Project Grant R21AI183178

DETERMINANTS OF ACUTE AND PERSISTENT ZIKA VIRUS SHEDDING IN SEMEN - PROJECT SUMMARY ZIKA VIRUS (ZIKV) CAUSES CONGENITAL ZIKV SYNDROME IN INFANTS EXPOSED IN UTERO FOLLOWING MATERNAL ZIKV INFECTION VIA MOSQUITO BITE OR SEXUAL TRANSMISSION DURING PREGNANCY. IT IS ESTIMATED THAT 3-23% OF ZIKV CASES ARE DUE TO SEXUAL TRANSMISSION. MORE THAN HALF OF ZIKV-INFECTED MEN SHED ZIKV IN SEMEN, AND SOME MEN SHED ZIKV IN SEMEN FOR UP TO 6 MONTHS POST-DISEASE ONSET. IN HUMAN SEMEN, ZIKV INFECTS SEVERAL CELL TYPES INCLUDING EPITHELIAL CELLS AND IMMUNE CELLS. USING A MOUSE MODEL, INFECTION OF EPIDIDYMIS EPITHELIAL CELLS HAS BEEN SHOWN TO BE CRITICAL FOR ACUTE ZIKV SHEDDING IN SEMEN. THE ROLE OF IMMUNE CELLS IN ZIKV SHEDDING IN SEMEN IS UNKNOWN, THOUGH WE DETECTED PERSISTENT ZIKV INFECTION WITHIN THE EPIDIDYMAL LUMEN, RATHER THAN THE EPIDIDYMIS EPITHELIUM, SUGGESTING IMMUNE CELLS MAY BE CRITICAL FOR PERSISTENT ZIKV SHEDDING. THE MOST CLOSELY RELATED VIRUS TO ZIKV, SPONDWENI VIRUS (SPOV), IS POORLY SHED IN SEMEN DESPITE DISSEMINATION TO THE MALE REPRODUCTIVE TRACT. ZIKV CONTAINING THE PRM/E STRUCTURAL GENES FROM SPOV IS SHED IN SEMEN AT SIGNIFICANTLY LOWER LEVELS DURING ACUTE INFECTION. THE LONG-TERM GOAL OF THIS PROJECT IS TO UNDERSTAND THE MECHANISMS OF ZIKV SHEDDING IN SEMEN AND SEXUAL TRANSMISSION. THE OBJECTIVES OF THIS STUDY ARE TO IDENTIFY THE VIRAL FACTORS AND CELL TYPES THAT CONTRIBUTE TO ACUTE AND PERSISTENT VIRAL SHEDDING IN SEMEN. THE HYPOTHESIS IS THAT ZIKV SHEDDING IN SEMEN IS DRIVEN BY VIRAL STRUCTURAL GENES AND PERSISTENT INFECTION OF IMMUNE CELLS IN THE MALE REPRODUCTIVE TRACT. TWO SPECIFIC AIMS WILL ADDRESS THIS HYPOTHESIS: 1) DETERMINE THE ROLE OF VIRAL STRUCTURAL GENES IN ACUTE ZIKV SHEDDING IN SEMEN; AND 2) DETERMINE THE ROLE OF IMMUNE CELLS IN PERSISTENT ZIKV SHEDDING IN SEMEN. IN THE FIRST AIM, WE WILL USE OUR REVERSE GENETICS SYSTEM TO GENERATE A REVERSE GENETICS SYSTEM FOR SPOV AND A CHIMERA CONTAINING ZIKV PRM/E STRUCTURAL GENES WITHIN SPOV. VIRUSES WILL BE USED TO INFECT MALE MICE, AND VIRAL SHEDDING IN EJACULATES WILL BE MEASURED. CELL TYPES DIFFERENTIALLY INFECTED WITHIN THE EPIDIDYMIS EPITHELIUM WILL BE IDENTIFIED. IN THE SECOND AIM, MACROPHAGES WILL BE DEPLETED FROM ZIKV-INFECTED MICE DURING ACUTE AND PERSISTENT INFECTION. VIRAL SHEDDING IN SEMEN AND INFECTION OF THE EPIDIDYMIS WILL BE MEASURED. THE RESEARCH PROPOSED HERE IS INNOVATIVE BECAUSE IT ASSESSES EJACULATES, WHICH ARE BIOLOGICALLY RELEVANT SAMPLES FOR SEXUAL TRANSMISSION, AND A NOVEL ROLE FOR IMMUNE CELLS IN ZIKV SEXUAL TRANSMISSION. UPON SUCCESSFUL COMPLETION OF THE PROPOSED RESEARCH, THE ANTICIPATED CONTRIBUTION OF THIS WORK WILL BE THE IDENTIFICATION OF THE FLAVIVIRUS FACTORS THAT DICTATE VIRAL SHEDDING IN SEMEN AND THE CELL TYPES THAT CONTRIBUTE TO VIRAL SHEDDING IN SEMEN. THIS CONTRIBUTION IS EXPECTED TO BE SIGNIFICANT BECAUSE IT WILL INCREASE OUR UNDERSTANDING OF HOW VIRUSES ARE SEXUALLY TRANSMITTED.