In this paper we perform an integrative investigation of the Eubranchus odhneri -Eubranchus sanjuanensis species complex. Several specimens morphologically similar to E. sanjuanensis were collected in the Sea of Japan. However, preliminary molecular data indicated that they were closer to the Arctic species E. odhneri. For our analysis we obtained new data on mitochondrial COI and 16S rRNA, and nuclear histone H3 and 18S rRNA molecular markers to reconstruct their phylogeny (Bayesian Inference and Maximum Likelihood approaches), delimit species with ASAP, bPTP and GMYC analyses and estimate divergence times of putative species. We also studied morphological and ecological features including external morphology, colouration, jaw and radular morphology, configuration of the reproductive system, ultrastructure of cnidosacs, and determination of possible diet preferences. Our results show that specimens from the Sea of Japan represent a new species described herein, Eubranchus malakhovi sp.n. It differs both morphologically and genetically from its closely related species, the Arctic E. odhneri and the North-East Pacific E. sanjuanensis. The new species is closely related to E. odhneri with low mtDNA divergence, and our divergence time estimations suggest these species split ~1.5 Mya.
Conformity of trans-Arctic Coryphellidae genera with the most recent revised taxonomy for the group was tested. Both morphological and molecular data were used to compare more than 200 specimens from various localities in the North Pacific, North Atlantic and Arctic Oceans. The molecular study included four markers (COI, 16S, H3 and 28S) and implemented Bayesian and maximum likelihood phylogenetic approaches. Species delimitation was investigated with automatic barcode gap discovery (ABGD), Poisson tree processes (PTP) and generalized mixed Yule coalescent (GMYC) methods. The inter- and intraspecific divergence were studied using the TCS-based haplotype analysis and calculation of p-distances. Morphological analysis included standard anatomical techniques and scanning electron microscopy. Based on integrative analysis, we conclude that Coryphella verrucosa and C. pseudoverrucosa represent a single species (C. verrucosa), which is highly heterogeneous at morphological and molecular levels. For Microchlamylla gracilis and M. amabilis, their synonymization under the name M. gracilis is suggested, but increased sample size is needed for further clarification of their identities. Finally, the amphiboreal species Borealea nobilis and the North Pacific B. sanamyanae are distinct species displaying secondary sympatry in the north-west Pacific. Based on these results, we propose a new classification system for the family Coryphellidae uniting all currently considered genera in the single genus Coryphella, and suggest a new approach for further revision of Flabellinidae s.l. taxa.
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