Validation of a novel method to create temporal records of hormone concentrations from the claws of ringed and bearded seals

Abstract

Ringed (Pusa hispida) and bearded seals (Erignathus barbatus) inhabit vast and often remote areas in the Arctic, making it difficult to obtain long-term physiological information concerning health and reproduction. These seals are experiencing climate-driven changes in their habitat that could result in physiological stress. Chronic physiological stress can lead to immunosuppression, decreased reproduction and decreased growth. Recently, keratin has become a popular matrix to measure steroid hormones, such as stress-related cortisol and reproduction-related progesterone. We developed and validated methods to extract cortisol and progesterone from the claws of adult female ringed (n = 20) and bearded (n = 3) seals using enzyme immunosorbent assays. As ringed and bearded seal claws grow, a pair of dark- and light-colored bands of keratin is deposited annually providing a guide for sampling. Two processing methods were evaluated, removal of claw material with a grinding bit or grinding followed by mechanical pulverization (102 paired samples from six claws, two each from three seals). Adding the mechanical pulverization step resulted in a 1.5-fold increase in hormone extraction. Progesterone from the proximal claw band was evaluated to biologically validate claw material as a measure of pregnancy in ringed seals (n = 14). Claws from pregnant seals had significantly higher claw progesterone concentrations than from non-pregnant seals. This suggests that the elevated progesterone associated with gestation was reflected in the claws, and that the most proximal claw band was indicative of pregnancy status at time of death. Thus, although the sample size was low and the collection dates unbalanced, this study demonstrates the potential to use claws to examine an extended time series (up to 12 yrs) of cortisol and progesterone concentrations in ringed and bearded seal claws.

Using claws to compare reproduction, stress and diet of female bearded and ringed seals in the Bering and Chukchi seas, Alaska, between 1953–1968 and 1998–2014

“…Therefore, using the ridges to identify dark bands allowed them to be consistently identified for sampling. Claw samples were processed using the same methods as in Karpovich et al (2020 ). Briefly, claws were soaked in deionized water to reveal the claw bands and, using a Dremel® tool, all dark bands were sampled once and all light bands were sampled twice (left and right halves when examining Fig.…”

“…The supernatant was stored at ≤ −20°C until hormone analysis. Progesterone and cortisol hormone assays were validated via parallelism and accuracy tests for bearded and ringed seal claws previously, as well as hormone extraction efficiency comparing two homogenization methods ( Karpovich et al 2020 ).…”

“… Bands in the claws of bearded and ringed seals are produced over time; light bands are produced in the late spring to late summer and the dark bands are produced in early fall to early spring ( McLaren 1958 ; Benjaminsen 1973 ; Karpovich et al 2020 ). The closer to the tip of the claw, the younger the age of the seal the claw band represents.…”

“…Front flipper claws from bearded and ringed seals archive hormones and stable isotopes deposited at the time of growth ( Carroll et al 2013 ; Karpovich et al 2020 ); similar to pinniped whiskers ( Karpovich et al 2018 ), baleen ( Hunt et al 2014 ) and baleen whale earplugs ( Trumble et al 2013 , 2018 ; Crain et al 2020 ). Furthermore, bearded and ringed seal claws form light and dark bands that together correspond to one year of claw growth, traditionally being used to ascertain minimum age of the individual ( McLaren 1958 ; Benjaminsen 1973 ; Ferreira et al 2011 ).…”

“…Furthermore, bearded and ringed seal claws form light and dark bands that together correspond to one year of claw growth, traditionally being used to ascertain minimum age of the individual ( McLaren 1958 ; Benjaminsen 1973 ; Ferreira et al 2011 ). Hormones and stable isotopes, once deposited, remain stored long-term in the claw for decades without degradation ( Carroll et al 2013 ; Karpovich et al 2020 ). To examine hormones and stable isotopes in samples from pre-1968 and post-1998, bearded and ringed seal claws were analyzed from female seals harvested in the Bering and Chukchi seas.…”