Biology and Fertility of Soils

, Volume 44, Issue 1, pp 113–119 | Cite as

Response of denitrifying communities to successive soil freeze–thaw cycles

  • Yosuke YanaiEmail author
  • Koki Toyota
  • Masanori Okazaki
Original Paper


The effect of soil freeze–thaw cycles on the denitrification potential was examined based on the C2H2 inhibition method. The gross N2O production curve of the soil sample (incubation with C2H2) showed minor changes between the freeze–thaw treatment and the unfrozen control. However, kinetics analysis revealed that the initial production rate, an indicator of the population density of denitrifying communities, decreased (P = 0.043) and the specific growth rate constant, an indicator of the activity of denitrifying communities, increased (P = 0.039) as a result of the freeze–thaw cycles in five of six soil samples examined. The increase in the specific growth rate constant suggested the stimulation of the activity of denitrifying communities that survived after the freeze–thaw cycles and may explain the minor suppression on the gross N2O production in spite of decreasing the population density of denitrifying communities that was suggested by the initial production rate. The net N2O production curve of the soil sample (incubation without C2H2) showed a remarkable change in one out of six soil samples, and in that one soil sample, N2O release to the atmosphere was largely stimulated (7.6 times) by the freeze–thaw cycles. However, the stimulation of the N2O release by the freeze–thaw cycles was even observed in two other selected soil samples (4.6 and 1.8 times), suggesting that an imbalance in the N2O-producing and N2O-reducing activities of denitrifying communities might complementally explain the N2O release stimulated by the freeze–thaw cycles.


Freeze–thaw cycles Soil Denitrifying communities C2H2 inhibition method N2O production 



The authors thank Dr. Y. Kurokawa and Dr. T. Ezawa for providing soil samples. A part of this research was funded by a Sasakawa Scientific Research Grant from The Japan Science Society (16-315), the TUA&T 21st Century COE Program (Evolution and Survival of Technology-based Civilization), and JSPS Research Fellowships for Young Scientists (17-6518).


