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. 1995 Mar;59(1):143–169. doi: 10.1128/mr.59.1.143-169.1995

Phylogenetic identification and in situ detection of individual microbial cells without cultivation.

R I Amann 1, W Ludwig 1, K H Schleifer 1
PMCID: PMC239358  PMID: 7535888

Abstract

The frequent discrepancy between direct microscopic counts and numbers of culturable bacteria from environmental samples is just one of several indications that we currently know only a minor part of the diversity of microorganisms in nature. A combination of direct retrieval of rRNA sequences and whole-cell oligonucleotide probing can be used to detect specific rRNA sequences of uncultured bacteria in natural samples and to microscopically identify individual cells. Studies have been performed with microbial assemblages of various complexities ranging from simple two-component bacterial endosymbiotic associations to multispecies enrichments containing magnetotactic bacteria to highly complex marine and soil communities. Phylogenetic analysis of the retrieved rRNA sequence of an uncultured microorganism reveals its closest culturable relatives and may, together with information on the physicochemical conditions of its natural habitat, facilitate more directed cultivation attempts. For the analysis of complex communities such as multispecies biofilms and activated-sludge flocs, a different approach has proven advantageous. Sets of probes specific to different taxonomic levels are applied consecutively beginning with the more general and ending with the more specific (a hierarchical top-to-bottom approach), thereby generating increasingly precise information on the structure of the community. Not only do rRNA-targeted whole-cell hybridizations yield data on cell morphology, specific cell counts, and in situ distributions of defined phylogenetic groups, but also the strength of the hybridization signal reflects the cellular rRNA content of individual cells. From the signal strength conferred by a specific probe, in situ growth rates and activities of individual cells might be estimated for known species. In many ecosystems, low cellular rRNA content and/or limited cell permeability, combined with background fluorescence, hinders in situ identification of autochthonous populations. Approaches to circumvent these problems are discussed in detail.

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Selected References

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  1. A unique symbiosis in the gut of tropical herbivorous surgeonfish (acanthuridae: teleostei) from the red sea. Science. 1985 Jul 5;229(4708):49–51. doi: 10.1126/science.229.4708.49. [DOI] [PubMed] [Google Scholar]
  2. Amann R. I., Binder B. J., Olson R. J., Chisholm S. W., Devereux R., Stahl D. A. Combination of 16S rRNA-targeted oligonucleotide probes with flow cytometry for analyzing mixed microbial populations. Appl Environ Microbiol. 1990 Jun;56(6):1919–1925. doi: 10.1128/aem.56.6.1919-1925.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Amann R. I., Krumholz L., Stahl D. A. Fluorescent-oligonucleotide probing of whole cells for determinative, phylogenetic, and environmental studies in microbiology. J Bacteriol. 1990 Feb;172(2):762–770. doi: 10.1128/jb.172.2.762-770.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Amann R. I., Stromley J., Devereux R., Key R., Stahl D. A. Molecular and microscopic identification of sulfate-reducing bacteria in multispecies biofilms. Appl Environ Microbiol. 1992 Feb;58(2):614–623. doi: 10.1128/aem.58.2.614-623.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Amann R. I., Zarda B., Stahl D. A., Schleifer K. H. Identification of individual prokaryotic cells by using enzyme-labeled, rRNA-targeted oligonucleotide probes. Appl Environ Microbiol. 1992 Sep;58(9):3007–3011. doi: 10.1128/aem.58.9.3007-3011.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Amann R., Springer N., Ludwig W., Görtz H. D., Schleifer K. H. Identification in situ and phylogeny of uncultured bacterial endosymbionts. Nature. 1991 May 9;351(6322):161–164. doi: 10.1038/351161a0. [DOI] [PubMed] [Google Scholar]
  7. Amikam D., Glaser G., Razin S. Mycoplasmas (Mollicutes) have a low number of rRNA genes. J Bacteriol. 1984 Apr;158(1):376–378. doi: 10.1128/jb.158.1.376-378.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Angert E. R., Clements K. D., Pace N. R. The largest bacterium. Nature. 1993 Mar 18;362(6417):239–241. doi: 10.1038/362239a0. [DOI] [PubMed] [Google Scholar]
