Uptake of vegetative cells of Azotobacter vinelandii into protoplasts of the mycorrhizal fungus Rhizopogon sp. can be induced by treatment with
polyethylene glycol (molecular weight, 6000). An L-form of the bacteria has been selected for within the differentiated fungal mycelium which is capable of
acetylene reduction and nitrogen fixation, as confirmed by
nitrogen-15 assays; this allows the fungus to grow on media lacking any combined
nitrogen. The fungus grows and reduces
acetylene on concentrations of
antibiotics that prevent the growth and activity of free-living Azotobacter. Electron microscopy has revealed modified mitochondrial forms or included bacterial L-forms surrounded by an extra fungal membrane within the hyphae of the modified strains.
Poly-beta-hydroxybutyric acid, a storage product of Azotobacter
cysts, has also been identified in the hyphae. This would appear to be the first report of the transgenosis for
acetylene reduction activity and nitrogen fixation into a eukaryote cell.