The Effects of Nonnutritive Sweeteners on the Cariogenic Potential of Oral Microbiome.

Nonnutritive sweeteners (NNSs) are sugar substitutes widely used to reduce the negative health effects of excessive sugar consumption. Dental caries, one of the most prevalent chronic diseases globally, results from a pathogenic biofilm with microecological imbalance and frequent exposure to sugars. Some research has shown that certain NNSs possess less cariogenic potential than sucrose, indicating their putative effect on oral microbiome. To uncover the alterations of acidogenic pathogens and alkali-generating commensals, as well as the biofilm cariogenic potential under the influence of NNSs, we selected four common NNSs (acesulfame-K, aspartame, saccharin, and sucralose) and established single-, dual-, and multispecies in vitro culture model to assess their effects on Streptococcus mutans (S. mutans) and/or Streptococcus sanguinis (S. sanguinis) compared to sucrose with the same sweetness. The results showed that NNSs significantly suppressed the planktonic growth, acid production, and biofilm formation of S. mutans or S. sanguinis compared with sucrose in single-species cultures. Additionally, decreased S. mutans/S. sanguinis ratio, less EPS generation, and higher pH value were observed in dual-species and saliva-derived multispecies biofilms with supplementary NNSs. Collectively, this study demonstrates that NNSs inhibit the cariogenic potential of biofilms by maintaining microbial equilibrium, thus having a promising prospect as anticaries agents.

Short-Chain N-Acylhomoserine Lactone Quorum-Sensing Molecules Promote Periodontal Pathogens in In Vitro Oral Biofilms.

Acylhomoserine lactones (AHLs), the quorum-sensing (QS) signals produced by a range of Gram-negative bacteria, are involved in biofilm formation in many pathogenic and environmental bacteria. Nevertheless, the current paradigm excludes a role of AHLs in dental plaque formation, while other QS signals, such as AI-2 and autoinducer peptides, have been demonstrated to play an important role in biofilm formation and virulence-related gene expression in oral pathogens. In the present work, we have explored the effect of externally added AHLs on in vitro oral biofilm models for commensal, cariogenic, and periodontal dental plaque. While little effect on bacterial growth was observed, some AHLs specifically affected the lactic acid production and protease activity of the biofilms. Most importantly, the analysis of bacterial diversity in the biofilms showed that the addition of C6-homoserine lactone (C6-HSL) results in a shift toward a periodontal bacterial composition profile by increasing the relative presence of the orange-complex bacteria Peptostreptococcus and Prevotella These results point to a relevant role of AHL-mediated QS in dental plaque formation and might be involved in the development of dysbiosis, the mechanism of which should be further investigated. This finding potentially opens new opportunities for the prevention or treatment of the periodontal disease.IMPORTANCE Dental plaque is omnipresent in healthy oral cavities and part of our commensal microbial colonization. At the same time, dental plaque is the cause of the most common human diseases, caries and gum disease. Dental plaque consists of billions of microbes attached to the surface of your teeth. Communication among these microbes is pivotal for development of these complex communities yet poorly studied in dental plaque. In the present study, we show that a specific communication molecule induces changes within the community related to the development of gum disease. This finding suggests that interfering with microbial communication may represent an interesting novel strategy to prevent gum disease that should be further investigated.

Effects of Norspermidine on Dual-Species Biofilms Composed of Streptococcus mutans and Streptococcus sanguinis.

