Skip to main content
SpringerLink
Log in

Problems and promises of assaying the genetic potential for nitrogen fixation in the marine environment

  • Published:
Microbial Ecology Aims and scope Submit manuscript

Abstract

Nitrogen fixation in the sea has attracted the attention of ecologists for decades. Much is known about the habitats in which it occurs and some of the factors that limit N2 fixation activity in different environments, but we still know little about the organisms that fix nitrogen, and what limits the growth and distribution of these organisms in marine environments. Molecular biology technological developments have provided tools for detecting and characterizing N2-fixing organisms in the environment. These techniques hold great promise for unraveling the mysteries and paradoxes of N2 fixation in the sea. In this review, we address the theoretical basis for the use of a molecular approach to N2 fixation, highlight the strengths and weaknesses of the approach, and provide case studies that demonstrate the potential contribution of molecular biology approaches to studies of N2 fixation in the sea.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Similar content being viewed by others

References

  1. Altabet MA, Francois R (1994) Sedimentary nitrogen isotopic ratio as a recorder of surface ocean nitrate utilization. Global Biogeochem Cycles 8:103–106

    Google Scholar 

  2. Altabet MA, Francois R, Murray DW Prell WL (1995) Climate-related variations in denitrification in denitrification in the abian Sea from sediment N-15/N-14 ratios. Nature 373:506–509

    Google Scholar 

  3. Amann RI, Ludwig W Schleifer KH (1995) Phylogenetic identification and in situ detection of individual microbial cells without cultivation. Microbiol Revs 59:143–169

    Google Scholar 

  4. Bebout BM, Fitzpatrick MW Paerl HW (1993) Identification of the sources of energy for N2 fixation and physiological characterization of nitrogen-fixing members of a marine microbial mat community. Appl Environ Microbiol 59:1495–1503

    Google Scholar 

  5. Bej AK, Mahbubani MH (1992) Applications of the polymerase chain reaction in environmental microbiology. PCR Meth Appl 1:151–159

    Google Scholar 

  6. Ben-Porath J, Zehr JP (1994) Detection and characterization of cyanobacterial nifH genes. Appl Environ Microbiol 60:880–887

    Google Scholar 

  7. Ben-Porath J, Carpenter EJ, Zehr JP (1993) Genotypic relationships in Trichodesmium (cyanophyceae) based on nifH sequence comparisons. J Phycol 29:806–810

    Google Scholar 

  8. Berger WH, Herguera JC (1992) Primary productivity and biogeochemical cycles in the sea. In: Falkowski PG, Woodhead AD (eds) Reading the sedimentary record of the ocean's productivity. Plenum Press, New York, pp 455–486

    Google Scholar 

  9. Berger WH, Smaetacek VS, Wefer G (1989) Productivity of the oceans: present and past. In: Berger WH, Smaetacek VS, Wefer G (eds) Ocean productivity and paleoproductivity—an overview. J. Wiley, Chichester, pp 1–34

    Google Scholar 

  10. Berner W Oeschger H, Stauffer B (1980) Information on the CO2 cycle from ice core studies. Radiocarbon 22:227–269

    Google Scholar 

  11. Bishop PE, Premakumar R (1992) Alternative N2 fixation systems. In: Stacey G, Burris RH, Evans HR (eds) Biological N2 fixation. Chapman and Hall, New York, pp 736–762

    Google Scholar 

  12. Bishop PA, MacDouglas SI, Wolfinger ED, Shermer CL (1990) N2 fixation: achievements and objectives. In: Gresshoff PM, Roth LE, Stacey G, Newton WE (eds) Genetics of alternative N2 fixation systems in Azotobacter vinelandii. Chapman and Hall, New York, pp 789–795

    Google Scholar 

  13. Broecker WS, Peng TH (1986) Carbon cycle: 1985. Glacial to interglacial changes in the operation of the global carbon cycle. Radicarbon 28:309–327

    Google Scholar 

  14. Burckle LH, Cirilli J (1987) Origin of diatom ooze belt in the Southern Ocean: implications for late quaternary paleooceanograph. Micropaleont 33:86–86

    Google Scholar 

  15. Cano RJ, Poinar H, Pieniazek N, Acra A, Poinar Jr, GO (1993) Amplification and sequencing of DNA from a 120–135 million-year-old weevil. Nature 363:536–539

    Google Scholar 

  16. Capone DG (1983) Benthic N2 fixation. In: Carpenter EJ, Capone DG (eds) Nitrogen in the marine environment. Academic Press, New York, pp 105–137

