Streptococcus Mutans Biofilm Formation Research Articles

Development of Streptococcus mutans biofilm in the presence of human colostrum and 3′-sialyllactose

Aim To evaluate the initial adhesion and formation of Streptococcus mutans biofilm in vitro in the presence of saliva, human colostrum and 3′-sialyllactose. Methods Human colostrum and salivas were collected from 30 mothers and newborn postpartum. Eighteen hours culture of S. mutans was treated with colostrum or 3′-sialyllactose in three different moments: before, during, and after 24 h from the microbial inoculation. Salivas were also tested in conjunction with colostrum. The assays were realized in sterile 96-well flat-bottom microtiter plates for 24 h. The biofilms were fixed, washed, stained with crystal violet, and extracted. Absorbance was measured to evaluate biofilm growth mass. Results Colostrum applied after and during the inoculation decreased biofilm formation when compared with the control (p < .05). The presence of saliva increased the biofilm biomass (p < .05). The application of 3′-sialyllactose reduced biofilm formation independently of moments of application (p < .05). Conclusion Saliva contributed to the proliferation of biofilm and colostrum did not prevent the initial adhesion, but interfered in the accumulation and development of microorganisms in biofilms. 3′-sialyllactose significantly decreased biofilm formation. This information expands the importance of colostrum as a potent oral antimicrobial biofluid.

Comparative Evaluation of Combinatory Interaction between Endocannabinoid System Compounds and Poly-L-lysine against Streptococcus mutans Growth and Biofilm Formation.

Endocannabinoid/endocannabinoid-like (EC/EC-like) are natural endogenous compounds which have been found to affect MRSA pathogenicity. Our previous studies showed that EC/EC-like was able to impair staphylococcal biofilm formation and maintenance as well as to alter biofilm-associated virulence factors. In the present study, we investigated the combinatory effect of the selected EC/EC-like with a natural antimicrobial agent, poly-L-lysine, on cariogenic bacteria Streptococcus mutans growth and biofilm formation. Among four tested EC/EC-like, only two, anandamide (AEA) and oleoylethanolamide (OEA), exhibited synergistic combinatory effect with poly-L-lysine against S. mutans. We attribute this distinct effect to differences in the fatty acid chain structure of the selected EC/EC-like compounds. Moreover, AEA exerted a specific antibiofilm mode of action against S. mutans by effecting total inhibition of biofilm formation while still allowing bacteria viability. Finally, we postulate that the presence of EC/EC-like and poly-L-lysine could enhance the permeability and efficacy of each other via hydrophobic and electrostatic interactions with the S. mutans membrane. In conclusion, we assume that a combination of endogenous natural compounds such as EC/EC-like and poly-L-lysine may benefit oral hygiene by preventing dental plaque.

A novel non-cytotoxic synthetic peptide, Pug-1, exhibited an antibiofilm effect on Streptococcus mutans adhesion.

The aim of this study was to evaluate the effects of synthetic peptides on the growth and biofilm formation of Streptococcus mutans and their cytotoxicity on a human keratinocyte cell line. Four pomegranate-derived peptides; Pug-1, Pug-2, Pug-3 and Pug-4 were synthesized. Streptococcus mutans (ATCC 25175 and DMST 41283) were used. The antibacterial and anti-adherence activities of the peptides were evaluated. The effect of the peptides on S. mutans glucosyltransferase (Gtf) activity was measured. The peptides' cytotoxicity was evaluated on a HaCaT cell line using an MTT assay. The bactericidal penetration depth of Pug-1 into the biofilm was investigated using confocal laser scanning microscopy (CLSM). Among the four peptides, Pug-1 (200µgml-1 ) exhibited antibacterial activity on S. mutans (DMST 41283). All peptides demonstrated anti-adherence activity. Streptococcus mutans Gtf activity was significantly inhibited by Pug-1. Interestingly, Pug-1 peptide was not cytotoxic to the HaCaT cell line. Pug-1 (100-200µgml-1 ) exhibited bactericidal penetration at approximately 50-100% of the biofilm depth. Therefore, Pug-1 might be a candidate of anticariogenic agent. SIGNIFICANCE AND IMPACT OF THE STUDY: Our data highlighted that pomegranate-derived antimicrobial peptides (AMPs) inhibit Streptococcus mutans adhesion, virulence-associated genes and enzymes. This in vitro study supports the hypothesis that pomegranate AMPs would be noncytotoxic to human keratinocytes, with a potent anticariogenic effect via an anti-adherence mode-of-action. The present study provides evidence for further investigation of these AMPs as therapeutic and preventive agents for dental caries.

