pulpal involvement by 7.2 weeks.

Metabolic activity of Streptococcus mutans biofilms and gene expression during exposure to xylitol and sucrose.

The objective of the study was to analyse Streptococcus mutans biofilms grown under different dietary conditions by using multifaceted methodological approaches to gain deeper insight into the cariogenic impact of carbohydrates. S. mutans biofilms were generated during a period of 24 h in the following media: Schaedler broth as a control medium containing endogenous glucose, Schaedler broth with an additional 5% sucrose, and Schaedler broth supplemented with 1% xylitol. The confocal laser scanning microscopy (CLSM)-based analyses of the microbial vitality, respiratory activity (5-cyano-2,3-ditolyl tetrazolium chloride, CTC) and production of extracellular polysaccharides (EPS) were performed separately in the inner, middle and outer biofilm layers. In addition to the microbiological sample testing, the glucose/sucrose consumption of the biofilm bacteria was quantified, and the expression of glucosyltransferases and other biofilm-associated genes was investigated. Xylitol exposure did not inhibit the viability of S. mutans biofilms, as monitored by the following experimental parameters: culture growth, vitality, CTC activity and EPS production. However, xylitol exposure caused a difference in gene expression compared to the control. GtfC was upregulated only in the presence of xylitol. Under xylitol exposure, gtfB was upregulated by a factor of 6, while under sucrose exposure, it was upregulated by a factor of three. Compared with glucose and xylitol, sucrose increased cell vitality in all biofilm layers. In all nutrient media, the intrinsic glucose was almost completely consumed by the cells of the S. mutans biofilm within 24 h. After 24 h of biofilm formation, the multiparametric measurements showed that xylitol in the presence of glucose caused predominantly genotypic differences but did not induce metabolic differences compared to the control. Thus, the availability of dietary carbohydrates in either a pure or combined form seems to affect the cariogenic potential of S. mutans biofilms.

Fermentative 2-carbon metabolism produces carcinogenic levels of acetaldehyde in Candida albicans.

UNLABELLED: Acetaldehyde is a carcinogenic product of alcohol fermentation and metabolism in microbes associated with cancers of the upper digestive tract. In yeast acetaldehyde is a by-product of the pyruvate bypass that converts pyruvate into acetyl-Coenzyme A (CoA) during fermentation. THE AIMS OF OUR STUDY WERE: (i) to determine the levels of acetaldehyde produced by Candida albicans in the presence of glucose in low oxygen tension in vitro; (ii) to analyse the expression levels of genes involved in the pyruvate-bypass and acetaldehyde production; and (iii) to analyse whether any correlations exist between acetaldehyde levels, alcohol dehydrogenase enzyme activity or expression of the genes involved in the pyruvate-bypass. Candida albicans strains were isolated from patients with oral squamous cell carcinoma (n = 5), autoimmune polyendocrinopathy-candidiasis-ectodermal dystrophy (APECED) patients with chronic oral candidosis (n = 5), and control patients (n = 5). The acetaldehyde and ethanol production by these isolates grown under low oxygen tension in the presence of glucose was determined, and the expression of alcohol dehydrogenase (ADH1 and ADH2), pyruvate decarboxylase (PDC11), aldehyde dehydrogenase (ALD6) and acetyl-CoA synthetase (ACS1 and ACS2) and Adh enzyme activity were analysed. The C. albicans isolates produced high levels of acetaldehyde from glucose under low oxygen tension. The acetaldehyde levels did not correlate with the expression of ADH1, ADH2 or PDC11 but correlated with the expression of down-stream genes ALD6 and ACS1. Significant differences in the gene expressions were measured between strains isolated from different patient groups. Under low oxygen tension ALD6 and ACS1, instead of ADH1 or ADH2, appear the most reliable indicators of candidal acetaldehyde production from glucose.

Structural investigation of an extracellular polysaccharide produced by the cariogenic bacterium Streptococcus mutans strain UA159.

The structure of an extracellular polysaccharide EPS159 produced from sucrose by Streptococcus mutans UA159 was investigated through the main oligosaccharides obtained from partial acid hydrolysis, monosaccharide/methylation analysis, and 1D/2D (1)H NMR spectroscopy. The results showed that EPS159 contained terminal, 3-substituted, 6-substituted, and 3,6-disubstituted α-D-glucopyranose residues in a molar percentage of 14, 18, 54, and 14%. The backbone of EPS159 was composed of →6)Glcp(1→ residues, and about 20% of the →6)Glcp(1→ residues was substituted at 3-OH by →3)Glcp(1→ and/or Glcp(1→ residues to form side chains. A composite model of EPS159, that includes all identified structural features, was formulated: [Formula, see text:].