Evaluation of a strategy for identifying nodulation competitiveness genes in Rhizobium leguminosarum biovar phaseoli

Rhizobium leguminosarum biovar phaseoli strain KIM5s is consistently much more competitive than strain CE3 in nodulation of beans (Phaseolus vulgaris L.) in the laboratory and in the field. To identify genes that contribute to the competitiveness of KIM5s, we transferred a cosmid library containing KIM5s DNA into CE3 and applied the transconjugants to bean plants to allow the plants to enrich for those with enhanced nodulation competitiveness. The nodule isolates were then applied to plants for further enrichment. Of 75 isolates from nodules sampled after the two enrichments, 9 were more competitive than CE3. For example, when outnumbered in the inocula 40-fold by a reference strain, these nine strains typically occupied 15-40% of the nodules compared with 0-3% for CE3. However, when these strains were cured of the cosmids, they remained highly competitive, demonstrating that the enhanced competitiveness of the strains was not associated with the cosmids. We found no evidence for cosmid insertion into the chromosome or for cosmid-induced genetic changes in these cured strains. We found some evidence suggesting that their altered competitiveness was due to spontaneous genetic changes that did not involve the cosmids. Although these highly competitive variants remain genetically uncharacterized, they may provide insight into bacterial traits that contribute to, or detract from, successful nodulation competitiveness.