Antagonist effect of probiotic bifidobacteria on biofilms of pathogens associated with periodontal disease.

The in vitro antagonist growth effect of bifidobacteria were evaluated on periodontal bacteria. Bifidobacterium longum, Bifidobacterium lactis and Bifidobacterium infantis biofilms were grown in single, double or triple combinations with putative periodontal pathogens P. gingivalis and F. nucleatum or beneficial bacteria S. oralis for 24, 72 and 168 h and the total counts were analyzed by checkerboard DNA-DNA hybridization. The results showed that B. infantis and B. lactis, as single species, demonstrated the best antagonist effect on F. nucleatum and P. gingivalis and no influence on S. oralis growth at 168 h. All the double combinations of bifidobacteria tested demonstrated an inhibitory effect on F. nucleatum (72 h) and P. gingivalis (168 h) and did not affect S. oralis counts at any time. In conclusion, B. lactis and B. infantis alone or in double combinations have antagonist effect on periodontopathogens biofilms, at different time points, and minimal influence on S. oralis growth.

Genetic and physiological effects of subinhibitory concentrations of oral antimicrobial agents on Streptococcus mutans biofilms.

Streptococcus mutans is the main etiological agent of dental caries because of its capacity to adhere to enamel structure and form biofilms. This study aimed to evaluate the effects of the anticariogenic agents - sodium fluoride (NaF) and chlorhexidine (CHX) - at levels below minimum inhibitory concentrations (sub-MICs) on the growth of planktonic cells and biofilms and on the expression of vicR and covR genes associated with the regulation of biofilm formation. MICs and minimum bactericidal concentrations (MBCs) of NaF and CHX were determined for S. mutans strains ATCC25175, UA159 and 3VF2. Growth curves were constructed for planktonic cells cultured in brain heart infusion (BHI) broth supplemented with NaF (0.125-0.75MIC) or CHX (0.25-0.75MIC). Biofilm formation assays were performed in microplates containing CHX or NaF at 0.5-1.0MIC and stained with violet crystal. Quantitative polymerase chain reaction determined the alterations in covR and vicR expression in cells exposed to antimicrobials at sub-MIC levels. NaF and CHX at sub-MIC levels affected the growth of planktonic cells of all three S. mutans strains, depending on the concentration tested. The biofilm formation in UA159 and 3VF2 was reduced by NaF at concentrations ≥0.5 MIC, while that of ATCC 25175 was reduced significantly irrespective of dose. In contrast, UA159 and 3VF2 biofilms were not affected by CHX at these levels, whereas those of ATCC 25175 were reduced significantly at all concentrations tested. Under sub-MIC conditions, CHX and (to a lesser degree) NaF increased vicR and covR expression in all three strains, although there were large differences between strains and treatment conditions employed. CHX and NaF at sub-MIC levels influence on the growth of S. mutans in planktonic and biofilm conditions and on transcript levels of biofilm-associated genes vicR and covR, in a dose-dependent manner.

Cytotoxicity and effects of curcumin and cinnamaldehyde hybrids on biofilms of oral pathogens.

The objective of the study was to evaluate the cytotoxicity and effect of curcumin-cinnamaldehyde hybrids (CCHs) on the biofilm of oral pathogens. Of the 18 hybrids tested, nine had an inhibitory effect on at least one of the bacterial species tested, with minimal inhibitory and bactericidal concentrations ranging from 9 to 625 µg ml-1. CCH 7 promoted a potent inhibitory effect against all the bacterial species tested and better compatibility than chlorhexidine (CHX). CCH 7 also presented a similar or improved effect over that of CHX, causing a reduction in bacterial metabolism and viability in single and dual-species biofilms. CCH 7 reduced by 86% and 34% the viability of multispecies biofilms formed by collection and clinical strains. It can be concluded that CCH 7 was cytocompatible at the minimal inhibitory concentration, presented anti-biofilm action against oral pathogens, and could act as an antimicrobial agent for application in endodontics.

