Introduction and expression of an anti-dipteran toxin gene from B. thuringiensis in nodulating Rhizobia

Rhizobium-based inoculants are widely used in leguminous crops. Improved strains with dual functions (i.e, nitrogen fixing and insecticidal) could be successfully applied to organic pest management. These could also be used as seed protectants, reducing the use of chemical insecticides. The cryIVB gene from B. thuringiensis israelensis coding for an anti-dipteran toxin was used as a model for expression. A highly stable plasmid vector (pTR101) was chosen in order to carry out nodulation experiments in the absence of selective pressure. Constructions were introduced into Rhizobium fredii 191 and meliloti 1021, through triparental matings with a helper plasmid. Expression was monitored by Western blotting of crude extracts using a specific antibody raised against recombinant CryIVB protein. Although rearrangements leading to deletions were observed, an immunoreactive polypeptide was reproducibly detected in both Rhizobia. The presence of the recombinant plasmids did not affect the nodulative capacity nor the growth of the inoculated plants.