Chapter 9 : Culture-Independent Microbiology

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The field of cultivation-independent microbiology has rapidly advanced over the past few years. The technological developments briefly described in this chapter have brought us the opportunity to study the enormous complexity of natural microbial communities in more comprehensive and complete terms. Because the basis of most cultivation-independent approaches in microbiology (except whole-cell in situ hybridization or exogenous isolation of mobile genetic elements) is DNA or RNA that is extracted from environmental matrices, the chapter is devoted to nucleic extraction. Two principal approaches to recover nucleic acids from environmental matrices exist. Microbial community structures are studied using multiphasic approaches by combining various methods. Although the kinds of bacterial populations present in an environmental sample are still explored best by the cloning of 16S rDNA genes or other phylogenetic markers, the temporal and spatial distribution of ribotypes can be followed best by molecular fingerprints. Information on the localization of respective ribotypes and their metabolic activities can be provided by whole-cell in situ hybridization. In addition, reporter genes are a powerful tool to study how microbes perceive their surroundings and how their metabolic activity relates to their spatial distribution. High-density DNA arrays that will allow monitoring gene content and expression—although still a methodological challenge—will provide new insights into complex microbial communities by linking information on structure and function. The advances of genomics strongly affect one`s understanding of microbes. In the future, advanced protein detection methods will become more important in addition to gene arrays in cultivation-independent microbiology.

Citation: Smalla K. 2004. Culture-Independent Microbiology, p 88-99. In Bull A (ed), Microbial Diversity and Bioprospecting. ASM Press, Washington, DC. doi: 10.1128/9781555817770.ch9
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Figure 1

DGGE and TGGE analysis of specific taxa to dissect complex communities and to detect less-abundant tibotypes. Experimental approach for the analysis of patterns of Betaproteobacteria (❰β-proteobacteria in the figure): in the first PCR a forward primer specific for Betaproteobacteria is used in combination with a universal primer to amplify Betaproteobacteria 16S rDNA from community DNA. The amplicons are used in a second PCR with a G + C-clamped bacterial primer in combination with a universal primer.

Citation: Smalla K. 2004. Culture-Independent Microbiology, p 88-99. In Bull A (ed), Microbial Diversity and Bioprospecting. ASM Press, Washington, DC. doi: 10.1128/9781555817770.ch9
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Figure 2

Seasonal dynamics of bacterial communities in the potato rhizosphere as revealed by DGGE analysis of 16S rDNA fragments amplified from community DNA. Nontransgenic (light gray) and transgenic (black) T4-lysozyme expressing Désirée. The arrow indicates Serratia ficaria. (Reprinted from .)

Citation: Smalla K. 2004. Culture-Independent Microbiology, p 88-99. In Bull A (ed), Microbial Diversity and Bioprospecting. ASM Press, Washington, DC. doi: 10.1128/9781555817770.ch9
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Figure 3

A polyphasic approach is required for the analysis of microbial communities.

Citation: Smalla K. 2004. Culture-Independent Microbiology, p 88-99. In Bull A (ed), Microbial Diversity and Bioprospecting. ASM Press, Washington, DC. doi: 10.1128/9781555817770.ch9
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1. Alm, E. W.,, and D. A. Stahl. 2000. Critical factors influencing the recovery and integrity of rRNA extracted from environmental samples: use of an optimized protocol to measure depth-related biomass distribution in freshwater sediments. J. Microbiol. Methods 40: 153 162.
2. Aim, E. W.,, D. Zheng,, and L. Raskin. 2000. The presence of humic substances and DNA in RNA extracts affects hybridization results. Appl. Environ. Microbiol. 66: 4547 4554.
3. Amann, R. I.,, W. Ludwig,, and K.-H. Schleifer. 1995. Phylogenetic identification and in situ detection of individual microbial cells without cultivation . Microbial Rev. 59: 143 169.
4. Andersen, J. B.,, C. Sternberg,, L. K. Poulsen,, S. P. Bjorn,, M. Givskov,, and S. Molin. 1998. New unstable variants of green fluorescent protein for studies of transient gene expression in situ. Appl. Environ. Microbiol. 64: 2240 2246.
5. Bakken, L. R.,, and V. Lindahl,. 1995. Recovery of bacterial cells from soil, p. 13 27. In J. T. Trevors, and J. D. Elsas (ed.), Nucleic Acids in the Environment. Springer-Verlag, Berlin, Germany.
