Local and global changes in cell density induce reorganisation of 3D packing in a proliferating epithelium.

Tissue morphogenesis is intimately linked to the changes in shape and organisation of individual cells. In curved epithelia, cells can intercalate along their own apicobasal axes, adopting a shape named 'scutoid' that allows energy minimization in the tissue. Although several geometric and biophysical factors have been associated with this 3D reorganisation, the dynamic changes underlying scutoid formation in 3D epithelial packing remain poorly understood. Here, we use live imaging of the sea star embryo coupled with deep learning-based segmentation to dissect the relative contributions of cell density, tissue compaction and cell proliferation on epithelial architecture. We find that tissue compaction, which naturally occurs in the embryo, is necessary for the appearance of scutoids. Physical compression experiments identify cell density as the factor promoting scutoid formation at a global level. Finally, the comparison of the developing embryo with computational models indicates that the increase in the proportion of scutoids is directly associated with cell divisions. Our results suggest that apico-basal intercalations appearing immediately after mitosis may help accommodate the new cells within the tissue. We propose that proliferation in a compact epithelium induces 3D cell rearrangements during development.

Top predator sea stars are the benthic equivalent to polar bears of the pelagic realm.

The marine pelagic compartment spans numerous trophic levels and consists of numerous reticulate connections between species from primary producers to iconic apex predators, while the benthic compartment is perceived to be simpler in structure and comprised of only low trophic level species. Here, we challenge this paradigm by illustrating that the benthic compartment is home to a subweb of similar structure and complexity to that of the pelagic realm, including the benthic equivalent to iconic polar bears: megafaunal-predatory sea stars.

Single-cell RNA-sequencing analysis of early sea star development.

Echinoderms represent a broad phylum with many tractable features to test evolutionary changes and constraints. Here, we present a single-cell RNA-sequencing analysis of early development in the sea star Patiria miniata, to complement the recent analysis of two sea urchin species. We identified 20 cell states across six developmental stages from 8 hpf to mid-gastrula stage, using the analysis of 25,703 cells. The clusters were assigned cell states based on known marker gene expression and by in situ RNA hybridization. We found that early (morula, 8-14 hpf) and late (blastula-to-mid-gastrula) cell states are transcriptionally distinct. Cells surrounding the blastopore undergo rapid cell state changes that include endomesoderm diversification. Of particular import to understanding germ cell specification is that we never see Nodal pathway members within Nanos/Vasa-positive cells in the region known to give rise to the primordial germ cells (PGCs). The results from this work contrast the results of PGC specification in the sea urchin, and the dataset presented here enables deeper comparative studies in tractable developmental models for testing a variety of developmental mechanisms.

The impact of acute low salinity stress on Antarctic echinoderms.

Climate change is causing increased coastal freshening in Antarctica, leading to reduced salinity. For Antarctica's endemic echinoderms, adapted to the stable polar environment, the impact of rapid reductions in coastal salinity on physiology and behaviour is currently unknown. Six common Antarctic echinoderms (the sea urchin Sterechinus neumayeri; the sea star Odontaster validus; the brittle star Ophionotus victoriae; and three sea cucumbers Cucumaria georgiana, Echinopsolus charcoti and Heterocucumis steineni), were directly transferred from ambient salinity (34.5‰) to a range of salinity dilutions (29-9‰) for 24 h. All species showed reduced activity and the establishment of a temporary osmotic gradient between coelomic fluid and external seawater. Most species exhibited a depression in oxygen consumption across tolerated salinities; however, at very low salinities that later resulted in mortality, oxygen consumption increased to levels comparable to those at ambient. Low salinity tolerance varied substantially between species, with O. victoriae being the least tolerant (24 h LC50 (lethal for 50% of animals) = 19.9‰) while E. charcoti and C. georgiana demonstrated the greatest tolerance (24 h LC50 = 11.5‰). These findings demonstrate the species-specific response of Antarctica's endemic echinoderms to short-term hypoosmotic salinity events, providing valuable insight into this phylum's ability to respond to an underreported impact of climate change.

