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Species delimitation is particularly challenging in Mediterranean nudibranchs because morphological distinction between closely related species is subtle and several cryptic species have been discovered. In this study, we apply an integrative approach to investigate taxonomic distinction of Flabellina cavolini and F. gaditana and assess their genetic and morphologic distinction. We combined a comprehensive phylogeographic survey from across their distribution range and compared genetic variation at mitochondrial and nuclear markers with the variation in colour pattern of cerata that is used to discriminate these two species. Phylogenetic, phylogeographic and species delimitation analyses consistently identify two evolutionary and taxonomic units that do not match with the morphospecies, demonstrating that Flabellina cavolini and F. gaditana are indeed two cryptic, closely related, species and that colour pattern of cerata is not a valid diagnostic character. This study emphasizes how the comparison of range‐wide patterns of phylogenetic and morphological variation, with the inclusion of samples from the type localities, is essential for species delimitation of Mediterranean nudibranchs. Moreover, this study demonstrates the occurrence of Flabellina gaditana in the Mediterranean Sea, allowing to extend the known distribution range of the species, that was previously restricted to the eastern Atlantic Ocean, as well as increasing our inventory of Mediterranean nudibranch diversity.

The genus Ciona is an interesting ‘taxonomic case’ because its evolutionary history and taxonomy have not yet been resolved completely. In this study, we present new findings, describing specimens of an unidentified Ciona species collected along the north-eastern coasts of Sardinia (Tyrrhenian Sea, Mediterranean Sea). Applying an integrative taxonomic approach, based on the joint examination of morphological and molecular traits, we identify these specimens as a new species, Ciona intermedia sp. nov. Morphological comparisons and peculiarities of the habitat first revealed that these Ciona specimens have intermediate characters compared with other Ciona species. Molecular characterization (based on three mitochondrial regions: two already used for discriminating Ciona cryptic species and a newly developed one) confirmed that our specimens could not be assigned to any previously molecularly-characterized species. Both molecular phylogenetic reconstructions and morphological data clearly indicate C. intermedia as sister clade of Ciona edwardsi. Our findings add further complexity to the taxonomy of Ciona, underlying the importance of an integrative taxonomic approach for the study of the evolutionary history of this enigmatic genus.

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