  1. Anderson JPE, Domsch KH (1975) Measurement of bacterial and fungal contributions to respiration of selected agricultural and forest soils. Can J Microbiol 21:314–322PubMedCrossRefGoogle Scholar
  2. Burton DL, Beauchamp EG (1994) Profile nitrous oxide and carbon dioxide concentrations in a soil subject to freezing. Soil Sci Soc Am J 58:115–122CrossRefGoogle Scholar
  3. Christensen S, Christensen BT (1991) Organic matter available for denitrification in different soil fractions: effect of freeze/thaw cycles and straw disposal. J Soil Sci 42:637–647CrossRefGoogle Scholar
  4. Christensen S, Tiedje JM (1990) Brief and vigorous N2O production by soil at spring thaw. J Soil Sci 41:1–4CrossRefGoogle Scholar
  5. Coyle CL, Zumft WG, Kroneck PMH, Körner H, Jakob W (1985) Nitrous oxide reductase from denitrifying Pseudomonas perfectomarina—purification and properties of a novel multi-copper enzyme. Eur J Biochem 153:459–467PubMedCrossRefGoogle Scholar
  6. Duxbury JM, Bouldin DR, Terry RE, Tate RL III (1982) Emissions of nitrous oxide from soils. Nature 298:462–464CrossRefGoogle Scholar
  7. Flessa H, Dörsch P, Beese F (1995) Seasonal variation of nitrous oxide and methane fluxes in differently managed arable soils in southern Germany. J Geophys Res 100:23115–23124CrossRefGoogle Scholar
  8. Goodroad LL, Keeney DR (1984) Nitrous oxide emissions from soils during thawing. Can J Soil Sci 64:187–194Google Scholar
  9. Groffman PM, Hardy JP, Nolan S, Fitzhugh RD, Driscoll CD, Fahey TJ (1999) Snow depth, soil frost, and nutrient loss in a northern hardwood forest. Hydrol Proc 13:2275–2286CrossRefGoogle Scholar
  10. Groffman PM, Driscoll CT, Fahey TJ, Hardy JP, Fitzhugh RD, Tierney GL (2001) Colder soils in a warmer world: a snow manipulation study in a northern hardwood forest ecosystem. Biogeochem 56:135–150CrossRefGoogle Scholar
  11. Holtan-Hartwig L, Bechmann M, Høyås TR, Linjordet R, Bakken LR (2002a) Heavy metals tolerance of soil denitrifying communities: N2O dynamics. Soil Biol Biochem 34:1181–1190CrossRefGoogle Scholar
  12. Holtan-Hartwig L, Dörsch P, Bakken LR (2002b) Low temperature control of soil denitrifying communities: kinetics of nitrous oxide production and reduction. Soil Biol Biochem 34:1797–1806CrossRefGoogle Scholar
  13. Koponen HT, Flöjt L, Martikainen PJ (2004) Nitrous oxide emissions from agricultural soils at low temperatures: a laboratory microcosm study. Soil Biol Biochem 36:757–766CrossRefGoogle Scholar
  14. Ludwig B, Wolf I, Teepe R (2004) Contribution of nitrification and denitrification to the emission of nitrous oxide in a freeze–thaw event in an agricultural soil. J Plant Nutr Soil Sci 167:678–684CrossRefGoogle Scholar
  15. Matsubara T, Sano M (1985) Isolation and some properties of a novel violet copper protein from a denitrifying bacterium, Alcaligenes sp. Chem Lett 1985:1053–1056CrossRefGoogle Scholar
  16. Müller C, Martin M, Stevens RJ, Laughlin RJ, Kammann C, Ottow JCG, Jäger H-J (2002) Processes leading to nitrous oxide emissions in grassland soil during freezing and thawing. Soil Biol Biochem 34:1325–1331CrossRefGoogle Scholar
  17. Öquist MG, Nilsson M, Sörensson F, Kaisimir-Klemedtsson Å, Persson T, Weslien P, Klemedtsson L (2004) Nitrous oxide production in a forest soil at low temperatures—processes and environmental controls. FEMS Microbiol Ecol 49:371–378CrossRefPubMedGoogle Scholar
  18. Röver M, Heinemeyer O, Kaiser E-A (1998) Microbial induced nitrous oxide emissions from an arable soil during winter. Soil Biol Biochem 30:1859–1865CrossRefGoogle Scholar
  19. Sehy U, Ruser R, Munch JC (2003) Nitrous oxide fluxes from maize field: relationship to yield, site-specific fertilization, and soil conditions. Agric Ecosys Environ 99:97–111CrossRefGoogle Scholar
  20. Sehy U, Dyckmans J, Ruser R, Munch JC (2004) Adding dissolved organic carbon to simulate freeze–thaw related N2O emissions from soil. J Plant Nutr Soil Sci 167:471–478CrossRefGoogle Scholar
  21. Stenström J, Hansen A, Svensson B (1991) Kinetics of microbial growth-associated production formation. Swed J Agr Res 21:55–62Google Scholar
  22. Teepe R, Ludwig B (2004) Variability of CO2 and N2O emissions during freeze–thaw cycles: results of model experiments on undisturbed forest-soil cores. J Plant Nutr Soil Sci 167:153–159CrossRefGoogle Scholar
  23. Teepe R, Brumme R, Beese F (2000) Nitrous oxide emissions from frozen soils under agricultural, fallow and forest land. Soil Biol Biochem 32:1807–1810CrossRefGoogle Scholar
  24. Teepe R, Brumme R, Beese F (2001) Nitrous oxide emissions from soil during freezing and thawing periods. Soil Biol Biochem 33:1269–1275CrossRefGoogle Scholar
  25. Tiedje JM (1994) Denitrifiers. In: Weaver RD, Angle JS, Bottomley PS (eds) Methods of soil analysis. Part 2—microbiological and biochemical properties. Soil Science Society of America, Wisconsin, pp 245–267Google Scholar
  26. van Bochove E, Prevost D, Pelletier F (2000) Effects of freeze–thaw and soil structure on nitrous oxide produced in a clay soil. Soil Sci Soc Am J 64:1638–1643CrossRefGoogle Scholar
  27. Wilhelm E, Battino R, Wilcock RJ (1977) Low-pressure solubility of gases in liquid water. Chem Rev 77:219–262CrossRefGoogle Scholar
  28. Yanai Y, Toyota K, Okazaki M (2004a) Effects of successive soil freeze–thaw cycles on soil microbial biomass and organic matter decomposition potential of soils. Soil Sci Plant Nutr 50:821–829Google Scholar
  29. Yanai Y, Toyota K, Okazaki M (2004b) Effects of successive soil freeze–thaw cycles on nitrification potential of soils. Soil Sci Plant Nutr 50:831–837Google Scholar
  30. Yoshinari T, Hynes R, Knowles R (1977) Acetylene inhibition of nitrous oxide reduction and measurement of denitrification and nitrogen fixation in soil. Soil Biol Biochem 9:177–183CrossRefGoogle Scholar
  31. Zumft W, Matsubara T (1985) A novel kind of multi-copper protein as terminal oxidereductase of nitrous oxide respiration in Pseudomonas perfectomarinus. FEBS Lett 148:107–112CrossRefGoogle Scholar

Copyright information

© Springer-Verlag 2007

Authors and Affiliations

  1. 1.Department of Environment-Symbiotic Production Systems, Division of Bio-Applications and Systems Engineering, Graduate School of Bio-Applications and Systems EngineeringTokyo University of Agriculture and TechnologyTokyoJapan
  2. 2.Division of Bio-Applications and Systems Engineering, Institute of Symbiotic Science and TechnologyTokyo University of Agriculture and TechnologyTokyoJapan

Personalised recommendations