Photodynamic therapy in root canals contaminated with Enterococcus faecalis using curcumin as photosensitizer

The aim of the study was to evaluate the photodynamic therapy (PDT) effect on root canals contaminated with Enterococcus faecalis using a light emitting diode (LED) light and a curcumin solution (CUR) as photosensitizer (PS). Eighty root canals from uniradicular human teeth were prepared with Protaper Universal rotary system and contaminated with E. faecalis for 21 days. They were divided as: GIa-PDT (CUR, pre-irradiation for 5 + 5 min of irradiation); GIb-PDT (CUR, pre-irradiation for 5 + 10 min of irradiation); GIIa-(CUR, pre-irradiation for 5 + 5 min without irradiation); GIIb-(CUR pre-irradiation for 5 + 10 min of irradiation); GIIIa-(physiological solution and irradiation for 5 min); and GIIIb-(physiological solution and irradiation for 10 min); positive and negative control groups. Collections from root canals were made at time intervals of 21 days after contamination, immediately after treatment, and 7 days after treatment, and submitted to colony forming units per milliter (CFU mL-1) counts. The data were submitted to ANOVA and Tukey multiple comparison tests, at a level of significance of 5 %. In the immediate post-treatment collection, group GIa showed greater bacterial reduction in comparison with GIIa, GIIb, GIIIa, GIIIb, and positive control (P < 0.05). At 7 days post-treatment, GIa showed significant bacterial reduction only in comparison with GIIIa (P < 0.05). Curcumin as sensitizer was effective by 5 min LED irradiation but not by 10 min irradiation PDT using LED light, and curcumin as PS was not effective in eliminating E. faecalis. No difference was observed for periods of irradiation.

In Vitro Alkaline pH Resistance of Enterococcus faecalis

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Effects of Ciprofloxacin-containing Scaffolds on Enterococcus faecalis Biofilms

Introduction: Antibiotic-containing polymer-based nanofibers (hereafter referred to as scaffolds) have demonstrated great potential for their use in regenerative endodontics from both an antimicrobial and cytocompatibility perspective. This study sought to evaluate in vitro the effects of ciprofloxacin (CIP)-containing polymer scaffolds against Enterococcus faecalis biofilms. Methods: Human mandibular incisors were longitudinally sectioned to prepare radicular dentin specimens. Sterile dentin specimens were distributed in 24-well plates and inoculated with E. faecalisfor biofilm formation. Infected dentin specimens were exposed to 3 groups of scaffolds, namely polydioxanone (PDS) (control), PDS + 5 wt% CIP, and PDS + 25 wt% CIP for 2 days. Colony-forming units (CFU/mL) (n = 10) and scanning electron microscopy (SEM) (n -= 2) were performed to quantitatively and qualitatively assess the antimicrobial effectiveness, respectively. Results: PDS scaffold containing CIP at 25 wt% showed maximum bacteria elimination with no microbial growth, differing statistically (P < .05) from the control (PDS) and from PDS scaffold containing CIP at 5 wt%. Statistical differences (P < .05) were also seen for the CFU/mL data between pure PDS (5.92-6.02 log CFU/mL) and the PDS scaffold containing CIP at 5 wt% (5.39 5.87 log CFU/mL). SEM images revealed a greater concentration of bacteria on the middle third of the dentin specimen. after 5 days of biofilm formation. On scaffold exposures, SEM images showed similar results when compared with the CFU/mL data. Dentin specimens exposed to PDS + 25 wt% CIP scaffolds displayed a practically bacteria-free surface. Conclusions: On the basis of the data presented, newly developed antibiotic-containing electrospun scaffolds hold promise as an intracanal medicament to eliminate biofilm/infection before regenerative procedures.

Different extracts of Zingiber officinale decrease Enterococcus faecalis infection in Galleria mellonella

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Estudo do pH e atividade antimicrobiana sobre Enterococcus faecalis de medicação intracanal à base de hidróxido de cálcio associada ao óleo de melaleuca, clorexidina ou farnesol

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Avaliação de propriedades físico-químicas e antimicrobianas sobre Enterococcus faecalis do mineral trióxido agregado associado a óleo de melaleuca ou farnesol

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Estudo do pH e atividade antimicrobiana sobre Enterococcus faecalis de medicação intracanal à base de hidróxido de cálcio associada ao óleo de melaleuca, clorexidina ou farnesol

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Avaliação de propriedades físico-químicas e antimicrobianas sobre Enterococcus faecalis do mineral trióxido agregado associado a óleo de melaleuca ou farnesol

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Antimicrobial activity and enterococcus faecalis biofilm formation on chlorhexidine varnishes