Finding food: how generalist predators use contact-chemosensory information to guide prey preferences.

Understanding the processes that guide carnivores in finding and selecting prey is a fundamental, unresolved challenge in sensory biology. To our knowledge, no published work has yet revealed the complete structural identities of compounds that cue preferences by generalist predators for different prey species. With this research imperative in mind, we determined the chemistry driving consumer preferences for live intact prey using two generalist predatory species (sea stars, Pisaster ochraceus; whelks, Acanthinucella spirata), along with two foundation prey species (mussels, Mytilus californianus; barnacles, Balanus glandula), inhabiting rocky, wave-swept shores. Each prey species is known to secrete either a 29.6 kDa (named 'KEYSTONEin') or a 199.6 kDa (named 'MULTIFUNCin') glycoprotein as a contact-chemical cue. Here, experimental manipulations utilized faux prey consisting of cleaned barnacle or mussel shells infused with KEYSTONEin, MULTIFUNCin or seawater (control) gels. Whelks exhibited a strong penchant for MULTIFUNCin over KEYSTONEin, irrespective of shell type. In contrast, sea stars generally preferred KEYSTONEin over MULTIFUNCin, but this preference shifted depending on the experimental context in which they encountered physical (shell) and chemical (glycoprotein) stimuli. This study ultimately demonstrates clear and contrasting chemical preferences between sea stars and whelks. It highlights the importance of experimental setting in determining chemical preferences. Finally, it shows that prey preferences by these predators hinge only on one or two contact-protein cues, without the need for quality coding via fluid-borne compounds, low-molecular-weight substances or mixture blends.

A reference genome for ecological restoration of the sunflower sea star, Pycnopodia helianthoides.

Wildlife diseases, such as the sea star wasting (SSW) epizootic that outbroke in the mid-2010s, appear to be associated with acute and/or chronic abiotic environmental change; dissociating the effects of different drivers can be difficult. The sunflower sea star, Pycnopodia helianthoides, was the species most severely impacted during the SSW outbreak, which overlapped with periods of anomalous atmospheric and oceanographic conditions, and there is not yet a consensus on the cause(s). Genomic data may reveal underlying molecular signatures that implicate a subset of factors and, thus, clarify past events while also setting the scene for effective restoration efforts. To advance this goal, we used Pacific Biosciences HiFi long sequencing reads and Dovetail Omni-C proximity reads to generate a highly contiguous genome assembly that was then annotated using RNA-seq-informed gene prediction. The genome assembly is 484 Mb long, with contig N50 of 1.9 Mb, scaffold N50 of 21.8 Mb, BUSCO completeness score of 96.1%, and 22 major scaffolds consistent with prior evidence that sea star genomes comprise 22 autosomes. These statistics generally fall between those of other recently assembled chromosome-scale assemblies for two species in the distantly related asteroid genus Pisaster. These novel genomic resources for P. helianthoides will underwrite population genomic, comparative genomic, and phylogenomic analyses-as well as their integration across scales-of SSW and environmental stressors.

Sulfated Polyhydroxysteroid Glycosides from the Sea of Okhotsk Starfish Henricia leviuscula spiculifera and Potential Mechanisms for Their Observed Anti-Cancer Activity against Several Types of Human Cancer Cells.