KR-12-a5 is a non-cytotoxic agent with potent antimicrobial effects against oral pathogens.

This study evaluated the cytotoxicity and antimicrobial activity of analogs of cationic peptides against microorganisms associated with endodontic infections. L-929 fibroblasts were exposed to LL-37, KR-12-a5 and hBD-3-1CV and chlorhexidine (CHX, control), and cell metabolism was evaluated with MTT. The minimal inhibitory concentration (MIC) and the minimal bactericidal/fungicidal concentration (MBC/MFC) of the peptides and CHX were determined against oral pathogens associated with endodontic infections. Enterococcus faecalis and Streptococcus mutans biofilms were cultivated in bovine dentin blocks, exposed to different concentrations of the most efficient antimicrobial peptide and analyzed by confocal laser scanning microscopy. CHX and peptides affected the metabolism of L-929 at concentrations > 31.25 and 500 µg ml-1, respectively. Among the peptides, KR-12-a5 inhibited growth of both the microorganisms tested with the lowest MIC/MBC/MFC values. In addition, KR-12-a5 significantly reduced E. faecalis and S. mutans biofilms inside dentin tubules. In conclusion, KR-12-a5 is a non-cytotoxic agent with potent antimicrobial and anti-biofilm activity against oral pathogens associated with endodontic infections.

Genotypic diversity and phenotypic traits of Streptococcus mutans isolates and their relation to severity of early childhood caries.

BACKGROUND: Early childhood caries (ECC) is an aggressive condition that can affect teeth of young children. This study aimed to evaluate genotypic diversity and phenotypic traits of S. mutans isolated from dental biofilms of children with different caries status in comparison with caries free (CF) children. METHODS: Streptococcus mutans strains were isolated from supragingival biofilm samples of CF, ECC and severe-ECC (S-ECC) children and genotyped by arbitrary-primer polymerase chain reaction - AP-PCR. S. mutans genotypes were tested for their ability to reduce the suspension pH through glycolysis, to tolerate extreme acid challenge and by their ability to form biofilm. Response variables were analyzed by ANOVA/Tukey or Kruskal-Wallis/Mann-Whitney tests at a 5% of significance. RESULTS: There was an increase in the prevalence of Streptococcus mutans in biofilms with the severity of dental caries. No differences in genotypic diversity and in acidogenicity of genotypes were found among CF, ECC and S-ECC children. S mutans strains with genotypes more characteristic for ECC and S-ECC children formed more biofilms than those identified in CF children. The strains isolated from S-ECC children were highly acid tolerant. CONCLUSION: Although S. mutans genotypic diversity was similar among the groups of children, phenotypic traits of S. mutans, especially the acid tolerance response, could explain the severity of early childhood caries.

Cytotoxicity and the effect of cationic peptide fragments against cariogenic bacteria under planktonic and biofilm conditions.

This study evaluated the cytotoxicity and effect of fragments derived from three oral cationic peptides (CP): LL-37, D6-17 and D1-23 against cariogenic bacteria under planktonic and biofilm conditions. For cytotoxicity analysis, two epithelial cell lines were used. The minimum inhibitory concentration and the minimal bactericidal concentration were determined for the CP fragments and the control (chlorhexidine-CHX) against cariogenic bacteria. The fractional inhibitory concentration was obtained for the combinations of CP fragments on Streptococcus mutans. Biofilm assays were conducted with the best antimicrobial CP fragment against S. mutans. The results indicated that D6-17 was not cytotoxic. D1-23, LL-37 and CHX were not cytotoxic in low concentrations. D1-23 presented the best bactericidal activity against S. mutans, S. mitis and S. salivarius. Combinations of CP fragments did not show a synergic effect. D1-23 presented a higher activity against S. mutans biofilm than CHX. It was concluded that D1-23 showed a substantial effect against cariogenic bacteria and low cytotoxicity.

