Plant and Soil

, Volume 415, Issue 1–2, pp 245–255 | Cite as

Wheat colonization by an Azospirillum brasilense ammonium-excreting strain reveals upregulation of nitrogenase and superior plant growth promotion

  • K. F. D. N. Santos
  • V. R. Moure
  • V. Hauer
  • A. R. S Santos
  • L. Donatti
  • C. W. Galvão
  • F. O. Pedrosa
  • E. M. Souza
  • R. Wassem
  • M. B. R. Steffens
Regular Article

Abstract

Aims

In this work, an ammonium-excreting strain (HM053) of A. brasilense was further characterized genetically and biochemically, and its abilities to colonize and promote wheat growth were determined.

Methods

Immunoblot, reverse transcription-qPCR, and DNA sequencing were used for HM053 characterization. To analyze wheat-A. brasilense interaction nifH::gusA fusions in the wild-type FP2 (FP2-7) and HM053 (HM053-36) backgrounds were employed.

Results

HM053 glutamine synthetase (GS) was not adenylylated in response to an ammonium shock or under any condition tested. Sequencing of the glnA gene revealed a substitution of a proline residue by a leucine at position 347 of the GS. Under axenic growth condition, HM053 was capable of colonizing the surface of wheat roots and increased by 30 and 49% the shoot and root dry weight, respectively, when compared with uninoculated plants, and by 30 and 31% when compared with the parental strain FP2. Although HM053-36 and FP2-7 showed GUS activity located mainly at lateral root emergence points, HM053-36 consistently showed stronger signals and expressed the nifH gene at a level 278 fold higher than strain FP2 in planta, according to qPCR data.

Conclusions

HM053, a spontaneous mutant in GS, increased wheat root and shoot dry weight when compared to the wild-type FP2. HM053 ability to excrete ammonium and fix nitrogen constitutively, even in the presence of high NH4 + concentration, could explain why this mutant has a higher potential to promote plant growth than FP2 and suggests HM053 as a potential nitrogen biofertilizer. However, HM053 should be tested under field conditions to evaluate its abilities to compete with indigenous microflora.

Keywords

Azospirillum brasilense Ammonium-excreting mutant Biofertilizer Glutamine synthetase glnA gene 

Abbreviations

PGPR

Plant-growth-promoting rhizobacterium (PGPR)

BNF

Biological nitrogen fixation

GS

Glutamine synthetase

GOGAT

Glutamate synthase

GUS

β-glucuronidase

CFU

Colony forming units

D.a.i

Days after inoculation

Notes

Acknowledgements

This work was supported by the National Institute of Science and Technology on Biological Nitrogen Fixation (INCT/CNPq). We thank Roseli Prado, Valter A. Baura, and Marilza Doroty Lamour for the technical assistance.

Supplementary material

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Figure S1 (TIFF 83470 kb)
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Figure S2 (TIFF 143630 kb)
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Figure S3 (TIFF 147708 kb)

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Copyright information

© Springer International Publishing Switzerland 2016

Authors and Affiliations

  • K. F. D. N. Santos
    • 1
  • V. R. Moure
    • 1
  • V. Hauer
    • 2
  • A. R. S Santos
    • 1
  • L. Donatti
    • 3
  • C. W. Galvão
    • 4
  • F. O. Pedrosa
    • 1
  • E. M. Souza
    • 1
  • R. Wassem
    • 2
  • M. B. R. Steffens
    • 1
  1. 1.Departamento de Bioquímica e Biologia MolecularUniversidade Federal do ParanáCuritibaBrazil
  2. 2.Departamento de GenéticaUniversidade Federal do ParanáCuritibaBrazil
  3. 3.Departamento de Biologia CelularUniversidade Federal do ParanáCuritibaBrazil
  4. 4.Departamento de Biologia Estrutural, Molecular e GenéticaUniversidade Estadual de Ponta GrossaPonta GrossaBrazil

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