Root Microbiota in Primary and Secondary Apical Periodontitis

Apical periodontitis is an inflammatory disease of the dental periradicular tissues triggered by bacteria colonizing necrotic root canals. Primary apical periodontitis results from the microbial colonization of necrotic pulp tissues. Secondary apical periodontitis results from a persistent infection of incorrectly treated root canals. The aim of this study was to characterize the microbiota present in primary and secondary intraradicular infections associated with apical periodontitis using 16S rRNA gene amplicon sequencing. Teeth exhibiting apical periodontitis with or without root canal treatment were extracted after informed consent. From each tooth, the intraradicular content as well as a dentin sample (control) were collected and subjected to DNA extraction. PCR amplicons of the V3–V4 region of the bacterial 16S rRNA gene were pooled and sequenced (2 × 300) on an Illumina MiSeq instrument. The bioinformatics analysis pipeline included quality filtering, merging of forward and reverse reads, clustering of reads into operational taxonomic units (OTUs), removal of putative contaminant OTUs and assigning taxonomy. The most prevalent and abundant OTU in both dentin and root canal samples was assigned to anaerobic bacterium Fusobacterium nucleatum. Multivariate analysis showed clustering of microbiota by sample type (dentin vs. intraradicular content) and, in root canals, by pathology (primary vs. secondary infection). The proportions of Enterococcus faecalis and F. nucleatum were, respectively, higher and lower when comparing secondary to primary infected root canals. Co-occurrence network analysis provided evidence of microbial interactions specific to the infection type. The identification of bacterial taxa differentially abundant in primary and secondary intraradicular infections may provide the basis for targeted therapeutic approaches aimed at reducing the incidence of apical periodontitis

Oxidative Stress in Bacteria Measured by Flow Cytometry

Cytométrie en flux pour la mesure du stress oxydatif dans les bactérie

Enzyme-mediated photoinactivation of Enterococcus faecalis using Rose Bengal-acetate

Rose Bengal-acetate (RB-Ac) is a pro-photosensitizer claimed to diffuse into target cells, where the acetate groups are hydrolyzed and the photosensitizing properties of Rose Bengal (RB) are restored. Despite promising results on tumor cells, the interaction of RB-Ac with bacteria has never been investigated. This study aimed to assess the interaction of RB-Ac with Enterococcus faecalis and to evaluate its potential use in antimicrobial photodynamic therapy (aPDT). Spectrofluorometry was used to assess the ability of E. faecalis to hydrolyze the RB-Ac compound. Fluorescence microscopy was employed to observe the distribution and to evaluate the cellular uptake of the RB produced. The antibacterial efficiency of RB-Ac-mediated aPDT was assessed by flow cytometry in combination with the LIVE/DEAD® staining. Results showed that RB-Ac was successfully hydrolyzed in the presence of E. faecalis cells. The RB produced appeared to incorporate the membrane of bacteria. Higher concentrations of RB-Ac resulted in higher incorporation of RB. The blue-light irradiation of RB-Ac-treated samples significantly reduced bacterial viability. Less than 0.01% of E. faecalis survived after incubation with 200 μM RB-Ac during 900 min and blue-light activation. The current report indicates that E. faecalis cells can hydrolyze the RB-Ac compound to produce active RB. The use of RB-Ac did not appear to allow cytoplasmic internalization of the RB produced, which rather incorporated the membrane bilayers of E. faecalis. The use of RB-Ac did not provide additional advantages over RB in terms of PS localization. Nonetheless, sufficient RB was produced and incorporated into the membranes of bacteria to elicit effective aPDT

Influence of different surface treatments on marginal adaptation in enamel and dentin

To compare marginal adaptation in enamel and dentin after different surface treatments before and after long-term simultaneous thermal and mechanical stresses in a mixed Class V restoration

Influence of high-power LED on composite hardness

To compare the surface hardness of a composite (Z250, 3MEspe) in standardized cavities, 6mm in diameter, in extracted human teeth after polymerization with A. high-power LED for 10s direct and 20s indirect (Freelight II, 3MEspe), B. conventional halogen for 20s direct and 40s indirect (Trilight, 3MEspe) and C. high power halogen for 10s direct and 20s indirect (Optilux 501 Turbo, SDSKerr)