Antimicrobial activity of conventional and plant-extract disinfectant solutions on microbial biofilms on a maxillofacial polymer surface.

STATEMENT OF PROBLEM: Dentists often note problems with infection in patients with maxillofacial prostheses. Conventional disinfection protocols are not always effective and may alter the properties of the polymer used in the prosthesis. Thus, the search for improved disinfection methods is important. PURPOSE: The purpose of this in vitro study was to evaluate and compare the antimicrobial activity of conventional disinfectant solutions (water and neutral soap and 4% chlorhexidine) and plant extracts (Cymbopogon nardus and Hydrastis canadensis) on specimens of maxillofacial silicone contaminated with Candida albicans and Staphylococcus aureus biofilms. MATERIAL AND METHODS: Seventy-two silicone (MDX4-4210) specimens were fabricated (5×2 mm) and sterilized. Thirty-six were contaminated with C albicans (10(6) cells/mL) and 36 with S aureus (10(8) cells/mL) to evaluate the antimicrobial activity of the cleaning protocols. After incubation (37°C/72 hours), the specimens were divided into 5 groups: not disinfected (positive control), soaking in saline solution for 10 minutes, soaking in 4% chlorhexidine for 10 minutes, soaking in C nardus for 10 minutes, soaking in H canadensis for 10 minutes, and washing by hand with water and neutral soap for 30 seconds. The viability of cells was evaluated by XTT (2,3-bis-(2-methoxy-4-nitro-5-sulfophenyl)-2H-tetrazolium-5-carboxanilide) assay and by scanning electron microscope analysis. The results were analyzed by ANOVA and the Tukey HSD test (α=.05). RESULTS: All disinfection solutions provided a statistically significant reduction in biofilm viability compared with the control group for both microorganisms (P<.05). Washing with water and neutral soap was significantly more effective in reducing biofilm viability than immersion in the disinfection solutions, with persistence of viable microorganisms between 1.05% for C albicans and 0.62% for S aureus after this cleaning protocol. Photomicrographs revealed that 4% chlorhexidine altered the surface of the polymer. CONCLUSIONS: Within the limitations of this in vitro study, it was concluded that the cleaning protocols with different disinfectant solutions produced a significant reduction in the viability of C albicans and S aureus biofilms on the silicone polymer. Washing with water and neutral soap was the most effective protocol against both microorganisms.

Synergistic antimicrobial potential of EGCG and fosfomycin against biofilms associated with endodontic infections.

OBJECTIVE: This study aimed to evaluate the cytotoxicity and synergistic effect of epigallocatechin gallate (EGCG) and fosfomycin (FOSFO) on biofilms of oral bacteria associated with endodontic infections. METHODOLOGY: This study determined minimum inhibitory and bactericidal concentration (MIC/MBC) and fractionated inhibitory concentration (FIC) of EGCG and FOSFO against Enterococcus faecalis, Actinomyces israelii, Streptococcus mutans, and Fusobacterium nucleatum. Monospecies and multispecies biofilms with those bacteria formed in polystyrene microplates and in radicular dentin blocks of bovine teeth were treated with the compounds and control chlorhexidine (CHX) and evaluated by bacterial counts and microscopy analysis. Toxicity effect of the compounds was determined on fibroblasts culture by methyl tetrazolium assays. RESULTS: The combination of EGCG + FOSFO demonstrated synergism against all bacterial species, with an FIC index ranging from 0.35 to 0.5. At the MIC/FIC concentrations, EGCG, FOSFO, and EGCG+FOSFO were not toxic to fibroblasts. EGCG+FOSFO significantly reduced monospecies biofilms of E. faecalis and A. israelli, whereas S. mutans and F. nucleatum biofilms were eliminated by all compounds. Scanning electron microscopy of multispecies biofilms treated with EGCG, EGCG+FOSFO, and CHX at 100x MIC showed evident biofilm disorganization and substantial reduction of extracellular matrix. Confocal microscopy observed a significant reduction of multispecies biofilms formed in dentin tubules with 84.85%, 78.49%, and 50.6% of dead cells for EGCG+FOSFO, EGCG, and CHX at 100x MIC, respectively. CONCLUSION: EGCG and fosfomycin showed a synergistic effect against biofilms of oral pathogens related to root canal infections without causing cytotoxicity.

