Aim To investigate the influence of biofilm structure on the biofilm removal capacity of endodontic irrigants and to study changes in the architecture of the remaining biofilms. Methodology Streptococcus oralis J22 and Actinomyces naeslundii T14V‐J1 were cocultured under different growth conditions on saliva‐coated hydroxyapatite discs. A constant depth film fermenter (CDFF) was used to grow steady‐state 4‐day biofilms. Biofilms were grown under static conditions for 4 and 10 days within a confined space. Twenty microlitres of 2% NaOCl, 2% Chlorhexidine (CHX), 17% Ethylene‐diamine‐tetra‐acetic acid (EDTA) and buffer were applied statically on the biofilms for 60 s. Biofilm removal was evaluated with optical coherence tomography (OCT). Post‐treated biofilms were assessed via low load compression testing (LLCT) and Confocal laser scanning microscopy (CLSM). Optical coherence tomography data were analysed through a two‐way analysis of variance (ANOVA). Low load compression testing and CLSM data were analysed through one‐way ANOVA and Dunnett's post hoc test. The level of significance was set at a < 0.05. Results The initial biofilm structure affected the biofilm removal capacity of the irrigants. NaOCl demonstrated the greatest chemical efficacy against the biofilms and was significantly more effective on the static than the CDFF biofilms (P < 0.001). CHX was ineffective and caused a rearrangement of the biofilm structure. Ethylene‐diamine‐tetra‐acetic acid exhibited a distinct removal effect only on the CDFF biofilms. Biofilm age influenced the structure of the remaining biofilms. The 4‐day grown remaining biofilms had a significantly different viscoelastic pattern compared to the respective 10‐day grown biofilms (P ≤ 0.01), especially in the NaOCl‐treated group. Confocal laser scanning microscopy analysis confirmed the CHX‐induced biofilm structural rearrangement. Conclusions Biofilm structure is an influential factor on the chemical efficacy of endodontic irrigants. Optical coherence tomography allows biofilm removal characteristics to be studied. NaOCl should remain the primary irrigant. Ethylene‐diamine‐tetra‐acetic acid was effective against cell‐rich/EPS‐poor biofilms. Chlorhexidine did not remove biofilm, but rather rearranged its structure.

The results obtained in this study suggest that students who used the DLO performed better than those who used conventional methods. This suggests that the DLO may be a useful teaching tool for dentistry undergraduates, on distance learning courses and as a complementary tool in face-to-face teaching.

Aim To study the influence of time and volume of 2% sodium hypochlorite (NaOCl) on biofilm removal and to investigate the changes induced on the biofilm architecture. Steady‐state, dual‐species biofilms of standardized thickness and a realistic contact surface area between biofilms and NaOCl were used. Methodology Streptococcus oralis J22 and Actinomyces naeslundii T14V‐J1 biofilms were grown on saliva‐coated hydroxyapatite discs within sample holders in the Constant Depth Film Fermenter (CDFF) for 96 h. Two per cent NaOCl was statically applied for three different time intervals (60, 120 and 300 s) and in two different volumes (20 and 40 μL) over the biofilm samples. The diffusion‐driven effects of time and volume on biofilm disruption and dissolution were assessed with Optical Coherence Tomography (OCT). Structural changes of the biofilms treated with 2% NaOCl were studied with Confocal Laser Scanning Microscopy (CLSM) and Low Load Compression Testing (LLCT). A two‐way analysis of variance (2‐way anova) was performed, enabling the effect of each independent variable as well as their interaction on the outcome measures. Results Optical coherence tomography revealed that by increasing the exposure time and volume of 2% NaOCl, both biofilm disruption and dissolution significantly increased. Analysis of the interaction between the two independent variables revealed that by increasing the volume of 2% NaOCl, significant biofilm dissolution could be achieved in less time. Examination of the architecture of the remaining biofilms corroborated the EPS‐lytic action of 2% NaOCl, especially when greater volumes were applied. The viscoelastic analysis of the 2% NaOCl‐treated biofilms revealed that the preceding application of higher volumes could impact their subsequent removal. Conclusions Time and volume of 2% NaOCl application should be taken into account for maximizing the anti‐biofilm efficiency of the irrigant and devising targeted disinfecting regimes against remaining biofilms.

Aim To investigate the anti‐biofilm efficacy and working mechanism of several NaOCl concentrations on dual‐species biofilms of different architecture as well as the changes induced on the architecture of the remaining biofilms. Methodology Streptococcus oralis J22 and Actinomyces naeslundii T14V‐J1 were co‐cultured under different growth conditions on saliva‐coated hydroxyapatite discs. A constant‐depth film fermenter (CDFF) was used to grow steady‐state, four‐day mature biofilms (dense architecture). Biofilms were grown under static conditions for 4 days within a confined space (less dense architecture). Twenty microlitres of buffer, 2‐, 5‐, and 10% NaOCl were applied statically on the biofilms for 60 s. Biofilm disruption and dissolution, as well as bubble formation, were evaluated with optical coherence tomography (OCT). The viscoelastic profile of the biofilms post‐treatment was assessed with low load compression testing (LLCT). The bacteria/extracellular polysaccharide (EPS) content of the biofilms was examined through confocal laser scanning microscopy (CLSM). OCT, LLCT and CLSM data were analysed through one‐way analysis of variance (ANOVA) and Tukey’s HSD post‐hoc test. Linear regression analysis was performed to test the correlation between bubble formation and NaOCl concentration. The level of significance was set at a < 0.05. Results The experimental hypothesis according to which enhanced biofilm disruption, dissolution and bubble formation were anticipated with increasing NaOCl concentration was generally confirmed in both biofilm types. Distinct differences between the two biofilm types were noted with regard to NaOCl anti‐biofilm efficiency as well as the effect that the several NaOCl concentrations had on the viscoelasticity profile and the bacteria/EPS content. Along with the bubble generation patterns observed, these led to the formulation of a concentration and biofilm structure‐dependent theory of biofilm removal. Conclusions Biofilm architecture seems to be an additional determining factor of the penetration capacity of NaOCl, and consequently of its anti‐biofilm efficiency.

The aim of the study was to assess the penetrability of two endodontic sealers (AH Plus and MTA Fillapex) into dentinal tubules, submitted to endodontic treatment and subsequently to endodontic retreatment. Thirty ex vivo incisors were prepared using ProTaper rotary system up to F3 instrument and divided in three groups according to the endodontic sealer used for root canal filling: AH Plus (AHP), MTA Fillapex (MTAF), and control group (CG) without using EDTA previously to the root canal filling. Rhodamine B dye (red) was incorporated to the sealers in order to provide the fluorescence which will enable confocal laser scanning microscopy (CLSM) assessment. All specimens were filled with gutta-percha cones using the lateral compaction technique. The specimens were submitted to endodontic retreatment using ProTaper Retreatment system, re-prepared up to F5 instruments and filled with gutta-percha cones and the same sealer used during endodontic retreatment. Fluorescein dye (green) was incorporated to the sealer in order to distinguish from the first filling. The roots were sectioned 2 mm from the apex and assessed by CLSM. No difference was found between the two experimental groups (P > 0.05). On the other hand, in the control group the sealers were not capable to penetrate into dentinal tubules after endodontic treatment (P > 0.05). In retreatment cases, none of the sealers were able to penetrate into dentin tubules. It can be concluded that sealer penetrability is high during endodontic treatment. However, MTA Fillapex and AH Plus do not penetrate into dentinal tubules after endodontic retreatment.