Topology Testing and Demographic Modeling Illuminate a Novel Speciation Pathway in the Greater Caribbean Sea Following the Formation of the Isthmus of Panama.

Recent genomic analyses have highlighted the prevalence of speciation with gene flow in many taxa and have underscored the importance of accounting for these reticulate evolutionary processes when constructing species trees and generating parameter estimates. This is especially important for deepening our understanding of speciation in the sea where fast moving ocean currents, expanses of deep water, and periodic episodes of sea level rise and fall act as soft and temporary allopatric barriers that facilitate both divergence and secondary contact. Under these conditions, gene flow is not expected to cease completely while contemporary distributions are expected to differ from historical ones. Here we conduct range-wide sampling for Pederson's cleaner shrimp (Ancylomenes pedersoni), a species complex from the Greater Caribbean that contains three clearly delimited mitochondrial lineages with both allopatric and sympatric distributions. Using mtDNA barcodes and a genomic ddRADseq approach, we combine classic phylogenetic analyses with extensive topology testing and demographic modeling (10 site frequency replicates x 45 evolutionary models x 50 model simulations/replicate = 22,500 simulations) to test species boundaries and reconstruct the evolutionary history of what was expected to be a simple case study. Instead, our results indicate a history of allopatric divergence, secondary contact, introgression, and endemic hybrid speciation that we hypothesize was driven by the final closure of the Isthmus of Panama and the strengthening of the Gulf Stream Current ~3.5 million years ago. The history of this species complex recovered by model-based methods that allow reticulation differs from that recovered by standard phylogenetic analyses and is unexpected given contemporary distributions. The geologically and biologically meaningful insights gained by our model selection analyses illuminate what is likely a novel pathway of species formation not previously documented that resulted from one of the most biogeographically significant events in Earth's history.

Sea anemone (Anthozoa, Actiniaria) diversity in Mo'orea (French Polynesia).

Sea anemones (Order Actiniaria) are a diverse group of marine invertebrates ubiquitous across marine ecosystems. Despite their wide distribution and success, a knowledge gap persists in our understanding of their diversity within tropical systems, owed to sampling bias of larger and more charismatic species overshadowing cryptic lineages. This study aims to delineate the sea anemone diversity in Mo'orea (French Polynesia) with the use of a dataset from the Mo'orea Biocode's "BioBlitz" initiative, which prioritized the sampling of more cryptic and understudied taxa. Implementing a target enrichment approach, we integrate 71 newly sequenced samples into an expansive phylogenetic framework and contextualize Mo'orea's diversity within global distribution patterns of sea anemones. Our analysis corroborates the presence of several previously documented sea anemones in French Polynesia and identifies for the first time the occurrence of members of genera Andvakia and Aiptasiomorpha. This research unveils the diverse sea anemone ecosystem in Mo'orea, spotlighting the area's ecological significance and emphasizing the need for continued exploration. Our methodology, encompassing a broad BLAST search coupled with phylogenetic analysis, proved to be a practical and effective approach for overcoming the limitations posed by the lack of comprehensive sequence data for sea anemones. We discuss the merits and limitations of current molecular methodologies and stress the importance of further research into lesser-studied marine organisms like sea anemones. Our work sets a precedent for future phylogenetic studies stemming from BioBlitz endeavors.

Towards the Exploration and Evolution of Insulin-like Venoms in Actiniaria (Sea anemones).

Recent studies have elucidated the diversity of genes encoding venom in Sea anemones. However, most of those genes are yet to be explored in an evolutionary context. Insulin is a common peptide across metazoans and has been coopted into a predatory venom in many venomous lineages. In this study, we focus on the diversity of insulin-derived venoms in Sea anemones and on elucidating their evolutionary history. We sourced data for 34 species of Sea anemones and found sequences belonging to two venom families which have Insulin PFAM annotations. Our findings show that both families have undergone duplication events. Members of each of the independently evolving clades have consistent predicted protein structures and distinct dN/dS values. Our work also shows that sequences allied with VP302 are part of a multidomain venom contig and have experienced a secondary gain into the venom system of cuticulate Sea anemones.