6. Bale, M. J.,, M. J. Day,, and J. C. Fry. 1988. Novel method for studying plasmid transfer in undisturbed river epilithon. Appl. Environ. Microbiol. 54: 2756 2758.
7. Boon, N.,, W. de Windt,, W. Verstraate,, and E. M. Top. 2002. Evaluation of nested PCR-DGGE (denaturing gradient gel electrophoresis) with group-specific 16S rRNA primers for the analysis of bacterial communities from different wastewater treatment plants. FEMS Microbiol. Ecol. 39: 101 112.
8. Borneman, J. 1999. Culture-independent identification of microorganisms that respond to specified stimuli. Appl. Environ. Microbiol. 65: 3398 3400.
9. Boschker, H. T. S.,, and J. J. Middelburg. 2002. Stable isotopes and biomarker in microbial ecology . FEMS Microbiol. Ecol. 1334: 1 12.
10. Boschker, H. T. S.,, S. C. Nold,, P. Wellsbury,, D. Bos,, W. de Graaf,, R. Pel,, R. J. Parkes,, and T. E. Cappenburg. 1998. Direct linking of microbial populations to specific biogeochemical processes by 13C-labelling of biomarkers. Nature 392: 801 804.
11. Christensen, B. B.,, C. Sternberg,, and S. Molin. 1996. Bacterial plasmid conjugation on semi-solid surfaces monitored with the green fluorescent protein (Gfp) from Aequorea victoria as a marker. Gene 173: 59 65.
12. Christensen, B. B.,, C. Sternberg,, J. B. Andersen,, L. Eberl,, S. Moeller,, M. Givskov,, and S. Molin. 1998. Establishment of new traits in a microbial biofilm community . Appl. Environ. Microbiol. 64: 2247 2255.
13. Dahlberg, C.,, M. Bergström,, and M. Hermansson. 1998. In situ detection of high levels of horizontal plasmid transfer in marine bacterial communities. Appl. Environ. Microbiol. 64: 2670 2675.
14. Dahllöf, I.,, H. Baillie,, and S. Kjelleberg. 2000. rpoB-based microbial community analysis avoids limitations inherent in 16S rRNA gene intraspecies heterogeneity. Appl. Environ. Microbiol. 66: 3376 3380.
15. Daims, H.,, J. L. Nielsen,, P. H. Nielsen,, K.-H. Schleifer,, and M. Wagner. 2001. In situ characterization of Nitrosospira-like nitrite-oxidizing bacteria active in wastewater treatment plants. Appl. Environ. Microbiol. 67: 5273 5284.
16. Duarte, G. F.,, A. S. Rosado,, L. Seldin,, A. C. Keijzer-Wolters,, and J. D. van Elsas. 1998. Extraction of ribosomal RNA and genomic DNA from soil for studying the diversity of the indigenous microbial community. J. Microbiol. Method. 32: 21 29.
17. Duineveld, B. M.,, G. A. Kowalchuk,, A. Keijzer,, J. D. van Elsas,, and J. A. van Veen. 2001. Analysis of bacterial communities in the rhizosphere of chrysanthemum via denaturing gradient gel electrophoresis of PCR-amplified 16S rRNA as well as DNA fragments coding for 16S rRNA . Appl. Environ. Microbiol. 67: 172 178.
18. Fægri, A.,, V. L. Torsvik,, and J. Goksøyr. 1977. Bacterial and fungal activities in soil: separation of bacteria and fungi by a rapid fractionated centrifugation technique. Soil Biol. Biochem. 9: 105 112.
19. Felske, A.,, A. Wolterink,, R. van Lis,, and A. D. L. Akkermans. 1998. Phylogeny of the main bacterial 16S rRNA sequences in Drentse A grassland soils (The Netherlands). Appl. Environ. Microbiol. 64: 871 879.
20. Fuchs, B. M.,, K. Syutsubo,, W. Ludwig,, and R. Amann. 2001. In situ accessibility of Escherichia coli 23S rRNA to fluorescently labeled probes . Appl. Environ. Microbiol. 67: 961 968.
21. Gieseke, A.,, U. Purkhold,, M. Wagner,, R. Amann,, and A. Schramm. 2001. Community structure and activity dynamics of nitrifying bacteria in a phosphate-removing biofilm. Appl. Environ. Microbiol. 67: 1351 1362.
