The effect of a chemomechanical protocol to eliminate microorganisms from pulpectomized primary teeth: three case reports

Introduction: Description of the bacterial community before and after chemomechanical preparation (CP) with the removal of a smear layer (SL) in pulpectomized primary teeth has been little reported. Objective: These case reports describe the presence of total microorganisms and Enterococcus faecalis in root canals of primary incisors before and after CP with SL removal. Case Reports: Microbiological samples were collected from the root canals of three children (3.66±0.58 years old) with necrosis (n=2) and irreversible pulpal inflammation (n=1) in maxillary primary incisors. After teeth isolation with a rubber dam and antisepsis of the operative field, the sample collections were performed with sterile absorbent paper cones before and after the CP, which included irrigation with 2.5% sodium hypochlorite followed by 6% citric acid to remove the SL. The collected samples were analysed immediately at the end of the clinical procedures. The plates were incubated anaerobically for 48 hours at 37°C. The results were expressed as colony forming units (CFU)/mL. Results: Two of the three teeth showed total microorganisms before the CP. One incisor had no microorganisms in the initial collection. No CFU was counted in the samples collected after CP. Moreover, E. faecalis was not observed any time, either before or after the CP. Conclusions: E. faecalis was not detected in any sample, yet two of the three root canals had microorganisms before CP. In cases where microorganisms were initially found, 100% elimination was observed after the applied protocol.

Introdução: A descrição da comunidade bacteriana antes e após o preparo químico-mecânico (PQM) com remoção da smear layer (SL) em dentes decíduos pulpectomizados tem sido pouco relatada. Objetivo: Esses relatos de casos descrevem a presença de microrganismos totais e Enterococcus faecalis em canais radiculares de incisivos decíduos antes e após PQM com remoção de SL. Relatos dos Casos: Amostras microbiológicas foram coletadas do canal radicular de três crianças (3,66 ± 0,58 anos) com necrose (n = 2) e inflamação pulpar irreversível (n= 1) em incisivos decíduos superiores. Após o isolamento dos dentes com dique de borracha e antissepsia do campo operatório, as coletas das amostras foram realizadas com cones de papel absorvente estéril antes e após o PQM, que incluiu irrigação com hipoclorito de sódio 2,5% seguido de ácido cítrico 6% para retirada do SL. As amostras coletadas foram analisadas imediatamente ao final dos procedimentos clínicos. As placas foram incubadas em anaerobiose durante 48 horas a 37°C. Os resultados foram expressos em unidades formadoras de colônias (UFC)/mL. Resultados: Dois dos três dentes apresentaram microrganismos totais antes do PQM. Um incisivo não apresentava microrganismos na coleta inicial. Nenhuma UFC foi contada nas amostras coletadas após o PQM. Além disso, o E. faecalis não foi observado nenhum momento, nem antes, nem depois do PQM. Conclusão: Não foi detectado E. faecalis em nenhuma amostra, porém dois dos três canais radiculares apresentavam microrganismos antes do PQM. Nos casos em que foram encontrados microrganismos inicialmente, observou-se 100% de eliminação após o protocolo aplicado.

Microbiological Aspects of Traumatic Injuries.

After traumatic injuries to teeth, microorganisms may invade the compromised pulp tissue and initiate pulp infection and periapical inflammation. In addition to bone resorption that typically accompanies pulp necrosis, root resorption frequently occurs. Root resorption has several variants that may occur shortly after the trauma or at a later stage. The pathological changes seen after traumatic injuries to teeth are invariably linked to the presence of microbial irritants. The presence of bacterial biofilms in the dental pulp space can be treated with regenerative or therapeutic endodontic procedures. However, necrosis of periodontal ligament is usually terminal for the tooth involved. In this review, the sources of bacteria after traumatic injuries are discussed. The types and role of microorganisms involved in the pathogenesis of endodontic pathosis after traumatic injuries are presented, and contemporary approaches for the management of these conditions are reviewed. Contemporary antimicrobial strategies are discussed. The rationale for the use of systemic and topical antimicrobials is presented. Finally, novel approaches to the use of antimicrobial therapies, particularly in regenerative procedures, are reviewed.

Microbiological aspects of traumatic injuries.

After traumatic injuries to teeth, microorganisms may invade the compromised pulp tissue and initiate pulp infection and periapical inflammation. In addition to bone resorption that typically accompanies pulp necrosis, root resorption frequently occurs. Root resorption has several variants that may occur shortly after the trauma or at a later stage. The pathological changes seen after traumatic injuries to teeth are invariably linked to the presence of microbial irritants. The presence of bacterial biofilms in the dental pulp space can be treated with regenerative or therapeutic endodontic procedures. However, necrosis of periodontal ligament is usually terminal for the tooth involved. In this review, the sources of bacteria after traumatic injuries are discussed. The types and role of microorganisms involved in the pathogenesis of endodontic pathosis after traumatic injuries are presented, and contemporary approaches for the management of these conditions are reviewed. Contemporary antimicrobial strategies are discussed. The rationale for the use of systemic and topical antimicrobials is presented. Finally, novel approaches to the use of antimicrobial therapies, particularly in regenerative procedures, are reviewed.

