Isolation and identification of root associated diazotrophs

Diazotrophs have been isolated from the rhizosphere or roots of plants by many workers. To recognize a certain diazotroph as the most abundant bacterium at a certain site or as the principal agent responsible for N₂-fixation is much more difficult. It is probable that many diazotrophs, including possibly the most efficient ones, have not been identified yet. The use of proper selective media which simulate the environment of the various diazotrophs in situ has led to the discovery of 10 new root-associated diazotrophs, three of them during 1986/1987 (Azospirillum halopraeferans, Herbaspirillum seropedicae and the recently proposed Acetobacter diazotrophicus). The importance of using a variety of carbon substrates in the growth media with pH indicators, and the use of N-free semi-solid media, is discussed. Recognition of plant-bacteria interactions requires, in addition to the identification of the bacteria, the demonstration of effects of the plant on the bacteria and of the bacteria on the plant. Confirmation of the identity of diazotrophs responsible for response of plants to inoculation must be made in experiments with strains labelled with antibiotic resistance or other markers. If establishment of the inoculated strain is demonstrated in plants grown in ¹₅N-labelled soil, the ¹₅N enrichment of the plants will reveal if any observed responses in N yield are due to N₂-fixation or increased soil/fertilizer-N uptake.