Manipulation of the seagrass-associated microbiome reduces disease severity.

Host-associated microbes influence host health and function and can be a first line of defence against infections. While research increasingly shows that terrestrial plant microbiomes contribute to bacterial, fungal, and oomycete disease resistance, no comparable experimental work has investigated marine plant microbiomes or more diverse disease agents. We test the hypothesis that the eelgrass (Zostera marina) leaf microbiome increases resistance to seagrass wasting disease. From field eelgrass with paired diseased and asymptomatic tissue, 16S rRNA gene amplicon sequencing revealed that bacterial composition and richness varied markedly between diseased and asymptomatic tissue in one of the two years. This suggests that the influence of disease on eelgrass microbial communities may vary with environmental conditions. We next experimentally reduced the eelgrass microbiome with antibiotics and bleach, then inoculated plants with Labyrinthula zosterae, the causative agent of wasting disease. We detected significantly higher disease severity in eelgrass with a native microbiome than an experimentally reduced microbiome. Our results over multiple experiments do not support a protective role of the eelgrass microbiome against L. zosterae. Further studies of these marine host-microbe-pathogen relationships may continue to show new relationships between plant microbiomes and diseases.

Predictable Changes in Eelgrass Microbiomes with Increasing Wasting Disease Prevalence across 23° Latitude in the Northeastern Pacific.

Predicting outcomes of marine disease outbreaks presents a challenge in the face of both global and local stressors. Host-associated microbiomes may play important roles in disease dynamics but remain understudied in marine ecosystems. Host-pathogen-microbiome interactions can vary across host ranges, gradients of disease, and temperature; studying these relationships may aid our ability to forecast disease dynamics. Eelgrass, Zostera marina, is impacted by outbreaks of wasting disease caused by the opportunistic pathogen Labyrinthula zosterae. We investigated how Z. marina phyllosphere microbial communities vary with rising wasting disease lesion prevalence and severity relative to plant and meadow characteristics like shoot density, longest leaf length, and temperature across 23° latitude in the Northeastern Pacific. We detected effects of geography (11%) and smaller, but distinct, effects of temperature (30-day max sea surface temperature, 4%) and disease (lesion prevalence, 3%) on microbiome composition. Declines in alpha diversity on asymptomatic tissue occurred with rising wasting disease prevalence within meadows. However, no change in microbiome variability (dispersion) was detected between asymptomatic and symptomatic tissues. Further, we identified members of Cellvibrionaceae, Colwelliaceae, and Granulosicoccaceae on asymptomatic tissue that are predictive of wasting disease prevalence across the geographic range (3,100 kilometers). Functional roles of Colwelliaceae and Granulosicoccaceae are not known. Cellvibrionaceae, degraders of plant cellulose, were also enriched in lesions and adjacent green tissue relative to nonlesioned leaves. Cellvibrionaceae may play important roles in disease progression by degrading host tissues or overwhelming plant immune responses. Thus, inclusion of microbiomes in wasting disease studies may improve our ability to understand variable rates of infection, disease progression, and plant survival. IMPORTANCE The roles of marine microbiomes in disease remain poorly understood due, in part, to the challenging nature of sampling at appropriate spatiotemporal scales and across natural gradients of disease throughout host ranges. This is especially true for marine vascular plants like eelgrass (Zostera marina) that are vital for ecosystem function and biodiversity but are susceptible to rapid decline and die-off from pathogens like eukaryotic slime-mold Labyrinthula zosterae (wasting disease). We link bacterial members of phyllosphere tissues to the prevalence of wasting disease across the broadest geographic range to date for a marine plant microbiome-disease study (3,100 km). We identify Cellvibrionaceae, plant cell wall degraders, enriched (up to 61% relative abundance) within lesion tissue, which suggests this group may be playing important roles in disease progression. These findings suggest inclusion of microbiomes in marine disease studies will improve our ability to predict ecological outcomes of infection across variable landscapes spanning thousands of kilometers.

Coastal urbanization influences human pathogens and microdebris contamination in seafood.

