ABSTRACT
In an era of rapid environmental change and increasing human presence, researchers need efficient tools for tracking contaminants to monitor the health of Antarctic flora and fauna. Here, we examined the utility of leopard seal whiskers as a biomonitoring tool that reconstructs time-series of significant ecological and physiological biomarkers. Leopard seals (Hydrurga leptonyx) are a sentinel species in the Western Antarctic Peninsula due to their apex predator status and top-down effects on several Antarctic species. However, there are few data on their contaminant loads. We analyzed leopard seal whiskers (n = 18 individuals, n = 981 segments) collected during 2018-2019 field seasons to acquire longitudinal profiles of non-essential (Hg, Pb, and Cd) and essential (Se, Cu, and Zn) trace elements, stable isotope (áº15N and áº13C) values and to assess Hg risk with Se:Hg molar ratios. Whiskers provided between 46 and 286 cumulative days of growth with a mean ~ 125 days per whisker (n = 18). Adult whiskers showed variability in non-essential trace elements over time that could partly be explained by changes in diet. Whisker Hg levels were insufficient (<20 ppm) to consider most seals being at "high" risk for Hg toxicity. Nevertheless, maximum Hg concentrations observed in this study were greater than that of leopard seal hair measured two decades ago. However, variation in the Se:Hg molar ratios over time suggest that Se may detoxify Hg burden in leopard seals. Overall, we provide evidence that the analysis of leopard seal whiskers allows for the reconstruction of time-series ecological and physiological data and can be valuable for opportunistically monitoring the health of the leopard seal population and their Antarctic ecosystem during climate change.
Subject(s)
Mercury , Seals, Earless , Trace Elements , Animals , Antarctic Regions , Ecosystem , Isotopes/analysis , Mercury/analysis , Trace Elements/analysis , Vibrissae/chemistryABSTRACT
The Pacific walrus (Odobenus rosmarus divergens) is an iconic Arctic marine mammal and an important resource to many Alaska Natives. A decrease in sea ice habitat and unknown population numbers has led to concern of the long-term future health of the walrus population. There is currently no clear understanding of how walrus physiology might be affected by a changing Arctic ecosystem. In this study, steroid hormone concentrations (progesterone, testosterone, cortisol and estradiol) were analysed in walrus bones collected during archaeological [3585-200 calendar years before present (BP)], historical [1880-2006 common era (CE)] and modern (2014-2016 CE) time periods, representing ~ 3651 years, to track changes in reproductive activity and cortisol concentrations (biomarker of stress) over time. Our results show that modern walrus samples have similar cortisol concentrations (median = 43.97 ± standard deviation 904.38 ng/g lipid) to archaeological walruses (38.94 ± 296.17 ng/g lipid, P = 0.75). Cortisol concentrations were weakly correlated with a 15-year average September Chukchi Sea ice cover (P = 0.002, 0.02, r 2 = 0.09, 0.04, for females and males, respectively), indicating a possible physiological resiliency to sea ice recession in the Arctic. All steroid hormones had significant negative correlations with mean walrus population estimates from 1960 to 2016 (P < 0.001). Progesterone in females and testosterone in males exhibited significant correlations with average September Chukchi Sea ice cover for years 1880-2016 (P < 0.001 for both, r2 = 0.34, 0.22, respectively). Modern walruses had significantly lower (P = < 0.001) reproductive hormone concentrations compared with historic walruses during times of rapid population increase, indicative of a population possibly at carrying capacity. This is the first study to apply bone as a tool to monitor long-term changes in hormones that may be associated with changes in walrus population size and sea ice cover.
ABSTRACT
Keratinized tissues, including whiskers, are ideal for acquiring a record of physiological parameters. Most tissues provide a snapshot of physiological status; however, whiskers may support longitudinal sampling for reproductive and stress-related hormones, if hormones are incorporated as whiskers grow and concentrations change with physiological state. Whiskers from female Steller sea lions (Eumetopias jubatus) and northern fur seals (Callorhinus ursinus) were serially sectioned and pulverized and steroid hormones were extracted. Standard methods were used to validate enzyme immunoassay kits for cortisol, progesterone, 17ß-estradiol and testosterone. All hormones were measurable in whisker segments from both species with progesterone concentrations showing cyclical patterns, which appear to signify previous pregnancies or luteal phases. Yearly progesterone concentrations were greater in years a pup was produced compared with years when no pup was observed. Free-ranging female Steller sea lions had reproductive rates between 0 and 1.0 (0.53 ± 0.33, n = 12) using a yearly progesterone concentration of 30 pg/mg or greater to classify a reproductive year as producing a pup and below 30 pg/mg as non-reproductive. Cortisol concentrations were greater near the root and rapidly declined, lacking any obvious patterns, throughout the rest of the whisker. Progesterone and testosterone concentrations were able to help determine sex of unknown individuals. Immunohistochemistry revealed that steroid hormones most likely do not leach out of whiskers based on the deposition patterns of progesterone and cortisol being present throughout the whisker length. Overall, measuring steroid hormones in whiskers can reveal individual reproductive histories over multiple years in sea lions and fur seals. Cyclical patterns of δ15N were useful for identifying periods of up to ~10 years of growth within whiskers, and measuring both stable isotopes and hormones may be useful for differentiating periods of active gestation from diapause and potentially track multi-year reproductive histories of female otariids.
