In acid soils, Rhizobium favelukesii strains, known as Oregon-like strains, are a potential risk for alfalfa production given their parasitic behaviour. In this study, we isolated five parasitic strains (ORY1 to ORY5) from alfalfa nodules grown in Uruguayan acid soils, with a 99.7% and a 100% 16S rRNA gene sequence identity to R. favelukesii type strain of LUP83. The BOX profiles of the five isolates showed two different patterns, suggesting some diversity among these acid-tolerant isolates. The genome sequence analysis of R. favelukesii strains ORY1, LPU83, and Or191 showed that they have around 87.5% of common coding genes, including the symbiotic genes. Moreover, the phylogenetic analysis of ORY1 symbiotic genes nifH, nifD, nifK, nodA, nodB, and nodD were related to the symbiotic genes of E. meliloti. We teste ORY1 competitiveness by inoculating seeds with 99:1 and 1:99 ratios of ORY1::gusA/E. meliloti U143. In both treatments, ORY1::gusA occupied more than 50% of nodules, evidencing its high competitiveness. However, the aerial biomass in these treatments was remarkably different, suggesting that the nodules induced by the efficient strain are essential to provide enough N for optimal plant growth. These findings support the needing of inoculating in areas where inefficient strains are likely to be present. Finally, we found three genes that encode amino acid sequences for domains of M16 peptidases (with homology to bacterial hrrP and sapA genes), two of them were contiguous and located in an accessory plasmid, whereas the other one was a chromosomal gene. These genes are likely to be involved in the parasitic behaviour of ORY1 strain.

In Uruguayan soils, populations of native and naturalized rhizobia nodulate white clover. These populations include efficient rhizobia but also parasitic strains, which compete for nodule occupancy and hinder optimal nitrogen fixation by the grassland. Nodulation competitiveness assays using gusA-tagged strains proved a high nodule occupancy by the inoculant strain U204, but this was lower than the strains with intermediate efficiencies, U268 and U1116. Clover biomass production only decreased when the parasitic strain UP3 was in a 99:1 ratio with U204, but not when UP3 was at equal or lower numbers than U204. Based on phylogenetic analyses, strains with different efficiencies did not cluster together, and U1116 grouped with the parasitic strains. Our results suggest symbiotic gene transfer from an effective strain to U1116, thereby improving its symbiotic efficiency. Genome sequencing of U268 and U204 strains allowed us to assign them to species Rhizobium redzepovicii, the first report of this species nodulating clover, and Rhizobium leguminosarun, respectively. We also report the presence of hrrP- and sapA-like genes in the genomes of WSM597, U204, and U268 strains, which are related to symbiotic efficiency in rhizobia. Interestingly, we report here chromosomally located hrrP-like genes.