Comparative Molecular Analysis of Gram-Negative Bacteria in Primary Teeth with Irreversible Pulpitis or Periapical Pathology.

Purpose: The purpose of this study was to compare the Gram-negative pathogens identified in the root canals of primary teeth with irreversible inflammatory pulpitis and in teeth showing apical periodontitis. Methods: Samples were collected from 123 root canals of primary teeth from three- to seven-year-old patients. Root canals were assigned to either group one (irreversible inflammatory pulpitis; n equals 63) or group two (pulp necrosis and apical periodontitis; n equals 60). Total number of cells of selected Gram-negative microorganisms was determined by the checkerboard DNA-DNA hybridization technique. Demographic data were compared using either chi-squared or t tests. Total numbers of microorganisms were compared using the Mann-Whitney test (α equals 0.05). Results: There were no significant intergroup differences in gender, age, and tooth group distribution (P>0.05). Among the 123 samples, 17 were discarded due to salivary contamination. The total numbers of Prevotella nigrescens, Treponema denticola, Fusobacterium nucleatum polymorphum, Fusobacterium nucleatum spp nucleatum, Aggregatibacter actinomycetemcomitans serotype a, Aggregatibacter actinomycetemcomitans serotype b, Porphyromonas gingivalis, Prevotella intermedia, and Prevotella melaninogenica were higher in teeth with apical periodontitis compared to those with irreversible inflammatory pulpitis (P<0.05). Conclusion: Higher numbers of Gram-negative bacteria were found in teeth with apical periodontitis compared to teeth with irreversible in- flammatory pulpitis.

Efficacy of various intracanal medicaments against aerobic and facultative anaerobic microorganism found in human primary teeth with necrotic pulp: A randomized clinical trial.

AIM: The aim of this in vivo study is to evaluate the antimicrobial efficacy of calcium hydroxide (Ca(OH)2), chlorhexidine (CHX), and metronidazole gel as intracanal medicaments against aerobic and facultative anaerobic microorganisms found in root canals of human primary teeth with necrotic pulp. SETTING AND DESIGN: It is a double-blinded randomized clinical trial. MATERIALS AND METHODS: Pulp canals of 45 single-rooted primary maxillary anterior teeth with pulp necrosis in 34 children were included in the study. They were divided into three groups of 15 samples each: Group I: Ca(OH)2; Group II: 1% CHX gel; and Group III: 1% metronidazole gel. Microbial count was obtained from each tooth at two different stages - (1) after instrumentation and (2) after placement of the medication. Statistical analysis using the SPSS 10.0 software program (SPSS Inc., Chicago, IL, USA) with Wilcoxon signed-rank test after grouping the samples was performed. RESULTS: Ca(OH)2, 1% CHX gel, and 1% metronidazole gel were ineffective in completely eliminating aerobic and facultative anaerobic microorganism from root canal of human primary teeth with necrotic pulp. CONCLUSION: None of the commonly used intracanal medicaments, that is Ca(OH)2, 1% CHX gel, and 1% metronidazole gel, was effective in completely eliminating aerobic and facultative anaerobic microorganism from root canal of human primary teeth with necrotic pulp. Ineffectiveness of these medicaments against aerobic and facultative anaerobic microorganism has opened new door of research regarding the manner of bacterial growth in unfavorable environmental and nutritional conditions, the way root canal environment favors biofilm formation and the use of suitable intracanal medicaments against single and multispecies biofilms.

The Radiopacity and Antimicrobial Properties of Different Radiopaque Double Antibiotic Pastes Used in Regenerative Endodontics.