Venoms for all occasions: The functional toxin profiles of different anatomical regions in sea anemones are related to their ecological function.

The phylum Cnidaria is the oldest extant venomous group and is defined by the presence of nematocysts, specialized organelles responsible for venom production and delivery. Although toxin peptides and the cells housing nematocysts are distributed across the entire animal, nematocyte and venom profiles have been shown to differ across morphological structures in actiniarians. In this study, we explore the relationship between patterns of toxin expression and the ecological roles of discrete anatomical structures in Telmatactis stephensoni. Specifically, using a combination of proteomic and transcriptomic approaches, we examined whether there is a direct correlation between the functional similarity of regions and the similarity of their associated toxin expression profiles. We report that the regionalization of toxin production is consistent with the partitioning of the ecological roles of venom across envenomating structures, and that three major functional regions are present in T. stephensoni: tentacles, epidermis and gastrodermis. Additionally, we find that most structures that serve similar functions not only have comparable putative toxin profiles but also similar nematocyst types. There was no overlap in the putative toxins identified using proteomics and transcriptomics, but the expression patterns of specific milked venom peptides were conserved across RNA-sequencing and mass spectrometry imaging data sets. Furthermore, based on our data, it appears that acontia of T. stephensoni may be transcriptionally inactive and only mature nematocysts are present in the distal portions of the threads. Overall, we find that the venom profile of different anatomical regions in sea anemones varies according to its ecological functions.

Population genomics for symbiotic anthozoans: can reduced representation approaches be used for taxa without reference genomes?

Population genetic studies of symbiotic anthozoans have been historically challenging because their endosymbioses with dinoflagellates have impeded marker development. Genomic approaches like reduced representation sequencing alleviate marker development issues but produce anonymous loci, and without a reference genome, it is unknown which organism is contributing to the observed patterns. Alternative methods such as bait-capture sequencing targeting Ultra-Conserved Elements are now possible but costly. Thus, RADseq remains attractive, but how useful are these methods for symbiotic anthozoan taxa without a reference genome to separate anthozoan from algal sequences? We explore this through a case-study using a double-digest RADseq dataset for the sea anemone Bartholomea annulata. We assembled a holobiont dataset (3854 loci) for 101 individuals, then used a reference genome to create an aposymbiotic dataset (1402 loci). For both datasets, we investigated population structure and used coalescent simulations to estimate demography and population parameters. We demonstrate complete overlap in the spatial patterns of genetic diversity, demographic histories, and population parameter estimates for holobiont and aposymbiotic datasets. We hypothesize that the unique combination of anthozoan biology, diversity of the endosymbionts, and the manner in which assembly programs identify orthologous loci alleviates the need for reference genomes in some circumstances. We explore this hypothesis by assembling an additional 21 datasets using the assembly programs pyRAD and Stacks. We conclude that RADseq methods are more tractable for symbiotic anthozoans without reference genomes than previously realized.

Never, Ever Make an Enemy Out of an Anemone: Transcriptomic Comparison of Clownfish Hosting Sea Anemone Venoms.

Sea anemones are predatory marine invertebrates and have diverse venom arsenals. Venom is integral to their biology, and is used in competition, defense, and feeding. Three lineages of sea anemones are known to have independently evolved symbiotic relationships with clownfish, however the evolutionary impact of this relationship on the venom composition of the host is still unknown. Here, we investigate the potential of this symbiotic relationship to shape the venom profiles of the sea anemones that host clownfish. We use transcriptomic data to identify differences and similarities in venom profiles of six sea anemone species, representing the three known clades of clownfish-hosting sea anemones. We recovered 1121 transcripts matching verified toxins across all species, and show that hemolytic and hemorrhagic toxins are consistently the most dominant and diverse toxins across all species examined. These results are consistent with the known biology of sea anemones, provide foundational data on venom diversity of these species, and allow for a review of existing hierarchical structures in venomic studies.

Comparison of sequence-capture and ddRAD approaches in resolving species and populations in hexacorallian anthozoans.

