**AWARDS ISSUED PRIOR TO JANUARY 20, 2025, WERE FUNDED UNDER PREVIOUS ADMINISTRATIONS AND MAY NOT REFLECT THE PRIORITIES AND POLICIES OF THE CURRENT ADMINISTRATION.** AS CASH, FORAGE, AND COVER CROPS, LEGUMES PLAY A CRUCIAL ROLE IN GLOBAL NUTRITION AND FOOD SECURITY. LEGUMES ESTABLISH BENEFICIAL ASSOCIATIONS WITH RHIZOBIAL BACTERIA WHO PERFORM BIOLOGICAL NITROGEN FIXATION (BNF)--PROVIDING NITROGEN (N) FERTILIZER TO PLANTS WITHOUT THE ECONOMIC AND GREENHOUSE GAS EMISSION COSTS OF CHEMICAL N INPUTS. HOWEVER, THIS MUTUALISM IS FAR FROM OPTIMIZED IN AGRICULTURAL SYSTEMS. CROP BENEFITS ARE HIGHLY VARIABLE AND CONTEXT-DEPENDENT AND EXISTING INOCULANTS OFTEN FAIL TO SURVIVE OR COMPETE IN AGRICULTURAL SOILS. IN THE NORTHEASTERN UNITED STATES, THE PERENNIAL FORAGE AND COVER CROP ALFALFA IS A CRITICAL COMPONENT OF CONVENTIONAL AND ORGANIC AGROECOSYSTEMS. THIS PROPOSAL LEVERAGES EXISTING ALFALFA VARIETY TRIALS TO ASSESS THE ABILITY OF ALFALFA (MEDICAGO SATIVA) TO STEER THE EVOLUTION OF THEIR MUTUALISTIC PARTNERS IN THE FACE OF ENVIRONMENTAL VARIATION. BY APPLYING NOVEL SEQUENCING TECHNOLOGIES PREVIOUSLY VALIDATED IN MEDICAGO TRUNCATULA TO FIELD TRIALS, WE WILL MEASURE THE EXTENT AND GENOMIC BASIS OF RHIZOBIAL ADAPTATION TO ALFALFA VARIETIES ACROSS SEASONS AND YEARS. BY CONDUCTING PAIRED MULTI- AND SINGLE-STRAIN GREENHOUSE EXPERIMENTS USING ISOLATES FROM THE TRIAL, WE WILL ASSESS STRAIN COMPETITION AND PLANT N-FIXING BENEFITS TO ENABLE SCREENING FOR ALFALFA VARIETIES' ABILITY TO REWARD HELPFUL RHIZOBIA. THIS PROPOSAL WILL BENEFIT 1) LEGUME BREEDERS BY ESTABLISHING A SCREENING PIPELINE AND 2) ORGANIC AND CONVENTIONAL GROWERS BY CONTRIBUTING TO THE SELECTION OF LEGUME VARIETIES. ULTIMATELY, LEVERAGING LEGUME HOSTS TO SELECT ON EXISTING RHIZOBIA REPRESENTS A NOVEL, COST-EFFECTIVE AVENUE TO PROMOTE EFFICIENT N-FIXATION, THEREBY SUPPORTING ROBUST YIELDS, IMPROVING SOIL HEALTH, AND DECREASING CHEMICAL N-INPUTS AND GREENHOUSE GAS EMISSIONS.