Gambierdiscus species diversity and community structure in St. Thomas, USVI and the Florida Keys, USA.

Ciguatera Poisoning (CP) is a widespread and complex poisoning syndrome caused by the consumption of fish or invertebrates contaminated with a suite of potent neurotoxins collectively known as ciguatoxins (CTXs), which are produced by certain benthic dinoflagellates species in the genera Gambierdiscus and Fukuyoa. Due to the complex nature of this HAB problem, along with a poor understanding of toxin production and entry in the coral reef food web, the development of monitoring, management, and forecasting approaches for CP has lagged behind those available for other HAB syndromes. Over the past two decades, renewed research on the taxonomy, physiology, and toxicology of CP-causing dinoflagellates has advanced our understanding of the species diversity that exists within these genera, including identification of highly toxic species (so called "superbugs") that likely contribute disproportionately to ciguatoxins entering coral reef food webs. The recent development of approaches for molecular analysis of field samples now provide the means to investigate in situ community composition, enabling characterization of spatio-temporal species dynamics, linkages between toxic species abundance and toxin flux, and the risk of ciguatoxin prevalence in fish. In this study we used species-specific fluorescent in situ hybridization (FISH) probes to investigate Gambierdiscus species composition and dynamics in St. Thomas (USVI) and the Florida Keys (USA) over multiple years (2018-2020). Within each location, samples were collected seasonally from several sites comprising varying depths, habitats, and algal substrates to characterize community structure over small spatial scales and across different host macrophytes. This approach enabled the quantitative determination of communities over spatiotemporal gradients, as well as the selective enumeration of species known to exhibit high toxicity, such as Gambierdiscus silvae. The investigation found differing community structure between St. Thomas and Florida Keys sites, driven in part by differences in the distribution of toxin-producing species G. silvae and G. belizeanus, which were present throughout sampling sites in St. Thomas but scarce or absent in the Florida Keys. This finding is significant given the high toxicity of G. silvae, and may help explain differences in fish toxicity and CP incidence between St. Thomas and Florida. Intrasite comparisons along a depth gradient found higher concentrations of Gambierdiscus spp. at deeper locations. Among the macrophytes sampled, Dictyota may be a likely vector for toxin transfer based on their widespread distribution, apparent colonization by G. silvae, and palatability to at least some herbivore grazers. Given its ubiquity throughout both study regions and sites, this taxa may also serve as a refuge, accumulating high concentrations of Gambierdiscus and other benthic dinoflagellates, which in turn can serve as source populations for highly palatable and ephemeral habitats nearby, such as turf algae. These studies further demonstrate the successful application of FISH probes in examining biogeographic structuring of Gambierdiscus communities, targeting individual toxin-producing species, and characterizing species-level dynamics that are needed to describe and model ecological drivers of species abundance and toxicity.

Optimizing marine macrophyte capacity to locally ameliorate ocean acidification under variable light and flow regimes: Insights from an experimental approach.

The urgent need to remediate ocean acidification has brought attention to the ability of marine macrophytes (seagrasses and seaweeds) to take up carbon dioxide (CO2) and locally raise seawater pH via primary production. This physiological process may represent a powerful ocean acidification mitigation tool in coastal areas. However, highly variable nearshore environmental conditions pose uncertainty in the extent of the amelioration effect. We developed experiments in aquaria to address two interconnected goals. First, we explored the individual capacities of four species of marine macrophytes (Ulva lactuca, Zostera marina, Fucus vesiculosus and Saccharina latissima) to ameliorate seawater acidity in experimentally elevated pCO2. Second, we used the most responsive species (i.e., S. latissima) to assess the effects of high and low water residence time on the amelioration of seawater acidity in ambient and simulated future scenarios of climate change across a gradient of irradiance. We measured changes in dissolved oxygen, pH, and total alkalinity, and derived resultant changes to dissolved inorganic carbon (DIC) and calcium carbonate saturation state (Ω). While all species increased productivity under elevated CO2, S. latissima was able to remove DIC and alter pH and Ω more substantially as CO2 increased. Additionally, the amelioration of seawater acidity by S. latissima was optimized under high irradiance and high residence time. However, the influence of water residence time was insignificant under future scenarios. Finally, we applied predictive models as a function of macrophyte biomass, irradiance, and residence time conditions in ambient and future climatic scenarios to allow projections at the ecosystem level. This research contributes to understanding the biological and physical drivers of the coastal CO2 system.

