Currently, between one-third and two-thirds of marine species may be undescribed, and previous estimates of there being well over one million marine species appear highly unlikely. More species than ever before are being described annually by an increasing number of authors. If the current trend continues, most species will be discovered this century.

We present the most extensive phylogenetic analysis to date, to our knowledge, of higher-level nemertean relationships, based on sequence data from four different genes (the nuclear genes for nuclear large subunit rRNA (28S rRNA) and histone H3 (H3), and the mitochondrial genes for mitochondrial large subunit rRNA (16S rRNA) and cytochrome c oxidase subunit I (COI)). Well-supported clades are, in general, compatible with earlier, more limited, analyses, and current classification is largely in agreement with our results, although there are some notable exceptions. Bdellonemertea (represented by Malacobdella) is found to be a part of Monostilifera, and Polystilifera is the monophyletic sister group to Monostilifera. Cratenemertidae is the sister group to the remaining monostiliferans (including Malacobdella), a group to which we apply the new name Distromatonemertea. Heteronemertea is monophyletic and forms a clade with Hubrechtella; for this clade we introduce the name Pilidiophora. Finally, Pilidiophora and Hoplonemertea (with Malacobdella) form a monophyletic group, and we introduce the name Neonemertea to refer to this group. Palaeonemertea is found to be non-monophyletic and basal among nemerteans.

BackgroundMany marine meiofaunal species are reported to have wide distributions, which creates a paradox considering their hypothesized low dispersal abilities. Correlated with this paradox is an especially high taxonomic deficit for meiofauna, partly related to a lower taxonomic effort and partly to a high number of putative cryptic species. Molecular-based species delineation and barcoding approaches have been advocated for meiofaunal biodiversity assessments to speed up description processes and uncover cryptic lineages. However, these approaches show sensitivity to sampling coverage (taxonomic and geographic) and the success rate has never been explored on mesopsammic Mollusca.ResultsWe collected the meiofaunal sea-slug Pontohedyle (Acochlidia, Heterobranchia) from 28 localities worldwide. With a traditional morphological approach, all specimens fall into two morphospecies. However, with a multi-marker genetic approach, we reveal multiple lineages that are reciprocally monophyletic on single and concatenated gene trees in phylogenetic analyses. These lineages are largely concordant with geographical and oceanographic parameters, leading to our primary species hypothesis (PSH). In parallel, we apply four independent methods of molecular based species delineation: General Mixed Yule Coalescent model (GMYC), statistical parsimony, Bayesian Species Delineation (BPP) and Automatic Barcode Gap Discovery (ABGD). The secondary species hypothesis (SSH) is gained by relying only on uncontradicted results of the different approaches (‘minimum consensus approach’), resulting in the discovery of a radiation of (at least) 12 mainly cryptic species, 9 of them new to science, some sympatric and some allopatric with respect to ocean boundaries. However, the meiofaunal paradox still persists in some Pontohedyle species identified here with wide coastal and trans-archipelago distributions.ConclusionsOur study confirms extensive, morphologically cryptic diversity among meiofauna and accentuates the taxonomic deficit that characterizes meiofauna research. We observe for Pontohedyle slugs a high degree of morphological simplicity and uniformity, which we expect might be a general rule for meiofauna. To tackle cryptic diversity in little explored and hard-to-sample invertebrate taxa, at present, a combined approach seems most promising, such as multi-marker-barcoding (i.e., molecular systematics using mitochondrial and nuclear markers and the criterion of reciprocal monophyly) combined with a minimum consensus approach across independent methods of molecular species delineation to define candidate species.

Open-ocean environments provide few obvious barriers to the dispersal of marine organisms. Major currents and/or environmental gradients potentially impede gene flow. One system hypothesized to form an open-ocean dispersal barrier is the Antarctic Polar Front, an area characterized by marked temperature change, deep water, and the high-flow Antarctic Circumpolar current. Despite these potential isolating factors, several invertebrate species occur in both regions, including the broadcast-spawning nemertean worm Parborlasia corrugatus. To empirically test for the presence of an open-ocean dispersal barrier, we sampled P. corrugatus and other nemerteans from southern South America, Antarctica, and the sub-Antarctic islands. Diversity was assessed by analyzing mitochondrial 16S rRNA and cytochrome c oxidase subunit I sequence data with Bayesian inference and TCS haplotype network analysis. Appropriate neutrality tests were also employed. Although our results indicate a single well-mixed lineage in Antarctica and the sub-Antarctic, no evidence for recent gene flow was detected between this population and South American P. corrugatus. Thus, even though P. corrugatus can disperse over large geographical distances, physical oceanographic barriers (i.e. Antarctic Polar Front and Antarctic Circumpolar Current) between continents have likely restricted dispersal over evolutionary time. Genetic distances and haplotype network analysis between South American and Antarctic/ sub-Antarctic P. corrugatus suggest that these two populations are possibly two cryptic species.

