Experimental Passage of St. Louis Encephalitis Virus In Vivo in Mosquitoes and Chickens Reveals Evolutionarily Significant Virus Characteristics

Abstract

St. Louis encephalitis virus (SLEV; Flaviviridae, flavivirus) was the major cause of epidemic flaviviral encephalitis in the U.S. prior to the introduction of West Nile virus (WNV) in 1999. However, outbreaks of SLEV have been significantly more limited then WNV in terms of levels of activity and geographic dispersal. One possible explanation for these variable levels of activity is that differences in the potential for each virus to adapt to its host cycle exist. The need for arboviruses to replicate in disparate hosts is thought to result in constraints on both evolution and host-specific adaptation. If cycling is the cause of genetic stability observed in nature and arboviruses lack host specialization, then sequential passage should result in both the accumulation of mutations and specialized viruses better suited for replication in that host. Previous studies suggest that WNV and SLEV differ in capacity for both genetic change and host specialization, and in the costs each accrues from specializing. In an attempt to clarify how selective pressures contribute to epidemiological patterns of WNV and SLEV, we evaluated mutant spectra size, consensus genetic change, and phenotypic changes for SLEV in vivo following 20 sequential passages via inoculation in either Culex pipiens mosquitoes or chickens. Results demonstrate that the capacity for genetic change is large for SLEV and that the size of the mutant spectrum is host-dependent using our passage methodology. Despite this, a general lack of consensus change resulted from passage in either host, a result that contrasts with the idea that constraints on evolution in nature result from host cycling alone. Results also suggest that a high level of adaptation to both hosts already exists, despite host cycling. A strain significantly more infectious in chickens did emerge from one lineage of chicken passage, yet other lineages and all mosquito passage strains did not display measurable host-specific fitness gains. In addition, increased infectivity in chickens did not decrease infectivity in mosquitoes, which further contrasts the concept of fitness trade-offs for arboviruses.

Specific and Nonspecific Host Adaptation during Arboviral Experimental Evolution

“…Using virus production as a surrogate, most of the time there is evidence of adaptation [Chen et al, 2003;Ciota et al, 2007bCiota et al, , 2008Ciota et al, , 2009Coffey et al, 2008;Cooper and Scott, 2001;Llewellyn et al, 2002;Moutailler et al, 2011;Romanova et al, 2007;Vasilakis et al, 2009], but occasionally there are no measurable changes [Ciota et al, 2007c]. It is possible that continuous passages result in no fitness changes if the virus is already highly adapted to the environment.…”

“…Tradeoffs for other arboviral species where these studies exist are unpredictable. For instance, vector-adapted RRV, SFV, VSV and RRV [Novella et al, 1995a;Peleg, 1971;Singh et al, 1971;Taylor and Marshall, 1975] replicate poorly in mammalian environments, but WNV and SLEV do not lose fitness in bypassed avian cells [Ciota et al, 2008[Ciota et al, , 2009. A simple explanation for the lack of expected tradeoffs could be that cell culture does not provide the environmental complexity found in live vectors and hosts, which would include additional components such as antiviral molecules.…”

West Nile Virus Experimental Evolution in vivo and the Trade-off Hypothesis

“…In vivo studies with flaviviruses have also been difficult to reconcile with the trade-off hypothesis. For example, chick-specialized SLEV showed increased infectivity in chicks but was unchanged in mosquitoes, while mosquito-specialized virus was unchanged in both systems [13]. Conversely, serial passage of WNV in mosquitoes resulted in faster replication and higher peak titers in mosquitoes with no significant cost to replication in live chicks [14].…”

Genetic Determinants of Differential Oral Infection Phenotypes of West Nile and St. Louis Encephalitis Viruses in Culex spp. Mosquitoes

“…9,24 Although adaptation studies in Culex mosquitoes and chickens (Gallus gallus) have shown greater genetic plasticity of SLEV compared with WNV, 25,26 the viral genetic determinants that dictate the comparatively greater oral susceptibility for WNV are unknown. Because SLEV is an endemic North American flavivirus that generally elicits low titers in avian hosts, it is likely that this virus has adapted over time to infect North American Culex at low blood meal titers.…”