Title : Strategies to improve heavy metal tolerance in legumes: metal-tolerance genes and the role of tolerant rhizobia
Legumes play a key role in sustainable agriculture. Mineral nitrogen deficiency is an important limiting factor for plant growth in arid and semi-arid regions, and rhizobia-legume symbioses are the primary source of fixed nitrogen in such areas. The introduction of legumes and their nodulating rhizobia may have an important effect on the reclamation of degraded and polluted marginal soils for sustainable agriculture. Such recovery is becoming an urgent matter due to the increasing extension of affected lands and the ever-rising requirements for food and feed. Heavy metal contamination is increasing worldwide in both wild ecosystems and agricultural soils due to natural processes, but mostly to anthropic activities, and they can enter the food chain by being taken up by plants. It becomes of the greatest interest to obtain legume varieties and bacterial inocula with enhanced tolerance to heavy metals for use in soil reclamation, which can be achieved by traditional trait selection or by biotechnological procedures. We will present our results on the selection of tolerant legume cultivars and rhizobial strains, as well as our biotechnological approaches to obtain legumes and rhizobia with improved tolerance to heavy metals. Understanding the genetic basis of metal accumulation and tolerance in plants is important for reducing the uptake of toxic metals in crops, as well as for removing heavy metals from soils by means of phytoremediation. Following exposure of Medicago truncatula seedlings to cadmium (Cd) and mercury (Hg), we conducted a genome-wide association study (GWAS) using relative root growth (RRG) and leaf accumulation measurements that allowed us to identify genes involved in heavy metal tolerance and accumulation. It is known that metal-tolerant symbiotic rhizobia have the potential to increase legume metal tolerance. We isolated from Hg-contaminated soils several Ensifer medicae strains that nodulate M. truncatula. We assembled and annotated the genomes of those rhizobia strains that showed wide variation in tolerance to Hg, and found structural variations in mercury reductase (merA) and alkylmercury lyase (merB), which are involved in Hg detoxification, and entire mer operons that were associated with the most Hg-tolerant strains. Genes in the mer operons and duplicated merA copies throughout the genomes showed significantly higher gene expression in the tolerant vs. less tolerant rhizobia strains. In the most tolerant E. medicae strain, a whole mer operon was located in a large additional 71-kb plasmid, which was not present in any other strain. Plasmid transfer to non-tolerant bacterial strains arises as a possibility to obtain increased Hg tolerance.