RNA-based Assay Demonstrated Enterococcus faecalis Metabolic Activity after Chemomechanical Procedures
| Autor: | Ericka Tavares Pinheiro, S. R. L. Teixeira, Giulio Gavini, George Táccio de Miranda Candeiro, L. C. Prado, Regina Célia Furukava Shin, Marcia Pinto Alves Mayer |
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| Rok vydání: | 2014 |
| Předmět: |
DNA
Bacterial RNA Ribosomal RNA Biology 16S ribosomal RNA biology.organism_classification Real-Time Polymerase Chain Reaction Molecular biology Enterococcus faecalis Microbiology genomic DNA Real-time polymerase chain reaction Complementary DNA RNA Ribosomal 16S Humans Dental Pulp Cavity General Dentistry Ribosomal DNA |
| Zdroj: | Journal of endodontics. 41(9) |
| ISSN: | 1878-3554 |
| Popis: | Because ribosomal RNA (rRNA) indicates metabolic cell activity, this study aimed to evaluate the sensitivity of rRNA-based quantitative polymerase chain reaction (RT-qPCR) for the identification of active Enterococcus faecalis in root canals samples compared with a method based on ribosomal DNA (rDNA) (rRNA genes).Samples were taken from 18 teeth with persistent/secondary intraradicular infection before (S1) and after (S2) chemomechanical preparation. RNA and DNA were extracted, and complementary DNA was synthesized from RNA using RT-PCR. Complementary DNA and genomic DNA were subjected to quantitative polymerase chain reaction with primers complementary for E. faecalis 16S rRNA sequence.E. faecalis was detected in 77.8% and 72.2% of S1 samples using rRNA- and rDNA-based assays, respectively. In contrast, E. faecalis was detected in only 33.3% of S2 samples using rDNA as the template compared with 61.1% using the rRNA-based method. The median concentration of rRNA copies of E. faecalis was significantly higher than rDNA copies, indicating a higher sensitivity for the method targeting rRNA in both S1 (P .01) and S2 samples (P .05). After chemomechanical preparation, the number of rRNA and rDNA copies was significantly reduced (P .05). The high ratio of rRNA to rDNA copies in S2 samples suggested that active E. faecalis persisted in root canals after chemomechanical preparation.The RT-qPCR assay provides a sensitive method for the identification of active E. faecalis from endodontic samples. Furthermore, the rRNA-based assay indicated that E. faecalis viable cells persisted in treated root canals, suggesting that it may be a useful tool for monitoring microbial load during endodontic treatment. |
| Databáze: | OpenAIRE |
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