INTRODUCTION: We evaluated the radiopacity and antibacterial properties of various concentrations of double antibiotic paste (DAP) containing barium sulfate (BaSO4) or zirconium oxide (ZrO2) radiopaque agents. METHODS: The radiopacity of 1, 10, and 25 mg/mL DAP containing 30% (w/v) BaSO4 or ZrO2, DAP-free radiopaque pastes, and commercially available radiopaque calcium hydroxide (Ca[OH]2) were evaluated according to ISO 6876/2001 with slight modifications (n = 6 per group). Dentin samples (n = 70) infected anaerobically for 3 weeks with bacterial biofilms obtained from a root canal of an immature tooth with pulpal necrosis were treated with similar experimental pastes or received no treatment (n = 7). After 1 week, the pastes were rinsed off, and biofilm disruption assays were conducted. To show the residual antibacterial effects, sterile dentin samples (n = 70) were pretreated for 1 week with the same pastes (n = 7). The pastes were rinsed off, and the samples were immersed in phosphate-buffered saline for 24 hours and infected anaerobically with the same bacterial biofilm mentioned earlier for 3 weeks before conducting biofilm disruption assays. Sterile dentin blocks were used in both antibacterial analyses as negative control groups (n = 7). Wilcoxon rank sum tests were used for statistical analyses. RESULTS: No tested concentrations of BaSO4 DAP or ZrO2 DAP showed significant differences from Ca(OH)2 in radiopacity. However, all tested concentrations of BaSO4 DAP, ZrO2 DAP, and Ca(OH)2 exhibited significant direct antibacterial effects. ZrO2 DAP at 1 mg/mL and Ca(OH)2 did not show significant residual antibacterial effects. CONCLUSIONS: BaSO4 DAP at 1 mg/mL provided significantly superior residual antibacterial effects and comparable radiopacity with the commercially available Ca(OH)2.

Correlation between crestal alveolar bone loss with intracanal bacteria and apical lesion area in necrotic teeth.

OBJECTIVE: This study aimed to analyze the correlation between crestal alveolar bone loss with the presence of some bacterial species in root canals and the apical lesion area of necrotic teeth. DESIGN: Data from 20 patients with diagnosis of pulp necrosis and acute apical abscesses, without active periodontal diseases, were evaluated. Patients with history of antibiotic usage three months prior to the study, with exposed pulp cavity, and with probing depth >3 mm were not included. The root size, the distance between the bone crest to the tooth apex in the mesial and distal surfaces, and the apical lesion area were measured from standard periapical radiographies by a calibrated examiner. Root canal samples were collected using sterilized paper points. In multirooted teeth, the largest root canal was sampled. Culture, microbial isolation and identification by phenotypic methods were performed. Spearman correlation and exact Fischer test were calculated between higher/lower existing bone crests, according to the median and the presence of specific bacteria. RESULTS: No statistically significant differences were found between occurrence of pathogenic bacteria, such as Porphyromonas gingivalis, Porphyromonas endodontalis, and Prevotella intermedia, and groups with higher/lower degree of bone loss (p > 0.05). A negative significant correlation was found between Parvimonas micra and periodontal bone loss (p = 0.02). Additionally, no statistically significant association was found between crestal bone loss and the apical lesion area. CONCLUSIONS: It was concluded that, in patients without active periodontitis, the presence of pathogenic bacteria in the root canal was not correlated with periodontal bone loss.

Detection of the unknown components of the oral microflora of teeth with periapical radiolucencies in a Turkish population using next-generation sequencing techniques.

AIM: To detect the unknown components of the oral microbiome and the effects of root canal treatment in a Turkish population and to evaluate the changes in microbial diversity in the root canals before and after treatment. METHODOLOGY: Single-rooted central and lateral maxillary incisors with one canal were chosen from 20 patients. Baseline samples of intact intracanal microbiota were collected from 20 root canals of permanent teeth with necrotic pulps using sterile paper points. After root canal preparation, the root canals were filled with a calcium hydroxide paste for 7 days. Calcium hydroxide was removed from root canal with 2.5% sodium hypochlorite and 17% EDTA using passive ultrasonic irrigation (PUI). A second bacteriologic samples were taken with sterile paper points prior to root filling. The samples were processes with DNase-I treatment using next-generation sequencing (NGS). Reduction in bacterial numbers during root canal treatment was evaluated using real-time quantitative PCR (qPCR). All statistical analyses were conducted using the MINITAB 17 software (Minitab Ltd. Co., Coventry, UK). A one-sample t-test was used to analyse the data. Statistical significance was accepted at P < 0.05. RESULTS: Relative abundances of Mycoplasma sp., Paludibacter sp., Tannerella sp., Prevotella spp. and an uncultured species from the order Bacteroidales decreased with root canal preparation and medication (98.7%, 99.8%, 98.8%, 97.7% and 99.3%, respectively), whilst the relative abundances of Methylobacterium sp., Corynebacterium sp. and Streptococcus infantis increased (93.1%, 94.8% and 99.4%, respectively). Considerable numbers of Streptophyta species were detected before and after treatment. The ratio of Agrobacterium sp. in the treated teeth community and the ratio of order Streptophyta in the infected canals had negative correlations with the success of bacterial elimination. CONCLUSIONS: The combination of NGS and qPCR techniques resulted in detection of previously unknown components of the oral microbiome and the effects of root canal treatment on their relative abundance in a Turkish population.

Prevotella strains and lactamic resistance gene distribution in different oral environments of children with pulp necrosis.

