Novel Endosymbionts in Rhizarian Amoebae Imply Universal Infection of Unrelated Free-Living Amoebae by Legionellales

Legionellales-infected water is a frequent cause of local outbreaks of Legionnaires’ disease and Pontiac fever. Decontaminations are difficult because Legionellales reproduce in eukaryotic microorganisms (protists). Most often, Legionellales have been isolated from amoebae; however, the culture-based sampling methods are taxonomically biased. Sequencing studies show that amoebae in the cercozoan class Thecofilosea are dominant in soils and wastewater treatment plants, prompting us to screen their capability to serve as potential hosts of endosymbiotic bacteria. Environmental isolates of Thecofilosea contained a surprising richness of endosymbiotic Legionellales, including Legionella. Considering the widespread dispersal of Legionellales in apparently unrelated amoeboid protist taxa, it appears that the morphotype and not the evolutionary origin of amoebae determines their suitability as hosts for Legionellales. We further provide a protocol for gnotobiotic cultivation of Legionellales and their respective hosts, facilitating future genomic and transcriptomic research of host–symbiont relationships.