The present study aimed at investigating the influence of norspermidine on the formation of dual-species biofilms composed of Streptococcus mutans (S. mutans) and Streptococcus sanguinis (S. sanguinis). Crystal violet assay was conducted to assess the formation of single-species biofilms of S. mutans and S. sanguinis, and the growth curve was carefully observed to monitor the growth of these two species of bacteria. Fluorescence in situ hybridization (FISH) and MTT array were used to analyze the composition and metabolic activity of the dual-species biofilms, respectively. Extracellular polysaccharides (EPS)/bacteria staining, anthrone method, and scanning electron microscopy (SEM) imaging were conducted to study the synthesis of EPS by dual-species biofilms. Lactic acid assay and pH were measured to detect dual-species biofilm acid production. We found that norspermidine had different effects on S. mutans and S. sanguinis including their growth and biofilm formation. Norspermidine regulated the composition of the dual-species biofilms, decreased the ratio of S. mutans in dual-species biofilms, and reduced the metabolic activity, EPS synthesis, and acid production of dual-species biofilms. Norspermidine regulated dual-species biofilms in an ecological way, suggesting that it may be a potent reagent for controlling dental biofilms and managing dental caries.

Effect of Sweeteners on Root Dentine Demineralization Using a Microcosm Biofilm Model / Efecto de los Edulcorantes en la Desmineralización de la Dentina Radicular Utilizando un Modelo de Biofilm Microcosmo

ABSTRACT: The aim of the present study was to evaluate the effect of commercial sweeteners on root dentin demineralization using a microcosm biofilm model. Bovine dentin specimens with pre-determined surface hardness were randomized into six groups according to the studied sweeteners: sucralose, stevia, saccharin, aspartame. Sucrose was used as a positive control and an untreated group as a negative control. The specimens were submitted to biofilm development from one saliva donor and the cariogenic challenge occurred on subsequent five days, twice a day. At the end, the percentage of surface hardness loss (%SHL) and biomass was determined and submitted to ANOVA followed by Tukey's test. Sucrose presented the highest rate of demineralization, however, all sweeteners tested lead to a statistically higher root demineralization compared to the negative control (p <0.05). Sucrose caused greater demineralization in root dentin, however, the sweeteners were also able to induce it under this biofilm model.

RESUMEN: El objetivo del presente estudio fue evaluar el efecto de los edulcorantes comerciales en la desmineralización de la dentina radicular utilizando un modelo de biofilm microcosmo. Se asignaron al azar muestras de dentina bovina con una dureza de la superficie predeterminada de acuerdo con los edulcorantes estudiados: sucralosa, estevia, sacarina, aspartame. La sacarosa se utilizó como control positivo y un grupo no tratado como control negativo. Las muestras se enviaron al desarrollo de biopelículas de un donante de saliva y el desafío cariogénico se produjo en los siguientes cinco días, dos veces al día. Al final, se determinó el porcentaje de pérdida de dureza de la superficie (% PDS) y biomasa y se aplicó un estudio estadístico de ANOVA seguido de la prueba de Tukey. La sacarosa presentó la mayor tasa de desmineralización; sin embargo, todos los endulzantes probados condujeron a una desmineralización de la raíz estadísticamente mayor en comparación con el control negativo (p<0,05). La sacarosa causó una mayor desmineralización en la dentina de raíz, sin embargo, los edulcorantes también fueron capaces de inducirla bajo este modelo de biofilm.

Effects of Antimicrobial Peptide GH12 on the Cariogenic Properties and Composition of a Cariogenic Multispecies Biofilm.

