Cappable-Seq reveals the transcriptional landscape of stress responses in the bacterial endosymbiont Wolbachia

Bacterial endosymbionts are highly prevalent among invertebrate animals, in which they can confer fitness benefits such as pathogen defence and/or act as reproductive manipulators, inducing phenotypes including cytoplasmic incompatibility (CI). For the alpha-proteobacterium Wolbachia , its wide distribution among macroparasites and disease-transmitting arthropods coupled with mutualistic roles, reduction of vector competence, and CI have found recent applications in the control of several vector-borne tropical diseases. However, in common with other bacterial endosymbionts, which often lose regulatory elements during genomic erosion, the degree to which Wolbachia can respond to environmental or pharmacological stressors is poorly understood. Here, we apply Cappable-Seq methodology to achieve unprecedented depth and resolution of transcriptional start sites (TSS) in two Wolbachia strains ( w MelPop-CLA and w AlbB) that have been used to transinfect mosquitoes for arbovirus control. We exposed Wolbachia in mosquito cell lines to temperature stress (both strains) or antibiotics ( w AlbB only) and observed that all classes of TSS (including antisense) exhibited differential regulation, some of which were associated with mobile elements and may control ncRNA expression. Cappable-Seq also resolved the organisation of the bicistronic cifA/cifB operon that is responsible for inducing CI in Wolbachia hosts and shows great promise for revealing regulation of symbiont functions in whole invertebrates.