Engineering a culturable Serratia symbiotica strain for aphid paratransgenesis

Aphids are global agricultural pests and important models for bacterial symbiosis. To date, none of the native symbionts of aphids have been genetically manipulated, which limits our understanding of how they interact with their hosts. Serratia symbiotica CWBI-2.3T is a culturable, gut-associated bacterium isolated from the black bean aphid. Closely related Serratia symbiotica strains are facultative aphid endosymbionts that are vertically transmitted from mother to offspring during embryogenesis. We demonstrate that CWBI-2.3T can be genetically engineered using a variety of techniques, plasmids, and gene expression parts. Then, we use fluorescent protein expression to track the dynamics with which CWBI-2.3T colonizes the guts of multiple aphid species, and we measure how this bacterium affects aphid fitness. Finally, we show that we can induce heterologous gene expression from engineered CWBI-2.3T in living aphids. These results inform the development of CWBI-2.3T for aphid paratransgenesis, which could be used to study aphid biology and enable future agricultural technologies. IMPORTANCEInsects have remarkably diverse and integral roles in global ecosystems. Many harbor symbiotic bacteria, but very few of these bacteria have been genetically engineered. Aphids are major agricultural pests and an important model system for the study of symbiosis. This work describes methods for engineering a culturable aphid symbiont, Serratia symbiotica CWBI-2.3T. These approaches and genetic tools could be used in the future to implement new paradigms for the biological study and control of aphids.