Evaluation of Streptococcus mutans biofilms formed on fluoride releasing and non fluoride releasing resin composites

Objectives

The aim of this study was to evaluate the acid production, acid tolerance and composition of Streptococcus mutans biofilms formed on fluoride releasing and non fluoride releasing resin composites.

Methods

S. mutans biofilms were formed on saliva-coated discs prepared from fluoride releasing (Unifil Flow and F2000) or non fluoride releasing materials (Filtek Z350, GRADIA DIRECT and hydroxyapatite). To assess the level of acid production and acid tolerance, glycolytic pH drop and proton permeability assays were performed using 94 h old S. mutans biofilms. To evaluate the biofilm composition, the biomass (total dry-weight), colony forming unit (CFU), water-insoluble extracellular polysaccharides (EPS), water-soluble EPS and intracellular iodophilic polysaccharides (IPS) of 94 h old S. mutans biofilms were analysed. The amount of fluoride of old culture medium released from the materials during the experimental period was also determined. Each assay was performed in duplicate in at least four different experiments (n = 8).

Results

All biofilms showed similar initial rates of acid production (0.083–0.089 pH drop/min) and proton permeability (0.025–0.036 pH increase/min), irrespective of fluoride release from the materials. On the other hand, the amount of biomass, water-insoluble EPS and IPS of the biofilms on Unifil Flow, which releases a larger amount of fluoride in the early stages of biofilm formation, were significantly lower than those on the other materials (up to 27%, 38% and 36% reduction in biomass, water-insoluble and IPS, respectively).

Conclusions

Our finding suggests that fluoride releasing resin composites might contribute to the decrease in cariogenic composition of S. mutans biofilms if an appropriate amount of fluoride is released in the early stages of biofilm formation.     

Antimicrobial activity of nanoemulsion on cariogenic Streptococcus mutans

Objective

To assess antimicrobial activities of nanoemulsion (NE) to control the adhesion and biofilm formation by Streptococcus mutans by in vitro.

Design

In vitro antimicrobial susceptibility of nanoemulsion was determined as per National Committee for Clinical Laboratory Standards guidelines and agar diffusion, serial dilution technique for the determination of minimum inhibitory concentration and minimum bactericidal concentration (MIC/MBC). Efficacy was tested by kinetics of killing, biofilm assay and scanning electron microscopy.

Results

: NE concentrations ranging from 1:100 to 1:10,000 dilutions were effective against S. mutans as shown through MIC/MBC assays. NE showed antimicrobial activity against planktonic cells at high dilutions, confirmed by time kill studies. 4-day-old S. mutans biofilms were treated with NE; subsequent reductions of bacterial cell counts were noticed with decreasing dilutions. Staining of NE-treated biofilms with LIVE/DEAD BacLight resulted in dead cell areas of up to 48% in 1 min, 84% at 1 h and significant (<0.05) increases in dead cell counts at all time points. Damage to cell membranes and cell walls of S. mutans by NE was demonstrated using scanning electron microscopy (SEM).

Conclusion

These results suggest that nanoemulsion has effective antibacterial activity against S. mutans and may be a useful medication in the prevention of dental caries.     

The antimicrobial efficacy of Fructus mume extract on orthodontic bracket: A monospecies-biofilm model study in vitro

Objective

The aim of this study was to evaluate the antimicrobial efficacy of Traditional Chinese Medicine Fructus mume on a monospecies-biofilm model established on orthodontic brackets in vitro.

Materials and methods

The antimicrobial effect of Fructus mume aqueous extract on the planktonic Streptococcus mutans (S. mutans) was tested by microdilution method (MIC). The cell viability of S. mutans biofilm on Damon3 MX bracket (Ormco, USA) after exposed to Fructus mume extract was quantified by XTT reduction assay. Visualization of the samples was performed by fluorescence microscope and confocal laser scanning microscopy (CLSM).

