INVESTIGATING BLOOD LACTATE CONCENTRATION AS A PROGNOSTIC INDICATOR FOR BIRDS PRESENTING WITH BREVETOXICOSIS: 2020-2021.

Large blooms of the dinoflagellate Karenia brevis cause annual harmful algal bloom events, or "red tides" on Florida's Gulf Coast. Each year, the Clinic for the Rehabilitation of Wildlife (CROW) is presented with hundreds of cases of aquatic birds that exhibit neurologic clinical signs due to brevetoxicosis. Double-crested cormorants (Phalacrocorax auratus) are the most common species seen, and typically present with a combination of ataxia, head tremors, knuckling, and/or lagophthalmos. Blood lactate levels are known to increase in mammals for a variety of reasons, including stress, hypoxia, sepsis, and trauma, but there is limited literature on blood lactate values in avian species. The objective of this study was to determine the prognostic value of blood lactate concentration on successful rehabilitation and release of birds presenting with clinical signs consistent with brevetoxicosis. Blood lactate levels were collected on intake, the morning after presentation and initial therapy, and prior to disposition (release or euthanasia) from 194 birds (including 98 cormorants) representing 17 species during the 2020-2021 red tide season. Overall, mean blood lactate at intake, the morning after intake, and predisposition was 2.9, 2.8, and 3.2 mmol/L, respectively, for released birds across all species (2.9, 2.9, and 3.2 mmol/L for released cormorants); 3.4, 3.4, and 6.5 mmol/L for birds that died (4.0, 3.5, and 7.9 mmol/L for cormorants that died); and 3.1, 3.5, and 4.7 mmol/L for birds that were euthanized (3.5, 4.7, and 4.9 mmol/L for cormorants that were euthanized). On average, birds that died or were euthanized had an elevated lactate at all time points as compared to those that were released, but these results were not statistically significant (P = 0.13). These results indicate that blood lactate levels do not appear to be useful as a prognostic indicator for successful release of birds, including double-crested cormorants, affected by brevetoxicosis.

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.

Mortality in Common (Sterna hirundo) and Sandwich (Thalasseus sandvicensis) Terns Associated with Bisgaard Taxon 40 Infection on Marco Island, Florida, USA.

Widely distributed aquatic species such as terns are highly dependent on, and can serve as indicators of, the global health of marine and other aquatic environments. Documented mass mortality events in terns have been associated with anthropogenic, weather-related and, less commonly, infectious causes. This study describes a multispecies mortality event associated with brevetoxicosis and Bisgaard taxon 40-induced sepsis involving common (Sterna hirundo) and sandwich (Thalasseus sandvicensis) terns off the southwest coast of Florida, USA, in November and December 2018. During an approximately 6-8-week period, a large number of birds were found dead or displayed weakness, ataxia or other neurological signs. Many were admitted to a wildlife hospital for evaluation, but most died or were euthanized due to poor prognosis. Necropsy of 12 birds revealed minimal or non-specific gross lesions. Initial toxicology screening of tissues for brevetoxins revealed levels that could be consistent with brevetoxicosis. However, histology revealed multiorgan inflammation and necrosis associated with a gram-negative bacillus. A bacterium isolated on aerobic culture of liver and heart tissues was unidentifiable in the MALDI-TOF database. Subsequently, 16 S rRNA gene sequencing revealed that the isolate shared 99.33% homology with Bisgaard taxon 40 from the Pasteurellaceae family. While the source of the bacterium and potential association with brevetoxin exposure are unclear, histopathology suggests that the bacterium was the proximate cause of clinical signs and mortality in all birds examined as well as the scale of the mortality event. This report highlights the need to conduct detailed investigations into wildlife mortality events and expands on the current, limited knowledge of the effects of novel Pasteurellaceae bacteria on avian health.

High-resolution Spatiotemporal Dynamics of Harmful Algae in the Indian River Lagoon (Florida)-A Case Study of Aureoumbra lagunensis, Pyrodinium bahamense, and Pseudo-nitzschia.

