Filling the gaps – New molecular and morphological data of gregarine apicomplexans colonising freshwater invertebrates

Gregarines (Apicomplexa: Gregarinasina) are widespread protist symbionts of invertebrates, occupying roles across the symbiotic spectrum from mutualism to parasitism. Despite their ecological importance, they remain far less studied than other apicomplexans, leaving many aspects of their diversity, host specificity, and evolutionary history unresolved. This is particularly true for freshwater taxa for which only a handful of small subunit (SSU) rDNA sequences from species colonising freshwater hosts are available in public databases. In this study, we screened ten freshwater invertebrates (Arthropoda and Annelida) from streams and rivers in North-Rhine Westphalia, Germany, for gregarine infections. Nine eugregarine species were detected and described by combining light and electron microscopy with SSU rDNA sequencing data. We provide new host and locality records, ultrastructural observations, and molecular data for these gregarine species. The SSU phylogenetic analyses reveal a novel well-supported subclade within Gregarinoidea and support the reassignment of the family Metameridae to the Actinocephaloidea. Our findings expand the available molecular and morphological data for freshwater eugregarines and contribute to a clearer picture of their evolutionary relationships.