Accuracy of electronic apex locater-controlled handpieces

An in vitro model was used to test a hypothesis that the manual (apex locator) and motor-driven (apex locator-controlled handpiece) operating modes of 3 newly developed apex-locator-controlled handpiece devices (Dentaport ZX, Endomaster, XSmart Dual) give the same working length. The depth of penetration of the file into the root canal was measured using a digital micrometer and the distance of the tip of the file relative to the major root foramen was determined by scanning electron microscopy (SEM). In the manual mode, the XSmart Dual device reported significantly shorter working lengths than the Dentaport ZX or the Endomaster devices. In the motor-driven mode, the XSmart Dual device reported significantly longer working lengths than the Dentaport ZX but not the Endomaster. Most instruments driven by the handpieces were confined within the root canal and differences in working lengths between manual and motor-driven modes were small for all devices (tenths of millimeters). We concluded that although the motor-driven mode of these devices appeared to be clinically safe, measurements obtained in manual and motor-driven operating modes are not equivalent

Long-term sealing ability of Pulp Canal Sealer, AH-Plus, GuttaFlow and Epiphany

AIM: To evaluate the long-term sealing ability of four contemporary endodontic sealers [Pulp Canal Sealer (PCS), AH-Plus, GuttaFlow and Epiphany] using a fluid filtration technique. METHODOLOGY: The palatal roots of 40 human maxillary molar teeth were selected and the root canal was prepared using a crown-down technique (apical size 40, 6% taper). Roots were irrigated with 3% NaOCl, 17% EDTA solution and rinsed with distilled water. Canals were filled with either PCS, AH-Plus, GuttaFlow or Epiphany using a single-cone technique (n = 8). Twenty-four hours after filling, the roots were connected to an automatic flow-recording device (Flodec System) filled with double-distilled water under pressure (0.2 bar) to measure leakage. Flow rates were assessed at 6, 12 or 24-h and after 1-year of storage. RESULTS: None of the materials fully prevented fluid flow. Fluid flow decreased after 6 h and decreased further after 12 h. After 24 h, PCS and AH-Plus allowed significantly more fluid flow than GuttaFlow and Epiphany. After 1 year, PCS allowed significantly more fluid flow than the other materials. No significant changes in leakage occurred between 24 h and 1 year. CONCLUSIONS: GuttaFlow and Epiphany allowed less fluid movement along filled straight roots

Rose bengal uptake by E. faecalis and F. nucleatum and light-mediated antibacterial activity measured by flow cytometry

Antibacterial photodynamic therapy (aPDT) using rose bengal (RB) and blue-light kills bacteria through the production of reactive oxygen derivates. However, the interaction mechanism of RB with bacterial cells remains unclear. This study investigated the uptake efficiency and the antibacterial activity of blue light-activated RB against Enterococcus faecalis and Fusobacterium nucleatum. Spectrophotometry and epifluorescence microscopy were used to evaluate binding of RB to bacteria. The antibacterial activity of RB after various irradiation times was assessed by flow cytometry in combination with cell sorting. Uptake of RB increased in a concentration dependent manner in both strains although E. faecalis displayed higher uptake values. RB appeared to bind specific sites located at the cellular poles of E. faecalis and at regular intervals along F. nucleatum. Blue-light irradiation of samples incubated with RB significantly reduced bacterial viability. After incubation with 10μM RB and 240s irradiation, only 0.01% (±0.01%) of E. faecalis cells and 0.03% (±0.03%) of F. nucleatum survived after treatment. This study indicated that RB can bind to E. faecalis and F. nucleatum in a sufficient amount to elicit effective aPDT. Epifluorescence microscopy showed a yet-unreported property of RB binding to bacterial membranes. Flow cytometry allowed the detection of bacteria with damaged membranes that were unable to form colonies on agars after cell sorting