Transmission of West Nile Virus by Culex quinquefasciatus Say Infected with Culex Flavivirus Izabal

Abstract

The natural history and potential impact of mosquito-specific flaviviruses on the transmission efficiency of West Nile virus (WNV) is unknown. The objective of this study was to determine whether or not prior infection with Culex flavivirus (CxFV) Izabal altered the vector competence of Cx. quinquefasciatus Say for transmission of a co-circulating strain of West Nile virus (WNV) from Guatemala. CxFV-negative Culex quinquefasciatus and those infected with CxFV Izabal by intrathoracic inoculation were administered WNV-infectious blood meals. Infection, dissemination, and transmission of WNV were measured by plaque titration on Vero cells of individual mosquito bodies, legs, or saliva, respectively, two weeks following WNV exposure. Additional groups of Cx. quinquefasciatus were intrathoracically inoculated with WNV alone or WNV+CxFV Izabal simultaneously, and saliva collected nine days post inoculation. Growth of WNV in Aedes albopictus C6/36 cells or Cx. quinquefasciatus was not inhibited by prior infection with CxFV Izabal. There was no significant difference in the vector competence of Cx. quinquefasciatus for WNV between mosquitoes uninfected or infected with CxFV Izabal across multiple WNV blood meal titers and two colonies of Cx. quinquefasciatus (p>0.05). However, significantly more Cx. quinquefasciatus from Honduras that were co-inoculated simultaneously with both viruses transmitted WNV than those inoculated with WNV alone (p = 0.0014). Co-inoculated mosquitoes that transmitted WNV also contained CxFV in their saliva, whereas mosquitoes inoculated with CxFV alone did not contain virus in their saliva. In the sequential infection experiments, prior infection with CxFV Izabal had no significant impact on WNV replication, infection, dissemination, or transmission by Cx. quinquefasciatus, however WNV transmission was enhanced in the Honduras colony when mosquitoes were inoculated simultaneously with both viruses. Unlike most known flaviviruses (Family, Flaviviridae: Genus, Flavivirus), insect-only flaviviruses are a unique group of flaviviruses that only infect invertebrates. The study of insect-only flaviviruses has increased in recent years due to the discovery and characterization of numerous novel flaviviruses from a diversity of mosquito species around the world. The widespread discovery of these viruses has prompted questions regarding flavivirus evolution and the potential impact of these viruses on the transmission of flaviviruses of public health importance such as WNV. Therefore, we tested the effect of Culex flavivirus Izabal (CxFV Izabal), an insect-only flavivirus isolated from Culex quinquefasciatus mosquitoes in Guatemala, on the growth and transmission of a strain of WNV isolated concurrently from the same mosquito species and location. Prior infection of C6/36 (Aedes albopictus mosquito) cells or Cx. quinquefasciatus with CxFV Izabal did not alter the replication kinetics of WNV, nor did it significantly affect WNV infection, dissemination, or transmission rates in two different colonies of mosquitoes that were fed blood meals containing varying concentrations of WNV. These data demonstrate that CxFV probably does not have a significant effect on WNV transmission efficiency in nature.