Skip to main content
SpringerLink
Log in

The significance of endophytic and phyllospheric N2 fixation in forest trees: evidence from stable (15N) and radioactive (13N) tracer studies

  • Review
  • Published:
Trees Aims and scope Submit manuscript

Abstract

Key message

The proportional contribution of endophytic and phyllospheric diazotrophs to tree N nutrition can be quantified by 15N techniques.

Abstract

Observations of vigorous growth of Pinus spp. on unreclaimed gravel mining sites, together with measured net nitrogen (N) gains of 50 kg N ha–1 y–1 in sand culture, indicate the importance of alternative N sources when soil organic matter (SOM) cannot meet plant demand. Non-nodulated trees can respond positively to inoculation with N2 fixing bacteria under conditions of low N supply, suggesting that non-nodular symbiotic N2 fixation can make a significant contribution to tree nutrition, a well-established pathway for cereals, forage grasses and some industrial crops in N-limited environments. We reviewed the literature where non-isotopic approaches and techniques based on stable (15N) and radioactive (13N) tracers were used to estimate the relative contribution of endophytic and phyllospheric N2 fixation to the N economies of non-nodulated trees. The principal avenue of enquiry has so far involved determination of the response of trees to inoculation with diazotrophic bacteria. There is a need to obtain in situ measurements of the reliance of non-leguminous trees on endophytic and phyllospheric N2 fixation under a range of environmental and edaphic conditions. Current methodological obstacles have to be overcome to meet this challenge. Future research directions are suggested.

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

  • Amir HG, Shamsuddin ZH, Halimi MS, Ramlan MF, Marziah M (1999) Potential use of rhizobacteria for sustainable oil palm seedling production. Malaysian J Soil Sci 3:39–50

    Google Scholar 

  • Anand R, Chanway C (2013) N2-fixation and growth promotion in cedar colonized by an endophytic strain of Paenibacillus polymyxa. Biol Fertil Soils 49:235–239

    Article  CAS  Google Scholar 

  • Anand R, Grayston S, Chanway C (2013) N2-fixation and seedling growth promotion of lodgepole pine by endophytic Paenibacillus polymyxa. Microb Ecol 66:369–374

    Article  CAS  PubMed  Google Scholar 

  • Bal A, Chanway CP (2012a) Evidence of nitrogen fixation in lodgepole pine inoculated with diazotrophic Paenibacillus polymyxa. Botany 90:891–896

    Article  CAS  Google Scholar 

  • Bal A, Chanway CP (2012b) 15N foliar dilution of western red cedar in response to seed inoculation with diazotrophic Paenibacillus polymyxa. Biol Fertil Soils 48:967–971

    Article  Google Scholar 

  • Barros BGF, de Freitas ADS, Tabosa JN, De Lyra MCCP, Mergulhão ACES, da Silva AF, Oliveira WS, Fernandes-Júnior PI, Sampaio EVSP (2020) Biological nitrogen fixation in field-grown sorghum under different edaphoclimatic conditions is confirmed by N isotopic signatures. Nutr Cycl Agroecosyst 117:93–101

    Article  CAS  Google Scholar 

  • Bei Q, Liu G, Tang H, Cadisch G, Rasche F, Xie Z (2013) Heterotrophic and phototrophic 15N2 fixation and distribution of fixed 15N in a flooded rice–soil system. Soil Biol Biochem 59:25–31

    Article  CAS  Google Scholar 

  • Boddey RM, Knowles R (1987) Methods for quantification of nitrogen fixation associated with gramineae. CRC Crit Rev Plant Sci 6:209–266

    Article  CAS  Google Scholar 

  • Boddey RM, Peoples MB, Palmer B, Dart PJ (2000) Use of the 15N natural abundance technique to quantify biological nitrogen fixation by woody perennials. Nutr Cycl Agroecosyst 57:235–270

    Article  Google Scholar 

  • Bormann BT, Bormann FH, Bowden WB, Piece RS, Hamburg SP, Wang D, Snyder MC, Li CY, Ingersoll RC (1993) Rapid N2 fixation in pines, alder, and locust: evidence from the sandbox ecosystems study. Ecology 74:583–598

    Article  Google Scholar 

  • Chalk PM (1991) The contribution of associative and symbiotic nitrogen fixation to the nitrogen nutrition of non-legumes. Plant Soil 132:29–39

    Article  CAS  Google Scholar 

  • Chalk PM (2016) The strategic role of 15N in quantifying the contribution of endophytic N2 fixation to the N nutrition of non-legumes. Symbiosis 69:63–80

    Article  CAS  Google Scholar 

  • Chalk PM (2020) Whither the enigma of soil nitrogen balance sheets? Plant Soil 456:455–460