AIM: To determine the presence of Prevotella species, the cfxA/cfxA2, blaZ and blaTEM genes associated with resistance to lactamic agents in different oral niches of children with pulp necrosis. METHODOLOGY: Children with pulp necrosis in primary teeth had samples of saliva, supragingival, pulp chamber and root canal biofilms collected and tested for Prevotella species (P. intermedia, P. nigrescens, P. tannerae) and for beta-lactam resistance genes (cfxA/cfxA2, blaZ and blaTEM). The presence of bacterial DNA was examined through PCR, with a specific primer directed to the 16S rRNA gene. Specific primers were used to detect the Prevotella species and beta-lactam resistance genes. The chi-square test was used to analyse associations between the presence of bacteria and clinical variables. The Cochran's Q test was used to assess whether the proportion of gene detection is the same between different sites. RESULTS: Thirty-two teeth were sampled from 27 patients with a mean age of 5.5 years (±1.76). The total detection rate of Prevotella strains was 29.1%, 25%, 21.8% and 32.29% in saliva, supragingival, pulp chamber and root canal samples, respectively. P. nigrescens was the most commonly detected species in all oral niches. The previous use of antibiotics was associated with detection of P. nigrescens in saliva (P = 0.03). Pain was associated with the presence of P. nigrescens (P = 0.04) and P. tannerae (P = 0.01) in pulp chamber biofilm. blaTEM was detected in the four oral niches, being more frequent (23.8%) in supragingival biofilm (Cochran's Q test, P = 0.04). The presence of P. intermedia in SB and PC was associated with the detection of blaTEM in saliva (P = 0.04). The cfxA/cfxA2 and blaZ genes were not detected in any of the four oral niches. CONCLUSIONS: The oral cavity of children with pulp necrosis had a variable distribution of Prevotella strains in different niches. Saliva, supragingival biofilm, pulp chamber and root canals of primary teeth with necrotic pulps can harbour resistance genes to beta-lactams agents.

Presence of archaea and selected bacteria in infected root canal systems.

Infections of the root canal have polymicrobial etiology. The main group of microflora in the infected pulp is bacteria. There is limited data that archaea may be present in infected pulp tissue. The aim of this study was to check the prevalence of archaea in necrotic root canal samples obtained from patients with primary or post-treatment infection. The prevalence of selected bacteria species (Prevotella intermedia, Porphyromonas gingivalis, Tannerella forsythia, Treponema denticola, Synergistes sp.) in necrotic samples was evaluated as well. Sixty-four samples from root canal were collected for DNA and RNA extraction. A PCR assay based on the 16S rRNA gene was used to determine the presence of archaea and selected bacteria. Of the 64 samples, 6 were analyzed by semiquantitative reverse transcription PCR to estimate expression profiles of 16S rRNA, and another 9 were selected for direct sequencing. Archaea were detected in 48.4% samples. Statistical analysis indicated a negative association in coexistence between archaea and Treponema denticola (P < 0.05; Pearson's χ2 test). The main representative of the Archaea domain found in infected pulp tissue was Methanobrevibacter oralis. Archaea 16S rRNA gene expression was significantly lower than Synergistes sp., Porphyromonas gingivalis, and Tannerella forsythia (P < 0.05; Student's t test). Thus, it can be hypothesized that archaea may participate in the endodontic microbial community.

Efficacy of a single session protocol for endodontic treatment in primary teeth: in vivo study.

AIMS: To determine the efficacy of a single session protocol (SSP) in the reduction of septic content of primary teeth root canals and identify the persistence of bacterial species associated with unsuccessful treatment. METHODS: Primary teeth root canals (16) with pulp necrosis and peri-radicular lesions were treated. Samples were collected at baseline (T1), and after chemo-mechanical preparation, before filling (T2). Identification of the microorganisms was determined using checkerboard DNA-DNA hybridisation. STATISTICAL ANALYSIS: Wilcoxon test was applied for comparison of mean number of species, proportion and mean count of each species between the evaluation times. RESULTS: Significant reductions were found in the mean number of bacteria species between T1 and T2 (p < 0.05), but not for the reduction in proportion (p > 0.05). There was a reduction (6.0-4.6) of the mean number of species associated with failure, without statistical significance. CONCLUSION: The SSP was capable of significantly reducing the septic content, even though, many of the bacteria associated with failure persisted at the time of root canal filling.

Association between bacteria occurring in the apical canal system and expression of bone-resorbing mediators and matrix metalloproteinases in apical periodontitis.