Genome-level sequencing is the next step in understanding species-level relationships within Anthozoa (soft corals, anemones, stony corals, and their kin) as morphological and PCR-directed (single-locus) sequencing methods often fall short of differentiating species. The sea anemone genus Metridium is a common northern temperate sea anemone whose species are difficult to differentiate using morphology alone. Here we use Metridium as a case study to confirm the low level of information available in six loci for species differentiation commonly sequenced for Actiniaria and explore and compare the efficacy of ddRAD and sequence-capture methods in species-level systematics and biogeographic studies. We produce phylogenetic trees from concatenated datasets and perform DAPC and STRUCTURE analyses using SNP data. The six conventional loci are not able to consistently differentiate species within Metridium. The sequence-capture dataset resulted in high support and resolution for both current species and relationships between geographic areas. The ddRAD datasets displayed ambiguity among species, and support between major geographic groupings was not as high as the sequence-capture datasets. The level of resolution and support resulting from the sequence-capture data, combined with the ability to add additional individuals and expand beyond the genus Metridium over time, emphasizes the utility of sequence-capture methods for both systematics and future biogeographic studies within anthozoans. We discuss the strengths and weaknesses of the genomic approaches in light of our findings and suggest potential implications for the biogeography of Metridium based on our sampling.

Origin and evolution of the symbiosis between sea anemones (Cnidaria, Anthozoa, Actiniaria) and hermit crabs, with additional notes on anemone-gastropod associations.

The anemone-crab mutualism is ubiquitous in temperate and tropical marine environments. In this symbiosis, one or more anemones live on a shell inhabited by a hermit crab and reciprocal phoretic, trophic, and defensive benefits are exchanged between the partners. Sea anemone-hermit crab symbionts belong to three families: Hormathiidae (Calliactis and Paracalliactis), Sagartiidae (Carcinactis and Verrillactis), and Actiniidae (Stylobates). Hermit crabs establish most partnerships by detaching anemones and placing them on their shell; sea anemones can also mount shells unaided, triggered by a mollusc-derived substance in the periostracum of the shell. At least partial cooperation by the anemones is necessary for successful establishment of the symbiosis. Here, we expand the evolutionary framework for hormathiid symbionts by generating a phylogeny with at least one member of each actiniarian symbiotic genus with hermit crabs using five molecular markers (16S, 12S, 18S, 28S, CO3). We not only corroborated the results from a previous study by finding two origins of hermit crab symbiosis within Hormathiidae, but also found additional origins for hermit crab symbiosis within Actiniaria. We provide for the first time evidence of a close relationship between symbionts Carcinactis dolosa and V. paguri. The ability to secrete chitin by the ectoderm of the column is inferred to be broadly convergent within Actiniaria whereas the secretion of a chitinous carcinoecium by the pedal disc is a distinct but convergent morphological adaptation of several lineages within Actiniaria. Our finding of multiple origins for both the hermit crab and gastropod symbioses suggests that the shell-mounting behavior might only have been the precursor of the hermit crab association among Calliactis spp.

Genomic signatures of sympatric speciation with historical and contemporary gene flow in a tropical anthozoan (Hexacorallia: Actiniaria).

Sympatric diversification is recognized to have played an important role in the evolution of biodiversity. However, an in situ sympatric origin for codistributed taxa is difficult to demonstrate because different evolutionary processes can lead to similar biogeographic outcomes, especially in ecosystems that can readily facilitate secondary contact due to a lack of hard barriers to dispersal. Here we use a genomic (ddRADseq), model-based approach to delimit a species complex of tropical sea anemones that are codistributed on coral reefs throughout the Tropical Western Atlantic. We use coalescent simulations in fastsimcoal2 and ordinary differential equations in Moments to test competing diversification scenarios that span the allopatric-sympatric continuum. Our results suggest that the corkscrew sea anemone Bartholomea annulata is a cryptic species complex whose members are codistributed throughout their range. Simulation and model selection analyses from both approaches suggest these lineages experienced historical and contemporary gene flow, supporting a sympatric origin, but an alternative secondary contact model receives appreciable model support in fastsimcoal2. Leveraging the genome of the closely related Exaiptasia diaphana, we identify five loci under divergent selection between cryptic B. annulata lineages that fall within mRNA transcripts or CDS regions. Our study provides a rare empirical, genomic example of sympatric speciation in a tropical anthozoan and the first range-wide molecular study of a tropical sea anemone, underscoring that anemone diversity is under-described in the tropics, and highlighting the need for additional systematic studies into these ecologically and economically important species.

