AbstractProtists frequently host diverse bacterial symbionts, in particular those affiliated with the order Holosporales (Alphaproteobacteria). All characterised members of this bacterial lineage have been retrieved in obligate association with a wide range of eukaryotes, especially multiple protist lineages (e.g. amoebozoans, ciliates, cercozoans, euglenids, and nucleariids), as well as some metazoans (especially arthropods and related ecdysozoans). While the genus Paramecium and other ciliates have been deeply investigated for the presence of symbionts, known members of the family “Candidatus Paracaedibacteraceae” (Holosporales) are currently underrepresented in such hosts. Herein, we report the description of “Candidatus Intestinibacterium parameciiphilum” within the family “Candidatus Paracaedibacteraceae”, inhabiting the cytoplasm of Paramecium biaurelia. This novel bacterium is almost twice as big as its relative “Candidatus Intestinibacterium nucleariae” from the opisthokont Nuclearia and does not present a surrounding halo. Based on phylogenetic analyses of 16S rRNA gene sequences, we identified six further potential species-level lineages within the genus. Based on the provenance of the respective samples, we investigated the environmental distribution of the representatives of “Candidatus Intestinibacterium” species. Obtained results are consistent with an obligate endosymbiotic lifestyle, with protists, in particular freshwater ones, as hosts. Thus, available data suggest that association with freshwater protists could be the ancestral condition for the members of the “Candidatus Intestinibacterium” genus.