Spatiotemporal transitions in Pseudo-nitzschia species assemblages and domoic acid along the Alaska coast.

The toxic diatom genus Pseudo-nitzschia is distributed from equatorial to polar regions and is comprised of >57 species, some capable of producing the neurotoxin domoic acid (DA). In the Pacific Arctic Region spanning the Bering, Chukchi, and Beaufort seas, DA is recognized as an emerging human and ecosystem health threat, yet little is known about the composition and distribution of Pseudo-nitzschia species in these waters. This investigation characterized Pseudo-nitzschia assemblages in samples collected in 2018 during summer (August) and fall (October-November) surveys as part of the Distributed Biological Observatory and Arctic Observing Network, encompassing a broad geographic range (57.8° to 73.0°N, -138.9° to -169.9°W) and spanning temperature (-1.79 to 11.7°C) and salinity (22.9 to 32.9) gradients associated with distinct water masses. Species were identified using a genus-specific Automated Ribosomal Intergenic Spacer Analysis (ARISA). Seventeen amplicons were observed; seven corresponded to temperate, sub-polar, or polar Pseudo-nitzschia species based on parallel sequencing efforts (P. arctica, P. delicatissima, P. granii, P. obtusa, P. pungens, and two genotypes of P. seriata), and one represented Fragilariopsis oceanica. During summer, particulate DA (pDA; 4.0 to 130.0 ng L-1) was observed in the Bering Strait and Chukchi Sea where P. obtusa was prevalent. In fall, pDA (3.3 to 111.8 ng L-1) occurred along the Beaufort Sea shelf coincident with one P. seriata genotype, and south of the Bering Strait in association with the other P. seriata genotype. Taxa were correlated with latitude, longitude, temperature, salinity, pDA, and/or chlorophyll a, and each had a distinct distribution pattern. The observation of DA in association with different species, seasons, geographic regions, and water masses underscores the significant risk of Amnesic Shellfish Poisoning (ASP) and DA-poisoning in Alaska waters.

Paralytic shellfish toxins in Alaskan Arctic food webs during the anomalously warm ocean conditions of 2019 and estimated toxin doses to Pacific walruses and bowhead whales.

Climate change-related ocean warming and reduction in Arctic sea ice extent, duration and thickness increase the risk of toxic blooms of the dinoflagellate Alexandrium catenella in the Alaskan Arctic. This algal species produces neurotoxins that impact marine wildlife health and cause the human illness known as paralytic shellfish poisoning (PSP). This study reports Paralytic Shellfish Toxin (PST) concentrations quantified in Arctic food web samples that include phytoplankton, zooplankton, benthic clams, benthic worms, and pelagic fish collected throughout summer 2019 during anomalously warm ocean conditions. PSTs (saxitoxin equivalents, STX eq.) were detected in all trophic levels with concentrations above the seafood safety regulatory limit (80 µg STX eq. 100 g-1) in benthic clams collected offshore on the continental shelf in the Beaufort, Chukchi, and Bering Seas. Most notably, toxic benthic clams (Macoma calcarea) were found north of Saint Lawrence Island where Pacific walruses (Odobenus rosmarus) are known to forage for a variety of benthic species, including Macoma. Additionally, fecal samples collected from 13 walruses harvested for subsistence purposes near Saint Lawrence Island during March to May 2019, all contained detectable levels of STX, with fecal samples from two animals (78 and 72 µg STX eq. 100 g-1) near the seafood safety regulatory limit. In contrast, 64% of fecal samples from zooplankton-feeding bowhead whales (n = 9) harvested between March and September 2019 in coastal waters of the Beaufort Sea near Utqiagvik (formerly Barrow) and Kaktovik were toxin-positive, and those levels were significantly lower than in walruses (max bowhead 8.5 µg STX eq. 100 g-1). This was consistent with the lower concentrations of PSTs found in regional zooplankton prey. Maximum ecologically-relevant daily toxin doses to walruses feeding on clams and bowhead whales feeding on zooplankton were estimated to be 21.5 and 0.7 µg STX eq. kg body weight-1 day-1, respectively, suggesting that walruses had higher PST exposures than bowhead whales. Average and maximum STX doses in walruses were in the range reported previously to cause illness and/or death in humans and humpback whales, while bowhead whale doses were well below those levels. These findings raise concerns regarding potential increases in PST/STX exposure risks and health impacts to Arctic marine mammals as ocean warming and sea ice reduction continue.

