PT - JOURNAL ARTICLE AU - Harald R. Gruber-Vodicka AU - Nikolaus Leisch AU - Manuel Kleiner AU - Tjorven Hinzke AU - Manuel Liebeke AU - Margaret McFall-Ngai AU - Michael G. Hadfield AU - Nicole Dubilier TI - The <em>Trichoplax</em> microbiome: the simplest animal lives in an intimate symbiosis with two intracellular bacteria AID - 10.1101/568287 DP - 2019 Jan 01 TA - bioRxiv PG - 568287 4099 - http://biorxiv.org/content/early/2019/03/05/568287.short 4100 - http://biorxiv.org/content/early/2019/03/05/568287.full AB - Placozoa is an enigmatic phylum of simple, microscopic, marine metazoans. Although intracellular bacteria have been found in all members of this phylum, almost nothing is known about their identity, location and interactions with their host. We used metagenomic and metatranscriptomic sequencing of single host individuals, plus metaproteomic and imaging analyses, to show that the placozoan Trichoplax H2 lives in symbiosis with two intracellular bacteria. One symbiont forms a new genus in the Midichloriaceae (Rickettsiales) and has a genomic repertoire similar to that of rickettsial parasites, but does not appear to express key genes for energy parasitism. Correlative microscopy and 3-D electron tomography revealed that this symbiont resides in an unusual location, the rough endoplasmic reticulum of its host’s internal fiber cells. The second symbiont belongs to the Margulisbacteria, a phylum without cultured representatives and not known to form intracellular associations. This symbiont lives in the ventral epithelial cells of Trichoplax, likely metabolizes algal lipids digested by its host, and has the capacity to supplement the placozoan’s nutrition. Our study shows that even the simplest animals known have evolved highly specific and intimate associations with symbiotic, intracellular bacteria, and highlights that symbioses with microorganisms are a basal trait of animal life.