Specific DNA Sequences for Detection of Soil Bacteria

  • X. Nesme
  • C. Picard
  • P. Simonet
Part of the Springer Lab Manuals book series (SLM)


Soils are colonized by numerous microorganisms reaching up to 1011 bacterial cells per g soil (Torsvik et al. 1990). They also can be considered as a reservoir for biodiversity which remains largely unknown, since only 10%–20% of the species living in soil have been isolated and characterized (Ward et al. 1990). This includes nitrogen-fixing bacteria such as Rhizobium, Frankia or Azospirillum, plant pathogens such as Agrobacterium and various fungi, and free-living bacteria involved in various biological processes. Concern has also been voiced about the fate of genetically engineered microorganisms (GMO) released in soils. Consequently, soil microbial populations as well as their in situ activities have remained largely unknown, indicating the need for new techniques which can provide detection tests with high levels of specificity and sensitivity regardless of physical and chemical characteristics of the environment.


Humic Acid Target Sequence Soil Bacterium Much Probable Number Crown Gall 
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. Araujo RS, Maya-Flores J, Barnes-McConnell D, Yokoyama C, Dazzo FB, Bliss FA (1986). Semienclosed tube cultures of bean plants (Phaseolus vulgaris L.) for enumeration of Rhizobium phaseoli by the Most-Probable-Number technique. Appl Environ Microbiol 52: 954–956Google Scholar
  2. Bej AK, Steffan RJ, diCesare J, Haff L, Atlas RM (1990) Detection of coliform bacteria in water by polymerase chain reaction and genes probes. Appl Environ Microbiol 56: 307–314Google Scholar
  3. Bej AK, Steffan RJ, diCesare J, Haff L, Atlas RM (1991) Detection of coliform bacteria and Escherichia coli and Shigella spp. in water by using the polymerase chain reaction and gene probes for ucd. Appl Environ Microbiol 57: 1013–1017Google Scholar
  4. Bollet C, Gevaudan MC, de Lamballerie X, Zandotti C, de Mieco P (1991) A simple method for the isolation of chromosomal DNA from gram-positive or acid-fast bacteria. Nucleic Acids Res 19: 1955Google Scholar
  5. Bosco M, Fernandez MP, Simonet P, Materassi R, Normand P (1992) Evidence that some Frankia sp. strains are able to cross boundaries between Alnus and Elaeagnus host specificity groups. Appl Environ Microbiol 58: 1569–1576Google Scholar
  6. Bouzard H, Ouadah D, Krimi Z, Jones JB, Travato M, Petit A, Dessaux Y (1993) Correlative association between resident plasmids and the host chromosome in a diverse Agrobacterium soil population. Appl Environ Microbiol 49: 1310–1317Google Scholar
  7. Bruce KD, Hiorns WD, Hobman JL, Osbom AM, Strike P, Ritchie DA (1992) Amplification of DNA from native populations of soil bacteria by using the polymerase chain reaction. Appl Environ Microbiol 58: 3413–3416Google Scholar
  8. Chen Y, Senesi N, Schnitzer M (1977) Information provided on humic substances by E4/E6 ratios. Soil Sci Soc Am J 41: 352CrossRefGoogle Scholar
  9. Cochran WG (1950) Estimation of bacterial densities by means of the most probable number. Biometrics 1950: 105–116CrossRefGoogle Scholar
  10. Crozat Y, Cleyet-Marel JC, Corman A (1987) Use of fluorescent antibody technique to characterize equilibrium survival concentrations of Bradyrhizobium japonicum strains in soil. Biol Fertil Soils 4: 85–90Google Scholar
  11. Deragon JM, Sinnett D, Mitchell G, Potier M, Labuda D (1990) Use of Gamma irradiation to eliminate DNA contamination for PCR. Nucleic Acids Res 18: 6149CrossRefGoogle Scholar
  12. Gilliand G, Perrin ST, Bunn HF (1990) Competitive PCR for quantitation of mRNA. In: Innis MA, Gelfand DH, Sninsky JJ, White IJ (eds). PCR protocols. A guide to methods and amplifications. Academic, San Diego, pp 21–27Google Scholar
  13. Golsteyn Thomas EJ, King RK, Burchak J, GannonVPJ (1991) Sensitive and specific detection of Listeria monocytogenes in milk and ground beef with the polymerase chain reaction. Appl Environ Microbiol 57: 2576–2580Google Scholar
  14. Grosjean MC, Picard C, Nesme X, Nazaret S, Cournoyer B, Paget E, Simonet P, Normand P, Bardin R (1991) L’amplification enzymatique de l’ADN: un outil d’étude en écologie microbienne. Phytoma 430: 34Google Scholar
  15. Higuchi R, Dolliger G, Walsh PS, Griffith R (1992) Simultaneous amplification and detection of specific DNA sequences. Biotechnology 10: 413–417CrossRefGoogle Scholar
