Integrated multi-omics decipher the complex nodule microbiota and distinct Frankiaceae symbiotic traits in wild actinorhizal plants.

Actinorhizal plants are ecologically important pioneer species in temperate regions, capable of nitrogen-fixing root nodule symbiosis with Frankiaceae bacteria. Despite their significance within the nitrogen-fixing clades (NFC), multi-omics studies of actinorhizal symbiosis remain scarce. We profiled prokaryotic communities in the rhizosphere, root, and/or nodule compartments from five phylogenetically representative actinorhizal species, three legumes, and four nonnodulated NFC species using 16S rDNA sequencing. Transcriptomic and metagenomic analyses were performed on actinorhizal roots and nodules, respectively. Metagenome-assembled genomes revealed four novel Frankiaceae species. Frankiae relative abundance levels in nodules were generally lower than rhizobia in legumes. Actinorhizal nodules harbour diverse bacterial taxa, which exhibit predominantly positive interactions, with Frankiae forming a tightly interacting subgroup. Actinorhizal plants engage actively with soil microbiota, recruiting a specific rhizosphere community enriched with beneficial microbes, including ammonia-oxidising archaea. Many symbiotic mechanisms in nodulating host plants are conserved and derived from pre-existing molecular modules. Our analysis suggests the phosphoinositide signalling likely functions in actinorhizal symbiotic signal transduction. However, Frankiae exhibit fundamentally different symbiotic functional characteristics compared to rhizobia, reflecting less intimate symbiosis, which might favour the life-history strategies of temperate perennial actinorhizal plants.