Elsevier

Harmful Algae

Volume 109, November 2021, 102109
Harmful Algae

Original article
Acute oral toxicity and tissue residues of saxitoxin in the mallard (Anas platyrhynchos)

https://doi.org/10.1016/j.hal.2021.102109Get rights and content

Highlights

  • Median lethal dose of saxitoxin in mallards was estimated to be 167 µg kg−1.

  • Detection of saxitoxin in tissues, not including the gastrointestinal tract, was inconsistent in mallards that died acutely.

  • No specific abnormalities or tissue pathology was associated with saxitoxin exposure.

  • Weight loss was associated with saxitoxin exposure in surviving mallards.

Abstract

Since 2014, widespread, annual mortality events involving multiple species of seabirds have occurred in the Gulf of Alaska, Bering Sea, and Chukchi Sea. Among these die-offs, emaciation was a common finding with starvation often identified as the cause of death. However, saxitoxin (STX) was detected in many carcasses, indicating exposure of these seabirds to STX in the marine environment. Few data are available that describe the effects of STX in birds, thus presenting challenges for determining its contributions to specific mortality events. To address these knowledge gaps, we conducted an acute oral toxicity trial in mallards (Anas platyrhynchos), a common laboratory avian model, using an up-and-down method to estimate the median lethal dose (LD50) for STX. Using an enzyme-linked immunosorbent assay (ELISA), we tested select tissues from all birds and feces from those individuals that survived initial dosing. Samples with an ELISA result that exceeded approximately 10 µg 100 g−1 STX and randomly selected ELISA negative samples were further tested by high-performance liquid chromatography (HPLC). Tissues collected from mallards were also examined grossly at necropsy and then later by microscopy to identify lesions attributable to STX. The estimated LD50 was 167 µg kg−1 (95% CI = 69–275 µg kg−1). Saxitoxin was detected in fecal samples of all mallards tested for up to 48 h after dosing and at the end of the sampling period (7 d) in three birds. In those individuals that died or were euthanized <2 h after dosing, STX was readily detected throughout the gastrointestinal tract but only infrequently in heart, kidney, liver, lung, and breast muscle. No gross or microscopic lesions were observed that could be attributable to STX exposure. Given its acute toxicity, limited detectability, and frequent occurrence in the Alaska marine environment, additional research on STX in seabirds is warranted.

Introduction

Saxitoxin (STX) is a highly potent algal toxin that occurs in many parts of the world across a wide variety of aquatic habitats, including freshwater, marine, and saline waterbodies, and is known to cause illness in fish, birds, and mammals (Landsberg et al., 2014; Lefebvre et al., 2004; Lefebvre et al., 2005; Wiese et al., 2010). Saxitoxin and its 56 known congeners are a broad group of natural toxins commonly known as paralytic shellfish toxins (PSTs; Hall et al., 1990; Wiese et al., 2010). A single bloom event can harbor more than one PST, and STX, if present, is not always the dominant toxin (Kaas and Henriksen, 2000). These toxins can enter the food web when PST producing dinoflagellates or cyanobacteria are ingested by shellfish, copepods, or other invertebrates and these, in turn, are consumed by larger organisms. Ingestion of PST by mammal and bird species can result in muscular weakness, motor incoordination, respiratory paralysis, and death (Gibble and Hoover, 2018; Landsberg et al., 2014; Shumway et al., 2003).

Saxitoxin has been reported as a cause or suspected cause of death in waterbird and marine mammal mortality events; however, it is often difficult to establish causality among free-ranging populations (Ben-Gigirey et al., 2021; Starr et al., 2017; Van Dolah et al., 2003). Conclusions related to STX intoxication among wildlife are typically based on indirect evidence, including detection of STX in the environment and/or prey species, detection of STX in tissues or gut contents, and the absence of other potential causes of mortality (Armstrong et al., 1978; Coulson et al., 1968; Nisbet, 1983; Shearn-Bochsler et al., 2014; Starr et al., 2017). Recent, widespread seabird mortality events in Alaska, combined with detection of algal toxins in marine wildlife throughout the region prompted investigation of STX as a possible contributing factor (Lefebvre et al., 2016; U.S. Geological Survey, 2020; Van Hemert et al., 2020, 2021). In an assessment of algal toxins associated with a large die-off of common murres (Uria aalge) in the Gulf of Alaska in 2015–2016, STX was detected commonly in beach-cast carcasses as well as in apparently healthy birds, although overall concentrations were higher in samples from birds associated with the mortality event (Van Hemert et al., 2020). In 2017, a multispecies seabird die-off occurred in the Bering and Chukchi Seas during which 88% (14 of 16) of northern fulmars (Fulmarus glacialis) tested had detectable levels of STX in their tissues (Van Hemert et al., 2021). Although emaciation was a common finding, and starvation was identified as a likely cause of death in numerous avian mortality events in Alaskan marine ecosystems since 2015 (U.S. Geological Survey, 2020, Van Hemert et al., 2020, 2021), the frequent detection of STX in carcasses warranted additional investigation. Without knowledge of STX toxicity in birds, or how actual exposure amounts relates to measurable tissue toxin concentrations, the contributions of STX to the cause of these and other recent die-offs remains elusive.