Seafood is one of the leading imported products implicated in foodborne outbreaks worldwide. Coastal marine environments are being increasingly subjected to reduced water quality from urbanization and leading to contamination of important fishery species. Given the importance of seafood exchanged as a global protein source, it is imperative to maintain seafood safety worldwide. To illustrate the potential health risks associated with urbanization in a coastal environment, we use next-generation high-throughput amplicon sequencing of the 16S ribosomal RNA gene combined with infrared spectroscopy to characterize and quantify a vast range of potential human bacterial pathogens and microdebris contaminants in seawater, sediment and an important oyster fishery along the Mergui Archipelago in Myanmar. Through the quantification of >1.25 million high-quality bacterial operational taxonomic unit (OTU) reads, we detected 5459 potential human bacterial pathogens belonging to 87 species that are commonly associated with gut microbiota and an indication of terrestrial runoff of human and agricultural waste. Oyster tissues contained 51% of all sequenced bacterial pathogens that are considered to be both detrimental and of emerging concern to human health. Using infrared spectroscopy, we examined a total of 1225 individual microdebris particles, from which we detected 78 different types of contaminant materials. The predominant microdebris contaminants recovered from oyster tissues included polymers (48%), followed by non-native minerals (20%), oils (14%) and milk supplement powders (14%). Emerging technologies provide novel insights into the impacts of coastal development on food security and risks to human and environmental health.

Warming and pollutants interact to modulate octocoral immunity and shape disease outcomes.

Warming environments can alter the outcome of host-parasite relationships with important consequences for biodiversity. Warming often increases disease risk, and interactions with other environmental factors can intensify impacts by modifying the underlying mechanisms, such as host immunity. In coastal ecosystems, metal pollution is a pervasive stressor that influences disease and immunity in many organisms. Despite the crisis facing coral reefs, which stems in part from warming-associated disease outbreaks, the impacts of metal pollutants on scleractinian and octocoral disease are largely unknown. We investigated how warming oceans and copper pollution affect host immunity and disease risk for two diseases of the abundant Caribbean octocoral, the sea fan Gorgonia ventalina. Field surveys across a sediment copper concentration gradient in Puerto Rico, USA revealed that cellular immunity of sea fans increased by 12.6% at higher sediment copper concentrations, while recovery from multifocal purple spots disease (MFPS) tended to decrease. MFPS severity in the field increased at warmer sites. In a controlled laboratory experiment, sea fans were inoculated with live cultures of a labyrinthulid parasite to test the interactive effects of temperature and copper on immune activation. As in the field, higher copper induced greater immunity, but the factorial design of the experiment revealed that copper and temperature interacted to modulate the immune response to the parasite: immune cell densities increased with elevated temperature at lower copper concentrations, but not with high copper concentrations. Tissue damage was also greater in treatments with higher copper and warmer temperatures. Field and lab evidence confirm that elevated copper hinders sea fan immune defenses against damaging parasites. Temperature and copper influenced host-pathogen interactions in octocorals by modulating immunity, disease severity, and disease recovery. This is the first evidence that metal pollution affects processes influencing disease in octocorals and highlights the importance of immune mechanisms in environmentally mediated disease outbreaks. Although coral conservation efforts must include a focus on global factors, such as rapid warming, reducing copper and other pollutants that compromise coral health on a local scale may help corals fight disease in a warming ocean.

Increases and decreases in marine disease reports in an era of global change.

Outbreaks of marine infectious diseases have caused widespread mass mortalities, but the lack of baseline data has precluded evaluating whether disease is increasing or decreasing in the ocean. We use an established literature proxy method from Ward and Lafferty (Ward and Lafferty 2004 PLoS Biology2, e120 (doi:10.1371/journal.pbio.0020120)) to analyse a 44-year global record of normalized disease reports from 1970 to 2013. Major marine hosts are combined into nine taxonomic groups, from seagrasses to marine mammals, to assess disease swings, defined as positive or negative multi-decadal shifts in disease reports across related hosts. Normalized disease reports increased significantly between 1970 and 2013 in corals and urchins, indicating positive disease swings in these environmentally sensitive ectotherms. Coral disease reports in the Caribbean correlated with increasing temperature anomalies, supporting the hypothesis that warming oceans drive infectious coral diseases. Meanwhile, disease risk may also decrease in a changing ocean. Disease reports decreased significantly in fishes and elasmobranchs, which have experienced steep human-induced population declines and diminishing population density that, while concerning, may reduce disease. The increases and decreases in disease reports across the 44-year record transcend short-term fluctuations and regional variation. Our results show that long-term changes in disease reports coincide with recent decades of widespread environmental change in the ocean.

