Abstract Introduction Revascularization outcome depends on microbial elimination because apical repair will not happen in the presence of infected tissues. This study evaluated the microbial ...composition of traumatized immature teeth and assessed their reduction during different stages of the revascularization procedures performed with 2 intracanal medicaments. Methods Fifteen patients (7–17 years old) with immature teeth were submitted to the revascularization procedures; they were divided into 2 groups according to the intracanal medicament used: TAP group ( n = 7), medicated with a triple antibiotic paste, and CHP group ( n = 8), dressed with calcium hydroxide + 2% chlorhexidine gel. Samples were taken before any treatment (S1), after irrigation with 6% NaOCl (S2), after irrigation with 2% chlorhexidine (S3), after intracanal dressing (S4), and after 17% EDTA irrigation (S5). Cultivable bacteria recovered from the 5 stages were counted and identified by means of polymerase chain reaction assay (16S rRNA). Results Both groups had colony-forming unit counts significantly reduced after S2 ( P < .05); however, no significant difference was found between the irrigants (S2 and S3, P = .99). No difference in bacteria counts was found between the intracanal medicaments used ( P = .95). The most prevalent bacteria detected were Actinomyces naeslundii (66.67%), followed by Porphyromonas endodontalis, Parvimonas micra, and Fusobacterium nucleatum, which were detected in 33.34% of the root canals. An average of 2.13 species per canal was found, and no statistical correlation was observed between bacterial species and clinical/radiographic features. Conclusions The microbial profile of infected immature teeth is similar to that of primarily infected permanent teeth. The greatest bacterial reduction was promoted by the irrigation solutions. The revascularization protocols that used the tested intracanal medicaments were efficient in reducing viable bacteria in necrotic immature teeth.
Abstract Introduction The aim of this study was to investigate the prevalence of virulence factors and the antimicrobial resistance of Enterococcus faecalis isolates of teeth with failure of the ...endodontic treatment. Methods Twenty root canal samples were collected from teeth with apical periodontitis. E. faecalis was firstly identified based on phenotypic features and then by 16S ribosomal RNA gene sequencing. The antimicrobial susceptibility was determined by the minimum inhibitory concentration (MIC) of amoxicillin, amoxicillin + clavulanate, azithromycin, benzylpenicillin, ciprofloxacin, clindamycin, chloramphenicol, doxycycline, erythromycin, gentamicin, metronidazole, moxifloxacin, rifampicin, tetracycline, and vancomycin using the E test method. Virulence factors ( ace , asa , asa373 , cylA , efaA , esp , and gelE ) were detected by polymerase chain reaction assay. Results Amoxicillin + clavulanate was effective against all strains. Intermediate and total resistance was found against the majority of the tested antimicrobials. The susceptibility of some microorganisms to some antimicrobial agents changed according to the evaluation time. MIC50 and MIC90 also varied according to the evaluation time. In relation to the virulence factors of the E faecalis isolates, ace was detected in 100% of the strains, asa (60%), asa373 (15%), efaA (95%), esp (70%), and gelE (75%), whereas cylA was not detected. Conclusions It was concluded that E. faecalis isolates from persistent endodontic infections showed varied degrees of intermediate/total resistance to several antimicrobial agents, with amoxicillin + clavulanate being the most effective agent. Moreover, the strains showed different patterns for virulence gene detection.
Abstract Introduction The aim of the study was to evaluate the cytotoxicity, radiopacity, pH, and flow of a calcium silicate–based and an epoxy resin–based endodontic sealer, MTA Fillapex (Angelus, ...Londrina, PR, Brazil) and AH Plus (Dentsply, Konstanz, Germany), respectively. Methods Cytotoxicity, radiopacity, and flow evaluation were performed following ISO requirements. The pH level was measured at periods of 3, 24, 72, and 168 hours. Cytotoxicity was evaluated by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide assay to check the Balb/c 3T3 cells viability at 1- to 4-week periods. Data were statistically analyzed by analysis of variance and the Tukey test with a significance level of 5%. Results In all tested periods, MTA Fillapex was more cytotoxic than AH Plus ( P < .05). Although AH Plus presented higher radiopacity than MTA Fillapex ( P < .05), both sealers showed minimum required values. MTA Fillapex presented alkaline pH in all experimental times, whereas AH Plus cement showed a slightly neutral pH and a flow significantly lower than that of MTA Fillapex ( P < .05). Conclusions Although MTA Fillapex was more cytotoxic than AH Plus, it showed suitable physicochemical properties for an endodontic sealer.
