Phylogenomics of enigmatic crustacean y-larvae reveals multiple origins of parasitism in barnacles.

The deeper relationships of major thecostracan (barnacles sensu lato) groups remain contentious due to a lack of genome-scale data of enigmatic lineages. Commonly referred to as y-larvae, Facetotecta is one such example. The group remains one of the few invertebrate groups without known adult stages and almost entirely obscured biodiversity. Here, we obtained the first metatranscriptome of Facetotecta (comprising ∼3,600 pooled y-larvae), sequenced with real-time, single-molecule isoform sequencing. By combining these data with a taxon-dense, genome-scale data matrix of Pancrustacea, we take a tree-based approach to ortholog identification and place this enigmatic group among other crustaceans. Taking the same approach on a taxonomically narrower matrix comprising malacostracan outgroups and 14 representatives of Thecostraca, we obtain congruent results with almost ten times the number of orthologs. Combined with new ultrastructural evidence on the host attachment system in y-larvae, our data compellingly demonstrate that Facetotecta constitute an early-diverging sister group to Cirripedia and that parasitism, including the termination of the planktonic life cycle in amoeboid, unarthropodal stages, evolved convergently and independently between this group and the parasitic barnacles, Rhizocephala. Finally, to overcome issues inherent in our limited, single-taxon sampling of Facetotecta in the phylogenomic analyses, we use hundreds of vetted y-larva Sanger sequences across five marker genes to robustly estimate a phylogeny of Thecostraca. This convincingly supports the monophyly of Facetotecta and its sister relationship with Cirripedia. Taken together, these exercises allow us to resolve the position of Facetotecta compellingly and take us one step closer to understanding how parasitism evolved in this fascinating group of crustaceans.