Differences in proteome perturbations caused by the Wolbachia strain wAu suggest multiple mechanisms of Wolbachia-mediated antiviral activity.

Some strains of the inherited bacterium Wolbachia have been shown to be effective at reducing the transmission of dengue virus (DENV) and other RNA viruses by Aedes aegypti in both laboratory and field settings and are being deployed for DENV control. The degree of virus inhibition varies between Wolbachia strains. Density and tissue tropism can contribute to these differences but there are also indications that this is not the only factor involved: for example, strains wAu and wAlbA are maintained at similar intracellular densities but only wAu produces strong DENV inhibition. We previously reported perturbations in lipid transport dynamics, including sequestration of cholesterol in lipid droplets, with strains wMel/wMelPop in Ae. aegypti. To further investigate the cellular basis underlying these differences, proteomic analysis of midguts was carried out on Ae. aegypti lines carrying strains wAu and wAlbA: with the hypothesis that differences in perturbations may underline Wolbachia-mediated antiviral activity. Surprisingly, wAu-carrying midguts not only showed distinct proteome perturbations when compared to non-Wolbachia carrying and wAlbA-carrying midguts but also wMel-carrying midguts. There are changes in RNA processing pathways and upregulation of a specific set of RNA-binding proteins in the wAu-carrying line, including genes with known antiviral activity. Lipid transport and metabolism proteome changes also differ between strains, and we show that strain wAu does not produce the same cholesterol sequestration phenotype as wMel. Moreover, in contrast to wMel, wAu antiviral activity was not rescued by cyclodextrin treatment. Together these results suggest that wAu could show unique features in its inhibition of arboviruses compared to previously characterized Wolbachia strains.