Symbiont replacements reset the co-evolutionary relationship between insects and their heritable bacteria.

Auchenorrhynchan insects (Hemiptera) generally depend on two bacterial symbionts for nutrition. These bacteria experience extreme genome reduction and loss of essential cell functions that require direct host support, or the replacement of failing symbionts with more capable ones. However, it remains unclear how hosts adapt to integrate symbionts into their systems, particularly when they are replaced. Here, we comparatively investigated the evolution of host-support mechanisms in the glassy-winged sharpshooter, Homalodisca vitripennis (GWSS), and the aster leafhopper, Macrosteles quadrilineatus (ALF). ALF harbors the ancestral co-symbionts of the Auchenorrhyncha that have tiny genomes, Sulcia (190 kb) and Nasuia (112 kb). In GWSS, Sulcia retains an expanded genome (245 kb), but Nasuia was replaced by the more capable Baumannia (686 kb). To support their symbionts, GWSS and ALF have evolved novel mechanisms via horizontal gene transfer, gene duplication, and co-option of mitochondrial support genes. However, GWSS has fewer support systems targeting essential bacterial processes. In particular, although both hosts use ancestral mechanisms to support Sulcia, GWSS does not encode all of the same support genes required to sustain Sulcia-ALF or Nasuia. Moreover, GWSS support of Baumannia is far more limited and tailored to its expanded capabilities. Our results demonstrate how symbiont replacements shape host genomes and the co-evolutionary process.