Mesorhizobium species are soil bacteria (rhizobia) that form endosymbiotic relationships with legumes such as Biserrula pelecinus through the infection of plant roots and development of root nodules. Rhizobia differentiate into bacteroids and activation of nitrogenase enzyme occurs, allowing atmospheric nitrogen (N2) to be reduced to ammonia and secreted to the host plant.
In Mesorhizobium spp., genes essential to nodulation and nitrogen fixation are located on mobile chromosomal elements or “symbiosis islands”. Previous work at the Centre for Rhizobium Studies revealed that when the highly effective N2-fixing microsymbiont of B. pelecinus, M. ciceri WSM1271 was inoculated into a field site in Northam, it transferred its symbiotic island to non-symbiotic Mesorhizobia, giving rise to new B. pelecinus microsymbionts M. oppurtunistum WSM2073 and M. australicum WSM2075. Crucially, while all strains share identical symbiosis islands, their N2 fixation efficiency on B. pelecinus differs significantly. Analysis of the full genome sequences found that WSM1271 carries an accessory plasmid, pMESCI01, which is absent in WSM2073 and WSM2075. Initial bioinformatic analysis revealed that pMESCI01 contains some putative N2-fixation genes, which may be essential to highly effective symbioses with B. pelecinus.
This project will investigate the cause of N2 fixation efficiency differences between these three strains. First, WSM1271 will be cured of its accessory plasmid by cloning the repABC region from pMESC01 into sacB-containing vectors pJQ200SK and pSRK-nptII. Subsequent constructs will be mobilized into WSM1271 to cure due to plasmid incompatibly. Analysis of symbiotic phenotype of the resultant mutant against wild-type WSM1271 will take place. Second, the physiological nature of the differences in symbiotic effectiveness between all strains on B. pelecinus will be explored through measurement of rates of N2 fixation on a per plant, per nodule and per bacteroid basis, to determine at which stage of the symbiotic interaction efficiency differences lie.
Ultimately, the findings of this project will provide crucial data to understand the basis of suboptimal N2 fixation in legume-Mesorhizobium symbioses.