Culture and PCR analysis of bacteria in infected root canals

Traditional techniques have found associations of black-pigmented bacteria (BPB) with primary- and Enterococcus faecalis with secondary- endodontic infections. Objective: To investigate the presence of 4 BPB (Prevotella intermedia, P.i., P. nigrescens, P.n.; Porphyromonas gingivalis, P.g.; P. endodontalis, P.e.) and E. faecalis (E.f.) in primary and secondary root-infected canals with periapical lesions by both culture and molecular analysis. Methods: Microbial samples were taken from 100 canals, 50 with necrotic pulp tissues (NPT, primary infection) and 50 after failed endodontic treatment (FET, secondary infection). Strict anaerobic techniques were used for serial dilution, plating, incubation and identification. For molecular detection, the samples were analysed for the presence of 5 endodontic pathogens using species-specific primers and PCR. Results: Using culture, P.i./P.n. were isolated from 11 canals with NPT, and from 2 canals with FET; P.g. from 1 canal with NPT, P.e. from none of the canals investigated. E.f. were isolated from 2 canals with NPT and from 21 canals with FET. Using PCR, P.i. were isolated from 28 canals with NPT and 5 canals with FET; P.n. from 16 canals with NPT and 6 canals with FET; P.g. from 22 canals with NPT and 16 canals with TEP; P.e. from 14 canals with NPT and 11 canals with TEP; E.f. from 41 canals with NPT and 38 canals with FET. Using culture, significant associations were found between primary infection and the presence of P.i. (p<0.05) and between secondary infection and the presence of E.f. (p<0.0001). Using PCR, associations were found between primary infection and the presence of P.i. and P.n. (both p<0.05). Conclusions: These findings indicate that the microbial profiles obtained by cultivation differ from those obtained by molecular analysis, which is more efficient in the detection of these species than culture (FAPESP 00/13689-7, CNPq -520277/99-6 & CAPES – BEX 2528/02-9, NIH grant DE 10467)