Bacteria In Infected Root Canals Research Articles

Improving the microbial sampling and analysis of secondary infected root canals by passive ultrasonic irrigation

ObjectiveThe persistence of pathogenic microorganisms in root canals is the most common reason for the failure of root canal treatment and the necessity of a root filling treatment, which results in an uncertain prognosis due to technical complexity and the variety of highly adaptable microorganisms.This study evaluated the effect of passive ultrasonic irrigation (PUI) on the outcome of the microbial analysis of root canal-treated teeth with persistent or recurrent apical inflammation in vivo.Materials and methodsSample collection was performed after root filling removal (sample S1, control group) and after PUI with NaCl (sample S2) using sterile paper points. In total, 19 samples were obtained. Quantification was performed by means of serial dilution of the samples. Subcultivated pure cultures were identified using MALDI-TOF MS complemented by the Vitek-2-System or PCR, followed by sequencing of the 16S rRNA gene. The results of the samples (S1 and S2) were evaluated regarding their bacterial count and composition.ResultsThe total count of bacteria and the number of aerobic/facultative anaerobic microorganisms significantly increased in the S2-samples after application of PUI. The number of obligate anaerobic microorganisms showed an increase after PUI, although it was not significant. We detected 12 different aerobic/facultative anaerobic microorganisms before PUI, and in 21 cases after PUI. Two different obligate anaerobic microorganisms were found in S1 samples compared to nine different species in S2 samples.ConclusionsPUI is a powerful method for detaching bacteria in infected root canals and enables a more precise analysis of the etiology of persistent endodontic infections.Clinical relevanceThis study indicates that PUI exerts a positive cleansing effect and adds to the accessibility of microorganisms during the application of bactericidal rinsing solution in root canal treatments.

Microbiological analysis of primary infected root canals with symptomatic and asymptomatic apical periodontitis of young permanent teeth

Background/Aim: Understanding the composition of bacteria in infected root canals is important for ameliorating the treatment strategies that lead to the elimination of pathogens and infection control, but also prevent reinfection. Aim of this study was to investigate microbial composition of primary infected root canals with apical periodontitis of young permanent teeth, originating form school children in Serbia, and its association with clinical symptoms. Material and Methods: To determine the bacterial composition of infected root canals in children, 35 endodontic samples were obtained. The identification of cultured bacteria was performed by MALDI-TOF MS analysis. The presence or absence of clinical symptoms were recorded. Results: Facultative anaerobes were 2,2 times more frequent than obligate anaerobes. The most common facultative anaerobes belonged to following genera, Streptococcus (58 isolates), Actinomyces (10) and Enterococcus (8), while predominant obligate anaerobes, belonged to genera Veillonella (15), Prevotella (9) and Fusobacterium (8). The most common clinical isolates recovered from infected root canals with symptomatic apical periodontitis were Veillonella parvula (10) and Fusobacterium nucleatum (7), while from the asymptomatic ones, they were Streptococcus mitis/Streptococcus oralis (5). Prevalence of Parvimonas micra, Prevotella buccae and Streptococcus constellatus within the root canals might be associated to clinical symptoms. Conclusions: Species of genera Streptococcus and Veillonella were the most common isolates from primary infected root canals with apical periodontitis in Serbian school children. Facultative anaerobes were predominant over obligate anaerobes. The prevalence of obligate anaerobes was much higher in symptomatic compared to asymptomatic root canal infections. No specific bacterial strain might be associated to a single examined clinical symptom (pain, tenderness to percussion or swelling), but majority of the strains are associated to all of the examined three symptoms.

Investigation of different sodium hypochlorite volumes, concentrations and times of irrigation in endodontic therapy: a systematic review

Investigation of different sodium hypochlorite volumes, concentrations and times of irrigation in endodontic therapy: a systematic review

Combinations of bacterial species associated with symptomatic endodontic infections in a Chinese population.

