GLOBALLY, TREATED OR UNTREATED WASTEWATER AND BIOSOLIDS ARE USED TO IRRIGATE OR FERTILIZE SOILS, MAKING THEM THE RECIPIENTS, RESERVOIRS, AND SOURCES OF ANTIBIOTIC RESISTANCE, AND HENCE, AN EMERGING HUMAN HEALTH CONCERN. THERE IS A CRITICAL NEED TO DETERMINE THE ENVIRONMENTAL HOTSPOTS FOR ANTIBIOTIC RESISTANCE IN WASTEWATER TREATMENT SYSTEMS AND AGRICULTURAL SOILS. THE OVERALL OBJECTIVE FOR THIS RESEARCH IS TO IDENTIFY THE ENVIRONMENTAL HOTSPOTS THAT PROMOTE THE PERSISTENCE AND DISSEMINATION OF ENTERIC ANTIBIOTIC-RESISTANT BACTERIA IN OREGON WASTEWATERS, AGRICULTURAL SOILS, AND FOOD CROPS. THE CENTRAL HYPOTHESIS IS THAT THE EMERGENCE OF ANTIBIOTIC RESISTANCE IS IMPACTED BY SEASONAL, GEOGRAPHICAL, AND OTHER ENVIRONMENTAL STRESSORS. THE RATIONALE UNDERLYING THIS STUDY IS THAT, DETERMINING THE ENVIRONMENTAL HOTSPOTS FOR ANTIBIOTIC RESISTANCE WILL INFORM THE SCIENTIFIC KNOWLEDGE BASE ON THE EMERGENCE OF ANTIBIOTIC RESISTANCE, AND WILL IMPACT POLICIES, INTERVENTIONS, AND RISK ASSESSMENTS FOR WASTEWATER TREATMENT AND AGRICULTURAL PRACTICES. THE PD AND CO-PDS ARE WELL-POSITIONED TO UNDERTAKE THE PROPOSED WORK BECAUSE OF THEIR COLLECTIVE TRACK RECORD OF STUDYING INACTIVATION, GROWTH, AND PERSISTENCE OF ENTERIC BACTERIA IN SOILS, WATERS, WASTEWATER SYSTEMS, AND FOOD MATRICES.THE CENTRAL HYPOTHESIS WILL BE TESTED BY PURSUING TWO SPECIFIC AIMS. THE FIRST IS TO IDENTIFY THE IMPACT OF SEASONAL AND GEOGRAPHICAL VARIATIONS ON THE PREVALENCE OF ANTIBIOTICS AND ANTIBIOTIC RESISTANCE IN WASTEWATER TREATMENT SYSTEMS IN OREGON. ANTIBIOTICS AND ANTIBIOTIC-RESISTANT GENES WILL BE CHARACTERIZED IN WASTEWATER INFLUENT, THE TREATED EFFLUENT, AND BIOSOLIDS ACROSS SEASONS (TO ASSESS SEASONAL VARIATIONS SUCH AS RAIN, TEMPERATURE, AND FLU SEASON), AND GEOGRAPHIES (I.E., VARIATIONS IN THE COASTAL, VALLEY, AND HIGH DESERT REGIONS). THE SECOND IS TO DETERMINE THE ENVIRONMENTAL HOTSPOTS FOR THE EMERGENCE OF ANTIBIOTIC RESISTANCE IN SOILS AND FOOD CROPS AFTER WASTEWATER AND BIOSOLIDS APPLICATION. INA GREENHOUSE SETTING, ANTIBIOTIC RESISTANCE WILL BE EVALUATED UNDER DIFFERENT COMBINATIONS OF IRRIGATION WITH WASTEWATER, SOIL AMENDMENT WITH BIOSOLIDS DURING THE GROWTH SEASONS OF CARROTS.RESEARCH WILL FOCUS ON TRACKING THE FATE OF ANTIBIOTICS, ANTIBIOTIC-RESISTANT BACTERIA AND GENES IN WASTEWATER TREATMENT PLANTS ACROSS OREGON, AND IN SOILS IRRIGATED WITH WASTEWATER OR FERTILIZED WITH BIOSOLIDS IN A GREENHOUSE SETTING. METHODS WILL INCLUDE ADVANCED ANALYTICAL CHEMICAL METHODS, CULTURING OF ANTIBIOTIC-RESISTANT BACTERIA, PHENOTYPIC AND GENOTYPIC CHARACTERIZATION OF ANTIBIOTIC-RESISTANT BACTERIA, AND HIGH-THROUGHPUT DNA SEQUENCING. AS OUTCOMES OF THE PROPOSED INVESTIGATION, WE EXPECT TO HAVE IDENTIFIED HOW WASTEWATER AND BIOSOLIDS APPLICATION IN AGRICULTURE IMPACT THE PERSISTENCE OF ANTIBIOTIC-RESISTANT BACTERIA AND GENES, AND THEIR DISSEMINATION VIA THE TRANSFER OF GENETIC ELEMENTS IN AGRICULTURAL SOIL AND FOOD CROPS.THE PROPOSED PROJECT WILL TRANSFORM KNOWLEDGE ON HOTSPOTS FOR HUMANEXPOSURE TO ENTERIC ANTIBIOTIC-RESISTANT BACTERIA IN WASTEWATER TREATMENT SYSTEMS, AGRICULTURAL SOILS, AND FOOD CROPS. FINDINGS WILL IMPROVE OUR UNDERSTANDING OF THE EMERGENCE OF ANTIBIOTIC RESISTANCE BY IDENTIFYING THE ASSOCIATED ENVIRONMENTAL HOTSPOTS IN WASTEWATER TREATMENT SYSTEMS AND AGRICULTURAL SOILS. UNDERSTANDING THE ENVIRONMENTAL CONTRIBUTORS OF THE DEVELOPMENT OF RESISTANCE IN HUMAN PATHOGENS IS CRITICAL AS INFECTIONS RESISTANT TO ANTIBIOTICS LEAD TO THE RISE OF ILLNESS AND DEATH. EACH YEAR IN THE U.S., THERE ARE TWO MILLION INFECTIONS WITH ENTERIC BACTERIA RESISTANT TO ANTIBIOTICS, AND THERE ARE 23,000 DEATHS AS A DIRECT RESULT OF THESE INFECTIONS. THE PROPOSED PROJECT WILL LAY THE FOUNDATION FOR THE DEVELOPMENT OF EVIDENCE-BASED POLICIES ON WASTEWATER TREATMENT, AGRICULTURAL PRACTICES, AND FOOD SAFETY IN THE U.S. THE SUCCESSFUL COMPLETION OF THIS WORK WILL GENERATE A FRAMEWORK FOR DEVELOPING STRATEGIES TO DECREASE THE EMERGENCE AND SPREAD OF ILLNESS, DISABILITY, AND DEATHATTRIBUTABLE TO ENTERIC ANTIBIOTIC-RESISTANT INFECTIONS.