Amplicon-based next-generation sequencing for comparative analysis of root canal microbiome of teeth with primary and persistent/secondary endodontic infections.

OBJECTIVES: To compare the root canal microbiome profiles of primary and persistent/secondary infections using high-throughput sequencing with the help of a reliable bioinformatics algorithm. MATERIALS AND METHODS: Root canal samples of 10 teeth in the primary endodontic infection (PEI) group and 10 teeth in the persistent/secondary endodontic infection (SEI) group were included resulting in a total of 20 samples. After DNA extraction from the samples, sequencing was performed on the Illumina MiSeq platform. Pair-end Illumina reads were imported to QIIME 2; amplicon sequence variants (ASVs) generated by DADA2 were mapped to GreenGenes database. Weighted UniFrac distances were calculated and principal coordinates analysis (PCoA) was used to compare beta diversity patterns. The multiple response permutation procedure (MRPP), the analysis of similarities (ANOSIM), and permutational multivariate analysis of variance (adonis) were conducted for testing group differences. Linear discriminant analysis effect size (LEfSe) analysis was utilized to identify differentially abundant taxa between the groups. The linear discriminant analysis (LDA) score threshold was set to 4.0. RESULTS: Within the Gram-negative facultative anaerobic Gammaproteobacteria class outgroup, two orders (Pasteurellales, Vibrionales) and two families (Pasteurellaceae, Vibrionaceae) were significantly more abundant in the PEI group, whereas Gram-positive bacteria, Actinomycetales order, and Gram-positive anaerobic taxa, one genus (Olsenella) and one species (Olsenella uli), were identified as significantly more abundant in the SEI group. CONCLUSIONS: A few taxa were differentially abundant within either the PEI or SEI group. CLINICAL RELEVANCE: Reliable bioinformatic tools are needed to define microbial profiles of endodontic infections. Based on a limited number of samples, no distinct variation was determined between the bacterial diversity of initial and recurrent endodontic infections.

Microbiota of Endodontically Infected Primary and Permanent Teeth.

Purpose: Differences in the endodontic microbiome of permanent and primary teeth during the mixed dentition period are still unknown. The purpose of this study was to examine bacterial diversity in endodontically infected primary and permanent teeth using 16S rRNA gene sequencing and the QIIME 2 (Quantitative Insights Into Microbial Ecology 2) bioinformatics pipeline. Methods: Microbial samples from endodontically infected primary (n equals 15) and permanent (n equals 15) maxillary or mandibular molar teeth were subjected to next-generation sequencing analysis based on examination of the hypervariable V3 to V4 region of the 16S rRNA gene. Statistical analysis was performed using R software. Results: Of 1,664,926 reads and 2,237 operational taxonomic units, 14 phyla, 89 families, and 236 genera were identified. Firmicutes were the most commonly detected phyla in both endodontically infected primary and permanent root canals. Bacteroides and Proteobacteria were more common in primary teeth, whereas Actinobacteria and Verrucomicrobia were more common in permanent teeth. The overall canal microbiota composition was similar in endodontically infected primary and permanent teeth (P=0.338). Conclusions: This study provides a comprehensive assessment of microbiota composition in endodontically infected primary and permanent teeth and gives a deeper insight into the origin of the root canal infections.

Evaluation of dentinal defects during root canal preparation using thermomechanically processed nickel-titanium files.

OBJECTIVE: The aim of this study was to compare the incidence of root cracks after root canal instrumentation with thermomechanically processed nickel-titanium (Ni-Ti) files with different instrumentation kinematics. MATERIALS AND METHODS: A total of 150 extracted mandibular premolars with mature apices and straight root canals were divided into five groups and used in this study. In Group 1, 30 teeth were prepared using hand K-files and assigned to control group, Group 2 was instrumented using K3XF Rotary files (SybronEndo, Glendora, CA, USA) with continuous rotary motion. The teeth in Group 3 were instrumented by ProTaper Next (Dentsply Maillefer, Ballaigues, Switzerland) rotary files which make asymmetric rotary motion, In Group 4, teeth were instrumented by RECIPROC (VDW, Munich, Germany) with reciprocation motion and in Group 5, teeth were instrumented by Twisted File (TF) Adaptive (SybronEndo, Orange, CA, USA) files that use combination of continuous rotation and reciprocation motion (n = 30/per group). All the roots were horizontally sectioned 3, 6, and 9 mm from the apex with a low speed saw under water cooling. Then, the slices were examined through a stereomicroscope to determine the presence of dentinal microcracks. RESULTS: For the apical (3-mm) and coronal (9-mm) sections, the ProTaper Next and TF Adaptive produced significantly more cracks than the hand files, RECIPROC, and K3XF (P < 0.05). There was no significant difference between the experimental groups and control group at the 6-mm level (P > 0.05). CONCLUSIONS: Within the limitations of this in vitro study, all thermal-treated Ni-Ti instruments and hand files caused microcracks in root canal dentin.

The effects of different nickel-titanium instruments on dentinal microcrack formations during root canal preparation.

