Enterococcus Faecalis Biofilm Research Articles

Comparative evaluation of antimicrobial efficacy of chitosan nanoparticles and calcium hydroxide against endodontic biofilm of Enterococcus faecalis: An in vitro study

Aim: The aim of the study was to assess and evaluate the antimicrobial effectiveness of chitosan nanoparticles (CSNPs) with calcium hydroxide in the elimination of Enterococcus faecalis. Materials and Methods: Using the broth microdilution method, the minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of calcium hydroxide and CSNPs were measured. The antibiofilm effect of calcium hydroxide and CSNPs against E. faecalis biofilm was qualitatively analyzed using a crystal violet assay. A 7-day-old biofilms of E. faecalis grown on dentine discs were assigned to the following three groups (n = 11 dentine discs), normal saline (group I), calcium hydroxide (group II), and CSNPs (group III). Quantification of live and dead cells using confocal microscopy was done to evaluate the antibiofilm efficacy of the medicaments included in the study. Results: MIC of calcium hydroxide and CSNPs against E. faecalis was observed at 2.5 mg/mL and 0.31 mg/mL, respectively. MBC of calcium hydroxide and CSNPs was observed at 2.5 mg/mL and 0.31 mg/mL, respectively. Using Crystal Violet (CV) assay, calcium hydroxide and CSNPs showed biofilm inhibition at concentrations of 2.5 mg/mL and 0.625 mg/mL, respectively. Confocal laser scanning microscopy analysis found that both calcium hydroxide and CSNPs showed a significant decrease in viable cells at their MBC values compared to the control group’s normal saline. CSNPs showed a significantly lower percentage of live cells than calcium hydroxide (P < 0.05). Conclusion: The study results reveal that the antimicrobial efficacy of CSNPs is better than calcium hydroxide and normal saline against E. faecalis biofilm.

Differences of protein expression in enterococcus faecalis biofilm during resistance to environmental pressures.

Enterococcus faecalis biofilm was frequently found on the failed treated root canal wall, which survived by resisting disinfectant during endodontic treatment.Many researches have been conducted to explore the mechanisms of persistence of this pathogen in unfavorable conditions. However, no comprehensive proteomics studies have been conducted to investigate stress response in Enterococcus faecalis caused by alkali and NaOCl. Enterococcus faecalis (E.f) has been recognized as a main pathogen of refractory apical periodontitis, its ability to withstand environmental pressure is the key to grow in the environment of high alkaline and anti-bacterial drug that causes chronic infection in the root canal. This study aims to focus on the protein expression patterns of E.f biofilm under extreme pressure environment". Enterococcus faecalis biofilm model was established in vitro. Liquid Chromatograph-Mass Spectrometer (LC-MS/MS)-based label free quantitative proteomics approach was applied to compare differential protein expression under different environmental pressures (pH 10 and 5% sodium hypochlorite (NaOCl)). And then qPCR and Parallel Reaction Monitoring Verification (PRM) were utilized to verify the consequence of proteomics. The number of taxa in this study was higher than those in previous studies, demonstrating the presence of a remarkable number of proteins in the groups of high alkaline and NaOCl. Proteins involved in ATP-binding cassette (ABC) transporter were significantly enriched in experimental samples. We identified a total of 15 highly expressed ABC transporters in the high alkaline environment pressure group, with 7 proteins greater than 1.5 times. This study revealed considerable changes in expression of proteins in E.f biofilm during resistance to environmental pressures. The findings enriched our understanding of association between the differential expression proteins and environmental pressures.

Diacetylcurcumin: a novel strategy against Enterococcus faecalis biofilm in root canal disinfection.

Aim: We evaluated Diacetylcurcumin (DAC), a derivative of curcumin, for its antibacterial and antibiofilm properties against Enterococcus faecalis. Methods: Minimum inhibitory concentration (MIC) and minimum bactericidal concentration were determined, along with antibiofilm potential and toxicity in Galleria mellonella. Additionally, in silico computational analysis was performed to understand its mechanisms of action. Results & conclusion: DAC demonstrated significant antibacterial effects, with MIC and MBC values of 15.6 and 31.25μg/ml, respectively, and reduced biofilm formation. A synergistic effect, reducing biofilm by 77%, was observed when combined with calcium hydroxide. G. mellonella toxicity tests confirmed DAC's safety at tested concentrations, suggesting its potential for use in root canal disinfection products.