Cytotoxicity and Biomineralization Potential of Flavonoids Incorporated into PNVCL Hydrogels.

This study aimed to evaluate the effects of flavonoids incorporated into poly(N-vinylcaprolactam) (PNVCL) hydrogel on cell viability and mineralization markers of odontoblast-like cells. MDPC-23 cells were exposed to ampelopsin (AMP), isoquercitrin (ISO), rutin (RUT) and control calcium hydroxide (CH) for evaluation of cell viability, total protein (TP) production, alkaline phosphatase (ALP) activity and mineralized nodule deposition by colorimetric assays. Based on an initial screening, AMP and CH were loaded into PNVCL hydrogels and had their cytotoxicity and effect on mineralization markers determined. Cell viability was above 70% when MDPC-23 cells were treated with AMP, ISO and RUT. AMP showed the highest ALP activity and mineralized nodule deposition. Extracts of PNVCL+AMP and PNVCL+CH in culture medium (at the dilutions of 1/16 and 1/32) did not affect cell viability and stimulated ALP activity and mineralized nodules' deposition, which were statistically higher than the control in osteogenic medium. In conclusion, AMP and AMP-loaded PNVCL hydrogels were cytocompatible and able to induce bio-mineralization markers in odontoblast-cells.

Understanding the Predictive Potential of the Oral Microbiome in the Development and Progression of Early Childhood Caries.

BACKGROUND: Early childhood caries (ECC) is the most common chronic disease in young children and a public health problem worldwide. It is characterized by the presence of atypical and fast progressive caries lesions. The aggressive form of ECC, severe early childhood caries (S-ECC), can lead to the destruction of the whole crown of most of the deciduous teeth and cause pain and sepsis, affecting the child's quality of life. Although the multifactorial etiology of ECC is known, including social, environmental, behavioral, and genetic determinants, there is a consensus that this disease is driven by an imbalance between the oral microbiome and host, or dysbiosis, mediated by high sugar consumption and poor oral hygiene. Knowledge of the microbiome in healthy and caries status is crucial for risk monitoring, prevention, and development of therapies to revert dysbiosis and restore oral health. Molecular biology tools, including next-generation sequencing methods and proteomic approaches, have led to the discovery of new species and microbial biomarkers that could reveal potential risk profiles for the development of ECC and new targets for anti-caries therapies. This narrative review summarized some general aspects of ECC, such as definition, epidemiology, and etiology, the influence of oral microbiota in the development and progression of ECC based on the current evidence from genomics, transcriptomic, proteomic, and metabolomic studies and the effect of antimicrobial intervention on oral microbiota associated with ECC. CONCLUSION: The evaluation of genetic and proteomic markers represents a promising approach to predict the risk of ECC before its clinical manifestation and plan efficient therapeutic interventions for ECC in its initial stages, avoiding irreversible dental cavitation.

Microbiological Properties and Cytotoxicity of PNVCL Hydrogels Containing Flavonoids as Intracanal Medication for Endodontic Therapy.

