Multiple assessment methodologies in determining the antibiofilm actions of sodium hypochlorite mixed with clodronate or etidronate in endodontic irrigation.

This study aimed to use multiple methodologies, including a novel usage of scanning electron microscopy (SEM), to evaluate the antimicrobial actions of sodium hypochlorite (NaOCl) admixed with clodronate or etidronate in root canal irrigation. The study also examined the usefulness of colony counting as a biofilm assessment methodology. Seven day Enterococcus faecalis biofilms were grown on hydroxyapatite discs. The discs were disinfected with 0.26 M clodronate-5% NaOCl, 0.26 M etidronate-5% NaOCl, 5% NaOCl, or treated with phosphate buffered saline (PBS). Assessments were performed using colony counting, SEM and the XTT reduction assay. The XTT assessment used the same groups but with 2.5% NaOCl. For colony counting, bacteria were removed from the discs by vortex mixing, followed by plating. The discs were subsequently fixed for SEM imagining and evaluators scored the SEM micrographs for remaining bacteria. Antibiofilm actions were assessed with the Kruskal-Wallis and Dunn's multiple comparison tests. SEM micrographs and the XTT assay revealed no differences between the NaOCl controls and the clodronate or etidronate mixtures with NaOCl (P > 0.05). It was concluded that the chelator mixtures with NaOCl had antibiofilm actions comparable to NaOCl. Furthermore, vortex mixing incompletely removed biofilm from HA discs in the PBS controls and hence colony counting using E. faecalis biofilms on hydroxyapatite discs could not be used for intergroup comparisons involving PBS. Additionally, colony counting could not be used for comparisons between the NaOCl treatment groups because the removal of bacteria from the substrate by vortex mixing was affected by the irrigant type.

Assessment of the Antibacterial Effects of Bismuth Nanoparticles against Enterococcus faecalis.

INTRODUCTION: Enterococcus faecalis (E. faecalis) is the most important species in dentistry and plays a significant role in the etiology of persistent apical lesions after root canal treatment. Up to date, the intracanal application of 2% chlorhexidine for 7 days is the best way to eliminate E. faecalis. However, due to the ability of this bacterium to persist and survive in harsh environments, many studies have been directed towards finding an alternative strategy for prevention or eradication of it. This study was conducted to investigate the effect of bismuth nanoparticles on E. faecalis, as an etiologic factor in recurrent root canal infections. METHODS: Forty patients, referred to Endodontic Ward of Shiraz University of Medical Science for endodontic pretreatment, provided root canal samples. First, all samples were transferred in Enterococcosel broth and incubated. Then, samples which showed growth were plated on blood agar plates and incubated for further PCR procedure. Nanoparticle powder was dissolved in high-purity water, and the final concentration of bismuth nanoparticles (BiNPs) was measured by the spectrophotometer. Minimum inhibitory concentration (MIC) of BiNPs against E. faecalis was determined by microbroth dilution method according to methods for antimicrobial susceptibility tests. Also, bactericidal assays were conducted in Mueller-Hinton broth medium and reported as the concentration of BiNPs that reduced the viable bacterial count by 99.9%. RESULTS: Of all samples, 77.5% revealed the presence of E. faecalis by PCR. Also, E. faecalis growth inhibition was observed at concentrations ranging from 0.625 µg/ml to 20 µg/ml (geometric mean: 2.337 µg/ml), and the MBC values were between 1.25 µg/ml and 40 µg/ml (geometric mean: 4.781 µg/ml), which in comparison with chlorhexidine, these values were about one-eighth of chlorhexidine. CONCLUSION: The experimental data suggest that bismuth nanoparticles could be an interesting alternative to combat E. faecalis, which, in view of the advantages mentioned for bismuth nanoparticle like inhibiting Streptococcus mutans biofilm formation and higher antibacterial activity compared to chlorhexidine, can be suggested to be used in different fields of dentistry.

Beta-lactams susceptibility testing of penicillin-resistant, ampicillin-susceptible Enterococcus faecalis isolates: a comparative assessment of Etest and disk diffusion methods against broth dilution.