The Indian River Lagoon (IRL), located on the east coast of Florida, is a complex estuarine ecosystem that is negatively affected by recurring harmful algal blooms (HABs) from distinct taxonomic/functional groups. Enhanced monitoring was established to facilitate rapid quantification of three recurrent bloom taxa, Aureoumbra lagunensis, Pyrodinium bahamense, and Pseudo-nitzschia spp., and included corroborating techniques to improve the identification of small-celled nanoplankton (<10 µm in diameter). Identification and enumeration of these target taxa were conducted during 2015-2020 using a combination of light microscopy and species-specific approaches, specifically immunofluorescence flow cytometry as well as a newly developed qPCR assay for A. lagunensis presented here for the first time. An annual bloom index (ABI) was established for each taxon based on occurrence and abundance data. Blooms of A. lagunensis (>2×108 cells L-1) were observed in all six years sampled and across multiple seasons. In contrast, abundance of P. bahamense, largely driven by the annual temperature cycle that moderates life cycle transitions and growth, displayed a strong seasonal pattern with blooms (105-107 cells L-1) generally developing in early summer and subsiding in autumn. However, P. bahamense bloom development was delayed and abundance was significantly lower in years and locations with sustained A. lagunensis blooms. Pseudo-nitzschia spp. were broadly distributed with sporadic bloom concentrations (reaching 107 cells L-1), but with minimal concentrations of the toxin domoic acid detected (<0.02 µg L-1). In summer 2020, multiple monitoring tools characterized a novel nano-cyanobacterium bloom (reaching 109 cells L-1) that coincided with a decline in A. lagunensis and persisted into autumn. Statistical and time-series analyses of this spatiotemporally intensive dataset highlight prominent patterns in variability for some taxa, but also identifies challenges of characterizing mechanisms underlying more episodic yet persistent events. Nevertheless, the intersect of temperature and salinity as environmental proxies proved to be informative in delineating niche partitioning, not only in the case of taxa with long-standing data sets but also for seemingly unprecedented blooms of novel nanoplanktonic taxa.

An occurrence of neurotoxic shellfish poisoning by consumption of gastropods contaminated with brevetoxins.

Brevetoxins were confirmed in urine specimens from patients diagnosed with neurotoxic shellfish poisoning (NSP) after consumption of gastropods that were recreationally harvested from an area previously affected by a Karenia brevis bloom. Several species of gastropods (Triplofusus giganteus, Sinistrofulgur sinistrum, Cinctura hunteria, Strombus alatus, Fulguropsis spirata) and one clam (Macrocallista nimbosa) from the NSP implicated gastropod collection area (Jewfish Key, Sarasota Bay, Florida) were examined for brevetoxins using liquid chromatography-tandem mass spectrometry (LC-MS/MS) and enzyme-linked immunosorbent assay (ELISA). All gastropods and the clam were contaminated with brevetoxins. Composite B-type toxin concentrations in gastropods ranged from 1.1 to 198 µg BTX-3 equiv./g by ELISA, levels likely capable of causing NSP in consumers. Several brevetoxin metabolites previously characterized in molluscan shellfish were identified in these gastropods. Brevetoxin analog profiles by ELISA were similar in the gastropod species examined. This work documents the occurrence of NSP through consumption of a type of seafood not typically monitored in Florida to protect human health, demonstrating the need to better assess and communicate the risk of NSP to gastropod harvesters in Karenia brevis endemic areas.

Immune function in Trachemys scripta following exposure to a predominant brevetoxin congener, PbTx-3, as a model for potential health impacts for sea turtles naturally exposed to brevetoxins.