OBJECTIVE: The aim of the present study was to investigate the incidence of dentinal microcracks caused by different preparation techniques. MATERIALS AND METHODS: 120 extracted human mandibular incisor teeth were divided into five experimental groups and one control group (n = 20): Group 1: Hand preparation with balanced force technique up to #25 K-file. Group 2: Preparation with only ProTaper F2 instrument in a reciprocating movement. Group 3: Preparation with Reciproc R25 instrument in a reciprocating movement. Group 4: Preparation with ProTaper instruments up to F2 instrument. Group 5: Preparation with ProTaper Next instruments up to X2 instrument. No procedure was applied to control group. The roots were sectioned horizontally at 3, 6 and 9 mm from the apex and examined. Absence or presence of dentinal microcracks was noted. RESULTS: The Chi-square test was performed to compare the appearance of cracked roots between all groups. There were no significant differences among the groups (P > 0.05). CONCLUSIONS: In conclusion, except the hand file and control group, all experimental groups showed microcrack formations.

The effect of various backfilling techniques on the fracture resistance of simulated immature teeth performed apical plug with Biodentine.

AIM: To evaluate the fracture resistance of simulated immature teeth that had been backfilled using different materials after using Biodentine as the apical plug material. DESIGN: Seventy-five single-rooted teeth were divided into five groups (n = 15). The 15 teeth in group 1 served as a negative control group and received no treatment. The remaining 60 teeth were instrumented to a #6 Peeso reamer to obtain a standard internal diameter of 1.5 mm. The apical 4 mm of 60 teeth was filled with Biodentine. The backfilling was then performed on each group as follows: group 2--no backfilling (positive control), group 3--gutta-percha, group 4--fiber post, and group 5--Biodentine. Specimens were then subjected to fracture testing. The force required to fracture each specimen was recorded, and the data were statistically analyzed. RESULTS: The mean fracture values of groups 1 and 4 were significantly higher than groups 2, 3, and 5 (P < 0.05). The values of groups 3 and 5 were significantly higher than group 2 (P < 0.05). CONCLUSIONS: The backfilling with fiber post after an apical Biodentine plug provided the highest fracture resistance among all experimental groups.

Incidence of apical crack initiation and propagation during the removal of root canal filling material with ProTaper and Mtwo rotary nickel-titanium retreatment instruments and hand files.

INTRODUCTION: The aim of this study was to determine the incidence of crack initiation and propagation in apical root dentin after retreatment procedures performed by using 2 rotary retreatment systems and hand files with additional instrumentation. METHODS: Eighty extracted mandibular premolars with single canals were selected. One millimeter from the apex of each tooth was ground perpendicular to the long axis of the tooth, and the apical surface was polished. Twenty teeth served as the control group, and no preparation was performed. The remaining 60 teeth were prepared to size 35 with rotary files and filled with gutta-percha and AH Plus sealer. Specimens were then divided into 3 groups (n = 20), and retreatment procedures were performed with the following devices and techniques: ProTaper Universal retreatment files, Mtwo retreatment files, and hand files. After retreatment, the additional instrumentation was performed by using size 40 ProTaper, Mtwo, and hand files. Digital images of the apical root surface were recorded before preparation, after instrumentation, after filling, after retreatment, and after additional instrumentation. The images were then inspected for the presence of any new apical cracks and propagation. Data were analyzed with the logistic regression and Fisher exact tests. RESULTS: All experimental groups caused crack initiation and propagation after use of retreatment instruments. The ProTaper and Mtwo retreatment groups caused greater crack initiation and propagation than the hand instrument group (P < .05) after retreatment. Additional instrumentation with ProTaper and Mtwo instruments after the use of retreatment instruments caused crack initiation and propagation, whereas hand files caused neither crack initiation nor propagation (P < .05). CONCLUSIONS: This study showed that retreatment procedures and additional instrumentation after the use of retreatment files may cause crack initiation and propagation in apical dentin.

Effect of agitation of EDTA with 808-nanometer diode laser on removal of smear layer.

INTRODUCTION: Near-infrared diode lasers can be used for several applications, which range from disinfection to smear layer removal in endodontics. This study evaluated the efficacy of agitation of 15% EDTA with an 808-nm diode laser on removal of the smear layer. METHODS: Sixty extracted human maxillary central incisor teeth were instrumented up to ProTaper F4 (Dentsply Maillefer, Ballagues, Switzerland) and then randomly divided into 6 groups (n = 10 for each group) according to the different final irrigating protocols as follows: 5% sodium hypochlorite for 120 seconds performed with the NaviTip (Dentsply Maillefer, Ballaigues, Switzerland) (control group); 15% EDTA for 120 seconds performed with the NaviTip; and agitation of 15% EDTA with an 808-nm diode laser for 10, 20, 30, and 40 seconds. Specimens were observed under a scanning electron microscope, and open dentinal tubules were counted using Adobe Photoshop software (Adobe Systems, San Jose, CA). The data were analyzed with 1-way analysis of variance and Tukey post hoc tests (P = .05). RESULTS: The number of open dentinal tubules was higher in the middle thirds than in the apical thirds. The differences between the apical and middle thirds were statistically significant (P < .05). Statistically significant differences were also found between the control group and the other groups in both the middle and apical thirds of the root canals (P < .05). CONCLUSIONS: The results indicated that agitation of 15% EDTA with an 808-nm diode laser for 20 seconds was effective in removing the smear layer in the apical thirds of root canals.