Dental caries is a biofilm-mediated disease that occurs when acidogenic/aciduric bacteria obtain an ecological advantage over commensal species. In previous studies, the effects of the antimicrobial peptide GH12 on planktonic bacteria and monospecies biofilms were confirmed. The objectives of this study were to investigate the effects of GH12 on a cariogenic multispecies biofilm and to preliminarily explain the mechanism. In this biofilm model, Streptococcus mutans ATCC 70061 was the representative of cariogenic bacteria, while Streptococcus gordonii ATCC 35105 and Streptococcus sanguinis JCM 5708 were selected as healthy microbiota. The results showed that GH12 was more effective in suppressing S. mutans than the other two species, with lower MIC and minimal bactericidal concentration (MBC) values among diverse type strains and clinical isolated strains. Therefore, GH12, at no more than 8 mg/liter, was used to selectively suppress S. mutans in the multispecies biofilm. GH12 at 4 mg/liter and 8 mg/liter reduced the cariogenic properties of the multispecies biofilm in biofilm formation, glucan synthesis, and lactic acid production. In addition, GH12 suppressed S. mutans within the multispecies biofilm and changed the bacterial composition. Furthermore, 8 mg/liter GH12 showed a selective bactericidal impact on S. mutans, and GH12 promoted hydrogen peroxide production in S. sanguinis and S. gordonii, which improved their ecological advantages. In conclusion, GH12 inhibited the cariogenic properties and changed the composition of the multispecies biofilm through a two-part mechanism by which GH12 directly suppressed the growth of S. mutans as well as enhanced the ecological competitiveness of S. sanguinis and S. gordoniiIMPORTANCE Dental caries is one of the most prevalent chronic infectious diseases worldwide, with substantial economic and quality-of-life impacts. Streptococcus mutans has been considered the principal pathogen of dental caries. To combat dental caries, an antimicrobial peptide, GH12, was designed, and its antibacterial effects on planktonic S. mutans and the monospecies biofilm were confirmed. As etiological concepts of dental caries evolved to include microecosystems, the homeostasis between pathogenic and commensal bacteria and a selective action on cariogenic virulence have increasingly become the focus. The novelty of this research was to study the effects of the antimicrobial peptides on a controlled cariogenic multispecies biofilm model. Notably, the role of an antimicrobial agent in regulating interspecific competition and composition shifts within this multispecies biofilm was investigated. With promising antibacterial and antibiofilm properties, the use of GH12 might be of importance in preventing and controlling caries and other dental infections.

In situ Effect of Arginine-Containing Dentifrice on Plaque Composition and on Enamel Demineralization under Distinct Cariogenic Conditions.

There is limited evidence that arginine-containing fluoridated dentifrices (AFD) have a better anticaries effect than regular fluoridated dentifrices (FD), especially in subjects at a higher risk for caries development. This study aimed to assess the effect of AFD on enamel demineralization and on the microbial and biochemical compositions of biofilm formed under different frequencies of sucrose exposure. It consisted of an in situ split-mouth design, where 12 adult volunteers who used FD for at least 2 months prior to the beginning of this study wore acrylic palatal appliances containing 4 bovine enamel specimens (1 pair at each side of the appliance) during 2 phases of 14 days each. FD slurry (3×/day) and 20% sucrose solution (4× and 8×/day) were dripped on the specimens during the first experimental phase. The same volunteers then used AFD during a 2-month washout period, followed by a second experimental phase where the AFD slurry and sucrose solution were applied onto a new subset of specimens. The percentage of enamel surface hardness loss (%SHL), the lesion depth (LD), the integrated mineral loss (IML), microbial counts on biofilms, the biomass, and inorganic and insoluble extracellular polysaccharide (IEPS) biofilm concentrations were determined. Higher %SHL, biomass, and IEPS and lower fluoride values were found at sucrose 8×/day exposure. Lower IEPS were found in the presence of AFD compared to FD. Similar %SHL, LD, and IML values were found between FD and AFD, irrespectively of the cariogenic challenge. The results suggest that AFD have an anticaries effect similar to that of regular FD.

Sucrose induced dentin demineralization in a microcosm biofilm model / Desmineralización de la dentina inducida por sacarosa en un modelo de biofilm microcosmo