Use of d-galactose to regulate biofilm growth of oral streptococci

In the oral microbial community, commensals can compete with pathogens and reduce their colonization in the oral cavity. A substance that can inhibit harmful bacteria and enrich beneficial bacteria is required to maintain oral health. The purpose of this study was to examine the effect of d-galactose on the biofilm formation of the cariogenic bacteria Streptococcus mutans and oral commensal streptococci and to evaluate their use in solution and in paste form. Biofilms of S. mutans, Streptococcus oralis, and Streptococcus mitis were formed on saliva-coated glass slips in the absence or presence of d-galactose and evaluated by staining with 1 % crystal violet. d-Galactose significantly inhibited the biofilm formation of S. mutans at concentrations ranging from 2 μM to 200 mM but increased the biofilm formation of S. oralis and S. mitis at concentrations of 2–200 mM. d-Galactose significantly inhibited three glucosyltransferase genes, gtfB, gtfC, and gtfD. The effect of d-galactose in the form of solution and paste was evaluated using bovine teeth. Pretreatment with 100 mM d-galactose on bovine teeth resulted in significantly reduced S. mutans biofilm formation. Our results suggest that d-galactose can be a candidate substance for the development of oral hygiene products to prevent caries by inhibiting the biofilm formation of S. mutans and simultaneously increasing the biofilm formation of commensal oral streptococci.

Quantum Dots of Tantalum Oxide with an Imidazolium Ionic Liquid as Antibacterial Agent for Adhesive Resin.

To synthesize tantalum oxide quantum dots (Ta2O5QDs) using an imidazolium ionic liquid as a precursor and evaluate the effect of its addition to an experimental adhesive resin on the degree of conversion (DC) and antibacterial activity. Ta2O5QDs was synthesized from the hydrolysis of an imidazolium ionic liquid (1-n-decyl-3-methylimidazolium hexachlorotantalate [DMI.TaCl6]) and evaluated by transmission electron microscopy (TEM). The adhesive was formulated with 66.7 wt% bisphenol A-glycidyl methacrylate (bis-GMA), 33.3 wt% 2-hydroxyethyl methacrylate (HEMA) with a photoinitiator/co-initiator system. In one group, Ta2O5QDS was added to the adhesive at 1 wt% Ta2O5QDs) and one group remained without Ta2O5QDS as control (CTRL). The adhesives were evaluated for DC by Fourier Transform Infrared spectroscopy. Direct contact inhibition assay was used to evaluate the antibacterial activity of the experimental adhesive resins against biofilm formation and planktonic bacteria. Ta2O5QDs had a particle size distribution of 1.52 ± 0.73 nm. DC was 50.91 ± 4.94% for CTRL and 55.4 ± 4.58% for Ta2O5QDs (p = 0.310). Ta2O5QDs showed less Streptococcus mutans biofilm formation on adhesive surfaces (p = 0.013). There was no statistically significant difference in terms of antibacterial activity against planktonic bacteria (p = 0.079). Non-agglomerated Ta2O5QDs synthesized from an imidazolium ionic liquid provided antibacterial activity to the experimental adhesive resin against biofilm formation of Streptococcus mutans.

