**AWARDS ISSUED PRIOR TO JANUARY 20, 2025, WERE FUNDED UNDER PREVIOUS ADMINISTRATIONS AND MAY NOT REFLECT THE PRIORITIES AND POLICIES OF THE CURRENT ADMINISTRATION.** CROP PATHOGENS AND PESTS REDUCE APPROXIMATELY 20-30% ANNUAL YIELD OF AGRICULTURAL PRODUCTION. THIS HAS LED TO SUBSTANTIAL ECONOMIC LOSSES AND REDUCTION IN FOOD SECURITY AT NATIONAL AND GLOBAL LEVELS. PLANT-DERIVED EXTRACELLULAR VESICLES (EVS) SECRETED DURING PLANT-PATHOGEN INTERACTIONS GAIN MORE AND MORE ATTENTION TO SUPPRESS THE EXPRESSION OF PATHOGEN'S VIRULENCE-RELATED GENES. THE OVERARCHING GOAL OF THIS SEED GRANT PROPOSAL IS TO PROVIDE AGRICULTURE SECTORS IN THE USA WITH ENVIRONMENTALLY FRIENDLY APPROACHES FOR BETTER PEST AND DISEASE CONTROL, CROP PROTECTION, AND AGRICULTURAL PRODUCTION, USING A SUSTAINABLE ECO-FRIENDLY GENE-SILENCING APPROACH. AS THE FIRST STEP, WE WILL CONDUCT A PROOF-OF-CONCEPT STUDY USING BIODEGRADABLE CHITOSAN NANOPARTICLES TO INCORPORATE THE C1C2 DOMAIN OF LACTADHERIN (LACTC1C2) VIA BIOTIN-STREPTAVIDIN BINDING TO REPLACE LABOR- AND TIME-INTENSIVE DIFFERENTIAL ULTRACENTRIFUGATION FOR THE ISOLATION OF PHOSPHATIDYLSERINE-EXPOSING PLANT EVS. TO DO SO, WE WILL FIRST DESIGN, SYNTHESIZE, AND CHARACTERIZE NANOPARTICLES INCORPORATING CHITOSAN, BIOTIN-STREPTAVIDIN COMPLEX, AND LACTC1C2 FOR THE PURIFICATION OF PLANT EVS. THEN, THE EFFICACY OF PREVENTING PATHOGEN INFECTION USING PURIFIED PLANT EVS TETHERED WITH NANOPARTICLES WILL BE INVESTIGATED. THE SPECIFIC AIMS OF THIS SEED GRANT PROJECT ARE TO 1) CONSTRUCT EXPRESSION VECTORS CONTAINING LACTC1C2-STREPTAVIDIN FUSION GENE FOR BACTERIAL TRANSFORMATION, PROTEIN EXPRESSION AND PURIFICATION, 2) FABRICATE LACTC1C2-STREPTAVIDIN-BOUND CHITOSAN NANOPARTICLES ASSOCIATED WITH EVS SECRETED FROM ARABIDOPSIS THALIANA LEAVES INFECTED WITH NECROTROPHIC FUNGUS BOTRYTIS CINEREA, AND 3) EXAMINE THE EFFICACY OF EV-NANOPARTICLE HYBRIDS ON SILENCING VIRULENCE GENES OF B. CINEREA. THE SUCCESSFUL OUTCOME OF THIS PROJECT WILL LAY THE FRAMEWORK FOR PD TO BECOME COMPETITIVE FOR FUTURE GRANTS FROM AFRI. ?