Effects of Rhizobium tropici azide-resistant mutants on growth, nitrogen nutrition and nodulation of common bean (Phaseolus vulgaris L.)

Inoculation with Rhizobium tropici and Rhizobium freirei can partially supply the nitrogen demand required for normal development and high productivity of common bean (Phaseolus vulgaris L.). Here, a simple and low-cost approach was used to select R. tropici sodium azide-resistant mutants with improved efficiency as microsymbiont of common bean. We mutagenized R. tropici CIAT889 by treatment with methyl methanesulfonate (MMS) and selected azide-resistant mutants. Six mutants (AzR14, AzR15, AzR16, AzR17, AzR18, and AzR19) with varying levels of resistance were obtained and characterized for symbiotic traits and performance upon inoculation of common bean. Mutants grew and produced exopolysaccharides similar to the CIAT899 wildtype; however, swarming motility, biofilm, and auxin production were altered. About nodulation, shoot biomass, and nitrogen content, the performance of mutant strains was improved or similar to that of CIAT899. All mutants enhanced nodulation of common bean; furthermore, AzR14, AzR18, and AzR19 increased nodule biomass by 25%-36%. AzR18 and AzR19 increased shoot biomass by up to 13% compared to CIAT899, whereas AzR14, AzR15, AzR16, and AzR17 reduced biomass accumulation by 9%-19%. In addition, AzR18 and AzR19 showed higher motility or biofilm production than the other mutant strains, suggesting that these traits are involved in bacterial symbiotic effectiveness. Shoot N content do not differ between plants inoculated with the CIAT899 or mutant strains, although plants inoculated with AzR18, AzR19 and CIAT899 had increased the shoot N as compared to uninoculated plants grown at N-limiting conditions. Our results demonstrated the possibility to improve the performance of the R. tropici microsymbiont through induced mutagenesis followed by the selection of azide resistant mutants. This approach allowed to identify the AzR18 and AzR19 strains, which showed superior nodulation and symbiotic effectiveness than CIAT899 while providing equivalent amounts of N to the inoculated plants. We propose the AzR18 and AzR19 as promising candidates for developing new and more efficient inoculants for common bean, nevertheless further trials under field conditions are required to confirm their superior performance to the wild strain.