RBV Pig Project Summary/Abstract Hepatitis E virus (HEV) is an emerging zoonotic pathogen endemic in both developing countries and in developed countries including the United States (U.S.). HEV infects 20 million individuals annually, resulting in 3.3 million symptomatic cases with 44,000 deaths. In countries like the U.S., HEV prevalence is being recognized as a danger to immunocompromised individuals where the virus becomes chronic and can result in acute liver failure among other consequences. Approximately 58-92% of HEV-infected organ transplant recipients developed chronic HEV infection. Currently the recommended treatment for chronic HEV is ribavirin (RBV). There have already been clinical cases where RBV has failed to clear HEV infection, resulting in poor patient outcome. RBV treatment failure has been attributed to single nucleotide variant (SNV) HEV strains that can be circulating as quasispecies in the host that have shown resistance to RBV and/or enhanced replication. Furthermore, long- term replication of HEV in the chronically infected host has led to the identification of viral quasispecies that contained insertions of host ribosomal protein S17 (RPS17) or S19 (RPS19) sequences also leading to enhanced viral replication and increased host range. How naturally occurring HEV variants with enhanced replication, such as those harboring RPS17 or single nucleotide variants, contribute to viral pathogenesis is currently unknown. The immediate goal of this project is to adapt the existing immunosuppressed pig chronic HEV system to model RBV treatment failure and resulting disease severity due to variant HEV strains. In specific aim 1, we hypothesize that the immunosuppressed swine model in conjunction with suboptimal RBV treatment will mimic patient treatment in the clinical setting resulting in novel HEV variants and enhanced disease pathogenesis. To test this hypothesis, we will experimentally infect immunosuppressed pigs with wild type HEV or variant HEV, treating some groups with RBV. We can then sequence resulting HEV variants and compare disease severity between variant and wild type HEV groups. In specific aim 2, we hypothesize that SNV variants exhibiting enhanced growth potential in vitro will correlate to enhanced disease in vivo in part due to differential host immune responses. To test this hypothesis, we will characterize the immune response in pigs infected with wild type HEV, and variant HEV viruses from samples collected during specific aim 1. The long- term goal of this project is to determine viral and host interactions that dictate the outcome of infection by HEV and develop strategies and therapeutics that can mitigate poor outcomes. The results from this R21 will be important for future in-depth mechanistic studies of host factors in HEV infection, especially chronic and cross- species infection. Our specific aims and long-term objectives align well with NIH mission goals seeking to enhance health, lengthen life, and r...