    Article  CAS  Google Scholar 

  • Chalk PM, Craswell ET (2018) An overview of the role of 15N in quantifying biological N2 fixation (BNF) and BNF dynamics in agro-ecosystems. Symbiosis 75:1–16

    Article  CAS  Google Scholar 

  • Chalk PM, He J-Z, Peoples MB, Chen D (2017) 15N2 as a tracer of biological N2 fixation: a 75-year retrospective. Soil Biol Biochem 106:36–50

    Article  CAS  Google Scholar 

  • Chanway CP, Holl FB (1991) Biomass increase and associative nitrogen fixation of mycorrhizal Pinus contorta seedlings inoculated with a plant growth promoting Bacillus strain. Can J Bot 69:507–511

    Article  CAS  Google Scholar 

  • Doty SL, Sher AW, Fleck ND, Khorasani M, Bumgarner RE, Khan Z, Ko AW, Kim SH, DeLuca TH (2016) Variable nitrogen fixation in wild Populus. PLoS ONE 11:e0155979

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  • Forrester DI, Schortemeyer M, Stock WD, Bauhus J, Khanna PK, Cowie AL (2007) Assessing nitrogen fixation in mixed-and single-species plantations of Eucalyptus globulus and Acacia mearnsii. Tree Physiol 27:1319–1328

    Article  CAS  PubMed  Google Scholar 

  • Fürnkranz M, Wanek W, Richter A, Abell G, Rasche F, Sessitsch A (2008) Nitrogen fixation by phyllosphere bacteria associated with higher plants and their colonizing epiphytes of a tropical lowland rainforest of Costa Rica. ISME J 2:561–570

    Article  PubMed  CAS  Google Scholar 

  • Ishii S, Suzui N, Ito S, Ishioka NS, Kawachi N, Ohtake N, Ohyama T, Fujimaki S (2009) Real-time imaging of nitrogen fixation in an intact soybean plant with nodules using 13N-labeled nitrogen gas. Soil Sci Plant Nutr 55:660–666

    Article  CAS  Google Scholar 

  • Jones K (1970) Nitrogen fixation in the phyllosphere of the Douglas fir, Pseudotsuga douglasii. Ann Bot 34:239–244

    Article  Google Scholar 

  • Kasel MCK, Schueller MJ, Ferrieri RA (2010) Optimizing [13N] N2 radiochemistry for nitrogen-fixation in root nodules of legumes. J Labelled Comp Radiopharm 53:592–597

    Article  CAS  Google Scholar 

  • Knoth JL, Kim SH, Ettl GJ, Doty SL (2014) Biological nitrogen fixation and biomass accumulation within poplar clones as a result of inoculations with diazotrophic endophyte consortia. New Phytol 201:599–609

    Article  CAS  PubMed  Google Scholar 

  • Meeks JC, Steinberg NA, Enderlin CS, Joseph CM, Peters GA (1987) Azolla-Anabaena relationship: XIII. Fixation of [13N] N2. Plant Physiol 84:883–886

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Moyes AB, Kueppers LM, Pett-Ridge J, Carper DL, Vandehey N, O’Neil J, Frank AC (2016) Evidence for foliar endophytic nitrogen fixation in a widely distributed subalpine conifer. New Phytol 210:657–668

    Article  CAS  PubMed  Google Scholar 

  • Oses R, Frank AC, Valenzuela S, Rodríguez J (2018) Nitrogen fixing endophytes in forest trees. In: Pirttilä AM, Frank AC (eds) Endophytes of forest trees; biology and applications. Springer, Cham, pp 191–204

    Chapter  Google Scholar 

  • Padda KP, Puri A, Zeng Q, Chanway CP, Wu X (2017) Effect of GFP-tagging on nitrogen fixation and plant growth promotion of an endophytic diazotrophic strain of Paenibacillus polymyxa. Botany 95:933–942

    Article  CAS  Google Scholar 

  • Padda KP, Puri A, Chanway C (2019) Endophytic nitrogen fixation – a possible ‘hidden’source of nitrogen for lodgepole pine trees growing at unreclaimed gravel mining sites. FEMS Microbiol Ecol 95:fiz172

    Article  CAS  PubMed  Google Scholar 

  • Puri A, Padda KP, Chanway CP (2017) Beneficial effects of bacterial endophytes on forest tree species. In: Maheshwari DK, Annapurna K (eds) Endophytes: crop productivity and protection, vol 2. Springer, Cham, pp 111–132

    Chapter  Google Scholar 

  • Puri A, Padda KP, Chanway CP (2020a) Can naturally-occurring endophytic nitrogen-fixing bacteria of hybrid white spruce sustain boreal forest tree growth on extremely nutrient-poor soils? Soil Biol Biochem 140:107642