This study aimed to evaluate the accuracy of disk diffusion and Etest methods, compared to that of the broth dilution reference method for identifying beta-lactam susceptibilities of Penicillin-Resistant, Ampicillin-Susceptible Enterococcus faecalis (PRASEF) isolates. Fifty-nine PRASEF and 15 Penicillin-Susceptible, Ampicillin-Susceptible E. faecalis (PSASEF) clinical nonrepetitive isolates were evaluated. The effectiveness of five beta-lactams (ampicillin, amoxicillin, imipenem, penicillin, and piperacillin) was tested. All antimicrobial susceptibility tests were performed and interpreted according to the Clinical and Laboratory Standards Institute guidelines. Interpretative discrepancies, such as essential agreement, categorical agreement, and errors, were assessed. The acceptability was ≥ 90% for both categorical agreement and essential agreement. Etest proved to be an accurate method for testing beta-lactam susceptibilities of the emerging PRASEF isolates, disk diffusion presented poor performance, particularly for imipenem and piperacillin.

Assessment of Nitrofurantoin as an Experimental Intracanal Medicament in Endodontics.

Background and Objectives. Multiple antibacterial agents have been mixed and used as an intracanal medicament-like modified triple antibiotic paste (MTAP) to eliminate Enterococcus faecalis (EF), which has been most frequently identified in the cases of failed root canal treatment and periapical lesions. This study is aimed at using a single antibacterial agent, nitrofurantoin (Nit), as an experimental intracanal medicament paste against different clinical isolates of EF), which has been most frequently identified in the cases of failed root canal treatment and periapical lesions. This study is aimed at using a single antibacterial agent, nitrofurantoin (Nit), as an experimental intracanal medicament paste against different clinical isolates of Materials and Methods. Three strains of EF), which has been most frequently identified in the cases of failed root canal treatment and periapical lesions. This study is aimed at using a single antibacterial agent, nitrofurantoin (Nit), as an experimental intracanal medicament paste against different clinical isolates of n = 90), group M (MTAP) (n = 90), group M (MTAP) (n = 90), group M (MTAP) (EF), which has been most frequently identified in the cases of failed root canal treatment and periapical lesions. This study is aimed at using a single antibacterial agent, nitrofurantoin (Nit), as an experimental intracanal medicament paste against different clinical isolates of n = 90), group M (MTAP) (n = 90), group M (MTAP) (n = 90), group M (MTAP) (EF), which has been most frequently identified in the cases of failed root canal treatment and periapical lesions. This study is aimed at using a single antibacterial agent, nitrofurantoin (Nit), as an experimental intracanal medicament paste against different clinical isolates of. RESULTS: Nit could eradicate S1, S2, and S3 completely with concentrations of 6.25, 12.5, and 25 mg/mL, respectively, while MTAP showed complete eradication of the three strains only at 25 mg/mL. In all the groups, it was found that the CFU counts of EF), which has been most frequently identified in the cases of failed root canal treatment and periapical lesions. This study is aimed at using a single antibacterial agent, nitrofurantoin (Nit), as an experimental intracanal medicament paste against different clinical isolates of. CONCLUSION: At the concentration of 25 mg/mL, the Nit paste is effective in eradicating EF completely when it is used as an intracanal medicament.EF), which has been most frequently identified in the cases of failed root canal treatment and periapical lesions. This study is aimed at using a single antibacterial agent, nitrofurantoin (Nit), as an experimental intracanal medicament paste against different clinical isolates of.

Antimicrobial activity of bioactive glass S53P4 against representative microorganisms causing osteomyelitis - Real-time assessment by isothermal microcalorimetry.