Inhibitory activity of lupinifolin isolated from Derris reticulata stem against biofilm formation of Streptococcus mutans and Staphylococcus aureus

Background: Biofilm formation activity of pathogenic bacteria plays an important role in the pathogenesis and progression of various diseases caused by bacterial infections. It has been reported that lupinifolin, a major phytochemical isolated from Derris reticulata stem, possesses an antibacterial activity against Streptococcus mutans and Staphylococcus aureus. Nonetheless, its actions on biofilm formation properties of S. mutans and S. aureus have not been clearly established. ?Objectives: This study aimed to investigate the antibacterial and antibiofilm formation activities of lupinifolin derived from D. reticulata stem against S. mutans and S. aureus. Subjects and Methods: The minimum inhibitory concentration (MIC) was evaluated using the microbroth dilution method. The antibiofilm formation activity of lupinifolin was conducted at various incubation periods using the crystal violet biofilm formation assay. Results: The MICs of lupinifolin against S. mutans and S. aureus were 4 and 8 μg/mL, respectively. Lupinifolin at the concentrations of sub-MICs had significant inhibitory actions against both sucrose-dependent and sucrose-independent biofilm formations of S. mutans. The lowest median inhibitory concentrations (IC50s) were found at the incubation periods of 12 h (0.57 ± 0.08 μg/mL) and 20 h (0.21 ± 0.04 μg/mL) against sucrose-dependent and sucrose-independent S. mutans biofilm formations, respectively. In addition, at its sub-MICs, lupinifolin also produced a significant inhibition against S. aureus biofilm formation with the lowest IC50 of 0.22 ± 0.03 μg/mL observed at 6-h incubation. Conclusion: These results evidently indicated that lupinifolin can potentially be developed further as a natural product-derived antibiofilm-forming agent for the prevention and/or treatment of biofilm-associated bacterial infections.

In vitro evaluation of milk-based, soy-based, and amino acid-based infant formulas on Streptococcus mutans biofilm formation.

Infant formulas are based on milk, and the addition of simple carbohydrates as a caloric source, for infants. The carbohydrates added in infant formulas can cause a significant increase of Streptococcus mutans in the oral cavity of infants adding to their cariogenicity. The aim of the study was to assess and compare the biofilm formation in three commercially available infant formulas;which are based on milk, soy and amino acid. In vitro microbiological assay of Streptococcus mutans biofilm formation in milk based, soy based and amino acid based infant formulas. Twenty-four hour-cultured S. mutans and microtiter plates were used for analysis. At microtiter plate, 190 μL of modified TSB broth containing SBF, MBF, amino acid-based infant formulas, and dairy whitener as a positive control in five dilutions (1:05, 1:10, 1:20, 1:40, and 1:80) was added into respective wells. 10 μL of cultured S. mutans was inoculated into the wells and incubated at 37°C for 24 h. Biofilm was washed, fixed, and stained with crystal violet. The absorbance was measured to evaluate biofilm growth, which was read as optical densities in a spectrophotometer at 490 nm and was tabulated. Three infant formulas tested showed S. mutans biofilm growth. Minimal biofilm growth was observed in amino acid-based formula at 1:80 dilution, followed by MBF at 1:10 dilution and SBF at 1:80 dilution. Commercially available infant formulas favor S. mutans biofilm growth and can be cariogenic. Amino acid-based infant formula was found to have less S. mutans biofilm growth than MBF and SBF.

Effect of surface characteristic of different restorative materials containing glass ionomer on Streptococcus mutans biofilm.

The aims of this study were to evaluate the surface morphology and surface roughness of restorative materials containing glass ionomer, analyze Streptococcus mutans biofilm formation on the surface of materials, and determine the correlation between surface roughness and biofilm. Four restorative materials: resin-modified glass ionomer; giomer; amalgomer; and glass carbomer were used and for each material, 6 mm in diameter and 2 mm in thickness disc-shaped specimens were prepared to evaluate the surface morphology (n = 3), surface roughness (n = 16), and biofilm (n = 20). Surface morphology was analyzed with a scanning electron microscope. Surface roughness was evaluated via an atomic force microscope. The biofilm was evaluated by counting the colony-forming units. Surface roughness measurements were evaluated using a one-way analysis of variance and Tukey HSD test. Biofilm parameters were analyzed using the Kruskal-Wallis H and Mann-Whitney U test. Pearson's correlation test was used to determine the correlation between surface roughness and biofilm. While the highest roughness values were obtained for amalgomer and glass carbomer, the lowest roughness values belonged to giomer and resin-modified glass ionomer. Statistically significant differences in the number of adherent bacteria were observed between the materials only on day 1. No statistically significant correlation was determined between surface roughness and biofilm. The resin content and small filler particle size of material positively affect surface roughness. However, there is no direct relationship between surface roughness and biofilm.

