INFECTIOUS DISEASES OF POULTRY AND SWINE REMAIN A SIGNIFICANT THREAT TO ANIMAL WELFARE AND CONSUMER COST. ESPECIALLY IMPORTANT ARE THOSE INFECTIOUS DISEASES WITH ABUNDANT GENETIC VARIATION, WHICH MAKES THEM DIFFICULT TO DIAGNOSE AND CONTROL, FURTHER AMPLIFYING THE COSTS TO PRODUCERS AND CONSUMERS. FOR EXAMPLE, PORCINE REPRODUCTIVE AND RESPIRATORY SYNDROME VIRUS (PRRSV), WHICH RESULTS IN A WIDE VARIETY OF CLINICAL SIGNS, INCLUDING PNEUMONIA AND SMALL LITTER SIZES IS A VIRUS WITH ABUNDANT GENETIC VARIATION. THIS GENETIC VARIATION RESULTS IN POOR CROSS PROTECTION OF VACCINES AND IMPAIRS THE ACCURACY OF DIAGNOSTIC TESTS. SIMILARLY IN POULTRY, RESPIRATORY VIRUSES SUCH AS INFECTIOUS BRONCHITIS VIRUS (IBV) AND NEWCASTLE DISEASE VIRUS (NDV), BOTH OF WHICH CAUSE COUGHING AND OTHER RESPIRATORY SIGNS, DECREASED EGG PRODUCTION, ILL THRIFT, AND DEATH, ALSO HAVE NUMEROUS VARIANTS. THE DIFFICULTY IN DIAGNOSING AND MANAGING PRRSV, IBV, AND NDV RESULTS IN COSTS ESTIMATED IN THE BILLIONS OF DOLLARS PERYEAR. DEVELOPING DIAGNOSTIC TESTS THAT WORK ON ALL GENETIC VARIANTS OF THESE VIRUSES, AND THAT CAN IDENTIFY WHICH GENETIC VARIANT PRESENT WILL INCREASE PRODUCTION EFFICIENCY AND IMPROVE ANIMAL WELFARE. WE HYPOTHESIZE THAT NEW SEQUENCING TECHNOLOGY WILL PROVIDE A MORE PRECISE DIAGNOSTIC ASSAY, WHILE MAINTAINING TIMELINESS AND AFFORDABILITY. FURTHERMORE, WE HYPOTHESIZE THAT THIS TECHNOLOGY HAS THE POTENTIAL TO DETECT VIRUSES THAT ARE NOT SUSPECTED OF BEING IN THE SAMPLE. IN THIS PROJECT WE PROPOSE TO TEST THE ABILITY OF THIS NEW SEQUENCING TECHNOLOGY. FIRST, WE WILL DETERMINE BASIC QUALITIES (E.G., SENSITIVITY) OF THIS TECHNOLOGY USING PRRSV AND IBV. SECOND, WE WILL TEST THIS TECHNIQUE USING DIAGNOSTIC SAMPLES OBTAINED FROM REAL-WORLD, PRODUCTION FACILITIES, USING SAMPLES KNOWN TO BE POSITIVE FOR PRRSV, IBV, OR NDV. LASTLY, WE WILL TEST THE ABILITY OF THIS NOVEL TECHNIQUE TO IDENTIFY UNSUSPECTED VIRUSES. OUR PROJECT WILL DEVELOP NEW DIAGNOSTIC ASSAYS FOR PRRSV, IBV, AND NDV. FURTHERMORE, THE KNOWLEDGE OBTAINED FROM THIS PROPOSAL CAN BE APPLIED TO OTHER INFECTIOUS DISEASES OF ANIMALS AND HUMANS, INCLUDING VIRUSES SUCH AS INFLUENZA VIRUSES, BACTERIA LIKE SALMONELLA, AND YET-TO-BE DISCOVERED AGENTS.OBJECTIVES: THE OBJECTIVES ARE...DETERMINE THE INITIAL LIMIT OF DETECTION AND REPEATABILITY FOR POSITIVE-SELECTION, NANOPORE SEQUENCING TO DETECT AND GENOTYPE PORCINE REPRODUCTIVE AND RESPIRATORY SYNDROME VIRUS (PRRSV) AND INFECTIOUS BRONCHITIS VIRUS (IBV) FROM ORAL SAMPLESDETERMINE IF SEQUENTIAL, POSITIVE-SELECTION, NANOPORE SEQUENCING EFFICIENTLY DETECTS AND GENOTYPES PRRSV, IBV, AND NEWCASTLE DISEASE VIRUS (NDV) FROM ORAL DIAGNOSTIC SAMPLESDETERMINE IF VIRUS-TARGETED, POSITIVE-SELECTION, NANOPORE SEQUENCING CAN DETECT MIXED INFECTIONS OF TARGETED VIRUSES AND UNTARGETED MICROORGANISMS IN EXPERIMENTAL AND FIELD SAMPLES