A sensitive method for detecting Porphyromonas gingivalis by polymerase chain reaction and its possible clinical application

Background: It is useful for the clinical diagnosis of periodontitis to mon itor the colonization of periodontopathic bacteria in periodontal pockets. In this study, we attempted to establish and possibly identify the clinical application of a sensitive method to detect Porphyromonas gingivalis (Pg.) , one of the putative periodontopathic bacteria related to chronic periodon titis. Methods: Genomic DNA extracted from cultured Pg. 381 and clinically isolate d subgingival plaque samples were used as a template of polymerase chain re action (PCR). We designed primers to amplify the genomic DNA coding 40 kDa outer membrane protein (OMP), one of the unique proteins to all strains of Pg. The efficiency and specificity of amplification were evaluated by agaro se gel electrophoresis and subsequent Southern hybridization with a digoxyg enin-labeled oligonucleotide probe. Results: Fewer than 100 Pg. bacterial cells in the specimen were reproducib ly detected by PCR-hybridization assay. This PCR-hybridization assay was at least 100 times more sensitive than the conventional indirect immunofluore scence assay (IIF). Furthermore, the imaging analysis showed that there is a linear correlation between the strength of the signal and the cell number of Pg. from which the template DNA was extracted semiquantitatively. It is noteworthy that the PCR assay could also be applied to detect Pg. from cli nical plaque samples and that it was approximately 100 times more sensitive than a conventional IIF assay. Conclusion: The PCR assay established in this study can be a powerful tool to detect Pg. in periodontal pockets and monitor the colonization and/or re colonization of P.g. at the very early phase.