22. Gillespie, D. E.,, S. F. Brady,, A. D. Bettermann,, N. P. Cianciotto,, M. R. Liles,, M. R. Rondon,, J. Clardy,, R. M. Goodman,, and J. Handelsman. 2002. Isolation of antibiotics turbomycin A and B from a metagenomic library of soil microbial DNA. Appl. Environ. Microbiol. 68: 4301 4306.
23. Giovannoni, S. J.,, T. B. Britschgi,, C. L. Mover,, and K. G. Field. 1990. Genetic diversity in Sargasso Sea bacterioplankton. Nature 345: 60 63.
24. Gomes, N. C. M.,, H. Heuer,, J. Schönfeld,, R. Costa,, L. Hagler-Mendonca,, and K. Smalla. 2001. Bacterial diversity of the rhizosphere of maize ( Zea mays) grown in tropical soil studied by temperature gradient gel electrophoresis. Plant Soil 232: 167 180.
25. Götz, A.,, R. Pukall,, E. Smit,, E. Tietze,, R. Prager,, H. Tschäpe,, J. D. van Elsas,, and K. Smalla. 1996. Detection and characterization of broad-host-range plasmids in environmental bacteria by PCR. Appl. Environ. Microbiol. 62: 2621 2628.
26. Griffiths, R. I.,, A. S. Whiteley,, A. G. O'Donnell,, and M. J. Bailey. 2000. Rapid method for coextraction of DNA and RNA from natural environments for analysis of ribosomal DNA- and rRNA-based microbial community composition. Appl. Environ. Microbiol. 66: 5488 5491.
27. Heuer, H.,, R. M. Kroppenstedt,, J. Lottmann,, G. Berg,, and K. Smalla. 2002. Effects of T4 lysozyme release from transgenic potato roots on bacterial rhizosphere communities are negligible relative to natural factors. Appl. Environ. Microbiol. 68: 1325 1335.
28. Heuer, H.,, M. Krsek,, P. Baker,, K. Smalla,, and E. M. H. Wellington. 1997. Analysis of actinomycete communities by specific amplification of genes encoding 16S rRNA and gel-electrophoretic separation in denaturing gradients. Appl. Environ. Microbiol. 63: 3233 3241.
29. Heuer, H.,, G. Wieland,, J. Schönfeld,, A. Schönwälder,, N. C. M. Gomes,, and K. Smalla,. 2001. Bacterial community profiling using DGGE or TGGE analysis, p. 177 190. In P. Rouchelle (ed.), Environmental Molecular Microbiology: Protocols and Applications. Horizon Scientific Press, Wymondham, United Kingdom.
30. Hill, K. E.,, A. J. Weightman,, and J. C. Fry. 1992. Isolation and screening of plasmids from the epilithon which mobilize recombinant plasmid pDlO. Appl. Environ. Microbiol. 58: 1292 1300.
31. Holben, W. E.,, and D. Harris. 1995. DNA-based monitoring of total bacterial community structure in environmental samples . Mol. Ecol. 4: 627 631.
32. Holben, W. E.,, J. K. Jansson,, B. K. Chelm,, and J. M. Tiedje. 1988. DNA probe method for the detection of specific microorganisms in the soil bacterial community. Appl. Environ. Microbiol. 54: 703 711.
33. Hopkins, D. W.,, S. J. Macnaughton,, and A. G. O'Donnell. 1991. A dispersion and differential centrifugation technique for representatively sampling microorganisms from soil. Soil Biol. Biochem. 23: 217 225.
34. Hugenholtz, P.,, B. M. Goebel,, and N. R. Pace. 1998. Impact of culture-independent studies on the emerging phylogenetic view of bacterial diversity. J. Bacteriol. 180: 4765 4774.
35. Hurt, R. A.,, X. Qiu,, L. Wu,, Y. Roh,, A. V. Palumbo,, J. M. Tiedje,, and J. Zhou. 2001. Simultaneous recovery of RNA and DNA from soils and sediments. Appl. Environ. Microbiol. 67: 4495 4503.
36. Klappenbach, J. A.,, J. M. Dunbar,, and T. M. Schmidt. 2000. rRNA operon copy number reflects ecological strategies of bacteria. Appl. Environ. Microbiol. 66: 1328 1333.
