Aim To assess and compare the microbiome of paired root apices and periapical lesions from cases with failed endodontic treatment and to associate the microbiome and bacterial metabolic pathways in both sites with asymptomatic apical periodontitis (AAP) and symptomatic apical periodontitis (SAP), using next‐generation sequencing (NGS). Methodology Matched root apices and periapical lesions of patients with failed root canal treatments were surgically extracted. Specimens were cryopulverized, bacterial DNA was extracted and the V3–V4 hypervariable regions of the 16 S rRNA gene were amplified and sequenced using the Illumina Miseq platform. Diversity and community composition were studied in the paired samples, as well as in AAP and SAP cases. Diversity indices were compared in each case by means of the Wilcoxon matched‐pairs signed rank and Mann–Whitney U tests. Differences in the community composition were explored with multivariate statistical analysis and Linear discriminant analysis Effect Size (LEfSe). Bacterial functional study was performed through the Phylogenetic Investigation of Communities by Reconstruction of Unobserved States (PICRUSt) analysis. Results Twenty‐one paired apices and lesions were successfully sequenced and analysed, identifying a total of 21 phyla and 600 genera. A higher alpha‐diversity was observed in the periapical lesions, although no global differences in the community composition between the two sites were found (p = .87), the most prevalent genera being Fusobacterium, Porphyromonas and Streptococcus. Prevotella, Clostridiales_vadinBB60_group, Bosea, Phreatobacter, Afipia and Xanthobacteriaceae_unclassified were enriched in SAP samples, while Pseudopropionibacterium, Campylobacter and Peptoniphilus were significantly more abundant in AAP cases (p < .05). Metabolic pathways involved in the amino acid metabolism or degradation and flagellum assembly were more abundant in SAP samples, whereas glucose metabolism‐related pathways were associated with AAP. Conclusions The bacterial community composition was similar in the apices and periapical lesions. The microbiome was different in AAP and SAP samples, gram‐negative bacteria showing higher relative abundances in SAP cases. An association was observed between amino acid degradation and flagellum assembly pathways, and the development of tenderness to percussion or palpation.

AimTo evaluate in a laboratory setting the antibiofilm activity of several irrigating protocols including conventional irrigation, ultrasonic activation and XP‐endo Finisher, with a mixture of sodium hypochlorite and etidronic acid in infected isthmuses and root canals of extracted human mandibular molar teeth.MethodologyFifty‐six mesial roots of mandibular molars, half of them with a continuous isthmus from the cervical to the apical third between the two root canals (type 1), and the other half with a continuous isthmus from the cervical to the middle third and one canal in the apical third (type 2), were included. The root canals were contaminated for 7 days with an Enterococcus faecalis suspension. There were three experimental groups plus a control group (n = 7 per type of root canal anatomy). All the root canals, except for the control group that was not treated, were chemomechanically prepared and then assigned to one of the experimental groups according to the final adjunctive procedure: conventional irrigation, ultrasonic activation or XP‐endo Finisher activation. The irrigating solution used was a combination of 2.5% sodium hypochlorite and 9% etidronic acid, and the final protocols were applied for three cycles of 30 s with a 3 mL volume. The antibiofilm activity was evaluated at each location (root canal and isthmus) and third (cervical, middle and apical) using confocal laser scanning microscopy and the live/dead technique. Statistical analysis was performed using SPSS (descriptive statistics) and SUDAAN (P‐value calculations).ResultsRoot canals had significantly lower biovolume values than the isthmuses (P < 0.05). The biovolume in the root canals was significantly reduced in all the experimental groups in all the thirds except for conventional irrigation in the apical third (P > 0.05). In the cervical and middle thirds, ultrasonic activation was associated with the lowest biovolumes (P < 0.05), followed by XP‐endo Finisher. In the isthmus, disinfection was similar in all the thirds for all the protocols. Conventional irrigation was associated with intermediate values with no significant differences from the control group or from the activated protocols (P > 0.05), although the latter were significantly different from the control group (P < 0.05). No differences were found between ultrasonic activation and XP‐endo Finisher in the middle and apical thirds (P > 0.05) in the isthmuses.ConclusionsIn this laboratory study on extracted teeth, the isthmus was more difficult to disinfect than root canals. In the root canals, ultrasonic activation and XP‐endo Finisher had a greater effectiveness than conventional irrigation. In the isthmuses, no differences were observed between the two activation techniques and conventional irrigation.

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Aim: There is a need to explore new alternatives for root canal disinfection in regenerative endodontics, since the current strategies are far from ideal. Currently, the potential use of diclofenac (DC) is being investigated for controlling root canal infections. The objective was to evaluate the antimicrobial efficacy of novel DC-based hydrogels (DCHs) against polymicrobial biofilms grown in radicular dentine and root canals and to compare results with triantibiotic (TAH) and diantibiotic (DAH) hydrogels, and calcium hydroxide (Ca[OH] 2 ). Methodology: The in vitro antimicrobial activity of intracanal medicaments was evaluated against 3-week-old polymicrobial root canal biofilms grown on human radicular dentine. Dentine samples were obtained and randomly divided into the study groups (n = 4/group): (1) 1 mg/ml TAH; (2) 1 mg/ml DAH; (3) 5% diclofenac (DCH); (4) 2.5% DCH; (5) 1.25% DCH; (6) 1 mg/ml DAH + 5% DCH; (7) Ca(OH) 2 paste; (8) positive control. The microbial viability, in terms of percentage of intact cell membranes, was assessed after 7 days by confocal scanning laser microscopy (CSLM). The ex vivo efficacy of intracanal medications was evaluated in root canals infected with a polymicrobial suspension. Intracanal microbiological samples at baseline (S1) and 7 days post-treatment (S2) were taken; microbial quantification and cell viability were assessed by quantitative polymerase chain reaction (qPCR) and flow cytometry (FC). The mean Log 10 of bacterial DNA copies in root canal samples before (S1) and the Log 10 reduction of DNA copies S1-S2 in qPCR were recorded. The absolute value of total cells stained, and the percentage reduction of intact membrane cells after treatment (S1-S2), were analysed by FC. Global comparison was done using the Kruskal-Wallis test, whilst the Mann-Whitney U test was used for pair-by-pair comparison.Results: Confocal scanning laser microscopy analysis indicated that the greatest effectiveness was obtained with 5% DCH, showing significant differences with respect to the other groups (p < .001). In root canals, the highest Log 10 DNA reduction S1-S2 was obtained with 5% DCH and TAH, with no differences between them.