Identification and in Situ Detection of Intracellular Bacteria in the Environment

  • Bettina C. Brand
  • Rudolf I. Amann
  • Michael Steinert
  • Dorothee Grimm
  • Jörg Hacker
Part of the Subcellular Biochemistry book series (SCBI, volume 33)


Today it is generally accepted that our knowledge of bacterial diversity in the environment has been severely limited by the need to obtain pure cultures prior to characterization by testing for multiple physiological and biochemical properties. In addition, the morphology of microorganisms is in general too simple to serve as a basis for a reliable and proper classification; only in rare cases does it allow the in situ identification of individual population members by microscopy (Woese, 1987). Viable plate count or most probable-number techniques have been used for quantification of active cells in different environments but are always selective and can therefore not yield sufficient documentation of the true community structure (Table 1). For aquatic habitats as well as soils and sediments it has been frequently reported that direct microscopic counts exceed viable-cell counts by several orders of magnitude (Torsvik et al.,1990; Ferguson et al.,1984; Jones, 1977). This phenomenon is known as the “great plate count anomaly” described by Staley and Konopka (1985). Any estimation of the numbers of bacteria in the environment, whether they are pathogens, indicator organisms or genetically modified microorganisms, must allow for the fact that a proportion of the target organisms have entered the non-culturable but viable fraction of the microbial population. This accounts especially for bacterial endosymbionts colonizing free-living and parasitic protozoa although the roles such endosymbionts play in host survial, infectivity, and invasiveness are unclear (Fritsche et al., 1993).


Listeria Monocytogenes Bacterial Endosymbiont Nucleic Acid Probe Acanthamoeba Castellanii Paramecium Caudatum 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.


Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.


  1. Abu Kwaik, Y., Gao, L.Y., Harb, O.S., and Stone, B.J., 1997, Transcriptional regulation of the macrophage-induced gene (gspA) of Legionella pneumophila and phenotypic characterization of a null mutant, Mol. Microbiol. 24: 629–642.CrossRefGoogle Scholar
  2. Achi, R., Mata, L., Siles, X., and Lindberg, A.A., 1996, Immunomagnetic separation and detection show shigellae to be common faecal agents in children from urban marginal communities of Costa Rica, J. Infect. 32: 211–218.PubMedCrossRefGoogle Scholar
  3. Amann, R.I., 1995a, Fluorescently labeled, rRNA-targeted oligonucleotide probes in the study of microbial ecology, Mol. Ecot 4: 543–554.CrossRefGoogle Scholar
  4. Amann, R.I., 19956, In situ identification of microorganisms by whole cell hybridization with rRNA-targeted nucleic acid probes, in: Molecular Microbial Ecology Manual,(A.D.L. Akkerman, J.D. van Elsas, and F.J. de Bruijn, eds.), Kluwer Academic Publishers, Dordrecht, Netherlands, 3.3.6., p. 1–15.Google Scholar
  5. Amann, R.I., Binder, B.J., Olson, R.J., Chisholm, S.W., Devereux, R., and Stahl, D.A., 1990a. Combination of 16S rRNA-targeted oligonucleotide probes with flow cytometry for analyzing mixed microbial populations, Appl. Environ. Microbial. 56: 1919–1925.Google Scholar