Results

HPLC analysis revealed that the main compounds of Fructus mume are organic acids. The MIC of Fructus mume extract on the planktonic S. mutans was 50 mg/mL. The optical density (OD) values, measured by XTT reduction assay from S. mutans biofilms after 1-min exposure to different test agents, demonstrated that the cell viability of S. mutans biofilms exposed to 250 mg/mL Fructus mume extract < BHI (−) (p < 0.01). Microscope image showed that Fructus mume extract obviously increased the amount of dead bacteria on the surface of bracket.

Conclusion

Fructus mume extract showed antimicrobial effect on S. mutans biofilm on orthodontic bracket in vitro which may indicate its potential use as an oral antimicrobial agent for orthodontic patients.     

Effect of biofilm formation on virulence factor secretion via the general secretory pathway in Streptococcus mutans

Objectives

To investigate the role of SecA in protein secretion, and to evaluate the effect of biofilm formation on protein secretion in Streptococcus mutans.

Design

S. mutans strains UA159 and GS-5 were used in this study. Cells grown in biofilm and planktonic conditions were observed using immunogold electron microscopy. The mRNA levels of ftf, gtfB, gtfC, gtfD, Pac and secA were analysed in different growth conditions using real-time quantitative polymerase chain reaction. The levels of wall proteins and whole-cell protein extracts were examined using Western blot analysis.

Results

A microdomain colocalised with SecA and virulence factors such as Pac (AgI/II) and glucosyltransferase (GTF) was observed. The mRNA level of secA was upregulated in the biofilm condition. The level of protein expression of SecA and wall protein levels of GTF, fructosyltransferase (FTF) and Pac (AgI/II) in the biofilm condition were significantly higher than in the planktonic condition.

Conclusions

These data suggest that S. mutans utilises the Sec pathway to secrete virulence factor proteins such as Pac (AgI/II), GTF and FTF, and protein secretion occurred at a distinct microdomain. The level of SecA, the key factor in the Sec pathway, was influenced significantly by biofilm formation in S. mutans.     

In vitro study of the properties of Streptococcus mutans in starvation conditions

Objectives

The present study is to assess Streptococcus mutans survivability in different starvation conditions and to determine the resistance of starved S. mutans to lethal acid and two common anti-caries agents, sodium fluoride (NaF) and chlorhexidine acetate (CHX).

Methods

S. mutans survival rates in sterile water, PBS, sterile saliva, 1/5 strength BHI and BHI were determined at a given time by plate count of viable cell. The resistance of starved S. mutans and control S. mutans to four times the minimal bactericidal concentration (MBC) of NaF, two times the MBC of CHX and acid (pH 2.8) was evaluated and compared respectively. Furthermore, field emission scanning electron microscope (FE-SEM) was used to observe the morphologic characteristics of the starved S. mutans cells.

Results

S. mutans showed starvation tolerance under five different starvation conditions, but the bacterial survival rates were different at the same time points. The starved S. mutans exhibited significantly higher resistance (p < 0.05) to challenge by anti-caries agents and acid than the control S. mutans. Additionally, starvation resulted in the morphologic modification of S. mutans, and the disruptive degree depended on the change in time.

Conclusion

The present study indicates that S. mutans displays starvation tolerance, and starvation decreased the susceptibility of S. mutans to NaF, CHX and acid.     

Application of an active attachment model as a high-throughput demineralization biofilm model

Objectives

To investigate the potential of an active attachment biofilm model as a high-throughput demineralization biofilm model for the evaluation of caries-preventive agents.

Methods

Streptococcus mutans UA159 biofilms were grown on bovine dentine discs in a high-throughput active attachment model. Biofilms were first formed in a medium with high buffer capacity for 24 h and then subjected to various photodynamic therapies (PACT) using the combination of Light Emitting Diodes (LEDs, Biotable^®) and Photogem^®. Viability of the biofilms was evaluated by plate counts. To investigate treatment effects on dentine lesion formation, the treated biofilms were grown in a medium with low buffer capacity for an additional 24 h. Integrated mineral loss (IML) and lesion depth (LD) were assessed by transversal microradiography. Calcium release in the biofilm medium was measured by atomic absorption spectroscopy.