This study aimed to evaluate the cytotoxicity and microbiological properties of poly (N-vinylcaprolactam)-PNVCL hydrogels containing flavonoids as intracanal medication for endodontic therapy. Antimicrobial activity of ampelopsin (AMP), isoquercitrin and rutin was determined against Enterococcus faecalis, Actinomyces israelii, Lactobacillus casei, Streptococcus mutans, and Fusobacterium nucleatum by the microdilution method. After synthesis and characterization by rheology, PNVCL hydrogels were loaded with AMP and controls calcium hydroxide (CH) and chlorhexidine (CHX), and determined the compounds release profile. PNVCL+AMP, PNVCL+CH, PNVCL+CHX were evaluated on multi-species biofilms and analyzed by Scanning Electron Microscopy (SEM) and Confocal Laser Scanning Microscopy (CLSM). Cytotoxicity was determined after fibroblasts exposure to serial dilutions of AMP and PNVCL hydrogel extracts. AMP was effective against all of the bacteria tested, especially against S. mutans, A. israelli and F. nucleatum. SEM and CLSM analysis showed that PNVCL + AMP caused a significant decrease and disorganization of multi-species biofilms and reduction of intracanal viable cells, superior to the other groups. AMP affected fibroblast viability at concentrations above 0.125 mg/mL, and extracts of PNVCL+AMP showed low cytotoxicity. In conclusion, PNVCL containing AMP demonstrated cytocompatibility and potent effect against multi-species biofilms and could be potential intracanal medication for endodontic purposes.

Interplay Among the Oral Microbiome, Oral Cavity Conditions, the Host Immune Response, Diabetes Mellitus, and Its Associated-Risk Factors-An Overview.

This comprehensive review of the literature aimed to investigate the interplay between the oral microbiome, oral cavity conditions, and host immune response in Diabetes mellitus (DM). Moreover, this review also aimed to investigate how DM related risk factors, such as advanced age, hyperglycemia, hyperlipidemia, obesity, hypertension and polycystic ovary syndrome (PCOS), act in promoting or modifying specific mechanisms that could potentially perpetuate both altered systemic and oral conditions. We found that poorly controlled glycemic index may exert a negative effect on the immune system of affected individuals, leading to a deficient immune response or to an exacerbation of the inflammatory response exacerbating DM-related complications. Hyperglycemia induces alterations in the oral microbiome since poor glycemic control is associated with increased levels and frequencies of periodontal pathogens in the subgingival biofilm of individuals with DM. A bidirectional relationship between periodontal diseases and DM has been suggested: DM patients may have an exaggerated inflammatory response, poor repair and bone resorption that aggravates periodontal disease whereas the increased levels of systemic pro-inflammatory mediators found in individuals affected with periodontal disease exacerbates insulin resistance. SARS-CoV-2 infection may represent an aggravating factor for individuals with DM. Individuals with DM tend to have low salivary flow and a high prevalence of xerostomia, but the association between prevalence/experience of dental caries and DM is still unclear. DM has also been associated to the development of lesions in the oral mucosa, especially potentially malignant ones and those associated with fungal infections. Obesity plays an important role in the induction and progression of DM. Co-affected obese and DM individuals tend to present worse oral health conditions. A decrease in HDL and, an increase in triglycerides bloodstream levels seem to be associated with an increase on the load of periodontopathogens on oral cavity. Moreover, DM may increase the likelihood of halitosis. Prevalence of impaired taste perception and impaired smell recognition tend to be greater in DM patients. An important interplay among oral cavity microbiome, DM, obesity and hypertension has been proposed as the reduction of nitrate into nitrite, in addition to contribute to lowering of blood pressure, reduces oxidative stress and increases insulin secretion, being these effects desirable for the control of obesity and DM. Women with PCOS tend to present a distinct oral microbial composition and an elevated systemic response to selective members of this microbial community, but the association between oral microbiome, PCOS are DM is still unknown. The results of the studies presented in this review suggest the interplay among the oral microbiome, oral cavity conditions, host immune response and DM and some of the DM associated risk factors exist. DM individuals need to be encouraged and motivated for an adequate oral health care. In addition, these results show the importance of adopting multidisciplinary management of DM and of strengthening physicians-dentists relationship focusing on both systemic and on oral cavity conditions of DM patients.

Cytocompatibility and Synergy of EGCG and Cationic Peptides Against Bacteria Related to Endodontic Infections, in Planktonic and Biofilm Conditions.

