Actinomycetes different from Frankia induce effective nodulation in Alnus acuminata and can be genetically transformed

VALDÉS D; IMANISHI L; GABBARINI L; HUSS-DANELL K; WALL LG

Bariloche, Argentina

Congreso; 15th International Frankia and Actinorhizal Plants Meeting; 2008

Two actinomycetes isolated from one disinfected field nodule of Alnus acuminata, named AaI4 and AaI5, grow in the absence of N, reduce acetylene, have short and branched chains with bacillary spores, show segmented vegetative hyphae, and develop spherical structures like vesicles in N free medium. Fingerprint BOX PCR revealed diversity between the isolates and show identity between single-spore cultures and the original cultures suggesting homogeneity of the strains. The interaction with plants revealed that AaI4 and AaI5 deformed root hairs of A. acuminata roots, were able to induce cell division in the cortex and to develop scarce nodulation when inoculated at high doses, compared to control Frankia ArI3. The strain AaI5 could be re-isolated from nodules induced by this strain in Alnus acuminata seedlings. Either in free-living state or in planta both strains show positive acetylene reduction activity although attempts to reveal nif genes with different Co-inoculation assays suggest that the strains AaI4 and AaI5 can be physiologically complemented by a non-infective Frankia strain for nodulation of Alnus acuminata. Preliminary results using RFLP-PCR suggest the possibility of co-infection of nodules in the cases of co-inoculation of Alnus acuminata with the actinomycete AaI4 and Frankia BCU110501. Both strains of actinomycetes AaI4 and AaI5 could be transformed by electroporation with plasmid pIJ8641 and expressed gfp. On the basis of detailed morphological characterization of the isolates cultivated with or without N, and scanning electron microscopy analysis, we conclude that these isolates, AaI4 and AaI5 were not Frankia. We consider that these findings open the paradigm of the biology of actinorhizal symbiosis to new microorganisms.