  16. Hobbie JE, Daley RJ, Jasper S (1977) Use of Nucleopore filters for counting bacteria by fluorescence microscopy. Appl Environ Microbiol 54: 2859–2861Google Scholar
  17. Holben WE, Jansson JK, Chelm BK, Tiedje JM (1988) DNA probe method for the detection of specific microorganisms in the soil bacterial community. Appl Environ Microbiol 54: 703 - 711Google Scholar
  18. Horn GT, Richards B, Klinger KW (1989) Amplification of a highly polymorphic VNTR segment by the polymerase chain reaction. Nucleic Acids Res 16: 2140CrossRefGoogle Scholar
  19. Jacobsen CJ, Rasmussen OF (1992) Development and application of a new method to extract bacterial DNA from soil based on separation of bacteria from soil with cation-exchange resin. Appl Environ Microbiol 58: 2458–2462Google Scholar
  20. Jinno Y, Yoshiura K, Niikawa N (1990) Use of psoralen as extinguisher of contaminated DNA in PCR. Nucleic Acids Res 18: 6739CrossRefGoogle Scholar
  21. Kitchin PA, Szotyori Z, Fromholc C, Almond N (1990) Avoidance of false positives. Nature 344: 201CrossRefGoogle Scholar
  22. Liesack W, Ward N, Stackebrandt E (1991) Strategies for molecular microbial ecological studies. Actinomycete 2: 63–76Google Scholar
  23. Lorenz MG, Wackernagel W (1987) Adsorption of DNA to sand and variable degradation rate of adsorbed DNA. Appl Environ Microbiol 53: 2948–2952Google Scholar
  24. McGrady MH (1915) The numerical interpretation of fermentation-tube results. J Infect Dis 17: 183–212CrossRefGoogle Scholar
  25. Major JG (1992) A rapid PCR method of screening for small mutations. BioTechniques 12: 40–43Google Scholar
  26. Mullis KB, Faloona FA (1989) Specific synthesis of DNA in vitro via a polymerase catalysed chain reaction. Methods Enzymol 155: 335–350CrossRefGoogle Scholar
  27. Nannipieri P, Ciardi C, Badalucco L (1986) A method to determine soil DNA and RNA. Soil Biol Biochem 18: 275–281CrossRefGoogle Scholar
  28. Navarro E, Simonet P, Normand P, Bardin R (1992) Characterization of natural populations of Nitrobacter spp. using PCR/RFLP analysis of the ribosomal intergenie spacer. Arch Microbiol 157: 107–115Google Scholar
  29. Nazaret, S.,B. Cournoyer, P. Normand, and P. Simonet. 1991. Phylogenetic relationships among Frankia genomic species determined by use of amplified 16S rDNA sequences. J Bacteriol 173: 4072–4078Google Scholar
  30. Nesme X, Leclerc MC, Bardin R (1990) PCR detection of an original endosymbiont: the Ti plasmid of Agrobacterium tumefaciens. In: Nardon P, Gianinazzi V, Grenier AM, Margulis L, Smith DC (eds). Endocytobiology IV, INRA, Paris, pp 47–50Google Scholar
  31. Nick G, Paget E, Simonet P, Moiroud A, Normand P (1992) The nodular endophytes of Coriaria spp. from a distinct lineage within the genus Frankia. Mol Ecol 1: 175–181CrossRefGoogle Scholar
  32. Normand P, Cournoyer B, Nazaret S, Simonet P (1992) Analysis of a ribosomal RNA operon in the actinomycete Frankia. Gene 111: 119–124CrossRefGoogle Scholar
  33. Ogram A, Sayler GS, Barkay T (1987) The extraction and purification of microbial DNA from sediments. J Microb Methods 7: 57–66CrossRefGoogle Scholar
  34. Paget E, Jocteur Monrozier L, Simonet P (1992) Adsorption of DNA on clay minerals:protection against DNase I and influence on gene transfer. FEMS Microbiol Lett 97: 31–40CrossRefGoogle Scholar
  35. Picard C, Ponsonnet C, Paget E, Nesme X, Simonet P (1992) Detection and enumeration of bacteria in soil by direct DNA extraction and polymerase chain reaction. Appl Environ Microbiol 58: 2717–2722Google Scholar
  36. Picard C, Ponsonnet C, Recorbet G, Antonelli F, Simonet P, Nesme X (1994) Detection of pathogenic Agrobacterium in soils by PCR. Plant pathogenic bacteria. 8th International conference. Les Collogues, INRA-Editions, Paris, 66: 729–734Google Scholar
  37. Pillai SD, Josephson KL, Bailey RL, Gerba CP, Pepper IL (1991) Rapid method for processing soil samples for polymerase chain reaction amplification of specific gene sequences. Appl Environ Microbiol 57: 2283–2286Google Scholar
  38. Ponsonnet C, Nesme X (1994) Identification of Agrobacterium strains by PCR-RFLP analysis of pTi and chromosomal regions. Arch Microbiol 161: 300–309Google Scholar
  39. Postma J, Van Elsas JD, Govaert JM, Van Veen JA (1988) The dynamics of Rhizobium leguminosarum bv. trifolii introduced in soil as determinated by immunofluorescence and selective plating techniques. FEMS Microbiol Ecol 53: 251–260Google Scholar
