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Analysis of cariogenic potential of alternative milk beverages by in vitro Streptococcus mutans biofilm model and ex vivo caries model.

ABSTRACT: OBJECTIVE:To evaluate the potential of various alternative milk beverages to support bacterial biofilm formation and acid production and cause unbalanced demineralization. DESIGN:in vitro assays were used to examine the ability of the beverages to support Streptococcus mutans' biofilm formation and acid production from sugar fermentation and the capacity of the beverages to buffer pH changes. Biofilm formation was done using 96-well plate model. Acid production was measured using L-Lactate assay kit, and the buffering capacity was assessed by pH titration. For ex vivo caries model, enamel and dentine slabs and S. mutans biofilms were exposed to selected alternative milk beverages three times a day, 30?min each, and by the end of the experiments, slab's demineralization was assessed by loss of surface microhardness. RESULTS:Of the alternative milk beverages tested in this study, Original Almond consistently supported the most S. mutans biofilms, followed by Chocolate Cashew Milk, while the least biofilms were measured with Unsweetened Flax Milk. The most acids and the lowest culture pH were measured with Toasted Coconut Almond Milk, while the least buffering capacity was measured with Unsweetened Coconut Milk. The results of ex vivo caries model showed that like Bovine Whole Milk, repeated exposure to Original Almond led to significant enamel and dentine slab demineralization, when compared to those exposed to saline as a control (P?

SUBMITTER: Huang Y

PROVIDER: S-EPMC6629491 | biostudies-literature | 2019 Sep

REPOSITORIES: biostudies-literature

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Analysis of cariogenic potential of alternative milk beverages by in vitro Streptococcus mutans biofilm model and ex vivo caries model.

Huang Yan Y Thompson Tatyana T Wang Yapin Y Yu Qingzhao Q Zhu Lin L Xu Xiaoming X Wen Zezhang T ZT Townsend Janice A JA

Archives of oral biology 20190626

<h4>Objective</h4>To evaluate the potential of various alternative milk beverages to support bacterial biofilm formation and acid production and cause unbalanced demineralization.<h4>Design</h4>in vitro assays were used to examine the ability of the beverages to support Streptococcus mutans' biofilm formation and acid production from sugar fermentation and the capacity of the beverages to buffer pH changes. Biofilm formation was done using 96-well plate model. Acid production was measured using ...[more]

PMID: 31276938

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Project description:Streptococcus mutans is a Gram-positive, facultative anaerobic bacterium, which causes dental caries after forming biofilms on the tooth surface while producing organic acids that demineralize enamel and dentin. We observed that the polyunsaturated arachidonic acid (AA) (ω-6; 20:4) had an anti-bacterial activity against S. mutans, which prompted us to investigate its mechanism of action. The minimum inhibitory concentration (MIC) of AA on S. mutans was 25 μg/ml in the presence of 5% CO2, while it was reduced to 6.25-12.5 μg/ml in the absence of CO2 supplementation. The anti-bacterial action was due to a combination of bactericidal and bacteriostatic effects. The minimum biofilm inhibitory concentration (MBIC) was the same as the MIC, suggesting that part of the anti-biofilm effect was due to the anti-bacterial activity. Gene expression studies showed decreased expression of biofilm-related genes, suggesting that AA also has a specific anti-biofilm effect. Flow cytometric analyses using potentiometric DiOC2(3) dye, fluorescent efflux pump substrates, and live/dead SYTO 9/propidium iodide staining showed that AA leads to immediate membrane hyperpolarization, altered membrane transport and efflux pump activities, and increased membrane permeability with subsequent membrane perforation. High-resolution scanning electron microscopy (HR-SEM) showed remnants of burst bacteria. Furthermore, flow cytometric analysis using the redox probe 2',7'-dichlorofluorescein diacetate (DCFHDA) showed that AA acts as an antioxidant in a dose-dependent manner. α-Tocopherol, an antioxidant that terminates the radical chain, counteracted the anti-bacterial activity of AA, suggesting that oxidation of AA in bacteria leads to the production of cytotoxic radicals that contribute to bacterial growth arrest and death. Importantly, AA was not toxic to normal Vero epithelial cells even at 100 μg/ml, and it did not cause hemolysis of erythrocytes. In conclusion, our study shows that AA is a potentially safe drug that can be used to reduce the bacterial burden of cariogenic S. mutans.

