scholarly journals Combined Effect of Melittin and DNase on Enterococcus faecalis Biofilms and Its Susceptibility to Sodium Hypochlorite

Materials ◽  
2020 ◽  
Vol 13 (17) ◽  
pp. 3740
Author(s):  
Sujitha Ramaraj ◽  
Mi-Ah Kim ◽  
Vinicius Rosa ◽  
Prasanna Neelakantan ◽  
Won-Jun Shon ◽  
...  
Keyword(s):  
Enterococcus Faecalis ◽  
Sodium Hypochlorite ◽  
Laser Scanning ◽  
Formation Rate ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
P Value ◽  
Water Control ◽  
Sem Images ◽  
Root Canals

Biofilm communities are tolerant to antimicrobials and difficult to eradicate. This study aimed to investigate the effect of melittin, an antimicrobial peptide, either alone or in combination with deoxyribonuclease (DNase), an inhibitor of extracellular deoxyribonucleic acid (eDNA), against Enterococcus faecalis (E. faecalis) biofilms, and biofilm susceptibility to sodium hypochlorite (NaOCl). Biofilms of E. faecalis were developed in root canals of bovine teeth. The biofilms were treated with distilled water (control), melittin, DNase, or DNase+melittin. The antibiofilm effects of the treatments were analyzed using colony forming unit (CFU) assay, crystal violet staining, confocal laser scanning microscopy (CLSM), and field emission scanning electron microscope (FE-SEM). The susceptibility of DNase+melittin-treated biofilms to NaOCl (0%, 2.5% and 5%) was investigated by the CFU assay. The data were statistically analyzed using one-way analysis of variance, followed by Tukey’s test. A p-value of <0.05 was considered significant. Specimens treated with DNase+melittin showed a more significant decrease in the CFUs, eDNA level, and biofilm formation rate than those treated only with melittin or DNase (p < 0.05). CLSM analysis showed DNase+melittin treatment significantly reduced the volume of biofilms and extracellular polymeric substance compared to either treatment alone (p < 0.05). FE-SEM images showed a high degree of biofilm disruption in specimens that received DNase+melittin. 2.5% NaOCl in specimens pretreated with DNase+melittin showed higher antibacterial activity than those treated only with 5% NaOCl (p < 0.05). This study highlighted that DNase improved the antibiofilm effects of melittin. Moreover, DNase+melittin treatment increased the susceptibility of biofilms to NaOCl. Thus, the complex could be a clinical strategy for safer use of NaOCl by reducing the concentration.

scholarly journals Er,Cr:YSGG Laser-Activation Enhances Antimicrobial and Antibiofilm Action of Low Concentrations of Sodium Hypochlorite in Root Canals

Antibiotics ◽  
2019 ◽  
Vol 8 (4) ◽  
pp. 232
Author(s):  
Pablo Betancourt ◽  
Josep María Sierra ◽  
Octavi Camps-Font ◽  
Josep Arnabat-Domínguez ◽  
Miguel Viñas
Keyword(s):  
Confocal Laser Scanning Microscopy ◽  
Sodium Hypochlorite ◽  
Laser Scanning ◽  
Laser Scanning Microscopy ◽  
Confocal Laser Scanning ◽  
Scanning Microscopy ◽  
Confocal Laser ◽  
Root Canals ◽  
Low Concentrations ◽  
Scanning Electron