Phylogenetic relationships among the clownfish-hosting sea anemones.

The clownfish-sea anemone symbiosis has been a model system for understanding fundamental evolutionary and ecological processes. However, our evolutionary understanding of this symbiosis comes entirely from studies of clownfishes. A holistic understanding of a model mutualism requires systematic, biogeographic, and phylogenetic insight into both partners. Here, we conduct the largest phylogenetic analysis of sea anemones (Order Actiniaria) to date, with a focus on expanding the biogeographic and taxonomic sampling of the 10 nominal clownfish-hosting species. Using a combination of mtDNA and nuDNA loci we test (1) the monophyly of each clownfish-hosting family and genus, (2) the current anemone taxonomy that suggests symbioses with clownfishes evolved multiple times within Actiniaria, and (3) whether, like the clownfishes, there is evidence that host anemones have a Coral Triangle biogeographic origin. Our phylogenetic reconstruction demonstrates widespread poly- and para-phyly at the family and genus level, particularly within the family Stichodactylidae and genus Stichodactyla, and suggests that symbioses with clownfishes evolved minimally three times within sea anemones. We further recover evidence for a Tethyan biogeographic origin for some clades. Our data provide the first evidence that clownfish and some sea anemone hosts have different biogeographic origins, and that there may be cryptic species of host anemones. Finally, our findings reflect the need for a major taxonomic revision of the clownfish-hosting sea anemones.

Crowds Replicate Performance of Scientific Experts Scoring Phylogenetic Matrices of Phenotypes.

Scientists building the Tree of Life face an overwhelming challenge to categorize phenotypes (e.g., anatomy, physiology) from millions of living and fossil species. This biodiversity challenge far outstrips the capacities of trained scientific experts. Here we explore whether crowdsourcing can be used to collect matrix data on a large scale with the participation of nonexpert students, or "citizen scientists." Crowdsourcing, or data collection by nonexperts, frequently via the internet, has enabled scientists to tackle some large-scale data collection challenges too massive for individuals or scientific teams alone. The quality of work by nonexpert crowds is, however, often questioned and little data have been collected on how such crowds perform on complex tasks such as phylogenetic character coding. We studied a crowd of over 600 nonexperts and found that they could use images to identify anatomical similarity (hypotheses of homology) with an average accuracy of 82% compared with scores provided by experts in the field. This performance pattern held across the Tree of Life, from protists to vertebrates. We introduce a procedure that predicts the difficulty of each character and that can be used to assign harder characters to experts and easier characters to a nonexpert crowd for scoring. We test this procedure in a controlled experiment comparing crowd scores to those of experts and show that crowds can produce matrices with over 90% of cells scored correctly while reducing the number of cells to be scored by experts by 50%. Preparation time, including image collection and processing, for a crowdsourcing experiment is significant, and does not currently save time of scientific experts overall. However, if innovations in automation or robotics can reduce such effort, then large-scale implementation of our method could greatly increase the collective scientific knowledge of species phenotypes for phylogenetic tree building. For the field of crowdsourcing, we provide a rare study with ground truth, or an experimental control that many studies lack, and contribute new methods on how to coordinate the work of experts and nonexperts. We show that there are important instances in which crowd consensus is not a good proxy for correctness.

Phylogenetic analysis reveals an evolutionary transition from internal to external brooding in Epiactis Verrill (Cnidaria: Anthozoa: Actiniaria) and rejects the validity of the genus Cnidopus Carlgren.