    Google Scholar 

  17. Capone DG (1983) N2 fixation in seagrass communities. Mar Tech Soc J 17:32–37

    Google Scholar 

  18. Capone DG (1988) Benthic N2 fixation. In: Blackburn TH, Sorensen J (eds) Nitrogen cycling in coastal marine environments. John Wiley and Sons, pp 85–123

  19. Capone DG, Budin J (1982) N2 fixation associated with rinsed roots and rhizomes of the eelgrass, Zostera marina. Plant Physiol 70:1601–1604

    Google Scholar 

  20. Capone DG, Carpenter EJ (1982) N2 fixation in the marine environment. Science 217:1140–1142

    Google Scholar 

  21. Capone DG, Carpenter EJ (1982) A perfusion method for assaying microbial activities in estuarine sediments: Applicability to studies of N2 fixation by C2H2 reduction. Appl Environ Microbiol 43:1400–1405

    Google Scholar 

  22. Capone DG, O'Neil JM, Zehr J, Carpenter EJ (1990) Basis for diel variation in nitrogenase activity in the marine planktonic cyanobacterium Trichodesmium thiebautii. Appl Environ Microbiol 56:3532–3536

    Google Scholar 

  23. Carpenter EJ (1982) Physiology and ecology of marine planktonic Oscillatoria (Trichodesmium). Mar Biol Lett 4:69–85

    Google Scholar 

  24. Carpenter EJ, Romans K (1991) Major role of the cyanobacterium Trichodesmium in nutrient cycling in the North Atlantic Ocean. Science 254:1356–1358.

    Google Scholar 

  25. Chisholm FP, Frankel SL, Goericke R, Olson RJ, Palenik B, Waterbury, JB, West-Johnsrud L, Zettler ER (1992) Prochlorococcus marinus nov. gen. nov. sp.: an oxyphototrophic marine prokaryote containing divinylchlorophyll a and b. Arch Microbiol 157:297–300

    Google Scholar 

  26. 26.Codispotti L (1989) Phosphorus vs. nitrogen limitation of new and export production. In: Berger WH, Smetacek VS, Wefer G (eds) Productivity of the ocean: present and past. S Bernhard, Dahlem Konferenzen, pp 377–394

    Google Scholar 

  27. Currin CA, Paerl HW, Suba GK (1990) Immunofluorescence detection and characterization of N2-fixing microorganisms from aquatic environments. Limnol Oceanogr 35:59–71

    Google Scholar 

  28. Delwiche CC (1981) The nitrogen cycle and nitrous oxide. In: Delwiche CC (ed) Denitrification, nitrification, and atmospheric nitrous oxide. Wiley & Sons, New York, pp 1–15

    Google Scholar 

  29. Dugdale RC (1967) Nutrient limitation in the sea: Dynamics, identification and significance. Limnol Oceanogr 12:685–695

    Google Scholar 

  30. Fanning KA (1992) Nutrient provinces in the sea: concentration ratios, reaction rate ratios, and ideal covariation. J Geophysical Res 97:5693–5712

    Google Scholar 

  31. Felsenstein J (1993) PHYLIP-Phylogeny Inference Package (Version 3.5c). Distributed by the author. Department of Genetics, University of Washington, Seattle

    Google Scholar 

  32. Francois R, Altabet MA (1992) Glacial to interglacial changes in surface nitrate utilization in the Indian sector of the Southern Ocean as recorded by sediment d15N. Paleoceanography 7:589–606

    Google Scholar 

  33. Fujita Y, Takahashi Y, Chuganji M, Matsubara H (1992) The nifH-like (frxC) gene is involved in the biosynthesis of chlorophyll in the filamentous cyanobacterium Plectonema boryanum. Plant Cell Physiol 33:81–92

    Google Scholar 

  34. Georgiadis MM, Komiya H, Chakrabarti P, Woo D, Kornuc JJ, Rees DC (1992) Crystallographic structure of the nitrogenase iron protein from Azotobacter vinelandii. Science 257:1653–1659

    Google Scholar 

  35. Giovannoni SJ, Britschgi TB, Moyer CL, Field KG (1990) Genetic diversity in Sargasso Sea bacterioplankton. Nature 345:60–63

    Article  CAS  PubMed  Google Scholar 

  36. Gordon JK, Brill WJ (1974) Depression of nitrogenase synthesis in the presence of excess NH4 +. Biochem Biophys Res Comm 59:967–971