The phylogenetic relationships of selected members of the phylum Nemertea are explored by means of six markers amplified from the genomic DNA of freshly collected specimens (the nuclear 18S rRNA and 28S rRNA genes, histones H3 and H4, and the mitochondrial genes 16S rRNA and cytochrome c oxidase subunit I). These include all previous markers and regions used in earlier phylogenetic analyses of nemerteans, therefore acting as a scaffold to which one could pinpoint any previously published study. Our results, based on analyses of static and dynamic homology concepts under probabilistic and parsimony frameworks, agree in the non‐monophyly of Palaeonemertea and in the monophyly of Heteronemerta and Hoplonemertea. The position of Hubrechtella and the Pilidiophora hypothesis are, however, sensitive to analytical method, as is the monophyly of the non‐hubrechtiid palaeonemerteans. Our results are, however, consistent with the main division of Hoplonemertea into Polystilifera and Monostilifera, the last named being divided into Cratenemertea and Distromatonemertea, as well as into the main division of Heteronemertea into Baseodiscus and the remaining species. The study also continues to highlight the deficient taxonomy at the family and generic level within Nemertea and sheds light on the areas of the tree that require further refinement. © The Willi Hennig Society 2011.

BackgroundIt has been suggested that statistical parsimony network analysis could be used to get an indication of species represented in a set of nucleotide data, and the approach has been used to discuss species boundaries in some taxa.Methodology/Principal FindingsBased on 635 base pairs of the mitochondrial protein-coding gene cytochrome c oxidase I (COI), we analyzed 152 nemertean specimens using statistical parsimony network analysis with the connection probability set to 95%. The analysis revealed 15 distinct networks together with seven singletons. Statistical parsimony yielded three networks supporting the species status of Cephalothrix rufifrons, C. major and C. spiralis as they currently have been delineated by morphological characters and geographical location. Many other networks contained haplotypes from nearby geographical locations. Cladistic structure by maximum likelihood analysis overall supported the network analysis, but indicated a false positive result where subnetworks should have been connected into one network/species. This probably is caused by undersampling of the intraspecific haplotype diversity.Conclusions/SignificanceStatistical parsimony network analysis provides a rapid and useful tool for detecting possible undescribed/cryptic species among cephalotrichid nemerteans based on COI gene. It should be combined with phylogenetic analysis to get indications of false positive results, i.e., subnetworks that would have been connected with more extensive haplotype sampling.

Some of the most interesting and enigmatic cnidarians are classified within the hydrozoan subclass Trachylina. Despite being relatively depauperate in species richness, the clade contains four taxa typically accorded ordinal status: Actinulida, Limnomedusae, Narcomedusae and Trachymedusae. We bring molecular data (mitochondrial 16S and nuclear small and large subunit ribosomal genes) to bear on the question of phylogenetic relationships within Trachylina. Surprisingly, we find that a diminutive polyp form, Microhydrula limopsicola (classified within Limnomedusae) is actually a previously unknown life stage of a species of Stauromedusae. Our data confirm that the interstitial form Halammohydra sp. (Actinulida) is derived from holopelagic direct developing ancestors, likely within the trachymedusan family Rhopalonematidae. Trachymedusae is shown to be diphyletic, suggesting that the polyp stage has been lost independently at least two times within trachyline evolution. Narcomedusae is supported as a monophyletic group likely also arising from trachymedusan ancestors. Finally, some data, albeit limited, suggest that some trachyline species names refer to cryptic species that have yet to be sorted taxonomically.

The first description of the cleavage program of the palaeonemertean Carinoma tremaphoros (a member of a basal clade of the Nemertea) is illustrated by confocal microscopy and microinjection and compared to development of more derived nemerteans and other eutrochozoans (Annelida, Mollusca, Sipunculida and Echiurida). Lineage tracers were injected into individual blastomeres of C. tremaphoros at the 2-, 4-, 8- and 16-cell stage. Subsequent development was followed to the formation of simple (so-called planuliform) planktonic larvae to establish the ultimate fates of the blastomeres. Results of labeling experiments demonstrate that the development of C. tremaphoros bears closer similarity to other Eutrochozoa than development of a previously studied hoplonemertean (Nemertopsis bivittata) and a heteronemertean (Cerebratulus lacteus) in that the first cleavage plane bears an invariant relationship to the plane of bilateral symmetry of the larval body. Additionally, our cell-labeling experiments support the earlier suggestion that the transitory pre-oral belt of cells in the larvae of C. tremaphoros corresponds to the prototroch of other Eutrochozoa. A unique feature of development of C. tremaphoros includes the oblique orientation of the trochal lineages with respect to the anterior-posterior axis of the larva. The significance and application of cleavage characters such as presence of molluscan vs. annelid cross for phylogenetic analyses is reviewed. We argue that molluscan or annelid cross, neither of which are present in nemerteans, are merely two out of much greater variety of patterns created by the differences in the relative size and timing of formation of micromere quartets and none can be considered, by itself, as evidence of close phylogenetic relationship between phyla.