The aim of this in vitro study was to assess the root dentin demineralization caused by a microcosm biofilm model that has been exposed to sucrose in different ways. Materials and Methods: Saliva of two volunteers was inoculated into an artificial medium for biofilm growth and dentin blocks were immersed into these media. Dentin specimens were randomly exposed to one of the five experimental conditions: C (control group - no saliva inoculum or sucrose), 0S (saliva inoculum without sucrose, negative control), 3S (three daily one-minute immersions in 20 % sucrose), 6S (six daily one-minute immersions in 20 % sucrose), and CS (continuously immersed in 5 % sucrose). After five days, biofilm was collected to determine the concentration of intracellular and extracellular polysaccharides and the dentin surface hardness loss (SHL) was measured. The experiment was carried out in triplicate. Results: The dentin SHL was higher in groups that were exposed to sucrose (3S, 6S and CS) and there was a statistically significant difference between all groups (p<0.001). CS had higher concentrations of polysaccharides (p>0.001) and there was no statistically significant difference between the other groups (0S, 3S and 6S) (p>0.005). Conclusion: The microcosm biofilm model developed has the potential to produce root dentin demineralization at different exposures to sucrose.

El objetivo de esta investigación in vitro fue evaluar la desmineralización de la dentina radicular causada por un modelo de biofilm microcosmo que fue expuesto de diferentes maneras a la sacarosa. La saliva de dos voluntarios fue colocada en un medio artificial para crecimiento del biofilm y los bloques de dentina fueron sumergidos en estos medios. Al aza rlos bloques fueron expuestos a una de las cinco condiciones experimentales: C (grupo control ­ sin inoculación de saliva o sacarosa), 0S (inoculación de saliva sin sacarosa, control negativo), 3S (tres inmersiones diarias de un minuto en sacarosa a 20 %), 6S (seis inmersiones diarias de un minuto en sacarosa a 20 %), y CS (sumergidos continuamente en 5 % de sacarosa). Después de cinco días, el biofilm fue recogido para determinar la concentración de polisacáridos intracelulares y extracelulares y fue medida la pérdida de dureza superficial de la dentina (SHL). El experimento se repitió en tres ocasiones. La dentina SHL era mayor en los grupos que fueron expuestos a la sacarosa (3S, 6S E CS) y hubo diferencia estadísticamente significativa entre todos los grupos (P<0,001). CS presentó mayor concentración de polisacáridos (p<0,001) y no hubo diferencia estadísticamente significativa entre los demás grupos (0S, 3S E 6S) (p>0,005). El modelo del biofilm desarrollado tiene potencial para producir desmineralización de la dentina radicular en diferentes exposiciones a la sacarosa.

Effect of the association of maltodextrin and sucrose on the acidogenicity and adherence of cariogenic bacteria.

OBJECTIVE: The aim was to investigate the effect of maltodextrin and sucrose association on the acidogenic and adherence profiles of cariogenic bacteria. DESIGN: Streptococcus mutans (S. mutans) and Lactobacillus casei (L. casei) were cultivated in culture medium containing maltodextrin, sucrose, maltodextrin-sucrose mixture or glucose. Analyses of the acidogenicity and microbial adherence were conducted in triplicate for each microorganism and tested carbohydrate. RESULTS: For L. casei, maltodextrin, sucrose and maltodextrin-sucrose mixture showed lower acidogenic potential compared to glucose. When the microorganism was S. mutans, sucrose and maltodextrin-sucrose mixture presented higher acidogenic potential compared to maltodextrin and glucose. Microbial adherence analysis revealed higher adherence for S. mutans in presence of sucrose and maltodextrin-sucrose mixture compared to maltodextrin and glucose. For L. casei, all the carbohydrates showed similar adherence percentages. CONCLUSION: The addition of maltodextrin to sucrose does not increase the cariogenicity of sucrose in terms of acidogenicity and adherence of the cariogenic bacteria.

In situ bacterial accumulation on dental restorative materials. CLSM/COMSTAT analysis.