Bioactive glass (BAG) is a synthetic bone substitute with intrinsic antimicrobial properties, used for bone defect filling. We evaluated the antimicrobial activity of two formulations of BAG S53P4 against representative pathogens of osteomyelitis: Staphylococcus aureus, Staphylococcus epidermidis, Enterococcus faecalis, Escherichia coli and Candida albicans. Antimicrobial activity of BAG S53P4 was assessed by isothermal microcalorimetry, a highly sensitive assay measuring metabolic-related microbial heat production in real-time. Standard CFUs-counting was performed in parallel. BAG granules (diameter 500-800 µm) and powder (<45 µm) were evaluated in two concentrations (400 and 800 mg/ml). Isothermal microcalorimetry was performed in glass ampoules containing growth medium, BAG and test microorganism, heat production was measured for 24 h. BAG S53P4 inhibited heat production of most-tested microorganisms with heat reduction of 60%-98% compared to positive control after 24 h of exposure to the highest-tested concentration (800 mg/ml). BAG S53P4 in powder formulation (<45 µm) inhibited more microbial growth than in granule formulation (500-800 µm), with the exception of C. albicans for which both formulations presented similar inhibition rates ranging between 87 % and 97 %. The BAG inhibitory ratios estimated from the variation in the growth rate constants of each microorganism compared to the growth control ranged between 2.55 % and 100 %. Comparable results were obtained by CFUs-counting, with complete reduction in cell viability of most microorganisms after ≤ 24 h of microbial exposure to BAG S53P4 powder. In summary, BAG S53P4 demonstrated efficient inhibition of microbial growth, especially in powder formulation.

Assessment of Tedizolid In Vitro Activity and Resistance Mechanisms against a Collection of Enterococcus spp. Causing Invasive Infections, Including Isolates Requiring an Optimized Dosing Strategy for Daptomycin from U.S. and European Medical Centers, 2016 to 2018.

High-level aminoglycoside resistance was noted in 30.0% of Enterococcus faecalis and 25.2% of Enterococcus faecium isolates. Only 3.3% and 2.1% of E. faecalis isolates had elevated daptomycin MIC (≥2 mg/liter) and vancomycin resistance, respectively. In contrast, 37.4% to 40.3% of E. faecium isolates exhibited these phenotypes. Tedizolid inhibited 98.9% to 100.0% of enterococci causing serious invasive infections, including resistant subsets. Oxazolidinone resistance was mainly driven by G2576T; however, optrA and poxtA genes were also detected, including poxtA in the United States and Turkey.

Role of non-PTS dependent glucose permease (GlcU) in maintaining the fitness cost during acquisition of nisin resistance by Enterococcus faecalis.

Nisin is used for food preservation due to its antibacterial activity. However, some bacteria survive under the prevailing conditions owing to the acquisition of resistance. This study aimed to characterize nisin-resistant Enterococcus faecalis isolated from raw buffalo milk and investigate their fitness cost. FE-SEM, biofilm and cytochrome c assay were used for characterization. Growth kinetics, HPLC, qPCR and western blotting were performed to confer their fitness cost. Results revealed that nisin-resistant E. faecalis were morphologically different from sensitive strain and internalize more glucose. However, no significant difference was observed in the growth pattern of the resistant strain compared to that of the sensitive strain. A non-phosphotransferase glucose permease (GlcU) was found to be associated with enhanced glucose uptake. Conversely, Mpt, a major phosphotransferase system responsible for glucose uptake, did not play any role, as confirmed by gene expression studies and western blot analysis of HPr protein. The phosphorylation of His-15 residue of HPr phosphoprotein was reduced, while that of the Ser-46 residue increased with progression in nisin resistance, indicating that it may be involved in the regulation of pathogenicity. In conclusion, resistance imposes a significant fitness cost and GlcU plays a key role in maintaining the fitness cost in nisin-resistant variants.

Assessment of vancomycin resistance transfer among enterococci of clinical importance in milk matrix.