The onset and persistence of endodontic infections due to residual biofilm after chemical disinfection promotes secondary bacterial infection. Alternative methods to disinfect operated root canals are a matter of great interest. The aim was to evaluate the antibacterial effectiveness of sodium hypochlorite (NaOCl) at low concentrations activated by the Er,Cr:YSGG laser-activated irrigation (LAI) against 10-day-old intracanal Enterococcus faecalis biofilm. Biofilms were formed inside the root canals and divided into 7 groups (n13): 0.5% NaOCl + Er,Cr:YSGG; Saline + Er,Cr:YSGG; 0.5% NaOCl + syringe irrigation(SI); 2.5% NaOCl + SI; 5% NaOCl + SI; positive and negative controls. Bacterial survivors were counted and specimens visualized under scanning electron and confocal laser scanning microscopy. Treatments with 0.5% NaOCl + Er,Cr:YSGG and 2.5% NaOCl + SI gave a significant reduction in the number of CFU/mm2. Moreover, scanning electron microscopy and confocal laser scanning microscopy imaging confirmed and reinforced bacteriological data. Thus, Er,Cr:YSGG LAI proved to be able to improve the intracanal distribution of 0.5% NaOCl after 60 s of activation, reaching the same level of effectiveness than 2.5% NaOCl. This is regarded as of clinical interest, since working with lower concentrations may contribute to reduce undesired effects.

scholarly journals Effectiveness of chitosan-propolis nanoparticle against Enterococcus faecalis biofilms in the root canal

2020 ◽  
Vol 20 (1) ◽  
Author(s):  
Abhishek Parolia ◽  
Haresh Kumar ◽  
Srinivasan Ramamurthy ◽  
Fabian Davamani ◽  
Allan Pau
Keyword(s):  
Enterococcus Faecalis ◽  
Root Canal ◽  
Laser Scanning ◽  
Current Trend ◽  
Laser Scanning Microscopy ◽  
Successful Outcome ◽  
Confocal Laser ◽  
Internal Diameter ◽  
Sem Images ◽  
Group I

Abstract Background The successful outcome of endodontic treatment depends on controlling the intra-radicular microbial biofilm by effective instrumentation and disinfection using various irrigants and intracanal medicaments. Instrumentation alone cannot effectively debride the root canals specially due to the complex morphology of the root canal system. A number of antibiotics and surfactants are being widely used in the treatment of biofilms however, the current trend is towards identification of natural products in disinfection. The aim of the study was to determine the antibacterial effect of chitosan-propolis nanoparticle (CPN) as an intracanal medicament against Enterococcus faecalis biofilm in root canal. Methods 240 extracted human teeth were sectioned to obtain 6 mm of the middle third of the root. The root canal was enlarged to an internal diameter of 0.9 mm. The specimens were inoculated with E. faecalis for 21 days. Following this, specimens were randomly divided into eight groups (n = 30) according to the intracanal medicament placed: group I: saline, group II: chitosan, group III: propolis100 µg/ml (P100), group IV: propolis 250 µg/ml (P250), group V: chitosan-propolis nanoparticle 100 µg/ml (CPN100), group VI: chitosan-propolis nanoparticle 250 µg/ml (CPN250), group VII: calcium hydroxide(CH) and group VIII: 2% chlorhexidine (CHX) gel. Dentine shavings were collected at 200 and 400 μm depths, and total numbers of CFUs were determined at the end of day one, three and seven. The non-parametric Kruskal Wallis and Mann–Whitney tests were used to compare the differences in reduction of CFUs between all groups and probability values of p < 0.05 were set as the reference for statistically significant results. The scanning electron microscope (SEM) and confocal laser scanning microscopy (CLSM) were also performed after exposure to CPNs. The effectiveness of CPNs were also evaluated against E. faecalis isolated obtained from patients having failed root canal treatment. Results The treatments of chitosan, P100, P250, CPN100, CPN250, CH and 2% CHX reduced the CFUs significantly compared to saline (p < .05). On day one and three, at 200 and 400-μm, CPN250 showed significant reduction of CFUs compared to all other groups (p < .05), while CPN100 was significantly better than other groups (p < .05) except CPN250 and 2% CHX. On day seven, at 200-μm CPN250 showed significant reduction of CFUs compared to all other groups (p < .05) except CPN100 and CHX, while at 400 μm CPN250 showed similar effectiveness as CPN100, CH and 2% CHX. SEM images showed root canal dentin treated with CPN250 had less coverage with E. faecalis bacteria similarly, CLSM images also showed higher percentage of dead E. faecalis bacteria with CPN250 than to CPN100. Conclusion CPN250 was the most effective in reducing E. faecalis colonies on day one, three at both depths and at day seven CPN250 was equally effective as CPN100 and 2% CHX.

