OVERVIEW: THE TWO-SPOTTED SPIDER MITE, TETRANYCHUS URTICAE (TSSM) IS AN EXTREME POLYPHAGOUS AGRICULTURAL PEST THAT FEEDS ON OVER 1100 PLANT SPECIES INCLUDING MORE THAN 150 CROPS WORLDWIDE. THIS SPECIES IS A RECORD-BREAKER IN DEVELOPMENT OF PESTICIDE RESISTANCE AMONG ARTHROPOD PESTS. IN COMBINATION WITH ITS INCREASED OUTBREAKS, ADAPTATION TO NOVEL HOST PLANTS AND CROP DAMAGE UNDER CONDITIONS OF CLIMATE CHANGE TSSM BECOMES ONE OF WORLD-WIDE HIGH-RISK PEST THAT THREATENS GLOBAL CROP SECURITY. THE EXISTENCE OF UNPRECEDENTED QUALITY OF GENOMIC RESOURCES AND GENETIC TOOLS AVAILABLE IN THIS SPECIES COUPLED WITH STRONG INTERNATIONAL RESEARCH COMMUNITY (WITH 300 PAPERS PUBLISHED ANNUALLY) MAKES IT AN EXCELLENT PEST MODEL FOR DEVELOPMENT OF INNOVATIVE GENOMIC-BASED TECHNOLOGIES FOR PEST CONTROL. THE OVERALL AIM OF THE PROPOSAL IS TO DISSECT FUNCTION OF DETOXIFICATION ENZYMES IN ABILITY OF TSSM'S RAPID ADAPTATION TO DIFFERENT HOST PLANTS. OUR AIM IS TO UNDERSTAND THE ROLE OF TSSM'S OXYGENASES INVOLVED IN THE INTERPLAY BETWEEN HOST PLANTS DEFENSE (E.G. TOMATO AND ARABIDOPSIS) AND TSSM'S ABILITY TO OVERCOME DEFENSE RESPONSE AND ADOPT TO NEW HOST. IN THIS THIS PROPOSAL WE HAVE SELECTED TWO GROUPS OF ENZYMES: THOSE SPECIFICALLY ASSOCIATED WITH HOST SHIFT, AND ONES WHICH ARE OVEREXPRESSED AFTER ADAPTATION. MOLECULAR CHARACTERIZATION OF THE SELECTED PROTEINS WILL SPECIFICALLY FOCUS ON THE IDENTIFICATION OF THEIR NATURAL SUBSTRATES THAT ARE CURRENTLY UNKNOWN. MOREOVER, THEIR BIOLOGICAL ROLES WILL BE DETERMINED USING A MULTIDISCIPLINARY APPROACH INVOLVING BOTH IN VITRO AND IN VIVO MODELS. THEIR BIOCHEMICAL CHARACTERIZATION IN COMBINATION WITH THEIR TISSUE LOCALIZATION AND RNAI-BASED GENE SILENCING WILL DETERMINE THEIR ROLE IN HERBIVORE-PLANT PEST ADAPTATION. TOGETHER, THIS WILL PROVIDE NEW KNOWLEDGE ON TSSM DETOXIFICATION MECHANISMS AT A BIOCHEMICAL AND ORGANISMAL LEVEL AND GENERATE TOOLS AND KNOWLEDGE FOR DEVELOPMENT NEW PEST CONTROL APPROACHES BASED ON BIOCHEMISTRY ANDGENOMICS.INTELLECTUAL MERIT: PLANTS PRODUCE A VAST NUMBER OF SECONDARY METABOLITES, WHOSE COMPOSITION VARIES QUALITATIVELY AND QUANTITATIVELY WITHIN AND BETWEEN SPECIES. TSSM IS ABLE TO OVERCOME A PLETHORA OF PLANT DEFENSE SYSTEMS. MOREOVER, TSSM HAS THE HIGHEST INCIDENCE OF PESTICIDE RESISTANCE, RESULTING IN EXTREME DIFFICULTY OF CONTROLLING POPULATIONS BY CONVENTIONAL CHEMICAL PEST MANAGEMENT. ANALYSES OF THE TSSM GENOME HAVE REVEALED AN UNPRECEDENTED PROLIFERATION OF MULTI-GENE FAMILIES THAT ARE ASSOCIATED WITH THE XENOBIOTIC METABOLISM. TO ELUCIDATE THE BASES FOR TSSM XENOBIOTIC METABOLISM AND RESISTANCE, THE PROPOSED STUDIES FOCUS ON SUBSTRATE IDENTIFICATION (PLANT SECONDARY METABOLITES) OF TSSM PROTEINS MAINLY FROM THE CYTOCHROME P450 AND INTRADIOL RING-CLEAVAGE DIOXYGENASE FAMILIES. THESE PROTEINS HAVE BEEN IDENTIFIED TO ASSOCIATE WITH MITE'S ABILITY TO SHIFT HOSTS AND/OR DISPLAY RESISTANCE TO ACARICIDES. EFFECTS OF SILENCING GENES CODING FOR THE PROTEINS OF INTEREST AND THE RESULTING EFFECTS ON TSSM FITNESS AND SURVIVAL WILL BE EVALUATED WITH SEVERAL PLANT HOSTS USING RNAI-BASED GENE SILENCING. MOREOVER, THE SELECTED PROTEINS WILL BE LOCALIZED TO SPECIFIC TSSM TISSUES AND CELLS WILL LEAD TO UNDERSTANDING THE TSSM DETOXIFICATION MACHINERY AT THE CELLULAR LEVEL. IT IS EXPECTED THAT THE STUDIES OF TSSM XENOBIOTIC METABOLISM WILL NOT ONLY IMPROVE OUR UNDERSTANDING OF PLANT-HERBIVORE INTERACTIONS AND ADAPTATION AT THE MOLECULAR LEVEL, BUT WILL ALSO PROVIDE FOUNDATIONS FOR DEVELOPMENT OF NEW PEST CONTROL TOOLS AND TARGETS FOR TSSM CONTROL, INCLUDING THE DESIGN OF NEW PESTICIDES AND DEVELOPMENT OF IP FROM THIS PROJECT.BROADER IMPACTS: SOCIETAL BENEFITS OF THIS PROJECT RANGE FROM EDUCATION OF NEW GENERATIONS OF SCIENTISTS, BENEFITS TO HUMAN HEALTH AND FOOD PRODUCTION, REDUCTION OF ENVIRONMENTAL POLLUTION, TO INCREASED SUSTAINABILITY OF AGRICULTURE AND FOOD SECURITY. THE PROPOSED STUDIES WILL PAVE THE WAY FOR DEVELOPMENT OF NEW GENERATION OF ACARICIDES THAT WILLNOT HAVE DETRIMENTAL EFFECTS ON HUMAN HEALTH OR ENVIRONMENT. MOREOVER, AS THIS PROPOSAL FOCUSES ON DEVELOPMENT OF NEW CROP PROTECTION STRATEGIES AGAINST MITE PESTS, IT WILL HAVE A DIRECT IMPACT ON SUSTAINABILITY OF AGRICULTURE AND FOOD SECURITY. SIGNIFICANT EMPHASIS IS PLACED ON TRAINING OF NEW GENERATION OF SCIENTISTS. BOTH UNDERGRADUATE AND GRADUATE STUDENTS WILL BE INVOLVED IN THIS MULTIDISCIPLINARY PROJECT.