To use microarrays to detect 11 selected bacteria in infected root canals, revealing bacterial combinations that are associated with clinical symptoms and signs of primary endodontic infections in a Chinese population. DNA was extracted from 90 samples collected from the root canals of teeth with primary endodontic infections in a Chinese population, and the 16S rRNA gene was amplified by polymerase chain reaction (PCR). The PCR products were hybridized to microarrays containing specific oligonucleotide probes targeting 11 species, and the arrays were screened with a confocal laser scanner. Pearson's chi-squared test and cluster analysis were performed to investigate the associations between the bacterial combinations and clinical symptoms and signs using SAS 8.02. Seventy-seven samples (86%) yielded at least one of the 11 target species. Parvimonas micra (56%), Porphyromonas endodontalis (51%), Tannerella forsythia (48%), Prevotella intermedia (44%) and Porphyromonas gingivalis (37%) were the most prevalent taxa and were often concomitant. The following positive associations were found between the bacterial combinations and clinical features: P. endodontalis and T. forsythia with abscess; P. gingivalis and P. micra with sinus tract; P. gingivalis and P. endodontalis or P. micra and P. endodontalis with abscess and sinus tract; and the combination of P. endodontalis, P. micra, T. forsythia and P. gingivalis with sinus tract (P < 0.05). Various combinations of P. micra, P. endodontalis, T. forsythia and P. gingivalis may contribute to abscesses or sinus tracts of endodontic origin with bacterial synergism in a Chinese population.

In vitro activity of photoactivated disinfection using a diode laser in infected root canals

Objective. To investigate the lethal activity of photoactivated disinfection (PAD) on Enterococcus faecalis (ATCC 29212) and mixed populations of aerobic or anaerobic bacteria in infected root canals using a diode laser after the application of a photosensitizer (PS). Materials and methods. First, the bactericidal activity of a low power diode laser (200 mW) against E. faecalis ATCC 29212 pre-treated with a PS (toluidine blue) for 2 min were examined after different irradiation times (30 s, 60 s and 90 s). The bactericidal activity in the presence of human serum or human serum albumin (HSA) was also examined. Second, root canals were infected with E. faecalis or with mixed aerobic or anaerobic microbial populations for 3 days and then irrigated with 1.5% sodium hypochlorite and exposed to PAD for 60 s. Results. Photosensitization followed by laser irradiation for 60 s was sufficient to kill E. faecalis. Bacteria suspended in human serum (25% v/v) were totally eradicated after 30 s of irradiation. The addition of HSA (25 mg/ml or 50 mg/ml) to bacterial suspensions increased the antimicrobial efficacy of PAD after an irradiation time of 30 s, but no longer. The bactericidal effect of sodium hypochlorite was only enhanced by PAD during the early stages of treatment. PAD did not enhance the activity of sodium hypochlorite against a mixture of anaerobic bacteria. Conclusions. The bactericidal activity of PAD appears to be enhanced by serum proteins in vitro, but is limited to bacteria present within the root canal.

Rapid Quantification of Bacteria in Infected Root Canals Using Fluorescence Reagents and a Membrane Filter: A Pilot Study on Its Clinical Application to the Evaluation of the Outcomes of Endodontic Treatment

Objective. The bacterial examination has been performed during the course of the root canal treatment. In the present pilot study, the new developed method, using fluorescence reagents and a membrane filter, was applied to the detection and quantification of bacteria in infected root canals, in order to evaluate the outcomes of the treatment. Methods. Six infected root canals with periapical lesions from 5 subjects were included. Informed consent was obtained from all subjects (age ranges, 23–79 years). Samples from infected root canals were collected at the beginning of the treatment (termed #25 First), the end of the first day of treatment (termed #55 First), and the next appointment day (termed #55 Second). Then, the bacterial count (CFU) was measured using fluorescence reagents (4′,6′-diamidino-2-phenylindole and propidium iodide) and the polycarbonate membrane filter by Bioplorer. Results. The mean ± SD of CFU in the sample of “#25 First” was (1.0 ± 1.4) × 105. As the root canal treatment progressed, the CFU decreased as 7.9 × 103 (#55 First) and 4.3 × 102 (#55 Second). Conclusion. In the present pilot study, rapid detection and quantification of bacteria in infected root canals were found to be successfully performed using fluorescence reagents and a membrane filter (Bioplorer analysis).