Abstract Introduction The infectious content of root canals, including bacteria and lipoteichoic acid (LTA), cause injuries to the periapical tissues. The purpose of this clinical study was to ...quantify the levels of both LTA and cultivable bacteria at the different phases of endodontic retreatment (ER) of teeth with post-treatment apical periodontitis. It also aimed to investigate the presence of gram-positive microorganisms before and after chemomechanical preparation (CMP) and intracanal medication (ICM). Methods Twenty infected root canals of single-rooted teeth were randomly assigned into 2 groups according to the chemical substance used for CMP ( n = 10 per group): chlorhexidine (CHX) group, 2% CHX gel, and the sodium hypochlorite (NaOCl) group, 6% NaOCl. Root canal samples were taken using paper points before (S1) and after CMP (S2) and after 30 days of ICM with calcium hydroxide + 2% CHX gel (S3). Microorganisms were identified by the culture technique using biochemical tests. Cultivable bacteria were determined by counting the colony-forming unit. LTA levels were measured using the enzyme-linked immunosorbent assay (pg/mL). Results A total of 70 gram-positive species, out of 102 species isolated, were found in the root canals (54 in S1, 4 in S2, and 12 in S3). Enterococcus faecalis was the most frequent isolated taxon in all phases of the ER. LTA (574.0 ± 94.7) and cultivable bacteria (101.2 ± 79.2) were present in all S1 samples. CMP decreased the overall levels of cultivable bacteria by 99.4% and LTA by 24.8% ( P < .05), whereas the total overall reduction level of ICM on viable bacteria was 99.5% and on LTA it was 38.6% ( P < .05). CMP with 2% CHX gel (CHX group, 99.3%) was more effective ( P < .05) than 6% NaOCl (NaOCl group, 92.1%) on bacterial reduction. Likewise, ICM showed a 100% reduction in the CHX group and 98.5% in the NaOCl group. Regarding the reduction of LTA, CMP with 2% CHX gel (CHX group, 26.9%) was more effective ( P < .05) than 6% NaOCl (NaOCl group, 22.6%). In addition, ICM showed a 43.2% reduction in the CHX group and 36.2% in the NaOCl group ( P > .05). Conclusions The reduction rates of bacteria were higher than the LTA. Moreover, gram-positive microorganisms were present in all phases of the endodontic retreatment.
Abstract Introduction This study was conducted to evaluate the microbiomes of endodontic-periodontal lesions before and after chemomechanical preparation (CMP). Methods Clinical samples were taken ...from 15 root canals (RCs) with necrotic pulp tissues and from their associated periodontal pockets (PPs) ( n = 15) of teeth with endodontic-periodontal lesions before and after CMP. The Human Oral Microbe Identification using Next Generation Sequencing (NGS) protocol and viable culture were used to analyze samples from RCs and PPs. The Mann-Whitney U test and Benjamini-Hochberg corrections were performed to correlate the clinical and radiographic findings with microbial findings ( P < .05). Results Bacteria were detected in 100% of the samples in both sites (15/15) using NGS. Firmicutes was the most predominant phylum in both sites using both methods. The most frequently detected species in the RCs before and after CMP using NGS were Enterococcus faecalis , Parvimonas micra , Mogibacterium timidum , Filifactor alocis , and Fretibacterium fastidiosum . The species most frequently detected in the PPs before and after CMP using NGS were P. micra , E. faecalis , Streptococcus constellatus , Eubacterium brachy , Tannerella forsythia , and F. alocis. Associations were found between periapical lesions ≤2 mm and Desulfobulbus sp oral taxon 041 and with periodontal pockets ≥6 mm and Dialister invisius and Peptostreptococcus stomatis (all P < .05, found in the RCs before CMP). Conclusions It is concluded that the microbial community present in combined endodontic-periodontal lesions is complex and more diverse than previously reported. It is important to note that bacteria do survive in some root canals after CMP. Finally, the similarity between the microbiota of both sites, before and after CMP, suggests there may be a pathway of infection between the pulp and periodontium.
