Abstract
Harmful algal blooms (HAB), more commonly referred to as 'red tides' often produce toxins with deleterious health effects in marine organisms. Brevetoxins, produced by the dinoflagellate Karenia brevis, have been documented to kill large numbers of fish and are an important cause of morbidity and mortality of marine mammals.1 Bottlenose dolphins can be exposed to brevetoxin either by ingestion of contaminated food or by inhalation of aerosolized toxin. However, exposure to brevetoxin is not always lethal in exposed animals.1 Therefore, dolphins can be exposed to repeated/chronic, sub-lethal concentrations, which could impact their overall health. Brevetoxins are lipophilic, polyether neurotoxins that specifically bind to cell membranes and activate voltage gated sodium channels, leading to an influx of sodium into the cell.2,3 These voltage gated sodium channels have been described on immune cells such as lymphocytes, neutrophils, and natural killer (NK) cells, suggesting that brevetoxin may interact with immune cells and modulate their functions.4 The objectives of this study were to measure the changes in innate (phagocytosis, respiratory burst, NK cell activity) and adaptive (mitogen-induced B and T lymphocyte proliferation) immune functions upon in vitro exposure to brevetoxin using bottlenose dolphin peripheral blood immune cells. The direct effects of brevetoxin on immune functions were performed using increasing concentrations of brevetoxin (PbTx-3) (0, 0.01, 0.1, 1, 10, 100, 500, and 1000 nM). Brevetoxin significantly increased spontaneous T lymphocyte proliferation at 0.1-1000 nM compared to the unexposed control. Brevetoxin significantly increased suboptimal and optimal ConA-induced T lymphocyte proliferation at 0.01-100 nM and 0.1 nM respectively, as well as suboptimal and optimal LPS-induced B lymphocyte proliferation at 0.01-100 nM and 0.01-500 nM respectively. Neutrophil and monocyte respiratory burst were significantly increased at 500 and 1000 nM. There were no effects on neutrophil or monocyte phagocytosis, or NK cell activity. Importantly, these in vitro concentrations were within the range measured in the blood of dolphins during two unusual mortality events, suggesting that naturally exposed dolphins may be at risk for immunomodulation. We are currently trying to semi-quantify intracellular Pb-Tx by immunohistochemistry and immunofluorescence to determine if changes in immune functions are proportional to intracellular brevetoxin concentrations. Brevetoxin-induced immunomodulation may increase an animal's susceptibility to secondary bacterial, viral, or fungal infections. Understanding the risk for immunomodulation upon HAB toxin exposure will contribute in the health assessment and management of marine mammals, as well as guide veterinarians and wildlife rehabilitators in caring for and treating afflicted animals.
Acknowledgements
We would like to thank the US Navy Marine Mammal Program for providing blood samples. Funding for this project was provided by the Morris Animal Foundation.
References
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