Research Article
1 February 1990

Fluorescent-oligonucleotide probing of whole cells for determinative, phylogenetic, and environmental studies in microbiology

Abstract

Fluorescent-dye-conjugated oligonucleotides were used to classify 14 Fibrobacter strains by fluorescence microscopy. On the basis of partial 16S rRNA sequences of six Fibrobacter strains, four hybridization probes were designed to discriminate between the species Fibrobacter succinogenes and Fibrobacter intestinalis and to identify F. succinogenes subsp. succinogenes. After in situ hybridization to whole cells of the six sequenced strains, epifluorescence microscopy confirmed probe specificity. The four probes were then used to make presumptive species and subspecies assignments of eight additional Fibrobacter strains not previously characterized by comparative sequencing. These assignments were confirmed by comparative sequencing of the 16S rRNA target regions from the additional organisms. Single-mismatch discrimination between certain probe and nontarget sequences was demonstrated, and fluorescent intensity was shown to be enhanced by hybridization to multiple probes of the same specificity. The direct detection of F. intestinalis in mouse cecum samples demonstrated the application of this technique to the characterization of complex natural samples.

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Published In

cover image Journal of Bacteriology
Journal of Bacteriology
Volume 172Number 2February 1990
Pages: 762 - 770
PubMed: 1688842

History

Published online: 1 February 1990

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Authors

R I Amann
Department of Veterinary Pathobiology, University of Illinois, Urbana 61801.
L Krumholz
Department of Veterinary Pathobiology, University of Illinois, Urbana 61801.
D A Stahl
Department of Veterinary Pathobiology, University of Illinois, Urbana 61801.

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