The Biological Nitrogen Fixation process can be carried out only by procariotic microorganisms known as diazotrophs that can colonize the plant interior systemically without causing apparent damage to their hosts. Many endophytic diazotrophs can also produce phytohormones, promoting plant growth.
We have used the Salkowski assay to detect IAA (indolacetic acid) and its derivatives in tryptophan-containing culture supernatants of endophytic diazotrophs from grass, pineapple, banana and palm tree. Among these, there were bacteria belonging to the Azospirillum spp, Herbaspirillum spp, Ochrobactrum spp, Burkholderia spp. genera, and also strains of the symbiotic diazotrophs Rhizobium spp. and Sinorhizobium spp.
A total of 35 isolates (23 from banana; 7 from pineapple; and 5 from palm tree), 13 reference strains (3 from Herbaspirillum; 6 from Azospirillum; 3 from Rhizobium; and 1 from Sinorhizobium) were screened for IAA production. Herbaspirillum spp. produced about 18 to 25 ug of IAA/mL, Azospirillum spp. about 20 to 35 ug of IAA/mL, Rhizobium spp. 7 to 10 ug of IAA/mL, and Sinorhizobium 25 ug of IAA/mL. Roughly, the supernatant IAA detection accompanied the growth curve of the strains. Among diazotroph isolates, 24 produced detectable IAA concentrations in the supernatant, varying from 0.58 to 54.40 ug of IAA/mL. Azospirillum and Herbaspirillum like strains showed the highest production of IAA in a level similar to that of reference strains.
A growth curve for H. seropedicae, strain Z78, with IAA detection showed that IAA production begins when the bacteria enters its log phase, increasing rapidily. However, after that, the IAA concentration in the supernatant decreases slightly (even though the total amount remains high).
A Thin Layer Chromatography (TLC) of purified H. seropedicae strain M2 extract showed three resolved bands when the TLC was reveled with Salkowski reagent. One of the bands had similar migration with IAA standard. The others may be intermediates in the IAA production path way from tryptophan.