Characterization of Microbiota of Root Canal-Treated Teeth with Posttreatment Disease

Abstract

This study evaluated the microbiota of root canals undergoing retreatment. The most prevalent taxa detected by checkerboard included Propionibacterium species, Fusobacterium nucleatum, streptococci, and Pseudoramibacter alactolyticus. Quantitative real-time PCR detected Enterococcus faecalis and streptococci in 38% and 41% of the cases, comprising 9.76% and 65.78% of the total bacterial counts, respectively. The findings call into question the status of E. faecalis as the main pathogen and suggest that other species can be candidate pathogens associated with persistent/secondary endodontic infections. P osttreatment apical periodontitis is an inflammatory disease caused by a persistent/secondary infection of the dental root canal (8,11,13,15,26,30). Enterococcus faecalis has been the most frequently detected species in root canal-treated teeth as revealed by both culture and molecular studies (5,8,11,12,16,23,24,26,30,31), but recent studies have called into question its etiologic role in posttreatment disease (7,15,20,31). Moreover, taxa related to several other genera, including Streptococcus, Dialister, Fusobacterium, Filifactor, Parvimonas, Prevotella, Propionibacterium, and Pyramidobacter, have also been detected in treated teeth (5,8,11,12,15,20,22,26,27,29,30).The purpose of this molecular study was 2-fold. First, the samples from root canal-treated teeth with posttreatment apical periodontitis undergoing retreatment were surveyed for the presence of 28 bacterial taxa using the reverse capture-checkerboard DNA-DNA hybridization approach. Then, the total bacterial counts and the presence, levels, and proportions of E. faecalis and streptococci were determined by using a quantitative real-time PCR (qPCR) assay.Forty-two teeth were selected from patients (30 females and 12 males aged 16 to 70 years; mean age, 41 years) who had been referred for root canal retreatment to the Department of Endodontics, Estácio de Sá University. All the root canal-treated teeth were asymptomatic, showed radiographic evidence of apical periodontitis, and had had endodontic therapy completed more than 2 years earlier. All teeth were coronally restored and no direct exposure of the root canal filling material to the oral cavity was evident. The termini of the root canal fillings ranged from 0 to 4 mm short of the radiographic apex. No case was overfilled. The selected teeth showed an absence of periodontal pockets Ͼ4 mm in size. Approval for the study protocol was obtained from the Ethics Committee of the Está-cio de Sá University.The procedures for disinfecting the operative field and taking samples from treated root canals were as described previously (22, 26). Eight participants (out of 50 patients that started the experiment) were excluded because control samples from the operative field were positive for bacterial DNA as determined by broad-range PCR. DNA was extracted from clinical samples using the QIAamp DNA minikit (Qiagen, Valencia, CA), following the protocol recommended by the manufacturer. To maximize DNA extraction, ...

Efficacy and retrievability of root canal filling using calcium silicate‐based and epoxy resin‐based root canal sealers with matched obturation techniques

“…The residues of sealers after removal of filling materials may act as a continuous infection source to hide the microorganisms behind of them and in the dentinal tubules (21). As previously described, the retrievability was evaluated by in vitro test using extracted teeth and it was possible to compare three kinds of calcium silicate-based sealers with AH-plus sealer.…”

Comparison of the Bacterial Composition and Structure in Symptomatic and Asymptomatic Endodontic Infections Associated with Root-Filled Teeth Using Pyrosequencing

“…Secondary and persistent endodontic infections have mainly been studied using culture methods [2–4,8,10,13,14,16,17,19] and molecular methods such as species-specific PCR and real-time PCR, checkerboard hybridization and DGGE [5,7,15,20–24]. Only few open-ended 16S rDNA-based cloning studies have been published in recent years [6,25–27].…”

The Unique hmuY Gene Sequence as a Specific Marker of Porphyromonas gingivalis

“…Therefore, these data should be viewed with caution and the following reasons should be taken into consideration. Determination of absolute total bacterial cells by qPCR could be influenced by the variation in the number of 16S rRNA gene copies in a given species, since it has been shown that it may vary from 1 to 15 [67]–[72], and in this study we assumed the presence of 4 copies in all bacteria. Thus, total bacterial counts evaluated by the number of 16S rDNA may be fluctuated in polymicrobial samples.…”