Detection of oral bacteria on the tongue dorsum using PCR amplification of 16S ribosomal RNA and its association with systemic disease in middle-aged and elderly patients

Abstract

The association between oral health and systemic disease is recognized in the literature. The present study aimed to clarify the association between oral bacteria on the tongue dorsum and factors associated with oral health and systemic disease in middle-aged and elderly patients. The association between bacterial numbers, oral health status and systemic disease was preliminarily investigated in 70 patients (mean age, 69.5 years; range, 45-92 years) who visited the Department of Oral Health, Hiroshima University Hospital (Hiroshima, Japan). The bacterial 16S ribosomal RNA gene was employed to quantitate bacterial numbers using real-time polymerase chain reaction (PCR). PCR was also performed to detect the DNA of periodontal disease-related bacteria. Oral bacterial numbers were marginally negatively correlated with moisture levels on the tongue surface [Spearman's rank correlation coefficient (R)=-0.131, P=0.28). Subjects with bleeding on probing (BOP) or a ≥4 mm probing depth (PD) exhibited higher Porphyromonas gingivalis (P. gingivalis)-positive rates (50.0 and 51.1%, respectively) than those without BOP or a <4 mm PD (39.5 and 30.4%, respectively). Subjects with medical histories of hypertension, diabetes, stroke and heart disease exhibited a trend toward higher P. gingivalis-positive rates than those without such disorders. These findings indicated that the tongue moisture level may be associated with bacterial numbers on the tongue surface, while P. gingivalis on the tongue surface may be associated with systemic and periodontal diseases. Further investigation in a larger number of participants is necessary to clarify the correlation between bacterial numbers and systemic disease.

Keywords: 16S ribosomal RNA gene; bacterial number; periodontal disease-related bacteria; real-time polymerase chain reaction; tongue dorsum.

Figure 1.

Standard curve indicating Ct values vs. Staphylococcus aureus numbers (CFU/ml). Serial 10-fold dilutions of Staphylococcus aureus DNA (ranging from 10¹ to 10⁸ CFU/ml) were used to make a standard curve. CFU, colony forming units.

Figure 2.

Correlation between age and bacterial numbers on the tongue surface. Age was weakly positively correlated with bacterial numbers on the tongue surface.

Figure 3.

Correlation between moisture levels and bacterial numbers on the tongue surface. A weak negative relationship was indicated between oral bacterial numbers and moisture levels.

Figure 4.

PCR detection of periodontal disease-related bacteria on the tongue surface. PCR was performed to detect T. denticola, T. forsythia and P. gingivalis DNA. 10 µl of each 20-µl PCR product was separated on a 2% agarose gel. P. gingivalis, Porphyromonas gingivalis; T. forsythia, Tannerella forsythia; T. denticola, Treponema denticola; PCR, polymerase chain reaction.