Dissemination of vancomycin resistance in enterococci has been associated with horizontal transfer of mobile genetic elements. Aim of the study was to evaluate if milk matrix is a suitable environment to support transferability of vancomycin resistance (vanA) gene from clinical vancomycin-resistant Enterococcus faecium to vancomycin-sensitive Enterococcus faecalis. Enterococci strains were firstly screened for the presence of cpd (inducible sex pheromone determinant) gene, vanA and tetL genes (vancomycin and tetracycline resistance markers, respectively) and the gelE (extracellular metalloendopeptidase) gene to define the mating pairs. Based on these selection markers, we investigated the transferability of eight plasmid-borne vanA harbored by E. faecium (vanA+, cpd-, tetL- and gelE-) into two E. faecalis (vanA-, cpd+, tetL + and gelE+) recipient strains in milk matrix. The strains were mated in a 1:1 ratio in 7% reconstituted milk and incubated at 37 °C. Transconjugants emerged from all 16 matings within 2 h of incubation and were evidenced by dual antibiotic resistance (vancomycin and tetracycline). The vancomycin-resistance of trasconjugants was maintained even after ten subsequent passages on nonselective medium. Transconjugants were positive for vanA, tetL and gelE genes. This study indicates milk matrix as suitable environment to support gene exchange between Enterococcus species.

Resistance proportions for eight priority antibiotic-bacterium combinations in OECD, EU/EEA and G20 countries 2000 to 2030: a modelling study.

BackgroundAntimicrobial resistance is widely considered an urgent global health issue due to associated mortality and disability, societal and healthcare costs.AimTo estimate the past, current and projected future proportion of infections resistant to treatment for eight priority antibiotic-bacterium combinations from 2000 to 2030 for 52 countries.MethodsWe collated data from a variety of sources including ResistanceMap and World Bank. Feature selection algorithms and multiple imputation were used to produce a complete historical dataset. Forecasts were derived from an ensemble of three models: exponential smoothing, linear regression and random forest. The latter two were informed by projections of antibiotic consumption, out-of-pocket medical spending, populations aged 64 years and older and under 15 years and real gross domestic product. We incorporated three types of uncertainty, producing 150 estimates for each country-antibiotic-bacterium-year.ResultsAverage resistance proportions across antibiotic-bacterium combinations could grow moderately from 17% to 18% within the Organisation for Economic Co-operation and Development (OECD; growth in 64% of uncertainty sets), from 18% to 19% in the European Union/European Economic Area (EU/EEA; growth in 87% of uncertainty sets) and from 29% to 31% in Group of Twenty (G20) countries (growth in 62% of uncertainty sets) between 2015 and 2030. There is broad heterogeneity in levels and rates of change across countries and antibiotic-bacterium combinations from 2000 to 2030.ConclusionIf current trends continue, resistance proportions are projected to marginally increase in the coming years. The estimates indicate there is significant heterogeneity in resistance proportions across countries and antibiotic-bacterium combinations.

Assessment of antibacterial activity of 2.5% NaOCl, chitosan nano-particles against Enterococcus faecalis contaminating root canals with and without diode laser irradiation: an in vitro study.

Objective: This study was done to evaluate the antibacterial effect of chitosan nano-particles (CNPs) root canal irrigant as a new alternative to Sodium hypochlorite (NaOCl) for disinfection of root canals inoculated with Enterococcus faecalis, with and without laser activation.Methodology: Sixty single rooted human premolars were decoronated, prepared and had their apical foramina sealed. E. faecalis were incubated in the root canals for 15 days. The teeth were then randomly divided into two experimental groups (n = 30) according to the disinfection protocol used. In Group I: disinfection was performed using the irrigant solutions only (Saline, 2.5% NaOCl, CNPs). Whereas in Group II, disinfection was done using the same irrigants followed by Diode laser at (980-nm) at 2 W output for 5 × 5 s. Intra-canal bacterial samples were taken before and after canal disinfection to determine the CFU count.Results: In group I, 2.5% NaOCl was as effective as CNP in eradication and significantly more effective than Saline (p = 0.008) in eradication of E. faecalis. In Group II, either 2.5% NaOCl or CNP in combination with diode laser irradiation showed a similarly high effect in bacterial eradication.Conclusions: Within the parameters used in this study, a combination therapy consisting of irrigation followed by diode laser irradiation should be utilized as an effective treatment modality for eliminating E. faecalis from root canal systems.

The importance of adjusting for enterococcus species when assessing the burden of vancomycin resistance: a cohort study including over 1000 cases of enterococcal bloodstream infections.