Objective: To evaluate, in vitro, the antimicrobial activity and biofilm formation of three chlorhexidine varnishes in four Enterococcus faecalis strains: E. faecalis ATCC 29212, E. faecalis EF-D1 (from failed endodontic treatment), E. faecalis 072 (cheese) and E. faecalis U-1765 (nosocomial infection), and one Enterococcus durans strain (failed endodontic treatment). Study Design: The direct contact test was used to study the antimicrobial activity. Bacterial suspensions were exposed for one hour to EC40, Cervitec (CE) and Cervitec Plus (CEP) varnishes. "Eradication" was defined as 100% bacterial kill. The formation of enterococci biofilms was tested on the surface of the varnishes after 24 hours of incubation and expressed as percentage of biofilm reduction. Results: EC40 eradicated all strains except E. faecalis ATCC 29212, where 98.78% kill was achieved. CE and CEP showed antimicrobial activity against all the strains, but most clearly against E. durans and E. faecalis 072. EC40 completely inhibited the formation of biofilm of E. faecalis ATCC 29212, E. faecalis 072 and E. durans. CE and CEP led to over 92% of biofilm reduction, except in the case of E. faecalis U-1765 on CEP (76.42%). Conclusion: The three varnishes studied were seen to be effective in killing the tested strains of enterococci and in inhibiting the formation of biofilm, the best results being observed with EC40.

Capsule Locus Polymorphism among Distinct Lineages of Enterococcus faecalis Isolated from Canals of Root-filled Teeth with Periapical Lesions

Introduction: Although Enterococcus faecalis is a member of the normal microbiota, it is also a major cause of nosocomial infections. Some strains of E. faecalis produce capsule, which contributes to pathogenesis through evasion of host defenses, and its production is dependent on the capsule (cps) operon polymorphism. This study investigated cps locus polymorphism in distinct lineages of E. faecalis isolated from canals of root-filled teeth with periapical lesions. Methods: Twenty-two E. faecalis isolates were evaluated regarding the cps operon polymorphism and genetic diversity. The 3 known CPS types were determined by polymerase chain reaction. This information was correlated with multilocus sequence typing data, which were used to define genetic lineages. Results: cpsA and cpsB were the only detected genes within the cps operon in 62.5% of E. faecalis strains (14/22), indicative of genotype CPS 1, which lacks capsule expression. The essential genes in the cps operon for capsule production were detected in the remaining strains, whereas 3 belonged to genotype CPS 5 and 5 strains to genotype CPS 2. A total of 14 sequence types (STs) were resolved in 22 E. faecalis isolates. Comparison with the E. faecalis international multilocus sequence typing database revealed that 9 STs were previously found, and that the 5 STs were novel. Conclusions: Certain E. faecalis genotypes from canals of root-filled teeth with periapical lesions belong to lineages associated with capsule expression and production of multiple virulence factors, which might account for their increased pathogenic potential. (J Endod 2012;38:58-61)

In vitro enterococcus faecalis biofilm formation on five adhesive systems

Objective: To determine the E. faecalis biofilm formation on the surface of five adhesive systems (AS) and its relationship with roughness. Study Design: The formation of E. faecalis biofilms was tested on the surface of four dual-cure AS: AdheSE DC, Clearfil DC Bond, Futurabond DC and Excite DSC and one light-cure antimicrobial AS, Clearfil Protect Bond, after 24 hours of incubation, using the MBEC high-throughput device. Results: E. faecalis biofilms grew on all the adhesives. The least growth of biofilm was on Excite DSC, Clearfil Protect Bond, and the control. Futurabond DC resulted in the greatest roughness and biofilm amount. There was a close relationship between the quantity of biofilm and roughness, except for Clearfil Protect Bond, which showed little biofilm but high roughness. Conclusion: None of the tested AS prevented E. faecalis biofilm formation, although the least quantity was found on the surface of Clearfil Protect Bond.

Comparative analysis of Enterococcus Faecalis biofilm formation on different substrates

Introduction: The aim of this study was to compare Enterococcus faecalis biofilm formation on different substrates. Methods: Cell culture plates containing growth medium and E. faecalis (ATCC 29212) were used to grow biofilm on bovine dentin, gutta-percha, hydroxyapatite, or bovine bone. Substrates were incubated at 37 C for 14 or 21 days, and the medium was changed every 48 hours. After the growth induction periods, specimens (n = 5 per group and per induction period) were stained by using Live/Dead, and the images were analyzed under a confocal microscope. The total biovolume (mm3), live bacteria biovolume (mm3), and substrate coverage (%) were quantified by using the BioImage_L software. Results obtained were analyzed by nonparametric tests (P = .05). Results: Biofilm formation was observed in all groups. Gutta-percha had the lowest total biovolume at 14 days (P < .05) and hydroxyapatite the highest at 21 days (P < .05). No significant difference was observed in green biovolume at 14 days. At 21 days, however, hydroxyapatite had the highest volume (P < .05). The percentages of coverage were similar among all substrates at 21 days (P > .05), but at 14 days, bovine bone presented the highest coverage (P < .05). Conclusions: E. faecalis was capable of forming biofilm on all substrates during both growth periods; hydroxyapatite presented the highest rates of biofilm formation. The type of substrate influenced the biofilm characteristics, according to the parameters evaluated