    Article  CAS  Google Scholar 

  • Puri A, Padda KP, Chanway CP (2020b) Evaluating lodgepole pine endophytes for their ability to fix nitrogen and support tree growth under nitrogen-limited conditions. Plant Soil 455:271–287

    Article  CAS  Google Scholar 

  • Spriggs AC, Stock WD, Dakora FD (2003) Influence of mycorrhizal associations on foliar δ15N values of legume and non-legume shrubs and trees in the fynbos of South Africa: implications for estimating N2 fixation using the 15N natural abundance method. Plant Soil 255:495–502

    Article  CAS  Google Scholar 

  • Tang Q, Puri A, Padda KP, Chanway CP (2017) Biological nitrogen fixation and plant growth promotion of lodgepole pine by an endophytic diazotroph Paenibacillus polymyxa and its GFP-tagged derivative. Botany 95:611–619

    Article  CAS  Google Scholar 

  • Unkovich M (2013) Isotope discrimination provides new insight into biological nitrogen fixation. New Phytol 198:643–646

    Article  CAS  PubMed  Google Scholar 

  • Unkovich M, Herridge D, James EK, Giller K, Peoples MB (2020) Reliable quantification of N2 fixation by non-legumes remains problematic. Nutr Cycl Agroecosyst 118:223–225

    Article  Google Scholar 

  • Unkovich M, Herridge D, Peoples M, Cadisch G, Boddey R, Giller K, Alves B, Chalk P (2008) Measuring plant-associated nitrogen fixation in agricultural systems. Australian Centre for International Agricultural Research, Canberra

    Google Scholar 

  • Van Langenhove L, Depaepe T, Verryckt LT, Fuchslueger L, Donald J, Leroy C, Moorthy SMK, Gargallo-Garriga A, Ellwood MF, Verbeeck H, Van Der Straeten D, Peñuelas J, Janssens IA (2021) Comparable canopy and soil free-living nitrogen fixation rates in a lowland tropical forest. Sci Total Environ 754:142202

    Article  PubMed  CAS  Google Scholar 

  • Wang X, Bei Q, Yang W, Zhang H, Hao J, Qian L, Feng Y, Xie Z (2020) Unveiling of active diazotrophs in a flooded rice soil by combination of NanoSIMS and 15N2-DNA-stable isotope probing. Biol Fertil Soils 56:1189–1199

    Article  CAS  Google Scholar 

  • Witty JF, Minchin FR (1988) Measurement of nitrogen fixation by the acetylene reduction assay; myths and mysteries. In: Beck DP, Materon LA (eds) Nitrogen fixation by legumes in Mediterranean agriculture. Developments in Plant and Soil Sciences, vol 32. Springer, Dordrecht, pp 331–344

    Chapter  Google Scholar 

  • Wolk CP, Austin SM, Bortins J, Galonsky A (1974) Autoradiographic localization of 13N after fixation of 13N-labeled nitrogen gas by a heterocyst-forming blue-green alga. J Cell Biol 61:440–453

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Yang H, Puri A, Padda KP, Chanway CP (2016) Effects of Paenibacillus polymyxa inoculation and different soil nitrogen treatments on lodgepole pine seedling growth. Can J for Res 46:816–821

    Article  CAS  Google Scholar 

  • Yin YG, Ishii S, Suzui N, Igura M, Kurita K, Miyoshi Y, Nagasawa N, Taguchi M, Kawachi N (2019) On-line rapid purification of [13N] N2 gas for visualization of nitrogen fixation and translocation in nodulated soybean. Appl Radiat Isot 151:7–12

    Article  CAS  PubMed  Google Scholar 

Download references

Funding

No funding was received to assist with the preparation of this manuscript.

Author information

Authors and Affiliations

  1. Faculty of Veterinary and Agricultural Sciences, The University of Melbourne, Parkville, Victoria, 3010, Australia

    Phillip M. Chalk, Shu K. Lam & Deli Chen

Authors
  1. Phillip M. Chalk
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Shu K. Lam
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Deli Chen
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Phillip M. Chalk.

Ethics declarations

Conflict of interest

The authors declare that they have no conflict of interest.

Additional information

Communicated by R. Guy.

Publisher's Note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Chalk, P.M., Lam, S.K. & Chen, D. The significance of endophytic and phyllospheric N2 fixation in forest trees: evidence from stable (15N) and radioactive (13N) tracer studies. Trees 36, 1179–1184 (2022). https://doi.org/10.1007/s00468-021-02256-y

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00468-021-02256-y

Keywords

Search

Navigation