Constructing an Antibiofouling and Mineralizing Bioactive Tooth Surface to Protect against Decay and Promote Self-Healing.

Numerous methods have been investigated to manage dental caries, one of the top three diseases threatening human health as reported by the World Health Organization. An innovative strategy was proposed to prevent dental caries and achieve self-healing of the decayed tooth, and a novel bioactive peptide was designed and synthesized to construct an antibiofouling and mineralizing dual-bioactive tooth surface. Compared to its original endogenous peptide, the synthesized bioactive peptide showed statistically significantly higher binding affinity to the tooth surface, stronger suppression of demineralization, and a certain promotion of tooth remineralization. The abilities of the peptide to inhibit Streptococcus mutans (S. mutans) biofilm formation and S. mutans adhesion on the tooth surface were not affected after synthesis. Biocompatibility tests revealed the safety of the synthesized bioactive peptide. Interaction mechanisms between the synthesized bioactive peptide and tooth surface were also explained by molecular dynamic simulation analysis. In summary, the synthesized bioactive peptide could be applied safely to prevent dental caries and effectively induce in situ self-healing remineralization for treatment of the decayed tooth.

The inhibitory effects of polypyrrole on the biofilm formation of Streptococcus mutans.

Streptococcus mutans primary thrives on the biofilm formation on the tooth surface in sticky biofilms and under certain conditions can lead to carious lesions on the tooth surface. To search for a new preventive material for oral biofilm-associated diseases, including dental caries, we investigated the effects of polypyrrole, which contains an electrochemical polymer and causes protonation and incorporation of anion under low pH condition, on the biofilm formation of S. mutans and other streptococci. In this study, polypyrrole was applied in biofilm formation assays with the S. mutans strains UA159 and its gtfB and gtfC double mutant (gtfBC mutant), S. sanguinis, S. mitis and S. gordonii on human saliva and bovine serum albumin-coated 96-well microtiter plates in tryptic soy broth supplemented with 0.25% sucrose. The effects of polypyrrole on biofilm formation were quantitatively and qualitatively observed. High concentrations of polypyrrole significantly inhibited the biofilm formation of S. mutans UA159 and S. sanguinis. As an inhibition mechanism, polypyrrole attached to the surface of bacterial cells, increased chains and aggregates, and incorporated proteins involving GTF-I and GTF-SI produced by S. mutans. In contrast, the biofilm formation of gtfBC mutant, S. sanguinis, S. mitis and S. gordonii was temporarily induced by the addition of low polypyrrole concentrations on human saliva-coated plate but not on the uncoated and bovine serum albumin-coated plates. Moreover, biofilm formation depended on live cells and, likewise, specific interaction between cells and binding components in saliva. However, these biofilms were easily removed by increased frequency of water washing. In this regard, the physical and electrochemical properties in polypyrrole worked effectively in the removal of streptococci biofilms. Polypyrrole may have the potential to alter the development of biofilms associated with dental diseases.

Rechargeable anti-microbial adhesive formulation containing cetylpyridinium chloride montmorillonite

Long-term anti-bacterial effect is a desired ability of any dental material in combating tooth caries as one of the most common and widespread persistent diseases today. Among several cationic quaternary ammonium compounds with antiseptic properties, cetylpyridinium chloride (CPC) is often used in mouthrinses and toothpastes. In this study, we incorporated CPC in a soft phyllosilicate mineral (clay), referred to as montmorillonite (Mont), to enable gradual CPC release with rechargeability. Besides measuring CPC release and recharge, we examined the anti-bacterial effect, cytotoxicity and bonding effectiveness of five experimental adhesive formulations, prepared by adding 1 and 3 wt% CPC_Mont, 3 wt% Mont (without CPC), and 1 and 3 wt% CPC (without Mont) to the commercial adhesive Clearfil S3 Bond ND Quick (‘C-S3B’; Kuraray Noritake). Strong inhibition of Streptococcus mutans biofilm formation by CPC_Mont adhesives was confirmed by optical density and SEM. CPC release from CPC_Mont adhesives was higher and lasted longer than from CPC adhesives, while CPC_Mont adhesives could also be recharged with CPC upon immersion in 2 wt% CPC. In conclusion, CPC_Mont technology rendered adhesives anti-bacterial properties with recharge ability, this without reducing its bonding potential, neither increasing its cytotoxicity. Statement of significanceDental caries is one of the most prevalent chronic diseases in the population worldwide and is the major cause of tooth loss. In this study, we developed cetylpyridinium chloride (CPC) loaded montmorillonite (CPC-Mont) with a long-term antibacterial efficacy to prevent caries. CPC is an antibacterial agent approved by FDA, used as an OTC drug and contained in oral hygiene aids. CPC-Mont was incorporated in a dental adhesive to gradually release CPC. CPC_Mont technology rendered adhesives anti-bacterial properties with rechargeability, this without reducing its bonding potential, neither increasing its cytotoxicity.