Project description:Streptococcus mutans is a Gram-positive bacterium highly associated with dental caries, and it has a strong biofilm-forming ability, especially in a sugar-rich environment. Many strategies have been undertaken to prevent dental caries by targeting these bacteria. Recently, we observed that a sustained-release varnish containing triclosan and cannabidiol (CBD) was more efficient than each compound alone in preventing biofilm formation by the fungus Candida albicans, which is frequently involved in oral infections together with S. mutans. It was therefore inquiring to study the effect of this drug combination on S. mutans. We observed that the combined treatment of triclosan and CBD had stronger anti-bacterial and anti-biofilm activity than each compound alone, thus enabling the use of lower concentrations of each drug to achieve the desired effect. The combined drug treatment led to an increase in the SYTO 9low, propidium iodide (PI)high bacterial population as analyzed by flow cytometry, indicative for bacteria with disrupted membrane. Both triclosan and CBD induced membrane hyperpolarization, although there was no additive effect on this parameter. HR-SEM images of CBD-treated bacteria show the appearance of elongated and swollen bacteria with several irregular septa structures, and upon combined treatment with triclosan, the bacteria took on a swollen ellipse and sometimes oval morphology. Increased biofilm formation was observed at sub-MIC concentrations of each compound alone, while combining the drugs at these sub-MIC concentrations, the biofilm formation was prevented. The inhibition of biofilm formation was confirmed by CV biomass staining, MTT metabolic activity, HR-SEM and live/dead together with exopolysaccharide (EPS) staining visualized by spinning disk confocal microscopy. Importantly, the concentrations required for the anti-bacterial and anti-biofilm activities toward S. mutans were non-toxic to the normal Vero epithelial cells. In conclusion, the data obtained in this study propose a beneficial role of combined triclosan/CBD treatment for potential protection against dental caries.

Project description:The aim of the study was to verify the hypothesis about differences in sweet taste perception in the group of preschool children with and without caries, and to determine its relationship with cariogenic microbiota and the frequency of sweets consumption in children. The study group included of 63 children aged 2-6 years: 32 with caries and 31 without caries. The study consisted of collecting questionnaire data and assessment of dental status using the decayed, missing, filled in primary teeth index (dmft) and the International Caries Detection and Assessment System (ICDAS II). The evaluation of sweet taste perception was carried out using a specific method that simultaneously assessed the level of taste preferences and the sensitivity threshold for a given taste. The microbiological analysis consisted of the assessment of the quantitative and qualitative compositions of the oral microbiota of the examined children. The sweet taste perception of children with caries was characterized by a lower susceptibility to sucrose (the preferred sucrose solution concentration was >4 g/L) compared to children without caries (in the range ? 4 g/L, p = 0.0015, chi-square test). A similar relationship was also observed for frequent snacking between meals (p = 0.0038, chi-square test). The analysis of studied variables showed the existence of a strong positive correlation between the perception of sweet taste and the occurrence and intensity of the cariogenic process (p = 0.007 for dmft; and p = 0.012 for ICDAS II), as well as the frequency of consuming sweets (p ? 0.001 for frequent and repeated consumption of sweets during the day, Spearman test) in children with caries. Additionally, children with an elevated sucrose taste threshold were more than 10-times more likely to develop S. mutans presence (OR = 10.21; 95% CI 3.11-33.44). The results of this study suggest the future use of taste preferences in children as a diagnostic tool for the early detection of increased susceptibility to caries through microbial dysbiosis towards specific species of microorganisms.

Project description:Background:It is cleared that some probiotic strains inhibit biofilm formation of oral bacteria, but its mechanisms are not clearly understood yet. It is proposed that one of the mechanisms can be biosurfactant production, a structurally diverse group of surface-active compounds synthesized by microorganisms. Hence, this study focused on the evaluation of the anti-biofilm and antiadhesive activities of the L. rhamnosus derived-biosurfactant against Streptococcus mutans and its effect on gtfB/C and ftf genes expression level. Materials and Methods:In this in vitro study Lactobacillus rhamnosus ATCC7469 overnight culture was used for biosurfactant production. The biosurfactant effect on the surface tension reduction was confirmed by drop collapse method. Chemical bonds in the biosurfactant were identified by Fourier transform infrared (FTIR). Anti-biofilm and antiadhesive activities of the biosurfactant were determined on glass slides and in 96-well culture plates, respectively. The effect of the biosurfactant on gtfB/C and ftf genes expression level was also investigated after biofilm formation, total RNA extraction, and reverse transcription by quantitative real-time reverse transcriptase polymerase chain reaction (PCR) assay (quantitative PCR). The data were assessed by one-way analysis of variance in the Tukey-Kramer postdeviation test for all pairs. P < 0.05 was considered statistically significant. Results:The FTIR results of biosurfactant showed that it was protein rich. It also showed anti-biofilm formation activity on the glass slide and antiadhesive activity till 40% on microtiter plate wells. It also showed a significant reduction (P < 0.05) in gtfB/C and ftf genes expression level. Conclusion:L. rhamnosus-derived biosurfactant exhibits a significant inhibitory effect on biofilm formation ability of S. mutans due to downregulation of biofilm formation associated genes, gtfB/C and ftf. L. rhamnosus-derived biosurfactant with substantial antiadhesive activity is suitable candidates for use in new generations of microbial antiadhesive agents.

Dataset Information

Analysis of cariogenic potential of alternative milk beverages by in vitro Streptococcus mutans biofilm model and ex vivo caries model.

Publications

Analysis of cariogenic potential of alternative milk beverages by in vitro Streptococcus mutans biofilm model and ex vivo caries model.

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