Analysis of genetic lineages and their correlation with virulence genes in enterococcus faecalis clinical isolates from root canal and systemic infections

Introduction: Enterococcus faecalis is a member of the mammalian gastrointestinal microbiota but has been considered a leading cause of hospital-acquired infections. In the oral cavity, it is commonly detected from root canals of teeth with failed endodontic treatment. However, little is known about the virulence and genetic relatedness among E. faecalis isolates from different clinical sources. This study compared the presence of enterococcal virulence factors among root canal strains and clinical isolates from hospitalized patients to identify virulent clusters of E. faecalis. Methods: Multilocus sequence typing analysis was used to determine genetic lineages of 40 E. faecalis clinical isolates from different sources. Virulence clusters were determined by evaluating capsule (cps) locus polymorphisms, pathogenicity island gene content, and antibiotic resistance genes by polymerase chain reaction. Results: The clinical isolates from hospitalized patients formed a phylogenetically separate group and were mostly grouped in the clonal complex 2, which is a known virulent cluster of E. faecalis that has caused infection outbreaks globally. The clonal complex 2 group comprised capsule-producing strains harboring multiple antibiotic resistance and pathogenicity island genes. On the other hand, the endodontic isolates were more diverse and harbored few virulence and antibiotic resistance genes. In particular, although more closely related to isolates from hospitalized patients, capsuleproducing E. faecalis strains from root canals did not carry more virulence/antibiotic genes than other endodontic isolates. Conclusions: E. faecalis isolates from endodontic infections have a genetic and virulence profile different from pathogenic clusters of hospitalized patients’ isolates, which is most likely due to niche specialization conferred mainly by variable regions in the genome.

Effect of photodynamic therapy with different formulations of methylene blue in teeth contaminated by Enterococcus faecalis

The aim of this study was to compare the disinfection of dentine using photodynamic therapy with methylene blue in different formulations. Thirty bovine teeth roots were autoclaved and incubated with a suspension of Enterococcus faecalis. The specimen were randomly divided into three groups: G1, the roots were filled with 10 mM methylene blue dissolved in water; G2, the roots were filled with 10 mM methylene blue dissolved in a mixture of glycerol: ethanol: water; G3, roots filled with 100 mM methylene blue dissolved in water. The groups were irradiated with a 660 nm diode laser with an output power of 100 mW for 4 min, energy density of 850 J/cm2 and after this procedure, the sensitizer was removed and microbial samples were collected from within the root canals. The samples were plated on mEnterococcus to count the colony-forming units (CFU/mL). The means were: Group 1=513×103, Group 2=1431×103 and Group 3=2.96×103. The statistical analysis detected higher disinfection achieved by G3 when compared with groups G1 and G2, and no significant difference between the groups G1 and G2 (P>0.05). The increase of the concentration of methylene blue dye achieved higher disinfection in photodynamic therapy.

Quantitative detection of Staphylococcus aureus, Enterococcus faecalis and Pseudomonas aeruginosa in human oral epithelial cells from subjects with periodontitis and periodontal health

Epithelial cells in oral cavities can be considered reservoirs for a variety of bacterial species. A polymicrobial intracellular flora associated with periodontal disease has been demonstrated in buccal cells. Important aetiological agents of systemic and nosocomial infections have been detected in the microbiota of subgingival biofilm, especially in individuals with periodontal disease. However, non-oral pathogens internalized in oral epithelial cells and their relationship with periodontal status are poorly understood. The purpose of this study was to detect opportunistic species within buccal and gingival crevice epithelial cells collected from subjects with periodontitis or individuals with good periodontal health, and to associate their prevalence with periodontal clinical status. Quantitative detection of total bacteria and Staphylococcus aureus, Pseudomonas aeruginosa and Enterococcus faecalis in oral epithelial cells was determined by quantitative real-time PCR using universal and species-specific primer sets. Intracellular bacteria were visualized by confocal microscopy and fluorescence in situ hybridization. Overall, 33 % of cell samples from patients with periodontitis contained at least one opportunistic species, compared with 15 % of samples from healthy individuals. E. faecalis was the most prevalent species found in oral epithelial cells (detected in 20.6 % of patients with periodontitis, P = 0.03 versus healthy individuals) and was detected only in cells from patients with periodontitis. Quantitative real-time PCR showed that high levels of P. aeruginosa and S. aureus were present in both the periodontitis and healthy groups. However, the proportion of these species was significantly higher in epithelial cells of subjects with periodontitis compared with healthy individuals (P = 0.016 for P. aeruginosa and P = 0.047 for S. aureus). Although E. faecalis and P. aeruginosa were detected in 57 % and 50 % of patients, respectively, with probing depth and clinical attachment level ≥6 mm, no correlation was found with age, sex, bleeding on probing or the presence of supragingival biofilm. The prevalence of these pathogens in epithelial cells is correlated with the state of periodontal disease.