COLLABORATIVE RESEARCH: THE SABOTEUR'S TOOLS IN ACTION: CYTOPLASMIC INCOMPATIBILITY GENES OF THE BACTERIAL ARTHROPOD SYMBIONT CARDINIUM HERTIGII -MOST INSECTS CARRY BACTERIA THAT ARE INHERITED FROM THEIR MOTHERS AND LIVE WITHIN THEIR CELLS. SOME OF SYMBIOTIC BACTERIA MANIPULATE THEIR INSECT HOSTS? REPRODUCTION IN WAYS THAT IMPROVES THE REPRODUCTION OF HOST INSECTS CARRYING THE BACTERIA. SOME OF THESE SYMBIONTS SABOTAGE HOST SPERM SUCH THAT FERTILIZED EGGS LAID BY FEMALES WITHOUT THE BACTERIUM DIE EARLY IN LIFE (?CYTOPLASMIC INCOMPATIBILITY,? OR ?CI?). OF THE FIVE BACTERIA KNOWN TO CAUSE CI, WOLBACHIA IS BEST STUDIED, YET CARDINIUM HERTIGII, THE FOCUS OF THIS STUDY, CAUSES CI WITHOUT THE SAME GENES WOLBACHIA USES. THE OVERALL GOAL OF THIS PROPOSAL IS TO DISCOVER THE MOLECULAR MECHANISM BY WHICH CARDINIUM CAUSES CI. THE PROJECT IS EXPECTED TO HAVE IMPORTANT BENEFITS: CARDINIUM TARGETS ANIMAL CELL DIVISION, A FUNDAMENTAL PROCESS THAT CAN BE BETTER UNDERSTOOD WHEN AGENTS THAT INTERFERE WITH IT ARE STUDIED. IN ADDITION, CI-CAUSING BACTERIA MAY BE USED FOR PEST OR VECTOR MANAGEMENT. THE CI-CAUSING WOLBACHIA REDUCES HOST SUSCEPTIBILITY TO VIRUSES OF INSECTS THAT CARRY IT, AND IS CURRENTLY BEING INTRODUCED TO MOSQUITO POPULATIONS AROUND THE WORLD TO REDUCE VECTOR-BORNE VIRAL DISEASES. THIS PROJECT WILL ALSO ENGAGE ELEMENTARY, HIGH SCHOOL AND UNDERGRADUATE STUDENTS IN SCIENTIFIC EDUCATION AND RESEARCH THROUGH OUTREACH PROGRAMS AND RESEARCH OPPORTUNITIES AT ALL THREE INSTITUTIONS. FURTHERMORE, A CITIZEN SCIENCE PROJECT LED BY THE UNIVERSITY OF ARIZONA AND NORTH CAROLINA STATE UNIVERSITY WILL ENGAGE AMATEUR ENTOMOLOGISTS IN RESEARCH THROUGH THEIR SUPPORT IN COLLECTING AND REARING PARASITIC WASPS OF WHITEFLIES TO CENSUS FOR CARDINIUM. MATERNALLY INHERITED BACTERIAL SYMBIONTS OF ARTHROPODS THAT MANIPULATE HOST REPRODUCTION PROFOUNDLY INFLUENCE HOST BIOLOGY AND EVOLUTION. SYMBIONTS CAUSING CI SABOTAGE HOST SPERM SUCH THAT FERTILIZED EGGS OF UNINFECTED FEMALES DIE IN EARLY EMBRYOGENESIS. THE INDEPENDENT EVOLUTION OF CI IN WOLBACHIA AND CARDINIUM REPRESENTS A REMARKABLE CASE OF CONVERGENT EVOLUTION OF A COMPLEX TRAIT. IN RECENT NSF-SUPPORTED WORK, WE IDENTIFIED TWO PUTATIVE CARDINIUM CI GENES IN A CARDINIUM STRAIN THAT INFECTS A PARASITIC WASP OF WHITEFLIES, ENCARSIA SUZANNAE. IN THREE OBJECTIVES WE AIM TO DETERMINE THE MECHANISM OF CI IN CARDINIUM: 1. DISSECT THE MECHANISM OF CARDINIUM CEPER1 CI CANDIDATE GENES USING AN INTEGRATED STUDY OF LOCALIZATION, DISCOVERY OF INTERACTING HOST AND CARDINIUM PROTEINS, AND MUTATION ANALYSIS. 2. EXPRESS GENES IN DROSOPHILA TO VERIFY THE ROLE OF THESE CANDIDATES IN CI. 3. IDENTIFY CI GENES OF CEINA3 CAUSING CI IN E. PARTENOPEA AND COMPARE TO CEPER1. CARDINIUM PRODUCES A VIRTUALLY IDENTICAL CI PHENOTYPE TO WOLBACHIA WITH DIFFERENT GENES. STUDY OF CARDINIUM CI GENES IN THE NON-MODEL ENCARSIA AS WELL AS HETEROLOGOUS MODELS YEAST AND DROSOPHILA WILL INCREASE OUR UNDERSTANDING OF BOTH CARDINIUM AND WOLBACHIA, BY SHOWING US WHICH PROCESSES ARE COMMON AND WHICH ARE UNIQUE, AND BY SHEDDING LIGHT ON CORE HOST PROCESSES VULNERABLE TO MANIPULATION. GIVEN THE INTEREST OF THE SCIENTIFIC AND PUBLIC HEALTH COMMUNITIES IN SYMBIONT-BASED PEST OR VECTOR-MANAGEMENT STRATEGIES, A DEEPER UNDERSTANDING OF HOST-SYMBIONT INTERACTIONS IN THE COMPARATIVELY UNDERSTUDIED CARDINIUM MAY ALSO LEAD TO NEW CONTEXTS FOR APPLICATIONS, OR FOR APPLICATIONS IN WHICH WOLBACHIA IS NOT EFFECTIVE. THIS AWARD REFLECTS NSF'S STATUTORY MISSION AND HAS BEEN DEEMED WORTHY OF SUPPORT THROUGH EVALUATION USING THE FOUNDATION'S INTELLECTUAL MERIT AND BROADER IMPACTS REVIEW CRITERIA.