Abstract

As top predators with relatively long life spans, marine mammals are vulnerable to the chronic effects of pollution, contaminants and emerging pathogens. Studies that evaluate these effects in marine mammal populations are scarce due to the lack of baseline data related to physiology and health of these animals. This study proposes implementation of a molecular assay that can be used as a powerful tool for health assessment studies due to its excellent sensitivity and specificity requiring minimal amount of tissue sample. Paired blood and skin samples were collected from wild belugas (Delphinapterus leucas) following live capture-release health assessments and subsistence hunts in Alaska. Quantification of ten selected gene products (cytokines, acute-phase and stress-response proteins) were carried out by using real-time polymerase chain reaction (RT-PCR) with SYBR-Green technology in both blood and skin, and baseline levels were established in two populations of wild belugas in Alaska, as well as in resident whales in Mystic Aquarium, CT. In general, the blood samples obtained from wild belugas showed significantly higher (p<0.05) levels expression for inflammatory cytokines interferon gamma (IFNγ), transforming growth factor-beta (TGFβ), and the stress/immune response markers aryl hydrocarbon receptor (AHR), heat-shock protein (HSP70) when compared to aquarium whales, suggesting their differentiated health status and continuous exposure to stressors in a natural environment. Among the two wild populations, subsistence-hunted whales showed a significant (p<0.001) downregulation of interleukin-10 (IL10) when compared to live captured-released whales. Individual gene expression profiles for both populations of whales showed the largest overall variability in gene expression for interleukin-12 (IL12), IFNγ, HSP70 and cyclooxygenase (COX2) genes, indicating interindividual variability in type and degree of response to various stressors. Expression of the cytokines IL10, IL12 and IFNγ were significantly lower in skin samples as compared to blood (p<0.001), indicating lower copy numbers. While expression of COX2 and HSP70 were significantly higher (p<0.05) in skin than in blood for both populations, expression of AHR, glucocorticoid receptor (Nr3c1) and TGFβ genes were at comparable levels in paired blood and skin samples. The results obtained from the tested biomarkers will be further investigated for correlations with the paired blood hematology and catecholamine measurements. Given their potential to provide information about the physiological status of marine mammals, these markers are therefore suggested as promising biomarkers of health in both blood and skin samples. Overall, the gene expression profiles of the markers identified in this study look promising to reveal the health and immune status of marine mammals, and can be used as indicators of environmental and anthropogenic stressors.

Acknowledgements

The authors wish to thank to the Bristol Bay and Point Lay field teams, Bristol Bay Native Association for their collaboration and help in sampling. The authors would like to give special thanks to Mystic Aquarium animal care and veterinary team, the beluga husbandry staff and research staff for helping to provide samples. The authors are also very thankful to the Bristol Bay Marine Mammal Council, the Alaska Beluga Whale Committee, Bristol Bay Native Association, Dr. Robert Suydam and The Department of Wildlife Management in North Slope Borough (Barrow, AK) and the community of Point Lay, AK for their permission and collaboration to carry out this field work. The authors are truly grateful to Georgia Aquarium for providing financial support for Bristol Bay field team. This study was funded by Office of Naval Research (ONR Award no: N00014-14-1-0411). Sampling of belugas in Bristol Bay, AK was conducted under National Marine Fisheries Service Scientific Research Permit #782-1719 (for the year 2008) and #14245 (for the years 2012–2014). Sampling of live-captured belugas in Point Lay, AK was conducted under Alaska Department of Fish and Game Research Permit #14610, and of subsistence-hunted whales was conducted under permit #17350 by the Department of Wildlife Management, North Slope Borough, AK.

* Presenting author