Submergence of Western equine encephalitis virus: Evidence of positive selection argues against genetic drift and fitness reductions

Understanding the circumstances under which arboviruses emerge is critical for the development of targeted control and prevention strategies. This is highlighted by the emergence of chikungunya and Zika viruses in the New World. However, to comprehensively understand the ways in which viruses emerge and persist, factors influencing reductions in virus activity must also be understood. Western equine encephalitis virus (WEEV), which declined during the late 20(th) century in apparent enzootic circulation as well as equine and human disease incidence, provides a unique case study on how reductions in virus activity can be understood by studying evolutionary trends and mechanisms. Previously, we showed using phylogenetics that during this period of decline, six amino acid residues appeared to be positively selected. To assess more directly the effect of these mutations, we utilized reverse genetics and competition fitness assays in the enzootic host and vector (house sparrows and Culex tarsalis mosquitoes). We observed that the mutations contemporary with reductions in WEEV circulation and disease that were non-conserved with respect to amino acid properties had a positive effect on enzootic fitness. We also assessed the effects of these mutations on virulence in the Syrian-Golden hamster model in relation to a general trend of increased virulence in older isolates. However, no change effect on virulence was observed based on these mutations. Thus, while WEEV apparently underwent positive selection for infection of enzootic hosts, residues associated with mammalian virulence were likely eliminated from the population by genetic drift or negative selection. These findings suggest that ecologic factors rather than fitness for natural transmission likely caused decreased levels of enzootic WEEV circulation during the late 20(th) century. Author summary Equally important to understanding how arboviruses emerge is understanding the conditions in which they experience reductions in activity. Western equine encephalitis virus (WEEV) provides a unique case study due to its reduction in equine and human incidence and wildlife transmission activity during the second half of the twentieth century. Despite those reductions, we identified six amino acid substitutions that appeared to increase fitness in avian hosts and/or mosquito vectors. We also found no effect of these mutations on mammalian virulence. Our results suggest that ecological factors likely explain the reduction in WEEV activity, even in the face of adaptive evolution.