Evidence for massive and recurrent toxic blooms of Alexandrium catenella in the Alaskan Arctic.

Among the organisms that spread into and flourish in Arctic waters with rising temperatures and sea ice loss are toxic algae, a group of harmful algal bloom species that produce potent biotoxins. Alexandrium catenella, a cyst-forming dinoflagellate that causes paralytic shellfish poisoning worldwide, has been a significant threat to human health in southeastern Alaska for centuries. It is known to be transported into Arctic regions in waters transiting northward through the Bering Strait, yet there is little recognition of this organism as a human health concern north of the Strait. Here, we describe an exceptionally large A. catenella benthic cyst bed and hydrographic conditions across the Chukchi Sea that support germination and development of recurrent, locally originating and self-seeding blooms. Two prominent cyst accumulation zones result from deposition promoted by weak circulation. Cyst concentrations are among the highest reported globally for this species, and the cyst bed is at least 6× larger in area than any other. These extraordinary accumulations are attributed to repeated inputs from advected southern blooms and to localized cyst formation and deposition. Over the past two decades, warming has likely increased the magnitude of the germination flux twofold and advanced the timing of cell inoculation into the euphotic zone by 20 d. Conditions are also now favorable for bloom development in surface waters. The region is poised to support annually recurrent A. catenella blooms that are massive in scale, posing a significant and worrisome threat to public and ecosystem health in Alaskan Arctic communities where economies are subsistence based.

Development of fluorescence in situ hybridization (FISH) probes to detect and enumerate Gambierdiscus species.

Ciguatera poisoning (CP) is a syndrome caused by the bioaccumulation of lipophilic ciguatoxins in coral reef fish and invertebrates, and their subsequent consumption by humans. These phycotoxins are produced by Gambierdiscus spp., tropical epiphytic dinoflagellates that live on a variety of macrophytes, as well as on dead corals and sand. Recent taxonomic studies have identified novel diversity within the Gambierdiscus genus, with at least 18 species and several sub-groups now identified, many of which co-occur and differ significantly in toxicity. The ability to accurately and quickly distinguish Gambierdiscus species in field samples and determine community composition and abundance is central to assessing CP risk, yet most Gambierdiscus species are indistinguishable using light microscopy, and other enumeration methods are semi-quantitative. In order to investigate the spatial and temporal dynamics of Gambierdiscus species and community toxicity, new tools for species identification and enumeration in field samples are needed. Here, fluorescence in situ hybridization (FISH) probes were designed for seven species commonly found in the Caribbean Sea and Pacific Ocean, permitting their enumeration in field samples using epifluorescence microscopy. This technique enables the assessment of community composition and accurate determination of cell abundances of individual species. Molecular probes detecting G. australes, G. belizeanus, G. caribaeus, G. carolinianus, G. carpenteri, and the G. silvae/G. polynesiensis clade were designed using alignments of large subunit ribosomal RNA (rRNA) sequences. These probes were tested for specificity and cross-reactivity through experiments in which field samples were spiked with known concentrations of Gambierdiscus cultures, and analyzed to confirm that Gambierdiscus can be successfully detected and enumerated by FISH in the presence of detritus and other organisms. These probes were then used to characterize Gambierdiscus community structure in field samples collected from the Florida Keys and Hawai'i, USA. The probes revealed the co-occurrence of multiple species at each location. Time-series FISH analyses of samples collected from the Florida Keys quantified seasonal shifts in community composition as well as fluctuations in overall Gambierdiscus cell abundance. Application of species-specific FISH probes provides a powerful new tool to those seeking to target individual Gambierdiscus species, including significant toxin-producers, in field populations. Moving forward, analysis of Gambierdiscus community composition across multiple environments and over time will also allow species dynamics to be linked to environmental parameters, improving our ability to understand and manage the current and changing risks of CP worldwide.