Effect of different polishing techniques for composite resin materials on surface properties and bacterial biofilm formation

ObjectivesBoth direct and indirect techniques are used for composite resin material (CRM) restorations. Polishing processes are needed in both techniques after intraoral adjustment. However, it is unclear as to which polishing technique should be preferred with respect to decreasing biofilm. The purpose of thisin vitro study was to evaluate the surface properties and Streptococcus mutans biofilm formation on direct and indirect CRMs after using different polishing techniques. MethodsTwo CRMs (direct and indirect) and four polishing techniques (aluminium oxide discs, diamond polishing paste, aluminium oxide polishing paste, and silicon carbide brush) were evaluated. The specimens were prepared for taking scanning electron microscopy images (n = 2) and determining surface roughness, surface free energy, and bacterial biofilm formation (BBF) with colony-forming unit counting and confocal laser scanning microscopy assays (n = 7). The data were analysed using two-way analysis of variance with Bonferroni as a post hoc test and Pearson’s correlation (p < .05). ResultsThe surface roughness values in the control group were higher than those in the diamond polishing paste group (p = 0.025), but the values in the aluminium oxide polishing paste and silicon carbide brush groups were comparable with those in the control group (p = 0.156 and p = 1.000, respectively). The highest surface free energy values were recorded in the silicon carbide brush group (p < 0.001), whereas there were no differences found among the other groups (p > 0.05). The highest BBF was seen in the silicon carbide brush (p < 0.001) and direct CRM (p < 0.001) groups. ConclusionBBF on the surface of direct CRMs differed from that on indirect CRMs after polishing the surface. The tested polishing techniques significantly influenced surface properties and BBF. Clinical significanceIn situations that require the intraoral adjustment of CRMs, polishing with a diamond polishing paste seems to be a good option to polish the surface of both direct and indirect CRMs because the diamond polishing paste results better in terms of decreasing biofilm formation and improving surface properties.

Evaluation of Alternative Methods to Assess the Biological Properties of Propolis on Metabolic Activity and Biofilm Formation in Streptococcus mutans.

Several biological activities have been reported for the Chilean propolis, among their antimicrobial and antibiofilm properties, due to its high polyphenol content. In this study, we evaluate alternative methods to assess the effect of Chilean propolis on biofilm formation and metabolic activity of Streptococcus mutans (S. mutans), a major cariogenic agent in oral cavity. Biofilm formation was studied by using crystal violet and by confocal microscopy. The metabolic activity of biofilm was evaluated by MTT and by flow cytometry analysis. The results show that propolis reduces biofilm formation and biofilm metabolic activity in S. mutans. When the variability of the methods to measure biofilm formation was compared, the coefficient of variation (CV) fluctuated between 12.8 and 23.1% when using crystal violet methodology. On the other hand, the CV ranged between 2.2 and 3.3% with confocal microscopy analysis. The CV for biofilm's metabolic activity measured by MTT methodology ranged between 5.0 and 11.6%, in comparison with 1.9 to 3.2% when flow cytometry analysis was used. Besides, it is possible to conclude that the methods based on colored compounds presented lower precision to study the effect of propolis on biofilm properties. Therefore, we recommend the use of flow cytometry and confocal microscopy in S. mutans biofilm analysis.

