Teeth with vital pulps and stage III periodontitis unresponsive to therapy exhibit a pulpal inflammatory profile similar to symptomatic irreversible pulpitis.

AIM: The aim of this study is to investigate the expression of inflammatory biomarkers (TNF-α, IL-10, IL-1ß) and the pulpitis-associated miRNA (miR-30a-5p and miR-128-3p) in pulp tissue samples from unrestored teeth with a vital normal pulp (NP), teeth with symptomatic irreversible pulpitis (IP) and in unrestored teeth with periodontal disease, unresponsive to periodontal therapy, and a vital pulp (EP). METHODOLOGY: Thirty patients were included in this observational study (10 teeth with NP, 10 teeth with IP, 10 teeth with EP). Dental pulp tissues samples were collected from patients during root canal treatment (RCT). RNA was extracted and qRT-PCR of target genes (tumour necrosis factor [TNF]-α, interleukin [IL]-1ß, IL-10) and miRNAs (has-miR-30a-5p, has-miR-128-3p) performed to assess the expression profile. Fold-change in expression was calculated using the formula 2-(ΔCt(Exp)-ΔCt(Ctrl)). One-way anova with post-hoc Tukey's was used to determine significant differences between groups. The significance level was set at 5% (p < .05). All teeth were also followed up clinically for 1 year and evaluated for a range of endodontic and periodontal-related outcomes. RESULTS: All investigated genes significantly increased in expression and miRNAs significantly decreased in expression in the IP and EP groups compared with the NP group (p < .05). With regards to TNF-α and IL-1ß there were no significant differences in expression between the IP and EP groups (p > .05), whereas IL-10 expression levels were significantly reduced in the EP compared with the IP group (p < .05). Both miR-30a-5p and miR-128-3p showed significantly reduced expression in both IP and EP lesions, compared with NP (p < .05); however, no significant differences in miRNA expression were observed between IP and EP groups (p > .05). One year after root canal treatment and periodontal maintenance, tooth mobility and probing depth were significantly reduced in the EP group (p < .05). CONCLUSION: Pulp tissues from teeth with IP and EP presented similar levels of altered inflammatory markers compared with NP. TNF-α, IL-10, IL-1ß cytokines and miRNAs (miR-30a-5p and miR-128-3p) are potential objective biomarkers to indicate pulpal inflammatory status, aiding diagnosis and directing clinical decision-making. RCT may be beneficial to improve stage III periodontitis unresponsive to non-surgical periodontal treatment, but further research is required.

Effect of supplementary sodium hypochlorite agitation techniques on an ex vivo oral multispecies biofilm during passive disinfection of simulated immature roots.

AIM: To compare the effect of different sodium hypochlorite (NaOCl) agitation techniques on an ex vivo oral multispecies biofilm during passive disinfection of simulated immature roots. METHODOLOGY: Extracted human teeth were prepared to simulate immature roots. They were infected with a dental plaque-derived multispecies biofilm and cultured for 14 days. The roots were randomly designated into four groups: (1) negative control (PBS), (2) 1.5% NaOCl (CNI), (3) CNI + Ultrasonic activation (UA), (4) CNI + EasyClean agitation (ECA), (5) CNI + XP-endo finisher agitation (XPF), and (6) positive control (6% NaOCl). Biofilm samples were collected from the root canals and used to determine the number of viable cells (colony-forming units), scanning electron microscopy, and 16S rRNA gene sequencing. The mean colony-forming units per mL (CFU/mL) were analysed using One-way anova. 16S rRNA sequencing data were analysed for alpha (observed OTUs, Shannon index, and Chao1) and beta diversity (Bray-Curtis dissimilarities). The LEfSe analysis was used to determine the effect of treatment procedures on the abundance of root canal microbiota. The significance was set at .05. RESULTS: PBS and CNI samples had significantly higher CFU/mL counts than UA, ECA, XPF, and 6% NaOCl samples (p < .05). The pre-treatment, PBS, and CNI groups had significantly greater alpha diversity than the UA, ECA, XPF, and 6% NaOCl groups (p < .05). NaOCl agitation groups and the 6% NaOCl group achieved a more pronounced reduction in bacteria from the genera Fusobacterium, Actinomyces, Porphyromonas, and Capnocytophaga. CONCLUSIONS: The effectiveness of passive disinfection protocols was enhanced by NaOCl agitation techniques, suggesting that this supplementary method can improve the outcome of revitalization procedures.

Photodynamic therapy in the root canal treatment of primary teeth: A systematic review of clinical trials.