Antibacterial effect of propolis nanoparticles against Enterococcus faecalis biofilm in the root canal

2019 ◽  
Author(s):  
Abhishek Parolia ◽  
Haresh Kumar Kumar ◽  
Srinivasan Ramamurthy Ramamurthy ◽  
Allan Pau
Keyword(s):  
Enterococcus Faecalis ◽  
Root Canal ◽  
Laser Scanning ◽  
Antibacterial Effect ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Internal Diameter ◽  
Sem Images ◽  
Group I ◽  
Significant Difference

Abstract Background: To determine the antibacterial effect of propolis nanoparticles (PNs) as an endodontic irrigant against Enterococcus faecalis biofilm in root canal. Methods: PNs were prepared by ultrasonication and the particle size distribution and polydispersity index were determined by dynamic light scattering using Zetasizer Nano S90. 210 extracted human teeth were sectioned to obtain 6mm of the middle third of the root. The root canal was enlarged to an internal diameter of 0.9mm. The specimens were inoculated with E. faecalis for 21 days. Following this, specimens were randomly divided into seven groups having 30 dentinal blocks in each group including group I: saline, group II: propolis 100µg/ml, group III: propolis 300µg/ml, group IV: propolis nanoparticle 100µg/ml, group V: propolis nanoparticle 300µg/ml, group VI: 6% sodium hypochlorite, group VII: 2% chlorhexidine. Dentine shavings were collected at 200 and 400 μm depths, and total numbers of CFUs were determined at the end of one, five, and ten minutes. The non-parametric Kruskal Wallis and Mann-Whitney tests were used to compare the differences in reduction of CFUs between all groups and probability values of P < 0.05 were set as the reference for statistically significant results. The scanning electron microscope and confocal laser scanning microscopy were also performed after exposure to PNs. Results: PN300 was significantly more effective in reducing CFUs compared to all other groups (p <0.05) except 6% NaOCl and 2% CHX (p >0.05) at all-time intervals and both depths. At five minutes, 6% NaOCl and 2 % CHX were the most effective in reducing CFUs (p <0.05) however, no significant difference was found in between PN300, 6% NaOCl and 2 % CHX at 10 minutes (p >0.05). SEM images also showed the maximum reduction of E. faecalis with PN300, 6% NaOCl and 2% CHX (>90 %) at five and ten minutes. CLSM images showed the number of dead cells in dentin was highest with PN300 (>90%) compared to PN100 (>40%) and saline (all live cells). Conclusion: PN300 was equally effective as 6% NaOCl, and 2% CHX in reducing E. faecalis CFUs after one minute, five and ten minutes at both depths.

scholarly journals Effect of reciprocating instrumentation on chlorhexidine substantivity on human dentin: chemical analysis followed by confocal laser microscopy

2021 ◽  
Vol 37 ◽  
pp. e37038
Author(s):  
Matheus Albino Souza ◽  
Fernanda Duda Bonatto ◽  
Afonso Cristiano Fleck da Silva ◽  
Ezequiel Santin Gabrielli ◽  
Felipe Trentin Motter ◽  
...  
Keyword(s):  
High Performance ◽  
Laser Scanning ◽  
Ex Vivo ◽  
Experimental Tests ◽  
Laser Scanning Microscopy ◽  
Live Cells ◽  
Confocal Laser ◽  
Water Control ◽  
Human Teeth ◽  
Root Canals

