Caught in transition: facultative intracellularity and genome evolution of Symbiopectobacterium in Rhodnius species

Abstract Blood-feeding insects typically depend on obligate intracellular bacterial symbionts that provide essential B vitamins absent from vertebrate blood. In contrast, kissing bugs (Triatominae) have long been considered atypical because they rely primarily on extracellular gut-associated bacteria. Recent reports of the genus Symbiopectobacterium in Rhodnius species raise questions about the diversity and evolution of symbiosis in these insects. Here, we investigate the distribution, genome evolution, and tissue localization of Symbiopectobacterium in the genus Rhodnius . Using comparative metagenomics, phylogenomics, fluorescence in situ hybridization, transmission electron microscopy, and hemolymph screening, we characterize a Symbiopectobacterium genome from Rhodnius prolixus and assess its occurrence across publicly available datasets representing multiple Rhodnius species. The R. prolixus strain possesses a large, highly dynamic genome enriched in mobile genetic elements, pseudogenes, and remnants of secretion systems, while retaining biosynthetic pathways for several B vitamins. Comparative analysis revealed variation in genome reduction among Rhodnius -associated strains, suggesting ongoing and potentially independent transitions toward host-restricted symbiosis. Localization analyses detected Symbiopectobacterium intracellularly within posterior midgut epithelial cells and occasionally in the hemolymph, consistent with a facultative intracellular lifestyle. However, no bacteriomes or stable intracellular structures were observed. Together, these findings indicate that Symbiopectobacterium represents an intermediate stage in the transition from environmentally associated bacteria to obligate intracellular mutualists in Triatominae.