Previous STX dosing studies in birds have been even more limited than field studies. When European starlings (Sturnus vulgaris) were fed clams (Saxidomus gigantea) containing STX, mortality was observed in 1 of 6 birds (Kvitek and Beitler, 1988). No mortality was reported in free-ranging glaucous-winged gulls (Larus glaucescens) after ingesting STX-contaminated shellfish, but regurgitation of the contaminated material and aversion towards future ingestion was noted (Kvitek, 1991).

The primary objective of this study was to evaluate toxicity of STX in birds by estimating the acute oral median lethal dose (LD50) of STX in mallards (Anas platyrhynchos). Secondarily, we evaluated associated tissue pathology, examined distribution and concentration of toxin in tissues, and assessed elimination of STX in fecal material of birds experimentally exposed to STX. These results will be useful for assessing the threat of STX to waterbirds and other wildlife and provide important baseline information for future experimental studies examining the toxicity and pharmacokinetics of STX in seabirds.

Section snippets

Study animals

Fourteen female mallards approximately 10–12 weeks of age were acquired from a commercial vendor in Iowa, United States. Females were used to reduce the potential of introducing another variable (sex) to this study, and females generally have lower LD50 values then males which allows for a more conservative LD50 estimate (reviewed in Bruce, 1985). These birds were hatched and raised indoors for 4 to 6 weeks then moved to outdoor freshwater ponds and provided with supplemental food until

Behavioral observations

Clinical signs of acute toxin ingestion in mallards included head shaking (11/11 mallards), excessive drinking (10/11), and regurgitation (6/11; not recorded in five birds that died acutely). Among survivors (dosed birds that did not die acutely), wing twitching and settling (lifting and dropping wings back into place; 3/5) and tail ‘wagging’ (3/5) behaviors were recorded. None of these behaviors were identified among control birds. Of mallards that died acutely, death occurred between 13 and

Evaluation of LD50

Seabirds and other waterbirds are commonly exposed to STX (Gibble and Hoover 2018; Shumway et al., 2003). Periodically, mortality in birds has been attributed to saxitoxicosis (Shearn-Bochsler et al., 2014; Shumway et al., 2003; Van Hemert et al., 2020), although there had previously been little quantitative information available to interpret findings of STX in birds. In this study, the toxicity of STX was investigated in mallards and the LD50 was estimated to be 167 µg kg−1 (95%

Conclusions

This study demonstrated acute mortality from STX ingestion in mallards and identified potentially greater sensitivity as compared to laboratory mice. Clinical signs of acute toxicity included head shaking, excessive drinking, and regurgitating; among dosed birds that survived, we also observed wing twitching and settling and tail ‘wagging.’ We did not observe gross or histopathological lesions associated with STX toxicity, and detection of STX in tissues other than the gastrointestinal tract

Author declaration

The data contained in this manuscript is original and has not been previously published nor is it being considered for publication elsewhere.

Declaration of Competing Interest

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Acknowledgments

We thank Dr. E. Falendysz, D. Calhoun, K. Werner, H. Lamb, and the rest of the animal care staff for providing veterinary and daily care of the animals and S. Steinfeldt and D. Johnson for assistance with necropsies and sample processing. Dr. K. Kuletz provided invaluable guidance and support in the conception and development of this project. This work was funded by the USGS Ecosystems Mission Area Biological Threats and Invasive Species Research (Fish and Wildlife Disease and Invasive Species)

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