A single cryptomonad cell harbors a complex community of organelles, bacteria, a phage, and selfish elements

Symbiosis between prokaryotes and microbial eukaryotes (protists) has broadly impacted both evolution and ecology. Endosymbiosis led to mitochondria and plastids, the latter spreading across the tree of eukaryotes by subsequent rounds of endosymbiosis. Present-day endosymbionts in protists remain both common and diverse, although what function they serve is often unknown. Here, we describe a highly complex community of endosymbionts and a bacteriophage (phage) within a single cryptomonad cell. Cryptomonads are a model for organelle evolution because their secondary plastid retains a relict endosymbiont nucleus, but only one previously unidentified Cryptomonas strain (SAG 25.80) is known to harbor bacterial endosymbionts. We carried out electron microscopy and FISH imaging as well as genomic sequencing on Cryptomonas SAG 25.80, which revealed a stable, complex community even after over 50 years in continuous cultivation. We identified the host strain as Cryptomonas gyropyrenoidosa, and sequenced genomes from its mitochondria, plastid, and nucleomorph (and partially its nucleus), as well as two symbionts, Megaira polyxenophila and Grellia numerosa, and one phage (MAnkyphage) infecting M. polyxenophila. Comparing closely related endosymbionts from other hosts revealed similar metabolic and genomic features, with the exception of abundant transposons and genome plasticity in M. polyxenophila from Cryptomonas. We found an abundance of eukaryote-interacting genes as well as many toxin-antitoxin systems, including in the MAnkyphage genome that also encodes several eukaryotic-like proteins. Overall, the Cryptomonas cell is an endosymbiotic conglomeration with seven distinct evolving genomes that all show evidence of inter-lineage conflict but nevertheless remain stable, even after more than 4,000 generations in culture.