Comparative Evaluation of Ultrasonic and Sonic Irrigant Activation Systems: Assessing Extrusion Risk, Debridement, and Biofilm Removal in Distinct Apical Preparation Sizes

This study aims to compare the effectiveness of ultrasonically and sonically activated irrigation in terms of extrusion risk, root canal debridement, and biofilm removal, considering distinct apical preparation sizes, through an ex vivo study in human teeth. Instrumented teeth, to an apical size of 35/.06 or 50/.06, were assigned to three different irrigation procedures: ultrasonically activated irrigation, sonically activated irrigation, and conventional manual irrigation. Apical extrusion risk was evaluated by quantifying irrigant and debris extrusion (n = 10/group). Debris evaluation and smear layer removal from the root canal wall were conducted by scanning electron microscopy (SEM) (n = 5/group), and the elimination of a mature biofilm of Enterococcus faecalis was assessed through resazurin assay and SEM (n = 10/group). For statistical analyses, Student’s paired t-test and the ANOVA with post-hoc Tukey were used. Activated irrigations exhibited a higher risk of extrusion for the larger apical size, while the risk for manual irrigation remained independent of the apical size. Substantially fewer residual debris and smear layers were observed after the activation of the irrigant, and there was a notable enhancement in biofilm elimination compared to manual irrigation (p < 0.05). Notably, the effectiveness of both activated irrigations was more pronounced in root canals prepared to a size 50/.06, with ultrasonic activation showing enhanced improvements. The findings of this study underscore the substantial impact of both ultrasonically and sonically activated irrigation on the effectiveness of root canal disinfection and debridement. This impact is especially prominent with larger apical size, albeit accompanied by an increased risk of extrusion.

Development of Chlorhexidine-loaded Lipid Nanoparticles Incorporated in a Bioceramic Endodontic Sealer

IntroductionThis study aimed to assess BioRoot RCS (BR) incorporating liposomal chlorhexidine digluconate (CHX) for its antibacterial activity, drug release capacity, and physicochemical properties. MethodsDrug release of CHX liposomal formulations in combination with BR was evaluated spectrophotometrically and through mathematical release models for 30 days. A selected combination was evaluated for antimicrobial properties against Enterococcus faecalis biofilm growth on human dentin. Cytotoxicity was assessed following the ISO 10993-5:2019 standard on days 1, 3, and 7. Physicochemical properties were evaluated through setting time, Fourier transform infrared spectroscopy, solubility, contact angle, and film thickness. ResultsFrom BR, liposomal CHX released up to 7-fold higher CHX than CHX solution (P < .05), following a triphasic drug release pattern compared to the CHX solution, which followed a quasi-Fickian diffusion. BR combined with a selected liposomal CHX completely inhibited E. faecalis biofilm growth compared to the combination of BR with CHX solution and the control group (P < .05). Liposomal CHX decreased the contact angle (P < .05) and solubility but increased cytotoxicity (P < .05) of BR, staying above the ISO threshold. None of the other physicochemical characteristics tested differed from BR (P > .05). ConclusionThis liposomal formulation improved CHX release from BR, enhancing the antibacterial effectiveness. It presents a promising approach for local antibiofilm therapy in endodontics without substantially altering the physicochemical characteristics of BR.

Comparative Evaluation of Antibacterial Efficacy, Molecular Docking of Ethanolic Extract of Blackseed, Seaweed and Calcium Hydroxide Intracanal Medicament with Enterococcus Faecalis Antigens