Abstract Introduction This study investigated the bacterial community involved in primary endodontic diseases, evaluated its ability to activate the macrophage Toll-like receptor 4 receptor through ...p38 mitogen-activated protein kinase (MAPK) and nuclear factor kappa B (NF-κB) signaling pathways, and determined the levels of endotoxins and interleukins (interleukin IL-6 and -10) produced by endodontic content-stimulated macrophages. Methods Samples were taken from 21 root canals by using sterile/apyrogenic paper points. Raw 264.7 macrophages were stimulated with root canal contents. Checkerboard DNA-DNA hybridization was used for bacterial analysis and the limulus amebocyte lysate assay for endotoxin measurement; p38 MAPK and NF-κB activation was determined by Western blot analysis. IL-6 and IL-10 were measured using the enzyme-linked immunosorbent assay. Results Bacteria and endotoxins were detected in 100% of the samples (21/21). The most frequently observed species were Parvimonas micra (16/21, 76%), Fusobacterium nucleatum ssp. nucleatum (15/21, 71%), and Porphyromonas endodontalis (14/21, 66%). Correlations were found between endotoxins and IL-6 and IL-10 ( P < .05); p38 phosphorylation had a peak at 60 minutes, and NF-κB was quickly activated after 10 minutes of stimulation. Conclusions It was concluded that the complex bacterial community was shown to be a potent activator of TLR-4 determined by the p38 MAPK and NF-κB signaling pathways, culminating in a high antigenicity against macrophages through the levels of IL-6 and IL-10, all significantly affected by endotoxin levels.
Abstract Background This study was performed to determine which of the quantitative methods, namely, chromogenic endpoint, chromogenic kinetic, and turbidimetric kinetic ones, best fit for the ...analysis of primary endodontic infections. Methods Twenty-one root canals with apical periodontitis were sampled with paper points. The same sample was analyzed by means of the endpoint chromogenic Limulus amebocyte lysate (LAL) assay (QCL), quantitative kinetic chromogenic LAL assay (KQCL), and kinetic turbidimetric LAL assay (Turbidimetric). Results All three LAL methods were effective in the recovery of endotoxin from root canal infection. Regardless of the method tested, endotoxin was detected in 100% of the root canals (21/21). The KQCL assay yielded a median value of endotoxin of 7.49 EU/mL, close to and not significantly different from those for the turbidimetric test (9.19 EU/mL) (both kinetic methods) ( p > 0.05). In contrast, the endpoint QCL showed a median value of 34.20 EU/mL ( p < 0.05). The comparison of the three methods revealed that both turbidimetric and KQCL methods were more precise, with best reproducibility (the coefficient variation between analysis of the root canal and its duplicate was lower than 10%). The inhibition/enhancement assay indicated a good interaction between the root canal samples with the turbidimetric method. Conclusion This study has revealed that quantitative kinetic-turbidimetric and kinetic-chromogenic LAL methods are best fitted for the analysis of endotoxins in root canal infection, both being more precise and allowing better reproducibility compared with the endpoint-QCL assay.