Reproductive behaviors in the sea anemone genus Epiactis provide an opportunity for investigating the evolution of reproductive phenomena such as brooding and sex allocation (hermaphroditic vs. gonochoric) in a group of closely related and easily accessible species. However, given its broad geographic distribution, the striking diversity in reproductive behaviors, and the lack of synapomorphy for the genus, the monophyly of Epiactis is questionable. Here we perform phylogenetic analyses to test the monophyly of Epiactis and the validity of Cnidopus, which consists entirely of species once assigned to Epiactis. We use the large number of brooding species in Epiactis to investigate evolutionary patterns in brooding modes and characteristics associated with them. We find a monophyletic group of North Pacific Epiactis species: this group includes the type species of the genus and species that brood internally or externally, and that are hermaphroditic or gonochoric. Based on the results, we reject the genus Cnidopus because its circumscription renders Epiactis sensu stricto paraphyletic. Ancestral character state reconstruction indicates that in the North Pacific, externally brooding species evolved from internally brooding ancestors and that sex allocation is highly labile. Species relationships in Epiactis and Aulactinia appear to conform to geographic patterns more strongly than to taxonomic hypotheses. Contrary to expectations based on other invertebrates, we fail to find a strong correlation between brooding and hermaphroditism.

A RNA-seq approach to identify putative toxins from acrorhagi in aggressive and non-aggressive Anthopleura elegantissima polyps.

BACKGROUND: The use of venom in intraspecific aggression is uncommon and venom-transmitting structures specifically used for intraspecific competition are found in few lineages of venomous taxa. Next-generation transcriptome sequencing allows robust characterization of venom diversity and exploration of functionally unique tissues. Using a tissue-specific RNA-seq approach, we investigate the venom composition and gene ontology diversity of acrorhagi, specialized structures used in intraspecific competition, in aggressive and non-aggressive polyps of the aggregating sea anemone Anthopleura elegantissima (Cnidaria: Anthozoa: Hexacorallia: Actiniaria: Actiniidae). RESULTS: Collectively, we generated approximately 450,000 transcripts from acrorhagi of aggressive and non-aggressive polyps. For both transcriptomes we identified 65 candidate sea anemone toxin genes, representing phospholipase A2s, cytolysins, neurotoxins, and acrorhagins. When compared to previously characterized sea anemone toxin assemblages, each transcriptome revealed greater within-species sequence divergence across all toxin types. The transcriptome of the aggressive polyp had a higher abundance of type II voltage gated potassium channel toxins/Kunitz-type protease inhibitors and type II acrorhagins. Using toxin-like proteins from other venomous taxa, we also identified 612 candidate toxin-like transcripts with signaling regions, potentially unidentified secretory toxin-like proteins. Among these, metallopeptidases and cysteine rich (CRISP) candidate transcripts were in high abundance. Furthermore, our gene ontology analyses identified a high prevalence of genes associated with "blood coagulation" and "positive regulation of apoptosis", as well as "nucleoside: sodium symporter activity" and "ion channel binding". The resulting assemblage of expressed genes may represent synergistic proteins associated with toxins or proteins related to the morphology and behavior exhibited by the aggressive polyp. CONCLUSION: We implement a multifaceted approach to investigate the assemblage of expressed genes specifically within acrorhagi, specialized structures used only for intraspecific competition. By combining differential expression, phylogenetic, and gene ontology analyses, we identify several candidate toxins and other potentially important proteins in acrorhagi of A. elegantissima. Although not all of the toxins identified are used in intraspecific competition, our analysis highlights some candidates that may play a vital role in intraspecific competition. Our findings provide a framework for further investigation into components of venom used exclusively for intraspecific competition in acrorhagi-bearing sea anemones and potentially other venomous animals.

Bellactis lux n. sp. (Cnidaria: Anthozoa: Actiniaria: Aiptasiidae), a new sea anemone from the Gulf of Mexico.

Here we describe a new species of sea anemone from the family Aiptasiidae based on specimens collected from the Gulf of Mexico (USA: Florida & Alabama). Accounts of this species have been known since the early 1990s, primarily from an underwater field guide and hobbyist aquarium literature under the name Lightbulb Anemone. We describe it as a new species from the genus Bellactis based on anatomy, histology, and cnidom. Members of this species are small in size, with a smooth, typically contracted column divided into regions based on color and bearing rows of two or three elevated cinclides in the mid column. Their tentacles are distinctive, translucent, distally inflated and can be bulbous in shape, with sub annular rings. This description synthesizes information about Bellactis and contextualizes what is known about its diversity in light of other members of the Aiptasiidae.