  6. Amann, R.I., Krumholz, L., and Stahl, D.A., 1990b, Fluorescent-oligonucleotide probing of whole cells for determinative, phylogenetic, and environmental studies in microbiology, J. Bacterio1. 172: 762–770.Google Scholar
  7. Amann, R., Springer, N., Ludwig, W, GSrtz, H.-D., and Schleifer, K.-H., 1991, Identification in situ and phylogeny of uncultured bacterial endosymbionts, Nature (London) 351: 161–164.CrossRefGoogle Scholar
  8. Amann, R.I., Stromley, J., Devereux, R., Key, R., and Stahl, D.A., 1992a, Molecular and microscopic identification of sulfate-reducing bacteria in multispecies biofllms, Appl. Environ. Microbiol. 58: 614–623.PubMedGoogle Scholar
  9. Amann, R., Zarda, B., Stahl, D.A., and Schleifer, K.-H., 1992b, Identification of individual prokaryotic cells by using enzyme-labeled, rRNA-targeted oligonucleotide probes, Appt Environ. Microbiol. 58: 3007–3011.Google Scholar
  10. Amann, R., Ludwig, W, and Schleifer, K.-H., 1995, Phylogenetic identification and in situ detec-tion of individual microbial cells without cultivation, Microbiol. Rev. 59: 143–169.PubMedGoogle Scholar
  11. Amann, R., Snaidr, J., Wagner, M., Ludwig, W, and Schleifer, K.-H., 1996, In situ visualization of high genetic diversity in a natural microbial community, J. Bacteria 178: 3496–3500.Google Scholar
  12. Amann, R., Springer, N., Schönhuber, W., Ludwig, W., Schmidt, E.N., Müller, K.-D., and Michel, R., 1997, Obligate intracellular bacterial parasites of acanthamoebae related to Chlamydia spp., Appl. Environ. Microbiol. 63: 115–121.PubMedGoogle Scholar
  13. Arn_ heim, N., and Ehrlich, H.A., 1992, Polymerase chain reaction strategy, Annu. Rev. Biochem. 61: 131–169.PubMedCrossRefGoogle Scholar
  14. Aßmus, B., Hutzler, P., Kirchhof, G., Amann,R., Lawrence, J.R., and Hartmann, A., 1995, In situ localization of Azospirillum brasilense in the rhizosphere of wheat using fluorescently labeled, rRNA-targeted oligonucleotide probes and scanning confocal laser microscopy, Appl. Environ. Microbiol. 61: 1013–1019.Google Scholar
  15. Barbaree, J.M., Fields, B.S., Feeley, J.C., Gorman, G. W, and Martin, W.T., 1986, Isolation of protozoa from water associated with a legionellosis outbreak and demonstration of intracellular multiplication of Legionella pneumophila, Appl. Environ. Microbiol. 51: 422–424.PubMedGoogle Scholar
  16. Barker, J., and Brown, M.R.W., 1994, Trojan horses of the microbial world: protozoa and the survival of bacterial pathogens in the environment, Microbiol. 140: 1253–1259.CrossRefGoogle Scholar
  17. Barker, J., Scaife, H., and Brown, M.R.W., 1995, Intraphagocytic growth induces an antibiotic-resistant phenotype of Legionella pneumophila, Antimicrob. Agents Chemother. 39: 2684–2688.PubMedCrossRefGoogle Scholar
  18. Bermudez, L.E., 1994, Immunobiology of Mycobacterium avium infection, Eur. J. Clin. Microbiol. Infect. Dis. 13: 1000–1006.PubMedCrossRefGoogle Scholar
  19. Birtles, R.J., Rowbotham, T.J., Storey, C., Marrie, T.J., and Raoult, D., 1997, Chlamydia-like obligate parasite of free-living amoebae, Lancet 349: 925–926.PubMedCrossRefGoogle Scholar
  20. Brand, B.C., and Hacker, J., 1996, The biology of Legionella infection., in: Host response to intracellular pathogens, ( S.H.E. Kaufmann, ed.), R.G. Landes Company, Austin, pp. 291–312.Google Scholar
  21. Briglia, M., Eggen, R.I.L., DeVos, W.M., and Van Elsas, J.D., 1996, Rapid and sensitive method for the detection of Mycobacterium chlorophenolicum PCP-1 in soil based on 16S rRNA gene-targeted PCR, Appl. Environ. Microbiol. 62: 1478–1480.PubMedGoogle Scholar