Host size and proximity to diseased neighbours drive the spread of a coral disease outbreak in Hawai'i.

Understanding how disease risk varies over time and across heterogeneous populations is critical for managing disease outbreaks, but this information is rarely known for wildlife diseases. Here, we demonstrate that variation in host and pathogen factors drive the direction, duration and intensity of a coral disease outbreak. We collected longitudinal health data for 200 coral colonies, and found that disease risk increased with host size and severity of diseased neighbours, and disease spread was highest among individuals between 5 and 20 m apart. Disease risk increased by 2% with every 10 cm increase in host size. Healthy colonies with severely diseased neighbours (greater than 75% affected tissue) were 1.6 times more likely to develop disease signs compared with colonies with moderately diseased neighbours (25-75% affected tissue). Force of infection ranged from 7 to 20 disease cases per 1000 colonies (mean = 15 cases per 1000 colonies). The effective reproductive ratio, or average number of secondary infections per infectious individual, ranged from 0.16 to 1.22. Probability of transmission depended strongly on proximity to diseased neighbours, which demonstrates that marine disease spread can be highly constrained within patch reefs.

Plastic waste associated with disease on coral reefs.

Plastic waste can promote microbial colonization by pathogens implicated in outbreaks of disease in the ocean. We assessed the influence of plastic waste on disease risk in 124,000 reef-building corals from 159 reefs in the Asia-Pacific region. The likelihood of disease increases from 4% to 89% when corals are in contact with plastic. Structurally complex corals are eight times more likely to be affected by plastic, suggesting that microhabitats for reef-associated organisms and valuable fisheries will be disproportionately affected. Plastic levels on coral reefs correspond to estimates of terrestrial mismanaged plastic waste entering the ocean. We estimate that 11.1 billion plastic items are entangled on coral reefs across the Asia-Pacific and project this number to increase 40% by 2025. Plastic waste management is critical for reducing diseases that threaten ecosystem health and human livelihoods.

Octocoral co-infection as a balance between host immunity and host environment.

Co-infection is the reality in natural populations, but few studies incorporate the players that matter in the wild. We integrate the environment, host demography, two parasites, and host immunity in a study of co-infection to determine the drivers of parasite interactions. Here, we use an ecologically important Caribbean sea fan octocoral, Gorgonia ventalina, that is co-infected by a copepod and a labyrinthulid protist. We first expanded upon laboratory studies by showing that immune suppression is associated with the labyrinthulid in a natural setting. Histological analyses revealed that immune cells (amoebocytes) were significantly suppressed in both labyrinthulid infections and co-infections relative to healthy sea fans, but remained unchanged in copepod infections. However, surveys of natural coral populations demonstrated a critical role for the environment and host demography in this co-infection: the prevalence of copepod infections increased with sea fan size while labyrinthulid prevalence increased with water depth. Although we predicted that immune suppression by the labyrinthulid would facilitate copepod infection, the two parasites did not co-occur in the sea fans more often than expected by chance. These results suggest that the distinct ecological drivers for each parasite overwhelm the role of host immune suppression in determining the distribution of parasites among hosts. This interplay of the environment and parasite-mediated immune suppression in sea fan co-infection provides insights into the factors underlying co-occurrence patterns in wild co-infections. Moving forward, simultaneous consideration of co-occurring parasites, host traits, and the environmental context will improve the understanding of host - parasite interactions and their consequences.

Seagrass ecosystems reduce exposure to bacterial pathogens of humans, fishes, and invertebrates.

Plants are important in urban environments for removing pathogens and improving water quality. Seagrass meadows are the most widespread coastal ecosystem on the planet. Although these plants are known to be associated with natural biocide production, they have not been evaluated for their ability to remove microbiological contamination. Using amplicon sequencing of the 16S ribosomal RNA gene, we found that when seagrass meadows are present, there was a 50% reduction in the relative abundance of potential bacterial pathogens capable of causing disease in humans and marine organisms. Moreover, field surveys of more than 8000 reef-building corals located adjacent to seagrass meadows showed twofold reductions in disease levels compared to corals at paired sites without adjacent seagrass meadows. These results highlight the importance of seagrass ecosystems to the health of humans and other organisms.