    Google Scholar 

  37. Guerinot ML, Colwell RR (1985) Enumeration, isolation, and characterization of N2-fixing bacteria from seawater. Appl Environ Microbiol 50:350–355

    Google Scholar 

  38. Hartwig EO, Stanley SO (1978) N2 fixation in Atlantic deep-sea and coastal sediments. Deep-Sea Res 25:411–417

    Google Scholar 

  39. Haselkorn R, Buikema WJ (1992) N2 fixation in cyanobacteria. In: Stacey G, Burns RH, Evans HJ (eds) Biological N2 fixation. Chapman and Hall, New York, pp 166–190

    Google Scholar 

  40. Hecky RE, Kilham R (1988) Nutrient limitation of phytoplankton in freshwater and marine environments: a review of recent evidence on the effects of enrichment. Limnol Oceanogr 33:796–822

    Google Scholar 

  41. Hecky RE, Campbell P, Hendzell LL (1993) The stoichiometry of carbon, nitrogen, and phosphorus in particulate matter of lakes and oceans. Limnol Oceanogr 38:709–924

    Google Scholar 

  42. Howarth RW, Marino R, Cole JJ (1988) N2 fixation in freshwater estuarine and marine ecosystems. 2. Biogeochemical controls. Limnol Oceanogr 33:688–701

    Google Scholar 

  43. Jasper JP, Gagosian RB (1989) Glacial-interglacial climatically forced d13C variations in sedimentary organic matter. Nature 342:60–62

    Google Scholar 

  44. Kawai A, Sugahara I (1971) Microbiological studies on N2 fixation in aquatic environments. III. On the nitrogen-fixing bacteria in offshore regions. Bull Jpn Soc Sci Fish 37:981–985

    Google Scholar 

  45. King GM, Carlton RG, Sawyer TE (1990) Anaerobic metabolism and oxygen distribution in the carbonate sediments of a submarine canyon. Mar Ecol Prog Ser 58:275–285

    Google Scholar 

  46. Kirshtein JD, Paerl HW, Zehr J (1991) Amplification, cloning, and sequencing of a nifH segment from aquatic microorganisms and natural communities. Appl Environ Microbiol 57:2645–2650

    Google Scholar 

  47. Klump V, Martens C (1983) Benthic nitrogen regeneration. In: Carpenter EJ, Capone DG (eds) Nitrogen in the marine environment. Academic Press, New York, pp 411–457

    Google Scholar 

  48. Lindahl T (1993) Instability and decay of the primary structure of DNA. Nature 362:709–715

    Article  CAS  PubMed  Google Scholar 

  49. Lyle MW, Prahl FG, Sparrow MA (1992) Upwelling and productivity changes inferred from a temperature record in the central equatorial Pacific. Nature 355:812–815

    Google Scholar 

  50. Maidak BL, Larsen N, McCaughey MJ, Overbeek R, Olsen GJ, Fogel K, Blandy J, Woese CR (1994) The ribosomal database project. Nucleic Acids Res 22:3485–3487

    Google Scholar 

  51. Martinez L, Silver MW, King JM, Alldredge AL (1983) N2 fixation by floating diatom mats: a source of new nitrogen to oligotrophic ocean waters. Science 221:152–154

    Google Scholar 

  52. McElroy MB (1983) Marine biological controls on atmospheric CO2 and climate. Nature 302:328–329

    Google Scholar 

  53. Merrick MJ (1992) Regulation of N2 fixation genes in free-living and symbiotic bacteria. In: Stacey G, Burris RH, Evans HJ (eds) Biological N2 fixation. Chapman and Hall, New York, pp 835–876

    Google Scholar 

  54. Michaels A, and 8 others (1996) Inputs, losses, and transformations of nitrogen and phosphorus in the deep North Atlantic Ocean. Biogeochemistry in press.

