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Mendoza-Suárez, Marcela A.; Geddes, Barney A.; Sánchez-Cañizares, Carmen; Ramírez-González, Ricardo H.; Kirchhelle, Charlotte; Jorrin, Beatriz; Poole, Philip S.
Proceedings of the National Academy of Sciences - PNAS, 05/2020, Volume: 117, Issue: 18Journal Article
Legumes tend to be nodulated by competitive rhizobia that do not maximize nitrogen (N₂) fixation, resulting in suboptimal yields. Rhizobial nodulation competitiveness and effectiveness at N₂ fixation are independent traits, making their measurement extremely time-consuming with low experimental throughput. To transform the experimental assessment of rhizobial competitiveness and effectiveness, we have used synthetic biology to develop reporter plasmids that allow simultaneous high-throughput measurement of N₂ fixation in individual nodules using green fluorescent protein (GFP) and barcode strain identification (Plasmid ID) through next generation sequencing (NGS). In a proof-of-concept experiment using this technology in an agricultural soil, we simultaneously monitored 84 different Rhizobium leguminosarum strains, identifying a supercompetitive and highly effective rhizobial symbiont for peas. We also observed a remarkable frequency of nodule coinfection by rhizobia, with mixed occupancy identified in ~20% of nodules, containing up to six different strains. Critically, this process can be adapted to multiple Rhizobium-legume symbioses, soil types, and environmental conditions to permit easy identification of optimal rhizobial inoculants for field testing to maximize agricultural yield.
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