Identification of Bacteria Enduring Endodontic Treatment Procedures by a Combined Reverse Transcriptase-Polymerase Chain Reaction and Reverse-Capture Checkerboard Approach

Introduction: This study identified the bacterial taxa enduring endodontic treatment procedures by using a combined 165 ribosomal RNA-based reverse-transcriptase polymerase chain reaction (RT-PCR) and reverse-capture checkerboard hybridization approach. Methods: Samples were taken from infected canals of 15 teeth with apical periodontitis before treatment (S1), after chemo-mechanical preparation with NaOCl as the irrigant (S2), and after interappointment medication with a calcium hydroxide paste (S3). Bacterial presence was first screened by a DNA-based single PCR assay. RNA extracts were subjected to RT-PCR, and the resulting products were surveyed for the presence of 28 targeted taxa by using the checkerboard method. Results: Bacteria were found in all Si samples. Detectable levels of bacterial ribosomal RNA, used as an indicator of viability, were observed in 60% of the cases after chemomechanical preparation and 53% after intracanal medication. The most prevalent taxa in Si were Olsenella uli (67%), Pyramidobacter piscolens (60%), Streptococcus species (53%), and Bacteroidetes clone X083 (53%). Streptococcus species (47%), Fusobacterium nucleatum (40%), and O. uli (33%) prevailed in S2, whereas Streptococcus species (47%), Pro propioni-bacterium acnes (27%), and O. uli (27%) were the most frequent taxa in S3. Conclusions: The present study with a combined molecular approach revealed that bacterial diversity was overall markedly reduced by treatment procedures. Although bacterial taxa more frequently identified in post-treatment samples emerge as potential risk factors for persistent disease, this remains to be determined by longitudinal studies. (J Endod 2010;36:45-52)