Results

Compared to the water treated control group, significant reduction in viability of S. mutans biofilms was observed when the combination of LEDs and Photogem^® was applied. LEDs or Photogem^® only did not result in biofilm viability changes. Similar outcomes were also found for dentine lesion formation. Significant lower IML and LD values were only found in the group subjected to the combined treatment of LEDs and Photogem^®. There was a good correlation between the calcium release data and the IML or LD values.

Conclusions

The high-throughput active attachment biofilm model is applicable for evaluating novel caries-preventive agents on both biofilm and demineralization inhibition. PACT had a killing effect on 24 h S. mutans biofilms and could inhibit the demineralization process.     

Inhibitory effects of fruit berry extracts on Streptococcus mutans biofilms

Dark‐colored fruit berries are a rich source of polyphenols that could provide innovative bioactive molecules as natural weapons against dental caries. High‐quality extracts of cranberry, blueberry, and strawberry, and a combination of the three berry extracts (Orophenol), were used to treat 24‐h‐old Streptococcus mutans biofilms. The grown biofilms were treated with the berry extracts at concentrations ranging from 62.5 to 500 μg ml^?1. Treated biofilms were assessed for metabolic activity, acidogenicity, biovolumes, structural organization, and bacterial viability. The biofilms treated with the cranberry and Orophenol extracts exhibited the most significant reductions in metabolic activity, acid production, and bacterial/exopolysaccharide (EPS) biovolumes, while their structural architecture appeared less compact than the control‐treated biofilms. The blueberry extract produced significant reductions in metabolic activity and acidogenicity only at the highest concentration tested, without significantly affecting bacterial/EPS biovolumes or biofilm architecture. Strawberry extracts had no significant effects on S. mutans biofilms. None of the berry extracts were bactericidal for S. mutans. The results indicate that cranberry extract was the most effective extract in disrupting S. mutans virulence properties without significantly affecting bacterial viability. This suggests a potential ecological role for cranberry phenols as non‐bactericidal agents capable of modulating pathogenicity of cariogenic biofilms.     

Corrosion behaviour of titanium in the presence of Streptococcus mutans

Objective

The main aim of this in vitro study was to evaluate the influence of Streptococcus mutans on the corrosion of titanium.

Methods

S. mutans biofilms were formed on commercially pure titanium (CP-Ti) square samples (10 mm × 10 mm × 1 mm) using a culture medium enriched with sucrose. Open circuit potential (OCP) and electrochemical impedance spectroscopy (EIS) measurements were used to evaluate the corrosion behaviour of CP-Ti in the presence of S. mutans in Fusayama's artificial saliva. The corrosion of biofilm-free CP-Ti samples was also evaluated in artificial saliva. Biofilms biomass was measured by spectrophotometry, using crystal violet staining, after 1, 2 and 7 days.

Results

The OCP values recorded on CP-Ti in the presence of S. mutans (−0.3 ± 0.02 V vs. SCE) was lower than those on biofilm-free CP-Ti (−0.1 ± 0.01 V vs. SCE) after 2 h of immersion in artificial saliva (p < 0.05). That reveals a high reactivity of titanium in presence of S. mutans. Impedance spectra revealed the formation of a compact passive film on titanium in artificial saliva or in the presence of a 2 days old S. mutans biofilm even though the corrosion resistance of CP-Ti has decreased in presence of a S. mutans biofilm.

Conclusion

The presence of bacterial colonies, such as S. mutans, negatively affected the corrosion resistance of the titanium.     

Evaluation of Streptococcus mutans adhesion to fluoride varnishes and subsequent change in biofilm accumulation and acidogenicity

Objectives

The aim of this study was to evaluate Streptococcus mutans adhesion to fluoride varnishes and subsequent change in biofilm accumulation and acidogenicity.