  9. Arnoldi J., Schlüter C., Duchrow M., Hübner L., Ernst M., Teske A., Flad H. D., Gerdes J., Böttger E. C. Species-specific assessment of Mycobacterium leprae in skin biopsies by in situ hybridization and polymerase chain reaction. Lab Invest. 1992 May;66(5):618–623. [PubMed] [Google Scholar]
  10. Ashkin A., Dziedzic J. M., Yamane T. Optical trapping and manipulation of single cells using infrared laser beams. Nature. 1987 Dec 24;330(6150):769–771. doi: 10.1038/330769a0. [DOI] [PubMed] [Google Scholar]
  11. Aznar R., Ludwig W., Amann R. I., Schleifer K. H. Sequence determination of rRNA genes of pathogenic Vibrio species and whole-cell identification of Vibrio vulnificus with rRNA-targeted oligonucleotide probes. Int J Syst Bacteriol. 1994 Apr;44(2):330–337. doi: 10.1099/00207713-44-2-330. [DOI] [PubMed] [Google Scholar]
  12. Bauman J. G., Bentvelzen P. Flow cytometric detection of ribosomal RNA in suspended cells by fluorescent in situ hybridization. Cytometry. 1988 Nov;9(6):517–524. doi: 10.1002/cyto.990090602. [DOI] [PubMed] [Google Scholar]
  13. Betzl D., Ludwig W., Schleifer K. H. Identification of lactococci and enterococci by colony hybridization with 23S rRNA-targeted oligonucleotide probes. Appl Environ Microbiol. 1990 Sep;56(9):2927–2929. doi: 10.1128/aem.56.9.2927-2929.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Braun-Howland E. B., Danielsen S. A., Nierzwicki-Bauer S. A. Development of a rapid method for detecting bacterial cells in situ using 16S rRNA-targeted probes. Biotechniques. 1992 Dec;13(6):928–934. [PubMed] [Google Scholar]
  15. Brenner D. J., O'Connor S. P., Winkler H. H., Steigerwalt A. G. Proposals to unify the genera Bartonella and Rochalimaea, with descriptions of Bartonella quintana comb. nov., Bartonella vinsonii comb. nov., Bartonella henselae comb. nov., and Bartonella elizabethae comb. nov., and to remove the family Bartonellaceae from the order Rickettsiales. Int J Syst Bacteriol. 1993 Oct;43(4):777–786. doi: 10.1099/00207713-43-4-777. [DOI] [PubMed] [Google Scholar]
  16. Britschgi T. B., Giovannoni S. J. Phylogenetic analysis of a natural marine bacterioplankton population by rRNA gene cloning and sequencing. Appl Environ Microbiol. 1991 Jun;57(6):1707–1713. doi: 10.1128/aem.57.6.1707-1713.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Brosius J., Dull T. J., Sleeter D. D., Noller H. F. Gene organization and primary structure of a ribosomal RNA operon from Escherichia coli. J Mol Biol. 1981 May 15;148(2):107–127. doi: 10.1016/0022-2836(81)90508-8. [DOI] [PubMed] [Google Scholar]
  18. Bull A. T., Goodfellow M., Slater J. H. Biodiversity as a source of innovation in biotechnology. Annu Rev Microbiol. 1992;46:219–252. doi: 10.1146/annurev.mi.46.100192.001251. [DOI] [PubMed] [Google Scholar]
  19. Burggraf S., Mayer T., Amann R., Schadhauser S., Woese C. R., Stetter K. O. Identifying members of the domain Archaea with rRNA-targeted oligonucleotide probes. Appl Environ Microbiol. 1994 Sep;60(9):3112–3119. doi: 10.1128/aem.60.9.3112-3119.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Button D. K., Schut F., Quang P., Martin R., Robertson B. R. Viability and isolation of marine bacteria by dilution culture: theory, procedures, and initial results. Appl Environ Microbiol. 1993 Mar;59(3):881–891. doi: 10.1128/aem.59.3.881-891.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Cary S. C., Giovannoni S. J. Transovarial inheritance of endosymbiotic bacteria in clams inhabiting deep-sea hydrothermal vents and cold seeps. Proc Natl Acad Sci U S A. 1993 Jun 15;90(12):5695–5699. doi: 10.1073/pnas.90.12.5695. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. DeLong E. F. Archaea in coastal marine environments. Proc Natl Acad Sci U S A. 1992 Jun 15;89(12):5685–5689. doi: 10.1073/pnas.89.12.5685. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. DeLong E. F., Wickham G. S., Pace N. R. Phylogenetic stains: ribosomal RNA-based probes for the identification of single cells. Science. 1989 Mar 10;243(4896):1360–1363. doi: 10.1126/science.2466341. [DOI] [PubMed] [Google Scholar]
  24. Delong E. F., Frankel R. B., Bazylinski D. A. Multiple evolutionary origins of magnetotaxis in bacteria. Science. 1993 Feb 5;259(5096):803–806. doi: 10.1126/science.259.5096.803. [DOI] [PubMed] [Google Scholar]
