Transformation of the Sex-Manipulative Endosymbiont Spiroplasma poulsonii and Persisting Hurdles for Functional Genetic Studies.

Insects are frequently infected by bacterial symbionts that greatly affect their physiology and ecology. Most of these endosymbionts are, however, barely tractable outside their native host, rendering functional genetics studies difficult or impossible. is a facultative bacterial endosymbiont of that manipulates the reproduction of its host by killing its male progeny at the embryonic stage. , although a very fastidious bacterium, is closely related to pathogenic species that are cultivable and genetically modifiable. In this work, we present the transformation of with a plasmid bearing a fluorescence cassette, leveraging techniques adapted from those used to modify the pathogenic species We demonstrate the feasibility of transformation and discuss approaches for mutant selection and fly colonization, which are persisting hurdles that must be overcome to allow functional bacterial genetics studies of this endosymbiont Dozens of bacterial endosymbiont species have been described and estimated to infect about half of all insect species. However, only a few them are tractable , which hampers our understanding of the bacterial determinants of the host-symbiont interaction. Developing a transformation method for is a major step toward genomic engineering of this symbiont, which will foster basic research on endosymbiosis. This could also open the way to practical uses of endosymbiont engineering through paratransgenesis of vector or pest insects.