ABSTRACT
Biological time series datasets provide an unparalleled opportunity to investigate regional and global changes in the marine environment. Baleen whales are long-lived sentinel species and an integral part of the marine ecosystem. Increasing anthropogenic terrestrial and marine activities alter ocean systems, and such alterations could change foraging and feeding behavior of baleen whales. In this study, we analyzed δ13C and δ15N of baleen whale earplugs from three different species (N = 6 earplugs, n = 337 laminae) to reconstruct the first continuous stable isotope profiles with a six-month resolution. Results of our study provide an unprecedented opportunity to assess behavioral as well as ecological changes. Abrupt shifts and temporal variability observed in δ13C and δ15N profiles could be indicative of behavior change such as shift in foraging location and/or trophic level in response to natural or anthropogenic disturbances. Additionally, five out of six individuals demonstrated long-term declining trends in δ13C profiles, which could suggest influence of emission of depleted 13CO2 from fossil fuel combustion referred to as the Suess effect. After adjusting the δ13C values of earplugs for the estimated Suess effect and re-evaluating δ13C profiles, significant decline in δ13C values as well as different rate of depletion suggest contribution of other sources that could impact δ13C values at the base of the food web.
Subject(s)
Ecosystem , Whales , Animals , Carbon Isotopes/analysis , Ear Protective Devices , Nitrogen Isotopes/analysisABSTRACT
Assessing the physiological impact of stressors in pinnipeds is logistically challenging, and many hormones are altered by capture and handling, limiting the utility of metabolically active tissues. Hair is increasingly being used to investigate stress-related and reproductive hormones in wildlife populations due to less-invasive collection methods, being metabolically inert once grown and containing multiple biomarkers of ecological interest. We validated enzyme immunoassays for measuring aldosterone, cortisol, corticosterone, and testosterone in lanugo (natal hair grown in utero) samples collected from Steller sea lions (Eumetopias jubatus), California sea lions (Zalophus californianus), and northern fur seals (Callorhinus ursinus). We applied laboratory validation methods including recovery of added mass, parallelism and dilution linearity. We found no effects due to differences in alcohol- versus detergent-based cleaning methods. Further, there were no significant differences in hormone concentrations in hair samples collected immediately after the molt and the subsequent samples collected over 1 year, indicating steroid hormones are stable once deposited into pinniped hair. We found no sex differences in any hormone concentrations, likely due to the lanugo being grown in utero and influenced by maternal hormone concentrations. For Steller sea lion and California sea lion pups, we found hormone concentrations significantly differed between rookeries, which warrants future research. Hair provides a novel tissue to explore the intrinsic or extrinsic drivers behind hormone measurements in otariids, which can be paired with multiple health-related metrics to further investigate possible drivers of physiological stress.
ABSTRACT
RATIONALE: A liquid chromatography/tandem mass spectrometry (LC/MS/MS) method was validated and utilized to measure and analyze four steroid hormones related to stress and reproduction in individual samples from a novel tissue, Pacific walrus (Odobenus rosmarus divergens, herein walrus) bone. This method determines steroid hormone concentrations in the remote walrus population over millennia from archaeological (>200 bp), historical (200-20 bp), and modern (2014-2016) time periods. METHODS: Lipids were extracted from walrus bone collected from these periods using methanol before LC/MS/MS analysis. Isotopically labeled internal standards for each target hormone were added to every sample. Analytical and physiological validations were performed. Additionally, a tissue comparison was done among paired walrus bone, serum, and blubber samples. A rapid resolution liquid chromatography system coupled to a QqQ mass spectrometer was used to analyze all samples after derivatization for progesterone, testosterone, cortisol, and estradiol concentrations. Multiple reaction monitoring was used for MS analysis and data were acquired in positive electrospray ionization mode. RESULTS: Progesterone, testosterone, cortisol, and estradiol were linear along their respective standard calibration curves based on their R2 values (all > 0.99). Accuracy ranged from 93-111% for all hormones. The recovery of extraction, recovery of hormones without matrix effect, was 92-101%. The overall process efficiency of our method for measuring hormones in walrus bone was 93-112%. Progesterone and testosterone concentrations were not affected by reproductive status among adult females and males, respectively. However, estradiol was different among pregnant and non-pregnant adult females. Overall, steroid hormones reflect a long-term reservoir in cortical bone. This method was also successfully applied to walrus bone as old as 3585 bp. CONCLUSIONS: LC/MS/MS analysis of bone tissue (0.2-0.3 g) provides stress and reproductive data from elusive walruses that were alive thousands of years ago. Based on physiological validations, tissue comparison, and published literature, steroid hormone concentrations measured in walrus cortical bone could represent an accumulated average around a 10-20-year time span. By investigating how stress and reproductive physiology may have changed over the past ~3000 years based on bone steroid hormone concentrations, this method will help answer how physiologically resilient walruses are to climate change in the Arctic.