  25. DiChristina T. J., DeLong E. F. Design and application of rRNA-targeted oligonucleotide probes for the dissimilatory iron- and manganese-reducing bacterium Shewanella putrefaciens. Appl Environ Microbiol. 1993 Dec;59(12):4152–4160. doi: 10.1128/aem.59.12.4152-4160.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Distel D. L., DeLong E. F., Waterbury J. B. Phylogenetic characterization and in situ localization of the bacterial symbiont of shipworms (Teredinidae: Bivalvia) by using 16S rRNA sequence analysis and oligodeoxynucleotide probe hybridization. Appl Environ Microbiol. 1991 Aug;57(8):2376–2382. doi: 10.1128/aem.57.8.2376-2382.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Distel D. L., Lane D. J., Olsen G. J., Giovannoni S. J., Pace B., Pace N. R., Stahl D. A., Felbeck H. Sulfur-oxidizing bacterial endosymbionts: analysis of phylogeny and specificity by 16S rRNA sequences. J Bacteriol. 1988 Jun;170(6):2506–2510. doi: 10.1128/jb.170.6.2506-2510.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Distel D. L., Wood A. P. Characterization of the gill symbiont of Thyasira flexuosa (Thyasiridae: Bivalvia) by use of polymerase chain reaction and 16S rRNA sequence analysis. J Bacteriol. 1992 Oct;174(19):6317–6320. doi: 10.1128/jb.174.19.6317-6320.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Edman J. C., Kovacs J. A., Masur H., Santi D. V., Elwood H. J., Sogin M. L. Ribosomal RNA sequence shows Pneumocystis carinii to be a member of the fungi. Nature. 1988 Aug 11;334(6182):519–522. doi: 10.1038/334519a0. [DOI] [PubMed] [Google Scholar]
  30. Embley T. M., Finlay B. J., Brown S. RNA sequence analysis shows that the symbionts in the ciliate Metopus contortus are polymorphs of a single methanogen species. FEMS Microbiol Lett. 1992 Oct 1;76(1-2):57–61. doi: 10.1016/0378-1097(92)90363-s. [DOI] [PubMed] [Google Scholar]
  31. Embley T. M., Finlay B. J., Thomas R. H., Dyal P. L. The use of rRNA sequences and fluorescent probes to investigate the phylogenetic positions of the anaerobic ciliate Metopus palaeformis and its archaeobacterial endosymbiont. J Gen Microbiol. 1992 Jul;138(7):1479–1487. doi: 10.1099/00221287-138-7-1479. [DOI] [PubMed] [Google Scholar]
  32. Fenchel T., Perry T., Thane A. Anaerobiosis and symbiosis with bacteria in free-living ciliates. J Protozool. 1977 Feb;24(1):154–163. doi: 10.1111/j.1550-7408.1977.tb05294.x. [DOI] [PubMed] [Google Scholar]
  33. Fenchel T., Ramsing N. B. Identification of sulphate-reducing ectosymbiotic bacteria from anaerobic ciliates using 16S rRNA binding oligonucleotide probes. Arch Microbiol. 1992;158(6):394–397. doi: 10.1007/BF00276298. [DOI] [PubMed] [Google Scholar]
  34. Ferguson R. L., Buckley E. N., Palumbo A. V. Response of marine bacterioplankton to differential filtration and confinement. Appl Environ Microbiol. 1984 Jan;47(1):49–55. doi: 10.1128/aem.47.1.49-55.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Finlay B. J., Embley T. M., Fenchel T. A new polymorphic methanogen, closely related to Methanocorpusculum parvum, living in stable symbiosis within the anaerobic ciliate Trimyema sp. J Gen Microbiol. 1993 Feb;139(2):371–378. doi: 10.1099/00221287-139-2-371. [DOI] [PubMed] [Google Scholar]
  36. Flärdh K., Cohen P. S., Kjelleberg S. Ribosomes exist in large excess over the apparent demand for protein synthesis during carbon starvation in marine Vibrio sp. strain CCUG 15956. J Bacteriol. 1992 Nov;174(21):6780–6788. doi: 10.1128/jb.174.21.6780-6788.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Fox G. E., Wisotzkey J. D., Jurtshuk P., Jr How close is close: 16S rRNA sequence identity may not be sufficient to guarantee species identity. Int J Syst Bacteriol. 1992 Jan;42(1):166–170. doi: 10.1099/00207713-42-1-166. [DOI] [PubMed] [Google Scholar]
  38. Fry N. K., Rowbotham T. J., Saunders N. A., Embley T. M. Direct amplification and sequencing of the 16S ribosomal DNA of an intracellular Legionella species recovered by amoebal enrichment from the sputum of a patient with pneumonia. FEMS Microbiol Lett. 1991 Oct 1;67(2):165–168. doi: 10.1016/0378-1097(91)90348-e. [DOI] [PubMed] [Google Scholar]
  39. Fuhrman J. A., McCallum K., Davis A. A. Novel major archaebacterial group from marine plankton. Nature. 1992 Mar 12;356(6365):148–149. doi: 10.1038/356148a0. [DOI] [PubMed] [Google Scholar]
  40. Fuhrman J. A., McCallum K., Davis A. A. Phylogenetic diversity of subsurface marine microbial communities from the Atlantic and Pacific Oceans. Appl Environ Microbiol. 1993 May;59(5):1294–1302. doi: 10.1128/aem.59.5.1294-1302.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Gebhart C. J., Barns S. M., McOrist S., Lin G. F., Lawson G. H. Ileal symbiont intracellularis, an obligate intracellular bacterium of porcine intestines showing a relationship to Desulfovibrio species. Int J Syst Bacteriol. 1993 Jul;43(3):533–538. doi: 10.1099/00207713-43-3-533. [DOI] [PubMed] [Google Scholar]