This study evaluated the cytocompatibility and antimicrobial/antibiofilm effects of epigallocatechin-3-gallate (EGCG) associated with peptide LL-37 and its analogue KR-12-a5 against oral pathogens. The effect of the compounds on metabolism of fibroblasts was evaluated by methyltetrazolium assays. Antimicrobial activity of the compounds was evaluated on Streptococcus mutans, Enterococcus faecalis, Actinomyces israelii, and Fusobacterium nucleatum under planktonic conditions, on single- and dual-species biofilms and E. faecalis biofilms in dentinal tubules and analyzed by bacterial counts and confocal microscopy. Data were statistically analyzed considering p < 0.05. EGCG and peptide combinations were not toxic to fibroblasts. KR-12-a5 showed synergistic or addictive effects with EGCG and LL-37 against all bacteria tested. However, EGCG associated with KR-12-a5 demonstrated the highest bactericidal activity on all bacteria tested, at lower concentrations. In single-species biofilms, EGCG + KR-12-a5 eliminated S. mutans and A. israelii and reduced E. faecalis and F. nucleatum counts around 5 log CFU/mL. EGCG + KR-12-a5 reduced E. faecalis (-3.93 log CFU/mL) and eliminated S. mutans in dual-species biofilms. No growth of E. faecalis and significant reduction in A. israelii (-6.24 log CFU/mL) and F. nucleatum (-4.62 log CFU/mL) counts were detected in dual-species biofilms. The combination of EGCG and KR-12-a5 led to 88% of E. faecalis dead cells inside dentin tubules. The association of EGCG and KR-12-a5 was cytocompatible and promoted synergistic effect against biofilms of bacteria associated with endodontic infections.

Effect of S. mutans combinations with bifidobacteria/lactobacilli on biofilm and enamel demineralization.

The present study evaluated the ability of Bifidobacterium and Lactobacillus species associated with streptococci to increase insoluble extracellular polysaccharide (EPS) production and initial caries lesion progression. Bovine enamel blocks (n = 190; 4 mm x 4 mm) were prepared, selected according to initial surface hardness (SH), and divided into two groups: a) double combinations: S. mutans with Bifidobacterium or Lactobacillus, and b) triple combinations: S. mutans and S. sobrinus with Bifidobacterium or Lactobacillus species. The blocks were exposed to the bacterial associations for 7 days. Subsequently, quantity of EPS from biofilms and caries lesion depth were determined by means of colorimetric and cross-sectional enamel hardness (ΔKHN) analysis. The data were submitted to one-way analysis of variance, followed by the Bonferroni test (p < 0.05). S. mutans with B. animalis or B. dentium produced a higher quantity of EPS; S. mutans + B. animalis led to the highest ∆KHN. S. mutans + S. sobrinus + B. longum induced greater EPS and ∆KHN values. In conclusion, associations of B. animalis and B. longum with streptococci promoted EPS production and caries lesion progression.

Antibiofilm and cytotoxic effect of 3,3'-dihydroxycurcumin (DHC) as photosensitizer agent in antimicrobial photodynamic therapy for endodontic purposes.