Methods

After producing fluoride varnish-coated hydroxyapatite discs using Fluor Protector (FP), Bifluorid 12 (BIF), Cavity Shield (CASH), or Flor-Opal Varnish White (FO), S. mutans biofilms were formed on the discs. To assess S. mutans adhesion to the discs, 4-h-old biofilms were analysed. To investigate the change in biofilm accumulation during subsequent biofilm formation, the biomass, colony forming units (CFU), and water-insoluble extracellular polysaccharides (EP) of 46-, 70-, and 94-h-old biofilms were analysed. To investigate the change in acidogenicity, pH values of the culture medium were determined during the experimental period. The amount of fluoride in the culture medium was also determined during the experimental period.

Results

BIF, CASH, and FO affected S. mutans adhesion (67–98% reduction) and subsequent biofilm accumulation in 46-, 70-, and 94-h-old biofilms. However, the reducing effect of the fluoride varnishes on the biomass, CFU count, water-insoluble EP amount, and acid production rate of the biofilms decreased as the biofilm age increased. These results may be related to the fluoride-release pattern of the fluoride varnishes. Of the fluoride varnishes tested, FO showed the highest reducing effect against the bacterial adhesion and subsequent biofilm accumulation.

Conclusions

Our findings suggest that if the results of these experiments are extrapolable to the in vivo situation, then reduced clinical benefit of using fluoride varnishes may occur with time.

Clinical significance

Fluoride varnish application can affect cariogenic biofilm formation but the anti-biofilm activity may be reduced with time.     

Effects of periodic fluoride treatment on fluoride ion release from fresh orthodontic adhesives

Objective

Periodic fluoride treatment may contribute to the ability of fresh orthodontic adhesives to provide long-term F⁻ release. The effects of periodic fluoride treatment on the amount of F⁻ release from fresh orthodontic adhesives was investigated.

Methods

F⁻ release was measured from a nonfluoride-releasing composite, a fluoride-releasing composite, a polyacid-modified composite (compomer), and two resin-modified glass-ionomer cements (RMGICs) at 1, 2, and 5 days after one of the following treatments: 225 ppm F⁻ solution, 900 ppm F⁻ solution, acidulated phosphate fluoride gel (APF), fluoridated dentifrice, and deionised water (control). F⁻ release was measured in a 5-day cycle, which was repeated 9 consecutive times. The amount of F⁻ release for each group was analysed using the repeated measures analysis of variance. Statistical significance was set at a level of α = 0.05.

Results

Periodic fluoride treatment temporarily increased F⁻ release in fresh fluoride-releasing orthodontic adhesives, but not in fresh nonfluoride-releasing composite. The order of effective fluoride-release was RMGICs > compomer > fluoride-releasing composite > nonfluoride-releasing composite. The application of APF or 900 ppm F⁻ solution was the most effective way to maintain F⁻ release from fresh orthodontic adhesives. However, the amount of F⁻ release gradually decreased with increasing specimen age.

Conclusion

Given the difficulty of routine use of APF at home, the results of this study show that a combination of RMGICs and high-dose fluoride mouth rinse is the most effective protocol to maintain F⁻ release from fresh orthodontic adhesives.

Clinical significance

Most studies have investigated fluoride-uptake abilities using aged materials in which fluoride had been lost for at least 1 month. This study has found that periodic fluoride treatment altered the conventional F⁻ release pattern of fresh fluoride-releasing materials and type of fluoride-containing medium plays a more critical role in fluoride recharging of the materials than fluoride concentration. This study will help clinicians to find the most effective fluoride treatment protocol of fresh materials.     

Correlation of biological properties with glucan-binding protein B expression profile in Streptococcus mutans clinical isolates

Objective

Streptococcus mutans is known to be a primary causative agent of dental caries and its surface proteins have been investigated to specify their association with its virulence. Amongst those, 4 glucan-binding proteins (Gbps) are considered to be important factors due to their glucan-binding properties, of which GbpB has been shown to participate in cell-wall construction and cell separation.

Design

We examined clinical isolates of S. mutans collected from the oral cavities of Japanese and Finnish subjects, and focused on the association of their GbpB expression profiles and biological properties related to virulence.

Results

Western blot analysis of GbpB expression by the isolates revealed a variety of patterns. Strains that showed single and multiple bands were used to designate S and M type strains, respectively, whilst those with no GbpB expression were classified as N type. The distribution of GbpB expression patterns was shown to be quite different between the Japanese and Finnish isolates. Furthermore, the chain length and doubling time of the N type in both populations were significantly longer than those of the other types.

