Use of quantitative PCR to evaluate methods of bacteria sampling in periodontal patients

Periodontal disease is associated with specific periodontal pathogens and may persist as gingivitis or progress to more severe disease. The bacteria involved in disease initiation and progression have not been identified. We used quantitative polymerase chain reaction (PCR) to compare the levels of Porphyromonas gingivalis, Tannerella forsythia, and Treponema denticola, and total bacteria detected by different sampling methods. On the basis of the results of clinical examinations, 57 patients were divided into 3 groups: healthy group (group A), gingivitis group (group B), and periodontitis group (group C). Bacterial samples were collected from saliva, mouthwash, and by paper-point sampling of gingival crevicular fluid (GCF), and the samples were analyzed with quantitative PCR targeting 16S rRNA. The numbers of total bacteria in samples of GCF, saliva, and mouthwash were 10⁵ to 10⁶, 10⁸, and 10⁷, respectively, per milliliter. The number of P. gingivalis in GCF samples was lower than 10 in group A; however, in groups B and C, the values were 10³ and 10⁴, respectively, indicating that the number of P. gingivalis increased with worsening clinical status. Findings were similar in the samples of saliva and mouthwash. The numbers of T. forsythia showed a pattern similar to that of P. gingivalis in all 3 samples. These results suggest that saliva and mouthwash samples are clinically useful for bacterial testing of periodontal diseases by quantitative PCR. In addition, mouthwash sampling is more feasible and straightforward than saliva sampling.