Three new monosulfated polyhydroxysteroid glycosides, spiculiferosides A (1), B (2), and C (3), along with new related unsulfated monoglycoside, spiculiferoside D (4), were isolated from an ethanolic extract of the starfish Henricia leviuscula spiculifera collected in the Sea of Okhotsk. Compounds 1-3 contain two carbohydrate moieties, one of which is attached to C-3 of the steroid tetracyclic core, whereas another is located at C-24 of the side chain of aglycon. Two glycosides (2, 3) are biosides, and one glycoside (1), unlike them, includes three monosaccharide residues. Such type triosides are a rare group of polar steroids of sea stars. In addition, the 5-substituted 3-OSO3-α-L-Araf unit was found in steroid glycosides from starfish for the first time. Cell viability analysis showed that 1-3 (at concentrations up to 100 µM) had negligible cytotoxicity against human embryonic kidney HEK293, melanoma SK-MEL-28, breast cancer MDA-MB-231, and colorectal carcinoma HCT 116 cells. These compounds significantly inhibited proliferation and colony formation in HCT 116 cells at non-toxic concentrations, with compound 3 having the greatest effect. Compound 3 exerted anti-proliferative effects on HCT 116 cells through the induction of dose-dependent cell cycle arrest at the G2/M phase, regulation of expression of cell cycle proteins CDK2, CDK4, cyclin D1, p21, and inhibition of phosphorylation of protein kinases c-Raf, MEK1/2, ERK1/2 of the MAPK/ERK1/2 pathway.

Complete Mitochondrial Genomes and Phylogenetic Analysis of Genus Henricia (Asteroidea: Spinulosida: Echinasteridae).

The genus Henricia is known to have intraspecific morphological variations, making species identification difficult. Therefore, molecular phylogeny analysis based on genetic characteristics is valuable for species identification. We present complete mitochondrial genomic sequences of Henricia longispina aleutica, H. reniossa, and H. sanguinolenta for the first time in this study. This study will make a significant contribution to our understanding of Henricia species and its relationships within the class Asteroidea. Lengths of mitochondrial genomes of the three species are 16,217, 16,223, and 16,194 bp, respectively, with a circular form. These genomes contained 13 protein-coding genes, two ribosomal RNA genes, 22 transfer RNA genes, and a D-loop. The gene order and direction aligned with other asteroid species. Phylogenetic relationship analysis showed that our Henricia species were in a monophyletic clade with other Henricia species and in a large clade with species (Echinaster brasiliensis) from the same family. These findings provide valuable insight into understanding the phylogenetic relationships of species in the genus Henricia.

A conserved node in the regulation of Vasa between an induced and an inherited program of primordial germ cell specification.

Primordial germ cells (PGCs) are specified by diverse mechanisms in early development. In some animals, PGCs are specified via inheritance of maternal determinants, while in others, in a process thought to represent the ancestral mode, PGC fate is induced by cell interactions. Although the terminal factors expressed in specified germ cells are widely conserved, the mechanisms by which these factors are regulated can be widely diverse. Here we show that a post-translational mechanism of germ cell specification is conserved between two echinoderm species thought to employ divergent germ line segregation strategies. Sea urchins segregate their germ line early by an inherited mechanism. The DEAD-box RNA - helicase Vasa, a conserved germline factor, becomes enriched in the PGCs by degradation in future somatic cells by the E3-ubiquitin-ligase Gustavus (Gustafson et al., 2011). This post-translational activity occurs early in development, substantially prior to gastrulation. Here we test this process in germ cell specification of sea star embryos, which use inductive signaling mechanisms after gastrulation for PGC fate determination. We find that Vasa-GFP protein becomes restricted to the PGCs in the sea star even though the injected mRNA is present throughout the embryo. Gustavus depletion, however, results in uniform accumulation of the protein. These data demonstrate that Gustavus-mediated Vasa turnover in somatic cells is conserved between species with otherwise divergent PGC specification mechanisms. Since Gustavus was originally identified in Drosophila melanogaster to have similar functions in Vasa regulation (Kugler et al., 2010), we conclude that this node of Vasa regulation in PGC formation is ancestral and evolutionarily transposable from the ancestral, induced PGC specification program to an inherited PGC specification mechanism.

Optimizing CRISPR/Cas9-based gene manipulation in echinoderms.