Sortase A-mediated modification of the Streptococcus mutans transcriptome and virulence traits.

Sortase A contributes to adhesion and biofilm formation of Streptococcus mutans by anchoring surface proteins like P1 onto the cell wall, and few other functional characterization has been annotated to this protein and its coding gene srtA. In this study we investigated that whether srtA deletion would affect S. mutans virulence determinants in addition to adhesion and further explored whether these effects were caused due to changes in S. mutans genomic transcription. We used acid-killing assays, glycolytic rate assessments, and exopolysaccharide (EPS) formation tests to detect whether srtA deletion influenced S. mutans acid tolerance/production and glucan formation. Comparisons between RNA-sequencing data from both the exponential and stationary phases of UA159 and the srtA-deleted strain were made to determine the impact of srtA knockout on S. mutans genomic transcription. Results of our assays indicated that S. mutans aciduricity was enhanced in the srtA deleted strain when bacterial cells were directly subjected to pH 2.8, but the enhancement was repressed when the acid tolerance response was induced in advance. The srtA mutation strain exhibited reduced EPS formation in mature biofilms. SrtA deletion led to pleiotropic changes in the S. mutans transcriptome with a growth phase-dependent pattern. The affected genes mainly included those involved in aciduricity, carbohydrate transport, and EPS formation. It was concluded that S. mutans srtA exhibited multiple effects on the virulence traits of this pathogen, including acid tolerance and glucan formation, and that these alterations could be partially due to transcriptional changes upon loss of srtA.

Theaflavin-3,3'-Digallate Suppresses Biofilm Formation, Acid Production, and Acid Tolerance in Streptococcus mutans by Targeting Virulence Factors.

As one of the most important cariogenic pathogens, Streptococcus mutans has strong abilities to form biofilms, produce acid and tolerate acid. In present study, we found that theaflavin-3,3′-digallate (TF3) had an inhibitory effect on S. mutans UA159 in vitro. Visualized by field emission-scanning electron microscopy, the suppressed formation of S. mutans biofilms grown with TF3 at sub-inhibitory concentrations could be attributed to the reduced biofilm matrix, which was proven to contain glucans and extracellular DNA (eDNA). Glucan-reduced effect of TF3 was achieved by down-regulating expression levels of gtfB, gtfC, and gtfD encoding glucosyltransferases. Besides, TF3 reduced eDNA formation of S. mutans by negatively regulating lrgA, lrgB, and srtA, which govern cell autolysis and membrane vesicle components. Furthermore, TF3 also played vital roles in antagonizing preformed biofilms of S. mutans. Bactericidal effects of TF3 became significant when its concentrations increased more than twofold of minimum inhibitory concentration (MIC). Moreover, the capacities of S. mutans biofilms to produce acid and tolerate acid were significantly weakened by TF3 at MIC. Based on real-time PCR (RT-PCR) analysis, the mechanistic effects of TF3 were speculated to comprise the inhibition of enolase, lactate dehydrogenase, F-type ATPase and the agmatine deiminase system. Moreover, TF3 has been found to downregulate LytST, VicRK, and ComDE two component systems in S. mutans, which play critical roles in the regulatory network of virulence factors. Our present study found that TF3 could suppress the formation and cariogenic capacities of S. mutans biofilms, which will provide new strategies for anti-caries in the future.

Experimental composites containing quaternary ammonium methacrylates reduce demineralization at enamel-restoration margins after cariogenic challenge

