** AWARDS ISSUED PRIOR TO JANUARY 20, 2025, WERE FUNDED UNDER PREVIOUS ADMINISTRATIONS AND MAY NOT REFLECT THE PRIORITIES AND POLICIES OF THE CURRENT ADMINISTRATION.** CITRUS IS ONE OF THE MOST IMPORTANT FRUIT CROPS IN THE U.S. AND IN THE WORLD WITH MANY CULTIVATED VARIETIES. THE PRODUCTION OF CITRUS IN THE U.S. HAS BEEN DEVASTED BY THE BACTERIAL DISEASES HUANGLONGBING (HLB), AND TO SOME EXTENT BY CITRUS CANKER. WE HAVE GENERATED MULTIPLE CITRUS GENOME-EDITED LINES TARGETING SIX SELECTED HOST PLANT SUSCEPTIBILITY (S) GENES TO ACHIEVE RESISTANCE AGAINST THESE DEVASTATING BACTERIAL DISEASES. BECAUSE COMMERCIAL CITRUS HAS TO BE CLONALLY PROPAGATED, GENE-EDITED CITRUS PLANTS WILL HAVE STABLY INCORPORATED CRISPR REAGENTS IN THEIR GENOMES THROUGH GENERATIONS. THIS INDICATES THAT THERE IS A POSSIBILITY OF CONTINUED CRISPR ACTIVITY AFFECTING BOTH GENOTYPE AND PHENOTYPE OF GENE-EDITED CITRUS LINES. THEREFORE, RISK-ASSESSMENT OF GENE-EDITING TECHNOLOGY FOR ENGINEERING CITRUS SPECIES IS IMPERATIVE TO MEET THE URGENT NEED FROM THE U.S. CITRUS INDUSTRY. WE PROPOSE TO PERFORM COMPREHENSIVE EVALUATION OF GENOME-WIDE OFF TARGET EFFECTS AND ASSOCIATED PHENOTYPES IN THESE CITRUS GENE-EDITED LINES COMPARED TO NON-MODIFIED CONTROLS IN FIELD STUDIES. FURTHER, WE WILL ALSO EVALUATE THE STABILITY OF EDITS AND PHENOTYPES OVER MULTIPLE YEARS UNDER FIELD CONDITIONS. THE RESULTS FROM THIS PROJECT WILL PROVIDE CITRUS GERMPLASM TO COMBAT BACTERIAL DISEASES AND ASSIST REGULATORY AGENCIES TO EFFECTIVELY AND COMPREHENSIVELY ASSESS THE RISKS OF GENE-EDITING IN CITRUS AT BOTH GENOTYPIC AND PHENOTYPIC LEVELS.