Taxonomic Study of Shallow-Water Octopuses (Cephalopoda: Octopodidae) in Japan and Adjacent Waters using Mitochondrial Genes with Perspectives on Octopus DNA Barcoding

A recent study revised the generic level taxonomy of the family Octopodidae, and several species groups that were previously included in Octopus are now recognized as independent genera. However, knowledge regarding phylogenetic relationships among taxa in the family Octopodidae is still poor. The phylogenetic relationships of shallow-water benthic octopus species in Japan and adjacent waters were examined in the present study using mitochondrial DNA cytochrome c oxidase subunits I and III (COI and COIII, respectively) sequence data, and the taxonomic statuses of the octopus species were compared to those derived by morphological identification. We obtained COI (657 bp) and COIII (464 bp) sequences from 45 taxa assigned to 34 species of seven genera. Samples used were voucher specimens stored in the National Museum of Nature and Science, Tokyo. Sequences of the genes of both Haliphron atlanticus and Tremoctopus violaceus from GenBank were included as outgroup taxa. Additionally, sequences of the genes of Octopus vulgaris obtained from Mediterranean coast of France, which is type locality of this species, was also included from GenBank in order to discuss about status of Japanese O. vulgaris. Through phylogenetic analyses, we recognized Callistoctopus and a distinct group including four Octopus species and an Enteroctopus species, which was designated the "Enteroctopus group" in the present study, with high bootstrap support. Amphioctopus was not completely distinguished because it sometimes formed a clade together with Hapalochlaena, which shows morphological affinities with Amphioctopus. Abdopus O. laqueus O. cyanea and O. vulgaris O. oliveri were recognized as distinct groups, but the phylogenetic support for these groups was weak. For identifying species using short DNA sequences, sequence divergences were calculated using COI sequences. Sequences within and among species were generally divergent enough to discriminate the species, but several overlaps with intra- and interspecific divergence caused by poor taxonomic resolution were also observed. Although performing species identification using DNA sequences from shallow-water benthic octopus species is premature, the results in this study indicated the possible application of COI sequences in octopus species identification, thereby providing a preliminary dataset for future DNA barcoding of octopus.