A method for screening Phaseolus vulgaris L. germplasm for preferential nodulation with a selected Rhizobium etli strain

As part of a breeding program to improve the nitrogen-fixing symbiosis between common bean (Phaseolus vulgaris) and Rhizobium etli, we developed a rapid screen for common bean accessions that preferentially nodulate with KIM5s, a high nitrogen fixing strain of R. etli. We constructed a mutant of KIM5s that did not fix nitrogen (Fix⁻) but was otherwise indistinguishable from KIM5s. We screened plants for symptoms of nitrogen deficiency when grown in a Honduran soil containing indigenous common bean-nodulating rhizobia (10⁴ per gram) and KM6001, the Fix⁻ mutant of KIM5s (10⁴/seedling added 7 days after planting). Leaf color was scored on a scale of 1 to 5, in which 1 was dark green and 5 was bright yellow. Of 820 genetically diverse accessions of P. vulgaris screened, 51 were scored 1, 626 were scored 2 or 3, and 143 were scored 4 or 5. Selfed seed was produced from common bean plants of the accessions scored 1, 4 or 5. Twenty-four accessions that scored 1, and 58 that scored 4 or 5 were screened in soil containing indigenous rhizobia and the wild type KIM5s (Fix⁺), and nodule occupancy was determined by antibiotic resistance. On the 24 common bean accessions that were scored 1, KIM5s occupied 0-6% of the nodules, on 26 of the accessions that were scored 4 or 5, KIM5s occupied 90%-100% of the nodules, and on the remaining 34 that scored 4 or 5, there was a distribution of nodule occupancy. Foliar color was highly correlated with nodule occupancy (r = 0.786, p = 0.01). The results indicate that the rapid visual screen using the Fix⁻ mutant accurately identified common bean accessions that preferentially nodulate with the wild-type KIM5s (Fix⁺) strain in soil containing indigenous rhizobia. This screen will facilitate introduction of the preferential nodulation trait into superior cultivars and provides the foundation for studies of the genetic basis of preferential nodulation.