Many species of marine life in southwestern Florida, including sea turtles, are impacted by blooms of the toxic dinoflagellate, Karenia brevis. Sublethal exposure to toxins produced by K. brevis has been shown to impact sea turtle health. Since all sea turtles in the Gulf of Mexico have protected status, a freshwater turtle, Trachemys scripta, was used as a model for immune system effects following experimental exposure to a predominant brevetoxin congener in K. brevis blooms, PbTx-3. Exposure to PbTx-3 was oral or intratracheal and health effects were assessed using a suite of immune function parameters: innate immune function (phagocytosis, plasma lysozyme activity), adaptive immune function (lymphocyte proliferation), and measures of oxidative stress (superoxide dismutase (SOD) and glutathione-S-transferase (GST) activity in plasma). Inflammation was also measured using plasma protein electrophoresis. In addition, differential expression of genes in peripheral blood leukocytes was determined using suppression subtractive hybridization followed by real-time PCR of specific genes. The primary immune effects of sublethal brevetoxin exposure in T. scripta following PbTx-3 administration, appear to be an increase in oxidative stress, a decrease in lysozyme activity, and modulation of immune function through lymphocyte proliferation responses. Plasma protein electrophoresis showed a decreased A:G ratio which may indicate potential inflammation. Genes coding for oxidative stress, such as thioredoxin and GST, were upregulated in exposed animals. That sublethal brevetoxin exposures impact immune function components suggests potential health implications for sea turtles naturally exposed to toxins. Knowledge of physiological stressors induced by brevetoxins may contribute to the ultimate goal of developing directed treatment strategies in exposed animals for reduced mortality resulting from red tide toxin exposure in sea turtles.

INTRAVENOUS LIPID EMULSION TREATMENT REDUCES SYMPTOMS OF BREVETOXICOSIS IN TURTLES (TRACHEMYS SCRIPTA).

Harmful algal blooms (HABs) occur when excess nutrients allow dinoflagellates to reproduce in large numbers. Marine animals are affected by blooms when algal toxins are ingested or inhaled. In the Gulf of Mexico, near annual blooms of Karenia brevis release a suite of compounds (brevetoxins) that cause sea turtle morbidity and mortality. The primary treatment at rehabilitation facilities for brevetoxin-exposed sea turtles is supportive care, and it has been difficult to design alternative treatment strategies without an understanding of the effects of brevetoxins in turtles in vivo. Previous studies using the freshwater turtle as a model species showed that brevetoxin-3 impacts the nervous and muscular systems, and is detoxified and eliminated primarily through the liver, bile, and feces. In this study, freshwater turtles (Trachemys scripta) were exposed to brevetoxin (PbTx-3) intratracheally at doses causing clear systemic effects, and treatment strategies aimed at reducing the postexposure neurological and muscular deficits were tested. Brevetoxin-exposed T. scripta displayed the same behaviors as animals admitted to rehabilitation centers for toxin exposure, ranging from muscle twitching and incoordination to paralysis and unresponsiveness. Two treatment regimes were tested: cholestyramine, a bile acid sequestrant; and an intravenous lipid emulsion treatment (Intralipidt) that provides an expanded circulating lipid volume. Cholestyramine was administered orally 1 hr and 6 hr post PbTx-3 exposure, but this regime failed to increase toxin clearance. Animals treated with Intralipid (100 mg/kg) 30 min after PbTx-3 exposure had greatly reduced symptoms of brevetoxicosis within the first 2 hr compared with animals that did not receive the treatment, and appeared fully recovered within 24 hr compared with toxin-exposed control animals that did not receive Intralipid. The results strongly suggest that Intralipid treatment for lipophilic toxins such as PbTx-3 has the potential to reduce morbidity and mortality in HAB-exposed sea turtles.

Assessing Karenia brevis red tide as a mortality factor of sea turtles in Florida, USA.

Data on Karenia brevis red tides (≥105 cells l-1) and on dead or debilitated (i.e. stranded) Kemp's ridleys Lepidochelys kempii, loggerheads Caretta caretta, green turtles Chelonia mydas, hawksbills Eretmochelys imbricata, and leatherbacks Dermochelys coriacea documented in Florida during 1986-2013 were evaluated to assess red tides as a sea turtle mortality factor. Unusually large numbers of stranded sea turtles were found coincident with red tides primarily along Florida's Gulf coast but also along a portion of Florida's Atlantic coast. These strandings were mainly adult and large immature loggerheads and Kemp's ridleys, and small immature green turtles and hawksbills. Unusually large numbers of stranded leatherbacks never coincided with red tide. For the 3 most common species, results of stranding data modeling, and of investigations that included determining brevetoxin concentrations in samples collected from stranded turtles, all indicated that red tides were associated with greater and more frequent increases in the numbers of stranded loggerheads and Kemp's ridleys than in the number of stranded green turtles. The mean annual number of stranded sea turtles attributed to K. brevis red tide was 80 (SE = 21.6, range = 2-338). Considering typical stranding probabilities, the overall mortality was probably 5-10 times greater. Red tide accounted for a substantial portion of all stranded loggerheads (7.1%) and Kemp's ridleys (17.7%), and a smaller portion of all stranded green turtles (1.6%). Even though K. brevis red tides occur naturally, the mortality they cause needs to be considered when managing these threatened and endangered species.

