Nodulation competitiveness as a requisite for improved rhizobial inoculants of Trifolium pratense

Red clover (Trifolium pratense L.) is used in the improvement of grasslands in Uruguay and has been inoculated with commercial strain U204 of Rhizobium leguminosarum bv trifolii since 1970s. Native-naturalized rhizobia strains present in soil are the basis for selecting and developing new inoculants. With this aim, we evaluated the diversity of red clover rhizobia in Uruguayan red clover pastures both historically inoculated with U204 and non-inoculated ones. Thirty-eight different enterobacterial repetitive intergenic consensus (ERIC) PCR genomic fingerprints were identified, albeit surprisingly only one of 80 isolates showed an ERIC profile similar to U204. Under controlled conditions, red clover plants inoculated with one of the native isolates, strain 317, produced more biomass than those inoculated with the commercial U204. ERIC-PCR was also used to show that strain 317 competed for nodulation better than U204 in a field with previous history of inoculation. Moreover, both U204 and 317 were tagged with a gusA reporter gene and their competitiveness for nodulation assessed in various soil types. Again, strain 317 appeared more competitive than U204, particularly in soils with previous history of inoculation. Our results reinforce the long-known idea of assessing the actual needs of inoculation of legumes in different soils and suggest that the indigenous isolate 317 is an effective and competitive strain that can be used for development of a new red clover inoculant.