Abstract

Beluga whales (Delphinapterus leucas) are exposed to many environmental and anthropogenic stressors, including loss of sea ice due to climate change, oil and gas exploration, shipping traffic, and pollution. The response to stressors is mediated by the hypothalamic pituitary adrenal (HPA) axis which releases glucocorticoids (GC) into the circulatory system that suppress unnecessary bodily functions and shift resources to quickly adjust to changes in the environment.^1,2 Although the stress response is necessary in the short term, continued GC release can have detrimental effects, including compromised immunity and low reproductive fitness.³ Therefore, GC are often used to measure stress and to assess population health.⁴

GC are metabolized by the liver and excreted with the feces.⁵ The resulting fecal GC metabolites (FGM) make an ideal biomarker for quantifying stress, and have been used to quantify endocrine activity in many animals, including wild cetaceans.^6,7 In addition, FGM analysis provides a more accurate estimation of overall hormone levels, as a single sample contains multiple hours of hormone accumulation, versus blood that that provides only a snapshot at time of sampling.^8,9

Aquarium belugas offer an opportunity for fecal sampling through positive behavioral training, which can be used to validate assays for the quantification of biomarkers of health, including FGM. FGM analysis also provides an opportunity for long-term monitoring of wild belugas, including endangered Cook Inlet belugas.

In this study, a commercially available enzyme immunoassay (EIA) was validated for measuring FGM in beluga fecal extracts. Biological validation was carried out with beluga whales (n=2) housed at Mystic Aquarium to show that FGM levels increase in response to an external challenge. Samples collected before and after an out of water examination (OWE) show a spike in FGM concentration (15–60 ng/g) within 4–7 h before returning to baseline levels (∼10 ng/g) by 48 h post-examination.

Longitudinal analysis of fecal samples collected from 3 aquarium belugas reveal a seasonal variation, with winter having the highest FGM levels (74.82 ng/g) and spring having the lowest (28.14 ng/g). Establishing the normal ranges of FGM in aquarium belugas during important lifecycle stages (e.g., breeding) will provide important information for husbandry and veterinary staff caring for these animals. Data from samples collected from subsistence hunted belugas from Point Lay, Alaska reveal significant differences between sex, age, and reproductive status, with pregnant females having significantly higher FGM levels (94.55 ng/g) than adult males (14.18 ng/g). Notably, a pregnant beluga carrying a full-term fetus had the highest FGM concentration (131.19 ng/g). In addition, the potential for FGM analysis as a diagnostic monitoring tool was demonstrated in the case of a rescued beluga calf from Cook Inlet. Significant variation in FGM levels (Avg=21.83 ng/g) was observed over the successful 3-month rehabilitation, however, all were within the range observed in healthy aquarium belugas.

This study has demonstrated that FGM analysis can be used to monitor endocrine activity in aquarium belugas, as well as wild beluga populations, thus targeting and playing a key role in conservation strategies for the survival of the species.

Acknowledgements

The authors would like to thank the Mystic Aquarium Arctic Coast husbandry team for their effort in collecting fecal samples, Brandy Hebert for her assistance in the laboratory, the Community of Point Lay, Alaska. for support and assistance in coordination of sample collection under NMFS permit #17350. Thanks to Katie Graham and the New England Aquarium for the use of their equipment. Special thanks to the collaborative efforts of staff caring for the rescued beluga calf for collecting fecal samples at Alaska SeaLife Center under MMHSRP MMPA/ESA permit #18786-03. This work was carried out under ONR grant #N00014-11-1-0437 and NSF REU grant #1658663.

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