  42. Gersdorf H., Meissner A., Pelz K., Krekeler G., Göbel U. B. Identification of Bacteroides forsythus in subgingival plaque from patients with advanced periodontitis. J Clin Microbiol. 1993 Apr;31(4):941–946. doi: 10.1128/jcm.31.4.941-946.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Gersdorf H., Pelz K., Göbel U. B. Fluorescence in situ hybridization for direct visualization of gram-negative anaerobes in subgingival plaque samples. FEMS Immunol Med Microbiol. 1993 Mar;6(2-3):109–114. doi: 10.1111/j.1574-695X.1993.tb00312.x. [DOI] [PubMed] [Google Scholar]
  44. Giovannoni S. J., Britschgi T. B., Moyer C. L., Field K. G. Genetic diversity in Sargasso Sea bacterioplankton. Nature. 1990 May 3;345(6270):60–63. doi: 10.1038/345060a0. [DOI] [PubMed] [Google Scholar]
  45. Giovannoni S. J., DeLong E. F., Olsen G. J., Pace N. R. Phylogenetic group-specific oligodeoxynucleotide probes for identification of single microbial cells. J Bacteriol. 1988 Feb;170(2):720–726. doi: 10.1128/jb.170.2.720-726.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  46. Giovannoni S. J., DeLong E. F., Schmidt T. M., Pace N. R. Tangential flow filtration and preliminary phylogenetic analysis of marine picoplankton. Appl Environ Microbiol. 1990 Aug;56(8):2572–2575. doi: 10.1128/aem.56.8.2572-2575.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  47. Göbel U. B., Geiser A., Stanbridge E. J. Oligonucleotide probes complementary to variable regions of ribosomal RNA discriminate between Mycoplasma species. J Gen Microbiol. 1987 Jul;133(7):1969–1974. doi: 10.1099/00221287-133-7-1969. [DOI] [PubMed] [Google Scholar]
  48. Hahn D., Amann R. I., Ludwig W., Akkermans A. D., Schleifer K. H. Detection of micro-organisms in soil after in situ hybridization with rRNA-targeted, fluorescently labelled oligonucleotides. J Gen Microbiol. 1992 May;138(5):879–887. doi: 10.1099/00221287-138-5-879. [DOI] [PubMed] [Google Scholar]
  49. Hahn D., Amann R. I., Zeyer J. Detection of mRNA in streptomyces cells by whole-cell hybridization with digoxigenin-labeled probes. Appl Environ Microbiol. 1993 Aug;59(8):2753–2757. doi: 10.1128/aem.59.8.2753-2757.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  50. Hahn D., Amann R. I., Zeyer J. Whole-Cell Hybridization of Frankia Strains with Fluorescence- or Digoxigenin-Labeled, 16S rRNA-Targeted Oligonucleotide Probes. Appl Environ Microbiol. 1993 Jun;59(6):1709–1716. doi: 10.1128/aem.59.6.1709-1716.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  51. Haygood M. G., Distel D. L. Bioluminescent symbionts of flashlight fishes and deep-sea anglerfishes form unique lineages related to the genus Vibrio. Nature. 1993 May 13;363(6425):154–156. doi: 10.1038/363154a0. [DOI] [PubMed] [Google Scholar]
  52. Hicks R. E., Amann R. I., Stahl D. A. Dual staining of natural bacterioplankton with 4',6-diamidino-2-phenylindole and fluorescent oligonucleotide probes targeting kingdom-level 16S rRNA sequences. Appl Environ Microbiol. 1992 Jul;58(7):2158–2163. doi: 10.1128/aem.58.7.2158-2163.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  53. Jarvis E. D., Widom R. L., LaFauci G., Setoguchi Y., Richter I. R., Rudner R. Chromosomal organization of rRNA operons in Bacillus subtilis. Genetics. 1988 Nov;120(3):625–635. doi: 10.1093/genetics/120.3.625. [DOI] [PMC free article] [PubMed] [Google Scholar]
  54. Jurtshuk R. J., Blick M., Bresser J., Fox G. E., Jurtshuk P., Jr Rapid in situ hybridization technique using 16S rRNA segments for detecting and differentiating the closely related gram-positive organisms Bacillus polymyxa and Bacillus macerans. Appl Environ Microbiol. 1992 Aug;58(8):2571–2578. doi: 10.1128/aem.58.8.2571-2578.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  55. Kane M. D., Poulsen L. K., Stahl D. A. Monitoring the enrichment and isolation of sulfate-reducing bacteria by using oligonucleotide hybridization probes designed from environmentally derived 16S rRNA sequences. Appl Environ Microbiol. 1993 Mar;59(3):682–686. doi: 10.1128/aem.59.3.682-686.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  56. Kemp P. F., Lee S., Laroche J. Estimating the growth rate of slowly growing marine bacteria from RNA content. Appl Environ Microbiol. 1993 Aug;59(8):2594–2601. doi: 10.1128/aem.59.8.2594-2601.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  57. Kerkhof L., Ward B. B. Comparison of Nucleic Acid Hybridization and Fluorometry for Measurement of the Relationship between RNA/DNA Ratio and Growth Rate in a Marine Bacterium. Appl Environ Microbiol. 1993 May;59(5):1303–1309. doi: 10.1128/aem.59.5.1303-1309.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  58. Khalfan H., Abuknesha R., Rand-Weaver M., Price R. G., Robinson D. Aminomethyl coumarin acetic acid: a new fluorescent labelling agent for proteins. Histochem J. 1986 Sep;18(9):497–499. doi: 10.1007/BF01675617. [DOI] [PubMed] [Google Scholar]
