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Rhizobium galegae strain NBIMTC2250 contains two plasmids one of which (the larger one) is the symbiotic plasmid. To determine the participation of the smaller plasmid in cellular functions and in the symbiosis, we used a positive selection system for isolation of derivatives cured of this plasmid. The derivatives did not show differences from the wild type with respect to the following free-living cellular properties: antibiotic-resistance pattern, carbohydrates and organic acid utilization, growth rate and colony morphology in pure culture, lipopolysaccharide (LPS) profile, motility and sensitivity to extreme temperatures, which indicated that these functions were not plasmid encoded. Some of the cured derivatives differed from the wild type in the symbiotic characteristics, and the possible reason for the differences was discussed. In these cases reintroduction of the plasmid did not restore the derivatives to a wild type but the plasmid-free and plasmid-recontaining derivatives had similar symbiotic phenotype. Hybridization study showed that the smaller plasmid had been lost integrally in all cured derivatives. These results and the fact that almost half of the plasmid-cured derivatives have the symbiotic characteristics similar to those of the parent strain suggested that the smaller plasmid of strain Rhizobium galegae NBIMTC2250 did not participate in the symbiotically important functions of this strain. |
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