Developing new tools for improved symbiont-mediated RNAi in the Western honey bee

Insect symbiont engineering is a biotechnological platform with enormous potential for basic science and applications in entomology. In the past few years these tools have been used to implement symbiont-mediated RNA interference (smRNAi) in the honey bee Apis mellifera. For this purpose, the bacterium Snodgrassella alvi, a symbiont associated with the honey bee gut, is engineered to produce and deliver double-stranded RNA (dsRNA). This approach has been leveraged to protect bee health. Our research group is currently working on the optimization and design of new smRNAi tools for functional genomics in honey bees and expanding these technologies to other insect-symbiont systems. We have demonstrated straightforward methods for engineering the chromosome of S. alvi based on electroporation and homologous recombination. By using these methods, we inactivated RNAse III in S. alvi to reduce degradation of dsRNA within the bacterial cell. This new S. alvi strain still maintains its ability to colonize the bee's digestive tract to levels similar to its wild type. Additionally, we have developed a genetically encoded dsRNA biosensor for optimizing smRNAi approaches. The biosensor uses RNA-binding protein domains coupled with bimolecular fluorescence complementation for in vivo quantification of dsRNA production in bacteria. Taken together, these new developments complement the Bee microbiome Tool Kit (BTK) and open opportunities for expanding smRNAi to other arthropods.