  59. Kogure K., Simidu U., Taga N. A tentative direct microscopic method for counting living marine bacteria. Can J Microbiol. 1979 Mar;25(3):415–420. doi: 10.1139/m79-063. [DOI] [PubMed] [Google Scholar]
  60. Kogure K., Simidu U., Taga N. Distribution of viable marine bacteria in neritic seawater around Japan. Can J Microbiol. 1980 Mar;26(3):318–323. doi: 10.1139/m80-052. [DOI] [PubMed] [Google Scholar]
  61. Kramer J. G., Singleton F. L. Measurement of rRNA Variations in Natural Communities of Microorganisms on the Southeastern U.S. Continental Shelf. Appl Environ Microbiol. 1993 Aug;59(8):2430–2436. doi: 10.1128/aem.59.8.2430-2436.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  62. Lane D. J., Stahl D. A., Olsen G. J., Heller D. J., Pace N. R. Phylogenetic analysis of the genera Thiobacillus and Thiomicrospira by 5S rRNA sequences. J Bacteriol. 1985 Jul;163(1):75–81. doi: 10.1128/jb.163.1.75-81.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  63. Larsen N., Olsen G. J., Maidak B. L., McCaughey M. J., Overbeek R., Macke T. J., Marsh T. L., Woese C. R. The ribosomal database project. Nucleic Acids Res. 1993 Jul 1;21(13):3021–3023. doi: 10.1093/nar/21.13.3021. [DOI] [PMC free article] [PubMed] [Google Scholar]
  64. Liesack W., Stackebrandt E. Occurrence of novel groups of the domain Bacteria as revealed by analysis of genetic material isolated from an Australian terrestrial environment. J Bacteriol. 1992 Aug;174(15):5072–5078. doi: 10.1128/jb.174.15.5072-5078.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  65. Lim E. L., Amaral L. A., Caron D. A., DeLong E. F. Application of rRNA-based probes for observing marine nanoplanktonic protists. Appl Environ Microbiol. 1993 May;59(5):1647–1655. doi: 10.1128/aem.59.5.1647-1655.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  66. Mann S., Sparks N. H., Board R. G. Magnetotactic bacteria: microbiology, biomineralization, palaeomagnetism and biotechnology. Adv Microb Physiol. 1990;31:125–181. doi: 10.1016/s0065-2911(08)60121-6. [DOI] [PubMed] [Google Scholar]
  67. Manz W., Szewzyk U., Ericsson P., Amann R., Schleifer K. H., Stenström T. A. In situ identification of bacteria in drinking water and adjoining biofilms by hybridization with 16S and 23S rRNA-directed fluorescent oligonucleotide probes. Appl Environ Microbiol. 1993 Jul;59(7):2293–2298. doi: 10.1128/aem.59.7.2293-2298.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  68. McSweeney C. S., Mackie R. I., Odenyo A. A., Stahl D. A. Development of an oligonucleotide probe targeting 16S rRNA and its application for detection and quantitation of the ruminal bacterium Synergistes jonesii in a mixed-population chemostat. Appl Environ Microbiol. 1993 May;59(5):1607–1612. doi: 10.1128/aem.59.5.1607-1612.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  69. Moench T. T., Konetzka W. A. A novel method for the isolation and study of a magnetotactic bacterium. Arch Microbiol. 1978 Nov 13;119(2):203–212. doi: 10.1007/BF00964274. [DOI] [PubMed] [Google Scholar]
  70. Munson M. A., Baumann P., Clark M. A., Baumann L., Moran N. A., Voegtlin D. J., Campbell B. C. Evidence for the establishment of aphid-eubacterium endosymbiosis in an ancestor of four aphid families. J Bacteriol. 1991 Oct;173(20):6321–6324. doi: 10.1128/jb.173.20.6321-6324.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  71. Muyzer G., de Waal E. C., Uitterlinden A. G. Profiling of complex microbial populations by denaturing gradient gel electrophoresis analysis of polymerase chain reaction-amplified genes coding for 16S rRNA. Appl Environ Microbiol. 1993 Mar;59(3):695–700. doi: 10.1128/aem.59.3.695-700.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  72. Namba S., Oyaizu H., Kato S., Iwanami S., Tsuchizaki T. Phylogenetic diversity of phytopathogenic mycoplasmalike organisms. Int J Syst Bacteriol. 1993 Jul;43(3):461–467. doi: 10.1099/00207713-43-3-461. [DOI] [PubMed] [Google Scholar]
  73. Neefs J. M., Van de Peer Y., De Rijk P., Chapelle S., De Wachter R. Compilation of small ribosomal subunit RNA structures. Nucleic Acids Res. 1993 Jul 1;21(13):3025–3049. doi: 10.1093/nar/21.13.3025. [DOI] [PMC free article] [PubMed] [Google Scholar]
