** AWARDS ISSUED PRIOR TO JANUARY 20, 2025, WERE FUNDED UNDER PREVIOUS ADMINISTRATIONS AND MAY NOT REFLECT THE PRIORITIES AND POLICIES OF THE CURRENT ADMINISTRATION.** INFECTIOUS DISEASES REMAIN A SIGNIFICANT THREAT TO ANIMAL WELFARE AND CONSUMER COST; THUS, DEVELOPING EFFICIENT METHODS TO DIAGNOSE AND CHARACTERIZE THESE INFECTIOUS AGENTS IS IMPORTANT FOR CONTROL. IN POULTRY, IMPROVING DIAGNOSTIC ASSAYS FOR RNA VIRUSES, SUCH AS AVIAN REOVIRUS (ARV), AVIAN CORONAVIRUS (ACOV, CAUSATIVE AGENT OF INFECTIOUS BRONCHITIS; IB), AND AVIAN ASTROVIRUSES (AASTVS), IS A KEY AREA IDENTIFIED BY NATIONAL POULTRY PRODUCER GROUPS. THE DIFFICULTY IN DIAGNOSING AND MANAGING THESE VIRAL DISEASES RESULTS IN COSTS ESTIMATED IN THE BILLIONS OF DOLLARS PER YEAR. DEVELOPING DIAGNOSTIC TESTS THAT WORK ON ALL GENETIC VARIANTS OF THESE VIRUSES, AND THAT CAN IDENTIFY WHICH GENETIC VARIANT IS 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. FIRST, WE WILL DETERMINE THE FEASIBILITY OF USING THE NEWEST METHODS TO CREATE SEQUENCING LIBRARIES FOR ARV, ACOV, AND AASTV. SECOND, WE WILL TEST THIS TECHNIQUE USING DIAGNOSTIC SAMPLES OBTAINED FROM REAL-WORLD, VETERINARY DIAGNOSTIC LABS, USING SAMPLES KNOWN TO BE POSITIVE FOR ARV, ACOV, AND/OR AASTV. LASTLY, WE WILL DETERMINE IF THESE ASSAYS CAN BE MULTIPLEXED TO INCREASE THE EFFICIENCY OF THE ASSAYS EVEN FURTHER. OUR PROJECT WILL DEVELOP NEW DIAGNOSTIC ASSAYS FOR ARV, ACOV, AND AASTV. 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.