37. Koizumi, Y.,, J. J. Kelly,, T. Nakagawa,, H. Urakawa,, S. El-Fantroussi,, S. Al-muzaini,, M. Fukui,, Y. Urushigawa,, and D. Stahl. 2002. Parallel characterization of anaerobic toluene- and ethylbenzene-degrading microbial consortia by PCR-denaturing gradient gel electrophoresis, RNA-DNA membrane hybridiza- tion, and DNA microarray technology. Appl. Environ. Microbiol. 68: 3215 3225.
38. Lee, N.,, P. H. Nielsen,, P. H. Andreasen,, S. Juretschko,, J. L. Nielsen,, K.-H. Schleifer,, and M. Wagner. 1999. Combination of fluorescent in situ hybridization and microautoradiography—a new tool for structure-function analyses in microbial ecology. Appl. Environ. Microbiol. 65: 1289 1297.
39. Leveau, J. H. J.,, and S. E. Lindow. 2002. Bioreporters in microbial ecology. Curr. Opin. Microbol. 5: 259 265.
40. Liesack, W.,, and P. F. Dunfield,. 2002. Biodiversity in soils: use of molecular methods for its characterization, p. 528 544. In G. Bitton (ed.), Encyclopedia of Environmental Microbiology. Wiley & Sons Inc., New York, N. Y..
41. Liu, W.-T.,, T. L. Marsh,, H. Cheng,, and L. J. Forney. 1997. Characterization of microbial diversity by determining terminal restriction fragment length polymorphisms of genes encoding 16S rRNA. Appl. Environ. Microbiol. 63: 4516 4522.
42. Loy, A.,, A. Lehner,, N. Lee,, J. Adamczyk,, H. Meier,, J. Ernst,, K.-H. Schleifer,, and M. Wagner. 2002. Oligonucleotide microarray for 16S rRNA gene-based detection of all recognized lineages of sulfate-reducing prokaryotes in the environment. Appl. Environ. Microbiol. 68: 5064 5081.
43. Manefield, M.,, A. S. Whiteley,, R. I. Griffiths,, and M. J. Bailey. 2002. RNA stable isotope probing, a novel means of linking microbial community function to phylogeny. Appl. Environ. Microbiol. 68: 5367 5373.
44. McCaig, A. E.,, L. A. Glover,, and J. Prosser. 1999. Molecular analysis of bacterial community structure and diversity in unimproved and improved upland grass pastures. Appl. Environ. Microbiol. 65: 1721 1730.
45. Miller, D. N.,, J. E. Bryant,, E. L. Madsen,, and W. C. Ghiorse. 1999. Evaluation and optimization of DNA extraction and purification procedures for soil and sediment samples. Appl. Environ. Microbiol. 65: 4715 4724.
46. Molin, S.,, and M. Givskov. 1999. Application of molecular tools for in situ monitoring of bacterial growth activities . Environ. Microbiol. 1: 383 391.
47. Moré, M. I.,, J. B. Herrick,, M. C. Silva,, W. C. Ghiorse,, and E. L. Madsen. 1994. Quantitative cell lysis of indigenous microorganisms and rapid extraction of microbial DNA from sediment. Appl. Environ. Microbiol. 60: 1572 1580.
48. Muyzer, G.,, and K. Smalla. 1998. Application of denaturing gradient gel electrophoresis (DGGE) and temperature gradient gel electrophoresis (TGGE) in microbial ecology. Antonie Leeuwenhoek 73: 127 141.
49. Muyzer, G.,, E. C. de Waal,, and A. G. Uitterlinden. 1993. Profiling of complex microbial populations by denaturing gradient gel electrophoresis analysis of polymerase chain reaction-amplified genes encoding for 16S rRNA. Appl. Environ. Microbiol. 59: 695 700.
50. Nielsen, J. L.,, S. Juretschko,, M. Wagner,, and P. H. Nielsen. 2002. Abundance and phylogenetic affiliation of iron reducers in activated sludge as assessed by fluorescence in situ hybridization and microautoradiography. Appl. Environ. Microbiol. 68: 4629 4636.
51. Nogales, B.,, E. R. B. Moore,, E. Llobet-Brossa,, R. Rossello-Mora,, R. Amann,, and K. N. Timmis. 2001. Combined use of 16S ribosomal DNA and 16S rRNA to study the bacterial community of polychlorinated biphenyl-polluted soil. Appl. Environ. Microbiol. 67: 1874 1884.
