Single-cell phylogenomics identifies major groups of marine eugregarine endosymbionts (Apicomplexa)

Gregarines are a large group of apicomplexan parasites that infect a wide range of invertebrate hosts, including diverse and speciose groups, such as annelids and arthropods. Marine eugregarines represent the majority of gregarine diversity, but remain poorly understood, especially their deepest phylogenetic relationships. To expand knowledge of marine eugregarine diversity and their evolutionary history, we surveyed marine invertebrates, with a particular focus on annelids, across multiple locations in British Columbia, Canada. From this effort, we obtained high-quality, single-cell transcriptomes from 20 different species of marine eugregarines, including nine previously described species and 11 novel ones, which more than doubles the amount of phylogenomic data for the group. These data, which comprehensively represent the known diversity of marine gregarines in annelid hosts, allowed us to construct an expanded phylogenetic tree based on small subunit ribosomal DNA sequences and a phylogenomic tree inferred from 142 proteins and 44,802 amino acid sequences. Our analyses show that marine eugregarines form six major lineages, five of which include species infecting annelid hosts: The Ancoroidea, Lecudinoidea, Loxomorphoidea n. superfam., Paralecudinoidea n. superfam., and a distinct lineage represented by Belladina schistomeringa n. gen. et sp. These findings contribute to ongoing efforts to build a robust molecular phylogenetic framework for gregarine diversity and refine gregarine classification. However, some of the deepest evolutionary relationships among these superfamilies remain unresolved, highlighting the need for expanded taxon sampling to better capture the true diversity of eugregarine parasites.