José F. Siqueira

Photodynamic Therapy with Two Different Photosensitizers as a Supplement to Instrumentation/Irrigation Procedures in Promoting Intracanal Reduction of Enterococcus faecalis

INTRODUCTIONnThis in vitro study aimed to investigate the antibacterial effects of photodynamic therapy (PDT) with methylene blue (MB) or toluidine blue (TB) (both at 15 microg/mL) as a supplement to instrumentation/irrigation of root canals experimentally contaminated with Enterococcus faecalis.nnnMETHODSnSeventy extracted teeth had their root canals contaminated with an endodontic strain of E. faecalis for 7 days, instrumented with nickel-titanium instruments and irrigated either with 2.5% NaOCl or with 0.85% NaCl, and then randomly distributed into four experimental groups: MB/NaOCl (PDT with MB and NaOCl as the irrigant), TB/NaOCl (PDT with TB and NaOCl as the irrigant), MB/NaCl (PDT with MB and NaCl as the irrigant), and TB/NaCl (PDT with TB and NaCl as the irrigant). For PDT, the photosensitizer remained in the canal for 2 minutes before exposed to red light emitted from a diode laser for 4 minutes. Samples were taken before and after instrumentation/irrigation and following the specific PDT procedure for each group, plated onto Mitis-salivarius agar and the colony forming units counted.nnnRESULTSnRegardless of the irrigant used (NaOCl or NaCl), instrumentation significantly reduced bacterial counts in comparison to the baseline (p < 0.001). NaOCl as the irrigant was significantly more effective than NaCl, and this difference persisted after PDT, irrespective of the photosensitizer used (p < 0.05). PDT with either MB or TB did not significantly enhance disinfection after chemomechanical preparation using NaOCl as irrigant (p > 0.05). No significant differences were observed between the two photosensitizers (p > 0.05).nnnCONCLUSIONnThese in vitro results suggest that PDT with either MB or TB may not exert a significant supplemental effect to instrumentation/irrigation procedures with regard to intracanal disinfection. Further adjustments in the PDT protocol may be required to enhance predictability in bacterial elimination before clinical use is recommended.

Ability of Chemomechanical Preparation with Either Rotary Instruments or Self-adjusting File to Disinfect Oval-shaped Root Canals

INTRODUCTIONnOval-shaped root canals might represent a great challenge for proper disinfection. This study compared the capability of a newly developed instrument, the self-adjusting file (SAF), and rotary nickel-titanium (NiTi) instrumentation to eliminate Enterococcus faecalis populations from long oval root canals of extracted human teeth. As a secondary purpose, the ability of a modification in sampling technique to recover bacteria lodged in recesses of oval canals was evaluated.nnnMETHODSnLong oval canals from mandibular incisors and maxillary second premolars were infected with E. faecalis (ATCC 29212) for 30 days and then randomly distributed into 2 experimental groups. In group 1, canals were prepared up to a 40/04 rotary BioRaCe instrument by using irrigation with NaviTip needles; in group 2, canals were prepared by using the SAF system with continuous irrigation. NaOCl and ethylenediaminetetraacetic acid were used as irrigants. Bacteriologic samples were taken before (S1) and after preparation (S2a and S2b).nnnRESULTSnReduction in the bacterial populations was highly significant in both groups (P < .001). Preparation of long oval canals with the SAF was significantly more effective than rotary NiTi instrumentation in reducing intracanal E. faecalis counts (P = .01). Frequency of positive cultures in S2 samples was 11 of 20 (55%) for rotary instrumentation and 4 of 20 (20%) for SAF instrumentation (P = .048). S2b samples (modified method) yielded more positive samples than S2a (12/40 vs 5/40), but this difference reached no statistical significance (P > .05).nnnCONCLUSIONSnThe SAF system was significantly more effective than rotary NiTi instrumentation used with syringe/needle irrigation in disinfecting long oval root canals in vitro. A modified sampling technique might be necessary for oval canals.

Molecular Detection of Black-Pigmented Bacteria in Infections of Endodontic Origin

A 16S rDNA-directed polymerase chain reaction method was used to assess the occurrence of four black-pigmented anaerobic rods in root canal infections. Samples were obtained from 54 infected teeth. Ten cases were diagnosed as acute periradicular abscesses. DNA was extracted from the samples and analyzed using a polymerase chain reaction-based identification assay. The method allowed detection of black-pigmented bacteria anaerobes in 59.3% of the examined teeth. Twelve cases yielded more than one black-pigmented species. In general Porphyromonas endodontalis was found in 42.6%, Porphyromonas gingivalis in 27.8%, Prevotella nigrescens in 7.4%, and Prevotella intermedia in 5.6% of the cases. P. endodontalis was found in 70% of the pus samples, P. gingivalis in 40%, and P. intermedia in 10%. P. gingivalis was always found associated with P. endodontalis in abscessed teeth. P. nigrescens was not found in any pus sample. The high prevalence of P. endodontalis and P. gingivalis suggests that they can play an important role in the pathogenesis of periradicular diseases.