Background: Infections caused by vancomycin-resistant enterococci (VRE) are on the rise worldwide. Few studies have tried to estimate the mortality burden as well as the financial burden of those infections and found that VRE are associated with increased mortality and higher hospital costs. However, it is unclear whether these worse outcomes are attributable to vancomycin resistance only or whether the enterococcal species (Enterococcus faecium or Enterococcus faecalis) play an important role. We therefore aimed to determine the burden of enterococci infections attributable to vancomycin resistance and pathogen species (E. faecium and E. faecalis) in cases of bloodstream infection (BSI). Methods: We conducted a retrospective cohort study on patients with BSI caused by Enterococcus faecium or Enterococcus faecalis between 2008 and 2015 in three tertiary care hospitals. Data was collected on true hospital costs (in €), length of stay (LOS), basic demographic parameters, and underlying diseases including the results of the Charlson comorbidity index (CCI). We used univariate and multivariable regression analyses to compare risk factors for in-hospital mortality and length of stay (i) between vancomycin-susceptible E. faecium- (VSEm) and vancomycin-susceptible E. faecalis- (VSEf) cases and (ii) between vancomycin-susceptible E. faecium- (VSEm) and vancomycin-resistant E. faecium-cases (VREm). We calculated total hospital costs for VSEm, VSEf and VREm. Results: Overall, we identified 1160 consecutive cases of BSI caused by enterococci: 596 (51.4%) cases of E. faecium BSI and 564 (48.6%) cases of E. faecalis BSI. 103 cases of E. faecium BSI (17.3%) and 1 case of E. faecalis BSI (0.2%) were infected by vancomycin-resistant isolates. Multivariable analyses revealed (i) that in addition to different underlying diseases E. faecium was an independent risk factor for in-hospital mortality and prolonged hospital stay and (ii) that vancomycin-resistance did not further increase the risk for the described outcomes among E. faecium-isolates. However, the overall hospital costs were significantly higher in VREm-BSI cases as compared to VSEm- and VSEf-BSI cases (80,465€ vs. 51,365€ vs. 31,122€ p < 0.001). Conclusion: Our data indicates that in-hospital mortality and infection-attributed hospital stay in enterococci BSI might rather be influenced by Enterococcus species and underlying diseases than by vancomycin resistance. Therefore, future studies should consider adjusting for Enterococcus species in addition to vancomycin resistance in order to provide a conservative estimate for the burden of VRE infections.

The effect of sublethal photodynamic therapy on the expression of Enterococcal surface protein (esp) encoding gene in Enterococcus faecalis: Quantitative real-time PCR assessment.

BACKGROUND: During antimicrobial photodynamic therapy (aPDT) in the treatment of endodontic intracanal infection, it is extremely likely that microorganisms would be exposed to sub-lethal doses of PDT (sPDT). Although sPDT would not result in microorganism death, it can considerably influence microbial virulence. This study evaluated the effect of sPDT on gene expression of Enterococci surface protein (esp) as a major virulence factor for biofilm formation in Enterococcus faecalis. MATERIALS AND METHODS: The lethal and sub-lethal potential of aPDT using indocyanine green (ICG), toluidine blue O (TBO), and methylene blue (MB), as the photosensitizers (PSs) and 660, 635, and 810 nm diode laser against E. faecalis was analyzed using colony-forming unit assays. Considering sub-lethal doses of PSs and photo-irradiation time of the diode laser, the expression of esp was evaluated through quantitative Real-time PCR (qRT-PCR). RESULTS: ICG at a concentration of 31.2-1000 µg/mL and both TBO and MB at 6.2-100 µg/mL significantly reducedE. faecalis growth. 660, 635, and 810 nm diode laser with energy density of 93.75-140.62, 137.5-206.25, and 31.25-62.5 J/cm2, reduced the bacterial count. ICG with a concentration of 15.6 µg/mL and irradiation time of 0.5 min, TBO with a concentration of 3.1 µg/mL and irradiation time of 3 min, and MB with a concentration of 3.1 µg/mL and irradiation time of 3 min were found as a sPDT dose against E. faecalis. The esp expression was significantly reduced at sub-lethal doses of ICG compared to TBO (2 fold) and MB (2.4 fold). CONCLUSION: Although all tested PSs showed bacterial reduction, ICG may be considered as the best PS in treating endodontic infection due to its higher efficacy in reduction of esp expression which has a main role in biofilm formation of E. faecalis.

