title: Sequence-Specific Fidelity Alterations Associated with West Nile Virus Attenuation in Mosquitoes.
abstract: High rates of error-prone replication result in the rapid accumulation of genetic diversity of RNA viruses. Recent studies suggest that mutation rates are selected for optimal viral fitness and that modest variations in replicase fidelity may be associated with viral attenuation. Arthropod-borne viruses (arboviruses) are unique in their requirement for host cycling and may necessitate substantial genetic and phenotypic plasticity. In order to more thoroughly investigate the correlates, mechanisms and consequences of arbovirus fidelity, we selected fidelity variants of West Nile virus (WNV; Flaviviridae, Flavivirus) utilizing selection in the presence of a mutagen. We identified two mutations in the WNV RNA-dependent RNA polymerase associated with increased fidelity, V793I and G806R, and a single mutation in the WNV methyltransferase, T248I, associated with decreased fidelity. Both deep-sequencing and in vitro biochemical assays confirmed strain-specific differences in both fidelity and mutational bias. WNV fidelity variants demonstrated host-specific alterations to replicative fitness in vitro, with modest attenuation in mosquito but not vertebrate cell culture. Experimental infections of colonized and field populations of Cx. quinquefaciatus demonstrated that WNV fidelity alterations are associated with a significantly impaired capacity to establish viable infections in mosquitoes. Taken together, these studies (i) demonstrate the importance of allosteric interactions in regulating mutation rates, (ii) establish that mutational spectra can be both sequence and strain-dependent, and (iii) display the profound phenotypic consequences associated with altered replication complex function of flaviviruses.
authors.affiliation: The Arbovirus Laboratory, Wadsworth Center, New York State Department of Health, Slingerlands, New York, United States of America.
authors.forename: Greta A
authors.initials: GA
authors.lastname: Van Slyke
authors.affiliation: Department of Biochemistry and Molecular Biology, The Pennsylvania State University, University Park, Pennsylvania, United States of America.
authors.forename: Jamie J
authors.initials: JJ
authors.lastname: Arnold
authors.affiliation: Department of Biochemistry and Molecular Biology, The Pennsylvania State University, University Park, Pennsylvania, United States of America.
authors.forename: Alex J
authors.initials: AJ
authors.lastname: Lugo
authors.affiliation: The Arbovirus Laboratory, Wadsworth Center, New York State Department of Health, Slingerlands, New York, United States of America.
authors.forename: Sara B
authors.initials: SB
authors.lastname: Griesemer
authors.affiliation: Department of Biochemistry and Molecular Biology, The Pennsylvania State University, University Park, Pennsylvania, United States of America.
authors.forename: Ibrahim M
authors.initials: IM
authors.lastname: Moustafa
authors.affiliation: The Arbovirus Laboratory, Wadsworth Center, New York State Department of Health, Slingerlands, New York, United States of America; Department of Biomedical Sciences, State University of New York at Albany School of Public Health, Albany, New York, United States of America.
authors.forename: Laura D
authors.initials: LD
authors.lastname: Kramer
authors.affiliation: Department of Biochemistry and Molecular Biology, The Pennsylvania State University, University Park, Pennsylvania, United States of America.
authors.forename: Craig E
authors.initials: CE
authors.lastname: Cameron
authors.affiliation: The Arbovirus Laboratory, Wadsworth Center, New York State Department of Health, Slingerlands, New York, United States of America; Department of Biomedical Sciences, State University of New York at Albany School of Public Health, Albany, New York, United States of America.
authors.forename: Alexander T
authors.initials: AT
authors.lastname: Ciota
mesh_terms: D000818:Animals; D001483:Base Sequence; D009033:Culicidae / Q000821:virology*; D014644:Genetic Variation / Q000235:genetics*; D054884:Host-Pathogen Interactions / Q000235:genetics; D009154:Mutation / Q000235:genetics; D012324:RNA-Dependent RNA Polymerase / Q000235:genetics; D014779:Virus Replication / Q000235:genetics*; D014902:West Nile virus / Q000235:genetics*
chemical_list: D012324:RNA-Dependent RNA Polymerase