ABSTRACT Aim: To evaluate the inhibitory effect of ethanolic extract blackseed, seaweed, and calcium hydroxide intracanal medicament with Enterococcus faecalis biofilm. To study the binding interaction between the active components of blackseed and seaweed against the enterococcal surface protein of (E. faecalis) by molecular docking. Materials and Methods: The ethanolic extracts of blackseed and seaweed were prepared using the Soxhlet apparatus. They were divided into three groups, namely, |Group I: Calcium hydroxide, Group II: Blackseed, and Group III: Seaweed. The antibacterial activity of the three groups was detected employing various concentrations ranging from 250, 125, and 62.5 μg/ml and based on the zone of inhibition. The inhibitory potential of medicaments to inhibit E. faecalis growth at various stages and kinetics plate were assessed following biofilm architecture evaluation by crystal violet biofilm assay. With the Swissdock suite, the molecular docking procedure was carried out. PyMOL version 4.1.5 was the program used for visualization. Since enterococcal surface protein (Esp) is primarily involved in the formation of biofilms, it was chosen as the target protein of E. faecalis. Based on their chromatographic investigations, Group II Thymoquinone (TQ) and Group III Ledenoxide were chosen as ligands. Results: The percentage of inhibition of E. faecalis biofilm was analyzed as statistically significant observed within groups. On post-hoc analysis, significant differences were present between the groups (P &lt; 0.05). Molecular docking reveals binding energies of thymoquinone (Group II) and ledenoxide (Group III) against the enterococcal surface protein of E. faecalis were −6.90 Kcal/mol and −6.44 Kcal/mol, respectively. Conclusion: Compared to seaweed, black seed extract exhibited higher antibacterial activity against the E. faecalis biofilm in microbial inhibition and molecular interaction.

In vitro antibiofilm effect of different irradiation doses in infected root canal model

BackgroundEradication of endodontic biofilms from the infected root canal system is still the main concern in endodontics. In this study, the role of the power density parameter in the efficacy of antimicrobial photodynamic therapy (PDT) with toluidine blue O (TBO) and phycocyanin (PC) activated by a 635 nm diode laser (DL) against Enterococcus faecalis biofilm in the root canal model was investigated. Materials and methodsThe E. faecalis biofilm in the root canal was treated with TBO and PC with different power densities (636, 954, 1273, and 1592 W/cm2). The untreated biofilm represented the control group. After the treatments, the biofilms were analyzed based on the number of colonies per milliliter. ResultsTBO and PC activated with 635 nm DL with a power density of 1592 W/cm2 were more efficient in removing E. faecalis biofilms within the root canals than those with a power density of 636 W/cm2 (p = 0.00). ConclusionThe light power density optimized the bacterial reduction of E. faecalis biofilms in the root canal spaces. These results provide information on the decisive parameters for performing PDT on intracanal biofilms.

The effect of root canal preparation tapers on planktonic bacteria and biofilm reduction in the apical third: A correlative microtomography and microbiological laboratory study.

To evaluate the influence of different preparation tapers on the reduction in planktonic bacteria and biofilms of Enterococcus faecalis and Candida albicans in the apical third (4 mm) of the mesial roots of mandibular molars, correlating decontamination with canal shape. After microtomography analysis for morphological standardization of the canals, 48 mandibular molar roots, each containing two canals (96 canals), were contaminated with E. faecalis and C. albicans and divided into four groups (n = 11) for canal instrumentation using ProDesign Logic 2 files with different tapers G (.03): # 25.03; G (.04): # 25.04; G (.05): # 25.05; and G (.06): # 25.06 and irrigation with 2.5% sodium hypochlorite. Four roots were examined under a scanning electron microscope (SEM) to qualitatively assess biofilm formation. Eight roots were used as the negative control group (samples were not contaminated). Bacteriological samples were taken exclusively from the apical third of the roots before and after chemical-mechanical preparation and bacterial counts were determined (CFU/mL). The final micro-CT scan was used to quantify the volume variation and unprepared canal area in the apical third. Statistical analysis was performed using the Kruskal-Wallis, Student-Newman-Keuls and Wilcoxon tests for analysis of microbiological data. anova and the Tukey or Games-Howell test were used for analysis of micro-CT data and Spearman's test for correlations (α = 5%). All groups showed a significant reduction in bacteria (p < .05), with no statistically significant difference between groups. There was no significant difference in per cent volume increase between groups. The unprepared area (Δ%) was affected by the file used (p = .026) and was significantly lower for G (.06) compared to G (.03). There was no statistically significant correlation among bacterial reduction, volume and unprepared area (p > .05). The different preparation tapers influenced root canal shaping in the apical third but did not improve decontamination in this region.