  74. Neimark H., Kirkpatrick B. C. Isolation and characterization of full-length chromosomes from non-culturable plant-pathogenic Mycoplasma-like organisms. Mol Microbiol. 1993 Jan;7(1):21–28. doi: 10.1111/j.1365-2958.1993.tb01093.x. [DOI] [PubMed] [Google Scholar]
  75. O'Neill S. L., Giordano R., Colbert A. M., Karr T. L., Robertson H. M. 16S rRNA phylogenetic analysis of the bacterial endosymbionts associated with cytoplasmic incompatibility in insects. Proc Natl Acad Sci U S A. 1992 Apr 1;89(7):2699–2702. doi: 10.1073/pnas.89.7.2699. [DOI] [PMC free article] [PubMed] [Google Scholar]
  76. Oliver J. D., Nilsson L., Kjelleberg S. Formation of nonculturable Vibrio vulnificus cells and its relationship to the starvation state. Appl Environ Microbiol. 1991 Sep;57(9):2640–2644. doi: 10.1128/aem.57.9.2640-2644.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  77. Olsen G. J., Lane D. J., Giovannoni S. J., Pace N. R., Stahl D. A. Microbial ecology and evolution: a ribosomal RNA approach. Annu Rev Microbiol. 1986;40:337–365. doi: 10.1146/annurev.mi.40.100186.002005. [DOI] [PubMed] [Google Scholar]
  78. Olsen G. J., Woese C. R. Ribosomal RNA: a key to phylogeny. FASEB J. 1993 Jan;7(1):113–123. doi: 10.1096/fasebj.7.1.8422957. [DOI] [PubMed] [Google Scholar]
  79. Poulsen L. K., Ballard G., Stahl D. A. Use of rRNA fluorescence in situ hybridization for measuring the activity of single cells in young and established biofilms. Appl Environ Microbiol. 1993 May;59(5):1354–1360. doi: 10.1128/aem.59.5.1354-1360.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  80. Ramsing N. B., Kühl M., Jørgensen B. B. Distribution of sulfate-reducing bacteria, O2, and H2S in photosynthetic biofilms determined by oligonucleotide probes and microelectrodes. Appl Environ Microbiol. 1993 Nov;59(11):3840–3849. doi: 10.1128/aem.59.11.3840-3849.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  81. Raskin L., Poulsen L. K., Noguera D. R., Rittmann B. E., Stahl D. A. Quantification of methanogenic groups in anaerobic biological reactors by oligonucleotide probe hybridization. Appl Environ Microbiol. 1994 Apr;60(4):1241–1248. doi: 10.1128/aem.60.4.1241-1248.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  82. Raskin L., Stromley J. M., Rittmann B. E., Stahl D. A. Group-specific 16S rRNA hybridization probes to describe natural communities of methanogens. Appl Environ Microbiol. 1994 Apr;60(4):1232–1240. doi: 10.1128/aem.60.4.1232-1240.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  83. Regnery R. L., Anderson B. E., Clarridge J. E., 3rd, Rodriguez-Barradas M. C., Jones D. C., Carr J. H. Characterization of a novel Rochalimaea species, R. henselae sp. nov., isolated from blood of a febrile, human immunodeficiency virus-positive patient. J Clin Microbiol. 1992 Feb;30(2):265–274. doi: 10.1128/jcm.30.2.265-274.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  84. Relman D. A., Lepp P. W., Sadler K. N., Schmidt T. M. Phylogenetic relationships among the agent of bacillary angiomatosis, Bartonella bacilliformis, and other alpha-proteobacteria. Mol Microbiol. 1992 Jul;6(13):1801–1807. doi: 10.1111/j.1365-2958.1992.tb01352.x. [DOI] [PubMed] [Google Scholar]
  85. Relman D. A., Loutit J. S., Schmidt T. M., Falkow S., Tompkins L. S. The agent of bacillary angiomatosis. An approach to the identification of uncultured pathogens. N Engl J Med. 1990 Dec 6;323(23):1573–1580. doi: 10.1056/NEJM199012063232301. [DOI] [PubMed] [Google Scholar]
  86. Relman D. A., Schmidt T. M., MacDermott R. P., Falkow S. Identification of the uncultured bacillus of Whipple's disease. N Engl J Med. 1992 Jul 30;327(5):293–301. doi: 10.1056/NEJM199207303270501. [DOI] [PubMed] [Google Scholar]
  87. Reysenbach A. L., Giver L. J., Wickham G. S., Pace N. R. Differential amplification of rRNA genes by polymerase chain reaction. Appl Environ Microbiol. 1992 Oct;58(10):3417–3418. doi: 10.1128/aem.58.10.3417-3418.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  88. Roller C., Wagner M., Amann R., Ludwig W., Schleifer K. H. In situ probing of gram-positive bacteria with high DNA G + C content using 23S rRNA-targeted oligonucleotides. Microbiology. 1994 Oct;140(Pt 10):2849–2858. doi: 10.1099/00221287-140-10-2849. [DOI] [PubMed] [Google Scholar]
  89. Roszak D. B., Colwell R. R. Survival strategies of bacteria in the natural environment. Microbiol Rev. 1987 Sep;51(3):365–379. doi: 10.1128/mr.51.3.365-379.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  90. Roszak D. B., Grimes D. J., Colwell R. R. Viable but nonrecoverable stage of Salmonella enteritidis in aquatic systems. Can J Microbiol. 1984 Mar;30(3):334–338. doi: 10.1139/m84-049. [DOI] [PubMed] [Google Scholar]
