Clinical ResearchApical Root Canal Microbiota as Determined by Reverse-capture Checkerboard Analysis of Cryogenically Ground Root Samples from Teeth with Apical Periodontitis
Section snippets
Specimen Collection and Processing
DNA extracts from samples collected for a previous investigation (5) were stored and available for reanalysis in this study. The examined material consisted of 17 extracted teeth randomly collected from 14 patients in the Oral Surgery Clinic, Estácio de Sá University, Rio de Janeiro. All teeth had necrotic pulps, radiographic evidence of apical periodontitis and extensive caries lesions, and were extracted for prosthetic reasons or by request of the patient. The study protocol was approved by
Results
Sterility control samples taken from the external root surface after disinfection yielded no PCR product. Negative PCR controls showed the predicted results. All apical and middle/coronal samples were positive for the presence of bacteria as determined by PCR with universal 16S rRNA gene primers.
All samples were also positive for bacteria in the reverse-capture checkerboard assay as revealed by the 2 universal probes. The results revealed that 27 and 20 taxon-specific probes tested reactive
Discussion
Only 2 studies have been published that use molecular methods to identify bacteria located in the apical root canal. One study (19) used a conventional species-specific PCR assay directed to 10 bacterial species, whereas the other (18) used the same reverse-capture checkerboard approach as used in the present study to detect 28 target species/phylotypes. The main difference between those studies and the present one was that a cryogenic grinding approach was used to pulverize root samples so the
Acknowledgments
This study was supported by grants from Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq), and Fundação Carlos Chagas Filho de Amparo à Pesquisa do Estado do Rio de Janeiro (FAPERJ), Brazilian Governmental Institutions. The authors are grateful to Marlei Gomes da Silva for technical assistance and Dr Rodrigo Soares de Moura Neto from the Institute for Research and Expertise in Forensic Genetics of the State Police Academy Sylvio Terra, Rio de Janeiro, for assistance with the
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