Efficacy of the GentleWave System in the removal of biofilm from the mesial roots of mandibular molars before and after minimal instrumentation: An exvivo study.

To compare the efficacy of Enterococcus faecalis biofilm removal using the GentleWave System (GWS) (Sonendo Inc, CA) on non-instrumented versus minimally instrumented root canal systems. Thirty-four mandibular molars were autoclaved and allocated to four groups: Negative control (n = 5); positive control (n = 5); Group 1: non-instrumentation + GWS (NI + GWS) (n = 12); and Group 2: minimal instrumentation + GWS (MI + GWS) (n = 12). Of 34 samples, 24 samples with Vertucci type 2 configuration within the mesial root of each sample were allocated to Groups 1 and 2 and then matched based on the working length and root canal configuration. After inoculation of samples with E. faecalis for 3 weeks, the GWS was used on Group 1 without any instrumentation and Group 2 after instrumentation of mesial canals until size 20/06v. CFU and SEM analysis were used. Log10 (CFU/mL) from the positive control, and Group 1 and 2 were 7.41 ± 0.53, 3.41 ± 1.54, and 3.21 ± 1.54, respectively. Both groups showed a statistically significant difference in the reduction of viable E. faecalis cells compared to the positive control (Group 1 [p = .0001] and Group 2 [p < .0001]), whilst showing no significant difference between the two tested groups (p < .05). The use of GWS on the non-instrumented root canal system could be an effective disinfection protocol in removing the biofilm without dentin debris formation.

Antibacterial biofilm efficacy of calcium hydroxide loaded on Gum Arabic nanocarrier: an in-vitro study

BackgroundAn innovative intracanal medication formulation was introduced in the current study to improve the calcium hydroxide (Ca(OH)2) therapeutic capability against resistant Enterococcus faecalis (E. faecalis) biofilm. This in-vitro study aimed to prepare, characterize, and evaluate the antibacterial efficiency of Ca(OH)2 loaded on Gum Arabic (GA) nanocarrier (Ca(OH)2-GA NPs) and to compare this efficiency with conventional Ca(OH)2, Ca(OH)2 nanoparticles (NPs), GA, and GA NPs.Materials and methodsThe prepared nanoparticle formulations for the tested medications were characterized using Transmission Electron Microscope (TEM) and Fourier-Transform Infrared Spectroscopy (FTIR). 141 human mandibular premolars were selected, and their root canals were prepared. Twenty-one roots were then sectioned into 42 tooth slices. All prepared root canals (n = 120) and teeth slices (n = 42) were divided into six groups according to the intracanal medication used. E. faecalis was inoculated in the samples for 21 days to form biofilms, and then the corresponding medications were applied for 7 days. After medication application, the residual E. faecalis bacteria were assessed using CFU, Q-PCR, and SEM. Additionally, the effect of Ca(OH)2-GA NPs on E. faecalis biofilm genes (agg, ace, and efaA) was investigated using RT-PCR. Data were statistically analyzed at a 0.05 level of significance.ResultsThe synthesis of NPs was confirmed using TEM. The results of the FTIR proved that the Ca(OH)2 was successfully encapsulated in the GA NPs. Ca(OH)2-GA NPs caused a significant reduction in the E. faecalis biofilm gene expression when compared to the control (p < 0.001). There were significant differences in the E. faecalis CFU mean count and CT mean values between the tested groups (p < 0.001) except between the Ca(OH)2 and GA CFU mean count. Ca(OH)2-GA NPs showed the least statistical E. faecalis mean count among other groups. SEM observation showed that E. faecalis biofilm was diminished in all treatment groups, especially in the Ca(OH)2-GA NPS group when compared to the control group.ConclusionsCa(OH)2 and GA nanoparticles demonstrate superior anti-E. faecalis activity when compared to their conventional counterparts. Ca(OH)2-GA NPs showed the best antibacterial efficacy in treating E. faecalis biofilm. The tested NP formulations could be considered as promising intracanal medications.

