**AWARDS ISSUED PRIOR TO JANUARY 20, 2025, WERE FUNDED UNDER PREVIOUS ADMINISTRATIONS AND MAY NOT REFLECT THE PRIORITIES AND POLICIES OF THE CURRENT ADMINISTRATION.** HUMANITY DEPENDS ON WHEAT, WHICH IS THE MOST WIDELY GROWN CROP IN THE WORLD. WHEAT PROVIDES 17% OF THE DAILY PROTEIN AND 20% OF THE FOOD CALORIES FOR 4.5 BILLION PEOPLE. THE TOTAL VALUE OF THE U.S. WHEAT CROP IN 2019 WAS $9 BILLION, AND ABOUT HALF OF THE CROP IS EXPORTED. THE DANGEROUS NEWLY-EMERGED WHEAT BLAST DISEASE, CAUSED BY THE NOTORIOUS SEED-BORNE FUNGUS MAGNAPORTHE ORYZAE, HAS PROVEN HARD TO CONTROL WHEN WEATHER CONDITIONS FAVOR DISEASE, OFTEN RESULTING IN DEVASTATING YIELD AND QUALITY LOSSES. THE MAJOR GOAL OF THIS PROJECT IS TO PREVENT THE EXOTIC WHEAT BLAST FUNGUS FROM ESTABLISHING IN THE U.S. WHEAT CROP. ADDITIONALLY, WHEAT BLAST HAS MAJOR NATIONAL SECURITY IMPLICATIONS FOR THE U.S. BASED ON ITS POTENTIAL FOR SERIOUS SOCIO-ECONOMIC AND GEOPOLITICAL CONSEQUENCES IN AREAS OF THE WORLD WITH DANGEROUS CONFLICT AND STRIFE. ALSO, AS A PATHOGEN THAT MOVES IN SEED OR GRAIN, THE EXOTIC WHEAT BLAST FUNGUS HAS POTENTIAL TO DISRUPT WORLD TRADE THROUGH PROTECTIONIST TRADE BARRIERS THATHAVE NEGATIVE IMPACTS ON EXPORT MARKETS AND THE WORLD ECONOMY. IN ADDITION TO DIRECTLY THREATENING THE U.S., THE WHEAT BLAST SYSTEM ALSO ILLUSTRATES COMPLICATIONS WITH DIAGNOSING AND CONTROLLING MULTIPLE NEW CROP DISEASES THAT EMERGE FROM 'HOST JUMP' EVENTS IN WHICH KNOWN PATHOGENS SUDDENLY EXPAND TO INFECT A NEW HOST. FIRST, EMERGING DISEASES CAUSED BY NEW SUBPOPULATIONS OF ESTABLISHED PATHOGENS PRESENT SPECIAL CHALLENGES FOR DIAGNOSTICS, BECAUSE IT IS NECESSARY TO DISCRIMINATE THE DANGEROUS NEW POPULATION FROM EXISTING POPULATIONS ALREADY IN THE COUNTRY. THAT IS, DIAGNOSTICS MUST DIFFERENTIATE INDIVIDUALS WITHIN A SPECIES AND NOT JUST THAT SPECIES. FOR WHEAT BLAST, THE EXOTIC SUBPOPULATION (KNOWN AS THE TRITICUM OR WHEAT POPULATION) WAS FIRST REPORTED IN BRAZIL IN 1985 BEFORE SPREADING WITHIN SOUTH AMERICA AND THEN CROSSING OCEANS TO ESTABLISH IN SOUTH ASIA IN 2016 AND IN AFRICA IN 2017. WITH PAST NIFA SUPPORT, WE HAVE TRACKED THE EVOLUTION OF THE EXOTIC WHEAT BLAST POPULATION (NOT IN THE U.S.) FROM A FOUNDING POPULATION THAT ALSO GAVE RISE TO THE SERIOUS EMERGING TURFGRASS DISEASE KNOWN AS GRAY LEAF SPOT (GLS) THAT IS WELL ESTABLISHED IN THE U.S. SINCE IT WAS FIRST REPORTED IN PENNSYLVANIA IN 1992. THE NATIVE GLS FUNGUS HAS GROWN AS A THREAT TO THE TURF/GOLF INDUSTRIES SINCE THAT FIRST REPORT ON PERENNIAL RYEGRASS AND ON PASTURE AND FORAGE GRASSES THAT ARE CRITICAL TO THE