PURPOSE: To evaluate the influence of different restorative materials on the biofilm structure accumulated in situ. METHODS: 15 discs of each material (ceramic; resin composite; resin-modified and conventional glass-ionomers; amalgam) were adapted to palatal devices in order to accumulate biofilm in situ, under a cariogenic challenge (20% sucrose solution, 10x/day). After 7 days, the specimens were carefully removed and visualized by confocal laser scanning microscopy (CLSM). The images were analyzed qualitatively (descriptive analysis about cell viability and architecture) and quantitatively using COMSTAT software (area, bio-volume, mean thickness, maximum thickness and roughness coefficient of the biofilm). The statistical analysis was performed by using the Kolmogorov-Smirnov and Kruskal-Wallis tests (P ≤ 5%). RESULTS: The medians of the biofilm parameters analyzed showed no statistical difference regarding different materials. However, qualitatively, glass-ionomer cements and amalgam showed visually a prevalence of non-viable cells forming small clusters distributed by the biofilm, and voids were presented in smaller proportion in the biofilm volume compared to composite and ceramic.

An in vitro study to determine the effect of Terminalia chebula extract and its formulation on Streptococcus mutans.

AIM: Many weapons are available in the arsenal of a dental professional to combat dental caries, which is almost ubiquitously present. From a public health perspective, most of these weapons are far from being an ideal drug. Hence, there is a demand for better and effective antibacterial agents. This factor stimulated the process of the present study. The aim of the study was to determine the effect of ethanol extract of Terminalia chebula on Streptococcus mutans. MATERIALS AND METHODS: Dried ripe fruits of Terminalia chebula were procured and powdered. Physical tests were done to estimate purity of the fruit powder. Hydroethanolic and aqueous extracts were prepared according to standard procedures. Minimum inhibitory concentration of the extracts was determined by tube dilution method and confirmed by agar dilution method. The effect of the hydroethanolic extract on sucrose induced adhesion, glucan-induced aggregation and on glycolysis of Streptococcus mutans was also assessed. Preservative, gelling agent and sweetener were added in suitable quantities to the ethanol extract, and mouthrinse was formulated. Minimum inhibitory concentration of the formulation was also determined. RESULTS: Yield was better in case of aqueous extract. The Minimum inhibitory concentration of hydroethanolic extract was determined to be 2.5%. Minimum inhibitory concentration of the aqueous extract was determined to be 10%. Hydroethanolic extract of Terminalia chebula (2.5%) inhibited sucrose induced adherence and aggregation of Streptococcus mutans in vitro. CONCLUSION: The mouthrinse formulated from ethanol extract of Terminalia chebula demonstrated substantial antibacterial activity and could be used as an effective anticaries agent. CLINICAL SIGNIFICANCE: Terminalia chebula mouthrinse can be effectively used in clinical practice as an anticaries mouthrinse with additional benefit being that it is safe and economical.

Effects of heat-inactivated Bifidobacterium BB12 on cariogenicity of Streptococcus mutans in vitro.

OBJECTIVE: Since some probiotic bacteria are cariogenic themselves, their suitability for caries management is questionable. Inactivated bacteria or their supernatants have been found to exert probiotic effects, whilst having several advantages compared with living bacteria. We hypothesized that viable and heat-inactivated Bifidobacterium animalis BB12 reduces the cariogenicity of Streptococcus mutans (SM) in vitro. DESIGN: We assessed mono- and mixed species biofilms of SM and viable or heat-inactivated BB12. Biofilms were grown in a continuous-culture-system under cariogenic conditions on smooth proximal enamel or cavitated dentine. For each of eight experimental subsets (4 biofilms×2 hard-tissue conditions), a total of 32 specimens was used. After 10 days, bacterial numbers of 12 biofilms per group were analysed, and all specimens submitted to transversal microradiography. RESULTS: Mineral loss was higher in cavitated dentine than smooth enamel for all biofilms (p<0.001, t-test). BB12-monospecies biofilms induced significantly less mineral loss than SM in both enamel (p<0.05) and dentine (p<0.001). Viable BB12 did not significantly reduce cariogenicity of SM (p>0.05), whilst heat-inactivated BB12 decreased cariogenicity of SM in dentinal cavities (p<0.01). Bacterial numbers were higher on dentine than enamel (p<0.05), but not significantly influenced by biofilm species (p>0.05). CONCLUSIONS: Heat-inactivated BB12 reduced the cariogenicity of SM in dentinal cavities in vitro. Inactivated probiotics might be suitable for caries control.

