Diverse toxin repertoire but limited metabolic capacities inferred from the draft genome assemblies of three <i>Spiroplasma</i> (Citri clade) strains associated with <i>Drosophila</i>

<i>Spiroplasma</i> (class <i>Mollicutes</i>) is a diverse wall-less bacterial genus whose members are strictly dependent on eukaryotic hosts (mostly arthropods and plants), with which they engage in pathogenic to mutualistic interactions. <i>Spiroplasma</i> are generally fastidious to culture <i>in vitro</i>, especially those that are vertically transmitted by their hosts, which include flies in the genus <i>Drosophila</i>. <i>Drosophila</i> has been invaded by at least three independent clades of <i>Spiroplasma</i>: Poulsonii (the best studied, contains reproductive manipulators and defensive mutualists associated with two major clades of <i>Drosophila</i> and has amongst the highest substitution rates within bacteria), Citri (restricted to the <i>repleta</i> group of <i>Drosophila</i>) and Ixodetis. We report the first genome drafts of <i>Drosophila</i>-associated Citri clade <i>Spiroplasma</i>: strain <i>s</i>Moj from <i>Drosophila mojavensis</i>, strain <i>s</i>Ald-Tx from <i>Drosophila aldrichi</i> from Texas (newly discovered; also associated with <i>Drosophila mulleri</i>) and strain <i>s</i>Hy2 from <i>Drosophila hydei</i> (the only <i>Drosophila</i> species known to naturally also harbour a Poulsonii clade strain, thereby providing an arena for horizontal gene transfer). Compared to their Poulsonii clade counterparts, we infer that the three Citri clade strains have the following: (1) equal or worse DNA repair abilities; (b) more limited metabolic capacities, which may underlie their comparatively lower titres and transmission efficiency; and (c) similar content of toxin domains, including at least one ribosome-inactivating protein, which is implicated in the Poulsonii-conferred defence against natural enemies. As a byproduct of our phylogenomic analyses and exhaustive search for certain toxin domains in public databases, we document the toxin repertoire in close relatives of <i>Drosophila</i>-associated <i>Spiroplasma</i>, and in a very divergent newly discovered lineage (i.e. 'clade X'). Phylogenies of toxin-encoding genes or domains imply substantial exchanges between closely and distantly related strains. Surprisingly, despite encoding several toxin genes and achieving relatively high prevalences in certain natural populations (<i>s</i>Ald-Tx in this study; <i>s</i>Moj in prior work), fitness assays of <i>s</i>Moj (this study) and <i>s</i>Ald-Tx (prior work) in the context of wasp parasitism fail to detect a beneficial effect to their hosts. Thus, how Citri clade strains persist in their <i>Drosophila</i> host populations remains elusive.