  22. Centers for Disease Control and Prevention, 1993, Initial therapy for tuberculosis in the era of multidrug resistance, MMWR. 42 (RR-7): 1–8.Google Scholar
  23. Chantier, S., and McIllmurray, M.B., 1988, Labeled antibody methods for detection and identification of microorganisms, Methods in Microbiology 19: 273–332.CrossRefGoogle Scholar
  24. Cirillo, J.D., Falkow, S., and Thomkins, L.S.,1994, Growth of Legionella pneumophila in Acanthamoeba castellanii enhances invasion, Infect. Immun. 62: 3254–3261.Google Scholar
  25. Cirillo, J.D., Falkow, S., Tompkins, L.S., and Bermudez, L.E., 1997, Interaction of Mycobacterium avium with environmental amoebae enhances virulence, Infect. Immun. 65: 3759–3767.PubMedGoogle Scholar
  26. Crawford, J.T., and Bates, J.H., 1986, Analysis of plasmids in Mycobacterium aviumintracellulare isolates from persons with acquired immunodeficiency syndrome, Am. Resp. Dis. 134: 659–661.Google Scholar
  27. DeLong, E.E, Wickham, G.S., and Pace, N.R., 1989, Phylogenetic stains: ribosomal RNA-based probes for the identification of single microbial cells, Science 243: 1360–1363.PubMedCrossRefGoogle Scholar
  28. Devereux, R., Kane, M.D., Winfrey, J., and Stahl, D.A., 1992, Genus-and group-specific hybridization probes for determinative and environmental studies of sulfate-reducing bacteria, System. Appl. Microbiol. 15: 601–610.CrossRefGoogle Scholar
  29. Eckert, K.A., and Kunkel,T.A., 1991, DNA polymerase fidelity and the polymerase chain reaction, PCR Methods Appl. 1: 17–24.PubMedCrossRefGoogle Scholar
  30. Embley, T.M., Finlay, B.J., and Brown, S., 1992a, RNA sequence analysis shows that the symbionts in the ciliate Metopus contortus are polymorphs of a single methanogen species, FEMS Microbiol. Lett. 97: 57–62.CrossRefGoogle Scholar
  31. Embley, T.M., Finlay, B.J., Thomas, R.H., and Dyal, P.L., 1992b, 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. 138: 1479–1487.PubMedCrossRefGoogle Scholar
  32. Essig, A., Heinemann, M., Simnacher, U., and Marre, R., 1997, Infection of Acanthamoeba castellanii by Chlamydia pneumoniae, Appl. Environ. Microbiol. 63: 1396–1399.PubMedGoogle Scholar
  33. Falkinham, J.O., 1996, Epidemiology of infection by nontuberculous Mycobacteria, Clin. MicrobioL. Rev. 9: 177–215.Google Scholar
  34. Farber, J.M., and Peterkin, P.I., 1991. Listeria monocytogenes, a food-borne pathogen, Microbiol. Rev. 55: 476–511.Google Scholar
  35. Ferguson, R.L., Buckley, E.N., and Palumbo, A.V., 1984, Response of marine bacterioplankton to differential filtration an confinement, AppL Environ. MicrobioL 47: 49–55.Google Scholar
  36. Frank, J.F., Gillett, R.A.N., and Ware, G.O., 1990, Association of Listeria spp. contamination in the dairy processing plant environment with the presence of staphylococci, J. Food Prot. 53: 928–932.Google Scholar
  37. Fritsche, T.R., Gautom, R.K., Seyedirashti, S., Bergeron, D.L., and Lindquist, T.D. 1993, Occurence of bacterial endosymbionts in Acanthamoeba spp. isolated from corneal and environmental specimens and contact lenses, J. Clin. Microbiol. 31: 1122–1126.PubMedGoogle Scholar
  38. Gilbert, R.J., Miller, K.L., and Roberts, D., 1989, L. monocytogenes and chilled foods, Lancet 1: 383–384.PubMedCrossRefGoogle Scholar