BACKGROUND: Dental caries is the most common oral disease worldwide, and it is estimated to affect 2.3 billion people, with at least 530 million of them being schoolchildren with decayed primary teeth. This condition can rapidly evolve into irreversible pulp inflammation and pulp necrosis and thus requiring endodontic intervention. Photodynamic therapy (PDT) is a supplementary method to conventional pulpectomy and is used to improve the protocol used for disinfection. AIM: The main objective of this study was to evaluate through a systematic review the efficacy of supplementary PDT on the pulpectomy of primary teeth. This review was registered a priori on the PROSPERO database (CRD42022310581). DESIGN: Two independent and blinded reviewers carried out a comprehensive search in five databases: PubMed, Cochrane, Scopus, Embase, and Web of Science. Eligible studies were randomized and nonrandomized clinical trials that evaluated in vivo microbiological load or clinical outcomes after using supplementary PDT in infected primary teeth. RESULTS: After the selection process, four studies met the inclusion criteria and were included in this study. Data regarding the sample characteristics and PDT protocols were retrieved. All included trials used phenothiazinium salts as photosensitizer agents. Only one study observed a significant difference in the in vivo microbiological load reduction outcome when performing PDT on primary teeth. The remaining studies all discussed the possible benefits of this intervention; however, none observed a significant difference in this outcome. CONCLUSION: In this systematic review, moderate-to-low certainty of the available evidence was observed, and thus, no significant conclusions can be drawn from the findings.

Effectiveness of adjunct therapy for the treatment of apical periodontitis: A systematic review and meta-analysis.

BACKGROUND: Adjunct therapy refers to any intracanal procedure going beyond chemomechanical preparation with instruments and traditionally delivered irrigants (excluding interim dressings). It is not clear whether and which of these adjunct therapies have a significant impact on the outcome of root canal treatment [healing of apical periodontitis (AP) and other patient-related outcomes]. OBJECTIVES: This systematic review aimed to analyse available evidence on the effectiveness of adjunct therapy for the treatment of AP in permanent teeth, according to a population, intervention, comparison, outcome, time and study design framework formulated a priori by the European Society of Endodontology. METHODS: Five electronic databases (PubMed, Embase, Scopus, Cochrane and Web of Science) were searched up to October 2021 to identify clinical studies comparing adjunct therapy to no adjunct therapy in adult patients with AP. Animal studies, reviews, studies with less than 10 patients per arm and studies with a follow-up time of less than 1 year, or less than 7 days for postoperative pain, were excluded. The quality of the included studies was appraised by the appropriate tools [Risk of Bias 2 (RoB2) for randomized clinical trials (RCTs) and Newcastle-Ottawa Scale for observational studies]. Meta-analysis was performed using a random-effects model. The certainty of the evidence was assessed using the Grading of Recommendations, Assessment, Development and Evaluation (GRADE) approach. RESULTS: Fourteen studies (13 RCTs and one retrospective cohort) fulfilled the inclusion criteria for this review. They evaluated different types of adjunct therapy: antimicrobial photodynamic therapy (aPDT; three studies), diode laser canal irradiation (3), Nd:YAG laser canal irradiation (2), Er;Cr:YSGG laser canal irradiation (1), ozone therapy (2) and ultrasonically activated irrigation (UAI) (4). Radiographical healing was reported in seven studies, but meta-analysis was only possible for UAI (two studies), showing no statistically significant difference in healing after 12 months. Pain after 7 days was reported in seven studies. Meta-analysis on three studies that used aPDT and on two studies using diode laser irradiation showed no significant difference in the prevalence of pain after 7 days between the control and adjunct therapy. According to RoB2 tool, six studies had a high risk of bias, five studies had some concerns, and two studies low risk of bias. The GRADE assessment revealed a very low strength of evidence for diode laser, and low strength of evidence for PDT, ozone and UAI studies. DISCUSSION: The included studies displayed significant heterogeneity in terms of type of adjunct therapy, technical details per adjunct therapy, outcome reporting and several combinations of these, limiting the potential for meta-analysis. CONCLUSIONS: There is insufficient evidence to recommend any adjunctive therapy for the treatment of apical periodontitis. REGISTRATION: Prospero CRD42021261869.

Microbiota present in combined endodontic-periodontal diseases and its risks for endocarditis.