The present research analyzed the reciprocating instrumentation associated to chlorhexidine (CHX) substantivity as its correlation with E. faecalis viability in ex vivo root canals. Eighty extracted single-rooted human teeth were used, being 40 to high-performance liquid chromatography (HPLC) and 40 to confocal laser scanning microscopy (CLSM). In both, teeth were decoronated and the cervical third was prepared. In the CLSM analysis, the root canals were inoculated with E. faecalis for 14 days. Samples were divided into 4 groups (n=10) according to instrumentation technique: no instrumentation and irrigation with distilled water (control); manual instrumentation (K-File); rotary instrumentation (ProTaper Next); and reciprocating instrumentation (Reciproc R25). Two percent chlorhexidine was applied as irrigating substance in experimental groups. Longitudinal grooves resulted in 2 halves root and 20 proof bodies in each group. Samples were divided by chance in two groups (n=10) and the outcomes were evaluated after two days and one week. The retained chlorhexidine and live cells after instrumentation techniques in each evaluation time was measured by HPLC and CLSM, respectively. Specific analysis was applied for experimental tests (p≤0.05). Both rotary as well as reciprocating techniques significantly reduced the amount of chlorhexidine on dentin in all observation periods (p<0.05). After evaluation times, all experimental groups presented lower live cells compared to control, but without statistically difference. Intragroup comparisons in times of evaluation showed no differences in instrumentation techniques, in chlorhexidine retention and number of live cells (p>0.05). Reciprocating instrumentation does not interfere on chlorhexidine substantivity.

scholarly journals Use of electromagnetic stimulation on an Enterococcus faecalis biofilm on root canal treated teeth in vitro

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Beatriz H. D. Panariello ◽  
Justin K. Kindler ◽  
Kenneth J. Spolnik ◽  
Ygal Ehrlich ◽  
George J. Eckert ◽  
...  
Keyword(s):  
Enterococcus Faecalis ◽  
Root Canal ◽  
Laser Scanning ◽  
Confocal Imaging ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Microscopy Imaging ◽  
Electromagnetic Stimulation ◽  
Root Canal Disinfection

AbstractRoot canal disinfection is of utmost importance in the success of the treatment, thus, a novel method for achieving root canal disinfection by electromagnetic waves, creating a synergistic reaction via electric and thermal energy, was created. To study electromagnetic stimulation (EMS) for the disinfection of root canal in vitro, single rooted teeth were instrumented with a 45.05 Wave One Gold reciprocating file. Specimens were sterilized and inoculated with Enterococcus faecalis ATCC 29,212, which grew for 15 days to form an established biofilm. Samples were treated with 6% sodium hypochlorite (NaOCl), 1.5% NaOCl 1.5% NaOCl with EMS, 0.9% saline with EMS or 0.9% saline. After treatments, the colony forming units (CFU) was determined. Data was analyzed by Wilcoxon Rank Sums Test (α = 0.05). One sample per group was scored and split for confocal laser scanning microscopy imaging. There was a significant effect with the use of NaOCl with or without EMS versus 0.9% saline with or without EMS (p = 0.012 and 0.003, respectively). CFUs were lower when using 0.9% saline with EMS versus 0.9% saline alone (p = 0.002). Confocal imaging confirmed CFU findings. EMS with saline has an antibiofilm effect against E. faecalis and can potentially be applied for endodontic disinfection.

scholarly journals Effect of Sodium Hypochlorite and Benzalkonium Chloride on the Structural Parameters of the Biofilms Formed by Ten Salmonella enterica Serotypes

Pathogens ◽  
2019 ◽  
Vol 8 (3) ◽  
pp. 154 ◽  
Author(s):  
Rosa Capita ◽  
Silvia Fernández-Pérez ◽  
Laura Buzón-Durán ◽  
Carlos Alonso-Calleja
Keyword(s):  
Sodium Hypochlorite ◽  
Salmonella Enterica ◽  
Benzalkonium Chloride ◽  
Laser Scanning ◽  
Surface Coverage ◽  
Structural Parameters ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Low Concentrations ◽  
Practical Field

