Critical parameters in facilitating the evolution of N₂-fixing symbiosis between diazotrophs and cereals

The key features of an experimental model for achieving significant biological N₂ fixation by associations between 'Azospirillum' and cereals are suggested to include adequate colonization, endophytic in nature to ensure both access to carbon substrates and suitable microaerobic oxygen conditions, and a means of ensuring adequate transfer of newly fixed nitrogen to the host plant. In our research program, we have exploited the property of synthetic plant hormones such as 2,4-dichlorophenoxyacetic acid to enhance access of azospirilla to protected niches such as the base of modified lateral roots ('para'-nodules) and channels between cortical cells. This approach has demonstrated the possibility of achieving such colonization. By using 'nifH-lacZ' fusions of 'A. brasilense' strains, we have been able to show a relationship between 'nifH' expression, oxygen pressure, and the magnitude of acetylene reduction rates in such associations. In addition, the significance of the 'flcA' (controlling flocculation) gene in effective colonization by regulating the expression of exopolysaccharides and the conversion of vegetative cells of azospirilla to cysts has been shown by the use of 'flcA'-mutants.