POULTRY INDUSTRY HAS BEEN SEVERELY IMPACTED SINCE 2014 BY THE OUTBREAKS OF THE HIGH PATHOGENIC AVIAN INFLUENZA (HPAI) VIRUSES, AND HUNDREDS OF MILLIONS OF CHICKENS AND TURKEYS HAVE BEEN SLAUGHTERED DUE TO THE HPAI OUTBREAKS IN THE UNITED STATES. SINCE 2021 A NOVEL TYPE OF HPAI H5N1 VIRUSES EMERGED IN NORTHAMERICA, ORIGINATED FROM EUROASIAN CONTINENTS, AND SPREAD TO HUNDREDS OF BOVINE CATTLE HERDS IN THE SPRING OF 2024, WHICH FURTHER DEVASTATED COMMERICAL AND BACKYARD POULTRY AS WELL AFTER FURTHER GENETIC MUTATIONS OCCURRED. FEW STUDIES HAVE BEEN CARRIED OUT TO UNDERSTAND HOW THE NEW HPAI H5N1 VIRUESES TRANSMIT AND CAUSE DISEASES IN POULTRY, AND NO VACCINES HAVE BEEN AVAILABLE TO STOP THESE OUTBREAKS. IN THIS PROPOSAL WE WILL COMARARTIVELY STUDY THREE H5N1 VIRUSES, THAT BELONG TO THE SAME 2.3.4.4B CLADE BUT FALL IN DIFFERENT EVOLUTIONARY STAGES, AND EXPLORE THEIR INFECTION, TRANSMISSION, AND THE DISEASE MECHANISMS IN CHICKENS AND TURKEYS. WE ALSO PROPOSE TO DEVELOP A NOVEL MICRORNA-BASED LIVE VACCINE TO CONTROL THE ONGOING OUTBREAKS OF THESE VIRUSES IN POULTRY. THE RESULT OF THIS STUDY WILL HELP US FURTHER UNDERSTAND THE NATURE OF THE NOVEL HPAI VIRUSES AND THE MECHANISMS OF THE INFECTIONS CAUSED BY THESE VIRUSES IN POULTRY. IT IS EXPECTED THAT MORE EFFECTIVE MEASURES AND POLICY RECOMMENDATIONS WILL ARISE FROM THIS STUDY FOR BETTER CONTROLLING THE SPREAD AND OUTBREAKS OF THE 2.3.4.4B CLADE HPAI IN POULTRY.