Antimicrobial photodynamic therapy assessment of three indocyanine green-loaded metal-organic frameworks against Enterococcus faecalis.

BACKGROUND: Antimicrobial photodynamic therapy (aPDT) has emerged as one of the promising non-invasive adjuvant treatments of endodontic infections. The key part of this technique is application of an optimized nontoxic photosensitizer (PS), like indocyanine green (ICG) which when activated by light can destroy bacterial contaminants. Notwithstanding all featured properties of ICG, this PS mainly suffers from the lack of stability and concentration-dependent aggregation. A variety of nanomaterials (NMs) has been widely exploited to improve the stability and efficiency of ICG. The objective of this study was to evaluate and compare the efficiency of three high capacious metal organic frameworks (MOFs) to produce MOF-ICG as novel PSs improving ICG loading, stability and antimicrobial activity. This is first report on ICG-loaded MOFs for aPDT against endodontic infections. MATERIALS AND METHODS: Three different nano-MOFs were synthesized (denoted as Fe-101, Al-101 and Fe-88), and employed for ICG loading (MOF-ICG). The stability of immobilized ICG, antimicrobial and anti-biofilm properties of MOF-ICG against Enterococcus faecalis (E. faecalis) as one of the main factors of endodontic infections as well as expression ratio of the esp gene in E. faecalis were evaluated. RESULTS: Fe-101 and Al-101 showed acceptable ICG loading (ICG loading capacity of 16.93 ± 0.32 and 18.17 ± 0.31, respectively) as well as considerable enhanced aqueous stability (percent of degradation were only 14% and 17%, respectively) in comparison to free ICG (percent of degradation was 95%) after 10 days. ICG-free MOFs could surprisingly suppress the viability of E. faecalis after laser irradiation up to 18.1%, 28.8%, and 38.3% for Al-101, Fe-88 and Fe-101, respectively. ICG loaded MOFs mediated aPDT could significantly reduce the count of E. faecalis to 60.72%, 45.12%, and 62.67%, respectively (p < 0.05). The Fe-88-ICG-PDT, Fe-101-ICG-PDT and Al-101-ICG-PDT considerably dropped the biofilm formation of E. faecalis by 37.54%, 47.01% and 53.68% (p < 0.05). The expression of esp gene was also remarkably declined to 4.4-, 6.0- and 6.2-fold after aPDT in the presence of Fe-88-ICG, Al-101-ICG and Fe-101-ICG, respectively (P < 0.05). CONCLUSION: Owing to the significant features of the Fe-101 including acceptable ICG loading and stability, as well as reasonable antimicrobial effect after ICG loading in comparison to free ICG, it could be considered as a promising nano-PSs in aPDT to remove E. faecalis.

A novel high-resolution melting analysis approach for rapid detection of vancomycin-resistant enterococci.