Conclusion

Our results suggest variations in S. mutans GbpB expression patterns, which may have relationships with the virulence of S. mutans.     

Antiadherent activity of Schinus terebinthifolius and Croton urucurana extracts on in vitro biofilm formation of Candida albicans and Streptococcus mutans

Objective

To evaluate the antiadherent property of crude, methanol and acetate methanol extract fractions from Schinus terebinthifolius and Croton urucurana in hydroalcoholic (HA) and dimethylsulfoxide (DMSO) solvents on in vitro biofilms formed by Streptococcus mutans and Candida albicans strains.

Design

The minimal concentration of adherence (MICA) was determined to evaluate the antiadherent potential of extracts on the in vitro biofilm formation. The extracts of plants were subjected to thin layer chromatography (TLC) in order to detect what class of compounds was responsible for the antiadherent activity. Data were estimated by analysis of variance (ANOVA) complemented by Tukey test level of significance set at 5%.

Results

Both plants demonstrated inhibition of S. mutans and C. albicans on in vitro biofilm formation. The biofilms of C. albicans were more efficiently inhibited by the S. terebinthifolius fraction of acetate–methanol and methanol in hydroalcoholic solvents (p < 0.05). The S. mutans biofilms adherence was best inhibited by the S. terebinthifolius crude extract and its methanolic fraction, both in hydroalcoholic solvent (p < 0.05). TLC of crude extracts and fractions of S. terebinthifolius detected the presence of several active compounds, including phenolic compounds, anthraquinones, terpenoids, and alkaloids. C. urucurana extracts confirmed activity for both microorganisms (p < 0.05). However, higher concentrations were needed to achieve antiadherent activity, mainly to inhibit in vitro biofilm formation of C. albicans.

Conclusion

The antiadherent potential of both plants on in vitro biofilms formed by C. albicans and S. mutans were confirmed, suggesting the importance of studies about these extracts for therapeutic prevention of oral diseases associated with oral biofilms.     

Acidogenicity of dual-species biofilms of bifidobacteria and Streptococcus mutans

Objective

The aim of this study was to evaluate the acidogenicity of dual-species biofilms of bifidobacteria and Streptococcus mutans.

Materials and methods

The following strains were tested: Bifidobacterium dentium DSM20436, Parascardovia denticolens DSM10105, and Scardovia inopinata DSM10107. Streptococcus mutans UA159 and Lactobacillus acidophilus ATCC4356 were used as control. Bifidobacteria were studied planktonically as they were not able to form monospecies biofilm, they were grown in biofilms associated with S. mutans. Endogenous polysaccharide reserves of cultures at log phase were depleted. Standardized suspensions of the microorganisms were incubated in growth media supplemented with 10 mM glucose, lactose, raffinose, glucose, or xylitol. S. mutans biofilms were grown on glass cover slips for 24 h to which bifidobacteria were added. After 24 h, the dual-species biofilms were exposed to the same carbon sources, and after 3 h, the pH of spent culture media and concentrations of organic acids were measured. Statistical analyses were carried out using ANOVA and Tukey’s test (α = 0.05).

Results

A higher pH drop was observed when S. mutans was associated with P. denticolens or S. inopinata, in either planktonic or biofilm cultures, than with S. mutans alone. Bifidobacteria showed a higher pH drop in the presence of raffinose than S. mutans or L. acidophilus.

Conclusions

Dual-species biofilms of bifidobacteria and S. mutans produced more acid and greater pH drops than biofilms of S. mutans alone.

Clinical relevance

New insights on the complex process of caries pathogenicity contribute to the establishment of preventive and therapeutic measures, in particular in specific cases, such as in early childhood caries.