Comparative evaluation of the auxiliary effect of different herbs to enhance the antibacterial efficacy of calcium hydroxide against Enterococcus faecalis biofilm – An in vitro study

Aim: The increase in potential side effects and safety concerns of conventional medicaments has led to the recent popularity of herbal alternative medications. The herbal products are known for their high antimicrobial activity, biocompatibility, anti-inflammatory, and antioxidant properties. The present study was designed to compare the effectiveness of calcium hydroxide (CH) in combination with various herbs as intracanal medicaments against Enterococcus faecalis biofilm. Methods: Forty extracted single-rooted maxillary teeth were decoronated and sectioned into two halves. They were later infected with E. faecalis to form a biofilm for 21 days. Subsequently, the root samples were divided into a control group and three experimental groups – the control group was treated with CH powder mixed with distilled water and the experimental Group 1 was treated with CH powder and aloe vera gel placed over PerioCol-guided tissue regeneration (GTR) membrane; Group 2 was treated with CH powder mixed with coronil, and incorporated with aloe vera gel; and Group 3 was treated with CH powder mixed with Ajmodadi Churna incorporated with aloe vera gel. At the end of 7 and 14 days, the antimicrobial efficacy of medicaments against E. faecalis biofilm was assessed at the depths of 200–400 μm. Kruskal–Wallis test and post hoc analysis – Mann–Whitney test were used for statistical analysis. Results: Ajmodadi Churna was most effective followed by coronil and CH placed over the PerioCol GTR membrane. Conclusion: Ajmodadi Churna exhibited superior results. Coronil displayed good efficacy against E. faecalis in long-term use and thus could be used as an effective intracanal medicament. Among the Ayurvedic preparations, both hold a promising future but to implement their use as sole intracanal medicaments clinically, further in vivo and long-term studies are warranted.

Eradication of mature biofilm from the isthmus region using Er, Cr: YSGG Laser as an activator of 2% chlorhexidine gluconate.

Challenges in the root canal system, such as isthmus, may limit the action of endodontic equipment and irrigation solutions, so using laser agitation is recommended to upgrade the removal of microbial biofilm. The objective of this study is to assess the effectiveness of the laser in photon-induced photoacoustic streaming protocol agitation of 2% chlorohexidine gluconate in removing mature biofilm in complex root canal systems. Seventy-five mesial roots of the lower first and second molars were separated and cultivated with Enterococcus faecalis bacteria for 30 days (except for the negative control group samples). The samples were divided into four groups (n=15), one group acted as a positive control, other groups were irrigated with 2% chlorohexidine gluconate, some of them were agitated with a passive ultrasonic device, while the other samples were agitated by an Erbium, chromium-doped yttrium, scandium, gallium, and garnet laser in photon-induced photoacoustic streaming protocol. An atomic force microscope was used as a new method to get the results in the isthmus area; A scanning electron microscope was also used in the study to examine the samples before and after the treatment. Statistical Package for Social Sciences software was used to collect and analyze data, and two-way ANOVA was used to compare the means of the test groups. According to the atomic force microscope and scanning electron microscope analyses, Erbium laser and passive ultrasonic activation groups showed higher antimicrobial efficacy than the syringe irrigation group (p<0.05). According to the study's results, the agitation of chlorohexidine gluconate fluid by Erbium laser in photon-induced photoacoustic streaming at 0.75 W offers better Enterococcus faecalis biofilm removal in the difficult-to-reach areas of lower first and second molars. Key words:Atomic force microscope, 2% chlorhexidine gluconate, Enterococcus faecalis, Erbium, chromium-doped yttrium, scandium, gallium, garnet laser, passive ultrasonic.