The influence of the strain on the ability of Salmonella enterica to form biofilms on polystyrene was investigated by confocal laser scanning microscopy. The effects of sodium hypochlorite with 10% active chlorine (SHY; 25,000, 50,000, or 100,000 ppm), and benzalkonium chloride (BZK; 1000, 5000, or 10,000 ppm) on twenty-four-hour-old biofilms was also determined. The biofilms of ten Salmonella enterica isolates from poultry (S. Agona, S. Anatum, S. Enteritidis, S. Hadar, S. Infantis, S. Kentucky, S. Thompson, S. Typhimurium, monophasic variant of S. Typhimurium 1,4,(5),12:i:-, and S. Virchow) were studied. Biofilms produced by S. Anatum, S. Hadar, S. Kentucky, and S. Typhimurium showed a trend to have the largest biovolume and the greatest surface coverage and thickness. The smallest biofilms (P < 0.01) in the observation field (14.2 × 103 µm2) were produced by S. Enteritidis and S. 1,4,(5),12:i:- (average 12.9 × 103 ± 9.3 × 103 µm3) compared to the rest of the serotypes (44.4 × 103 ± 24.7 × 103 µm3). Biovolume and surface coverage decreased after exposure for ten minutes to SHY at 50,000 or 100,000 ppm and to BZK at 5000 or 10,000 ppm. However, the lowest concentrations of disinfectants increased biovolume and surface coverage in biofilms of several strains (markedly so in the case of BZK). The results from this study suggest that the use of biocides at low concentrations could represent a public health risk. Further research studies under practical field conditions should be appropriate to confirm these findings.

scholarly journals Percentage of Gutta-Percha-, Sealer-, and Void-Filled Areas in Oval-Shaped Root Canals Obturated with Different Filling Techniques: A Confocal Laser Scanning Microscopy Study

2020 ◽  
Vol 14 (01) ◽  
pp. 008-012
Author(s):  
Vinicio Hidemitsu Goto Hirai ◽  
Ricardo Machado ◽  
Maria Carolina Lucato Budziak ◽  
Lucila Piasecki ◽  
Alexandre Kowalczuck ◽  
...  
Keyword(s):  
Confocal Laser Scanning Microscopy ◽  
Laser Scanning ◽  
Continuous Wave ◽  
Laser Scanning Microscopy ◽  
Confocal Laser Scanning ◽  
Scanning Microscopy ◽  
Confocal Laser ◽  
Root Canals ◽  
Gutta Percha

Abstract Objective This study compared different obturation techniques, analyzing percentage of areas filled with gutta-percha, sealer, and voids (PGFA, PSFA, and PVFA, respectively) in oval-shaped root canals. Materials and Methods A total of 60 extracted human mandibular central incisors were decoronated, instrumented, and irrigated using the same protocol. After drying, the root canal was filled with AH Plus labeled with 0.1% rhodamine B dye using a Lentulo spiral. The filling procedure was performed by dividing the teeth into four groups according to the respective technique: G1, cold lateral condensation; G2, continuous wave of condensation; G3, modified cold lateral condensation using an F3 master cone; and G4, modified continuous wave of condensation using an ISO (International Organization for Standardization) sized 30 gutta-percha cone. Then, slices measuring 1.5 mm in thickness were obtained 3 and 6 mm from the apex and evaluated by confocal laser scanning microscopy to determine PGFA, PSFA, and PVFA. Statistical Analysis The data were analyzed statistically with analysis of variance and Games-Howell’s tests (p = 0.05). Results The groups showed no significant differences in the apical third (3 mm from the apex). In the middle third (6 mm from the apex), G3 and G1 showed higher PGFA and PVFA, respectively. G3 showed lower PSFA than G2 and G4. Both cold techniques (G1 and G3) promoted lower PSFA than both warm techniques (G2 and G4). Conclusions Notwithstanding the limitations of this in vitro study, PGFA, PSFA, and PVFA ranged significantly only in the middle third, as observed by the different filling techniques. Higher PGFA and PVFA values were obtained for G3 and G1, respectively. Both cold techniques promoted lower PSFA than both warm techniques.