Effects of Electropolishing Surface Treatment on the Cyclic Fatigue Resistance of BioRace Nickel-Titanium Rotary Instruments

INTRODUCTIONnThis study evaluated the influence of electropolishing surface treatment on the number of cycles to fracture of BioRace rotary nickel-titanium endodontic instruments.nnnMETHODSnBioRace size BR5C instruments with or without electropolishing surface treatment were used in an artificial curved canal under rotational speed of 300 rpm until fracture. Fractured surfaces and the helical shafts of fractured instruments were analyzed by scanning electron microscopy (SEM).nnnRESULTSnPolished instruments displayed a significantly higher number of cycles to fracture when compared with nonpolished instruments (P < .001). Actually, the number of cycles to fracture of a polished BR5C instrument was 124% higher than that of a nonpolished instrument. SEM analysis showed that the fractured surface of both polished and nonpolished BR5C instruments had ductile morphologic characteristics. Evaluation of the separated fragments after cyclic fatigue testing showed the presence of microcracks near the fracture surface. Polished instruments exhibited fine cracks that assumed an irregular path (zigzag crack pattern), whereas nonpolished instruments showed cracks running along the machining grooves.nnnCONCLUSIONSnElectropolishing surface treatment of BioRace endodontic instruments significantly increased the cyclic fatigue resistance.

Supplementing the Antimicrobial Effects of Chemomechanical Debridement with Either Passive Ultrasonic Irrigation or a Final Rinse with Chlorhexidine: A Clinical Study

INTRODUCTIONnThe ability of 2 different approaches to supplement the antimicrobial effects of chemomechanical debridement in infected root canals was compared in vivo.nnnMETHODSnSamples from necrotic root canals of teeth with apical periodontitis were taken at the baseline (S1), after preparation with rotary nickel-titanium BioRaCe instruments and 2.5% NaOCl irrigation (S2), and then after either passive ultrasonic irrigation (PUI) for activation of NaOCl (n = 13) or a final rinse with 2% chlorhexidine (CHX) (n = 14) (S3). The incidence of positive culture for bacteria and fungi as well as positive broad-range polymerase chain reaction (PCR) results for bacteria, fungi, and archaea was determined.nnnRESULTSnAll S1 samples were positive for bacteria in all methods. Fungi were not detected, and archaea occurred in only one S1 sample. Treatment procedures were significantly effective in reducing the incidence of positive culture and PCR results. Although both supplementary approaches reduced the incidence of positive bacteriologic results when compared with postinstrumentation samples, reduction was not statistically significant (P > .05). There was no significant difference for intergroup comparisons either (P > .05).nnnCONCLUSIONSnAlthough supplementary disinfection with either PUI or a final rinse with CHX can reduce the number of cases with positive culture and PCR results for bacteria, many cases still remain with detectable bacteria in the main root canal. Research on alternative or supplementary antimicrobial methods or substances should be encouraged.

Apical Root Canal Microbiota as Determined by Reverse-capture Checkerboard Analysis of Cryogenically Ground Root Samples from Teeth with Apical Periodontitis

INTRODUCTIONnBacteria located in the apical root canal system potentially participate in the pathogenesis of apical periodontitis. Detection and identification of apical bacteria can be compromised because of limitations in conventional sampling and identification procedures. This study identified several bacterial taxa in the apical and middle/coronal segments of primarily infected root canal system by using pulverized root segments and a culture-independent molecular method.nnnMETHODSnSeventeen extracted teeth with attached apical periodontitis lesions were sectioned to obtain 2 root fragments (apical and middle/coronal segments). Root fragments were cryogenically ground, and DNA was extracted from samples. After multiple displacement amplification, DNA from samples was used as template in a reverse-capture checkerboard hybridization assay targeting 28 bacterial taxa.nnnRESULTSnBacterial DNA was detected in all samples. The most prevalent taxa in the apical root canal system were Olsenella uli (76.5%), Prevotella baroniae (71%), Porphyromonas endodontalis (65%), Fusobacterium nucleatum (53%), and Tannerella forsythia (47%). O. uli, P. endodontalis, and Propionibacterium acnes were as frequently detected in apical samples as they were in middle/coronal samples. P. baroniae, T. forsythia, and F. nucleatum were found more frequently in the apical part of the canal as compared with matched coronal segments. Streptococcus species were more prevalent in middle/coronal samples. The median and mean of shared bacterial taxa between matched apical and middle/coronal segments were 27% and 41%, respectively.nnnCONCLUSIONSnSeveral candidate endodontic pathogens were very prevalent in the apical root canal system. The apical microbiota was usually complex and differed in species composition when compared with the microbiota of middle/coronal samples from the same tooth.

Characterization of virulence factors and clonal diversity of Enterococcus faecalis isolates from treated dental root canals.