Evaluation of Antimicrobial Efficacy of Silver Nanoparticles and Chitosan Nanoparticles against Enterococcus Faecalis and Candida Albicans Biofilm (An in vitro Study)

Evaluation of Antimicrobial Efficacy of Silver Nanoparticles and Chitosan Nanoparticles against Enterococcus Faecalis and Candida Albicans Biofilm (An in vitro Study)

In vitro evaluation of three engineered multispecies endodontic biofilms on a dentinal disk substrate

The aim of this study was the development of a complex multispecies endodontic biofilm using Candida albicans, Proteus mirabilis and Pseudomonas aeruginosa on a biofilm of Enterococcus faecalis in a dentinal substrate design. The endodontic pathology is a biofilm-mediated infection, and the aim of root canal therapy is to reduce, as much as possible, the bacterial population. Thus, it is important to develop a laboratory endodontic biofilm to test the effect of new irrigation and obturation techniques on reduction of bacterial count. The culture of Enterococcus faecalis from ATCC 29212 began with aerobic cultivation on blood agar, followed by transfer to Brain Heart Infusion (BHI) broth with 5% sucrose. Incubation occurred in a shaker at 37 °C for 24 h, followed by an additional 24-h static phase. After 10 d, Proteus mirabilis, Pseudomonas aeruginosa, and Candida albicans were introduced sequentially in three distinct groups. Group 1: the order of addition was Candida albicans, Proteus mirabilis, and Pseudomonas aeruginosa; Group 2: the order was Pseudomonas aeruginosa, Candida albicans, and Proteus mirabilis; and Group 3: Proteus mirabilis, Pseudomonas aeruginosa, and Candida albicans. After 16 days, the biofilm was carefully extracted, transferred to sterile BHI, and dissected using a sterile needle technique. Subsequently, an optical density test, bacterial counts, and colony enumeration were performed on various agar plates. Group 2 in which Pseudomonas aeruginosa was added directly after Enterococcus faecalis followed by Candida albicans and Proteus mirabilis showed significantly greater total bacterial count than the other two groups.

Establishment and inactivation of mono-species biofilm in a semipilot-scale water distribution system using nanocomposite of silver nanoparticles/montmorillonite loaded cationic chitosan

This study presents a novel approach in the synthesis and characterization of nanocomposites comprising cationic chitosan (CCS) blended with varying concentrations of silver nanoparticles/montmorillonite (AgNPs/MMT). AgNPs/MMT was synthesized using soluble starch as a reducing and stabilizing agent. Subsequently, nanocomposites, namely CCS/AgMMT-0, CCS/AgMMT-0.5, CCS/AgMMT-1.5, and CCS/AgMMT-2.5, were developed by blending 2.5 g of CCS with 0, 0.5, 1.5, and 2.5 g of AgNPs/MMT, respectively, and the corresponding nanocomposites were prepared using ball milling technique. Transmission electron microscopy (TEM) analysis revealed the formation of nanocomposites that exhibiting nearly spherical morphologies. Dynamic light scattering (DLS) measurements displayed average particle sizes of 1183 nm, 131 nm, 140 nm, and 188 nm for CCS/AgMMT-0, CCS/AgMMT-0.5, CCS/AgMMT-1.5, and CCS/AgMMT-2.5, respectively. The narrow polydispersity index (~0.5) indicated uniform particle size distributions across the nanocomposites, affirming monodispersity. Moreover, the zeta potential values exceeding 30 mV across all nanocomposites that confirmed their stability against agglomeration. Notably, CCS/AgMMT-2.5 nanocomposite exhibited potent antibacterial and antibiofilm properties against diverse pipeline materials. Findings showed that after 15 days of incubation, the highest populations of biofilm cells, Pseudomonas aeruginosa biofilm, developed over UPVC, MDPE, DCI, and SS, with corresponding HPCs of 4.79, 6.38, 8.81, and 7.24 CFU/cm2. The highest cell densities of Enterococcus faecalis biofilm in the identical situation were 4.19, 5.89, 8.12, and 6.9 CFU/cm2. The nanocomposite CCS/AgMMT-2.5 exhibited the largest measured zone of inhibition (ZOI) against both P. aeruginosa and E. faecalis, with measured ZOI values of 19 ± 0.65 and 17 ± 0.21 mm, respectively. Remarkably, the research indicates that the youngest biofilm exhibited the most notable rate of inactivation when exposed to a dose of 150 mg/L, in comparison to the mature biofilm. These such informative findings could offer valuable insights into the development of effective antibiofilm agents and materials applicable in diverse sectors such as water treatment facilities, medical devices, and industrial pipelines.

