This project sought to understand the outcomes of predator-disease dynamics by exploring a recent pandemic that decimated 90% of ochre sea stars (Pisaster ochraceus) in the eastern North Pacific in 2013. The research team explored how recovery may depend upon often difficult-to-see processes such as the interplay of migration and natural selection in marine species. While the population of sea stars is currently rebounding due to several years of unusually high recruitment, the sea star wasting disease continues to persist at low levels. This project aimed to determine the genetic consequences of the pandemic and subsequent recovery.

Understanding the consequences of large-scale pandemics in the broader contexts of geographic heterogeneity and chronic changes in ocean pH and temperature is an emerging contemporary issue. This project employed long-term characterization of population dynamics and genetic consequences of a sea star wasting disease (SSWD) outbreak, which caused median 90% mortality in Pisaster ochraceus populations in the northeastern Pacific, to estimate potential long-term consequences for the species. While the largest recorded influx of new recruits occurred in 2014-2016, it is unknown where they originated from, whether recruits and surviving adults remained susceptible to the disease, which persisted at low levels, and for how long these dynamics might continue. This long-term dataset provides a unique opportunity for exploring the short and long term repercussions of such large-scale disease outbreaks and the population dynamics that they precipitate. This project built on long-term field studies of wild populations to describe host population dynamics, the disease, and genomic diversity. The goal was to discover genetic variation associated with SSWD and to dissociate that variation from population genomic effects attributable to abiotic environmental variation. Objectives were: (1) Census P. ochraceus at 24 sites throughout its range to describe population dynamics, the prevalence of SSWD, and measure abiotic variables. (2) Conduct laboratory experiments coupled with RNAseq analyses to determine loci differentially regulated during exposure to SSWD, temperature, salinity, and pCO2 anomalies. (3) Map ddRAD, RNAseq, and candidate loci under selection to a P. ochraceus genome. (4) Conduct range-wide population genomic analyses for 3 years to assess genetic (SNP) variation among wild-caught specimens with, versus without, SSWD across a geographic mosaic of abiotic variation. (5) Explore links between SSWD and candidate loci, such as EF1A. These analyses described the immediate genomic consequences of the disease outbreak, the population dynamics that the outbreak set in motion, and the interplay of factors and mechanisms – such as disease, temperature, migration, selection – that affected these changes. The results advanced understanding of general processes and interactions that shape population genomic structure in coastal ecosystems, providing resources to inform future research and applications in design of management strategies for coastal living resources.

Significant Results:

The results of experiments brought about considerable revision to planned experimental design to address hypotheses of SSWD etiology.

In Y1, based on an early common garden incubation which resulted in 100% wasting in the absence of external stimuli, we performed experiments to test the following hypotheses:  Wasting is related to substances that accumulate in response to containment; Wasting is the result of emersion in warmer air temperatures and potential effects of desiccation; Exposure to non-specific compounds from wasting stars would result in wasting; Abrasion or contact between individuals would result in wasting through epidermal damage. We accepted all hypotheses, since for: Faster wasting was observed in aquariums with longer water residence times than those with shorter residence times; Desiccation resulted in faster development of SSWD lesions compared to specimens that were not subject to desiccation under the same aquarium water residence times; Specimens treated with proteinase-k treated asteroid tissue homogenates wasted more slowly than those with untreated tissue homogenates (but note that both treatments wasted significantly faster than completely untreated specimens i.e. controls); and co-housed specimens wasted more quickly than those in individual incubations.

These results led to a further temporal study of wasting in the absence of stimuli, where we tracked microbiome and virome changes in control, artificial scar, and SSWD lesion margin tissues over time in 2 specimens that wasted and 4 that did not waste during the course of the experiment. This observational study showed that copiotrophic taxa (notably the orders Alteromonadales) increased dramatically in all specimens during the experiment, and that upon lesion formation in the SSWD-affected specimens there was an increase in microaerophiles and facultative anaerobes. Furthermore, between the time of lesion formation and animal death, strict anaerobes, including Desulfuromonadales, became abundant. Together with the results of the first hypothesis the results suggested that wasting reflected an increasingly suboxic environment on animal surfaces and led to a further hypothesis: SSWD is a response of asteroids to suboxic conditions that occur as a consequence of bacterial respiration at the animal-water interface. The cause of enhanced bacterial respiraton in aquarium studies may have been accumulation of organic matter from stressed organisms (e.g. mucus production), but results from earlier work in this project and previous challenge experiments also suggested that decaying animal-derived DOM may have stimulated OM pools. Examination of temporal patterns of SSWD in concert with chlorophyll a at a single site exhibited coherence between annual phytoplankton maxima and SSWD. Furthermore, inter-species variation in SSWD susceptibility may have related to the shape and structure of these taxa, since flatter/less rugose species (e.g. Dermasterias imbricata) experienced considerably less SSWD than more rugose species (e.g. Pisaster ochraceus). These observations led to the hypothesis that copiotroph proliferation is a consequence of excess organic matter availability, and may be related to variable physical properties and respiratory activities of affected species.