The impact of new technology can be appreciated by how broadly it is used. Investigators that previously relied only on pharmacological approaches or the use of morpholino antisense oligonucleotide (MASO) technologies are now able to apply CRISPR-Cas9 to study biological problems in their model organism of choice much more effectively. The transitions to new CRISPR-based approaches could be enhanced, first, by standardized protocols and education in their applications. Here we summarize our results for optimizing the CRISPR-Cas9 technology in a sea urchin and a sea star, and provide advice on how to set up CRISPR-Cas9 experiments and interpret the results in echinoderms. Our goal through these protocols and sharing examples of success by other labs is to lower the activation barrier so that more laboratories can apply CRISPR-Cas9 technologies in these important animals.

Sea stars resist wasting through active immune and collagen systems.

Epidemics are becoming more common and severe, however, pinpointing the causes can be challenging, particularly in marine environments. The cause of sea star wasting (SSW) disease, the ongoing, largest known panzootic of marine wildlife, is unresolved. Here, we measured gene expression longitudinally of 24 adult Pisaster ochraceus sea stars, collected from a recovered site, as they remained asymptomatic (8 individuals) or naturally progressed through SSW (16 individuals) in individual aquaria. Immune, tissue integrity and pro-collagen genes were more highly expressed in asymptomatic relative to wasting individuals, while hypoxia-inducible factor 1-α and RNA processing genes were more highly expressed in wasting relative to asymptomatic individuals. Integrating microbiome data from the same tissue samples, we identified genes and microbes whose abundance/growth was associated with disease status. Importantly, sea stars that remained visibly healthy showed that laboratory conditions had little effect on microbiome composition. Lastly, considering genotypes at 98 145 single-nucleotide polymorphism, we found no variants associated with final health status. These findings suggest that animals exposed to the cause(s) of SSW remain asymptomatic with an active immune response and sustained control of their collagen system while animals that succumb to wasting show evidence of responding to hypoxia and dysregulation of RNA processing systems.

Coelomic fluid of asteroid echinoderms: Current knowledge and future perspectives on its utility for disease and mortality investigations.

Coelomic fluid surrounds the internal organs of asteroid echinoderms (asteroids, otherwise known as sea stars or starfish) and plays an essential role in the immune system, as well as in the transport of respiratory gases, nutrients, waste products, and reproductive mediators. Due to its importance in physiology and accessibility for nonlethal diagnostic sampling, coelomic fluid of asteroids provides an excellent sample matrix for health evaluations and can be particularly useful in disease and mortality investigations. This is especially important in light of recent increases in the number of affected individuals and species, larger geographic scope, and increased observed frequency of sea star wasting events compared with historic accounts of wasting. This review summarizes the current knowledge about coelomocytes, the effector cell of the asteroid immune system; coelomic fluid electrolytes, osmolality, acid-base status and respiratory gases, and microbiota; and genomic, transcriptomic, and proteomic investigations of coelomic fluid. The utility of coelomic fluid analysis for assessing stressor responses, diseases, and mortality investigations is considered with knowledge gaps and future directions identified. This complex body fluid provides an exciting opportunity to increase our understanding of this unique and ecologically important group of animals.

Sex-specific expression of pheromones and other signals in gravid starfish.

