1887

Abstract

This paper describes a quantitative fluorescent hybridization (FISH) assay for the differential identification of and in clinical samples, and compares its performance with less discriminatory culture and quantitative immunofluorescence (IF) assays. Fluorescence-labelled oligonucleotide probes directed to specific 16S rRNA sequences of , , and were hybridized under stringent conditions with cultured reference strains or plaque samples from deep periodontal pockets. Probe specificity was defined with strains from multiple oral species. The lower detection level of the assays was approximately 3×10 target cells per ml of plaque-sample suspension. , , and were detected in plaques with prevalences of 69, 67, 0 and 28%, respectively. On average, 39×10 , 31×10 and 56×10 cells were counted per positive sample. All three species were found almost exclusively in dense mixed aggregates. Quantitative FISH data agreed satisfactorily with corresponding IF data (=0711). Both FISH and IF enumerations of the sum of and markedly exceeded the c.f.u. counts of black-pigmented colonies in -free cultured subgingival plaques. The results demonstrate the validity of this new assay. Unlike established IF, culture, PCR or checkerboard DNA hybridization assays, this FISH assay differentiates quantitatively between and , provides visual accuracy control, and offers insights into the spatial distribution of the target cells within a clinical sample.

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2002-05-01
2019-10-17
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