Effects of lactose-containing stevioside sweeteners on dental biofilm acidogenicity.

The aim of this study was to evaluate the effect of a commercial lactose-containing stevioside sweetener on biofilm acidogenicity in vivo. Nine volunteers refrained from brushing their teeth for 3 days in five phases. On the 4th day of each phase, the pH of the biofilm was measured by the "Strip method". Interproximal plaque pH was measured before and up to 60 minutes after a 10 mL mouthrinse for 1 minute with the test solutions: I - sweetener with 93% lactose and 7% stevioside; II - sweetener with 6.8% saccharin, 13.6% cyclamate, and 0.82% stevioside; III - 18% sucrose solution (positive control); IV - mineral water (negative control); and V- 93% lactose solution. The results revealed that the most pronounced pH fall was found with sucrose (positive control), followed by the 93% lactose solution, the sweetener with lactose + stevioside, the sweetener with saccharin + cyclamate + stevioside, and finally water (negative control). According to the area under the curve, the two sweeteners containing stevioside were significantly different, and the sweetener with lactose + stevioside was significantly different from water but not from sucrose. The critical pH for dentin demineralization (pH ≤ 6.5) was reached by all volunteers after rinsing with sucrose solution, lactose solution, and the stevioside + lactose sweetener. Analysis of the data suggests that lactose-containing stevioside sweeteners may be cariogenic, especially to dentin.

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.

A 12-week clinical study assessing the clinical effects on plaque metabolism of a dentifrice containing 1.5% arginine, an insoluble calcium compound and 1,450 ppm fluoride.

PURPOSE: To evaluate the clinical effect on plaque metabolism of a dentifrice containing 1.5% arginine, an insoluble calcium compound and 1,450 ppm fluoride compared to a commercially available dentifrice containing 1,450 ppm fluoride in a silica base. METHODS: A 12-week, parallel, randomized, double-blind study using 48 subjects was conducted at the Colgate-Palmolive Technology Center (Piscataway, NJ, USA). One group used a test dentifrice containing 1.5% arginine, an insoluble calcium compound and 1,450 ppm fluoride as sodium monofluorophosphate (MFP), and the other group used a commercial silica dentifrice with 1,450 ppm fluoride as sodium fluoride (NaF) as a control. Plaque metabolism analyses were conducted at baseline and after 2, 4, 6, 8, and 12 weeks of assigned product use. The plaque analyses included pH measurements before and after a sucrose rinse, ammonia production and lactic acid production. RESULTS: Subjects using the test dentifrice had significantly higher plaque pH values before (P< or = 0.01) and after (P< or = 0.045) a sucrose challenge than those using the commercially available control dentifrice. Subjects using the test dentifrice also produced higher levels of ammonia and lower levels of lactic acid compared to subjects using the control dentifrice.

Effect of high-fructose corn syrup on Streptococcus mutans virulence gene expression and on tooth demineralization.

High-fructose corn syrup-55 (HFCS-55) has been widely welcomed in recent years as a substitute for sucrose on the basis of its favourable properties and price. The objective of this study was to determine the influence of HFCS-55 on the expression of Streptococcus mutans UA159 virulence genes and on tooth demineralization. Real-time reverse-transcription PCR (real-time RT-PCR) and microhardness evaluations were performed to examine gene expression and enamel demineralization, respectively, after treatment with HFCS-55 and/or sucrose. Significant up-regulation of glucosyltransferase B (gtfB) by HFCS-55 was found. A mixture of HFCS-55 and sucrose could positively enhance expression of glucan-binding protein (gbp) genes. Regarding acidogenicity, expression of the lactate dehydrogenase (ldh) gene was unaffected by HFCS-55. A notable finding in this study was that 5% HFCS-55 significantly enhanced expression of the intracellular response gene of the two-component VicRK signal transduction system (vicR). Demineralization testing showed that the microhardness of teeth decreased by a greater extent in response to HFCS-55 than in response to sucrose. The results indicate that HFCS-55 can enhance S. mutans biofilm formation indirectly in the presence of sucrose and that HFCS-55 has a more acidogenic potential than does sucrose. Summing up the real-time PCR and demineralization results, HFCS-55 appears to be no less cariogenic than sucrose in vitro - at least, not under the conditions of our experiments.