Saxitoxin Poisoning in Green Turtles (Chelonia mydas) Linked to Scavenging on Mass Mortality of Caribbean Sharpnose Puffer Fish (Canthigaster rostrata-Tetraodontidae).

Fish within the family Tetraodontidae are potential sources of both endogenous tetrodotoxins (TTXs) and dietary derived saxitoxins (STXs). Ingestion of fish tissues containing these toxins by other vertebrates can lead to severe illness and death. The Caribbean sharpnose puffer (Canthigaster rostrata) is a widespread tetraodontid species within the western Atlantic. Mass settlement of juveniles into foraging habitats have been associated with large-scale puffer fish mortality events. In 2013, 2014, and 2017, puffer mortality events on the southern Caribbean coast of Costa Rica were also associated with strandings of green turtles (Chelonia mydas) found to have fed on C. rostrata. Stranded sea turtles were found dead without apparent cause or alive with severe neurological signs that resolved during short periods of captivity. Puffer fish and turtle organ samples were analyzed for both TTXs and STXs. Concentrations of TTXs were extremely low in the fish (0.5-0.7 µg/g) and undetectable in turtle stomach contents. However, concentrations of STXs in whole fish (16.6-47.5 µg STX-eq/g) exceeded the 0.8 µg STX-eq/g human seafood safety threshold for STXs by orders of magnitude. Saxitoxins were also detected in samples of stomach contents (ingested fish), brain, lung, kidney, and serum from three affected turtles. Study results indicate that saxitoxicosis resulting from opportunistic foraging on C. rostrata during fish mortality events may be a significant factor in episodic stranding of green sea turtles in this region.

Brevetoxin exposure in sea turtles in south Texas (USA) during Karenia brevis red tide.

Five green (Chelonia mydas) and 11 Kemp's ridley (Lepidochelys kempii) sea turtles found dead, or that died soon after stranding, on the southern Texas (USA) coast during 2 Karenia brevis blooms (October 2015, September-October 2016) were tested for exposure to brevetoxins (PbTx). Tissues (liver, kidney) and digesta (stomach and intestinal contents) were analyzed by ELISA. Three green turtles found alive during the 2015 event and 2 Kemp's ridley turtles found alive during the 2016 event exhibited signs of PbTx exposure, including lethargy and/or convulsions of the head and neck. PbTx were detected in 1 or more tissues or digesta in all 16 stranded turtles. Detected PbTx concentrations ranged from 2 to >2000 ng g-1. Necropsy examination and results of PbTx analysis indicated that 10 of the Kemp's ridleys and 2 of the green turtles died from brevetoxicosis via ingestion. This is the first documentation of sea turtle mortality in Texas attributed to brevetoxicosis.

Role of Biomarkers in Monitoring Brevetoxins in Karenia brevis Exposed Shellfish.