  39. Glöckner, F.O.,Amann, R., Alfreider,A., Pernthaler, J., Psenner, R.,1rebesius, K., and Schleifer, K.-H., 1996, An optimized in situ hybridization protocol for planktonic bacteria, Syst. Appl. Microbiol. 19: 403–406.Google Scholar
  40. Grange, J.M., 1991, Environmental mycobacteria and human disease, Lepr. Rev. 62: 353–361.PubMedGoogle Scholar
  41. Grimm, D., Merkert, H., Ludwig, W, Schleifer, K.-H., Hacker, J., and Brand, B.C., 1998, Spe-cific detection of Legionella pneumophila: construction of a new 16S rRNA-targeted oligonuclotide probe, Appl. Environ. MicrobioL 64: 2686–2690.PubMedGoogle Scholar
  42. Gutell, R.R., Larsen, N., and Woese, C.R., 1994, Lessons from an evolving rRNA: 16S and 23S rRNA structures from a comparative perspective, Microbiol. Rev. 58: 10–26.PubMedGoogle Scholar
  43. Hahn, D., Amann, R.I., Ludwig, W., Akkermans, A.D.L., and Schleifer, K.-H., 1992, Detection of micro-organisms in soil after in situ hybridization with rRNA-targeted, fluorescently labelled oligonucleotides, J. Gen. Microbiol. 138: 879–887.PubMedCrossRefGoogle Scholar
  44. Holben, W.E., Jansson, J.K., Chelm, B.K., and Tiedje, J.M., 1988, DNA probe method for the detection of specific microorgnisms in the soil community, Appt Environ. Microbiol. 54: 703–711.Google Scholar
  45. Horn, M., Wagner, M., Fritsche, T., and Schleifer, K.-H., 1998, Phylogenetic studies on Acanthamoeba and nonculturable bacterial endosymbionts using 18S and 16S rDNA sequence analysis. VAAM, General Meeting, Frankfurt, Germany.Google Scholar
  46. Horwitz, M.A., and Silverstein, S.C., 1980, Legionnaires’ disease bacterium (Legionella pneu- mophila) multiplies intracellularly in human monocytes, J. Clin. Invest. 66: 441–450.PubMedCrossRefGoogle Scholar
  47. Hussong, D., Colwell, R.R., O’Brien, M., Weiss, E., Pearson, A.D., Weiner, R.M., and Burge, W.D., 1987, Viable Legionella pneumophila not detectable by culture on agar media, Bio/Technology 5: 947–950.CrossRefGoogle Scholar
  48. Islam, M.S., Hasan, M.K., Miah, M.A., Sur, G.C., Felsenstein, A., Venkatesan, M., Sack, R.B., and Albert, M.J., 1993, Use of the polymerase chain reaction and fluorescent-antibody methods for detecting viable but nonculturable Shigella dysenteriae type 1 in laboratory microcosms, Appt Environ. MicrobioL 59: 536–540.Google Scholar
  49. Jepras, R.I., Fitzgeorge, R.B., and Baskerville, A., 1985, A comparison of virulence of two strains of Legionella pneumophila based on experimental aerosol infection of guinea pigs, J. Hyg. 95: 29–38.PubMedCrossRefGoogle Scholar
  50. Jones, J.G., 1977, The effects of environmental factors on estimated viable and total populations of planktonic bacteria in lakes and experimental enclosures, Freshwater Biol. 7: 67–91.CrossRefGoogle Scholar
  51. Khan, M.U.; Curlin, G.T., and Huq, M.I., 1979, Epidemiology of Shigella dysenteriae type 1 infections in Dacca [sic] urban area, Trop. Geogr. Med. 31: 213–223.PubMedGoogle Scholar
  52. King, C.H., Shotts, E.B., Wooley, R.E., and Porter, K.G., 1988, Survival of coliforms and bacterial pathogens within protozoa during chlorination, Appl. Environ. Microbiol. 54: 3023–3033.PubMedGoogle Scholar
  53. King, W., Raposa, S., Warshaw, J., Johnson, A., Halbert, D., and Klinger, J.D., 1989, A new colorimetric nucleic acid hybridization assay for Listeria in foods, Int. J. Food Microbio!. 8: 225–232.CrossRefGoogle Scholar