How Do Phages Disrupt the Structure of Enterococcus faecalis Biofilm?

Biofilms are composed of multicellular communities of microbial cells and their self-secreted extracellular polymeric substances (EPS). The viruses named bacteriophages can infect and lyze bacterial cells, leading to efficient biofilm eradication. The aim of this study was to analyze how bacteriophages disrupt the biofilm structure by killing bacterial cells and/or by damaging extracellular polysaccharides, proteins, and DNA. The use of colorimetric and spectrofluorimetric methods and confocal laser scanning microscopy (CLSM) enabled a comprehensive assessment of phage activity against E. faecalis biofilms. The impact of the phages vB_Efa29212_2e and vB_Efa29212_3e was investigated. They were applied separately or in combination on 1-day and 7-day-old biofilms. Phages 2e effectively inhibited the growth of planktonic cells with a limited effect on the biofilm. They did not notably affect extracellular polysaccharides and proteins; however, they increased DNA levels. Phages 3e demonstrated a potent and dispersing impact on E. faecalis biofilms, despite being slightly less effective than bacteriophages 2e against planktonic cells. Phages 3e reduced the amount of extracellular polysaccharides and increased eDNA levels in both 1-day-old and 7-day-old biofilm cultures. Phage cocktails had a strong antimicrobial effect on both planktonic and biofilm-associated bacteria. A significant reduction in the levels of polysaccharides, proteins, and eDNA in 1-day-old biofilm samples was noted, which confirms that phages interfere with the structure of E. faecalis biofilm by killing bacterial cells and affecting extracellular polymer levels.

In Vitro Assessment of SWEEPS and Antimicrobial Photodynamic Therapy Alone or in Combination for Eradicating Enterococcus faecalis Biofilm in Root Canals.

This study investigates the efficacy of antimicrobial photodynamic therapy (aPDT) using riboflavin and a blue diode laser (BDL), combined with shock wave-enhanced emission photoacoustic streaming (SWEEPS), against Enterococcus faecalis. A total of 48 extracted single-rooted human teeth were used. The root canals were instrumented, sealed at their apices, had the smear layer removed, and then underwent autoclave sterilization. Subsequently, each canal was inoculated with E. faecalis bacterial suspension and allowed to incubate for ten days. After confirming the presence of biofilms through scanning electron microscopy (SEM) in three teeth, the remaining teeth were randomly allocated into nine groups, each containing five teeth: control, 5.25% sodium hypochlorite (NaOCl), BDL, SWEEPS + normal saline, SWEEPS + NaOCl, riboflavin, riboflavin + SWEEPS, riboflavin + BDL, and riboflavin + BDL + SWEEPS. After the treatment, the numbers of colony-forming units (CFUs)/mL were calculated. The data were analysed using one-way ANOVA followed by Tukey's test for comparisons. All groups, with the exception of the BDL group, exhibited a significant reduction in E. faecalis CFU/mL when compared to the control group (p < 0.001). The difference in CFU/mL value between riboflavin + BDL + SWEEPS and riboflavin + SWEEPS was significant (p = 0.029), whereas there was no significant difference between riboflavin + BDL + SWEEPS and riboflavin + BDL (p = 0.397). Moreover, there was no statistically significant difference between the riboflavin + SWEEPS group and the riboflavin + BDL group (p = 0.893). The results demonstrated that combining the SWEEPS technique with riboflavin as a photosensitizer activated by BDL in aPDT effectively reduced the presence of E. faecalis in root canals.

Investigating the effect of Er,Cr:YSGG laser agitation of sodium hypochlorite on the removal of mature biofilm in the complex root canal systems using atomic force microscopy.