In Y2 we addressed our second hypothesis in an experiment examining the development of SSWD, bacterial abundance and bacterial assemblage composition on surfaces in response to experimentally depleted O₂. We observed SSWD only in deleted O₂ conditions; no specimen developed SSWD in normoxic conditions. The abundance of bacteria on surfaces in suboxic treatments was significantly lower than in normoxic controls (suboxic conditions reduce aerobic bacterial growth rates, and anaerobes generally experience slower growth rates), and communities shifted randomly between individuals, but no single organizational unit corresponded with treatment. These results confirm that SSWD is related to suboxic conditions. Following this, we addressed our hypothesis in an experiment where Pisaster ochraceus was amended with 3 organic matter sources (peptone, a phytoplankton culture, and particulate matter collected from near the experiment), and its impacts on bacterial+archaeal microbial assembles and bacterial abundance observed by 16S rRNA amplicon sequencing and epifluorescence microscopy. SSWD generated more quickly in specimens that were treated with peptone and with Dunaliella tertiolecta treatment. Bacterial abundance showed a strong increase in the first phase of the experiment followed by a strong decrease over time in affected specimens, which reflected shifts in assemblage composition from copiotroph to microaerophile to anaerobic taxa. We next examined the rugosity of affected and less-affected asteroid species by computed tomography and micro-computed tomography, and compared measured respiration rates (RR), and RR as a ratio of theoretical RR (based on diffusive flux across membranes) and found that SSWD affected specimens were more rugose, and had both higher RR and RR:Theoretical RR. Taken together, these results suggest that SSWD is a stress response to bacterial proliferation on animal respiratory surfaces, which is most clearly tied to the genesis of suboxic conditions through bacterial respiration. The likely cause of enhanced bacterial respiration is excess organic matter availability through phytoplankton or macroalgal exudates or by proximity to decaying animal carcasses.

In Y3, we examined genome variation in SSaDV and other densoviruses through intensive (n = 660 specimens) and geographically extensive (global) survey. Amongst specimens, 22% of asymptomatic and 22% of SSWD-affected individuals were positive for SSaDV by PCR, and sequence analyses of these amplicons revealed >98% identity based on non-structural gene 1. Geographic and re-analyzed data from 2013-2014 revealed 29 densoviral genomes within asteroids, mostly within asymptomatic specimens. Observation of RNA viruses during temporal survey (i.e. Y1) revealed over 48 ssRNA and densoviral genomes were in Santa Cruz specimens in 2018, none of which could be assigned uniquely to SSWD lesion margin tissues. Collectively these results suggest that densoviruses, and indeed most viruses, are widespread in asteroids and not associated with SSWD.

Analysis of tissue samples of Pisaster ochraceus collected in Y2 (i.e. suboxic and organic matter enrichment experiments) for differential loci expression is ongoing and will proceed as a part of NSF-1737091 (i.e. by John Wares and graduate student Paige Duffin). These will then be related to loci identified in post-2013/2014 surviving populations and juveniles identified as part of NSF-1737381.

Work completed on this project:

Santa Cruz Experiments Summer 2018

Bodega Bay Experiments Summer 2019

Sitka and Santa Cruz Imaging Study Collection 2019

Publications on this project to date:

Aquino CA, Besemer RM, DeRito CM, Kocian J, Porter IR, Raimondi P, Rede JE, Schiebelhut LM, Sparks JP, Wares JP, Hewson I (2021) “Evidence that microorganisms at the animal-water interface drive sea star wasting disease” Frontiers in Microbiology. DOI: https://www.frontiersin.org/articles/10.3389/fmicb.2020.610009/

Hewson I, Aquino CA, DeRito CM (2020) “Virome variation during sea star wasting disease progression in Pisaster ochraceus (Asteroidea, Echinodermata)” Viruses 12: 1332

Hewson I, Johnson MR, Tibbetts I (2020) “An unconventional flavivirus and other RNA viruses in the sea cucumber (Holothuroidea; Echinodermata) virome” Viruses 12: 1057

Jackson EW, Wilhelm RC, Johnson MR, Lutz HL, Danforth I, Gaydos JK, Hart MW, Hewson I (2020) “Diversity of sea star-associated densoviruses and transcribed endogenized viral elements of densovirus origin” Journal of Virology DOI: 10.1128/JVI.01594-20

Mordecai GJ, Hewson I (2020) “Coronaviruses in the sea” Frontiers in Microbiology. https://doi.org/10.3389/fmicb.2020.01795

Jackson EW, Pepe-Ranney C, Johnson MR, Distel D, Hewson I (2020) “A highly prevalent and pervasive densovirus discovered among sea stars from the North American Atlantic Coast” Applied and Environmental Microbiology

Hewson I (2019) “Technical pitfalls that bias comparative microbial community analyses of aquatic diseases” Diseases of Aquatic Organisms. 137: 109-124

Hewson I, Sullivan B, Jackson EW, Xu Q, Long H, Lin C, Quijano Cardé EM, Seymour J, Siboni N, Jones MRL, Sewell MA (2019) “Perspective: Something old, something new? Review of wasting and other mortality in Asteroidea (Echinodermata)” Frontiers in Marine Science https://doi.org/10.3389/fmars.2019.00406

Jackson EW, Pepe-Ranney C, Debenport SJ, Buckley DH, Hewson I (2018) “The microbial landscape of sea star and anatomical and interspecies variability of their microbiome” Frontiers in Microbiology. https://doi.org/10.3389/fmicb.2018.0182

Hewson, I, Bistolas, KSI, Quijano Cardé, EM, Button JB, Foster PJ, Flanzenbaum JM, Kocian J, Lewis CK (2018) “Investigating the complex association between viral ecology, environment and North Pacific sea star wasting” Frontiers in Marine Science 5:77