Monitoring and management programs for marine toxins in seafood depend on efficient detection tools for their success in protecting public health. Here we review current methods of detection for neurotoxic shellfish poisoning (NSP) toxins, and current knowledge in brevetoxin metabolism in shellfish. In addition, we discuss a novel approach to developing monitoring tools for NSP toxins in molluscan shellfish. NSP is a seafood-borne disease caused by the consumption of brevetoxin-contaminated shellfish. Brevetoxins are a suite of cyclic polyether compounds found in blooms of the marine dinoflagellate Karenia brevis (K. brevis) and are potent neurotoxins. Preventive controls for NSP in the U.S. currently rely upon environmental monitoring of K. brevis blooms and assessment of their shellfish toxicity by mouse bioassay. The mouse bioassay for NSP approved by National Shellfish Sanitation Program was developed in the 1960s when very little information on the structural and toxicological properties of brevetoxins in algae and shellfish was available. Alternative methods to mouse bioassay based on current scientific knowledge in the area are needed for monitoring NSP toxins. It is now established that brevetoxins are metabolized extensively in shellfish. Algal brevetoxins undergo oxidation and reduction, as well as conjugation with fatty acids and amino acids in shellfish. Recently, three metabolites have been identified as biomarkers of brevetoxin exposure and toxicity in Eastern oyster (Crassostrea virginica) and hard clam (Mercenaria sp.). The role of these biomarkers in monitoring NSP toxins in K. brevis exposed molluscan shellfish is reviewed. Comparisons of biomarker levels by liquid chromatography-mass spectrometry (LC-MS) with composite toxin as measured by enzyme linked immunosorbent assay (ELISA), and shellfish toxicity by mouse bioassay, support the application of these biomarkers as a dynamic and powerful approach for monitoring brevetoxins in shellfish and prevention of NSP.

GREEN TREE FROG ( HYLA CINEREA) AND GROUND SQUIRREL ( XEROSPERMOPHILUS SPILOSOMA) MORTALITY ATTRIBUTED TO INLAND BREVETOXIN TRANSPORT AT PADRE ISLAND NATIONAL SEASHORE, TEXAS, USA, 2015.

: On 16 September 2015, a red tide ( Karenia brevis) bloom impacted coastal areas of Padre Island National Seashore Park, Texas, US. Two days later and about 0.9 km inland, 30-40 adult green tree frogs ( Hyla cinerea) were found dead after displaying tremors, weakness, labored breathing, and other signs of neurologic impairment. A rainstorm accompanied by high winds, rough surf, and high tides, which could have aerosolized brevetoxin, occurred on the morning of the mortality event. Frog carcasses were in good body condition but contained significant brevetoxin in tissues. Tissue brevetoxin was also found in two dead or dying spotted ground squirrels ( Xerospermophilus spilosoma) and a coyote ( Canis latrans) found in the area. Rainwater collected from the location of the mortality event contained brevetoxin. Green tree frog and ground squirrel mortality has not been previously attributed to brevetoxin exposure and such mortality suggested that inland toxin transport, possibly through aerosols, rainfall, or insects, may have important implications for coastal species.

Tissue uptake, distribution and excretion of brevetoxin-3 after oral and intratracheal exposure in the freshwater turtle Trachemys scripta and the diamondback terrapin Malaclemys terrapin.

Harmful algal blooms (HABs) occur nearly annually off the west coast of Florida and can impact both humans and wildlife, resulting in morbidity and increased mortality of marine animals including sea turtles. The key organism in Florida red tides is the dinoflagellate Karenia brevis that produces a suite of potent neurotoxins referred to as the brevetoxins (PbTx). Despite recent mortality events and rehabilitation efforts, still little is known about how the toxin directly impacts sea turtles, as they are not amenable to experimentation and what is known about toxin levels and distribution comes primarily from post-mortem data. In this study, we utilized the freshwater turtle Trachemys scripta and the diamondback terrapin, Malaclemys terrapin as model organisms to determine the distribution, clearance, and routes of excretion of the most common form of the toxin, brevetoxin-3, in turtles. Turtles were administered toxin via esophageal tube to mimic ingestion (33.48µg/kg PbTx-3, 3×/week for two weeks for a total of 7 doses) or by intratracheal instillation (10.53µg/kg, 3×/week for four weeks for a total of 12 doses) to mimic inhalation. Both oral and intratracheal administration of the toxin produced a suite of behavioral responses symptomatic of brevetoxicosis. The toxin distributed to all organ systems within 1h of administration but was rapidly cleared out over 24-48h, corresponding to a decline in clinical symptoms. Excretion appears to be primarily through conjugation to bile salts. Histopathological study revealed that the frequency of lesions varied within experimental groups with some turtles having no significant lesions at all, while similar lesions were found in a low number of control turtles suggesting another common factor(s) could be responsible. The overall goal of this research is better understand the impacts of brevetoxin on turtles in order to develop better treatment protocols for sea turtles exposed to HABs.

