DNA Barcoding And Molecular Phylogeny Of Cephalopods From Chinese Waters

The extant cephalopods is the third largest molluscan class, which is economicallyimportant and highly valued marine animal in the world. Cephalopods is a highlypromising animals for mariculture because of such characters as extremely rapidgrowth, short life span and high nutritional qualities. So species identification andknowledge about distribution of this class help provide better informed conservationmeasures. Although morphology has been successfully employed to identify speciesand phylogenetic analysis in most cephalopods, some cephalopods are difficult todistinguish with traditional morphological identification owing to phenotypicvariation and the existence of closely related taxa. Therefore, a heuristic andhigh-throughput proxy, that bypasses problems that arise when there is littlemorphology left to interpret should be developed.DNA barcoding, as a new technology to aid in species identification, proposes asolution to these limitations. This molecular technology became an intriguing hotspotin the taxonomy research field since it was proposed. Over the last decade, DNAbarcoding has become a potentially useful tool to aid in identifying known speciesand discovering cryptic species and new species. Cytochrome c oxidase I (COI) DNAbarcoding has gained considerable momentum in many animal groups. The primaryaim of the current study is to test the effectiveness of COI DNA barcoding inidentifying cephalopod species from Chinese waters and analysis the phylogeny ofcephalopods based on COI and large ribosomal subunit (16S rRNA) genes.The main work and conclusion of the present study were as the following:1. The COI gene from126specimens of33species of cephalopods were sequenced.The pairwise intra-and interspecific distances were assessed. Intra-and interspecificdistances ranged from0to6.8%(average1.3%) and from6.2%to22.1%(average16.6%). High levels of genetic differentiation within Loliolus beka led to an overlapbetween intra-and interspecific distances. The barcoding gap was absent. Re-analysisafter exclusion of the individuals of L. beka, sequence distance within species did notexceed sequence distance among species and a clear gap was present. Relationshipsamong species were estimated by NJ and Bayesian phylogenetic analyses. Except L. beka, all species fell into reciprocally monophyletic with high bootstrap values usingNJ and Bayesian clustering metrics. The accordant results further demonstrating thatthe32species studied here could be identified to species-level by DNA barcodingtechniques based on COI. The one notable exception, L. beka, possessed twomitochondrial lineages in the construction of phylogenetic trees with high levels ofintraspecific distances, suggesting the occurrence of cryptic species. In this study,seven unidentified octopuses were divided into three undescribed species, beingconsistent with the morphology-based identification. This study confirms the efficacyof COI DNA barcoding for identifying species as well as discovering cryptic diversityof Chinese cephalopods.2. The16S rRNA genes from132specimens of34species of cephalopods weresequenced. Sequences of16other species in this class gathered from GenBank wereincluded. Based on COI and16S rRNA, we discussed the phylogenetic relationship ofCephalopoda. Two methods, NJ and BI, were adopted to construct the phylogenetictrees. All these analyses consistently showed that Cephalopoda, Nautiloidea,Coleoidea, Sepiida, Sepiolida, Octopoda, Loliginidae and Ommastrephidae aremonophyletic and Nautiloidea was basal. These results are consistent with traditionalmorphological classification. The differences between traditional morphologicalclassification and this study are mainly concentrated on two points.(1) All analyses inthis study showed that the Octopodidae is paraphyletic.(2) No analysis recoveredLoliginidae (Myopsida) sister to Ommastrephidae+Enoploteuthidea (Oegopida),failing to support Teuthoidea. Vampyromorpha has been highly controversial becauseit contains several autapomorphic characters which are not shared with octobrachiansor decabrachians. In this study, four phylogenetic trees formed two results, so thephylogenetic relationships between Vampyromorpha and other orders cannot beresolved. The phylogenetic analysis of intrafamilial level within cephalopods showedthat the phylogenetic relationships of Octopodidae and Uroteuthis were confused, andsupported the view that Sepia pharaoni might be a complex of closely related species.