Effect of Powassan virus genetic variation on replication in human cell culture and transmission by ticks

PROJECT SUMMARY Powassan virus (POWV) is a neuroinvasive tick-borne flavivirus that is maintained in nature by Ixodid ticks. Over the past decade, the number of human cases has quadrupled, and wildlife studies suggest that the intensity of enzootic transmission is also increasing. POWV is marked by diverse transmission cycles and disease outcomes. The virus consists of two distinct phylogenetic clades, and evidence of infection has been observed in several tick and vertebrate species. Experimental studies of the relationships between POWV genetic diversity, tick transmission, and disease outcome are currently lacking. The goal of this study is to determine phenotypic diversity of POWV in vitro, identify relevant genotypes of potential medical importance, and determine efficiency of virus dissemination in ticks. To facilitate this study, we have collected 47 environmental isolates spanning a wide geographic range in North America, from a number of tick and vertebrate sources, and spanning over 6 decades. We hypothesize that POWV is a phenotypically diverse virus that can differ in disease potential and ability to transmit. To test this hypothesis, we determine replication and cytopathic effect phenotypes in human and tick cell culture. In collaboration with the Broad Institute, we use POWV whole genome sequences to identify genetic variants that correlate with virus phenotype in cell culture. In our second aim, we use an innovative method of barcoding virus to investigate virus dissemination in 4 tick transmission routes: oral, vertical, transstadial, and co-feeding. This information is critical to understanding how POWV persists in the tick vector. The results of this proposed study contribute to our currently lacking knowledge of this emerging flavivirus. The following aims are proposed: Aim 1: Quantify POWV phenotypic diversity in cell culture and identify genetic variants of interest. Aim 2: Assess the effects of transmission route on Powassan virus population in Ixodes scapularis ticks.