52. Nogales, B.,, K. N. Timmis,, D. B. Nedwell,, and A. M. Osborn. 2002. Detection and diversity of expressed denitrification genes in estuarine sediments after reverse transcriptase-PCR amplification from mRNA. Appl. Environ. Microbiol. 68: 5017 5025.
53. Normander, B.,, N. B. Hendriksen,, and O. Nybroe. 1999. Green fluorescent protein-marked Pseudomonas fluorescens: localization, viability, and activity in the natural barley rhizosphere. Appl. Environ. Microbiol. 65: 4646 4651.
54. Nübel, U.,, B. Engelen,, A. Felske,, J. Snaidr,, A. Wiesenhuber,, R. I. Amann,, W. Ludwig,, and H. Backhaus. 1996. Sequence heterogeneities of genes encoding 16S rRNAs in Paenibacillus polymyxa detected by temperature gradient gel electrophoresis. J. Bacteriol. 178: 5636 5643.
55. Ogram, A.,, G. S. Sayler,, and T. J. Barkay. 1987. DNA extraction and purification from sediments. J. Microbiol. Methods 7: 57 66.
56. Oliver, J. D., 2000. Problems in detecting dormant (VBNC) cells and the role of DNA elements in this response, p. 1 15. In J. K. Jansson,, J. D. van Elsas,, and M. J. Bailey (ed.), Tracking Genetically-Engineered Microorganisms. Eurekah, Austin, Tex..
57. Osborn, A. M.,, E. R. B. Moore,, and K. N. Timmis. 2000. An evaluation of terminal-restriction fragment length polymorphism (T-RFLP) analysis for the study of microbial community structure and dynamics. Environ. Microbiol. 2: 39 50.
58. Pace, N. R. 1997. A molecular view of microbial diversity and the biosphere. Science 276: 734 740.
59. Pace, N. R.,, D. A. Stahl,, D. L. Lane,, and G. J. Olsen. 1986. The analysis of natural microbial populations by rRNA sequences. Adv. Microb. Ecol. 9: 1 55.
60. Radajewski, S.,, P. Ineson,, N. R. Parekh,, and J. C. Murrell. 2000. Stable-isotope probing as a tool in microbial ecology. Nature 403: 646 649.
61. Ramsing, N. B.,, M. Kühl,, and B. B. Jørgensen. 1993. Distribution of sulfate-reducing bacteria, O2, and H2S in photosynthetic biofilms determined by oligonucleotide probes and microelectrodes. Appl. Environ. Microbiol. 59: 3840 3849.
62. Rondon, M. R.,, P. R. August,, A. D. Bettermann,, S. F. Brady,, T. H. Grossman,, M. R. Liles,, K. A. Loiacono,, B. A. Lynch,, I. A. MacNeil,, M. S. Osburne,, J. Clardy,, J. Handelsman,, and R. M. Goodman. 2000. Cloning the soil metagenome: a strategy for accessing the genetic and functional diversity of uncultured microorganisms. Appl. Environ. Microbiol. 66: 2541 2547.
63. Roszak, D. B.,, and R. R. Colwell. 1987. Survival strategies of bacteria in the natural environment. Microbiol. Rev. 51: 365 379.
64. Schwieger, F.,, and C. C. Tebbe. 1998. A new approach to utilize PCR-single strand-conformation polymorphism for 16S rRNA gene-based microbial community analysis. Appl. Environ. Microbiol. 64: 4870 4876.
65. Schwieger, F.,, and C. C. Tebbe. 2000. Effect of field inoculation with Sinorhizobium meliloti L33 on the composition of bacterial communities in rhizospheres of a target plant ( Medicago saliva) and a non-target plant ( Chenopodium album)-linking of 16S rRNA gene-based single-strand conformation polymorphism community profiles to the diversity of cultivated bacteria. Appl. Environ. Microbiol. 66: 3556 3565.
66. Smalla, K.,, and P. Sobecky. 2002. The prevalence and diversity of mobile genetic elements in environmental bacteria assessed with new tools. FEMS Microbiol. Ecol. 42: 165 175.
67. Smalla, K.,, G. Wieland,, A. Buchner,, A. Zock,, J. Parzy,, S. Kaiser,, N. Roskot,, H. Heuer,, and G. Berg. 2001. Bulk and rhizosphere soil bacterial communities studied by denaturing gradient gel electrophoresis: plant-dependent enrichment and seasonal shifts revealed. Appl. Environ. Microbiol. 67: 4742 4751.