INTRODUCTION: Infective endocarditis (IE) is an inflammatory disease usually caused by bacteria that enter the bloodstream and establish infections in the inner linings or valves of the heart, including blood vessels. Despite the availability of modern antimicrobial and surgical treatments, IE continues to cause substantial morbidity and mortality. Oral microbiota is considered one of the most significant risk factors for IE. The objective of this study was to evaluate the microbiota present in root canal (RC) and periodontal pocket (PP) clinical samples in cases with combined endo-periodontal lesions (EPL) to detect species related to IE using NGS. METHODS: Microbial samples were collected from 15 RCs and their associated PPs, also from 05 RCs with vital pulp tissues (negative control, NC). Genomic studies associated with bioinformatics, combined with structuring of a database (genetic sequences of bacteria reported for infective endocarditis), allowed for the assessment of the microbial community at both sites. Functional prediction was conducted using PICRUSt2. RESULTS: Parvimonas, Streptococcus, and Enterococcus were the major genera detected in the RCs and PPs. A total of 79, 96, and 11 species were identified in the RCs, PPs, and NCs, respectively. From them, a total of 34 species from RCs, 53 from PPs, and 2 from NCs were related to IE. Functional inference demonstrated that CR and PP microbiological profiles may not be the only risk factors for IE but may also be associated with systemic diseases, including myocarditis, human cytomegalovirus infection, bacterial invasion of epithelial cells, Huntington's disease, amyotrophic lateral sclerosis, and hypertrophic cardiomyopathy. Additionally, it was possible to predict antimicrobial resistance variants for broad-spectrum drugs, including ampicillin, tetracycline, and macrolides. CONCLUSION: Microorganisms present in the combined EPL may not be the only risk factor for IE but also for systemic diseases. Antimicrobial resistance variants for broad-spectrum drugs were inferred based on PICRUSt-2. State-of-the-art sequencing combined with bioinformatics has proven to be a powerful tool for conducting studies on microbial communities and could considerably assist in the diagnosis of serious infections. CLINICAL RELEVANCE: Few studies have investigated the microbiota in teeth compromised by combined endo-periodontal lesions (EPL), but none have correlated the microbiological findings to any systemic condition, particularly IE, using NGS techniques. In such cases, the presence of apical periodontitis and periodontal disease can increase IE risk in susceptible patients.

Biocompatibility analysis in subcutaneous tissue and physico-chemical analysis of pre-mixed calcium silicate-based sealers.

OBJECTIVES: To evaluate the biocompatibility, physical and chemical properties of three pre-mixed calcium silicate-based sealers and an epoxy resin-based material were assessed. Pre-mixed sealers supposedly obtain water from the root canal moist to hydrate and set. MATERIALS AND METHODS: Polyethylene tubes were filled with the materials Bio-C Sealer Ion+, Bio-C Sealer, EndoSequence BC Sealer and AH Plus Jet, or left empty and surgically implanted in the subcutaneous tissue of Wistar rats. The animals were euthanised and the tubes and tissue were removed for histological analysis and scanning electron microscopy (SEM) coupled with energy-dispersive spectrometry (EDS). Materials' surface chemical characterisation was assessed using Raman spectroscopy and SEM/EDS. Flow, setting time (in two conditions), solubility, radiopacity and pH were also analysed. ANOVA and Bonferroni correction were performed for comparisons (P < 0.05). RESULTS: Inflammatory response observed in the tissues subsided from 7 to 30 days. Tungsten migration could be detected in the surrounding tissue following AH Plus Jet implantation. All calcium silicate-based sealers exhibited zirconium oxide (radiopacifier) and tricalcium silicate peaks before and after implantation. All materials exhibited flow values above 17 mm. An approximately tenfold difference was observed between the plaster- and metal-mould setting times of the calcium silicate cements indicating its sensitivity to moist variations and solubility above 8% was also observed for these materials. CONCLUSIONS: Pre-mixed materials exhibited variable setting time and solubility with a decreasing inflammatory response. CLINICAL RELEVANCE: The variable moist-dependant setting time with high solubility poses a concern for the clinical use of these pre-mixed sealers.

Microbiological profile of root canals indicated for endodontic retreatment due to secondary endodontic infections or for prosthetic reasons.

OBJECTIVE: The objective of this study was to evaluate the microbiological profile of root canals indicated for endodontic retreatment due to secondary endodontic infections evidenced by the presence of chronic apical periodontitis (G1) or for prosthetic reasons, without clinical and radiographic signs of endodontic reinfection (G2). METHODS: Microbiological samples were collected from thirty teeth (N=30) out of which 15 were indicated for retreatment due to the presence of chronic apical periodontitis (G1) and 15 were indicated exclusively for prosthetic reasons (G2). Samples were collected from root canals before (S1), after chemomechanical preparation (S2), and after 30 days of intracanal medication composed of calcium hydroxide and 2% chlorhexidine gel (S3). The molecular analysis was performed using Nested-PCR for the detection of 17 bacterial species. The efficacy of each stage of the retreatment in reducing the microbial load was verified by counting colony-forming units (CFU). The statistical analysis considered a significance level of 5%. RESULTS: The results showed that bacteria were detected in 100% of the cases in S1, in both groups, by Nested PCR. The most frequently found species in S1 in both groups were Enterococcus faecalis, Fusobacterium nucleatum, Porphyromonas gingivalis, Aggregatibacter actinomycetemcomitans, and Parvimonas micra. The microbial load of G1 was higher than G2 in the initial samples (S1). Endodontic retreatment was effective in reducing the microbial load in G1 and G2. Statistically significant associations were found between specific bacteria and clinical signs and symptoms. CONCLUSION: It was concluded that the microbial community present in the root canal of teeth indicated for endodontic retreatment is mixed and heterogeneous. G1 and G2 differ in the number of species per case and microbial load. CLINICAL RELEVANCE: Endodontic retreatment was effective in reducing the microbial load. Statistically significant associations were found between specific bacteria and clinical signs and symptoms.