Devastating Transboundary Impacts of Sea Star Wasting Disease on Subtidal Asteroids.

Sea star wasting disease devastated intertidal sea star populations from Mexico to Alaska between 2013-15, but little detail is known about its impacts to subtidal species. We assessed the impacts of sea star wasting disease in the Salish Sea, a Canadian / United States transboundary marine ecosystem, and world-wide hotspot for temperate asteroid species diversity with a high degree of endemism. We analyzed roving diver survey data for the three most common subtidal sea star species collected by trained volunteer scuba divers between 2006-15 in 5 basins and on the outer coast of Washington, as well as scientific strip transect data for 11 common subtidal asteroid taxa collected by scientific divers in the San Juan Islands during the spring/summer of 2014 and 2015. Our findings highlight differential susceptibility and impact of sea star wasting disease among asteroid species populations and lack of differences between basins or on Washington's outer coast. Specifically, severe depletion of sunflower sea stars (Pycnopodia helianthoides) in the Salish Sea support reports of major declines in this species from California to Alaska, raising concern for the conservation of this ecologically important subtidal predator.

Plant characteristics associated with widespread variation in eelgrass wasting disease.

Seagrasses are ecosystem engineers of essential marine habitat. Their populations are rapidly declining worldwide. One potential cause of seagrass population declines is wasting disease, which is caused by opportunistic pathogens in the genus Labyrinthula. While infection with these pathogens is common in seagrasses, theory suggests that disease only occurs when environmental stressors cause immunosuppression of the host. Recent evidence suggests that host factors may also contribute to disease caused by opportunistic pathogens. In order to quantify patterns of disease, identify risk factors, and investigate responses to infection, we surveyed shoot density, shoot length, epiphyte load, production of plant defenses (phenols), and wasting disease prevalence in eelgrass Zostera marina across 11 sites in the central Salish Sea (Washington state, USA), a region where both wasting disease and eelgrass declines have been documented. Wasting disease was diagnosed by the presence of necrotic lesions, and Labyrinthula cells were identified with histology. Disease prevalence among sites varied from 6 to 79%. The probability of a shoot being diseased was higher in longer shoots, in patches of higher shoot density, and in shoots with higher levels of biofouling from epiphytes. Phenolic concentration was higher in diseased leaves. We hypothesize that this results from the induction of phenols during infection. Additional research is needed to evaluate whether phenols are an adaptive defense against Labyrinthula infection. The high site-level variation in disease prevalence emphasizes the potential for wasting disease to be causing some of the observed decline in eelgrass beds.

Managing marine disease emergencies in an era of rapid change.

Infectious marine diseases can decimate populations and are increasing among some taxa due to global change and our increasing reliance on marine environments. Marine diseases become emergencies when significant ecological, economic or social impacts occur. We can prepare for and manage these emergencies through improved surveillance, and the development and iterative refinement of approaches to mitigate disease and its impacts. Improving surveillance requires fast, accurate diagnoses, forecasting disease risk and real-time monitoring of disease-promoting environmental conditions. Diversifying impact mitigation involves increasing host resilience to disease, reducing pathogen abundance and managing environmental factors that facilitate disease. Disease surveillance and mitigation can be adaptive if informed by research advances and catalysed by communication among observers, researchers and decision-makers using information-sharing platforms. Recent increases in the awareness of the threats posed by marine diseases may lead to policy frameworks that facilitate the responses and management that marine disease emergencies require.

Improving marine disease surveillance through sea temperature monitoring, outlooks and projections.

To forecast marine disease outbreaks as oceans warm requires new environmental surveillance tools. We describe an iterative process for developing these tools that combines research, development and deployment for suitable systems. The first step is to identify candidate host-pathogen systems. The 24 candidate systems we identified include sponges, corals, oysters, crustaceans, sea stars, fishes and sea grasses (among others). To illustrate the other steps, we present a case study of epizootic shell disease (ESD) in the American lobster. Increasing prevalence of ESD is a contributing factor to lobster fishery collapse in southern New England (SNE), raising concerns that disease prevalence will increase in the northern Gulf of Maine under climate change. The lowest maximum bottom temperature associated with ESD prevalence in SNE is 12 °C. Our seasonal outlook for 2015 and long-term projections show bottom temperatures greater than or equal to 12 °C may occur in this and coming years in the coastal bays of Maine. The tools presented will allow managers to target efforts to monitor the effects of ESD on fishery sustainability and will be iteratively refined. The approach and case example highlight that temperature-based surveillance tools can inform research, monitoring and management of emerging and continuing marine disease threats.

