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Science 10 March 1989:
Vol. 243. no. 4896, pp. 1360 - 1363
DOI: 10.1126/science.2466341
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Articles

Science, Vol 243, Issue 4896, 1360-1363
Copyright © 1989 by American Association for the Advancement of Science

articles

Phylogenetic stains: ribosomal RNA-based probes for the identification of single cells

EF DeLong, GS Wickham, and NR Pace

Department of Biology, Indiana University, Bloomington 47405.

Rapid phylogenetic identification of single microbial cells was achieved with a new staining method. Formaldehyde-fixed, intact cells were hybridized with fluorescently labeled oligodeoxynucleotides complementary to 16S ribosomal RNA (rRNA) and viewed by fluorescence microscopy. Because of the abundance of rRNA in cells, the binding of the fluorescent probes to individual cells is readily visualized. Phylogenetic identification is achieved by the use of oligonucleotides (length 17 to 34 nucleotides) that are complementary to phylogenetic group-specific 16S rRNA sequences. Appropriate probes can be composed of oligonucleotide sequences that distinguish between the primary kingdoms (eukaryotes, eubacteria, archaebacteria) and between closely related organisms. The simultaneous use of multiple probes, labeled with different fluorescent dyes, allows the identification of different cell types in the same microscopic field. Quantitative microfluorimetry shows that the amount of an rRNA-specific probe that binds to Escherichia coli varies with the ribosome content and therefore reflects growth rate.


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Combined Microautoradiography-16S rRNA Probe Technique for Determination of Radioisotope Uptake by Specific Microbial Cell Types In Situ.
C. C. Ouverney and J. A. Fuhrman (1999)
Appl. Envir. Microbiol. 65, 1746-1752
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Counting and Size Classification of Active Soil Bacteria by Fluorescence In Situ Hybridization with an rRNA Oligonucleotide Probe.
H. Christensen, M. Hansen, and J. Sørensen (1999)
Appl. Envir. Microbiol. 65, 1753-1761
   Abstract »    Full Text »
Physiological States of Individual Salmonella typhimurium Cells Monitored by In Situ Reverse Transcription-PCR.
K. Holmstrøm, T. Tolker-Nielsen, and S. Molin (1999)
J. Bacteriol. 181, 1733-1738
   Abstract »    Full Text »
Immunochemical Detection and Isolation of DNA from Metabolically Active Bacteria.
E. Urbach, K. L. Vergin, and S. J. Giovannoni (1999)
Appl. Envir. Microbiol. 65, 1207-1213
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Identification of a Novel Group of Bacteria in Sludge from a Deteriorated Biological Phosphorus Removal Reactor.
A. T. Nielsen, W.-T. Liu, C. Filipe, L. Grady Jr., S. Molin, and D. A. Stahl (1999)
Appl. Envir. Microbiol. 65, 1251-1258
   Abstract »    Full Text »
Enumeration and Detection of Anaerobic Ferrous Iron-Oxidizing, Nitrate-Reducing Bacteria from Diverse European Sediments.
K. L. Straub and B. E. E. Buchholz-Cleven (1998)
Appl. Envir. Microbiol. 64, 4846-4856
   Abstract »    Full Text »
Flow Cytometric Analysis of the In Situ Accessibility of Escherichia coli 16S rRNA for Fluorescently Labeled Oligonucleotide Probes.
B. M. Fuchs, G. Wallner, W. Beisker, I. Schwippl, W. Ludwig, and R. Amann (1998)
Appl. Envir. Microbiol. 64, 4973-4982
   Abstract »    Full Text »
Species-Specific Oligonucleotides for Enumeration of Pseudomonas putida F1, Burkholderia sp. Strain JS150, and Bacillus subtilis ATCC 7003 in Biodegradation Experiments.
N. M. DuTeau, J. D. Rogers, C. T. Bartholomay, and K. F. Reardon (1998)
Appl. Envir. Microbiol. 64, 4994-4999
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Growth Rate Regulation of rRNA Content of a Marine Synechococcus (Cyanobacterium) Strain.
B. J. Binder and Y. C. Liu (1998)
Appl. Envir. Microbiol. 64, 3346-3351
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Specific Ribosomal DNA Sequences from Diverse Environmental Settings Correlate with Experimental Contaminants.
M. A. Tanner, B. M. Goebel, M. A. Dojka, and N. R. Pace (1998)
Appl. Envir. Microbiol. 64, 3110-3113
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Seasonal and Spatial Variability of Bacterial and Archaeal Assemblages in the Coastal Waters near Anvers Island, Antarctica.
A. E. Murray, C. M. Preston, R. Massana, L. T. Taylor, A. Blakis, K. Wu, and E. F. DeLong (1998)
Appl. Envir. Microbiol. 64, 2585-2595
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Specific Detection of Legionella pneumophila: Construction of a New 16S rRNA-Targeted Oligonucleotide Probe.
D. Grimm, H. Merkert, W. Ludwig, K.-H. Schleifer, J. Hacker, and B. C. Brand (1998)
Appl. Envir. Microbiol. 64, 2686-2690
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Population Dynamics of Phenol-Degrading Bacteria in Activated Sludge Determined by gyrB-Targeted Quantitative PCR.
K. Watanabe, S. Yamamoto, S. Hino, and S. Harayama (1998)
Appl. Envir. Microbiol. 64, 1203-1209
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In Situ Gene Expression in Mixed-Culture Biofilms: Evidence of Metabolic Interactions between Community Members.
S. Møller, C. Sternberg, J. B. Andersen, B. B. Christensen, J. L. Ramos, M. Givskov, and S. Molin (1998)
Appl. Envir. Microbiol. 64, 721-732
   Abstract »    Full Text »
Multiple Evolutionary Origins of Magnetotaxis in Bacteria.
E. F. DeLong, R. B. Frankel, and D. A. Bazylinski (1993)
Science 259, 803-806
   Abstract »    PDF »


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