A continuum of ecology and evolution contributes to mutualism breakdown between legumes and rhizobia

Ecological and evolutionary processes jointly shape community responses to environmental perturbations, yet are often examined separately - even in microorganisms, where they occur on similar timescales and are linked by mobile genetic elements like plasmids. Mutualism breakdown is a pressing challenge to examine this intersection, as shifts in diverse microbial communities might coincide with evolutionary changes in mutualistic traits within a key species. Here we study the legume-rhizobium mutualism after 33 years of nitrogen fertilization. Pairing manipulative inoculation studies with long-read amplicon sequencing provides unprecedented resolution across biological scales: whole bacterial community, genus Rhizobium, individual Rhizobium ASVs, and symbiosis plasmids. Clover host plants in N-fertilized plots encounter microbial communities depauperate in their preferred symbiont but enriched in other Rhizobium taxa; moreover, their symbiotic nodules house low-quality symbiosis plasmids. Thus, concurrent ecological and evolutionary shifts make mutualists both rarer and inferior and feed back to decrease the benefits hosts receive from mutualism.