**AWARDS ISSUED PRIOR TO JANUARY 20, 2025, WERE FUNDED UNDER PREVIOUS ADMINISTRATIONS AND MAY NOT REFLECT THE PRIORITIES AND POLICIES OF THE CURRENT ADMINISTRATION.** SPOTTED WING DROSOPHILA IS A SMALL FRUIT FLY THAT CAN NOW BE COMMONLY FOUND ACROSS THE MAJOR FRUIT GROWING REGIONS OF THE UNITED STATES. THE MALE FRUIT FLIES CAN BE EASILY DIFFERENTIATED FROM OTHER FRUIT FLIES BY A DARK SPOT THAT IS ON EACH WING, WHICH GIVES THE COMMON NAME OF THE SPECIES. ALTHOUGH NOW COMMON, IT WAS NOT ALWAYS THIS WAY. SPOTTED WING DROSOPHILA IS A NATIVE OF ASIA AND WAS NOT FOUND ANYWHERE IN THE USA PRIOR TO 2008. FEMALES HAVE A HIGHLY DEVELOPED EGG LAYING APPARATUS WHICH MEANS THEY CAN INFEST A WIDE RANGE OF HOST FRUIT BEFORE HARVEST. GROWERS ARE CURRENTLY APPLYING BROAD SPECTRUM INSECTICIDES MORE FREQUENTLY THAN IN THE PAST TO PROTECT FRUIT, BUT THE EFFECTIVENESS OF THE SPRAYS IS WEATHER DEPENDENT. RESISTANCE TO INSECTICIDES HAS BEEN REPORTED IN CALIFORNIA. NEW CONTROL TACTICS ARE NEEDED.GENETIC TECHNOLOGIES OFFER A SPECIES-SPECIFIC ENVIRONMENTALLY-FRIENDLY METHOD FOR PEST CONTROL.WE HAVE DEVELOPED MALE-ONLY STRAINS OF SPOTTED WING DROSOPHILA THAT PRODUCE ONLY MALES ON DIET THAT LACKS THE ANTIBIOTIC TETRACYCLINE. WHEN MALES CARRYING THE FEMALE KILLING GENE MATE WITH A WILD TYPE FEMALE, ALL THEIR FEMALE OFFSPRING THAT INHERIT THE FEMALE KILLING GENE DIE. WE HAVE SHOWN THAT REPEATED RELEASES OF THE MALES LEAD TO SUPPRESSION OF LABORATORY CAGE POPULATIONS OF SPOTTED WING DROSOPHILA, DRIVING THE CAGES TO EXTINCTION. WE HAVE ALSO DEVELOPED A GENE DRIVE STRAIN OF SPOTTED WING DROSOPHILA WITH THE DRIVE TARGETING A GENE ESSENTIAL FOR FEMALE DEVELOPMENT. IF A MALE WITH THE GENE DRIVE MATES WITH A WILD TYPE FEMALE, ABOUT 70% OF THE FEMALE OFFSPRING INHERIT THE DISRUPTED FEMALE-ESSENTIAL GENE. WITHOUT DRIVE, ONLY 50% OF THE OFFSPRING WILL INHERIT THE DISRUPTED GENE. OUR FIRST OBJECTIVE IS TO IMPROVE DRIVE EFFICIENCY THROUGH MODIFICATIONS TO THE CRISPR/CAS9 SYSTEM THAT IS THE BASIS OF THE DRIVE. THE AIM IS TO INCREASE GERMLINE (TESTES) EXPRESSION OF CAS9 AND INCREASE THE FREQUENCY OF THE GENE COPYING MECHANISM THAT LEADS TO DRIVE. IF SUCCESSFUL, WE ANTICIPATE THAT MORE THAN 70% OF THE OFFSPRING OF DRIVE MALES WILL CARRY THE DISRUPTED FEMALE-ESSENTIAL GENE.THE OTHER OBJECTIVES OF OUR PROJECT DIRECTLY RELATE TO PROGRAM 3 (C) ASSESSING THE INFLUENCE OF GENETIC BACKGROUND ON THE EXPRESSION OF AND PHENOTYPES CONFERRED BY GENETIC MODIFICATION. RECENT WORK FROM OUR LAB AND OTHERS HAVE SHOWN THAT DIFFERENCES IN GENETIC BACKGROUND CAN PROFOUNDLY INFLUENCE THE EFFICACY OF MALE-ONLY AND GENE DRIVE STRAINS. FURTHER, GENOMIC ANALYSES INDICATE THAT THERE ARE SIGNIFICANT GENETIC DIFFERENCES BETWEEN SPOTTED WING DROSOPHILA POPULATIONS IN THE USA, PARTICULARLY EAST AND WEST COAST POPULATIONS. OBVIOUSLY, A MALE-ONLY STRAIN THAT IS ONLY EFFECTIVE IN NORTH CAROLINA WOULD NOT BE AS USEFUL AS A STRAIN THAT COULD SUPPRESS FLIES IN ANY REGION OF THE COUNTRY. WE WILL OBTAIN SPOTTED WING DROSOPHILA FROM DIFFERENT AREAS OF THE USA AND TEST THE POTENTIAL OF THE MALE ONLY AND GENE DRIVE STRAINS FOR POPULATION SUPPRESSION IN THE DIFFERENT GENETIC BACKGROUNDS. THE INFORMATION GAINED FROM THESE EXPERIMENTS WILL BE VALUABLE FOR REGULATORS IN ASSESSING FUTURE FIELD RELEASE(S) OF THE SPOTTED WING DROSOPHILA STRAINS.