There and Back Again: The Unexpected Journeys of Metridium de Blainville, 1824 between the Old Oceans and throughout the Modern World.

AbstractMembers of the sea anemone genus Metridium are abundant in temperate rocky habitats and fouling communities. Their biogeographic history is expected to reflect changes in currents and habitats that have influenced benthic communities, such as the climate-influenced changes that occurred during the Last Glacial Maximum. More recently, however, anthropogenic influences such as shipping transportation and the creation of artificial habitat have altered and affected the composition of modern-day marine communities. Here we use sequence-capture data to examine the genetic structure of Metridium across its shallow-water distribution to (1) evaluate species boundaries within Metridium, (2) elucidate the dispersal history of Metridium between and among oceans, and (3) assess the influence of anthropogenic movement on modern-day populations. We find strong evidence for two species within Metridium: M. farcimen and M. senile. Dispersal from the Pacific to the Atlantic included a subsequent isolation of a small population in or above the Bering Sea, which has presumably moved southward. Within the native range of M. senile, admixture is prevalent even between oceans as a result of anthropogenic activities. The nonnative populations in Chile and the Falkland Islands came from at least two distinct introduction events originating from both coasts of the United States in the North Pacific and North Atlantic Oceans. Hybridization between M. senile and M. farcimen is documented as occurring in anthropogenically influenced habitats. The heavy influence from anthropogenic activities will continue to impact our understanding of marine organisms, particularly within the native range and for those that are easily transported across long distances.

Single-cell atavism reveals an ancient mechanism of cell type diversification in a sea anemone.

Cnidocytes are the explosive stinging cells unique to cnidarians (corals, jellyfish, etc). Specialized for prey capture and defense, cnidocytes comprise a group of over 30 morphologically and functionally distinct cell types. These unusual cells are iconic examples of biological novelty but the developmental mechanisms driving diversity of the stinging apparatus are poorly characterized, making it challenging to understand the evolutionary history of stinging cells. Using CRISPR/Cas9-mediated genome editing in the sea anemone Nematostella vectensis, we show that a single transcription factor (NvSox2) acts as a binary switch between two alternative stinging cell fates. Knockout of NvSox2 causes a transformation of piercing cells into ensnaring cells, which are common in other species of sea anemone but appear to have been silenced in N. vectensis. These results reveal an unusual case of single-cell atavism and expand our understanding of the diversification of cell type identity.

Echinoderm phylogeny including Xyloplax, a progenetic asteroid.

Reconstruction of the phylogeny of the five extant classes of the phylum Echinodermata has proven difficult. Results concerning higher-level taxonomic relationships among echinoderms are sensitive to the choice of analytical parameters and methods. Moreover, the proposal of a putative sixth class based on a small enigmatic disc-shaped echinoderm, Xyloplax, from the deep seas of the Bahamas and New Zealand in the 1980s further complicated the problem. Although clearly an echinoderm, Xyloplax did not have clear affinity among known groups. Using molecular sequence and developmental data from recently collected Xyloplax adults and embryos, we show that rather than representing an ancient distinct lineage as implied by its status as a class, Xyloplax is simply a starfish that is closely related to the asteroid family Pterasteridae. Many members of the Pterasteridae and all Xyloplax inhabit deep or polar seas and brood young. Brooding pterasterids and Xyloplax hold their young in specialized adult chambers until the young reach an advanced juvenile stage after which they are released as free-living individuals. We hypothesize that the unique morphology of Xyloplax evolved via progenesis--the truncation of somatic growth at a juvenile body plan but with gonadal growth to maturity. Although the overall phylogeny of extant echinoderms remains sensitive to the choice of analytical methods, the placement of Xyloplax as sister to pterasterid asteroids is unequivocal. Based on this, we argue that the proposed class and infraclass status of Xyloplax should be suppressed.

Toward a natural classification: phylogeny of acontiate sea anemones (Cnidaria, Anthozoa, Actiniaria).