Evaluation of Antibacterial Effects by Atmospheric Pressure Nonequilibrium Plasmas against Enterococcus faecalis Biofilms In Vitro

IntroductionThe aim of this study was to evaluate the in vitro antibacterial activity by atmospheric pressure nonequilibrium plasmas (APNPs) as an effective approach against bacterial biofilms in root canal systems during endodontic therapy. MethodsSterile cover slips were placed into the wells of tissue culture plates to permit the formation of Enterococcus faecalis biofilms. Biofilms were treated for 5 minutes with APNPs or 2% chlorhexidine digluconate (CHX). The viability of biofilm bacteria was analyzed by staining and confocal laser scanning microscopy. In addition, infected single-rooted teeth were exposed to APNPs or 2% CHX for 5, 10, and 15 minutes. After treatment, the root canals were flushed, and the resulting suspensions were inoculated onto brain-heart infusion agar to assess bacterial survival. Finally, micro–computed tomography scanning was used to observe and verify the root canal systems relative to the antibacterial effects obtained. ResultsTreatment for 5 minutes with APNPs or 2% CHX killed the majority of bacteria in the E. faecalis biofilms. Moreover, APNP treatment was as effective as 2% CHX for inactivating bacteria in infected root canals (P > .05). Bacterial survival after treatment with APNPs or 2% CHX remarkably reduced with increasing exposure times (P < .05). There was no significant difference between bacterial survival in complex root canal systems and simple straight canals (P > .05). ConclusionsAPNPs can be an effective adjunct to standard endodontic antimicrobial treatment.

Cultivable bacteria in infected root canals as identified by 16S rRNA gene sequencing

Traditionally, cultivable bacteria isolated from infected root canals have been identified by phenotype-based methods. Because 16S ribosomal RNA (rRNA) gene sequencing has emerged as a more accurate and reliable tool for bacterial identification, the present study applied this approach to identify bacterial isolates recovered from the root canals of teeth with chronic apical periodontitis. Anaerobic techniques were used for culturing; identification of the isolates was carried out by polymerase chain reaction amplification and sequencing of the V5-V8 region of the 16S rRNA gene. Bacteria were found in all samples. The mean number of taxa per canal was 3.1, ranging from 2 to 8. The median number of cultivable bacterial cells in the root canals was 4.2 x 10(5), ranging from 2.8 x 10(3) to 3.3 x 10(7). Eighty-seven strains belonging to 52 bacterial taxa were identified. The most prevalent taxa were Fusobacterium nucleatum, Porphyromonas gingivalis, Pseudoramibacter alactolyticus, Micromonas micros and streptococci. The following bacterial phyla were represented in this study: Firmicutes (22 taxa, 46% of the identified isolates), Actinobacteria (14 taxa, 25.3% of the isolates), Bacteroidetes (eight taxa, 13.8% of the isolates), Fusobacteria (three taxa, 9.2% of the isolates) and Proteobacteria (five taxa, 5.7% of the isolates). Some of the isolates represented unnamed species not previously cultivated and characterized. In conclusion, our findings using a combined anaerobic culture-molecular identification approach confirmed the polymicrobial nature of primary endodontic infections with dominance of anaerobic bacteria. Notably several bacteria that are difficult or impossible to identify by phenotypic means were identified, including previously uncultivated taxa, cultivated-but-not-yet-characterized taxa and newly named species.

Effects of Chemomechanical Preparation With 2.5% Sodium Hypochlorite and Intracanal Medication With Calcium Hydroxide on Cultivable Bacteria in Infected Root Canals