BACKGROUND: Vancomycin-resistant enterococci (VRE) are resistant to most classes of antibiotics. Diagnosis of VRE using standard methods takes 2 to 5 days. Development of a rapid PCR-assay that detects and identifies resistant genes in bacteria would provide time-critical information on the presence of VRE in clinical samples allowing early treatment and management of infected patients. OBJECTIVES: Investigate the use of high resolution melting analysis (HRMA) and 16S-rRNA-PCR approach for rapid and cost-effective identification of VRE. DESIGN: Descriptive antibiotic susceptibility studies. SETTING: Manchester Academic Health Sciences Centre and School of Translational Medicine, University of Manchester, UK, and Department of Clinical Laboratory Sciences, Taibah University, Saudi Arabia. MATERIALS AND METHODS: PCR-HRMA using 16S-rRNA V1-primers was used to detect and identify VRE. DNA from different strains of vancomycin-resistant and -sensitive Enterococcus faecalis (VSE) and Enterococcus faecium were amplified using V1-primer followed by HRMA in a single run. Differentiation of VRE from VSE was based on curve shapes generated against reference organisms (Bacteroides fragilis). MAIN OUTCOMES MEASURES: Amplification curves and difference plots for VRE and VSE. RESULTS: Difference plots were generated for all vancomycin-resistant and -sensitive E faecalis and E faecium strains by subtracting their fluo.rescence melting profile from that of a reference-species B fragilis. A characteristic curve shape was produced by vancomycin-sensitive E faecalis and E faecium. However, vancomycin-resistant strains of these bacteria were associated with a markedly different curve shape facilitating a clear differentiation. CONCLUSION: The 16S-PCR-HRMA approach has the potential for detecting vancomycin-resistant E faecium and E faecalis. Data with VRE provide the basis for combining VRE identification with pathogens speciation in a rapid, cheap assay able to identify a pathogen as an Enterococcus and whether it is vancomycin-sensitive or -resistant E faecium or E faecalis in a single PCR and HRMA run. LIMITATIONS: Tested on specific, but not all, reference Enterococcus species and clinical isolates. CONFLICT OF INTEREST: None.

Ex vivo assessment of synergic effect of chlorhexidine for enhancing antimicrobial photodynamic therapy efficiency on expression patterns of biofilm-associated genes of Enterococcus faecalis.

BACKGROUND: It has clearly been demonstrated that Enterococcus faecalis, as a persistent microorganism, is the major agent in the etiopatogeny of endodontic infections. Recently, the limitations of conventional endodontic therapy have given rise to many attempts to introduce antimicrobial photodynamic therapy (aPDT) as an alternative treatment. The aim of this study was to analyze the ex vivo effect of aPDT in combination with 2.0% chlorhexidine (CHX) as a conventional therapy on colony count and expression patterns of genes associated with biofilm formation of E. faecalis. MATERIALS AND METHODS: A total of 125 extracted human single-rooted teeth were divide into six groups (A-F; n = 20) and were incubated with E. faecalis. Group A- photosensitizer (indocyanine green [ICG]); B- diode laser; C- aPDT; D- 2.0% CHX; E- aPDT with photosensitizer modified by 2.0% CHX; and F- control group (no procedure was performed). Five remaining teeth were used to confirm the presence of E. faecalis biofilm via scanning electron microscope. Counts of colony forming units (CFUs) in each group were evaluated separately and quantitative real-time PCR (qRT-PCR) was then applied to monitor genes expression of fsrC, efa, and gelE involved in E. faecalis biofilm. RESULTS: The results showed that none of the tested groups achieved eradication or inhibition of biofilm. On the other hand, aPDT + 2.0% CHX, 2.0% CHX, and ICG- mediated aPDT groups showed significantly less CFU/mL than ICG and diode laser groups. The group with the lowest CFU/mL count was the aPDT + 2.0% CHX, being statistically different from all other groups that could decrease the expression levels of efa, gelE, and fsrC genes 6.8-, 8.3-, and 12.1-fold, respectively. CONCLUSION: Based on the results, the synergism effect of ICG-aPDT with 2.0% CHX leads to modulation of the virulence of E. faecalis strains biofilm model by suppressing the expression of the genes associated with biofilm formation.

Zein based magnesium oxide nanoparticles: Assessment of antimicrobial activity for dental implications.

MgO nanoparticles have been recently discovered as an antibacterial, however, they limited by property degradation due to agglomeration. The addition of a coating agent, such as a zein polymer, is effective in preventing agglomeration without affecting nanosized properties. The aim of this study was to assess the antimicrobial property of MgO nanoparticles when coated with a zein polymer against several oral bacteria and fungi. This was done by utilizing various assessment techniques. The ultimate aim is to use these nanoparticles in dental preparations. The antimicrobial activity of zein-coated MgO nanoparticles at different concentrations of 0.5, 1 and 2% were tested against four different microorganisms: Staphylococcus aureus, Streptococcus mutans and Enterococcus faecalis (gram positive bacteria), and Candida albicans (as oral fungus). Two different techniques were utilized: the Kirby-Bauer test, and a modified direct contact test. The results indicated that the antibacterial effect of 1% or 2% zein-coated MgO nanowires were statistically significant (p<0.05) against the four organisms studied: S. mutans, S. aureus, E. faecalis and C. albicans. Zein-coated MgO nanoparticles are a new human friendly and potent antimicrobial agent that can be incorporated in the formulation of a variety of new dental materials and products that should provide improvements in dental care and oral health.

