Plant and Soil

, Volume 290, Issue 1–2, pp 245–257 | Cite as

NH 4 + triggers the synthesis and release of biological nitrification inhibition compounds in Brachiaria humidicola roots

  • G. V. Subbarao
  • H. Y. Wang
  • O. Ito
  • K. Nakahara
  • W. L. Berry
Original Paper


The release of chemical compounds from plant roots that suppress soil nitrification is termed biological nitrification inhibition (BNI). Determining the environmental factors that control the synthesis and release of BNI-compounds from Brachiaria humidicola (Rendle) Schweick, a tropical pasture grass that thrives on acid soils, is the focus of this investigation. Because the BNI trait is related to the N status of the plant, we investigated the possibility that the expression of this trait would be related to the forms of N found in the root environment. Plants were grown with two sources of N, NH 4 + or NO 3 for 60 days and the release of BNI-compounds monitored. Only plants grown with NH 4 + released BNI-compounds from roots. The presence of NH 4 + and possibly the secondary effect of its uptake (i.e., acidic pH) in the root environment significantly enhanced the release of BNI-compounds. Both the NH 4 + and NO 3 grown plants responded to the stimulus from NH 4 + in the root environment. BNI-compounds found in root tissue and their release were nearly three times greater in NH 4 + grown than from NO 3 grown plants. The BNI-compounds released from roots composed of at least three active components—Type-I (stable to pH changes from 3.0 to 10), Type-II (temporarily loses its inhibitory effect at a pH higher than a threshold pH of 4.5 and the inhibitory effect is reestablished when the root exudate pH is adjusted to <4.5) and Type-III (inhibitory effect is irreversibly lost if the pH of the root exudate reaches 10.0 or above). A major portion of BNI-compounds released in the presence of NH 4 + is of Type-I. In the absence of NH 4 + , mostly Type-II and Type-III BNI-compounds were released. The BNI-compounds inhibited the function of Nitrosomonas europaea through the blocking of both ammonia monooxygenase and hydroxylamino oxidoreductase pathways. These results indicate that the release of BNI-compounds from B. humidicola roots is a regulated function and that presence of NH 4 + in the root environment is necessary for the sustained synthesis and release of BNI.


Biological nitrification inhibition BNI-compounds Bioluminescence assay Nitrogen forms 



We are grateful to Dr Taro Iizumi (Kurita Water Industries Limited, Wakamtya, Japan) who has kindly provided us with the luminescent recombinant N. europaea strain for this research. We acknowledge the participation and help from our colleagues at CIAT (Cali, Colombia) during this study have also provided the seed material used for various experiments described in this manuscript. We are thankful to Dr Yiyong Zhu (Nanjing Agricultural University, Nanjing, China) for many thoughtful suggestions and inputs during the interpretation of the data. Also, we are grateful to Dr K.L. Sahrawat (ICRISAT, India), who has gone through our manuscript critically and offered many suggestions to improve the clarity of our presentation and interpretation of the data.


