** AWARDS ISSUED PRIOR TO JANUARY 20, 2025, WERE FUNDED UNDER PREVIOUS ADMINISTRATIONS AND MAY NOT REFLECT THE PRIORITIES AND POLICIES OF THE CURRENT ADMINISTRATION.** RECENTLY, THIS TEAM EXPERIMENTALLY DEMONSTRATED, FOR THE FIRST TIME, THAT HORIZONTAL GENE TRANSFER (HGT) EVENTS CAN INDEED OCCUR FROM GENETICALLY ENGINEERED (GE) CYANOBACTERIA TO A COMMON LABORATORY STRAIN OF E. COLI. THE NEXT IMPORTANT STEP IS TO ASSESS WHETHER HGT COULD OCCUR FROM GE CYANOBACTERIA TO OTHER MICROORGANISMS INCLUDING SOIL-ENVIRONMENTAL RELATED BACTERIA SUCH AS SERRATIA MARCESCENS, PSEUDOMONAS PUTIDA, BACILLUS LICHENIFORMIS AND RISK GROUP-2 ORGANISMS, SUCH AS STAPHYLOCOCCUS AUREUS, CAMPYLOBACTER JEJUNI, SALMONELLA SP, AND PSEUDOMONAS AERUGINOSA, WHICH ARE WIDESPREAD IN NATURE AND COULD POSE A SERIOUS HEALTH RISK TO HUMANS AND ANIMALS IF THEY WERE TO OBTAIN ANTIBIOTIC RESISTANCE THROUGH HGT FROM GE ORGANISMS. THEREFORE, IN THE PROPOSED PROJECT, THIS TEAM WILL MEASURE THE RATES OF HGT FROM GE CYANOBACTERIA TO SOIL-ENVIRONMENTAL BACTERIA/RISK GROUP-2 MICROORGANISMS. THE TEAM WILL ALSO CHARACTERIZE AND ELUCIDATE THE HGT MECHANISMS, SUCH AS TO WHETHER THE HGT IS THROUGH A DIRECT CELL-TO-CELL INTERACTION OR BY AN INDIRECT ROUTE OF CELL-TO-LIQUID ENVIRONMENT (EDNA)-TO-CELL PROCESS, USING A NEWLY DEVELOPED PROTOCOL FOR MEASURING HGT RATES FROM GE ALGAE TO NON-PHOTOSYNTHETIC ORGANISMS. THE TEAM WILL FURTHER ASSESS THE FATE/STABILITY OF THE TRANSGENES AND THE SURVIVABILITY OF GE CYANOBACTERIA IN COMPARISON WITH THEIR RELATED WILD-TYPE ORGANISMS. THE TEAM WILL ACHIEVE EXPERIMENTAL ASSESSMENT ON THE SURVIVABILITY AND HGT OF GE CYANOBACTERIA WITH THE NATIVE MICROORGANISMS IN SOIL-POND ENVIRONMENTAL WATER SAMPLES. THE PROPOSED PROJECT WILL DIRECTLY SUPPORT THE BIOTECHNOLOGY RISK ASSESSMENT RESEARCH GRANTS (BRAG) PROGRAM IN THE PROGRAM AREA 3. GENE TRANSFER BETWEEN GENETICALLY ENGINEERED ANIMALS, PLANTS, AND MICROORGANISMS AND RELATED WILD AND AGRICULTURAL ORGANISMS. THIS PROJECT WILL YIELD QUANTITATIVE RATES OF HGT FROM GE CYANOBACTERIA TO SOIL-ENVIRONMENTAL BACTERIA/RISK GROUP-2 MICROORGANISMS AND BETTER UNDERSTANDING OF POSSIBLE HGT ROUTES THAT WILL BE HIGHLY VALUABLE TO THE FEDERAL REGULATORY AGENCIES RELATED TO BIOTECHNOLOGY TO MAKE SCIENTIFIC EVIDENCE-BASED POLICY.