Acontia-nematocyst-dense, thread-like extensions of the mesenterial filaments-are the characteristic feature of the actiniarian group Acontiaria. Phylogenetic analyses have shown that acontiate taxa form a clade that also includes some taxa without acontia. We analyse five molecular markers from 85 actiniarians to explore the phylogenetic relationships among families in Acontiaria, including acontiate species assigned to other higher taxa and species without acontia that have been allied to Acontiaria. Based on our results, we redefine the group to accommodate those lineages that have lost acontia, and formalize it as superfamily Metridioidea. Based on stable and well supported clades, we resurrect Phelliidae and Amphianthidae, redefine Kadosactinidae and Actinoscyphiidae, and move two species to new genera: that previously termed Sagartiogeton erythraios belongs in Jasonactis gen. nov.; and that previously termed Anthosactis pearseae belongs in Ostiactis gen. nov., type genus of Ostiactinidae fam. nov. We also synonymized Halcampoididae and Halcampidae (as Halcampidae) and Andvakiidae and Isophelliidae (as Andvakiidae). The results of our phylogenetic analyses indicate that the diagnostic morphological characters used in the family-level taxonomy of acontiate actiniarians such as the nematocysts of the acontia, the marginal sphincter muscle, and mesenteries divisible into macro- and micro-cnemes, have to be revisited, as these features are highly homoplasious.

Phylogeny and taxonomy of Haloclavidae (Verrill, 1899) with a redescription of the parasitic, burrowing sea anemone, Peachia chilensis Carlgren, 1931.

Haloclavidae Verrill, 1899 is a family of burrowing sea anemones grouped within the superfamily Actinioidea (Rafinesque, 1815). Currently, it includes 30 species in 10 genera. Characters given for this family in descriptions of its taxa have not been consistent, with numerous exceptions to the expectations of the familial diagnosis. Previous phylogenetic analyses have shown that Haloclavidae is potentially a polyphyletic group, but resolution of relationships of the few representatives of Haloclavidae included in analyses has been problematic. Here we address questions of monophyly and affinity of Haloclavidae using three mitochondrial and two nuclear markers. We assess the monophyly of Haloclavidae in the context of all major lineages of Actiniaria Hertwig, 1882, emphasizing diversity of superfamily Actinioidea. We use parsimony-based character optimization to interpret the distribution of key traits in the superfamily. We find that Haloclavidae is not monophyletic and propose two new families, Peachiidae fam. nov. and Harenactidae fam. nov., while also retaining some species in the family Haloclavidae, so that taxonomy better reflects relationships and diversity of the group. In addition, we redescribe a species within the newly created Peachiidae, Peachia chilensis Carlgren, 1931. We use recent larval samples obtained in Antofagasta, Chile, and the histological slides from the original description to redescribe P. chilensis, to provide a complete account of cnidae, external, and internal morphology. Finally, we compare P. chilensis to other burrowing anemones found in Chile and provide an understanding of the genus Peachia that reflects recent phylogenetic perspective on diversity of anemones previously assigned to family Haloclavidae.

Freshwater mussels (Unionidae) brought into captivity exhibit up-regulation of genes involved in stress and energy metabolism.

Approximately two thirds of freshwater mussel species in the United States and Canada are imperiled, and populations are declining rapidly. Translocation and captive management are commonly used to mitigate losses of freshwater mussel biodiversity, but these conservation tools may result in decreased growth and increased mortality. This study uses RNA-Seq to determine how translocation into captivity affects gene expression in Amblema plicata. Mussels were collected from the Muskingum River in Ohio, USA and brought into a captive holding facility. RNA was extracted from gill tissue 11 months post translocation from mussels in captivity and the Muskingum River on the same day. RNA was sequenced on an Illumina HiSeq 2500, and differential expression analysis was performed on de novo assembled transcripts. More than 1200 transcripts were up-regulated in captive mussels, and 246 were assigned functional annotations. Many up-regulated transcripts were involved in energy metabolism and the stress response, such as heat shock proteins and antioxidants. More than 500 transcripts were down-regulated in captive mussels, and 41 were assigned functional annotations. We observed an over-representation of down-regulated transcripts associated with immune response. Our work suggests that A. plicata experienced moderate levels of stress and altered energy metabolism and immune response for at least 11 months post translocation into captivity.