Title:

Co-Inoculation of Copper-Stressed Alnus glutinosa Seedlings with Frankia sp. QA3 and Frankia inefficax EuI1c

Alders can colonize and reclaim mining and industrial sites, whose soils are contaminated by high concentrations of heavy metals. These plants form a nitrogen-fixing mutualistic relationship with bacteria in the genus Frankia. Frankia associate with actinorhizal plants from eight families and allow these plants to overcome a range of environmental stresses including heavy metals, high salinity, and drought. Phylogenetic analysis of the genus Frankia has identified four distinct clades. Each clade is distinguished by their plant host range. Alders tend to associate only with clade 1 Frankia strains. However, in studies at polluted sites, Frankia clade 3 and 4 strains were identified in alder nodules. We were interested in determining if an environmental factor may play a role in inducing plant host flexibility in the interaction between host plant and symbiont in heavy metal polluted soils. To investigate, ten-week-old alder seedlings were inoculated with either a clade 1 Frankia strain (QA3), a clade 4 Frankia strain (EuI1c), or co-inoculated with both strains. Alders in each inoculation condition were treated with either 1 mM copper chloride or left untreated. For all alders, nodule formation was counted, and plant health observed. For co-inoculated alder seedlings, population dynamics within nodules will be measured via qPCR. It is expected that EuI1c will be present within nodules, though likely at a lower abundance than QA3. Preliminary data indicates that co-inoculated copper-stressed plants produced more nodules per plant than co-inoculated plants without copper-stress or plants inoculated with only one strain under both conditions. These results suggest that co-inoculation with EuI1c and QA3 under copper-stress may increase plant nodule production and consequently, copper tolerance. Work is currently in progress to produce knockout mutants of EuI1c copper-tolerance genes to determine the mechanisms Frankia strains use to tolerate environmental stressors and establish symbiosis with their hosts under these conditions.

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