Ochre star mortality during the 2014 wasting disease epizootic: role of population size structure and temperature.

Over 20 species of asteroids were devastated by a sea star wasting disease (SSWD) epizootic, linked to a densovirus, from Mexico to Alaska in 2013 and 2014. For Pisaster ochraceus from the San Juan Islands, South Puget Sound and Washington outer coast, time-series monitoring showed rapid disease spread, high mortality rates in 2014, and continuing levels of wasting in the survivors in 2015. Peak prevalence of disease at 16 sites ranged to 100%, with an overall mean of 61%. Analysis of longitudinal data showed disease risk was correlated with both size and temperature and resulted in shifts in population size structure; adult populations fell to one quarter of pre-outbreak abundances. In laboratory experiments, time between development of disease signs and death was influenced by temperature in adults but not juveniles and adult mortality was 18% higher in the 19 °C treatment compared to the lower temperature treatments. While larger ochre stars developed disease signs sooner than juveniles, diseased juveniles died more quickly than diseased adults. Unusual 2-3 °C warm temperature anomalies were coincident with the summer 2014 mortalities. We suggest these warm waters could have increased the disease progression and mortality rates of SSWD in Washington State.

Linking sewage pollution and water quality to spatial patterns of Porites lobata growth anomalies in Puako, Hawaii.

Sewage pollution threatens the health of coastal populations and ecosystems, including coral reefs. We investigated spatial patterns of sewage pollution in Puako, Hawaii using enterococci concentrations and δ(15)N Ulva fasciata macroalgal bioassays to assess relationships with the coral disease Porites lobata growth anomalies (PGAs). PGA severity and enterococci concentrations were high, spatially variable, and positively related. Bioassay algal δ(15)N showed low sewage pollution at the reef edge while high values of resident algae indicated sewage pollution nearshore. Neither δ(15)N metric predicted PGA measures, though bioassay δ(15)N was negatively related to coral cover. Furthermore, PGA prevalence was much higher than previously recorded in Hawaii and the greater Indo-Pacific, highlighting Puako as an area of concern. Although further work is needed to resolve the relationship between sewage pollution and coral cover and disease, these results implicate sewage pollution as a contributor to diminished reef health.

Up in Arms: Immune and Nervous System Response to Sea Star Wasting Disease.

Echinoderms, positioned taxonomically at the base of deuterostomes, provide an important system for the study of the evolution of the immune system. However, there is little known about the cellular components and genes associated with echinoderm immunity. The 2013-2014 sea star wasting disease outbreak is an emergent, rapidly spreading disease, which has led to large population declines of asteroids in the North American Pacific. While evidence suggests that the signs of this disease, twisting arms and lesions, may be attributed to a viral infection, the host response to infection is still poorly understood. In order to examine transcriptional responses of the sea star Pycnopodia helianthoides to sea star wasting disease, we injected a viral sized fraction (0.2 µm) homogenate prepared from symptomatic P. helianthoides into apparently healthy stars. Nine days following injection, when all stars were displaying signs of the disease, specimens were sacrificed and coelomocytes were extracted for RNA-seq analyses. A number of immune genes, including those involved in Toll signaling pathways, complement cascade, melanization response, and arachidonic acid metabolism, were differentially expressed. Furthermore, genes involved in nervous system processes and tissue remodeling were also differentially expressed, pointing to transcriptional changes underlying the signs of sea star wasting disease. The genomic resources presented here not only increase understanding of host response to sea star wasting disease, but also provide greater insight into the mechanisms underlying immune function in echinoderms.

Whole transcriptome analysis reveals changes in expression of immune-related genes during and after bleaching in a reef-building coral.