Synthesis, molecular docking, antimicrobial, antioxidant and toxicity assessment of quinoline peptides.

A series of quinoline based peptides were synthesized by a one-pot reaction through Ugi-four component condensation of lipoic acid, cyclohexyl isocyanide, aniline derivatives and 2-methoxy quinoline-3-carbaldehyde derivatives under microwave irradiation. The products were obtained in excellent yields and high purity. Solvent optimization and the effect of microwave irradiation with various powers were also observed. All the synthesized compounds were characterized by FTIR, NMR spectral data and elemental analysis. A total of eight peptides were subjected to antimicrobial, antioxidant and toxicity evaluation. Among them, four peptides showed potential towards antibacterial screening with Bacillus cereus, Staphylococcus aureus, Escherichia coli, Enterococcus faecalis and Candida albicans, Candida utilis and three peptides showed antioxidant test positive (DPPH). Besides, toxicity of all the peptides were evaluated by using brine shrimp and it was observed that four peptides showed mortality rate less than 50% up to 48h. Molecular docking studies revealed that the higher binding affinity of the two peptides toward DNA gyrase than ciprofloxacin based on Libdock score. The described chemistry represents a facile tool to synthesize complex heterocycles of pharmaceutical relevance in a highly efficient and one-pot fashion. The advantages of this method are its green approach, inexpensive solvent, shorter reaction times and excellent yields.

Ex Vivo Assessment of Photodynamic Therapy in Achieving Microbial Reduction.

The study was to assess the effects of photodynamic therapy (PDT) on root canals contaminated by Enterococcus faecalis (E. faecalis). PDT was performed with 0.005% methylene blue dye and with a low-intensity red emission laser. Microbiological samples were collected before, immediately and 72 hours after instrumentation. Although PDT performed with the chosen light parameters promoted additional microbial reduction, this therapy did not achieve a statistically significant reduction of E. faecalis.

Molecular assessment of virulence determinants, hospital associated marker (IS16gene) and prevalence of antibiotic resistance in soil borne Enterococcus species.

Enterococci, no more regarded as GRAS (Generally Recognized As Safe) organism, are emerging as an important source of nosocomial infections worldwide. The main contributors in pathogenesis of enterococci are the presence of various virulent factors and antibiotic resistance genes. We aimed to examine the prevalence, dissemination, antibiotic resistance and virulent factors associated with enterococci from bulk soil (BS). A total of 372 enterococci were isolated from 500 soil samples. PCR was used to identify the isolates up to species level and for carriage of 16 virulence genes including hospital associated marker (i.e. IS16). E. faecium (77%), E. faecalis (10%), E. hirae (4%) and E. casseliflavus (1%) were the major species isolated. The efaAfs was the most dominant gene (100%), followed by gelE (78.9%), sprE (76.3%) and esp (13%) in E. faecalis isolates. The E. faecium carried largely efaAfm (86.8%) and acm (50.3%) genes. Presence of entP (10%), entA (8.3%) and entB (6.9%) genes was detected mostly in E. faecium, while enlA (18%) and ef1097 (2.6%) was only detected in E. faecalis isolates. 50% E. faecalis and 2% E. faecium isolates harbored IS16, while five E. faecalis harbored both IS16 and espTIM genes providing strong evidence about the presence of espTIM gene on 64 Kb pathogenicity island. BOX and RAPD PCR analysis revealed high degree of genetic variation within the species. Degree of resistance against 12 major antibiotics showed chloramphenicol as the most effective and meropenom as the least effective antibiotic. Presence of multiple antibiotic resistant, virulent and hospital associated enterococci in bulk soil represents a potential source for further dissemination to humans and animals and poses potential impact on public health.