Scaffolds for Dentin-Pulp Complex Regeneration.

Background and Objectives: Regenerative dentistry aims to regenerate the pulp-dentin complex and restore those of its functions that have become compromised by pulp injury and/or inflammation. Scaffold-based techniques are a regeneration strategy that replicate a biological environment by utilizing a suitable scaffold, which is considered crucial for the successful regeneration of dental pulp. The aim of the present review is to address the main characteristics of the different scaffolds, as well as their application in dentin-pulp complex regeneration. Materials and Methods: A narrative review was conducted by two independent reviewers to answer the research question: What type of scaffolds can be used in dentin-pulp complex regeneration? An electronic search of PubMed, EMBASE and Cochrane library databases was undertaken. Keywords including "pulp-dentin regeneration scaffold" and "pulp-dentin complex regeneration" were used. To locate additional reports, reference mining of the identified papers was undertaken. Results: A wide variety of biomaterials is already available for tissue engineering and can be broadly categorized into two groups: (i) natural, and (ii) synthetic, scaffolds. Natural scaffolds often contain bioactive molecules, growth factors, and signaling cues that can positively influence cell behavior. These signaling molecules can promote specific cellular responses, such as cell proliferation and differentiation, crucial for effective tissue regeneration. Synthetic scaffolds offer flexibility in design and can be tailored to meet specific requirements, such as size, shape, and mechanical properties. Moreover, they can be functionalized with bioactive molecules, growth factors, or signaling cues to enhance their biological properties and the manufacturing process can be standardized, ensuring consistent quality for widespread clinical use. Conclusions: There is still a lack of evidence to determine the optimal scaffold composition that meets the specific requirements and complexities needed for effectively promoting dental pulp tissue engineering and achieving successful clinical outcomes.

Chemomechanical preparation influences the microbial community and the levels of LPS, LTA and cytokines in combined endodontic-periodontal lesions: A clinical study.

BACKGROUND: This study was conducted to compare the microbiomes, the levels of lipopolysaccharides (LPS), lipoteichoic acid (LTA), and cytokines (interleukin [IL]-1ß and tumor necrosis factor-alpha [TNF-α]), before and after chemomechanical preparation (CMP) of the root canals (RC) and their associated periodontal pockets (PP) in teeth with combined EPL. MATERIALS: Samples were taken from 10 RC and PP, before and after CMP. The microbiomes (next-generation sequencing, V3-V4 hypervariable region of the 16S rRNA gene), microbiome diversity (bioinformatics analyses), LPS (limulus amebocyte lysate), LTA, IL-1ß, and TNF-α (ELISA) were evaluated. A statistical analysis was performed with significance level set at 5%. RESULTS: The most abundant phyla in both sites were Firmicutes and Proteobacteria. Comparative studies of bacterial genera species revealed that some increased and others decreased after CMP at both sites. A 3% reduction in Gram-negative bacteria (RC) and a 4% increase in Gram-positive bacteria (PP) were detected. LPS levels were 4.4 times higher in PP than in the RC. LTA was detected in all samples investigated. Higher levels of IL-1ß and TNF-α were detected in both sites at baseline. After CMP, LPS, LTA, IL-1ß and TNF-α were reduced in both sites. CONCLUSION: The microbial community in the RC and PP in teeth with combined EPL indicated a similarity between both sites. CMP effectively reduced the microbial load and the LPS levels from teeth with EPL, and consequently diminished the cytokine levels. The reduction in LTA levels in the RC and PP proved challenging.

Comparative analysis of bacterial content, levels of lipopolysaccharides and lipoteichoic acid in symptomatic and asymptomatic endodontic infections at different stages of endodontic treatment.

OBJECTIVES: This study aimed to detect the presence of specific bacteria and to evaluate the levels of lipopolysaccharides (LPS) and lipoteichoic acid (LTA) in symptomatic necrotic root canals associated with acute apical abscess (symptomatic group - GI). It also aimed to compare the findings with those presented by asymptomatic necrotic root canals (asymptomatic group - GII) in the different stages of the endodontic treatment. MATERIALS AND METHODS: Microbiological samples were collected from 20 root canals, including purulent collection from acute apical abscesses, before and after chemo-mechanical preparation (CMP) preparation (CMP) with chlorhexidine gel 2% and after 30 days of intracanal medication (ICM) with (Ca[OH]2) + chlorhexidine. The culture method was used to evaluate the efficacy of CMP and ICM. Nested PCR was used to detect the presence of 17 specific bacteria. Levels of LPS were measured by using limulus amebocyte lysate, whereas enzyme-linked immunosorbent assay was used for the quantification of LTA. RESULTS: CMP was effective in reducing the microbial load in both groups (P < 0.05). LPS levels were higher in GI than in GII (P < 0.05). There was a significant reduction in the LPS levels after CMP and ICM (P < 0.05) in GI and GII. LTA levels were significantly reduced in GI after ICM and in GII after CMP and ICM (both P < 0.05). Fusobacterium nucleatum and Enterococcus faecalis were frequently identified in both groups, alone or in combination with each other. CONCLUSION: Different species were detected in all stages of the endodontic treatment. CMP was able to reduce bacterial content and the levels of LPS, but not of LTA in the symptomatic group. High levels of LPS were correlated with spontaneous pain and pain to percussion in the symptomatic group. CLINICAL RELEVANCE: This clinical study showed that chemo-mechanical preparation was able to reduce bacterial load and levels of LPS, but not of LTA in the symptomatic group. Elevated levels of LPS were correlated with the presence of symptomatology.

