Establishment of in vivo and in vitro platforms for investigation of the effects of Wolbachia on the vector competence of Culex pipiens

Wolbachia is an intracellular bacterial symbiont found in nearly half of all arthropod species. In some mosquito vector species, it is known to exhibit repressive effects on viral replication and transmission. Sindbis virus (SINV) is an arthritogenic alphavirus that causes yearly human infections in Northern Europe. Its transmission cycle involves passerine birds and two main mosquito vectors, Culex pipiens and Cx. torrentium. Cx. pipiens is almost always infected with Wolbachia, yet the impact on its vector competence for SINV has never been investigated. The purpose of this study was to establish in vivo (Aim 1) and in vitro (Aims 2 and 3) platforms for the investigation of the effect of Wolbachia on vector competence and viral replication in mosquitoes of the Cx. pipiens complex. For Aim 1, a colony of lab-reared Cx. pipiens molestus was cleared of its natural Wolbachia infection through tetracycline treatment. For Aim 2, a proliferating primary cell culture was initiated from the eggs of Cx. pipiens molestus. For Aim 3, attempts were made to infect Hsu cells with Wolbachia from Cx. pipiens molestus ovaries with varying success. Protocols for all three primary aims have been established. Preliminary experiments suggest that Hsu cells do not support SINV replication. Additionally, the vector competence of Wolbachia-infected Cx. pipiens molestus for SINV has been investigated, however with inconclusive results. Continued maintenance of the Wolbachia-free mosquito line and primary cell culture is projected to allow future studies of Wolbachia-mediated phenotypes and its effect on virus transmission by Cx. pipiens