BACKGROUND: Many echinoderms form seasonal aggregations prior to spawning. In some fecund species, a spawning event can lead to population outbreaks with detrimental ecosystem impacts. For instance, outbreaks of crown-of-thorns starfish (COTS), a corallivore, can destroy coral reefs. Here, we examine the gene expression in gravid male and female COTS prior to spawning in the wild, to identify genome-encoded factors that may regulate aggregation and spawning. This study is informed by a previously identified exoproteome that attracts conspecifics. To capture the natural gene expression profiles, we isolated RNAs from gravid female and male COTS immediately after they were removed from the Great Barrier Reef.  RESULTS: Sexually dimorphic gene expression is present in all seven somatic tissues and organs that we surveyed and in the gonads. Approximately 40% of the exoproteome transcripts are differentially expressed between sexes. Males uniquely upregulate an additional 68 secreted factors in their testes. A suite of neuropeptides in sensory organs, coelomocytes and gonads is differentially expressed between sexes, including the relaxin-like gonad-stimulating peptide and gonadotropin-releasing hormones. Female sensory tentacles-chemosensory organs at the distal tips of the starfish arms-uniquely upregulate diverse receptors and signalling molecules, including chemosensory G-protein-coupled receptors and several neuropeptides, including kisspeptin, SALMFamide and orexin. CONCLUSIONS: Analysis of 103 tissue/organ transcriptomes from 13 wild COTS has revealed genes that are consistently differentially expressed between gravid females and males and that all tissues surveyed are sexually dimorphic at the molecular level. This finding is consistent with female and male COTS using sex-specific pheromones to regulate reproductive aggregations and synchronised spawning events. These pheromones appear to be received primarily by the sensory tentacles, which express a range of receptors and signalling molecules in a sex-specific manner. Furthermore, coelomocytes and gonads differentially express signalling and regulatory factors that control gametogenesis and spawning in other echinoderms.

Little evidence for genetic variation associated with susceptibility to sea star wasting syndrome in the keystone species Pisaster ochraceus.

The keystone species Pisaster ochraceus suffered mass mortalities along the northeast Pacific Ocean from Sea Star Wasting Syndrome (SSWS) outbreaks in 2013-2016. SSWS causation remains of debate, leading to concerns as to whether outbreaks will continue to impact this species. Considering the apparent link between ocean temperature and SSWS, the future of this species and intertidal communities remains uncertain. Surveys of co-occurring apparently normal and wasting P. ochraceus along the central Oregon coast in 2016 allowed us to address whether variation in disease status showed genetic variation that may be associated with differences in susceptibility to SSWS. We performed restriction site-associated DNA sequencing (2bRAD-seq) to genotype ~72,000 single nucleotide polymorphism (SNP) loci across apparently normal and wasting sea stars. Locus-specific analyses of differentiation (FST ) between disease-status groups revealed no signal of genetic differences separating the two groups. Using a multivariate approach, we observed weak separation between the groups, but identified 18 SNP loci showing highest discriminatory power between the groups and scanned the genome annotation for linked genes. A total of 34 protein-coding genes were found to be located within 15 kb (measured by linkage disequilibrium decay) of at least one of the 18 SNPs, and 30 of these genes had homologies to annotated protein databases. Our results suggest that the likelihood of developing SSWS symptoms does not have a strong genetic basis. The few genomic regions highlighted had only modest levels of differentiation, but the genes associated with these regions may form the basis for functional studies aiming to understand disease progression.

The first records of sea star wasting disease in Crossaster papposus in Europe.

Sea star wasting disease (SSWD) refers to a suite of gross pathological signs observed in Asteroidea species. It presents to varying degrees as abnormal posture, epidermal ulceration, arm autotomy and eversion of viscera. We report observations of SSWD in the sunstar Crossaster papposus, the first observations of its kind in Europe. While the exact cause of SSWD remains unknown, studies have proposed pathogenic and environmental-stress pathways for disease outbreaks. Although the present observations do not support a precise aetiology, the presence of SSWD in a keystone predator may have wide reaching ecological and management implications.

Nodal induces sequential restriction of germ cell factors during primordial germ cell specification.

Specification of the germ cell lineage is required for sexual reproduction in animals. The mechanism of germ cell specification varies among animals but roughly clusters into either inherited or inductive mechanisms. The inductive mechanism, the use of cell-cell interactions for germ cell specification, appears to be the ancestral mechanism in animal phylogeny, yet the pathways responsible for this process are only recently surfacing. Here, we show that germ cell factors in the sea star initially are present broadly, then become restricted dorsally and then in the left side of the embryo where the germ cells form a posterior enterocoel. We find that Nodal signaling is required for the restriction of two germ cell factors, Nanos and Vasa, during the early development of this animal. We learned that Nodal inhibits germ cell factor accumulation in three ways including: inhibition of specific transcription, degradation of specific mRNAs and inhibition of tissue morphogenesis. These results document a signaling mechanism required for the sequential restriction of germ cell factors, which causes a specific set of embryonic cells to become the primordial germ cells.