Identification of Fusobacterium nucleatum in primary and secondary endodontic infections and its association with clinical features by using two different methods.

OBJECTIVE: Fusobacterium nucleatum is an important oral pathogen involved in endodontic infections. This study aimed to assess the frequency of Fusobacterium nucleatum in primary and secondary endodontic infections and its associations with the clinical features in a Brazilian population by using both culture and nested PCR methods. METHODS: A total of 100 microbial samples from patients with primary (n=50) and secondary endodontic infections (n=50) were analyzed by using culture and nested PCR methods. Strict anaerobic techniques were used for culture and identification of F. nucleatum. The DNA extracted from the samples was analyzed for the presence of target species by using species-specific primers. RESULTS: Culture and nested PCR methods detected F. nucleatum, respectively, in 11/100 and 82/100 root canals. F. nucleatum was isolated by culture from 10/50 (20%) root canals with primary infections and from 1/50 (2%) root canal with secondary/persistent infections. Nested PCR detected F. nucleatum in 42/50 (84%) root canals with primary infections and in 40/50 (80%) root canals with secondary/persistent endodontic infections. F. nucleatum was associated with spontaneous pain, tenderness to percussion, pain on palpation, swelling, tooth mobility, wet root canals, hemorrhagic exudate, tooth decay, inadequate restoration, and poor endodontic filling. CONCLUSION: F. nucleatum was found in more cases of primary endodontic infections than in cases of secondary/persistent ones. A higher prevalence of F. nucleatum was detected by using the nested PCR method than by using culture. The presence of F. nucleatum in the root canals was associated with several clinical features. CLINICAL RELEVANCE: The high prevalence of F. nucleatum in the root canals detected by molecular methods, and its association with several clinical features reveals the importance of these species in the development of apical pathologies and reinforces the need of an endodontic treatment directed to bacterial elimination.

Microbiological analysis of endodontically treated teeth with apical periodontitis before and after endodontic retreatment.

OBJECTIVE: To characterize the microbiota of teeth with endodontic treatment failure by 16S ribosomal RNA genetic sequencing (GS) and PCR at the different phases of the endodontic retreatment and to associate the presence of specific bacteria with clinical and radiographic features in teeth with apical periodontitis. MATERIALS AND METHODS: Twenty infected root canals of single-rooted teeth were selected. Samples were collected with sterile paper points before chemo-mechanical preparation (CMP) (S1), after CMP (S2) and after 30 days of intracanal medication (ICM) (S3). Microbial identification was performed using GS and PCR. Tukey-Kramer post hoc test and post hoc ANOVA were used for intergroup analysis. Paired t test and repeated-measures ANOVA were applied for intragroup analysis, at a significance level of 5%. RESULTS: A total of 89 strains were identified using GS. Sixty-five strains were recovered in S1 and 15 strains in S2, and 9 strains remained in S3. Enterococcus faecalis was the most predominant bacteria. Gram-positive cocci bacteria predominated. Gram-negative species were also detected. Using species-specific PCR primers to detect seven species, the most prevalent ones at all the phases of the endodontic retreatment were E. faecalis and Porphyromonas gingivalis. However, Parvimonas micra and P. gingivalis were associated with previous pain, P. gingivalis was associated with tenderness to percussion and E. faecalis, Fusobacterium nucleatum and P. gingivalis were associated with periapical lesion > 3 mm. CONCLUSIONS: In conclusion, the microbiota of persistent infection is polymicrobial with predominance of E. faecalis and P. gingivalis in all phases of the endodontic retreatment, regardless of the method used for microbial identification. Associations were found between specific bacteria and clinical/radiographic features. CLINICAL RELEVANCE: The characterization of the bacteria present at all phases of the endodontic retreatment is important for the monitoring of the effectiveness of the techniques used and to better understand the susceptibility of these species to the disinfection agent used during the procedures.

Detection of Streptococcus mutans in symptomatic and asymptomatic infected root canals.

