Selective constraint and adaptive potential of West Nile virus within and among naturally infected avian hosts and mosquito vectors

Arthropod-borne viruses are among the most genetically constrained RNA viruses, yet they have a remarkable propensity to adapt and emerge. We studied wild birds and mosquitoes naturally infected with West Nile virus (WNV) in a 'hot spot' of virus transmission in Chicago, IL, USA. We generated full coding WNV genome sequences from spatiotemporally matched bird and mosquito samples using high-throughput sequencing, allowing a molecular evolutionary assessment with deep coverage. Mean among samples was 0.66 (±0.02 SE) and was bimodal, with mean nucleotide diversity being higher between samples (interhost  = 0.001;  = 0.024) than within them (intrahost  < 0.0001;  < 0.001). Eight genomic sites with  > 1.01 (in the , , , , and 5'-noncoding genomic regions) showed bird versus mosquito variant frequency differences of >30 per cent and/or polymorphisms fixed in ≥5 host or vector individuals, suggesting host tropism for these variants. However, phylogenetic analyses demonstrated a lack of grouping by bird or mosquito, most inter-sample differences were synonymous (mean interhost / = 0.04), and there was no significant difference between hosts and vectors in either their nucleotide diversities or levels of purifying selection (mean intrahost / = 0.28 in birds and / = 0.21 in mosquitoes). This finding contrasts with the 'trade-off' and 'selective sieve' hypotheses that have been proposed and tested in the laboratory, which predict strong host versus vector effects on WNV genetic variation, with heightened selective constraint in birds alternating with heightened viral diversity in mosquitoes. Overall, our data show WNV to be highly selectively constrained within and between both hosts and vectors but still able to vary at a limited number of sites across the genome. Such site-specific plasticity in the face of overall selective constraint may offer a mechanism whereby highly constrained viruses such as WNV and its relatives can still adapt and emerge.