  55. Mitsui A, Kumazawa S, Takahashi A, Ikemoto H, Cao S, Arai T (1986) Strategy by which nitrogen-fixing unicellular cyanobacteria grow photoautotrophically. Nature 323:720–722

    Google Scholar 

  56. Mortlock RA, Charles CD, Froelich PN, Zibello MA, Saltzman J, Hays JD, Burckle LH (1991) Evidence for lower productivity in the Antarctic Ocean during the last glaciation. Nature 351:220–223

    Google Scholar 

  57. Mullis KB, Faloona FA (1987) Specific synthesis of DNA in vitro via a polymerase-catalyzed chain reaction. Meth Enzymol 155:335–350

    Google Scholar 

  58. O'Donohue, MJ, Moriarty DJW, Mac Rae IC (1991) N2 fixation in sediments and the rhizosphere of the seagrass Zostera capricom. Microb Ecol 22:53–64

    Google Scholar 

  59. O'Neil JM, Capone DG (1989) Nitrogenase activity in tropical carbonate marine sediments. Mar Ecol Prog Ser 56:145–156

    Google Scholar 

  60. Olsen GJ, Woese CR, Overbeek R (1994) The winds of (evolutionary) change: breathing new life into microbiology. J Bacteriol 176:1–6

    Google Scholar 

  61. Pääbo S (1989) Ancient DNA: extraction, characterization, molecular cloning, and enzymatic amplification. Proc Natl Acad Sci USA 86:1939–1943

    Google Scholar 

  62. Paerl HW (1990) Physiological ecology and regulation of N2 fixation in natural waters. In: Marshall KC (ed) Advances in microbial ecology. Plenum Publishing, pp 305–344

  63. Paerl HW Bebout BM, Prufert LE (1989) Naturally occurring patterns of oxygenic photosynthesis and N2 fixation in a marine microbial mat: physiological and ecological ramifications. In: Cohen Y, Rosenberg E (eds) Microbial mats: physiological ecology of benthic microbial communities. Am Soc Microbiol, pp 326–341

  64. Parks RJ, Cragg BA, Bale SJ, Getliff JM, Goodman K, Rochelle PA, Fry JC, Weightman AJ, Harvey SM (1994) Deep bacterial biosphere in Pacific Ocean sediment. Nature 371:410–413

    Google Scholar 

  65. Patriquin D, Knowles R (1972) N2 fixation in the rhizosphere of marine angiosperms. Mar Biol 16:49–58

    Google Scholar 

  66. Pichard SL, Paul JH (1993) Gene expression per gene dose, a specific measure of gene expression in aquatic microorganisms. Appl Environ Microbiol 59:451–457

    Google Scholar 

  67. Pope MR, Murrell SA, Ludden PW (1985) Convalent modification of the iron protein of nitrogenase from Rhodospirillum rubrum by adenosine diphoshoribosylation of a specific arginine residue. Proc Natl Acad Sci USA 82:3173–3177

    Google Scholar 

  68. Postgate JR (1982) The fundaments of N2 fixation. Cambridge University Press, Cambridge, Massachusetts, pp 252

    Google Scholar 

  69. Redfield AC (1958) The biological control of chemical factors in the environment. Am Sci 46:205–222

    Google Scholar 

  70. Revsbech NP, Sorensen J, Blackburn TH, Lomholt J (1981) Distribution of oxygen in marine sediments measured with microelectrodes. Limnol Oceanogr 25:403–411

    Google Scholar 

  71. Reysenbach AL, Giver LJ, Wickham GS, Pace NR (1992) Differential amplification of rRNA genes by polymerase chain reaction. Appl Environ Microbiol 58:3417–3418

    Google Scholar 

  72. Roberts GP, Ludden PW (1992) N2 fixation by photosynthetic bacteria. In: Stacey G, Burris RH, Evans HJ (eds) Biological N2 fixation. Chapman and Hall, New York, pp 135–165

    Google Scholar 

  73. Rostek F, Ruhland G, Bassinot FC, Muller PJ, Labeyrle LD, Lancelot Y, Bard E (1993) Reconstructing sea surface temperature and salinity using δ18O and alkenone records. Nature 364:319–323

    Google Scholar 

  74. Ruvkun GB, Ausubel FM (1980) Interspecies homology of nitrogenase genes. Proc Natl Acad Sci USA 77:191–195

    Google Scholar 

  75. Saiki RK, Gelfand DH, Stoffel S, Scharf SJ, Higuchi R, Horn GT, Mullis KB, Erlich HA (1988) Primer-directed enzymatic amplification of DNA with a thermostable DNA polymerase. Science 239:487–491

    CAS  PubMed  Google Scholar 

  76. Sancetta C (1992) Primary production in the glacial North Atlantic and North Pacific oceans. Nature 360:249–251

    Google Scholar 

  77. Seitzinger SP, Garber JH (1987) N2 fixation and 15N2 calibration of the acetylene reduction assay in coastal marine sediments. Mar Ecol Prog Ser 37:65–73

