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Direct quantitative differentiation between Prevotella intermedia and Prevotella nigrescens in clinical specimens.


Gmür, R; Thurnheer, T (2002). Direct quantitative differentiation between Prevotella intermedia and Prevotella nigrescens in clinical specimens. Microbiology, 148(5):1379-1387.

Abstract

This paper describes a quantitative fluorescent in situ hybridization (FISH) assay for the differential identification of Prevotella intermedia and Prevotella nigrescens 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 P. intermedia, P. nigrescens, Prevotella pallens and Prevotella denticola 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 Prevotella species. The lower detection level of the assays was approximately 3x10(3) target cells per ml of plaque-sample suspension. P. intermedia, P. nigrescens, P. pallens and P. denticola were detected in plaques with prevalences of 69, 67, 0 and 28%, respectively. On average, 3.9 x 10(6) P. intermedia, 3.1 x 10(6) P. nigrescens and 5.6 x 10(5) P. denticola 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 (r=0.711). Both FISH and IF enumerations of the sum of P. intermedia and P. nigrescens markedly exceeded the c.f.u. counts of black-pigmented colonies in Porphyromonas gingivalis-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 P. intermedia and P. nigrescens, provides visual accuracy control, and offers insights into the spatial distribution of the target cells within a clinical sample.

This paper describes a quantitative fluorescent in situ hybridization (FISH) assay for the differential identification of Prevotella intermedia and Prevotella nigrescens 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 P. intermedia, P. nigrescens, Prevotella pallens and Prevotella denticola 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 Prevotella species. The lower detection level of the assays was approximately 3x10(3) target cells per ml of plaque-sample suspension. P. intermedia, P. nigrescens, P. pallens and P. denticola were detected in plaques with prevalences of 69, 67, 0 and 28%, respectively. On average, 3.9 x 10(6) P. intermedia, 3.1 x 10(6) P. nigrescens and 5.6 x 10(5) P. denticola 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 (r=0.711). Both FISH and IF enumerations of the sum of P. intermedia and P. nigrescens markedly exceeded the c.f.u. counts of black-pigmented colonies in Porphyromonas gingivalis-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 P. intermedia and P. nigrescens, provides visual accuracy control, and offers insights into the spatial distribution of the target cells within a clinical sample.

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23 citations in Web of Science®
27 citations in Scopus®
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Additional indexing

Item Type:Journal Article, refereed
Communities & Collections:04 Faculty of Medicine > Center for Dental Medicine > Institute of Oral Biology
Dewey Decimal Classification:610 Medicine & health
Language:English
Date:1 May 2002
Deposited On:11 Feb 2008 12:24
Last Modified:05 Apr 2016 12:19
Publisher:Society for General Microbiology
ISSN:1350-0872
Related URLs:http://mic.sgmjournals.org/cgi/content/full/148/5/1379?view=long&pmid=11988511
PubMed ID:11988511
Permanent URL: http://doi.org/10.5167/uzh-1684

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