ObjectiveThis study evaluated the influence of experimental composites containing quaternary ammonium monomers (QAM) at different concentrations and alkyl chains on demineralization at enamel-composite margins after cariogenic challenge. MethodsStandardized 4×4mm cavities were cut into 35 bovine enamel blocks, which were randomly divided into seven groups (n=5) and restored with the following experimental composites and commercial materials: (G12.5) – 5% dimethylaminododecyl methacrylate (DMADDM) with a 12-carbon alkyl chain (G12.10) – 10% DMADDM, (G16.5) – 5% dimethylaminohexadecyl methacrylate (DMAHDM) with a 16-carbon alkyl chain (G16.10) – 10% DMAHDM, (CG) – control group (without QAM), (GZ250) – commercial composite (Filtek Z250®), and (GIC) – glass ionomer cement (Maxxion R®). After restorative procedures, initial microhardness was measured and experimental composites were subjected to Streptococcus mutans biofilm formation for 48h. After cariogenic challenge, the samples were washed and microhardness was reassessed. A 3D non-contact profilometer was used to determine surface roughness and enamel demineralization was assessed by micro-CT. Microhardness results were analyzed by the Kruskal–Wallis and Mann-Whitney tests and micro-CT results were analyzed by Tukey’s HSD test (95% confidence interval). ResultsNone of the materials could prevent mineral loss at the enamel-restoration margins. The addition of 10% DMAHDM yielded the lowest, albeit statistically significant, mineral loss (p<0.05). 3D non-contact profilometry showed enamel surface roughness modification after biofilm exposure. The CG had the highest roughness values. Micro-CT analysis revealed mineral loss, except for GIC. SignificanceThe addition of 10% QAM with a 16-carbon chain in experimental composites reduced mineral loss at the enamel-restoration margins after cariogenic challenge.

Antimicrobial peptides Pep19–2.5 and Pep19-4LF inhibit Streptococcus mutans growth and biofilm formation

With this study, we investigated the effect of synthetic antimicrobial peptides Pep19–2.5 and Pep194LF alone or in combination with antibiotics on S. mutans growth and biofilm formation/disruption. We also examined the cytotoxic effect of each peptide on monocytes.S. mutans was cultured in the presence of different concentrations of each peptide. We showed that Pep19–2.5 and Pep19-4LF were able to significantly (p ≤ 0.01) inhibit the growth of S. mutans. The synthetic peptides also decreased biofilm formation by S. mutans. Furthermore, both peptides reduced the viability of S. mutans in already formed biofilms. The combination of each peptide with antibiotics (penicillin/streptomycin, P/S) produced additive interactions which inhibited S. mutans growth and biofilm formation. Pep19–2.5 and Pep19-4LF were nontoxic, as they did not decrease monocyte viability and did not increase the lactate dehydrogenase activity of the exposed cells. In conclusion, synthetic peptides Pep19–2.5 and Pep19-4LF did inhibit S. mutans growth and its capacity to form biofilm. Both peptides were found to be nontoxic to monocytes. These data provide new insight into the efficacy of synthetic peptides Pep19–2.5 and Pep19-4LF against S. mutans. These peptides may thus be useful in controlling the adverse effects of this cariogenic bacterium in human.

Rhamnus prinoides (gesho): A source of diverse anti-biofilm activity

Ethnopharmacological relevanceRhamnus prinoides (gesho) is an evergreen shrub from East Africa traditionally used for the treatment of illnesses including atopic dermatitis, ear, nose and throat infections, pneumonia, arthritis, brucellosis, flu, indigestion and fatigue. Aim of the studySeveral of the conditions for which gesho is traditionally used are associated with communities of surface-attached microorganisms, or biofilms. We hypothesized that gesho has anti-biofilm activity. The principal aim of this study was to evaluate gesho-associated anti-biofilm activity and identify active compounds. Materials and methodsLyophilized ethanol and aqueous extracts were prepared from dried Rhamnus prinoides stems and leaves. Biofilm inhibition was measured by crystal violet staining and subsequent viability assays were conducted on growth agar. Chemical fractionation, chemical testing, Fourier transform infrared spectroscopy (FTIR) and gas chromatography-mass spectrometry (GC-MS) were used to isolate and identify active compounds. ResultsLeaf and stem ethanol extracts significantly inhibited Staphylococcus aureus, Bacillus subtilis and Streptococcus mutans biofilm formation up to 99.9% and reduced planktonic cell growth up to 10 log units relative to untreated controls. The anti-biofilm activity of the ethanol stem extracts was due to a biocidal or bacteriostatic mechanism while bacteriostatic or anti-pathogenic mechanisms were attributed to the leaf ethanol extract. Gesho extracts showed activity against all three species tested but the treatment efficacy and mechanism were species dependent. Chemical fractionation and activity screens of the leaf ethanol extract identified ethyl 4-ethoxybenzoate and 4-hydroxy 4-methyl pentanone to be compounds with anti-biofilm activity. Ethyl 4-ethoxybenzoate activity was potentiated by DMSO. Notably, concentrations of both compounds were identified where biofilm formation was prevented without inhibition of cell growth; i.e. anti-pathogenic characteristics were evident. ConclusionGesho leaf ethanol extract contains chemicals with anti-biofilm and bactericidal activities. This work lends support to the traditional use of gesho for treating topical infections and warrants further investigation into Rhamnus prinoides as a source of antibacterial and anti-biofilm agents.