U.S. LIVESTOCK AND DAIRY INDUSTRIES. THE EXOTIC WHEAT POPULATION IS CLOSELY RELATED TO THE NATIVE GRAY LEAF SPOT POPULATION, COMPLICATING THE DISCOVERY OF DIAGNOSTIC PROBES TO DIFFERENTIATE EXOTIC FUNGUS FROM NATIVE STRAINS, AND GENOMES OF THESE TWO RECENTLY EMERGED PATHOGEN POPULATIONS ARE MIXTURES OF DNA SEGMENTS APPARENTLY INHERITED THROUGH SEXUAL RECOMBINATION WITH MANY OTHER HOST-ADAPTED POPULATIONS ALSO FOUND IN THE U.S. AND WORLD-WIDE. TO ADDRESS THIS FIRST CHALLENGE, IN OBJECTIVE 1, WE WILL DEVELOP A WIDELY APPLICABLE PLUG AND PLAY BIOINFORMATICS PIPELINE THAT WILL ALLOW NON-EXPERTS TO USE PATHOGEN GENOME SEQUENCES AND POPULATION BIOLOGY TO IDENTIFY SETS OF DIAGNOSTIC PROBES FOR DEFINITIVE PATHOGEN IDENTIFICATION FOR THE BLAST FUNGUS AS WELL AS OTHER IMPORTANT PATHOGEN SYSTEMS. IN OBJECTIVE 2, WE WILL USE THIS DIAGNOSTIC DISCOVERY PIPELINE TO IDENTIFY EFFECTIVE DIAGNOSTICS FOR THE EXOTIC WHEAT PATHOGENS AS WELL AS THE NATIVE GRAY LEAF SPOT PATHOGENS. OBJECTIVE 3 ADDRESSES A SECOND CHALLENGE WITH EMERGING PATHOGENS OF MAJOR CROP PLANTS. SPECIFICALLY, CONTROLLING EMERGING DISEASES IS A MOVING TARGET DUE TO CONTINUOUS ADAPTATION OF THE MICROBE TO BECOME A BETTER PATHOGEN ON THE NEW HOST. THAT IS, ONCE ESTABLISHING A FOOTHOLD IN A NEW CROP, THE NEWLY-EMERGED PATHOGEN EVOLVES TO GAIN AGGRESSIVENESS TOWARDS THAT HOST THROUGH SUCCESSIVE DISEASE CYCLES. WITH PREVIOUS NIFA SUPPORT, WE HAVE SHOWN THAT THE EXOTIC TRITICUM POPULATION IS STILL EVOLVING TO OVERCOME PREVIOUSLY IDENTIFIED RESISTANCE AND TO BECOME AN EVEN MORE AGGRESSIVE PATHOGEN OF WHEAT. THEREFORE, WE WILL CHARACTERIZE GENOME AND PATHOGENICITY CHARACTERISTICS OF TRITICUM ISOLATES FROM 2019 PLUS MORE RECENT ISOLATES IN ORDER TO TRACK CONTINUING EVOLUTION IN THE FIELD. PATHOGENICITY ON WHEAT AND OTHER POTENTIAL HOSTS WILL BE ASSAYED IN A BIOSAFELY LEVEL-3 CONTAINMENT LAB AT KANSAS STATE UNIVERSITY AND CONFIRMED IN FIELD STUDIES WITH COLLABORATORS IN BOLIVIA. PATHOGEN STRAINS MORE CLOSELY REPRESENTING THE DIVERSITY OF CURRENT TRITICUM PATHOGENS IN THE FIELD WILL BE INCORPORATED INTO OBJECTIVE 4 WITH A GOAL TO IDENTIFY NEW RESISTANCE TO STACK TOGETHER WITH THE ONLY CURRENTLY EFFECTIVE WHEAT BLAST RESISTANCE IN WHEAT. IN SUMMARY, WE WILL CREATE NEW TOOLS FOR DISEASE DETECTION AND CONTROL OF THE EXOTIC WHEAT BLAST FUNGUS WHILE ALSO UNDERSTANDING ONGOING PATHOGEN EVOLUTION IN THE FIELD. OUR PLUG AND PLAY BIOINFORMATICS PIPELINE AND KNOWLEDGE GAINED ON EMERGING PATHOGEN EVOLUTION WILL PROVIDE VALUE FOR MULTIPLE PLANT PATHOGEN SYSTEMS THAT IMPACT THE U.S. AND THE WORLD.