An attempted transinfection of Wolbachia in the Western honey bee (Apis mellifera)

Wolbachia, an intracellular endosymbiont found in up to 60% of arthropods, has been celebrated for its highly varied host-phenotype interactions. These effects are diverse, ranging from reproductive manipulations to obligate mutualisms and facultative symbiosis. These facultative effects include increased resistance to, and reduction in the ability to vector, a number of RNA viruses in insects. Artificial transinfection to mediate human vector-borne diseases such as Dengue fever and Zika virus in Aedes mosquitoes has had considerable success globally. However, using Wolbachia to mediate zoonotic disease directly in threatened species has not been examined. The Western honey bee (Apis mellifera) has shown significant global population declines across the US and Europe, suffering from a diverse range of pathogens, including viral RNA and parasite vector networks. Wolbachia infection in honey bees has only been detected once and its effects have not been investigated. Here, I present the first attempted transinfection of Wolbachia in the Western honey bee using established transinfection protocols.  The natural, but rarely found, Wolbachia infection reported in A. mellifera was examined against a robust phylogeny of all existing Wolbachia supergroups, a feat that has not been updated in the literature since 2015. I discovered Wolbachia infection in Ancistrocerus gazella, the European tube wasp, where it has never been observed. I isolated the natural Wolbachia strain hosted by Drosophila melanogaster (wMel ) and more than 1200 individuals from a range of honey bee life stages (from eggs to adults) were used as potential Wolbachia recipients using sound microinjection protocols. Additionally, I present a novel transinfection avenue utilizing artificial insemination and honey bee breeding using Wolbachia-inoculated drone semen.  When no individuals were successfully infected with Wolbachia in F0 or F1, I investigated the expression of several antimicrobial peptides to characterize the immune response in young larvae to Wolbachia microinjection. There was a significant upregulation of apidaecin when injected with live Wolbachia, but not heat-treated bacteria, which has never been reported in host immune response to Wolbachia previously. The findings presented in this study highlight the importance of Wolbachia strain selection, immune response to Wolbachia, and the potential requirement for cell line culture in future transinfection attempts into A. mellifera. These findings will help inform future transinfection attempts, which are encouraged.