Bioactive Antibacterial Modification of Orthodontic Microimplants Using Chitosan Biopolymer

Titanium alloy (Ti6Al4V)-based microimplants are extensively used as anchorage units in orthodontic treatment; however, they have a high failure rate (10 to 30%). This study aimed to enhance the survivability of these small implants by improving their bioactive and antibacterial properties, via modifying their surface with chitosan. For a stable covalent bond, microimplant surfaces were first functionalized with 3-triethoxysilylpropylamine (APTES) and then grafted either with a succinic acid or polyacrylic acid spacer, followed by attachment with chitosan. Validity tests were conducted on titanium alloy disk (TI) specimens treated with the same chemical process (TI specimens were used to avoid the practical difficulties in testing the small and complex-shaped microimplants). The results confirmed successful APTES functionalization and grafting of succinic- or polyacrylic acid. Anchoring of chitosan was confirmed via X-ray photoelectron spectroscopy (XPS) and confocal laser scanning microscopy (CLSM); furthermore, the anchoring ability of succinic acid, as a spacer, for chitosan was superior to that of polyacrylic acid. Chitosan-modified TI specimens (TI-SA-Ch) showed better biocompatibility with pre-osteoblastic MC3T3-E1 cells, as confirmed from their enhanced adhesion, proliferation, and from cell viability analysis. Biofilm formation of Streptococcus mutans and Streptococcus sobrinus was effectively reduced by 53% and 31%, respectively, on the surface of TI-SA-Ch. These results clearly indicated that chitosan modification might be a promising approach to enhance the bioactive- and antibacterial properties of orthodontic microimplants, and thus, increase their stability in the jaw bone in clinical practice.

Disruption of Streptococcus mutans Biofilm by Rhamnolipid Biosurfactant Secreted from Pseudomonas aeruginosa ATCC10145

Objectives: The study was to examine the effect of rhamnolipid biosurfactant on Streptococcus mutans biofilm formation on orthodontic appliances. Methods/Statistical Analysis: Several properties of S. mutans cells such as formation of biofilm, detachment ability, the surface properties were changed after three different concentrations of biosurfactant treatment which were 10 mg/ml, 5 mg/ml and 2.5 mg/ml. S. mutans biofilms were disrupted by rhamnolipids produced by Pseudomonas aeruginosa ATCC10145 at different concentrations and chlorhexidine digluconate 0.12% (v/v) was used as positive control. Findings: The biosurfactant showed some antimicrobial activities against S. mutans with Minimum Inhibitory Concentration (MIC) varied from 10 mg/ml to 0.01 mg/ml. MIC were observed at 1.25 mg/ml by microdilution 96-well plate method. The Minimum Bactericidal Concentration (MBC) was detected at a higher concentration of 10 mg/ml by plating onto agar. The best treatment of dental biofilm formation was determined at 10 mg/ml showing the highest percentage of biofilm detachment which was 89.53%. The biofilm development is commonly known as strongly dependent on hydrophobicity/hydrophilicity substrate properties using contact angle measurement. Higher percentage (89.53%) of biofilm detachment determined at polystyrene surface compared to stainless steel surface with the aid of rhamnolipid. Rhamnolipid displayed a significant potential as disrupting agents against established biofilms produced by several bacterial and fungal species. Application/Improvements: P. aeruginosa were successfully extracted their crude biosurfactant to treat the growth of biofilm. Future studies can be performed on specific mechanisms on how biosurfactant inhibit and disrupt biofilm growth. Keywords: Biofilm Formation, Biosurfactant, Pseudomonas Aeruginosa, Streptococcus mutans