Testate Amoebae Examined by Confocal and Two-Photon Microscopy: Implications for Taxonomy and Ecophysiology

2010 ◽  
Vol 16 (6) ◽  
pp. 735-746 ◽  
Author(s):  
Zuzana Burdíková ◽  
Martin Čapek ◽  
Pavel Ostašov ◽  
Jiří Machač ◽  
Radek Pelc ◽  
...  
Keyword(s):  
Laser Scanning ◽  
Three Dimensional ◽  
Environmental Scanning Electron Microscopy ◽  
Testate Amoebae ◽  
Shell Morphology ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Polarization Microscopy ◽  
Sem Images ◽  
Two Photon

AbstractTestate amoebae (TA) are a group of free-living protozoa, important in ecology and paleoecology. Testate amoebae taxonomy is mainly based on the morphological features of the shell, as examined by means of light microscopy or (environmental) scanning electron microscopy (SEM/ESEM). We explored the potential applications of confocal laser scanning microscopy (CLSM), two photon excitation microscopy (TPEM), phase contrast, differential interference contrast (DIC Nomarski), and polarization microscopy to visualize TA shells and inner structures of living cells, which is not possible by SEM or environmental SEM. Images captured by CLSM and TPEM were utilized to create three-dimensional (3D) visualizations and to evaluate biovolume inside the shell by stereological methods, to assess the function of TA in ecosystems. This approach broadens the understanding of TA cell and shell morphology, and inner structures including organelles and endosymbionts, with potential implications in taxonomy and ecophysiology.

Interspecies Interactions in Dual-Species Biofilms Formed by Psychrotrophic Bacteria and the Tolerance of Sessile Communities to Disinfectants

2020 ◽  
Vol 83 (6) ◽  
pp. 951-958 ◽  
Author(s):  
LEI YUAN ◽  
NI WANG ◽  
FAIZAN A. SADIQ ◽  
GUOQING HE
Keyword(s):  
Hydrogen Peroxide ◽  
Sodium Hypochlorite ◽  
Peracetic Acid ◽  
Laser Scanning ◽  
Structural Changes ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Interspecies Interactions ◽  
Mixed Species ◽  
Psychrotrophic Bacteria

ABSTRACT Biofilms on the surface of food processing equipment act as potential reservoirs of microbial contamination. Bacterial interactions are believed to play key roles in both biofilm formation and antimicrobial tolerance. In this study, Aeromonas hydrophila, Chryseobacterium oncorhynchi, and Pseudomonas libanensis, which were previously isolated from Chinese raw milk samples, were selected to establish two dual-species biofilm models (P. libanensis plus A. hydrophila and P. libanensis plus C. oncorhynchi) on stainless steel at 7°C. Subsequently, three disinfectants, hydrogen peroxide (100 ppm), peracetic acid (100 ppm), and sodium hypochlorite (100 ppm), were used to treat the developed sessile communities for 10 min. Structural changes after exposure to disinfectants were analyzed with confocal laser scanning microscopy. The cell numbers of both A. hydrophila and C. oncorhynchi recovered from surfaces increased when grown as dual species biofilms with P. libanensis. Dual-species biofilms were more tolerant of disinfectants than were each single-species biofilm. Peracetic acid was the most effective disinfectant for removing biofilms, followed by hydrogen peroxide and sodium hypochlorite. The results expand the knowledge of mixed-species biofilms formed by psychrotrophic bacteria and will be helpful for developing effective strategies to eliminate bacterial mixed-species biofilms. HIGHLIGHTS

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