  40. Richard GM (1974) Modifications of the diphenylamine reaction giving increased sensitivity and simplicity in the estimation of DNA. Anal Biochem 57: 369–376CrossRefGoogle Scholar
  41. Rochelle PA, Olson BH (1991) A simple technique for electroelution of DNA from environmental samples. Biotechniques 11: 724–728Google Scholar
  42. Ruano G, Fenton W, Kidd KK (1989) Biphasic amplification of very dilute DNA samples via “booster” PCR. Nucleic Acids Res 17: 5407CrossRefGoogle Scholar
  43. Sarkar G, Kapelner S, Sommer SS (1990) Formamide can dramatically improve the specificity of PCR. Nucleic Acids Res 18: 7465CrossRefGoogle Scholar
  44. Sarkar G, Sommer SS (1991) Parameters affecting susceptibility of PCR contamination to UV inactivation. BioTechniques 10: 591–593Google Scholar
  45. Selenska S, Klingmüller W (1991) DNA recovery and direct detection of Tn5 sequences from soil. Lett Appl Microbiol 13: 21–24CrossRefGoogle Scholar
  46. Sheng Zhu Y, Isaacs ST, Cimino GD, Hearst JE (1991) The use of exonuclease III for polymerase chain reaction sterilization. Nucleic Acids Res 19: 2511CrossRefGoogle Scholar
  47. Simonet P, Capellano A, Navarro E, Bardin R, Moiroud A (1984) An improved method for lysis of Frankia with achromopeptidase allows detection of new plasmids. Can J Microbiol 30: 1292–1295CrossRefGoogle Scholar
  48. Simonet P, Grosjean MC, Misra AK, Nazaret S, Cournoyer B, Normand P (1991) Frankia genus-specific characterization by polymerase chain reaction. Appl Environ Microbiol 57: 3278–3286Google Scholar
  49. Simonet P, Normand P, Moiroud A, Bardin R (1990) Identification of Frankia strains in nodules by hybridization of polymerase chain reaction products with strain-specific oligonucleotide probes. Arch Microbiol 153: 235–240CrossRefGoogle Scholar
  50. Smalla K, Cresswell N, Mendonca-Hagler LC, Wolters A, Van Elsas JD (1993) Rapid DNA extraction protocol from soil for polymerase chain reaction-mediated amplification. J Appl Bacteriol 674: 78–85Google Scholar
  51. Soltanpour PN, Jones JB, Workman SM (1982) Optical emission spectrometry. In Page AL, Miller RH, Keeney DR (eds) Methods of soil analysis. 2 Chemical and microbiological properties. Agronomy 9: 29–65Google Scholar
  52. Steffan RJ, Atlas RM (1988) DNA amplification to enhance detection of genetically engineered bacteria in environmental samples. Appl Environ Microbiol 54: 2185–2191Google Scholar
  53. Steffan RJ, Goksoyr J, Bej AK, Atlas RM (1988) Recovery of DNA from soils and sediments. Appl Environ Microbiol 54: 2908–2915Google Scholar
  54. Suggs SV, Hirose T, Miake T, Kawashima EH, Johnson MJ, Itakura K, Wallace RB (1981) ICN-UCLA Symp Mol Cell Biol 231: 683Google Scholar
  55. Tebbe CC, Vahjen W (1993) Interference of humic acids and DNA extracted directly from soil in detection and transformation of recombinant DNA from bacteria and a yeast. Appl Environ Microbiol 59: 2657–2665Google Scholar
  56. Torsvik VL (1980) Isolation of bacterial DNA from soil. Soil Biol Biochem 12: 15–21CrossRefGoogle Scholar
  57. Torsvik V, Goksoyr J, Daae FL (1990) High diversity in DNA of soil bacteria. Appl Environ Microbiol 56: 782–787Google Scholar
  58. Trevors JT (1992) DNA extraction from soil. Microb Releases 1: 3–9Google Scholar
  59. Tsai YL, Olson BJ (1991) Rapid method for direct extraction of DNA from soil and sediments. Appl Environ Microbiol 57: 1070–1074Google Scholar
  60. Tsai YL, Olson BH (1992) Rapid method for separation of bacterial DNA from humic substances in sediments for polymerase chain reaction. Appl Environ Microbiol 58: 2292–2295Google Scholar
  61. Ward DM. Weller R, Bateson MM (1990) 16S rRNA sequences reveal numerous uncultured microorganisms in a natural community. Nature 344: 63–65Google Scholar
  62. Weyant RS, Edmonds P, Swaminathan B (1990) Effect of ionic and nonionic detergents on the Taq polymerase. Biofeedback 9: 308–309Google Scholar
  63. Wu D.Y, Ugozzoli L, Pal BK, Qian J, Wallace RB (1991) DNA Cell Biol 10: 233–238CrossRefGoogle Scholar
  64. Yap EPH, McGee JOD (1991) Short PCR product yields improved by lower denaturation temperatures. Nucleic Acids Res 19: 1713CrossRefGoogle Scholar

Copyright information

© Springer-Verlag Berlin Heidelberg 1995

Authors and Affiliations

  • X. Nesme
  • C. Picard
  • P. Simonet

There are no affiliations available

Personalised recommendations