Abstract Introduction The smear layer adheres to dentinal surface, thus occluding the dentinal tubules. Because this layer disfavors the penetration of irrigant solutions and root canal fillings, it ...should be removed. The aim of this study was to compare the effectiveness of 37% phosphoric acid with that of 17% EDTA and 10% citric acid in the removal of smear layer. Materials and Methods Fifty-two maxillary single-rooted human canines were accessed and instrumented. Between each instrument used, the canals were irrigated with sodium hypochlorite. After instrumentation, the teeth were irrigated with distilled water and then divided into groups according to the time and substances employed. The substances used were 17% EDTA, 10% citric acid, and 37% phosphoric acid solution and gel. The experimental time periods were of 30 seconds, 1 minute, and 3 minutes. The samples were prepared and observed by means of scanning electron microscopy. Three photomicrographs (2,000×) were recorded for each sample regarding the apical, middle, and cervical thirds. A score system was used to evaluate the images. Results None of the substances analyzed in this study was effective for removing the smear layer at 30 seconds. In the 1-minute period, the phosphoric acid solution showed better results than the other substances evaluated. In the 3-minute period, all the substances worked well in the middle and cervical thirds although phosphoric acid solution showed excellent results even in the apical third. Conclusions These findings point toward the possibility that phosphoric acid solution could be a promising agent for smear layer removal.
Abstract Introduction Macrophages are highly activated by endodontic contents. This study investigated the correlation between different clinical signs/symptoms and radiographic features according to ...the levels of interleukin (IL)-1β, tumor necrosis factor α (TNF-α), IL-6, IL-10, prostaglandin E2 (PGE2 ), and their networks produced by endodontic content–stimulated macrophages collected from primary endodontic infection with apical periodontitis (PEIAP). Methods Samples were taken from 21 root canals with PEIAP by using paper points. The presence of exudate (EX), pain on palpation (POP), tenderness to percussion (TTP), and the size of the radiographic lesion (SRL) were recorded. Polymerase chain reaction (16S rDNA) was used for bacterial detection and limulus amebocyte lysate (LAL) assay for endotoxin measurement. Raw 264.7 macrophages were stimulated with bacterial contents during 24 hs. The amounts of IL-1β, TNF-α, IL-6, IL-10 and PGE2 were measured by enzyme-linked immunosorbent assay. Log-based data were correlated by multiple logistic regression ( P < .05). Results Bacteria and endotoxin were detected in 100% of the samples. IL-6 and TNF-α were positively correlated with SRL and EX, respectively ( P < .05). Clinical signs/symptoms and radiographic findings were set as dependent variables for EX-positive correlations between PGE2 , IL-1β, and TNF-α ( P < .05), whereas IL-6 and PGE2 were positively correlated to each other in POP but negatively correlated in SRL ( P < .05). When POP and TTP-POP were set as dependent variables, different cytokine networks were found. Conclusions Our findings suggest different roles for each cytokine in the development of apical periodontitis, whose effects of overlapping networks depend on the signs/symptoms and radiographic features found in endodontic infection.
Abstract Introduction The aim of this work was to characterize the by-products formed in the associations between the most commonly used irrigants in endodontic practice through electrospray ...ionization quadrupole time-of-flight mass spectrometry analyses. Methods Sodium hypochlorite (NaOCl) (0.16%, 1%, 2.5%, and 5.25%) was associated with 2% chlorhexidine (CHX) solution and gel, 17% EDTA, 10% citric acid, 37% phosphoric acid, saline solution, ethanol, and distilled water. CHX solution and gel were also associated with all above mentioned irrigants. The solutions were mixed in a 1:1 ratio, and electrospray ionization quadrupole time-of-flight mass spectrometry was used to characterize the precipitates when formed. Results CHX produced an orange-brown precipitate when associated with NaOCl from 1%–5.25% and an orange-white precipitate when associated with 0.16% NaOCl. When associated with EDTA, CHX produced a white milky precipitate, and when associated with saline solution and ethanol, a salt precipitation was produced. No precipitation was observed when CHX was associated with citric acid, phosphoric acid, or distilled water. In the NaOCl associations, precipitation occurred only when CHX was present. Conclusion The orange-brown precipitate observed in the association between CHX and NaOCl occurs because of the presence of NaOCl, an oxidizing agent causing chlorination of the guanidino nitrogens of the CHX. The precipitates formed in the reaction of CHX with EDTA, saline solution, and ethanol were associated with acid-base reactions, salting-out process, and lower solubility, respectively. NaOCl associated with EDTA, citric acid, and phosphoric acid leads mainly to chlorine gas formation. Intermediate flushes with distilled water seem to be appropriate to prevent or at least reduce formation of by-products.
PDF