  54. Kopczinsky, E.D., Bateson, M.M., and Ward, D.M., 1994, Recognition of chimeric small-subunit ribosomal DNAs composed of genes from uncultivated microorganisms, Appl. Environ. Microbiol. 60: 746–748.Google Scholar
  55. Kurtz, J.B., Bartlett, C.L.R., Newton, U.A., White, R.A., and Jones, N.L., 1982, Legionella pneumophila in cooling towers in London and a pilot trial of selected biocides, J. Hyg. 88: 369–381.Google Scholar
  56. Liesack, W., Weyland, H., and Stackebrandt, E., 1991, Potential risks of gene amplification by PCR as determined by 16S rDNA analysis of a mixed-culture of strict barophilic bacteria, Microb. Ecol. 21: 191–198.CrossRefGoogle Scholar
  57. Loessner, M.J., Rudolf, M., and Scherer, S., 1997, Evaluation of luciferase reporter bacteriophage A511::luxAB for detection of Listeria monocytogenes in contaminated foods, Appl. Environ. Microbiol. 63: 2961–2965.PubMedGoogle Scholar
  58. Ly, T.M.C., and Muller, H.E., 1990a, Interactions of Listeria monocytogenes, Listeria seeligeri and Listeria innocua with protozoans, J. Gen. Appl. Microbiol. 36: 143–150.CrossRefGoogle Scholar
  59. Ly, T.M.C., and Muller, H.E., 1990b. Ingested Listeria monocytogenes survive and multiply in protozoa, J. Med. Microbiol. 33: 51–54.PubMedCrossRefGoogle Scholar
  60. Maidak, B.L., Olsen, G.J., Larsen, N., Overbeek,R., McCaughey, M.J., and Woese, C.R., 1997, The RDP (Ribosomal Database Project), Nucleic Acids Res. 25: 109–110.PubMedCrossRefGoogle Scholar
  61. Manz, W., Amann, R., Ludwig, W., Wagner, M., and Schleifer, K.-H., 1992, Phylogenetic oligodeoxynucleotide probes for the major subclasses of proteobacteria: problems and solutions, System. Appl. Microbiol. 15: 593–600.CrossRefGoogle Scholar
  62. Manz, W., Szewzyk, U., Eriksson, P., Amann, R., Schleifer, K.H., and Stenström, T.-A., 1993, 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. 59: 2293–2298.Google Scholar
  63. Manz, W, Szewzyk, R., Szewzyk, U., Hutzler, P., Amann, R., and Schleifer, K.H., 1995, In situ identification of Legionellaceae using specific rRNA-targeted oligonucleotide probes and confocal laser scanning microscopy, Microbio. 141: 29–39.Google Scholar
  64. Menard, R., Dehio, C., and Sansonetti, P, 1996, Bacterial entry into epithelial cells: the paradigm of Shigella, Trends Microbiol. 4: 220–226.PubMedCrossRefGoogle Scholar
  65. Michel, R., Hauröder-Philippczyk, B., Müller, K.-D., and Weishaar, I., 1994, Acanthamoeba from human nasal mucosa infected with an obligate intracellular parasite, Eur. J. Parasitol. 30: 104–110.Google Scholar
  66. Neef, A., Zaglauer, A., Meier, H., Amann, R.; Lemmer, H., and Schleifer, K.H., 1996, Popula-tion analysis in a denitrifying sand filter: conventional and in situ identification of Para-coccus sp. in methanol-fed biofilms, Appl. Environ. Microbiol. 62: 4329–4339.PubMedGoogle Scholar
  67. Olsen, G.J., Lane, D.J., Giovannoni, S.J., Pace, N.R., and Stahl, D.A., 1986, Microbial ecology and evolution: a ribosomal RNA approach, Annu. Rev. Microbiol. 40: 337–365.PubMedCrossRefGoogle Scholar
  68. Ouverney, C.C., and Fuhrman, J.A., 1997, Increase in fluorescence intensity of 16S rRNA in situ hybridization in natural samples treated with chloramphenicol, Appl. Environ. Microbiol. 63: 2735–2740.PubMedGoogle Scholar
  69. Paszko-Kolva, C., Shahamat, M., and Colwell, R.R., 1992, Longterm survival of Legionella pneumophila serogroup 1 under low-nutrient conditions and associated morphological changes, FEMS MicrobioL EcoL 102: 45–55.CrossRefGoogle Scholar