OBJECTIVES: To investigate the presence of Streptococcus mutans in root canals of symptomatic necrotic teeth (SNT) and their associated acute apical abscesses (AAA) and in the root canals of asymptomatic necrotic teeth (ANT). It also aimed to investigate the presence of the cnm and cbm genes in specimens that harbored S. mutans. MATERIALS AND METHODS: DNA was extracted from samples collected from 10 patients presenting pulpal necrosis associated with radiographic evidence of apical periodontitis (ANT) and from 10 patients in need of endodontic therapy due to the presence of pulpal necrosis (SNT) and AAA. The control group consisted of 10 patients with teeth with normal vital pulp and requiring endodontic treatment for prosthetic reasons. The presence of S. mutans was detected by quantitative real-time-PCR (qPCR) using species-specific primers. Samples harboring S. mutans were further evaluated for the presence of CBP genes by qPCR as well. RESULTS: All studied sites showed a high prevalence of S. mutans, except the control group. Specifically, 60% of ANT and 70% of AAA/SNT paired samples were positive for S. mutans. The cnm gene was detected positive for S. mutans only in ANT samples (66.6%). The cbm gene was not detected in any of the investigated sites. CONCLUSIONS: S. mutans was found in high prevalence in both asymptomatic and symptomatic endodontic infections, including in abscesses, but it was not detected in the root canals of teeth with normal vital pulp. Interestingly, cnm+ S. mutans was only detected in asymptomatic/chronic primary endodontic infections associated with apical lesion. Therefore, it appears that cnm, and possibly other CBPs, may play an underestimated role in chronic endodontic infections. CLINICAL RELEVANCE: A high prevalence of Streptococcus mutans cnm+ gene was detected only in asymptomatic primary endodontic infections associated with apical lesion. Therefore, it appears that this collagen-binding protein gene plays an underestimated role in asymptomatic/chronic endodontic infections.

Prevalence of red and orange microbial complexes in endodontic-periodontal lesions: a systematic review and meta-analysis.

OBJECTIVE: This systematic review assesses the prevalence of microbial complexes in endodontic-periodontal lesion. MATERIALS AND METHODS: Nine databases were searched through August 2020. Experts were consulted to indicate additional studies. Studies were blindly selected by two reviewers based on pre-defined eligibility criteria. Studies that evaluated the prevalence of microbial orange and red complexes among patients with endodontic-periodontal lesion were considered eligible. Risk of bias was assessed using the Joanna Briggs Institute Critical Appraisal Checklist for Studies Reporting Prevalence Data. RESULTS: From 572 articles found on all databases, 11 clinical studies were finally included. The following microorganisms were investigated: P. gingivalis, T. forsythia, T. denticola, F. nucleatum, F. periodonticum, P. micra, P. intermedia, P. nigrescens, S. constellatus, C. gracilis, C. rectus, C. showae and E. nodatum. Considering the orange complex, P. micra, E. nodatum and S. constellatus were prevalent in both root canal and periodontal pockets. P. gingivalis and T. forsythia belonging to the red complex were prevalent only in periodontal pockets. The red complex microorganisms were not found very frequently in root canal. CONCLUSIONS: There is a similarity between the microbiome of root canal and periodontal pockets, with prevalence of the three microorganisms of the orange complex. Two microorganisms from the red complex were prevalent only in periodontal pockets. CLINICAL RELEVANCE: The prevalence of specific microorganisms in endodontic-periodontal lesion is important to understand the microbiological profile of the patients involved and to correlate it with possible clinical and repair conditions of this pathology.

Physicochemical, antimicrobial, and biological properties of White-MTAFlow.

OBJECTIVES: To evaluate a new material containing tantalum oxide as an alternative radiopacifier, and a water-based gel for hydration, in comparison with two calcium silicate-based cement: ProRoot MTA and Biodentine. MATERIALS AND METHODS: ProRoot MTA (Dentsply), Biodentine (Septodont), and a new hydraulic calcium silicate cement White-MTAFlow (Ultradent) (in 'thin' consistency) were characterized using X-ray diffraction (XRD), scanning electron microscopy (SEM), and energy-dispersive spectroscopy (EDS). The interaction with dentin was also assessed using SEM and EDS. Physical and chemical properties radiopacity, setting time, linear flow, volumetric central filling, and lateral flow, pH, and volume change were investigated together with the color luminosity (L) and color change (ΔE). The agar diffusion and direct contact antimicrobial activity, and methylthiazolyldiphenyl-tetrazolium-bromide (MTT) cytotoxicity using human fibroblast cells were also evaluated. Data were statistically analyzed at a 5% significance level. RESULTS: All materials were composed of tricalcium and dicalcium silicate but had different radiopacifiers, and calcium hydroxide (portlandite) deposition was detected in XRD analysis. White-MTAFlow exhibited radiopacity values in accordance with ISO standard, and the longest setting time. The water-based gel provided the highest linear flow, a comparable cavity central filling, and the highest groove-lateral flow in the volumetric flow analysis. White-MTAFlow exhibited an alkalinity reduction, and Biodentine, a progressive increase of pH values after 28 days. However, similar volume loss for White-MTAFlow was assessed in comparison to Biodentine after the 28-day immersion. White-MTAFlow showed the highest L value (91.5), and ProRoot MTA the lowest (78.1) due to dentin staining caused by bismuth migration. None of the materials exhibited inhibition halos against the tested bacteria, and similar turbidity values were obtained after 48 h in direct contact with E. faecalis, indicating an upregulation to bacterial growth. White-MTAFlow showed MTT cytocompatibility similarly to the control group. CONCLUSIONS: White-MTAFlow in 'thin' consistency presents comparable physicochemical, biological, and antimicrobial properties to ProRoot MTA and Biodentine, and does not cause color alteration in dentin. CLINICAL RELEVANCE: White-MTAFlow is a suitable material for use as reparative endodontic cement. Further studies considering its biocompatibility are necessary.