Cariogenicity of different commercially available bovine milk types in a biofilm caries model.

PURPOSE: This study's purpose was to assess the cariogenicity of commercial bovine milk types in an experimental biofilm/caries model. METHODS: Enamel and dentin slabs were used to grow biofilms of Streptococcus mutans UA159. Slabs/biofilms were exposed three times per day to commercial skim, semi-skim, whole, whole lactose-free, and whole with 10 percent sucrose-added bovine milk and to 10 percent sucrose and 0.9 percent sodium chloride as positive and negative caries-control, respectively. Biofilms were analyzed for bacterial counts, biomass, proteins, and polysaccharide production. Slab's demineralization was assessed by loss of surface microhardness and the biofilm acidogenicity by medium pH. RESULTS: Only whole and whole lactose-free milk kept pH above the demineralization threshold, inducing the lowest demineralization in both enamel and dentin (P<.05). Skim and semi-skim milk induced similar demineralization to the sucrose control, albeit slightly lower for semi-skim milk (P<.05). Whole and whole lactose-free milk produced lower biomass and less insoluble polysaccharides than the other treatments in enamel and dentin (P<.05). Adding 10 percent sucrose to whole milk turned it as cariogenic as 10 percent sucrose solution. CONCLUSION: Bovine whole milk seemed less cariogenic than sucrose and the other commercial milk types, but not anticariogenic. Fat content in milk seemed to reduce cariogenicity of the fluid.

Effect of water rinsing after acidulated phosphate fluoride gel on dental plaque acidity: an in situ study.

PURPOSE: This study's purpose was to determine whether rinsing with water or cleansing teeth after topical fluoride therapy affected plaque acidity. METHODS: This randomized, controlled, crossover, in situ study was conducted on 25 dental students who wore an acrylic mandibular appliance containing one enamel block. The patients were subjected to four treatment protocols involving acidulated phosphate fluoride (APF) application followed by rinsing the mouth with water: (1) after 30 minutes (APF-30); (2) after 15 minutes (APF-15); (3) immediately (APF-0); after cleansing the teeth with a cotton roll immediately following APF (APF-cotton); or (4) no fluoride therapy (control). After 48 hours, plaque pH was measured before and five, 10, 15, 20, and 30 minutes after rinsing with 10 percent sucrose solution. RESULTS: The least pH changes, the lowest pH drop, and the fastest pH recovery were found in the APF-30 and APF-15 groups, followed by the APF-0 and APF-cotton groups. CONCLUSIONS: Rinsing with water or cleansing the teeth with a cotton roll immediately after acidulated phosphate fluoride application can reduce its effect on dental plaque acidity; however, the inclusion of a controlled water rinse 15 minutes after APF gel application did not seem to influence the inhibitory effect of fluoride on plaque acidity.

Effect of cariogenic challenge on the stability of dentin bonds.