AIM: To evaluate the association between the presence of selected bacterial species/groups in the apical root canal and expression of mediators of soft and bone tissue destruction in apical periodontitis lesions. Relationships between bacteria and some other features of apical periodontitis were also investigated. METHODOLOGY: Seventeen freshly extracted teeth with pulp necrosis and apical periodontitis were included. The apical root segment was sectioned and cryopulverized; DNA was extracted and evaluated for the presence of 9 bacterial species/groups using real-time polymerase chain reaction. Lesions were processed for histopathological and immunohistochemical analyses, which targeted matrix metalloproteinase-2 (MMP-2) and -9 (MMP-9), receptor activator of NFκB (RANK), RANK ligand (RANKL) and osteoprotegerin (OPG). Associations of the target bacteria with expression of these mediators, presence of symptoms, lesion size and histopathological diagnosis were evaluated. Data were analysed using the chi-square, Fisher's exact, Mann-Whitney and Pearson tests. P values lower than 0.05 were considered significant. RESULTS: All pulverized apical root samples were positive for bacteria. The most prevalent taxa were Actinobacteria (53%), Streptococcus species (35%), Fusobacterium species and Parvimonas micra (18%). The target mediators exhibited a high mean expression in the lesions (MMP-2: 82%; MMP-9: 73%; RANK: 78%; RANKL; 81%; OPG; 83%). Mean RANKL:OPG ratio was significantly higher in granulomas than cysts (P < 0.05, Mann-Whitney test). Actinobacteria were associated with granulomas, higher MMP-2 expression, lower OPG expression, and higher RANKL:OPG ratio (P < 0.05 for all, Fisher's exact test or Mann-Whitney test). No other significant associations were found. CONCLUSION: Actinobacteria may play an important role in the active phase of soft and bone tissue destruction in apical periodontitis.

Clinical comparison of the effectiveness of 7- and 14-day intracanal medications in root canal disinfection and inflammatory cytokines.

OBJECTIVE: This clinical study compared the effectiveness of 7- and 14-day intracanal medications in the reduction of bacteria/endotoxins from primarily infected root canals and determined their antigenicity against macrophages through the levels of cytokines. METHODS: Seventy-two primarily infected teeth were randomly divided into six groups according to medication and time of application: 7-day groups = G1, Ca(OH)2 + saline solution (SSL); G2, Ca(OH)2 + 2% chlorhexidine (CHX) gel; and G3, 2% CHX gel and 14-day groups = G4, Ca(OH)2 + SSL; G5, Ca(OH)2 + 2% CHX gel; and G6, 2% CHX gel (all groups, n = 12). Bacterial and endotoxin samples were collected from root canals and inflammatory cytokines of macrophages supernatants. Culture techniques were used to determine bacterial counts and limulus amebocyte lysate (LAL) assay to quantify endotoxins. IL-1ß, TNF-α, and PGE2 were measured by ELISA-assay. RESULTS: With regard to the bacterial reduction, no differences were found between all protocols tested (p > 0.05). The CHX protocols (G3 and G6) exhibited the lowest effectiveness against endotoxins (p < 0.05). All protocols were effective in lowering the levels of IL-1ß, TNF-α, and PGE2 (p < 0.05), with no difference between the medications tested on days 7 or 14 (p > 0.05). Particularly, the 7-day CHX-protocol (G3) exhibited the lowest effectiveness in lowering the levels of most cytokines compared to the 14-day protocols (G6) (p < 0.05). CONCLUSIONS: All the 7- and 14-day intracanal medications were effective in reducing bacteria and endotoxins as well as in lowering the levels of inflammatory cytokines, with CHX showing limited effectiveness against endotoxins. Moreover, 7-day CHX-protocol exhibited the lowest effectiveness in lowering the levels of most cytokines compared to the 14-day protocols. CLINICAL SIGNIFICANCE: Seven-day CHX protocol is the less effective protocol and should be carefully applied by the clinician.

Effect of ultrasonic activation on the reduction of bacteria and endotoxins in root canals: a randomized clinical trial.

AIM: This randomized clinical trial aimed to compare the effectiveness of ultrasonic activation with that of nonactivated irrigation on the removal of bacteria and endotoxin from root canals. METHODOLOGY: Fifty patients with necrotic pulps and asymptomatic apical periodontitis were randomly allocated into two groups according to the final irrigation protocol after root canal preparation: Group UI - ultrasonic irrigation (n = 25) and Group NI - needle irrigation (n = 25). The root canals were medicated with calcium hydroxide for 14 days. Microbiological sampling was performed before (S1) and after the root canal preparation (S2), after the irrigation protocols (S3) and after the removal of the intracanal medication (S4). Total bacteria counts were determined by qPCR and the endotoxin levels by the limulus amebocyte lysate assay. Intragroup analyses were performed using the Wilcoxon test for related samples, whereas intergroup analyses were performed using the Mann-Whitney U-test (P < 0.05). RESULTS: All S1 samples were positive for bacteria, with median numbers of 1.49 × 106 and 8.55 × 105 bacterial cells for the UI and NI groups, respectively. This number significantly decreased in S2 samples (UI: 1.41 × 104 ; NI: 3.53 × 104 ; both with P < 0.001). After final irrigation protocols, there was a significant decrease in bacterial load from S2 to S3 samples in both groups (UI: 4.29 × 103 ; NI: 1.08 × 104 ; P < 0.01). Intergroup analysis revealed a significant difference between irrigation methods regarding bacterial counts in S3 samples (P < 0.05). In contrast, no significant differences were observed between groups for endotoxin levels (P > 0.05). CONCLUSIONS: Ultrasonic activation was more effective than nonactivated irrigation for reducing the number of bacteria but not the endotoxin levels in root canals of teeth with apical periodontitis.