Pathogenic potential of Enterococcus faecalis strains isolated from root canals after unsuccessful endodontic treatment.

OBJECTIVES: To evaluate strains of Enterococcus faecalis isolated from endodontic failures cases for (a) presence of virulence genes, namely, gelatinase production (gelE), surface protein (esp), collagen-binding adhesin (ace), cytolysin activator (cylA), E. faecalis antigen A (efaA) and aggregation substance (asa), all by using PCR; (b) biofilm formation capacity; and (c) activity of gelatinase and ß-lactamase. MATERIALS AND METHODS: Twenty-five strains of E. faecalis were tested. The DNA extracted from these strains was used for identification of virulence genes by PCR and 1% agarose gel. Biofilm formation was performed on polystyrene microplates by using the violet crystal staining method. For assessment of the gelatinase activity, inoculum of pure cultures was deposited in tubes containing gelatin and a nutrient broth, whereas nitrocefin disks were used to assess the ß-lactamase action. RESULTS: The virulence genes efaA and cylA were detected in 100% of the strains, whereas gelE was present in 84%, ace in 68%, esp in 56% and asa in 48%. Four strains had no biofilm formation, 17 had poor formation and four had moderate formation. Gelatinase production was observed in three strains and ß-lactamase resistance in five strains of E. faecalis. TOPIC: Diverse patterns of virulence gene detection were observed among the E. faecalis strains, with predominance of those capable of forming biofilm. A few strains have been found to hydrolyze gelatin proteins, whereas ß-lactamase resistance was detected in different isolates. CLINICAL RELEVANCE: To understand the influence of virulence factors in E. faecalis on the host heath status.

Evaluation of the presence of microorganisms from root canal of teeth submitted to retreatment due to prosthetic reasons and without evidence of apical periodontitis.

AIM: The objective of this study was to evaluate the presence of microorganisms in the root canals (RC) of teeth submitted to endodontic retreatment due to prosthetic reasons and without evidence of apical periodontitis. MATERIAL AND METHODS: Twenty teeth referred to endodontic retreatment due to prosthetic reasons and without evidence of apical periodontitis were included in this study. Gutta-percha (GP) was collected from each third of the RC. After GP removal, microbial samples were also collected from the full extension of RC using paper points and files. The samples were cultivated in Fastidious Anaerobe Agar and the colony-forming units (CFU/mL) were counted. They were also semi-quantitatively analyzed by checkerboard DNA-DNA hybridization using 40 DNA bacterial probes. RESULTS: Microorganisms were found in all samples. The coronal third of GP was more contaminated than the apical third (p ≤ 0.05). There was prevalence of Enterococcus hirae and Enterococcus faecalis in all RC thirds and also in the samples collected from the full extension of RCs. CONCLUSION: Microorganisms were found in all cases referred to endodontic retreatment due to prosthetic reasons and without evidence of apical periodontitis. Enterococcus was the genus most frequently detected. CLINICAL SIGNIFICANCE: An endodontic retreatment should be considered before replacing a prosthesis.

Periodontal and endodontic infectious/inflammatory profile in primary periodontal lesions with secondary endodontic involvement after a calcium hydroxide-based intracanal medication.

OBJECTIVE: The aim of the present study was to investigate the effects of a calcium hydroxide-based intracanal medication (ICM) on periodontal and endodontic infectious/inflammatory contents and on periodontal clinical parameters in teeth with primary periodontal lesion and secondary endodontic involvement. MATERIALS AND METHODS: Ten patients with abnormal pulp test results and deep probing depth derived from primary periodontal disease with secondary endodontic involvement were included. Samples were collected from root canals (RC) and periodontal pockets (PP) in order to investigate the microbiological status, levels of endotoxin (LPS), cytokines, and matrix metalloproteinases (MMP), before and after ICM. PCR was used for microbiological assessment. The kinetic-chromogenic LAL assay was used for LPS quantification. Quantikine ELISA kits were used for measurement of IL-1 α, IL-1 ß, TNF-α, PGE2, MMP-2, MMP-3, MMP-8, MMP-9, and MMP-13 levels. The statistical analyses were made using the Friedman and Wilcoxon tests (p < 0.05). T test was used to compare data on periodontal characteristics. RESULTS: ICM did not reduce the number of microorganisms in PP and RC, except for Fusobacterium nucleatum in RC. There was a significant reduction in LPS, MMPs, IL-1 ß, and TNF-α levels in PP after ICM. In RC, LPS, MMP13, PGE2, and IL-1ß levels remained unaltered (p > 0.05); however, the levels of the other MMPs and cytokines were reduced (p < 0.05). After 1 year of the root canal treatment, tooth mobility was significantly reduced (p ≤ 0.05). CONCLUSIONS: The use of a calcium hydroxide-based ICM showed positive effects for periodontal treatment prognosis, as it reduced LPS, cytokine, and MMP levels in periodontal pockets. CLINICAL SIGNIFICANCE: Patients presenting deep probing depth and undergoing periodontal treatment for at least 6 months, with no positive response to periodontal therapy, might benefit with the endodontic treatment.