BACKGROUND: Curcuminoids have been designed not only to improve chemical and metabolic stability of curcumin (CUR), but also to increase its antimicrobial activity, without effecting its ability as photosensitizer agent in antimicrobial photodynamic therapy (aPDT) with light emitting diode (LED). This study evaluated the antimicrobial and antibiofilm action of curcumin analog DHC (or 3,3'-dihydroxycurcumin), submitted or not to LED irradiation, on microorganisms of endodontic importance and its influence on fibroblasts viability. METHODS: DHC was synthetized by modified Pablon's methodology and the experiments were conducted under irradiation or not with indium gallium nitride-based LED (440-480nm, 100 mW/cm2, 0.78 cm2,60 s). The antimicrobial activity of CUR and DHC were determined by the Minimum Inhibitory and Bactericidal Concentration assays against Gram-positive and Gram-negative bacteria and the effect of both compounds on fibroblast viability was tested using colorimetric assays. They were also evaluated on 72h and 7days single-species biofilms and on 14 days multispecies biofilms formed inside dentin tubules by bacterial colonies counts and confocal microscopy, respectively. Data were analyzed statistically considering p<0.05. RESULTS: DHC had bactericidal effect against all bacteria tested higher than CUR, in planktonic conditions. CUR and DHC (at 39 and 19 µg/mL, respectively) were cytocompatible and LED irradiation reduced fibroblast viability, regardless of compound. CUR and DHC reduced the growth of single-species biofilms and the effect of aPDT was bacteria dependent. DHC reduced more than 70% of microorganisms from multispecies biofilms, superior to CUR effect. CONCLUSIONS: DHC showed low cytotoxicity and antibiofilm effect similar to curcumin, when submitted or not to aPDT, and could be further explored as a bioactive compound for endodontic purposes.

Inhibitory Effect of a KSL-W Peptide-Loaded Poloxamer 407-Based Microemulsions for Buccal Delivery on Fusobacterium nucleatum Biofilm.

KSL-W peptide has demonstrated antibacterial and antifungal activity and inhibitory effects against oral biofilm. This study aimed to check out the effect of chlorhexidine (CLX) or KSL-W peptide-loaded poloxamer 407-based microemulsions for buccal delivery on Fusobacterium nucleatum (F. nucleatum) biofilm. The formulation (F) containing 10% copolymer poloxamer 407 dispersion (1%), 40% oleic acid and 50% PPG-5-CETETH-20 was characterized by polarized light microscopy (PLM), small-angle X-ray scattering (SAXS), rheology, bioadhesive and syringeability; and in the treatment of a biofilm produced by F. nucleatum. The darkfield images obtained by PLM and the SAXS curves with an extended peak indicated that the system was characteristic of microemulsions. In a continuous analysis, microemulsions exhibited Newtonian behavior. In frequency, the oscillatory analysis profile presented predominantly viscous behavior. Bioadhesive force detected in the analysis of F (7.4 ± 1.81 mN˙ s) and syringeability (17.83± 5.97 N · mm) being adequate values for buccal administration. After 4 h, KSL-W-loaded F shown over 20% higher effectiveness than chlorhexidine-loaded microemulsions. In conclusion, the KSL-W-loaded microemulsions showed a considerable reduction in F. nucleatum biofilm formation and presented promising structural properties for buccal drug delivery.

Correction: Sealing agent reduces formation of single and dual-species biofilms of Candida albicans and Enterococcus faecalis on screw joints at the abutment/implant interface.

[This corrects the article DOI: 10.1371/journal.pone.0223148.].

Sealing agent reduces formation of single and dual-species biofilms of Candida albicans and Enterococcus faecalis on screw joints at the abutment/implant interface.

The aim of this research was to evaluate the efficacy of a commercial sealing agent at the abutment/implant interface against microleakage of single and dual-species biofilms of Candida albicans and Enterococcus faecalis into external hexagon (EH) and Morse taper (MT) prosthetic connections. A total of 216 samples of implants and their abutments were tested. Six groups (n = 36) were evaluated based on biofilm and period of incubation (7 and 14 days). The implant connections EH and MT (n = 18) were divided according to the use of the material (n = 9) (EH-T and MT-T: with the sealing agent; EH-C and MT-C: control). The biofilms were analyzed by microbial counting (CFU/mL) and SEM analysis and photographs of the material in the screw joints were also taken. Data were analyzed by Student t test, two-way ANOVA and Bonferroni test. For the single-species biofilms, there was a significant reduction in the growth of E. faecalis when compared MT-C and MT-T or EH-C and EH-T at 7 and 14 days. The same was observed for C. albicans biofilms. For dual-species biofilms of E. faecalis and C. albicans, the sealing agent was more effective in preventing microbial infiltration into the MT connection at 14 days, while microbial infiltration did not occur into EH connections even in absence of the sealing agent for both periods of evaluation. Overall, these data suggest that the presence of the sealing agent reduces or eliminates the microleakage of E. faecalis and C. albicans biofilms into the implants regardless of the period of incubation.