Disease-driven mass mortality event leads to widespread extirpation and variable recovery potential of a marine predator across the eastern Pacific.

The prevalence of disease-driven mass mortality events is increasing, but our understanding of spatial variation in their magnitude, timing and triggers are often poorly resolved. Here, we use a novel range-wide dataset comprised 48 810 surveys to quantify how sea star wasting disease affected Pycnopodia helianthoides, the sunflower sea star, across its range from Baja California, Mexico to the Aleutian Islands, USA. We found that the outbreak occurred more rapidly, killed a greater percentage of the population and left fewer survivors in the southern half of the species's range. Pycnopodia now appears to be functionally extinct (greater than 99.2% declines) from Baja California, Mexico to Cape Flattery, Washington, USA and exhibited severe declines (greater than 87.8%) from the Salish Sea to the Gulf of Alaska. The importance of temperature in predicting Pycnopodia distribution rose more than fourfold after the outbreak, suggesting latitudinal variation in outbreak severity may stem from an interaction between disease severity and warmer waters. We found no evidence of population recovery in the years since the outbreak. Natural recovery in the southern half of the range is unlikely over the short term. Thus, assisted recovery will probably be required to restore the functional role of this predator on ecologically relevant time scales.

Diversity of Sea Star-Associated Densoviruses and Transcribed Endogenous Viral Elements of Densovirus Origin.

A viral etiology of sea star wasting syndrome (SSWS) was originally explored with virus-sized material challenge experiments, field surveys, and metagenomics, leading to the conclusion that a densovirus is the predominant DNA virus associated with this syndrome and, thus, the most promising viral candidate pathogen. Single-stranded DNA viruses are, however, highly diverse and pervasive among eukaryotic organisms, which we hypothesize may confound the association between densoviruses and SSWS. To test this hypothesis and assess the association of densoviruses with SSWS, we compiled past metagenomic data with new metagenomic-derived viral genomes from sea stars collected from Antarctica, California, Washington, and Alaska. We used 179 publicly available sea star transcriptomes to complement our approaches for densovirus discovery. Lastly, we focus the study on sea star-associated densovirus (SSaDV), the first sea star densovirus discovered, by documenting its biogeography and putative tissue tropism. Transcriptomes contained only endogenized densovirus elements similar to the NS1 gene, while numerous extant densoviral genomes were recovered from viral metagenomes. SSaDV was associated with nearly all tested species from southern California to Alaska, and in contrast to previous work, we show that SSaDV is one genotype among a high diversity of densoviruses present in sea stars across the West Coast of the United States and globally that are commonly associated with grossly normal (i.e., healthy or asymptomatic) animals. The diversity and ubiquity of these viruses in sea stars confound the original hypothesis that one densovirus is the etiological agent of SSWS.IMPORTANCE The primary interest in sea star densoviruses, specifically SSaDV, has been their association with sea star wasting syndrome (SSWS), a disease that has decimated sea star populations across the West Coast of the United States since 2013. The association of SSaDV with SSWS was originally drawn from metagenomic analysis, which was further studied through field surveys using quantitative PCR (qPCR), with the conclusion that it was the most likely viral candidate in the metagenomic data based on its representation in symptomatic sea stars compared to asymptomatic sea stars. We reexamined the original metagenomic data with additional genomic data sets and found that SSaDV was 1 of 10 densoviruses present in the original data set and was no more represented in symptomatic sea stars than in asymptomatic sea stars. Instead, SSaDV appears to be a widespread, generalist virus that exists among a large diversity of densoviruses present in sea star populations.

A Highly Prevalent and Pervasive Densovirus Discovered among Sea Stars from the North American Atlantic Coast.

