Taqman Real Time PCR Assay for Desulfomicrobium Orale in Chronic Periodontal Lesions

Introduction: Sulfate-reducing bacteria as well as Desulfomicrobium orale (D. orale) have both been shown to play a potential etiopathogenetic role for periodontal disease.

Methods: From 15 treated periodontitis patients 45 subgingival biofilm isolates (SBIs) were obtained from residual periodontal pockets ˃5mm. Real-time PCR was performed using species-specific primers and a TaqMan probe. Absolute quantification was achieved by an external recombinant DNA-standard. Sign alignment of the 16S rRNA gene was performed using commercial algorithm. Furthermore, the correlation between probing depth (PD) or clinical attachment level (CAL) and the quantity of the target gene of D. orale were analyzed.

Results: The prevalence of D. orale within the SBIs was 100%. The mean target gene amount was 7.2E+04. Sign alignment of the sequence of the 16S rRNA gene showed high similarities to further Desulfomicrobium strains. The correlation of CAL and PD with the target gene count of D. orale was not significant (p> 0.05).

Conclusion: For the first time the absolute amount of subgingival D. orale was measured using real-time PCR technology in chronic periodontal lesions. This assay provides a culture independent risk monitoring of patients suffering from periodontitis by qualitative and quantitative analysis of D. orale.

Keywords: Sulfate-reducing bacteria; Real-time PCR; Absolute quantification; Periodontal disease, Desulfomicrobium orale