68. Staley, J. T.,, and A. Konopka. 1985. Measurement of in situ activities of nonphotosynthetic microorganisms in aquatic and terrestrial habitats. Annu. Rev. Microbiol. 39: 321 346.
69. Stoltzfus, J. R.,, J. K. Jansson,, and F. J. de Bruijn,. 2000. Using green fluorescent protein (GFP) as biomarker or bioreporter for bacteria, p. 101 116. In J. K. Jansson,, J. D. van Elsas,, and M. J. Bailey (ed.), Tracking Genetically-Engineered Microorganisms. Eurekah, Austin, Tex.
70. Tebbe, C. C.,, and W. Vahjen. 1993. Interference of humic acids and DNA extracted directly from soil in detection and transfor- mation of recombinant DNA from bacteria and a yeast. Appl. Environ. Microbiol. 59: 2657 2665.
71. Tebbe, C. C.,, A. Schmalenberger,, S. Peters,, and F. Schwieger,. 2001. Single-strand conformation polymorphism (SSCP) for microbial community analysis, p. 161 175. In P. Rouchelle (ed.), Environmental Molecular Microbiology: Protocols and Applications. Horizon Scientific Press, Wymondham, United Kingdom.
72. Top, E. M.,, W. E. Holben,, and L. J. Forney. 1995. Characterization of diverse 2,4-dichlorophenoxyacetic acid-degradative plasmids isolated from soil by complementation. Appl. Environ. Microbiol. 61: 1691 1698.
73. Torsvik, V. 1980. Isolation of bacterial DNA from soil. Soil Biol. Biochem. 12: 15 21.
74. Torsvik, V.,, J. Goksøyr,, and F. L. Daae. 1990. High diversity in DNA of soil bacteria. Appl. Environ. Microbiol. 56: 782 787.
75. Unge, A.,, and J. Jansson. 2001. Monitoring population size, activity, and distribution of gfp-luxAB-tagged Pseudomonas fluorescens SBW25 during colonization of wheat. Microbiol. Ecol. 41: 290 300.
76. Urbach, E.,, K. L. Vergin,, and S. J. Giovannoni. 1999. Immunochemical detection and isolation of DNA from metabolically active bacteria. Appl. Environ. Microbiol. 65: 1207 1213.
77. Van Elsas, J. D.,, K. Smalla,, and C. C. Tebbe,. 2000. Extraction and analysis of microbial community nucleic acids from environmental matrices, p. 29 51. In J. K. Jansson,, J. D. van Elsas,, and M. J. Bailey (ed.), Tracking Genetically-Engineered Microorganisms. Eurekah, Austin, Tex..
78. von Wintzingerode, F.,, U. B. Göbel,, and E. Stackebrandt. 1997. Determination of microbial diversity in environmental samples: pitfalls of PCR-based rRNA analysis. FEMS Microbiol. Rev. 21: 213 229.
79. Ward, D. M.,, M. M. Bateson,, R. Weller,, and A. L. Ruff-Roberts. 1990. 16S rRNA sequences reveal numerous microorganisms in a natural community. Nature 345: 63 65.
80. Weinbauer, M. G.,, I. Fritz,, D. F. Wenderoth,, and M. G. Höfle. 2002. Simultaneous extraction from bacterioplankton of total RNA and DNA suitable for quantitative structure and function analyses . Appl. Environ. Microbiol. 68: 1082 1087.
81. Woese, C. R. 1987. Bacterial evolution. Microbiol. Rev. 51: 221 271.
82. Wu, L.,, D. K. Thompson,, G. Li,, R. A. Hurt,, J. M. Tiedje,, and J. Zhou. 2001. Development and evaluation of functional gene arrays for detection of selected genes in the environment. Appl. Environ. Microbiol. 67: 5780 5790.
83. Yang, C.-H.,, and D. E. Crowley. 2000. Rhizosphere microbial community structure in relation to root location and plant iron nutritional status. Appl. Environ. Microbiol. 66: 345 351.
84. Yin, B.,, D. Crowley,, G. Sparovek,, W. J. de Melo,, and J. Borneman. 2000. Bacterial functional redundancy along a soil reclamation gradient. Appl. Environ. Microbiol. 66: 4361 4365.

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