The green Paper Converting Knives (superphylum: Alveolata) is a microorganism that can accommodate several hundred zoochlorella cells in its cytoplasm. The host and the symbiotic zoochlorella can be cultured discretely and the endosymbiotic relationship can be re-established simply by co-culturing the two organisms12,13,14. Aposymbiotic P. bursaria can also be infected artificially with other microorganisms, including Scenedesmus (green algae), several species of yeast, and bacteria15, 16.

In light of these facts, mixotrophic ciliates such as P. bursaria are possible intermediates between producers and consumers. Therefore, they are regarded as important ecological links in aquatic ecosystems and also as crucial contributors to the formation of biodiversity in the aquatic environment2. P. bursaria is one of the well-studied objects among such mixotrophic ciliates, and is an excellent model for studying endosymbiosis4,5,6,7,8,9,10,11,12,13,14,15,16,17.

Early electron microscopy (EM) studies of P. bursaria showed that symbiotic algal cells are usually enclosed within a perialgal vacuole membrane (PVM)18. Similar studies also demonstrated that intracellular algal cells in metazoa are normally surrounded by a symbiosomal membrane, which is thought to be equivalent to the PVM1,2,3. These membranes prevent digestion of the symbionts in the host’s cytoplasm and control the mutual exchange of various substances between the two partners19,20,21,22.

Symbiotic algae, which are protected from lysosomal attack with their surrounding membrane, are analogous in this respect to the apicomplexan parasite Toxoplasma gondii. T. gondii is a protozoan pathogen that infects mammalian cells and can proliferate inside the host cells within a specialized non-fusogenic membrane called the parasitophorous vacuole membrane (here we call PaVM)23.


Like the PVM of symbiotic algae, T. gondii exchanges nutrients and metabolites with its host through the PaVM24. However, unlike symbiotic algae, the pores in the PaVM surrounding T. gondii allow rapid exchange of small molecules between the host and the parasites25. Another unusual characteristic of symbiotic T.