Role of Two Flagella of Bradyrhizobium japonicum in Competition for Nodulation of Soybean

M. JULIA ALTHABEGOITI; JULIETA M. COVELLI; M. FLORENCIA LÓPEZ; ANÍBAL R. LODEIRO

San Miguel de Tucumán

Congreso; VII Congreso Argentino de Microbiología General; 2011

The agronomic use of Bradyrhizobium japonicum in symbiosis with soybean is via inoculation ofseeds or seed furrow with rhizobia. As a result, the roots of soybean plants develop nodulesthat once became occupied by B. japonicum can fix atmospheric N2 to NH4+, reducing it to an extent that would satisfy all the needs of the plant. Therefore, the capacity of rhizobia to movefrom the soil or from seeds to the sites of infection on the roots results determinant for theestablishment of the symbiosis and consequently motility of B. japonicum is very interesting tostudy.B. japonicum has two flagellar systems. One system is composed of a 33 kDa flagellin (thinfilament) and the other consists of a 65 kDa flagellin (thick filament); it has been proposed thatboth play different roles (Kanbe et al., 2007). From the parental strains B. japonicum LP 3004(derived from USDA 110, Sm-resistant) and LP 3008 (LP 3004 with an increased motility)(Althabegoiti et al., 2008), we generated mutants lacking thin flagellum (LP 6865 and LP 6866),mutants lacking thick flagellum (LP 5843 and LP 5844) and mutants that lack flagella (LP 6543y LP 6644), respectively. Strains were observed by TEM and mutations and morphology wereconfirmed for each type of flagellum. After that, liquid cultures of each mutant were observed ina phase-contrast microscope and only strains that have thick filament were capable of swimstraightly whereas strains with thin filament tumbled more frequently. Finally, those that nothave flagella did not swim.With the purpose of studying contribution of each flagellum in colonization of soybean rootscompetition experiments were done. Competitiveness was examinated at field capacity andsoybean was inoculated with mixtures containing each parental strain together with eachderived mutant. After twenty one days of growth in greenhouse the occupation of plant noduleswas determinated by their antibiotic resistance and a statistical analysis was done. As a result,we observed that mutants lacking thick flagellum and non-motile double mutants were lesscompetitive that parental strains, but there were not completely displaced. However, mutantslacking thin flagellum were more competitive for nodulation. This event suggests that thinfilament is not necessary for B. japonicum movement to the roots, showing a complex role ofthese flagellins in competitiveness. They could also have an additional function involved ininteraction with the plant, opening interesting aspects for further analysis.