  91. SCHAECHTER M., MAALOE O., KJELDGAARD N. O. Dependency on medium and temperature of cell size and chemical composition during balanced grown of Salmonella typhimurium. J Gen Microbiol. 1958 Dec;19(3):592–606. doi: 10.1099/00221287-19-3-592. [DOI] [PubMed] [Google Scholar]
  92. Saiki R. K., Gelfand D. H., Stoffel S., Scharf S. J., Higuchi R., Horn G. T., Mullis K. B., Erlich H. A. Primer-directed enzymatic amplification of DNA with a thermostable DNA polymerase. Science. 1988 Jan 29;239(4839):487–491. doi: 10.1126/science.2448875. [DOI] [PubMed] [Google Scholar]
  93. Salama M., Sandine W., Giovannoni S. Development and application of oligonucleotide probes for identification of Lactococcus lactis subsp. cremoris. Appl Environ Microbiol. 1991 May;57(5):1313–1318. doi: 10.1128/aem.57.5.1313-1318.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  94. Schleifer K. H., Stackebrandt E. Molecular systematics of prokaryotes. Annu Rev Microbiol. 1983;37:143–187. doi: 10.1146/annurev.mi.37.100183.001043. [DOI] [PubMed] [Google Scholar]
  95. Schmidt T. M., DeLong E. F., Pace N. R. Analysis of a marine picoplankton community by 16S rRNA gene cloning and sequencing. J Bacteriol. 1991 Jul;173(14):4371–4378. doi: 10.1128/jb.173.14.4371-4378.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  96. Schut F., de Vries E. J., Gottschal J. C., Robertson B. R., Harder W., Prins R. A., Button D. K. Isolation of Typical Marine Bacteria by Dilution Culture: Growth, Maintenance, and Characteristics of Isolates under Laboratory Conditions. Appl Environ Microbiol. 1993 Jul;59(7):2150–2160. doi: 10.1128/aem.59.7.2150-2160.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  97. Southwick P. L., Ernst L. A., Tauriello E. W., Parker S. R., Mujumdar R. B., Mujumdar S. R., Clever H. A., Waggoner A. S. Cyanine dye labeling reagents--carboxymethylindocyanine succinimidyl esters. Cytometry. 1990;11(3):418–430. doi: 10.1002/cyto.990110313. [DOI] [PubMed] [Google Scholar]
  98. Spring S., Amann R., Ludwig W., Schleifer K. H., van Gemerden H., Petersen N. Dominating role of an unusual magnetotactic bacterium in the microaerobic zone of a freshwater sediment. Appl Environ Microbiol. 1993 Aug;59(8):2397–2403. doi: 10.1128/aem.59.8.2397-2403.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  99. Springer N., Ludwig W., Amann R., Schmidt H. J., Görtz H. D., Schleifer K. H. Occurrence of fragmented 16S rRNA in an obligate bacterial endosymbiont of Paramecium caudatum. Proc Natl Acad Sci U S A. 1993 Nov 1;90(21):9892–9895. doi: 10.1073/pnas.90.21.9892. [DOI] [PMC free article] [PubMed] [Google Scholar]
  100. Springer N., Ludwig W., Drozański W., Amann R., Schleifer K. H. The phylogenetic status of Sarcobium lyticum, an obligate intracellular bacterial parasite of small amoebae. FEMS Microbiol Lett. 1992 Sep 15;75(2-3):199–202. doi: 10.1016/0378-1097(92)90403-b. [DOI] [PubMed] [Google Scholar]
  101. Stackebrandt E., Liesack W., Goebel B. M. Bacterial diversity in a soil sample from a subtropical Australian environment as determined by 16S rDNA analysis. FASEB J. 1993 Jan;7(1):232–236. doi: 10.1096/fasebj.7.1.8422969. [DOI] [PubMed] [Google Scholar]
  102. Stahl D. A., Flesher B., Mansfield H. R., Montgomery L. Use of phylogenetically based hybridization probes for studies of ruminal microbial ecology. Appl Environ Microbiol. 1988 May;54(5):1079–1084. doi: 10.1128/aem.54.5.1079-1084.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  103. Stahl D. A., Lane D. J., Olsen G. J., Pace N. R. Analysis of hydrothermal vent-associated symbionts by ribosomal RNA sequences. Science. 1984 Apr 27;224(4647):409–411. doi: 10.1126/science.224.4647.409. [DOI] [PubMed] [Google Scholar]
  104. Stahl D. A., Lane D. J., Olsen G. J., Pace N. R. Characterization of a Yellowstone hot spring microbial community by 5S rRNA sequences. Appl Environ Microbiol. 1985 Jun;49(6):1379–1384. doi: 10.1128/aem.49.6.1379-1384.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  105. Staley J. T., Konopka A. Measurement of in situ activities of nonphotosynthetic microorganisms in aquatic and terrestrial habitats. Annu Rev Microbiol. 1985;39:321–346. doi: 10.1146/annurev.mi.39.100185.001541. [DOI] [PubMed] [Google Scholar]
  106. Stouthamer R., Breeuwert J. A., Luck R. F., Werren J. H. Molecular identification of microorganisms associated with parthenogenesis. Nature. 1993 Jan 7;361(6407):66–68. doi: 10.1038/361066a0. [DOI] [PubMed] [Google Scholar]
