Acquisition of a Novel Sulfur-Oxidizing Symbiont in the Gutless Marine Worm Inanidrilus exumae

Gutless phallodrilines are marine annelid worms without a mouth or gut, which live in an obligate association with multiple bacterial endosymbionts that supply them with nutrition. In this study, we discovered an unusual symbiont community in the gutless phallodriline Inanidrilus exumae that differs markedly from the microbiomes of all 22 of the other host species examined. Comparative 16S rRNA gene sequence analysis and fluorescence in situ hybridization revealed that I. exumae harbors cooccurring gamma-, alpha-, and deltaproteobacterial symbionts, while all other known host species harbor gamma- and either alpha- or deltaproteobacterial symbionts. Surprisingly, the primary chemoautotrophic sulfur oxidizer “Candidatus Thiosymbion” that occurs in all other gutless phallodriline hosts does not appear to be present in I. exumae. Instead, I. exumae harbors a bacterial endosymbiont that resembles “Ca. Thiosymbion” morphologically and metabolically but originates from a novel lineage within the class Gammaproteobacteria. This endosymbiont, named Gamma 4 symbiont here, had a 16S rRNA gene sequence that differed by at least 7% from those of other free-living and symbiotic bacteria and by 10% from that of “Ca. Thiosymbion.” Sulfur globules in the Gamma 4 symbiont cells, as well as the presence of genes characteristic for autotrophy (cbbL) and sulfur oxidation (aprA), indicate that this symbiont is a chemoautotrophic sulfur oxidizer. Our results suggest that a novel lineage of free-living bacteria was able to establish a stable and specific association with I. exumae and appears to have displaced the “Ca. Thiosymbion” symbionts originally associated with these hosts. IMPORTANCE All 22 gutless marine phallodriline species examined to date live in a highly specific association with endosymbiotic, chemoautotrophic sulfur oxidizers called “Ca. Thiosymbion.” These symbionts evolved from a single common ancestor and represent the ancestral trait for this host group. They are transmitted vertically and assumed to be in transition to becoming obligate endosymbionts. It is therefore surprising that despite this ancient, evolutionary relationship between phallodriline hosts and “Ca. Thiosymbion,” these symbionts are apparently no longer present in Inanidrilus exumae. They appear to have been displaced by a novel lineage of sulfur-oxidizing bacteria only very distantly related to “Ca. Thiosymbion.” Thus, this study highlights the remarkable plasticity of both animals and bacteria in establishing beneficial associations: the phallodriline hosts were able to acquire and maintain symbionts from two very different lineages of bacteria, while sulfur-oxidizing bacteria from two very distantly related lineages were able to independently establish symbiotic relationships with phallodriline hosts.