Abstract
Stable endosymbiosis of a bacterium into a host cell promotes cellular and genomic complexity. The mealybug Planococcus citri has two bacterial endosymbionts; remarkably, the gammaproteobacterium Moranella endobia lives in the cytoplasm of the betaproteobacterium Tremblaya princeps. These two bacteria, along with genes horizontally transferred from other bacteria to the P. citri genome, encode complementary gene sets that form a complex metabolic patchwork. Here we test the stability of this three-way symbiosis by sequencing host-symbiont genome pairs for five diverse mealybug species. We find marked fluidity over evolutionary time: while Tremblaya is the result of a single infection in the ancestor of mealybugs, the innermost gammaproteobacterial symbionts result from multiple replacements of inferred different ages from related but distinct bacterial lineages. Our data show that symbiont replacement can happen even in the most intricate symbiotic arrangements, and that pre-existing horizontally transferred genes can remain stable on genomes in the face of extensive symbiont turnover.
