Hepatincolaceae (Alphaproteobacteria) are Distinct From Holosporales and Independently Evolved to Associate With Ecdysozoa

ABSTRACT The Hepatincolaceae (Alphaproteobacteria) are a group of bacteria that inhabit the gut of arthropods and other ecdysozoans, associating extracellularly with microvilli. Previous phylogenetic studies, primarily single‐gene analyses, suggested their relationship to the Holosporales, which includes intracellular bacteria in protist hosts. However, the genomics of Hepatincolaceae is still in its early stages. In this study, the number of available Hepatincolaceae genomes was increased to examine their evolutionary and functional characteristics. It was found that the previous phylogenetic grouping with Holosporales was incorrect due to sequence compositional biases and that Hepatincolaceae form an independent branch within the Hepatincolaceae. This led to a reinterpretation of their features, proposing a new evolutionary scenario that involves an independent adaptation to host association compared to the Holosporales, with distinct specificities. The Hepatincolaceae exhibit greater nutritional flexibility, utilising various molecules available in the host gut and thriving in anaerobic conditions. However, they have a less complex mechanism for modulating host interactions, which are likely less direct than those of intracellular bacteria. In addition, representatives of Hepatincolaceae show several lineage‐specific traits related to differences in host species and life conditions. The main differences between the ‘Ca. Hepatincolaceae’ and the Holosporales are their habitats and modes of existence. The former are non‐motile bacteria that exist extracellularly in the host gut lumen, associating with microvilli, while the latter are intracellular bacteria, primarily hosted by protists such as amoebae and ciliates, and often possess flagellar genes.