This clinical study was conducted to assess the bacterial reduction after chemomechanical preparation with 2.5% NaOCl as an irrigant and the additive antibacterial effect of intracanal dressing with calcium hydroxide. According to stringent inclusion criteria, 11 teeth with primary intraradicular infections and chronic apical periodontitis were selected and monitored in the study. Bacterial samples were taken at the baseline (before treatment) (S1), after chemomechanical preparation with 2.5% NaOCl as an irrigant (S2), and after a 7-day dressing with a calcium hydroxide paste in glycerin (S3). Cultivable bacteria recovered from infected root canals at the 3 stages were counted and identified by means of 16S rRNA gene sequencing analysis. At S1, all canals were positive for bacteria, with the mean number of 2.8 taxa per canal (range, 1-6). At S2, 5 cases (45.5%) still harbored cultivable bacteria, with 1 or 2 species per canal. At S3, bacteria were cultured from 2 cases (18.2%), with 1 species per positive case. There was no indication that any specific bacterial taxon was more resistant to treatment. A significant reduction in bacterial counts was observed between S1 and S2, and S1 and S3. However, no statistically significant difference was observed for comparisons involving S2 and S3 samples with regard to the number of cases yielding negative cultures (P = .18) or quantitative bacterial reduction (P = .19). It was concluded that the whole antibacterial protocol used in this study significantly reduced the number of bacteria in the canal and rendered most canals free of cultivable bacteria.

On the use of denaturing gradient gel electrophoresis approach for bacterial identification in endodontic infections

Bacteria in infected root canals of teeth evincing chronic apical periodontitis lesions were identified by a polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) approach. DNA was extracted from root canal samples, and part of the 16S rRNA gene of all bacteria was amplified by PCR and separated by DGGE, generating banding patterns representative of the community structure. Twenty visible bands were cut out of the gel, re-amplified, and sequenced to provide identification. Sequencing analysis revealed the presence of both cultivable and as-yet-uncultivated species in the samples analyzed, including representatives of the genera Fusobacterium, Bacteroides, Dialister, Synergistes, Prevotella, Eubacterium and Peptostreptococcus. Unambiguous identification was not always possible and the method's limitations are discussed. In general, the findings showed that PCR-DGGE can be useful for the identification of both cultivable and as-yet-uncultivated bacteria in endodontic infections.

Increased number of anaerobic bacteria in the infected root canal in type 2 diabetic rats

The purpose of this study was to investigate the relationship between type 2 diabetes mellitus and anaerobic bacteria detected in infected root canals. Normal Wistar rats (control) received a standard laboratory diet with water (group A), and GK rats (type 2 diabetes mellitus rats) a normal laboratory diet with water (group B) or a 30% sucrose solution (group C). Chemotaxis assay was conducted on polymorphonuclear leukocytes from the 3 groups, and the numbers of anaerobic bacteria in infected root canals were determined. In the chemotaxis assay on the polymorphonuclear leukocytes, the chemotactic response of cells in group C was lower than that for groups A and B (P < .01). As to bacteria detected in the root canal, obligate anaerobic bacteria which stained gram negative, were significantly more numerous in group C (P < .01) than in groups A and B. The metabolic condition produced by type 2 diabetes mellitus in rats might lower the general host resistance against bacterial infection.

Detection of black-pigmented bacteria in infected root canals

Black-pigmented bacteria have been implicated in the endodontic infections. This group of microorganisms includes Porphyromonas endodontalis, Porphyromonas gingivalis, Prevotella intermedia, and Prevotella nigrescens. The organisms display a wide variety of virulence factors that may be pertinent to acute endodontic infections. The aim of this study was to identify P. endodontalis, P. gingivalis, P. intermedia, and P. nigrescens by using special potency disk test, filter paper spot test, 165 rRNA gene-directed PCR, and API 32A. Microbial samples were collected from root canals of 33 intact teeth with necrotic pulp and/or apical periodontitis. Conventional laboratory methods were used for identification of the strains of black pigmented bacteria. Eighteen of 33 samples were positive for the growth of black-pigmented bacteria Five colonies were cultured from each pure cultured colonies from Brucella agar plate. Seventy seven colonies were positive for the growth of black-pigmented bacteria. Thirty three of 77(42.6%) were identifed as P. nigrescens, 10 of 77(12.9%)were P. gingivalis, 6 of 77(7.8%) were P. endodontalis, 10 of 77(12.9%) were P. intermedia. On the contrary the reference strains of P. nigrescens, experimental strains of P nigrescens was sensitive to kanamycin in special potency disk test. 165 rRNA gene PCR and API test after rapid presumptative identification methods, such as special potency disk test and filter paper spot test, would be accurate detection methods for black-pigemented bacteria.