Frequency of Porphyromonas gingivalis fimA in smokers and nonsmokers after periodontal therapy.

Porphyromonas gingivalis is one of the most important Gram-negative anaerobe bacteria involved in the pathogenesis of periodontitis. P. gingivalis has an arsenal of specialized virulence factors that contribute to its pathogenicity. Among them, fimbriae play a role in the initial attachment and organization of biofilms. Different genotypes of fimA have been related to length of fimbriae and pathogenicity of the bacterium. OBJECTIVES: The aim of this study was to identify 5 types of fimA genotype strains in smokers and nonsmokers with periodontitis, before and after periodontal therapy. MATERIAL AND METHODS: Thirty-one patients with periodontitis harboring P. gingivalis were selected: 16 nonsmokers (NS) and 15 smokers (SM). Clinical and microbiological parameters were evaluated at baseline and 3 months after periodontal treatment, namely: plaque index, bleeding on probe, probing depth, gingival recession and clinical attachment level. The frequency of P. gingivalis and fimA genotype strains were determined by polymerase chain reaction. RESULTS: Type I fimA was detected in the majority of SM and NS at baseline, and the frequency did not diminish after 3 months of treatment. The frequency of type II genotype was higher in SM than NS at baseline. After 3 months, statistical reduction was observed only for types II and V fimA genotypes in SM. The highest association was found between types I and II at baseline for NS (37.5%) and SM (53.3%). CONCLUSION: The most prevalent P. gingivalis fimA genotypes detected in periodontal and smoker patients were genotypes I and II. However, the presence of fimA genotype II was higher in SM. Periodontal treatment was effective in controlling periodontal disease and reducing type II and V P. gingivalis fimA.

Antimicrobial peptide-loaded liquid crystalline precursor bioadhesive system for the prevention of dental caries.

BACKGROUND: Anticaries agents must interfere with the adhesion of Streptococcus mutans and its proliferation in dental biofilm, without causing host toxicity and bacterial resistance. Natural substances, including cationic antimicrobial peptides (CAMPs) and their fragments, such as ß-defensin-3 peptide fragment (D1-23), have been widely studied. However, the chemical and physical stability of CAMPs may be compromised by external factors, such as temperature and pH, reducing the period of antimicrobial activity. METHODS: To overcome the aforementioned disadvantage, this study developed and character-ized a drug delivery system and evaluated the cytotoxicity and effect against S. mutans biofilm of a D1-23-loaded bioadhesive liquid crystalline system (LCS). LCS was composed of oleic acid, polyoxypropylene-(5)-polyoxyethylene-(20)-cetyl alcohol, Carbopol® 974P and Carbopol® 971P. LCS was analyzed by polarized light microscopy (PLM), rheology (viscoelasticity and flow properties) and in vitro bioadhesion. The viability of epithelial cells was evaluated. Minimal inhibitory concentration (MIC) and minimal bactericidal concentration (MBC) against S. mutans were determined for D1-23 for further evaluation of the effect against S. mutans biofilm after 4 and 24 h of exposure to treatments. RESULTS: PLM, rheology, and in vitro bioadhesion tests showed that both viscosity and bioadhesion of LCS increased after it was diluted with artificial saliva. D1-23-loaded LCS system presented better activity against S. mutans biofilm after 24 h when compared to 4 h of treatment, showing a cumulative effect. Neither LCS nor D1-23-loaded LCS presented toxicity on human epithelial cells. CONCLUSION: D1-23-loaded LCS is a promising drug delivery system for the prevention of dental caries.