Identificación del complejo rojo en la necrosis pulpar abierta y cerrada con técnicas moleculares / Red complex identification in exposed and unexposed necrotic pulp with molecular techniques

Objetivo: identificar el complejo rojo periodontal, formado por Porphyromonas gingivalis, Treponema denticola y Tannerella forsythia, en la infección endodóntica primaria de necrosis pulpar, con cámara abierta y cerrada, utilizando técnicas de reacción en cadena de la polimerasa. Materiales y métodos: se realizó la toma para reacción en cadena de la polimerasa en 27 dientes con necrosis pulpar, 13 con cámara pulpar abierta y 14 con cámara cerrada. Resultados: en las muestras de necrosis abierta se identificaron P. gingivalis en un 92 por ciento, T. denticola en un 76 por ciento, T. forsythia en un 76 por ciento y el complejo rojo en un 61 por ciento. Las tomas de necrosis cerrada mostraron P. gingivalis en un 78 por ciento y T. denticola en un 57 por ciento; no se identificaron T. forsythia ni el complejo rojo. El análisis estadístico evidenció diferencias significativas entre los dos grupos (P<0,05). Conclusión: el sinergismo de las tres bacterias que forman el complejo rojo agravaría la patogénesis de la infección endodóntica y permitiría relacionar la microbiología endodóntica con la microbiología de periodontitis crónica.

16S Ribosomal RNA Gene Sequencing to Evaluate the Effects of 6 Commonly Prescribed Antibiotics.

INTRODUCTION: The rapid antibiotic sensitivity test (RAST) is a novel in-office culture and sensitivity system for endodontic infections. The purpose of this research was to validate the RAST system as a viable, in-office alternative to antibiotic sensitivity testing using turbidity to determine antibiotic sensitivities of endodontic infections. METHODS: Aspirates were taken from the root canals of 9 necrotic human teeth at the initiation of root canal therapy. These samples were cultured in the RAST medium, and antibiotic sensitivity to 6 antibiotics was tested. Further analysis was performed using 16S ribosomal RNA (rRNA) gene sequencing. RESULTS: Thirty-one bacterial phyla were identified as well as 2 phyla of the kingdom Archaea. Augmentin (Dr. Reddy's Laboratories Ltd, Hyderabad, India) and ampicillin performed identically at 24 hours, inhibiting turbidity in 100% of the samples. At 48 hours in anaerobic conditions, Augmentin outperformed ampicillin by 13%. Ciprofloxacin was the least efficacious antibiotic. At 48 hours, only 22% of anaerobic ciprofloxacin cultures affectively inhibited bacterial growth. CONCLUSIONS: The RAST medium is a viable in-office alternative to antibiotic susceptibility testing in an off-site laboratory. It is able to support the growth of a wide variety of microorganisms in both aerobic and anaerobic environments, and, in combination with 16S rRNA gene sequencing, it led to the identification of a new archaebacterial phylum, Crenarchaeota, as part of the endodontic infection microbiome.

Identificación de microorganismos por reacción en cadena de la polimerasa en necrosis pulpar y periodontitis apical / Identification of microorganisms by polymerase chain reaction in pulp necrosis and apical periodontitis