  70. Peel, M., Donachle, W, and Shaw, A., 1988, Temperature-dependent expression of flagella of Listeria monocytogenes studied by electron microscopy, SDS-PAGE and western blotting, J. Gen. Microbiol. 143: 2171–2178.Google Scholar
  71. Peters, M., Muller, C., Rush-Gerdes, S., Seidel, C., Gobel, U., Pohle, H.D., and Ruf, B., 1995, Isolation of atypical mycobacteria from tap water in hospitals and homes: is this a possible source of disseminated MAC infection in AIDS patients?, J. Inf. 31: 39–40.CrossRefGoogle Scholar
  72. Pillay, D.G., Karas, A.J., and Sturm, A.W., 1997, An outbreak of Shiga bacillus dysentery in KwaZulu/Natal, South Africa, J. Infect. 34: 107–111.PubMedCrossRefGoogle Scholar
  73. Poulsen, L.K., Ballard, G., and Stahl, D.A., 1993, Use of rRNA fluorescence in situ hybridization for measuring the activity of single cells in young and established biofilms, AppL Environ. Microbiol. 59: 1354–1360.PubMedGoogle Scholar
  74. Rahman, I., Shahamat, M., Chowdhury, M.A.R., and Colwell, R.R., 1996, Potential virulence of viable but nonculturable Shigella dysenteriae type 1, Appl. Environ. MicrobioL 62: 115–120.PubMedGoogle Scholar
  75. Ramsing, N.B., Kühl, M., and Jorgensen, B.B., 1993, Distribution of sulfate-reducing bacteria, 02 and H2S in photosynthetic biofilms determined by oligonucleotide probes and micro-electrodes, Appl. Environ. Microbiol. 59: 3820–3849.Google Scholar
  76. Ren, T., and Frank, J.F., 1993, Susceptibility of starved planktonic and biofitm Listeria monocytogenes to quaternary ammonium sanitizer as determined by direct viable and agar plate counts, J. Food Prot. 56: 573–576.Google Scholar
  77. Rogers, J., and Keevil, C.W.,1992, Immunogold and fluorescein immunolabelling of Legionella pneumophila within an aquatic biofitm visualized by using episcopic differential interference contrast microscopy, AppL Environ. Microbio!. 58: 2326–2330.Google Scholar
  78. Roller, C., Wagner, M., Amann, R., Ludwig, W., and Schleifer, K.-H., 1994. In situ probing of gram-positive bacteria with high DNA G+C content using 23S rRNA-targeted oligonucleotides, Microbio. 140: 2849–2858.Google Scholar
  79. Rowe, B., and Gross, R.J., 1984, Shigella, in: Bergey’s manual of systematic bacteriology, ( N.R. Krieg, G. Holt, eds.) Williams and Wilkins, Baltimore, pp. 423–427.Google Scholar
  80. Rudi, K., Kroken, M., Dahlberg, O.J., Deggerdal, A., Jakobsen, K.S., and Larsen, F, 1997, Rapid, universal method to isolate PCR-ready DNA using magnetic beads, BioTechniques 22: 506–511.PubMedGoogle Scholar
  81. Ruf, B., Schürmann, D., Horbach, I., Fehrenbach, F.J., and Pohle, H.D., 1990, Prevalence and diagnosis of Legionella pneumonia: a 3-year prospective study with emphasis on application of urinary antigen detection, J. Infect. Dis. 162: 1341–1348.PubMedCrossRefGoogle Scholar
  82. Salfinger, M., and Pfyffer, G.E., 1994, The new diagnostic Mycobacteriology Laboratory, Eur. J. Clin. MicrobioL Infect. Dis. 13: 961–979.PubMedCrossRefGoogle Scholar
  83. Sansonetti, P., 1992, Molecular and cellular biology of Shigella flexneri invasiveness: from cell assay systems to shigellosis, Curr. Top. Microbiol. Immunol. 180: 1–19.Google Scholar
  84. Saylers, A.A., and Whitt, D.D., 1994, Bacterial pathogenesis, ASM, Washington, D.C., pp. 169–181.Google Scholar