Climate change is negatively affecting the stability of natural ecosystems, especially coral reefs. The dissociation of the symbiosis between reef-building corals and their algal symbiont, or coral bleaching, has been linked to increased sea surface temperatures. Coral bleaching has significant impacts on corals, including an increase in disease outbreaks that can permanently change the entire reef ecosystem. Yet, little is known about the impacts of coral bleaching on the coral immune system. In this study, whole transcriptome analysis of the coral holobiont and each of the associate components (i.e. coral host, algal symbiont and other associated microorganisms) was used to determine changes in gene expression in corals affected by a natural bleaching event as well as during the recovery phase. The main findings include evidence that the coral holobiont and the coral host have different responses to bleaching, and the host immune system appears suppressed even a year after a bleaching event. These results support the hypothesis that coral bleaching changes the expression of innate immune genes of corals, and these effects can last even after recovery of symbiont populations. Research on the role of immunity on coral's resistance to stressors can help make informed predictions on the future of corals and coral reefs.

Persistent shifts in Caribbean coral microbiota are linked to the 2010 warm thermal anomaly.

The response of corals to warm temperature anomalies includes changes in coral bacterial assemblages. There are clear differences between the microbiota of bleached and healthy corals. However, few studies have tracked the microbiota of individual colonies throughout a warming event. We used 454 pyrosequencing and repeated measures to characterize bacterial assemblages in 15 Gorgonia ventalina colonies before, during, 4 months after, and 1 year after the 2010 Caribbean warm thermal anomaly. In the latter three sampling times, the G. ventalina microbiota differed significantly from the microbiota of Orbicella faveolata colonies, which were sampled only at these three times. O. faveolata microbiota did not exhibit coordinated shifts through time. Notably, the microbiota of the repeatedly sampled G. ventalina colonies shifted persistently from before to during, after, and long after the warming event. The same pattern emerges from the norm of reaction for the individual G. ventalina colonies. This is the first study to show persistent shifts in coral microbiota in association with a warm thermal anomaly. Whether shifting microbiota is adaptive or maladaptive, the lasting change in bacterial assemblages following this warming event identifies a new way that coral microbiota shape the response of coral colonies under thermal stress.

Infectious diseases affect marine fisheries and aquaculture economics.

Seafood is a growing part of the economy, but its economic value is diminished by marine diseases. Infectious diseases are common in the ocean, and here we tabulate 67 examples that can reduce commercial species' growth and survivorship or decrease seafood quality. These impacts seem most problematic in the stressful and crowded conditions of aquaculture, which increasingly dominates seafood production as wild fishery production plateaus. For instance, marine diseases of farmed oysters, shrimp, abalone, and various fishes, particularly Atlantic salmon, cost billions of dollars each year. In comparison, it is often difficult to accurately estimate disease impacts on wild populations, especially those of pelagic and subtidal species. Farmed species often receive infectious diseases from wild species and can, in turn, export infectious agents to wild species. However, the impact of disease export on wild fisheries is controversial because there are few quantitative data demonstrating that wild species near farms suffer more from infectious diseases than those in other areas. The movement of exotic infectious agents to new areas continues to be the greatest concern.

Densovirus associated with sea-star wasting disease and mass mortality.

Populations of at least 20 asteroid species on the Northeast Pacific Coast have recently experienced an extensive outbreak of sea-star (asteroid) wasting disease (SSWD). The disease leads to behavioral changes, lesions, loss of turgor, limb autotomy, and death characterized by rapid degradation ("melting"). Here, we present evidence from experimental challenge studies and field observations that link the mass mortalities to a densovirus (Parvoviridae). Virus-sized material (i.e., <0.2 µm) from symptomatic tissues that was inoculated into asymptomatic asteroids consistently resulted in SSWD signs whereas animals receiving heat-killed (i.e., control) virus-sized inoculum remained asymptomatic. Viral metagenomic investigations revealed the sea star-associated densovirus (SSaDV) as the most likely candidate virus associated with tissues from symptomatic asteroids. Quantification of SSaDV during transmission trials indicated that progression of SSWD paralleled increased SSaDV load. In field surveys, SSaDV loads were more abundant in symptomatic than in asymptomatic asteroids. SSaDV could be detected in plankton, sediments and in nonasteroid echinoderms, providing a possible mechanism for viral spread. SSaDV was detected in museum specimens of asteroids from 1942, suggesting that it has been present on the North American Pacific Coast for at least 72 y. SSaDV is therefore the most promising candidate disease agent responsible for asteroid mass mortality.