Large-scale comparative genomics unravels great genomic diversity across the Rickettsia and Ca. Megaira genera and identifies Torix group as an evolutionarily distinct clade.

Rickettsia are intracellular bacteria originally described as arthropod borne pathogens that are emerging as a diverse group of often biologically important, non-pathogenic symbionts of invertebrates and microeukaryotes. However, sparse genomic resources for symbiotic strains and for the sister genus (Candidatus Megaira) inhibit our understanding of Rickettsia evolution and biology. Here, we present the first closed genomes of Ca. Megaira from an alga (Mesostigma viride), and Torix Rickettsia from midge (Culicoides impunctatus) and bed bug (Cimex lectularius) hosts. Additionally, we sequenced and constructed draft genomes for Ca. Megaira from another alga (Carteria cerasiformis), Transitional group Rickettsia from tsetse fly (Glossina morsitans submorsitans), and Torix Rickettsia from a spider mite (Bryobia graminum). We further extract 22 draft genomes from arthropod genome sequencing projects, including 1 Adalia, 4 Transitional, 1 Spotted Fever, 7 Torix, 7 Belli and the first Rhyzobius and Meloidae Rickettsia group genomes. We used new and existing Rickettsia genomes to estimate the phylogeny and metabolic potential across groups and reveal transitions in genomic properties. These data reveal Torix as unique amongst currently described Rickettsia, with highly distinct and diverse accessory genomes. We confirm the presence of a third subclade of Torix, previously only known from gene marker sequences. Further, Torix share an intact pentose phosphate pathway with Ca. Megaira, not observed in other Rickettsia. Considering the distinctness and diversity of Torix, we propose that the group be named Candidatus Tisiphia. The wide host range of Ca. Tisiphia symbionts necessitates onward research to understand the biological and physiological bases of Ca. Tisiphia-host interactions. Importance statementMembers of the genus Rickettsia were originally identified as causative agents of mammalian vector-borne disease. In the last 25 years we have recognised that many Rickettsia are arthropod symbionts, and sit alongside a sister taxon, Ca. Megaira, which are symbiotic associates of microeukaryotes. The lack of genomic information for symbiotic strains affects our ability to determine the evolutionary relationships between strains and understand the biological underpinnings of the different symbioses. We clarify these relationships by assembling 26 genomes of Rickettsia from understudied groups, and the first two Ca. Megaira, from various insects and microeukaryotes. Of note, the accessory genome diversity and broad host range of Torix Rickettsia parallels all other Rickettsia combined. This diversity, alongside the breadth of host species, make the Torix clade an important hidden player in invertebrate biology and physiology. We argue this clade should be given its own genus status, for which we propose Ca. Tisiphia.