NON-TECHNICAL SUMMARY 2017-05723WE ARE MAKING LIVE VACCINES TO PROTECT FISH FROM INFECTIOUS DISEASES THAT CAUSE SIGNIFICANT LEVELS OF DISEASE IN COMMERCIALLY GROWN FISH (CATFISH, TROUT, SALMON, TILAPIA, ETC.) AND ECONOMIC LOSSES TO THE AQUACULTURE INDUSTRY. INITIALLY WE WILL FOCUS ON DEVELOPING A VACCINE AGAINST WHITE SPOT DISEASE CAUSED BY A FISH PARASITE. THE WORLDWIDE DECLINE OF OCEAN FISHERIES STOCKS HAS PROVIDED IMPETUS FOR RAPID GROWTH OF AQUACULTURE, I.E., FISH, CRUSTACEAN, AND SHELLFISH FARMING. CURRENTLY THE AQUACULTURE INDUSTRY IS ONE OF THE MOST IMPORTANT SOURCES OF HUMAN FOOD AND IT HAS THE FASTEST GROWTH-RATE OF ALL ANIMAL-PRODUCING FOOD SECTORS. IN THE U.S., AQUACULTURE IS THE FASTEST GROWING ANIMAL FOOD-PRODUCING SEGMENT OF AGRICULTURE GENERATING ANNUAL REVENUE OF APPROXIMATELY $1.3 BILLION IN THE U.S. THE CONTRIBUTION OF AQUACULTURE TO THE GLOBAL FISH SUPPLY BY WEIGHT HAS INCREASED FROM 3.9 PERCENT IN 1970 TO 50 PERCENT IN 2008, WITH A VALUE OF $98.3 BILLION. IN 2008, IT WAS PREDICTED THAT THE ANNUAL GLOBAL CONSUMPTION OF SEAFOOD BY 2010 WOULD BE 110-120 MILLION METRIC TONS. IN FACT, THE LATEST STATISTICS SHOW THAT FISHERIES PRODUCTION FOR DIRECT HUMAN CONSUMPTION INCREASED TO MORE THAN 136 MILLION TONS IN 2012. THE NEED FOR INCREASED AQUACULTURE OUTPUT OVER THE NEXT 20 YEARS IS ESSENTIAL TO DEAL WITH HUMAN POPULATION GROWTH PROJECTIONS, ANTICIPATED ECONOMIC DEVELOPMENT, AND CONCERN OVER THE FUTURE SUSTAINABILITY OF CAPTURE FISHERIES. TODAY THE GLOBAL AQUACULTURE INDUSTRY IS CHALLENGED TO INCREASE SUSTAINABLE PRODUCTION, REDUCE ENVIRONMENTAL CONTAMINATION AND DIVERSIFY PRODUCTION. ECONOMIC LOSSES DUE TO INFECTIOUS DISEASES IN THE GLOBAL AQUACULTURE INDUSTRY ARE ESTIMATED TO BE $3 BILLION ANNUALLY. OUR OBJECTIVE IS TO PROVIDE A MEANS TO REDUCE THESE LOSSES AND RENDER PRODUCTION OF FISH MORE ECONOMICAL. OUR EFFORTS WILL ENHANCE THE SAFETY OF FISH FOR HUMAN CONSUMPTION AND ALSO REDUCE THE USE OF ANTIBIOTICS AND DRUGS IN THE AQUACULTURE INDUSTRY THAT WILL ALSO REDUCE SELECTION FOR DRUG-RESISTANT PATHOGENS THAT CONTAMINATE THE ENVIRONMENT AND COMPROMISE DEALING WITH BACTERIAL INFECTIOUS DISEASES OF ANIMALS AND HUMANS. TO ACHIEVE THESE OBJECTIVES WE WILL DESIGN AND CONSTRUCT STRAINS OF EDWARDSIELLA PISCICIDA THAT CAUSES A SEPTICEMIC INFECTION IN MANY FISH SPECIES TO RENDER IT HIGHLY IMMUNOGENIC BUT TOTALLY UNABLE TO CAUSE DISEASE. THE PROCEDURES DEVISED ARE NEW AND NOVEL AND USE STRATEGIES THAT HAVE HAMPERED SUCCESS WITH MANY PAST EFFORTS TO DEVELOP SAFE EFFICACIOUS LIVE VACCINES TO CONTROL INFECTIOUS DISEASES. THE VACCINE STRAINS ARE DESIGNED WITH GENETIC CONTROLS TO ENSURE THEIR SAFETY AND EFFICACY. THE VACCINES WILL BE ADMINISTERED TO YOUNG FISH (FINGERLINGS) BY BATH IMMERSION, WHICH IS MOST COST EFFECTIVE SINCE THERE IS NO NEED TO HANDLE INDIVIDUAL FISH OR USE INJECTION DEVISES. THE CONSTRUCTED VACCINES WILL ALSO EXHIBIT THE PROPERTY OF BIOLOGICAL CONTAINMENT SUCH THAT THEY CANNOT PERSIST IN FISH EXCEPT FOR THE TIME NEEDED TO INDUCE THE DESIRED IMMUNE RESPONSES AND WILL BE UNABLE TO SURVIVE ANYWHERE IN NATURE INCLUDING THE WATER ENVIRONMENTS IN WHICH THE IMMUNIZED FISH WILL LIVE.SPECIFIC OBJECTIVES FOR RESEARCH.SPECIFIC AIM 1: CONTINUE ISOLATION AND CHARACTERIZATION OF E. PISCICIDA MUTANT STRAINS TO BE CHARACTERIZED FOR PHENOTYPES INCLUDING ATTENUATION AND IMMUNOGENICITY, AS WELL AS CONSTRUCTION AND VALIDATION OF STRAINS WITH MULTIPLE MUTATIONS CONFERRING REGULATED DELAYED ATTENUATION AND REGULATED DELAYED SYNTHESIS OF PROTECTIVE ANTIGENS ENCODED BY CODON-OPTIMIZED SEQUENCES AND DELIVERED BY TYPE 2 OR 3 SECRETION AND/OR BY REGULATED DELAYED IN VIVO LYSIS.SPECIFIC AIM 2: CONTINUE TO IMPROVE RAEV STRAINS FOR SYNTHESIS AND DELIVERY OF ICH SURFACE ANTIGENS AND CONSTRUCT REGULATED DELAYED LYSIS VECTORS WITH OPERON FUSIONS ENCODING TWO OR THREE ICH SURFACE PROTEINS AND VALIDATE BIOSAFETY AND COMPLETE BIOLOGICAL CONTAINMENT PROPERTIES.SPECIFIC AIM 3: FULLY EVALUATE IMMUNOGENICITY OF OPTIMIZED RAEV-ICH STRAINS AND ABILITY TO INDUCE PROTECTIVE IMMUNITY OF LONG DURATION TO ICH CHALLENGE IN CHANNEL CATFISH.THIS IS A COLLABORATION BETWEEN THE UNIVERSITY OF FLORIDA AND THE UNIVERSITY OF GEORGIA.