OBJECTIVE: The oral environment is subject to biofilm accumulation and cariogenic challenge, and few studies exist on the effect of these factors on the bond strength of adhesive systems. The aim of this study was to test if the exposure of adhesive interfaces to cariogenic challenge under biofilm accumulation could promote higher degradation than the exposure to biofilm accumulation alone. MATERIAL AND METHODS: Five molars were ground until exposure of medium dentin and then restored (Single Bond 2 and Z250 3M ESPE). The tooth/resin sets were cut to obtain beam-shaped specimens, which were distributed according to the aging conditions (n=20): water for 24 h (control); biofilm under cariogenic challenge for 3, 5 or 10 days; biofilm without cariogenic challenge for 10 days; and water for 3 months. Microcosm biofilms were formed from human saliva and grown in a saliva analogue medium, supplemented or not with sucrose to promote cariogenic challenge. Specimens were tested for microtensile bond strength, and failure modes were classified using light microscopy. Bond strength data were analyzed using ANOVA and failure modes were analyzed using ANOVA on ranks (α=0.05). RESULTS: No significant differences in bond strength were detected among the aging methods (P=0.248). The aging period was associated with an increase in the frequency of adhesive failures for the groups aged for 10 days or longer (P<0.001). CONCLUSION: Aging leads to a higher prevalence of interfacial adhesive failures, although this effect is not associated with cariogenic challenge or reduction in bond strengths.

Cariogenic effects of probiotic Lactobacillus rhamnosus GG in a dental biofilm model.

Probiotic bacteria have been suggested to inhibit Streptococcus mutans (SM) and thus prevent dental caries. However, supporting evidence is weak and probiotic species might be cariogenic themselves. Thus, we compared and combined the probiotic Lactobacillus rhamnosus GG (LGG) with SM and analysed the resulting mineral loss (ΔZ) in dental tissues. We simulated three biofilm compositions (SM, LGG, SM × LGG), two lesion sites (smooth enamel, dentin cavity) and two nutrition supply frequencies (twice/day, 6 times/day) in a multi-station, continuous-culture biofilm model. A total of 240 bovine enamel and dentin samples were cut, polished and embedded. All experimental procedures were performed in independent duplicates, with 10 samples being allocated to each group for each experiment (final sample size n = 20/group). Biofilms were cultured on the specimens and supplied with 2% sucrose medium and artificial saliva in consecutive pulses. After 10 days, ΔZ and bacterial numbers were assessed. SM × LGG biofilms caused significantly increased ΔZ compared with SM or LGG biofilms (p < 0.01, Mann-Whitney test), and ΔZ was significantly increased in dentin cavities compared with smooth enamel lesions (p < 0.01). Bacterial numbers did not significantly differ between biofilms of different species (p > 0.05, ANOVA). Frequent nutrition supply significantly increased bacterial numbers (p < 0.01). Biofilms in dentin cavities compared to smooth enamel harboured significantly more bacteria (p < 0.05). LGG induced mineral loss especially in dentin cavities and under highly cariogenic conditions. LGG did not have inhibitory effects on SM, but rather contributed to the caries process in vitro.

Sucrose substitutes affect the cariogenic potential of Streptococcus mutans biofilms.

Streptococcus mutans is considered the primary etiologic agent of dental caries and contributes significantly to the virulence of dental plaque, especially in the presence of sucrose. To avoid the role of sucrose on the virulence factors of S. mutans, sugar substitutes are commonly consumed because they lead to lower or no production of acids and interfere with biofilm formation. This study aimed to investigate the contribution of sugar substitutes in the cariogenic potential of S. mutans biofilms. Thus, in the presence of sucrose, glucose, sucralose and sorbitol, the biofilm mass was quantified up to 96 h, the pH of the spent culture media was measured, the expression of biofilm-related genes was determined, and demineralization challenge experiments were conduct in enamel fragments. The presence of sugars or sugar substitutes profoundly affected the expression of spaP, gtfB, gtfC, gbpB, ftf, vicR and vicX in either biofilm or planktonic cells. The substitution of sucrose induced a down-regulation of most genes involved in sucrose-dependent colonization in biofilm cells. When the ratio between the expression of biofilm and planktonic cells was considered, most of those genes were down-regulated in biofilm cells in the presence of sugars and up-regulated in the presence of sugar substitutes. However, sucralose but not sorbitol fulfilled the purpose of reducing the cariogenic potential of the diet since it induced the biofilm formation with the lowest biomass, did not change the pH of the medium and led to the lowest lesion depth in the cariogenic challenge.