  85. Schönhuber, W, Fuchs, B., Juretschko, S., and Amann, R., 1997, Improved sensitivity of whole cell hybridization by the combination of horseradish peroxidase-labeled oligonucleotides and tyramide signal amplification, Appl. Environ. Microbio!. 63: 3268–3273.Google Scholar
  86. Schramm, A., Larsen, L.H., Revsbech, N.P., Ramsing, N.B., Amann, R., and Schleifer, K.-H., 1996, Structure and function of a nitrifying biofitm as determined by in situ hybridization and microelectrodes, Appl. Environ. Microbio!. 62: 4641–4647.Google Scholar
  87. Schuchat, A., Swaminathan, B., and Broome, C.V., 1991, Epidemiology of human listeriosis, Clin. Microbiol. Rev. 4: 169–183.PubMedGoogle Scholar
  88. Seeliger, H.P.R., and Jones, D., 1986, Listeria, in: Bergey’s Manual of Systematic Bacteriology, 2: 1235–1245.Google Scholar
  89. Somerville, C., Knight, I.T., Straube, W.L., and Colwell, R.R., 1989, Simple rapid method for the direct isolation of nucleic acids from aquatic environments, Appl. Environ. Microbiol. 55: 548–554.PubMedGoogle Scholar
  90. Springer, N., Ludwig, W, Drozanski, V., Amann, R., and Schleifer, K.-H., 1992, The phylogenetic status of Sarcobium lyticum, an obligate intracellular bacterial parasite of small amoebae, FEMS Microbiol. Leu. 96: 199–202.CrossRefGoogle Scholar
  91. Springer, N., Ludwig, W., Amann, R., Schmidt, H.J., Görtz, H.-D., and Schleifer, K.-H., 1993, Occurrence of fragmented 16S rRNA in an obligate bacterial endosymbiont of Paramecium caudatum, Proc. Natl. Acad. Sci. USA 90: 9892–9895.PubMedCrossRefGoogle Scholar
  92. Springer, N., Amann, R., Ludwig, W, Schleifer, K.-H., and Schmidt, H., 1996, Polynucleobacter necessarius, an obligate bacterial endosymbiont of the hypotrichous ciliate Euplotes aediculatus, is a member of the 0-subclass of Proteobacteria, FEMS Microb. Lett. 135: 333–336.Google Scholar
  93. Stahl, D.A., and Amann, R.I., 1991, Development and application of nucleic acid probes in bacterial systematics, in: Sequencing and Hybridization Techniques in Bacterial Systematics, ( E. Stackebrandt and M. Goodfellow, eds.), John Wiley and Sons, Chichester, England, pp. 205–248.Google Scholar
  94. Stahl, D.A., Flesher, B., Mansfield, H.R., Montgomery, L., 1988, The use of phylogenetically based hybridization probes for studies of ruminai microbial ecology, Appl. Environ. Microbiol. 54: 1079–1084.Google Scholar
  95. Staley, J.T., and Konopka, A., 1985, Measurement of in situ activities of nonphotosynthetic microorganisms in aquatic and terrestrial habitats, Annu. Rev. Microbiol. 39: 32–346.CrossRefGoogle Scholar
  96. Steinert, M., Emödy, L., Amann, R., and Hacker, J., 1997, Resuscitation of viable but nonculturable Legionella pneumophila Philadelphia JR32 by Acanthamoeba castellanii, Appl. Environ. Microbiol. 63: 2047–2053.Google Scholar
  97. Steinert, M., Birkness, K., White, E., Fields, B., and Quinn, F., 1998a, Mycobacterium avium bacilli grow saprozoically in coculture with Acanthamoeba polyphaga and survive within the cyst wall, Appl. Environ.Microbiol. 64: 2256–2261.Google Scholar
  98. Steinert, M., Birkness, K., White, E., Quinn, F., and Fields, B., 1998b, Survival of bacterial pathogens within Acanthamoeba polyphaga, 98th ASM General Meeting, Atlanta, Abstr. N49, p. 374.Google Scholar
  99. Strunk, O., Gross, O., Reichel, B., May, M., Hermann, S., Stuckman, N., Nonhoff, B., Lenke, M.,Ginhart, A., Vilbig, A., Ludwig, T., Bode, A., Schleifer, K.-H., and Ludwig, W, 1998, ARB: a software environment for sequence data, http://www.mikro.biologie. Scholar