Objetivos: identificar, mediante la técnica de reacción en cadena de la polimerasa, diez especies bacterianas procedentes de conductos radiculares necróticos, así como analizar su asociación con signos y síntomas de la periodontitis apical (dolor, exudado y movilidad), dentro de un área geográfica específica, Santiago de Compostela (noroeste de España). Métodos: se realizó un estudio descriptivo, en un plazo de recogida de muestras de un año. Se extrajeron 43 muestras de los conductos radiculares necróticos, a razón de una muestra por pacientes, quienes fueron examinados previamente para determinar la presencia de periodontitis apical radiográfica, de la cual fueron descritos sus signos y síntomas. Se utilizaron puntas de papel absorbentes estériles y se procedió a extraer de dichas muestras el ADN por medio de la técnica del fenol-cloroformo. El producto se amplificó por medio de la reacción en cadena de la polimerasa, usando cebadores específicos para diez microorganismos. El resultado se visualizó por medio de electroforesis, utilizando un transiluminador UV. Los resultados se analizaron estadísticamente, para así establecer qué microorganismos estaban presentes de forma individual y cuáles se presentaron formando asociaciones como el llamado red complex, en relación con los signos y síntomas presentes en la periodontitis apical. Resultados: Fusobacterium nucleatum fue el microorganismo con presencia de forma individual más frecuente (83,72 por ciento de las muestras). En cuanto al estudio de la asociación de los microorganismos con los signos y síntomas, Porphyromonas endodontalis presentó una asociación significativa con respecto al exudado (p< 0,05) y movilidad (p< 0,05). En cambio, Enterococcus spp. y Treponema denticola (p< 0,01) presentaron ambos una asociación significativa con respecto al dolor. En cuanto a la incidencia del red complex fue de 6 casos en total. Conclusiones: Fusobacterium nucleatum y Streptococcus spp. son los microorganismos identificados con mayor porcentaje en comparación a los restantes estudiados. Porphyromonas endodontalis es el microorganismo con mayor asociación estadística con respecto a los signos y síntomas de la periodontitis apical(AU)

Objectives: identify by polymerase chain reaction technique ten bacterial species obtained from necrotic root canals, and analyze their association with signs and symptoms of apical periodontitis (pain, exudate and mobility) in a specific geographic area: Santiago de Compostela (northwestern Spain). Methods: a descriptive study was conducted based on a one-year sample collection period. Forty-three samples were taken from necrotic root canals, at a rate of one sample per patient, who had been previously examined for radiographic apical periodontitis, of which the signs and symptoms were described. Sterile absorbent paper points were used to extract the DNA samples, applying the phenol-chloroform technique. The product was amplified by polymerase chain reaction, using specific primers for ten microorganisms. The result was visualized by electrophoresis using a UV transilluminator. Outcomes were analyzed statistically to determine which microorganisms were present individually and which formed associations such as the so-called red complex, according to the signs and symptoms present in the apical periodontitis. Results: Fusobacterium nucleatum was the most common individual microorganism (83.72 percent of the samples). As to the association of microorganisms with signs and symptoms, Porphyromonas endodontalis was found to be significantly associated with exudate (p< 0.05) and mobility (p< 0.05), while both Enterococcus spp. and Treponema denticola (p< 0.01) had a significant association with pain. Incidence of the red complex was 6 cases in all. Conclusions: Fusobacterium nucleatum and Streptococcus spp. were the microorganisms identified as displaying the highest percentages. Porphyromonas endodontalis showed the greatest statistical association with signs and symptoms of apical periodontitis(AU)

Identificación de microorganismos por reacción en cadena de la polimerasa en necrosis pulpar y periodontitis apical / Identification of microorganisms by polymerase chain reaction in pulp necrosis and apical periodontitis

Objetivos: identificar, mediante la técnica de reacción en cadena de la polimerasa, diez especies bacterianas procedentes de conductos radiculares necróticos, así como analizar su asociación con signos y síntomas de la periodontitis apical (dolor, exudado y movilidad), dentro de un área geográfica específica, Santiago de Compostela (noroeste de España). Métodos: se realizó un estudio descriptivo, en un plazo de recogida de muestras de un año. Se extrajeron 43 muestras de los conductos radiculares necróticos, a razón de una muestra por pacientes, quienes fueron examinados previamente para determinar la presencia de periodontitis apical radiográfica, de la cual fueron descritos sus signos y síntomas. Se utilizaron puntas de papel absorbentes estériles y se procedió a extraer de dichas muestras el ADN por medio de la técnica del fenol-cloroformo. El producto se amplificó por medio de la reacción en cadena de la polimerasa, usando cebadores específicos para diez microorganismos. El resultado se visualizó por medio de electroforesis, utilizando un transiluminador UV. Los resultados se analizaron estadísticamente, para así establecer qué microorganismos estaban presentes de forma individual y cuáles se presentaron formando asociaciones como el llamado red complex, en relación con los signos y síntomas presentes en la periodontitis apical. Resultados: Fusobacterium nucleatum fue el microorganismo con presencia de forma individual más frecuente (83,72 por ciento de las muestras). En cuanto al estudio de la asociación de los microorganismos con los signos y síntomas, Porphyromonas endodontalis presentó una asociación significativa con respecto al exudado (p< 0,05) y movilidad (p< 0,05). En cambio, Enterococcus spp. y Treponema denticola (p< 0,01) presentaron ambos una asociación significativa con respecto al dolor. En cuanto a la incidencia del red complex fue de 6 casos en total. Conclusiones: Fusobacterium nucleatum y Streptococcus spp. son los microorganismos identificados con mayor porcentaje en comparación a los restantes estudiados. Porphyromonas endodontalis es el microorganismo con mayor asociación estadística con respecto a los signos y síntomas de la periodontitis apical(AU)