     

Inhibitory effect of Lactobacillus salivarius on Streptococcus mutans biofilm formation

Dental caries arises from an imbalance of metabolic activities in dental biofilms developed primarily by Streptococcus mutans. This study was conducted to isolate potential oral probiotics with antagonistic activities against S. mutans biofilm formation from Lactobacillus salivarius, frequently found in human saliva. We analysed 64 L. salivarius strains and found that two, K35 and K43, significantly inhibited S. mutans biofilm formation with inhibitory activities more pronounced than those of Lactobacillus rhamnosus GG (LGG), a prototypical probiotic that shows anti‐caries activity. Scanning electron microscopy showed that co‐culture of S. mutans with K35 or K43 resulted in significantly reduced amounts of attached bacteria and network‐like structures, typically comprising exopolysaccharides. Spot assay for S. mutans indicated that K35 and K43 strains possessed a stronger bactericidal activity against S. mutans than LGG. Moreover, quantitative real‐time polymerase chain reaction showed that the expression of genes encoding glucosyltransferases, gtfB, gtfC, and gtfD was reduced when S. mutans were co‐cultured with K35 or K43. However, LGG activated the expression of gtfB and gtfC, but did not influence the expression of gtfD in the co‐culture. A transwell‐based biofilm assay indicated that these lactobacilli inhibited S. mutans biofilm formation in a contact‐independent manner. In conclusion, we identified two L. salivarius strains with inhibitory activities on the growth and expression of S. mutans virulence genes to reduce its biofilm formation. This is not a general characteristic of the species, so presents a potential strategy for in vivo alteration of plaque biofilm and caries.     

Microbicidal efficacy of thiocyanate hydrogen peroxide after adding lactoperoxidase under saliva loading in the quantitative suspension test

Objective

As shown in the quantitative suspension test adding lactoperoxidase to a thiocyanate (SCN⁻) hydrogen peroxide (H₂O₂) combination over the physiological saliva level has significant positive antimicrobial effects to a level of totally killing Streptococcus mutans, Streptococcus sanguinis, and Candida albicans. The aim of this study was to evaluate this positive effect under human saliva loading.

Methods

The bactericidal and fungicidal effect of lactoperoxidase was evaluated in a quantitative suspension test by using two test mixtures of a 2.0% thiocyanate and 1.2% hydrogen peroxide solution, one without (Group A) and one with (Group B) lactoperoxidase under saliva loading. Following the quantitative suspension tests (EN-13727/EN-13624), the growth of surviving bacteria and fungi in a nutrient broth was measured. The exposure times were restricted to 1, 3, 5, and 15 min. All statistical analyses were carried out with SPSS 11.5.

Results

In the quantitative suspension test, the combination of thiocyanate and hydrogen peroxide showed relatively low antimicrobial effectiveness on S. mutans, S. sanguinis, and C. albicans in the presence of human saliva at measured time points in comparison to the mixture with lactoperoxidase, which showed a high bactericidal activity within 15 min (S. mutans and S. sanguinis) and fungicidal activity within 3 min (C. albicans).

Conclusion

The antimicrobial effectiveness of the tested thiocyanate hydrogen peroxide combination was increased significantly by adding lactoperoxidase in the quantitative suspension test under human saliva loading.     

Clotrimazole and econazole inhibit Streptococcus mutans biofilm and virulence in vitro

Objective: The aim of this study was to determine the inhibitory effect of eight antifungal drugs on S. mutans growth, biofilm formation and virulence factors.MethodsThe actions of antifungal drugs on S. mutans were determined by recovery plates and survival kinetic curves. Biofilms were observed by scanning electron microscopy and the viable cells were recovered on BHI plates, meanwhile biofilms were stained by BacLight live/dead kit to investigate the biofilm viability. Bacteria/extracellular polysaccharides staining assays were performed to determine the EPS production of S. mutans biofilms. Acidogenicity and acidurity of S. mutans were determined using pH drop and acid tolerance assays, and the expression of ldh gene was evaluated using qPCR.ResultsWe found that clotrimazole (CTR) and econazole (ECO) showed antibacterial activities on S. mutans UA159 and S. mutans clinical isolates at 12.5 and 25 mg/L, respectively. CTR and ECO could also inhibit S. mutans biofilm formation and reduce the viability of preformed biofilm. CTR and ECO affected the live/dead ratio and the EPS/bacteria ratio of S. mutans biofilms. CTR and ECO also inhibited the pH drop, lactate acid production, and acid tolerance. The abilities of CTR and ECO to inhibit S. mutans ldh expression were also confirmed.ConclusionsWe found that two antifungal azoles, CTR and ECO, had the abilities to inhibit the growth and biofilm formation of S. mutans and more importantly, they could also inhibit the virulence factors of S. mutans.     