The etiology of sea star wasting syndrome is hypothesized to be caused by a densovirus, sea star-associated densovirus (SSaDV), that has previously been reported on the Pacific and Atlantic Coasts of the United States. In this study, we reevaluated the presence of SSaDV among sea stars from the North American Atlantic Coast and in doing so discovered a novel densovirus that we have named Asterias forbesi-associated densovirus (AfaDV), which shares 78% nucleotide pairwise identity with SSaDV. In contrast to previous studies, SSaDV was not detected in sea stars from the North American Atlantic Coast. Using a variety of PCR-based techniques, we investigated the tissue tropism, host specificity, and prevalence of AfaDV among populations of sea stars at five locations along the Atlantic Coast. AfaDV was detected in three sea star species (Asterias forbesi, Asterias rubens, and Henricia sp.) found in this region and was highly prevalent (>80% of individuals tested; n = 134), among sampled populations. AfaDV was detected in the body wall, gonads, and pyloric caeca (digestive gland) of specimens but was not detected in their coelomic fluid. A significant difference in viral load (copies mg-1) was found between tissue types, with the pyloric caeca having the highest viral loads. Further investigation of Asterias forbesi gonad tissue found germ line cells (oocytes) to be virus positive, suggesting a potential route of vertical transmission. Taken together, these observations show that the presence of AfaDV is not an indicator of sea star wasting syndrome because AfaDV is a common constituent of these animals' microbiome, regardless of health.IMPORTANCE Sea star wasting syndrome is a disease primarily observed on the Pacific and Atlantic Coasts of North America that has significantly impacted sea star populations. The etiology of this disease is unknown, although it is hypothesized to be caused by a densovirus, SSaDV. However, previous studies have not found a correlation between SSaDV and sea star wasting syndrome on the North American Atlantic Coast. This study suggests that this observation may be explained by the presence of a genetically similar densovirus, AfaDV, that may have confounded previous studies. SSaDV was not present in sea stars screened in this study, and instead, AfaDV was commonly found in sea star populations across the New England region, with no apparent signs of disease. These results suggest that sea star densoviruses may be common constituents of the animals' microbiome, and the diversity and extent of these viruses among wild populations may be greater than previously recognized.

An initial comparative genomic autopsy of wasting disease in sea stars.

Beginning in 2013, sea stars throughout the Eastern North Pacific were decimated by wasting disease, also known as "asteroid idiopathic wasting syndrome" (AIWS) due to its elusive aetiology. The geographic extent and taxonomic scale of AIWS meant events leading up to the outbreak were heterogeneous, multifaceted, and oftentimes unobserved; progression from morbidity to death was rapid, leaving few tell-tale symptoms. Here, we take a forensic genomic approach to discover candidate genes that may help explain sea star wasting syndrome. We report the first genome and annotation for Pisaster ochraceus, along with differential gene expression (DGE) analyses in four size classes, three tissue types, and in symptomatic and asymptomatic individuals. We integrate nucleotide polymorphisms associated with survivors of the wasting disease outbreak, DGE associated with temperature treatments in P. ochraceus, and DGE associated with wasting in another asteroid Pycnopodia helianthoides. In P. ochraceus, we found DGE across all tissues, among size classes, and between asymptomatic and symptomatic individuals; the strongest wasting-associated DGE signal was in pyloric caecum. We also found previously identified outlier loci co-occur with differentially expressed genes. In cross-species comparisons of symptomatic and asymptomatic individuals, consistent responses distinguish genes associated with invertebrate innate immunity and chemical defence, consistent with context-dependent stress responses, defensive apoptosis, and tissue degradation. Our analyses thus highlight genomic constituents that may link suspected environmental drivers (elevated temperature) with intrinsic differences among individuals (age/size, alleles associated with susceptibility) that elicit organismal responses (e.g., coelomocyte proliferation) and manifest as sea star wasting mass mortality.