The high prevalence of Enterococcus faecalis in root canal treated teeth with post-treatment disease, as evidenced by both molecular and traditional culturing methods, suggests that this species may be a key player in endodontic treatment failure. This study aimed to detect virulence factors by phenotypic and western blotting tests, and virulence genes by PCR from 20 clinical strains of E. faecalis isolated from treated root canals of teeth with (10) or without (10) apical periodontitis. Moreover, genomic diversity of these strains was assessed by pulsed-field gel electrophoresis (PFGE) and rep-PCR. All 20 strains presented the gelE gene (gelatinase), but 10 of them did not hydrolyze gelatin. Seven of the 10 gelatinase-producing isolates were recovered from root canals with lesions, which suggests a role for this virulence factor in the pathogenesis of post-treatment disease. The esp gene was expressed only in cases where gelatinase production was negative. The other virulence genes were found in 90% (efaA and ace genes), 45% (agg gene) and 95% (cpd gene) of the E. faecalis isolates. As for PFGE and rep-PCR, no specific clonal type of E. faecalis was found in association with teeth with or without disease, revealing the interindividual clonal diversity of endodontic infections.

Recontamination of coronally unsealed root canals medicated with camphorated paramonochlorophenol or calcium hydroxide pastes after saliva challenge

This in vitro study evaluated the ability of some medications to prevent recontamination of coronally unsealed root canals by bacteria from saliva. The medications tested were camphorated paramonochlorophenol (CPMC) applied in cotton pellets in the pulp chamber; calcium hydroxide/saline solution paste filling the root canal; and calcium hydroxide/CPMC/glycerin paste also filling the root canal. Medicated canals were exposed to saliva, and the number of days required for total recontamination to occur was recorded. Canals medicated with CPMC in cotton pellets were thoroughly recontaminated within an average of 6.9 days. Canals filled with calcium hydroxide/saline solution and calcium hydroxide/CPMC/glycerin showed entire recontamination within an average of 14.7 and 16.5 days, respectively. Calcium hydroxide pastes were significantly more effective than CPMC (p < 0.05).

Detection of antibiotic resistance genes in samples from acute and chronic endodontic infections and after treatment

OBJECTIVEnThe purpose of this study was twofold: survey samples from acute and chronic endodontic infections for the presence of genes encoding resistance to beta-lactams, tetracycline and erythromycin, and evaluate the ability of treatment to eliminate these genes from root canals.nnnDESIGNnDNA extracts from samples of abscess aspirates (n=25) and root canals of teeth with asymptomatic apical periodontitis (n=24) were used as template for direct detection of the genes blaTEM, cfxA, tetM, tetQ, tetW, and ermC using real-time polymerase chain reaction (PCR). Bacterial presence was determined using PCR with universal bacterial primers. Root canals of the asymptomatic cases were also sampled and evaluated after chemomechanical procedures using NiTi instruments with 2.5% NaOCl irrigation.nnnRESULTSnAll abscess and initial root canal samples were positive for bacteria. At least one of the target resistance genes was found in 36% of the abscess samples and 67% of the asymptomatic cases. The most prevalent genes in abscesses were blaTEM (24%) and ermC (24%), while tetM (42%) and tetW (29%) prevailed in asymptomatic cases. The blaTEM gene was significantly associated with acute cases (p=0.02). Conversely, tetM was significantly more prevalent in asymptomatic cases (p=0.008). Treatment eliminated resistance genes from most cases.nnnCONCLUSIONSnAcute and chronic endodontic infections harboured resistance genes for 3 classes of widely used antibiotics. In most cases, treatment was effective in eliminating these genes, but there were a few cases in which they persisted. The implications of persistence are unknown. Direct detection of resistance genes in abscesses may be a potential method for rapid diagnosis and establishment of proactive antimicrobial therapy.

Antibiotic resistance genes in anaerobic bacteria isolated from primary dental root canal infections

Fourty-one bacterial strains isolated from infected dental root canals and identified by 16S rRNA gene sequence were screened for the presence of 14 genes encoding resistance to beta-lactams, tetracycline and macrolides. Thirteen isolates (32%) were positive for at least one of the target antibiotic resistance genes. These strains carrying at least one antibiotic resistance gene belonged to 11 of the 26 (42%) infected root canals sampled. Two of these positive cases had two strains carrying resistance genes. Six out of 7 Fusobacterium strains harbored at least one of the target resistance genes. One Dialister invisus strain was positive for 3 resistance genes, and 4 other strains carried two of the target genes. Of the 6 antibiotic resistance genes detected in root canal strains, the most prevalent were blaTEM (17% of the strains), tetW (10%), and ermC (10%). Some as-yet-uncharacterized Fusobacterium and Prevotella isolates were positive for blaTEM, cfxA and tetM. Findings demonstrated that an unexpectedly large proportion of dental root canal isolates, including as-yet-uncharacterized strains previously regarded as uncultivated phylotypes, can carry antibiotic resistance genes.