Sublethal red tide toxin exposure in free-ranging manatees (Trichechus manatus) affects the immune system through reduced lymphocyte proliferation responses, inflammation, and oxidative stress.

The health of many Florida manatees (Trichechus manatus latirostris) is adversely affected by exposure to blooms of the toxic dinoflagellate, Karenia brevis. K. brevis blooms are common in manatee habitats of Florida's southwestern coast and produce a group of cyclic polyether toxins collectively referred to as red tide toxins, or brevetoxins. Although a large number of manatees exposed to significant levels of red tide toxins die, several manatees are rescued from sublethal exposure and are successfully treated and returned to the wild. Sublethal brevetoxin exposure may potentially impact the manatee immune system. Lymphocyte proliferative responses and a suite of immune function parameters in the plasma were used to evaluate effects of brevetoxin exposure on health of manatees rescued from natural exposure to red tide toxins in their habitat. Blood samples were collected from rescued manatees at Lowry Park Zoo in Tampa, FL and from healthy, unexposed manatees in Crystal River, FL. Peripheral blood leukocytes (PBL) isolated from whole blood were stimulated with T-cell mitogens, ConA and PHA. A suite of plasma parameters, including plasma protein electrophoresis profiles, lysozyme activity, superoxide dismutase (SOD) activity, and reactive oxygen/nitrogen (ROS/RNS) species, was also used to assess manatee health. Significant decreases (p<0.05) in lymphocyte proliferation were observed in ConA and PHA stimulated lymphocytes from rescued animals compared to non-exposed animals. Significant correlations were observed between oxidative stress markers (SOD, ROS/RNS) and plasma brevetoxin concentrations. Sublethal exposure to brevetoxins in the wild impacts some immune function components, and thus, overall health, in the Florida manatee.

Biomarkers of brevetoxin exposure and composite toxin levels in hard clam (Mercenaria sp.) exposed to Karenia brevis blooms.

Brevetoxins in clams (Mercenaria sp.) exposed to recurring blooms of Karenia brevis in Sarasota Bay, FL, were studied over a three-year period. Brevetoxin profiles in toxic clams were generated by ELISA and LC-MS. Several brevetoxin metabolites, as identified by LC-MS, were major contributors to the composite brevetoxin response of ELISA. These were S-desoxyBTX-B2 (m/z 1018), BTX-B2 (m/z 1034), BTX-B5 (m/z 911), open A-ring BTX-B5 (m/z 929), and BTX-B1 (m/z 1018). Summed values of these metabolites were highly correlated (R(2) = 0.9) with composite B-type brevetoxin measurements by ELISA. S-desoxyBTX-B2, BTX-B2, and BTX-B1 were the most persistent and detectable in shellfish for several months after dissipation of blooms. These metabolites were selected as LC-MS biomarkers of brevetoxin exposure and reflective of composite B-type brevetoxins in hard clam. ELISA and LC-MS values were moderately correlated with toxicity of the shellfish by mouse bioassay. ELISA and LC-MS methods offer rapid screening and confirmatory determination of brevetoxins, respectively, as well as toxicity assessment in clams exposed to K. brevis blooms.

Osmotic stress does not trigger brevetoxin production in the dinoflagellate Karenia brevis.

With the global proliferation of toxic harmful algal bloom species, there is a need to identify the environmental and biological factors that regulate toxin production. One such species, Karenia brevis, forms nearly annual blooms that threaten coastal regions throughout the Gulf of Mexico. This dinoflagellate produces brevetoxins, which are potent neurotoxins that cause neurotoxic shellfish poisoning and respiratory illness in humans, as well as massive fish kills. A recent publication reported that a rapid decrease in salinity increased cellular toxin quotas in K. brevis and hypothesized that brevetoxins serve a role in osmoregulation. This finding implied that salinity shifts could significantly alter the toxic effects of blooms. We repeated the original experiments separately in three different laboratories and found no evidence for increased brevetoxin production in response to low-salinity stress in any of the eight K. brevis strains we tested, including three used in the original study. Thus, we find no support for an osmoregulatory function of brevetoxins. The original publication also stated that there was no known cellular function for brevetoxins. However, there is increasing evidence that brevetoxins promote survival of the dinoflagellates by deterring grazing by zooplankton. Whether they have other as-yet-unidentified cellular functions is currently unknown.