Objectives: identify by polymerase chain reaction technique ten bacterial species obtained from necrotic root canals, and analyze their association with signs and symptoms of apical periodontitis (pain, exudate and mobility) in a specific geographic area: Santiago de Compostela (northwestern Spain). Methods: a descriptive study was conducted based on a one-year sample collection period. Forty-three samples were taken from necrotic root canals, at a rate of one sample per patient, who had been previously examined for radiographic apical periodontitis, of which the signs and symptoms were described. Sterile absorbent paper points were used to extract the DNA samples, applying the phenol-chloroform technique. The product was amplified by polymerase chain reaction, using specific primers for ten microorganisms. The result was visualized by electrophoresis using a UV transilluminator. Outcomes were analyzed statistically to determine which microorganisms were present individually and which formed associations such as the so-called red complex, according to the signs and symptoms present in the apical periodontitis. Results: Fusobacterium nucleatum was the most common individual microorganism (83.72 percent of the samples). As to the association of microorganisms with signs and symptoms, Porphyromonas endodontalis was found to be significantly associated with exudate (p< 0.05) and mobility (p< 0.05), while both Enterococcus spp. and Treponema denticola (p< 0.01) had a significant association with pain. Incidence of the red complex was 6 cases in all. Conclusions: Fusobacterium nucleatum and Streptococcus spp. were the microorganisms identified as displaying the highest percentages. Porphyromonas endodontalis showed the greatest statistical association with signs and symptoms of apical periodontitis(AU)

Effectiveness in the Removal of Endotoxins and Microbiological Profile in Primary Endodontic Infections Using 3 Different Instrumentation Systems: A Randomized Clinical Study.

INTRODUCTION: This clinical study was conducted to correlate the microbiological profile and levels of endotoxins found in primary endodontic infection with the presence of clinical features and to evaluate the removal of microorganisms and endotoxins using rotary, reciprocating, and hybrid systems for biomechanical preparation. METHODS: Thirty single root canals with primary endodontic infection were evaluated with signs and symptoms and were randomly divided into 3 groups according to the instrumentation system used (n = 10) as follows: rotary Mtwo instruments (VDW, Munich, Germany) with 8 files, the reciprocating Reciproc system (VDW) with a single file, and Genius hybrid instruments with 3 files (1 rotary and 2 reciprocating files) with irrigation using 24 mL 2.5% sodium hypochlorite. Samples were collected before (S1) and after instrumentation (S2) before being submitted to microbiological culture (colony-forming units/mL) and the checkerboard DNA-DNA hybridization test. Endotoxins were quantified using the limulus amebocyte lysate assay. RESULTS: Microbiological culture showed statistical differences in the reduction of colony-forming units/mL with all systems tested (P < .05), but no statistical difference was found among the groups. The most frequently detected species were Capnocytophaga ochracea (53%) and Fusobacterium nucleatum (53%) at S1 and F. nucleatum (50%) and Leptotrichia buccalis (50%) at S2. As for the reduction of endotoxins at S2, Mtwo presented the best results (95.05%) followed by the Genius (91.85%) and Reciproc (64.68%) groups, but no statistical difference was found among the groups. Previous pain, tenderness to percussion, and presence of a sinus tract were associated with specific microorganisms (P < .05). CONCLUSIONS: Signs and symptoms were correlated with microorganisms. Endodontic treatment was effective in reducing bacteria and endotoxins but was not capable of completely removing them from the root canal.

Relaciones endoperiodontales: una frontera transitable / Endoperiodontal relationships: a passable border

La existencia de una íntima conexión entre el conducto radicular y el ligamento periodontal, ha dado lugar a lo que se conoce como relaciones endoperiodontales. Esto se debe a la presencia de varias vías anatómicas de comunicación entre ambas entidades: el foramen apical, los forámenes laterales pertenecientes a conductos accesorios y conductillos dentinarios en zonas de ausencia del cemento dentario protector. Los microorganismos y sus agentes tóxicos tienen la capacidad de afectar ambos tejidos en esa interrelación física y biológica. El diagnóstico diferencial entre enfermedad endodóntica y periodontal es de vital importancia para la elección del tratamiento y el pronóstico a distancia.

The intimate connection between the root canal and theperiodontal ligament generates a concept that is known asendo-periodontal relationships. This is due to the presence ofseveral anatomical communications between them: apical foramen,lateral foramina and dentinal tubules denuded of theircementum coverage. Microorganisms and their toxic agentscan affect both tissues due to this physical and biological interrelation.Proper differential between endodontic and periodontaletiology is vital to the accurate choice of treatment andfor the long term prognosis.