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A positively selected mutation in the WNV 2K peptide confers resistance to superinfection exclusion in vivo.

ABSTRACT: Molecular epidemiologic studies of North American (NA) West Nile virus (WNV; Flaviviridae, Flavivirus) have documented the displacement of the introduced NY99 genotype with WN02. In addition, these studies have shown that particular substitutions are under positive selection. One occurs in the C-terminus of the NS4A coding sequence and results in a valine to methionine substitution at position nine of the 2K peptide. 2K-V9M confers the ability to overcome superinfection exclusion in vitro. We hypothesized that WNV strains bearing 2K-V9M have higher fitness than wildtype in Culex quinquefasciatus mosquitoes. Although infection rates and viral titers were not significantly different, virus dissemination rates were significantly higher with WNV 2K-V9M. As a super-infecting virus, WNV 2K-V9M was more successful than wildtype, however, in a mixed infection, 2K-V9M was not. These data support observations that 2K-V9M confers a context-specific selective advantage in mosquitoes and provides an in vivo mechanism for its positive selection.

SUBMITTER: Campbell CL

PROVIDER: S-EPMC4486337 | biostudies-literature | 2014 Sep

REPOSITORIES: biostudies-literature

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A positively selected mutation in the WNV 2K peptide confers resistance to superinfection exclusion in vivo.

Campbell Corey L CL Smith Darci R DR Sanchez-Vargas Irma I Zhang Bo B Shi Pei-Yong PY Ebel Gregory D GD

Virology 20140805

Molecular epidemiologic studies of North American (NA) West Nile virus (WNV; Flaviviridae, Flavivirus) have documented the displacement of the introduced NY99 genotype with WN02. In addition, these studies have shown that particular substitutions are under positive selection. One occurs in the C-terminus of the NS4A coding sequence and results in a valine to methionine substitution at position nine of the 2K peptide. 2K-V9M confers the ability to overcome superinfection exclusion in vitro. We hy ...[more]

PMID: 25104615

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Project description:West Nile virus (WNV) is the leading cause of mosquito-borne illness in the United States. There are currently no human vaccines or therapies available for WNV, and vector control is the primary strategy used to control WNV transmission. The WNV vector Culex tarsalis is also a competent host for the insect-specific virus (ISV) Eilat virus (EILV). ISVs such as EILV can interact with and cause superinfection exclusion (SIE) against human pathogenic viruses in their shared mosquito host, altering vector competence for these pathogenic viruses. The ability to cause SIE and their host restriction make ISVs a potentially safe tool to target mosquito-borne pathogenic viruses. In the present study, we tested whether EILV causes SIE against WNV in mosquito C6/36 cells and Culex tarsalis mosquitoes. The titers of both WNV strains-WN02-1956 and NY99-were suppressed by EILV in C6/36 cells as early as 48-72 h post superinfection at both multiplicity of infections (MOIs) tested in our study. The titers of WN02-1956 at both MOIs remained suppressed in C6/36 cells, whereas those of NY99 showed some recovery towards the final timepoint. The mechanism of SIE remains unknown, but EILV was found to interfere with NY99 attachment in C6/36 cells, potentially contributing to the suppression of NY99 titers. However, EILV had no effect on the attachment of WN02-1956 or internalization of either WNV strain under superinfection conditions. In Cx. tarsalis, EILV did not affect the infection rate of either WNV strain at either timepoint. However, in mosquitoes, EILV enhanced NY99 infection titers at 3 days post superinfection, but this effect disappeared at 7 days post superinfection. In contrast, WN02-1956 infection titers were suppressed by EILV at 7 days post-superinfection. The dissemination and transmission of both WNV strains were not affected by superinfection with EILV at either timepoint. Overall, EILV caused SIE against both WNV strains in C6/36 cells; however, in Cx. tarsalis, SIE caused by EILV was strain specific potentially owing to differences in the rate of depletion of shared resources by the individual WNV strains.

Project description:UnlabelledMany viruses have the capacity to prevent a cell from being infected by a second virus, often termed superinfection exclusion. Alphaherpesviruses, including the human pathogen herpes simplex virus 1 (HSV-1) and the animal herpesvirus pseudorabies virus (PRV), encode a membrane-bound glycoprotein, gD, that can interfere with subsequent virion entry. We sought to characterize the timing and mechanism of superinfection exclusion during HSV-1 and PRV infection. To this end, we utilized recombinant viruses expressing fluorescent protein (FP) markers of infection that allowed the visualization of viral infections by microscopy and flow cytometry as well as the differentiation of viral progeny. Our results demonstrated the majority of HSV-1- and PRV-infected cells establish superinfection exclusion by 2 h postinfection. The modification of viral infections by virion inactivation and phosphonoacetic acid, cycloheximide, and actinomycin D treatments indicated new protein synthesis is needed to establish superinfection exclusion. Primary infection with gene deletion PRV recombinants identified that new gD expression is not required to establish superinfection exclusion of a secondary viral inoculum. We also identified the timing of coinfection events during axon-to-cell spread, with most occurring within a 2-h window, suggesting a role for cellular superinfection exclusion during neuroinvasive spread of infection. In summary, we have characterized a gD-independent mechanism of superinfection exclusion established by two members of the alphaherpesvirus family and identified a potential role of exclusion during the pathogenic spread of infection.ImportanceSuperinfection exclusion is a widely observed phenomenon initiated by a primary viral infection to prevent further viruses from infecting the same cell. The capacity for alphaherpesviruses to infect the same cell impacts rates of interviral recombination and disease. Interviral recombination allows genome diversification, facilitating the development of resistance to antiviral therapeutics and evasion of vaccine-mediated immune responses. Our results demonstrate superinfection exclusion occurs early, through a gD-independent process, and is important in the directed spread of infection. Identifying when and where in an infected host viral genomes are more likely to coinfect the same cell and generate viral recombinants will enhance the development of effective antiviral therapies and interventions.

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A positively selected mutation in the WNV 2K peptide confers resistance to superinfection exclusion in vivo.

Publications

A positively selected mutation in the WNV 2K peptide confers resistance to superinfection exclusion in vivo.

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