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  • Quantitative real-time PCR using TaqMan and SYBR green for Actinobacillus actinomycetemcomitans, Porphyromonas gingivalis, Prevotella intermedia, tetQ gene and total bacteria
    Maeda, Hiroshi ...
    Accurate quantification of bacterial species in dental plaque is needed for microbiological diagnosis of periodontal diseases. The present study was designed to assess the sensitivity, specificity ... and quantitativity of the real-time PCR using the GeneAmp Sequence Detection System with two fluorescence chemistries. TaqMan probe with reporter and quencher dye, and SYBR Green dye were used for sources of the fluorescence. Primers and probes were designed for Actinobucillus actinomycetemcomitans, Porphyromanas gingiva/is, Prevotella intermedia and total bacteria based on the nucleotide sequences of the respective 16S ribosomal RNA genes. Since spread of antibiotic resistance genes is one of the crucial problems in periodontal therapy, quantitative detection of tetQ gene, which confers resistance to tetracycline, was included in the examination. The detection of P. gingivalis,P. intermedia and A. actinomycetemcomitans was linear over a range of 10-10 7 cells (10-10 7 copies for tetQ gene), while the quantitative range for total bacteria was 10 2-10 7 cells. Species-specific amplifications were observed for the three periodontal bacteria, and there was no significant difference between the TaqMan and SYBR Green chemistry in their specificity, quantitativity and sensitivity. The SYBR Green assay, which was simpler than TaqMan assay in its manipulations, was applied to the clinical plaque samples.The plaque samples were obtained from eight patients (eight periodontal pockets) before and 1 week after the local drug delivery of minocycline. Although the number of P. gingivalis, P. intermedia and A. actinomycetemcomitans markedly decreased after the antibiotic therapy in most cases, higher copy numbers of the tetQ gene were detectable. (Abstract truncated at 2000 characters).
    Source: FEMS Immunology and Medical Microbiology. - ISSN 0928-8244 (Letn. 39, 2003, str. 81-86)
    Type of material - article, component part
    Publish date - 2003
    Language - english
    COBISS.SI-ID - 19190489

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