Middle East Respiratory Syndrome Coronavirus (MERS-CoV) likely originated in bats and passed to humans through dromedary camels; however, the genetic mechanisms underlying cross-species adaptation remain poorly understood. Variation in the host receptor, dipeptidyl peptidase 4 (DPP4), can block the interaction with the MERS-CoV spike protein and form a species barrier to infection. To better understand the species adaptability of MERS-CoV, we identified a suboptimal species-derived variant of DPP4 to study viral adaption. Passaging virus on cells expressing this DPP4 variant led to accumulation of mutations in the viral spike which increased replication. Parallel passages revealed distinct paths of viral adaptation to the same DPP4 variant. Structural analysis and functional assays showed that these mutations enhanced viral entry with suboptimal DPP4 by altering the surface charge of spike. These findings demonstrate that MERS-CoV spike can utilize multiple paths to rapidly adapt to novel species variation in DPP4.

Filoviruses are strongly associated with several species of bats as their natural reservoirs. In this study, we determined the replication potential of all filovirus species: Marburg marburgvirus, Taï Forest ebolavirus, Reston ebolavirus, Sudan ebolavirus, Zaire ebolavirus, and Bundibugyo ebolavirus. Filovirus replication was supported by all cell lines derived from 6 Old and New World bat species: the hammer-headed fruit bat, Buettikofer's epauletted fruit bat, the Egyptian fruit bat, the Jamaican fruit bat, the Mexican free-tailed bat and the big brown bat. In addition, we showed that Marburg virus Angola and Ebola virus Makona-WPGC07 efficiently replicated at 37°C, 37°-41°C, or 41°C, contrary to the hypothesis that temporal elevation in temperature due to flight affects filovirus replication in bats.

The burden of ticks and the pathogens they carry is increasing worldwide. Powassan virus (POWV, Flaviviridae: Flavivirus), the only known North American tick-borne flavivirus, is of particular concern due to rising cases and the severe morbidity of POWV encephalitis. Here, we use a multifaceted approach to evaluate the emergence of lineage II POWV, known as deer tick virus (DTV), in parts of North America where human cases occur. We detected DTV-positive ticks from eight of twenty locations in the northeastern United States with an average infection rate of 1.4%. High-depth whole genome sequencing of eighty-four POWV and DTV samples allowed us to assess geographic and temporal phylodynamics. We observed both stable infection in the northeastern United States and patterns of geographic dispersal within and between regions. Bayesian skyline analysis demonstrated DTV population expansion over the last fifty years. This is concordant with the documented expansion of I. scapularis tick populations and suggests increasing risk of human exposure as the vector spreads. Finally, we isolated sixteen novel viruses in cell culture and demonstrated limited genetic change after passage, a valuable resource for future studies investigating this emerging virus.

Powassan virus (POWV) is a tick-borne flavivirus which has resulted in increasing human cases over the past two decades. Despite high prevalence in ticks and evidence of broad distribution in North America, fewer than 50 human cases are detected annually with evidence of undetected asymptomatic infections. Experimental studies of the relationships between POWV genetic diversity and disease potential are currently lacking. In the present study, sixteen isolates originating from 13 locations in the United States and Canada were used to assessin vitrophenotypic diversity in human neuronal cells. Broad differences in replication and cytopathic ability were observed between isolates, even amongst those in the same sublineage.In vitrophenotype was not associated with geographic or temporal location and could not be associated with specific genotypes. These results support the observation that the North American POWV population may be highly genetically and phenotypically diverse. The degree to whichin vitrophenotype reflects transmission and pathogenesis remains to be determined.

Monitoring pathogens in wildlife populations is imperative for effective management, and for identifying locations for pathogen spillover among wildlife, domestic species and humans. Wildlife pathogen surveillance is challenging, however, as sampling often requires the capture of a significant proportion of the population to understand host pathogen dynamics. To address this challenge, we assessed the ability to use hunter-collected teeth from puma across Colorado to recover genetic data of two feline retroviruses, feline foamy virus (FFV) and feline immunodeficiency virus (FIVpco) and show they can be utilized for this purpose. Comparative phylogenetic analyses of FIVpco and FFV from tooth and blood samples to previous analyses conducted with blood samples collected over a nine-year period from two distinct areas was undertaken highlighting the value of tooth derived samples. We found less FIVpco phylogeographic structuring than observed from sampling only two regions and that FFV data confirmed previous findings of endemic infection, minimal geographic structuring, and supported frequent cross-species transmission from domestic cats to pumas. Viral analysis conducted using intentionally collected blood samples required extensive financial, capture and sampling efforts. This analysis illustrates that viral genomic data can be cost effectively obtained using tooth samples incidentally-collected from hunter harvested pumas, taking advantage of samples collected for morphological age identification. This technique should be considered as an opportunistic method to provide broad geographic sampling to define viral dynamics more accurately in wildlife.

Powassan virus (POWV) is an emergent tick-borne encephalitis virus of Lyme disease endemic sites in North America. Due to range expansion and local intensification of deer tick vector (Ixodes scapularis) populations in the northeastern and upper midwestern U.S., encephalitis cases are increasingly being reported. A better understanding of the transmission cycle of POWV may allow for predicting the eventual public health burden. Recent phylogeographic analyses of POWV have identified geographical structuring, with well-defined northeastern and midwestern clades of the deer tick virus subtype (lineage II); sublineages exist within each clade. It may be that the local sublineages differ in their capacity to be transmitted by the deer tick vector. Accordingly, we determined whether there are strain-dependent differences in transmission. Five recent, low-passage POWV isolates were used to measure aspects of vector competence, using viremic and artificial infection methods. Infection rates in experimental ticks remained consistent between all five isolates tested, resulting in 12-20% infection rate and no clear differences in viral load. We conclude that there is a genotype independent ability of POWV to infect deer ticks, and that differences in transmission efficiency are not likely to serve as the basis for regional differences in apparent public health burden.