Endodontic infections caused by remaining biofilm following disinfection with chemical fluids encourage secondary bacterial infection; hence, employing laser pulses to activate the fluids is advised to improve microbial biofilm clearance. This study investigated the performance of Er,Cr:YSGG laser in photon-induced photoacoustic streaming (PIPS) agitation of 5.25% sodium hypochlorite (NaOCl) to enhance the removal of mature Enterococcus faecalis (E. faecalis) biofilms in complex root canal systems. The mesial roots of the lower first and second molars were separated and inoculated with E. faecalis bacteria for 30 days. The roots were irrigated with 5.25% NaOCl, some of them were agitated with passive ultrasonic irrigation (PUI), and the other roots were agitated by Er,Cr:YSGG laser using PIPS at 60 µs/pulse, 5 Hz, and 0.25, 0.5, 0.75, 1, and 1.25 W. An atomic force microscope (AFM) was used as a new method to obtain the results in the isthmus area; the results that have been obtained from each group were compared with each other. ANOVA was utilized to compare the means of the test groups. Based on the AFM and SEM analyses, laser agitation and passive ultrasonic activation groups have shown higher antimicrobial efficacy than the conventional syringe irrigation group (P<0.05). Based on the findings of this investigation, the agitation of 5.25% NaOCl solution by Er,Cr:YSGG laser in PIPS at (60 µs/pulse, 5 Hz, 1.25 W) offers better mature bacterial biofilm removal in the mesial root of lower human molars than the same irrigant with syringe irrigation and passive ultrasonic activation technique.

Evaluation of anti-biofilm effect of antimicrobial sonodynamic therapy-based periodontal ligament stem cell-derived exosome-loaded kojic acid on Enterococcus faecalis biofilm.

Introduction. Antimicrobial sonodynamic therapy (aSDT) is an approach that uses ultrasound waves (UWs) and a sonosensitizer to generate reactive oxygen species (ROS) to damage microbial cells in biofilms. Using nano-carriers, such as exosomes (Exos), to deliver the sonosensitizer can potentially enhance the effectiveness of aSDT.Hypothesis/Gap Statement. aSDT can downregulate the expression of gelE and sprE genes, increasing the production of endogenous ROS and degradation of pre-formed Enterococcus faecalis biofilms.Aim. This study investigated the anti-biofilm effect of aSDT-based periodontal ligament stem cell-derived exosome-loaded kojic acid (KA@PDL-Exo) on pre-formed E. faecalis biofilms in root canals.Methodology. Following the isolation and characterization of PDL-Exo, KA@PDL-Exo was prepared and confirmed. The minimal biofilm inhibitory concentration (MBIC) of KA, PDL-Exo, KA@PDL-Exo and sodium hypochlorite (NaOCl) was determined, and their anti-biofilm effects were assessed with and without UWs. The binding affinity of KA with GelE and SprE proteins was evaluated using in silico molecular docking. Additionally, the study measured the generation of endogenous ROS and evaluated changes in the gene expression levels of gelE and sprE.Results. The results revealed a dose-dependent decrease in the viability of E. faecalis cells within biofilms. KA@PDL-Exo was the most effective, with an MBIC of 62.5 µg ml-1, while NaOCl, KA and PDL-Exo had MBIC values of 125, 250 and 500 µg ml-1, respectively. The use of KA@PDL-Exo-mediated aSDT resulted in a significant reduction of the E. faecalis biofilm (3.22±0.36 log10 c.f.u. ml-1; P<0.05). The molecular docking analysis revealed docking scores of -5.3 and -5.2 kcal mol-1 for GelE-KA an SprE-KA, respectively. The findings observed the most significant reduction in gene expression of gelE and sprE in the KA@PDL-Exo group, with a decrease of 7.9- and 9.3-fold, respectively, compared to the control group (P<0.05).Conclusion. The KA@PDL-Exo-mediated aSDT was able to significantly reduce the E. faecalis load in pre-formed biofilms, decrease the expression of gelE and srpE mRNA, and increase the generation of endogenous ROS. These findings imply that KA@PDL-Exo-mediated aSDT could be a promising anti-biofilm strategy that requires additional in vitro and in vivo investigations.

Antibacterial Efficacy of Dual-dye and Dual Laser Photodynamic Therapy on Oral Biofilms of Enterococcus faecalis, Streptococcus mutans, and Prevotella intermedia: An In Vitro Study

Antibacterial Efficacy of Dual-dye and Dual Laser Photodynamic Therapy on Oral Biofilms of Enterococcus faecalis, Streptococcus mutans, and Prevotella intermedia: An In Vitro Study