Antimicrobial activity of Bifidobacterium spp. isolated from healthy adult Koreans against cariogenic microflora

Objective

Dental caries is the main common infectious disease in the human oral cavity. Streptococcus mutans and Streptococcus sobrinus were reported to be the most important etiological factors in human dental caries. Thus, we examined the inhibitory effects of Bifidobacterium spp. cells and culture supernatants against S. mutans and S. sobrinus, including Streptococcus gordonii, and Aggregatibacter actinomycetemcomitans, which is associated with periodontal disease.

Methods

Mutans streptococci or A. actinomycetemcomitans and lactic acid bacteria (LAB) were mixed in 1:1 ratio and then incubated for 90 min at 37 °C. After the incubation, the viability of mutans streptococci or A. actinomycetemcomitans was determined by plate count technique. We also investigated the morphological changes of S. mutans treated with LAB using scanning electron microscopy (SEM).

Results

In vitro viability of S. mutans, S. sobrinus, S. gordonii, and A. actinomycetemcomitans was affected by human intestinal LAB identified as Bifidobacterium adolescentis SPM1005 and Bifidobacterium longum SPM1207. Especially, B. adolescentis SPM1005 cells at 1.0 × 10⁸ CFU had a strong growth-inhibiting effect against S. mutans and induced a 64% loss of its viability (p < 0.05). In addition, swollen and disrupted S. mutans were observed after incubation with B. adolescentis SPM1005. However, the culture supernatant of this strain did not show such inhibitory activity.

Conclusion

B. adolescentis SPM1005 cells decreased the growth of S. mutans, which is a risk factor for dental caries. Therefore, we suggest that this Bifidobacterium strain may be a useful probiotic microorganism for prevention of dental caries that does not have adverse effects.     

Inhibitory effects of tea catechin epigallocatechin-3-gallate against biofilms formed from Streptococcus mutans and a probiotic lactobacillus strain

ObjectiveEffects of tea catechin epigallocatechin-3-gallate (EGCG) against biofilm formation by Streptococcus mutans and probiotic Lactobacillus casei in Yakult^® (LcY) were examined.DesignBiofilms were formed by S. mutans alone (Sm) and co-culture of S. mutans and LcY (Sm + LcY) in the absence or presence of EGCG. The biomass of biofilms, which were sonicated or not, was measured by the crystal violet assay. Biofilm morphology was observed by scanning electron microscopy. Bacterial viability and extracellular polysaccharides were determined by SYTO9/propidium iodide and dextran-conjugated fluorescein staining, respectively, and confocal microscopy. Gene expression of glucosyltransferase was determined by quantitative polymerase chain reaction.ResultsWhile 250 μg/ml EGCG significantly decreased the biomass and acid production of Sm biofilms, 500 μg/ml EGCG was required to inhibit Sm + LcY biofilm formation and acid production. EGCG decreased the amount of live bacteria present in both Sm and Sm + LcY biofilms. The level of dead bacteria in Sm + LcY biofilms was higher than in Sm biofilms when formed in the presence of 250 μg/ml EGCG. EGCG decreased levels of extracellular polysaccharides in Sm and Sm + LcY biofilms. The extent of biofilm removal by sonication was not different between Sm and Sm+LcY biofilms formed in the absence or presence of 62.5 or 125 μg/ml EGCG. The level of Sm gtfB and gtfD expression in Sm + LcY biofilms was higher than those in the Sm biofilms when formed in the presence of EGCG at 250 μg/ml.ConclusionThe results indicated that LcY might interfere the inhibitory effects of EGCG against biofilm formation by S. mutans.