  100. Szewzyk, U., Manz, W, Amann, R., Schleifer, K.-H., and Stenström, T.-A., 1994, Growth and in situ detection of a pathogenic Escherichia coli in biofilms of a heterotrophic water bacterium by use of 16S- and 23S-rRNA-directed fluorescent oligonucleotide probes, FEMS Microbiol. Ecol. 13: 169–175.Google Scholar
  101. Torsvik V., Goksoyr, J., and Daae, F.L., 1990, High diversity of DNA of soil bacteria, Appl. Environ. Microbiol. 56: 782–787.PubMedGoogle Scholar
  102. Van de Peer, Y., Jansen, J., De Rijk, P., and De Wachter, R., 1997, Database on the structure of small ribosomal subunit RNA, Nucleic Acids Res. 25: 111–116.PubMedCrossRefGoogle Scholar
  103. Wagner, M., Amann, R., Lemmer, H., and Schleifer, K.-H., 1993, Probing activated sludge with proteobacteria-specific oligonucleotides: inadequacy of culture-dependent methods for describing microbial community structure, Appl. Environ. Microbiol. 59: 1520–1525.PubMedGoogle Scholar
  104. Wagner, M., Schmid, M., Juretschko, S., Trebesius, K.-H., Bubert, A., Goebel, W., and Schleifer, K.-H., 1998, In situ detection of a virulence factor mRNA and 16S rRNA in Listeria monocytogenes, FEMS Microbiol. Lett. 160: 159–168.Google Scholar
  105. Wallner, G., Amann, R., and Beisker, W, 1993, Optimizing fluorescent in situ hybridization of suspended cells with rRNA-targeted oligonucleotide probes for the flow cytometric identification of microorganisms, Cytometry 14: 136–143.PubMedCrossRefGoogle Scholar
  106. Wang, G.C.Y., and Wang, Y., 1996, The frequency of chimeric molecules as a consequence of PCR co-amplification of 16S rRNA genes from different bacterial species, Microbiology 142: 1107–1114.PubMedCrossRefGoogle Scholar
  107. Wayne, L.G., 1994, Dormancy of Mycobacterium tuberculosis and latency of disease, Eur. J. Clin. Microbiol. Infect. Dis. 13: 908–914.PubMedCrossRefGoogle Scholar
  108. Wayne, L.G., and Kubica, G.P., 1986, Mycobacteriae, in: Bergey ‘s Manual of Systematic Bacteriology, 2: 1436–1457.Google Scholar
  109. Wintermeyer, E., Ludwig, B., Steinert, M., Schmitt, B., Fischer,G., and Hacker, J., 1995, Influence of site-specific altered Mip proteins on intracellular survival of Legionella pneumophila in eukaryotic cells, Infect. Immun. 63: 4576–4583.Google Scholar
  110. Woese, C.R., 1987, Bacterial evolution, Microbiol. Rev. 51: 221–271.PubMedGoogle Scholar
  111. Yan, W, Malik, M.N., Peterkin, P.I., and Sharpe, A.N., 1996, Comparison of the hydrophobic grid-membrane filter DNA probe method and the Health Protection Branch standard method for the detection of Listeria monocytogenes in foods, Int. J. Food Microbiol. 30: 379–84.PubMedCrossRefGoogle Scholar
  112. Zarda, B., Amann, R., Wallner,G., and Schleifer, K.-H., 1991, Identification of single bacterial cells using digoxigenin-labeled, rRNA-targeted oligonucleotides, J. Gen. Microbiol. 137: 2823–2830.Google Scholar

Copyright information

© Springer Science+Business Media New York 2000

Authors and Affiliations

  • Bettina C. Brand
    • 1
  • Rudolf I. Amann
    • 2
  • Michael Steinert
    • 1
  • Dorothee Grimm
    • 1
  • Jörg Hacker
    • 1
  1. 1.Institute for Molecular Biology of Infectious DiseasesUniversity of WürzburgWürzburgGermany
  2. 2.Dept. Molecular EcologyMax-Planck-Institute for Marine MicrobiologyBremenGermany

Personalised recommendations