Brevetoxin in blood, biological fluids, and tissues of sea turtles naturally exposed to Karenia brevis blooms in central west Florida.

In 2005 and 2006, the central west Florida coast experienced two intense Karenia brevis red tide events lasting from February 2005 through December 2005 and August 2006 through December 2006. Strandings of sea turtles were increased in the study area with 318 turtles (n = 174, 2005; n = 144, 2006) stranding between 1 January 2005 and 31 December 2006 compared to the 12-yr average of 43 +/- 23 turtles. Live turtles (n = 61) admitted for rehabilitation showed clinical signs including unresponsiveness, paresis, and circling. Testing of biological fluids and tissues for the presence of brevetoxin activity by enzyme-linked immunosorbent assay found toxin present in 93% (52 of 56) of live stranded sea turtles, and 98% (42 of 43) of dead stranded sea turtles tested. Serial plasma samples were taken from several live sea turtles during rehabilitation and toxin was cleared from the blood within 5-80 days postadmit depending upon the species tested. Among dead animals the highest brevetoxin levels were found in feces, stomach contents, and liver. The lack of significant pathological findings in the majority of animals necropsied supports toxin-related mortality.

Brevetoxicosis in seabirds naturally exposed to Karenia brevis blooms along the central west coast of Florida.

Harmful algal bloom events caused by the dinoflagellate Karenia brevis occurred along the central west Florida, USA, coast from February 2005 through December 2005 and from August 2006 through December 2006. During these events, from 4 February 2005 through 28 November 2006, live, debilitated seabirds admitted for rehabilitation showed clinical signs that included disorientation, inability to stand, ataxia, and seizures. Testing of blood, biologic fluids, and tissues for brevetoxin by enzyme-linked immunosorbent assay found toxin present in 69% (n=95) of rehabilitating seabirds. Twelve of the 19 species of birds had evidence of brevetoxin exposure. Commonly affected species included Double-crested Cormorants (Phalacrocorax auritus), Brown Pelicans (Pelecanus occidentalis), Great Blue Herons (Ardea herodias), and Common Loons (Gavia immer). Serial blood and fecal samples taken from several live seabirds during rehabilitation showed that brevetoxin was cleared within 5-10 days after being admitted to the rehabilitation facility, depending on the species tested. Among seabirds that died or were euthanized, the highest brevetoxin concentrations were found in bile, stomach contents, and liver. Most dead birds had no significant pathologic findings at necropsy, thereby supporting brevetoxin-related mortality.

Coyote (Canis latrans) and domestic dog (Canis familiaris) mortality and morbidity due to a Karenia brevis red tide in the Gulf of Mexico.

In October 2009, during a Karenia brevis red tide along the Texas coast, millions of dead fish washed ashore along the 113-km length of Padre Island National Seashore (PAIS). Between November 2009 and January 2010, at least 12 coyotes (Canis latrans) and three domestic dogs (Canis familiaris) died or were euthanized at PAIS or local veterinary clinics because of illness suspected to be related to the red tide. Another red tide event occurred during autumn 2011 and, although fewer dead fish were observed relative to the 2009 event, coyotes again were affected. Staff at PAIS submitted carcasses of four coyotes and one domestic dog from November 2009 to February 2010 and six coyotes from October to November 2011 for necropsy and ancillary testing. High levels of brevetoxins (PbTxs) were measured by enzyme-linked immunosorbent assay in seven of the coyotes and the dog, with concentrations up to 634 ng PbTx-3 eq/g in stomach contents, 545 ng PbTx-3 eq/g in liver, 195 ng PbTx-3 eq/g in kidney, and 106 ng PbTx-3 eq/mL in urine samples. Based on red tide presence, clinical signs, and postmortem findings, brevetoxicosis caused by presumptive ingestion of toxic dead fish was the likely cause of canid deaths at PAIS. These findings represent the first confirmed report of terrestrial mammalian wildlife mortalities related to a K. brevis bloom. The implications for red tide impacts on terrestrial wildlife populations are a potentially significant but relatively undocumented phenomenon.