  107. Torsvik V., Goksøyr J., Daae F. L. High diversity in DNA of soil bacteria. Appl Environ Microbiol. 1990 Mar;56(3):782–787. doi: 10.1128/aem.56.3.782-787.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  108. Tsien H. C., Bratina B. J., Tsuji K., Hanson R. S. Use of oligodeoxynucleotide signature probes for identification of physiological groups of methylotrophic bacteria. Appl Environ Microbiol. 1990 Sep;56(9):2858–2865. doi: 10.1128/aem.56.9.2858-2865.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  109. Unterman B. M., Baumann P., McLean D. L. Pea aphid symbiont relationships established by analysis of 16S rRNAs. J Bacteriol. 1989 Jun;171(6):2970–2974. doi: 10.1128/jb.171.6.2970-2974.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  110. Wagner M., Amann R., Lemmer H., Schleifer K. H. Probing activated sludge with oligonucleotides specific for proteobacteria: inadequacy of culture-dependent methods for describing microbial community structure. Appl Environ Microbiol. 1993 May;59(5):1520–1525. doi: 10.1128/aem.59.5.1520-1525.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  111. Wagner M., Erhart R., Manz W., Amann R., Lemmer H., Wedi D., Schleifer K. H. Development of an rRNA-targeted oligonucleotide probe specific for the genus Acinetobacter and its application for in situ monitoring in activated sludge. Appl Environ Microbiol. 1994 Mar;60(3):792–800. doi: 10.1128/aem.60.3.792-800.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  112. Wallner G., Amann R., Beisker W. Optimizing fluorescent in situ hybridization with rRNA-targeted oligonucleotide probes for flow cytometric identification of microorganisms. Cytometry. 1993;14(2):136–143. doi: 10.1002/cyto.990140205. [DOI] [PubMed] [Google Scholar]
  113. Ward D. M., Weller R., Bateson M. M. 16S rRNA sequences reveal numerous uncultured microorganisms in a natural community. Nature. 1990 May 3;345(6270):63–65. doi: 10.1038/345063a0. [DOI] [PubMed] [Google Scholar]
  114. Weisburg W. G., Tully J. G., Rose D. L., Petzel J. P., Oyaizu H., Yang D., Mandelco L., Sechrest J., Lawrence T. G., Van Etten J. A phylogenetic analysis of the mycoplasmas: basis for their classification. J Bacteriol. 1989 Dec;171(12):6455–6467. doi: 10.1128/jb.171.12.6455-6467.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  115. Weller R., Ward D. M. Selective Recovery of 16S rRNA Sequences from Natural Microbial Communities in the Form of cDNA. Appl Environ Microbiol. 1989 Jul;55(7):1818–1822. doi: 10.1128/aem.55.7.1818-1822.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  116. Werren J. H., Hurst G. D., Zhang W., Breeuwer J. A., Stouthamer R., Majerus M. E. Rickettsial relative associated with male killing in the ladybird beetle (Adalia bipunctata). J Bacteriol. 1994 Jan;176(2):388–394. doi: 10.1128/jb.176.2.388-394.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  117. Wilson K. H., Blitchington R., Frothingham R., Wilson J. A. Phylogeny of the Whipple's-disease-associated bacterium. Lancet. 1991 Aug 24;338(8765):474–475. doi: 10.1016/0140-6736(91)90545-z. [DOI] [PubMed] [Google Scholar]
  118. Woese C. R. Bacterial evolution. Microbiol Rev. 1987 Jun;51(2):221–271. doi: 10.1128/mr.51.2.221-271.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  119. Woese C. R., Kandler O., Wheelis M. L. Towards a natural system of organisms: proposal for the domains Archaea, Bacteria, and Eucarya. Proc Natl Acad Sci U S A. 1990 Jun;87(12):4576–4579. doi: 10.1073/pnas.87.12.4576. [DOI] [PMC free article] [PubMed] [Google Scholar]
  120. Woese C. R., Maniloff J., Zablen L. B. Phylogenetic analysis of the mycoplasmas. Proc Natl Acad Sci U S A. 1980 Jan;77(1):494–498. doi: 10.1073/pnas.77.1.494. [DOI] [PMC free article] [PubMed] [Google Scholar]
  121. Woese C. R., Stackebrandt E., Ludwig W. What are mycoplasmas: the relationship of tempo and mode in bacterial evolution. J Mol Evol. 1984;21(4):305–316. doi: 10.1007/BF02115648. [DOI] [PubMed] [Google Scholar]
  122. Woese C. R., Stackebrandt E., Macke T. J., Fox G. E. A phylogenetic definition of the major eubacterial taxa. Syst Appl Microbiol. 1985;6:143–151. doi: 10.1016/s0723-2020(85)80047-3. [DOI] [PubMed] [Google Scholar]
  123. Yang D., Oyaizu Y., Oyaizu H., Olsen G. J., Woese C. R. Mitochondrial origins. Proc Natl Acad Sci U S A. 1985 Jul;82(13):4443–4447. doi: 10.1073/pnas.82.13.4443. [DOI] [PMC free article] [PubMed] [Google Scholar]
  124. Zarda B., Amann R., Wallner G., Schleifer K. H. Identification of single bacterial cells using digoxigenin-labelled, rRNA-targeted oligonucleotides. J Gen Microbiol. 1991 Dec;137(12):2823–2830. doi: 10.1099/00221287-137-12-2823. [DOI] [PubMed] [Google Scholar]

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