    Google Scholar 

  78. Short FT (1983) The seagrass Zostera marina L.: plant morphology and bed structure in relation to sediment ammonium in Izembeck lagoon Alaska. Aquat Bot 16:149–161

    Google Scholar 

  79. Simonet P, Grosjean MC, Misra AK, Nazaret S, Cournoyer B, Normand P (1991) Frankia genus-specific characterization by polymerase chain reaction. Appl Environ Microbiol 57:3278–3286

    Google Scholar 

  80. Ueda T, Suga Y, Yahiro N, Matsuguchi T (1995) Remarkable N2-fixing bacterial diversity detected in rice roots by molecular evolutionary analysis of nifH gene sequences. J Bacteriol 177:1414–1417

    Google Scholar 

  81. Vitousek PM, Howarth RW (1991) Nitrogen limitation on land and in the sea: how can it occur? Biogeochemistry 13:87–115

    Google Scholar 

  82. Ward DM, Weller R, Bateson MM (1990) 16S rRNA sequences reveal numberous uncultured inhabitants in a natural community. Nature 345:63–65

    Article  CAS  PubMed  Google Scholar 

  83. Woese CR (1987) Bacterial evolution. Microbiol Rev 51:221–271

    CAS  PubMed  Google Scholar 

  84. Woodward SR, Weyand NJ, Bunnell M (1994) DNA sequence from Cretaceous period bone fragments. Nature 266:1229–1231

    Google Scholar 

  85. Wynn-Williams DD, Rhodes ME (1974) N2 fixation in seawater. J Appl Bacteriol 37:203–216

    Google Scholar 

  86. Yoch DC, Whiting GJ (1986) Evidence for NH4 + switch-off regulation of nitrogenase activity by bacteria in salt marsh sediments and roots of the grass Spartina alterniflora. Appl Environ Microbiol 51:143–149

    Google Scholar 

  87. Young JPW (1992) Phylogenetic classification of nitrogen-fixing organisms. In: Stacey G, Burris RH, Evans HJ (eds) Biological N2 fixation. Chapman and Hall, New York, pp 43–86

    Google Scholar 

  88. Zehr JP (1992) Molecular biology of nitrogen fixation in natural populations of marine cyanobacteria. In: Carpenter EJ, Capone DG, Rueter JG (eds) Marine pelagic cyanobacteria Trichodesmium and other diszotrophs. Kluwer Academic Publishers, Dordrecht, The Netherlands, pp 249–264

    Google Scholar 

  89. Zehr JP (1994) N2 fixation in the sea: why only trichodesmium? In: Joint I, Carr N, Falkowski P (eds) NATO ASI series. Springer-Verlag.

  90. Zehr JP, McReynolds LA (1989) Use of degenerate oligonucleotides for amplification of the nifH gene from the marine cyanobacterium Trichodesmium thiebautii. Appl Environ Microbiol 55:2522–2526

    Google Scholar 

  91. Zehr JP, Ohki K, Fujita Y (1991) Arrangement of nitrogenase structural genes in an aerobic filamentous nonheterocystous cyanobacterium. J Bacteriol 173:7055–7058

    Google Scholar 

  92. Zehr JP, Wyman M, Miller V, Duguay L, Capone DG (1993) Modification of the Fe protein of nitrogenase in natural populations of Trichodesmium thiebautii. Appl Environ Microbiol 59:669–676

    Google Scholar 

  93. Zehr JP, Mellon M, Braun S, Litaker W, Steppe T, Paerl HW (1995) Diversity of heterotrophic N2 fixation genes in a marine cyanobacterial mat. Appl Environ Microbiol 61:2527–2532

    Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Department of Biology MRC 303, Rensselaer Polytechnic Institute, 12180-3590, Troy, New York, USA

    J. P. Zehr

  2. Chesapeake Biological Laboratory, University of Maryland, 20688, Solomons, Maryland, USA

    D. G. Capone

Authors
  1. J. P. Zehr
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. D. G. Capone
    View author publications

    You can also search for this author in PubMed Google Scholar

Additional information

Correspondence to: J.P. Zehr.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Zehr, J.P., Capone, D.G. Problems and promises of assaying the genetic potential for nitrogen fixation in the marine environment. Microb Ecol 32, 263–281 (1996). https://doi.org/10.1007/BF00183062

Download citation

  • Received:

  • Accepted:

  • Issue Date:

  • DOI: https://doi.org/10.1007/BF00183062

Keywords

Search

Navigation