Effectiveness of calcium hydroxide-based intracanal medication on infectious/inflammatory contents in teeth with post-treatment apical periodontitis.

OBJECTIVE: The objective of this work was to investigate in vivo the effects of calcium hydroxide-based intracanal medication (ICM) on the levels of bacteria, pro-inflammatory cytokines (PICs), and matrix metalloproteinases (MMPs) in root canals and periradicular tissues of teeth with failure of the root canal treatment and apical periodontitis. MATERIALS AND METHODS: Twenty infected root canals of single-rooted teeth were randomly assigned into two groups according to the irrigant used for chemomechanical preparation (CMP) (n = 10 per group): G1 - 2% chlorhexidine (CHX) gel and G2 - 6% sodium hypochlorite (NaOCl). Root canal contents were taken by using paper points before CMP (S1) and after 30 days of calcium hydroxide-based ICM (S2). Microbial reduction was calculated by means of colony-forming unit count (CFU/mL), with PICs and MMPs (pg/mL) being measured by using enzyme-linked immunosorbent assay (ELISA). RESULTS: Culturable bacteria (101.2 ± 79.2), PICs (IL-1ß 1.2 ± 0.4 and TNF-α 8.8 ± 4.7), MMP-2 (803.7 ± 96.4), MMP-3 (453.9 ± 229.3), MMP-8 (245.9 ± 122.4), MMP-9 (129.4 ± 29.6), and MMP-13 (70.8 ± 12.8) were present in all S1 samples. After 30 days of ICM (S2), a 99.5% microbial reduction was observed, together with a significant reduction of PICs in all groups. Overall, it was observed a decrease in the levels of MMPs (S2), except MMP-13, which was found in increased levels after ICM (P < .05), independently of the groups. CONCLUSIONS: Calcium hydroxide-based intracanal medications have had a positive effect on the microbial reduction by decreasing the levels of PICs and MMPs. Both auxiliary chemical substances (i.e., 2% CHX and 6% NaOCl) presented similar effects when calcium hydroxide was used as intracanal medication. CLINICAL RELEVANCE: Teeth with failure of the root canal treatment and apical periodontitis, and consequently with high levels of bacteria, PIC, and MMP, may present a better prognosis after a 30 days of a calcium hydroxide-based ICM.

Clinical investigation of bacterial species and endotoxin in endodontic infection and evaluation of root canal content activity against macrophages by cytokine production.

INTRODUCTION: This study investigated the presence of different Gram-negative bacterial species and the levels of endotoxins found in primary endodontic infection (PEI), determining their stimulation ability against macrophages through the levels of interleukin (IL)-1, IL-6, IL-10, and tumor necrosis factor alpha (TNF-α), and evaluated their relationship with clinical and radiographic findings. MATERIAL AND METHODS: Samples were taken from 21 root canals with primary endodontic infection with apical periodontitis (PEIAP). Molecular techniques were used for bacterial detection. Limulus amebocyte lysate assay was used to measure endotoxins. Pro-inflammatory cytokines were measured by ELISA assay. RESULTS: All samples were positive for bacterial DNA (21/21). Prevotella nigrescens (57.2 %) was the most frequent species. Higher levels of endotoxins were found in teeth with pain on palpation and exudation (all p < 0.05). Positive correlations were found between endotoxins and the levels of TNF-α and IL-1ß, whereas a negative correlation was found between endotoxin and the amount of IL-10 (p < 0.05). Endotoxin levels were found to be a risk factor for exudation and increased the number of Gram-negative bacterial species for the presence of a larger area of bone destruction (all p < 0.05). CONCLUSION: A wide variety of Gram-negative bacterial species are involved in primary endodontic infection, with participation of different Treponema species. Thus, the levels of endotoxins and the number of Gram-negative bacteria species present in root canals were considered risk factors for the severity of endodontic infection. CLINICAL RELEVANCE: The present study revealed that Gram-negative bacterial species and endotoxins play an important role in the development of signs/symptoms and the severity of bone destruction, this knowledge is essential for the establishment of an effective therapy.