  1. Belser LW, Schmidt EL (1981) Inhibiting effect of nitrapyrin on three genera of ammonia oxidizing nitrifiers. Appl Environ Microbiol 41:819–821PubMedGoogle Scholar
  2. Dijkshoorn W (1973) Organic acids, and their role in ion uptake. In: Butler GW, Bailey RW (eds) Chemistry and biochemistry of herbage. Academic, London, pp 163–188Google Scholar
  3. Giles J (2005) Nitrogen study fertilizes fears of pollution. Nature 433:791PubMedCrossRefGoogle Scholar
  4. Glass ADM (2003) Nitrogen use efficiency of crop plants: physiological constraints upon nitrogen absorption. Crit Rev Plant Sci 22:453–470CrossRefGoogle Scholar
  5. Iizumi T, Nakamura K (1997) Cloning, nucleotide sequence, and regulatory analysis of the Nitrosomonas europaea dnaK gene. Appl Environ Microbiol 63:1777–1784PubMedGoogle Scholar
  6. Iizumi T, Mizumoto M, Nakamura K (1998) A bioluminescence assay using Nitrosomonas europaea for rapid and sensitive detection of nitrification inhibitors. Appl Environ Microbiol 64:3656–3662PubMedGoogle Scholar
  7. Ishikawa T, Subbarao GV, Ito O, Okada K (2003) Suppression of nitrification and nitrous oxide emission by the tropical grass Brachiaria humidicola. Plant Soil 255:413–419CrossRefGoogle Scholar
  8. Kowalchuk GA, Stephen JR (2001) Ammonia-oxidizing bacteria: a model for molecular microbial ecology. Annu Rev Microbiol 55:485–529PubMedCrossRefGoogle Scholar
  9. Lalonde S, Boles E, Hellmann H, Barker L, Patrick JW, Frommer WB, Ward JM (1999) The dual function of sugar carriers: transport and sugar sensing. Plant Cell 11:707–726PubMedCrossRefGoogle Scholar
  10. Lata JC, Degrange V, Raynaud X, Maron PA, Lensi R, Abbadie L (2004) Grass population control nitrification in Savanna soils. Funct Ecol 13:762–763CrossRefGoogle Scholar
  11. Leininger S, Urich T, Schloter M, Schwark L, Qi J, Nicol GW, Prosser JI, Schuster SC, Schleper C (2006) Archaea predominate among ammonia-oxidizing prokaryotes in soils. Nature 442:806–809PubMedCrossRefGoogle Scholar
  12. Marschner H, Haussling M, George E (1991) Ammonium and nitrate uptake rates and rhizosphere-pH in non-mycorrhizal roots of Norway spruce (Picea abies (L.) Karst.). Trees 5:14–21CrossRefGoogle Scholar
  13. McCarty GW (1999) Modes of action of nitrification inhibitors. Biol Fertil Soils 29:1–9CrossRefGoogle Scholar
  14. Miles JW, do Valle WCB, Rao IM, Euclides VPB (2004) Brachiaria grasses. In: Sollenberger LE, Moser L, Burson B (eds) Warm-season grasses. ASA/CSSA/SSSA, Madison, pp 745–783Google Scholar
  15. Norton JM, Alzerreca JJ, Suwa Y, Klotz MG (2002) Diversity of ammonia monooxygenase operon in autotrophic ammonia-oxidizing bacteria. Arch Microbiol 177:139–149PubMedCrossRefGoogle Scholar
  16. Palmgren MG, Harper JF (1999) Pumping with plant P-type ATPase. J Exp Bot 50:883–893CrossRefGoogle Scholar
  17. Prasad R, Power JF (1995) Nitrification inhibitors for agriculture, health, and the environment. Adv Agron 54:233–281CrossRefGoogle Scholar
  18. Rao IM, Zeigler RS, Vera RR, Sarkarung S (1993) Selection and breeding for acid-soil tolerance in crops: upland rice and tropical forages as case studies. Bioscience 43:454–465CrossRefGoogle Scholar
  19. Rao IM, Kerridge PC, Macedo M (1996) Adaptation to low fertility acid soils and nutritional requirements of Brachiaria. In: Miles JW, Maass BL, do Valle CB (eds) The biology, agronomy and improvement of Brachiaria. CIAT, Cali, Colombia, pp 53–71Google Scholar
  20. Raun WR, Johnson GV (1999) Improving nitrogen use efficiency for cereal production. Agron J 91:357–363CrossRefGoogle Scholar
  21. Raven JA (1986) Biochemical disposal of excess H+ in growing plants. New Phytol 104:175–206CrossRefGoogle Scholar
  22. Rice CW, Pancholy SK (1972) Inhibition of nitrification by climax ecosystem. Am J Bot 59:1033–1040CrossRefGoogle Scholar
  23. Robinson JB (1963) Nitrification in a New Zealand grassland soil. Plant Soil 14:173–183CrossRefGoogle Scholar
  24. Salsac L, Haillou S, Morot-Gaudry J, Lesaint C (1987) Nitrate and ammonium nutrition in plants. Plant Physiol Biochem 25:805–812Google Scholar
  25. Subbarao GV, Ito O, Sahrawat KL, Berry WL, Nakahara K, Ishikawa T, Watanabe T, Suenaga K, Rondon M, Rao IM (2006a) Scope and strategies for regulation of nitrification in agricultural systems—challenges and opportunities. Crit Rev Plant Sci 25:303–335CrossRefGoogle Scholar
  26. Subbarao GV, Ishikawa T, Ito O, Nakahara K, Wang HY, Berry WL (2006b) A bioluminescence assay to detect nitrification inhibitors released from plant roots—a case study with Brachiaria humidicola. Plant Soil. Scholar
  27. Sze H, Li X, Palmgren MG (1999) Energization of plant cell membranes by H+-pumping ATPases: regulation and biosynthesis. Plant Cell 11:677–689PubMedCrossRefGoogle Scholar
  28. Takaki H, Ikeda M, Yamada Y, Harada T (1968) Occurrence of glucosamine in higher plants. Soil Sci Plant Nutr (Japan) 14:56–61Google Scholar
  29. Vannelli T, Hooper AB (1992) Oxidation of nitrapyrin to 6-chloropicolinic acid by the ammonia-oxidizing bacterium Nitrosomonas europaea. Appl Environ Microbiol 58:2321–2325PubMedGoogle Scholar
  30. Woldendorp JW, Laanbroek HJ (1989) Activity of nitrifiers in relation to nitrogen nutrition of plants in natural ecosystems. Plant Soil 115:217–228CrossRefGoogle Scholar
  31. Yan F, Feuerle R, Schaffer S, Fortmeier H, Schubert S (1998) Adaptation of active proton pumping and plasmalemma ATPase activity of corn roots to low root medium pH. Plant Physiol 117:311–319PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media B.V. 2006

Authors and Affiliations

  • G. V. Subbarao
    • 1
  • H. Y. Wang
    • 2
  • O. Ito
    • 1
  • K. Nakahara
    • 3
  • W. L. Berry
    • 4
  1. 1.Crop Production and Environment DivisionJapan International Research Center for Agricultural Sciences (JIRCAS)IbarakiJapan
  2. 2.The State Key Laboratory of Soil and Sustainable AgricultureNanjingChina
  3. 3.Food Science and Technology DivisionJapan International Research Center for Agricultural Sciences (JIRCAS)IbarakiJapan
  4. 4.Department of Ecology and Evolutionary BiologyUniversity of CaliforniaLos AngelesUSA

Personalised recommendations