Longitudinal progesterone profiles in baleen from female North Atlantic right whales (Eubalaena glacialis) match known calving history

© The Author(s), 2016. This article is distributed under the terms of the Creative Commons Attribution License. The definitive version was published in Conservation Physiology 4 (2016): cow014, doi:10.1093/conphys/cow014.Reproduction of mysticete whales is difficult to monitor, and basic parameters, such as pregnancy rate and inter-calving interval, remain unknown for many populations. We hypothesized that baleen plates (keratinous strips that grow downward from the palate of mysticete whales) might record previous pregnancies, in the form of high-progesterone regions in the sections of baleen that grew while the whale was pregnant. To test this hypothesis, longitudinal baleen progesterone profiles from two adult female North Atlantic right whales (Eubalaena glacialis) that died as a result of ship strike were compared with dates of known pregnancies inferred from calf sightings and post-mortem data. We sampled a full-length baleen plate from each female at 4 cm intervals from base (newest baleen) to tip (oldest baleen), each interval representing ∼60 days of baleen growth, with high-progesterone areas then sampled at 2 or 1 cm intervals. Pulverized baleen powder was assayed for progesterone using enzyme immunoassay. The date of growth of each sampling location on the baleen plate was estimated based on the distance from the base of the plate and baleen growth rates derived from annual cycles of stable isotope ratios. Baleen progesterone profiles from both whales showed dramatic elevations (two orders of magnitude higher than baseline) in areas corresponding to known pregnancies. Baleen hormone analysis shows great potential for estimation of recent reproductive history, inter-calving interval and general reproductive biology in this species and, possibly, in other mysticete whales.This work was supported by the Eppley Foundation for Research, the National Oceanographic and Atmospheric Administration Marine Mammal Health and Stranding Program and the Woods Hole Oceanographic Institution Ocean Life Institute

Multi-year longitudinal profiles of cortisol and corticosterone recovered from baleen of North Atlantic right whales (Eubalaena glacialis)

© The Author(s), 2017. This is the author's version of the work. It is posted here under a nonexclusive, irrevocable, paid-up, worldwide license granted to WHOI. It is made available for personal use, not for redistribution. The definitive version was published in General and Comparative Endocrinology 254 (2017): 50-59, doi:10.1016/j.ygcen.2017.09.009.Research into stress physiology of mysticete whales has been hampered by difficulty in obtaining repeated physiological samples from individuals over time. We investigated whether multi-year longitudinal records of glucocorticoids can be reconstructed from serial sampling along full-length baleen plates (representing ~10 years of baleen growth), using baleen recovered from two female North Atlantic right whales (Eubalaena glacialis) of known reproductive history. Cortisol and corticosterone were quantified with immunoassay of subsamples taken every 4 cm (representing ~60 d time intervals) along a full-length baleen plate from each female. In both whales, corticosterone was significantly elevated during known pregnancies (inferred from calf sightings and necropsy data) as compared to intercalving intervals; cortisol was significantly elevated during pregnancies in one female but not the other. Within intercalving intervals, corticosterone was significantly elevated during the first year (lactation year) and/or the second year (post-lactation year) as compared to later years of the intercalving interval, while cortisol showed more variable patterns. Cortisol occasionally showed brief high elevations (“spikes”) not paralleled by corticosterone, suggesting that the two glucocorticoids might be differentially responsive to certain stressors. Generally, immunoreactive corticosterone was present in higher concentration in baleen than immunoreactive cortisol; corticosterone:cortisol ratio was usually >4 and was highly variable in both individuals. Further investigation of baleen cortisol and corticosterone profiles could prove fruitful for elucidating long-term, multi-year patterns in stress physiology of large whales, determined retrospectively from stranded or archived specimens.This work was supported by the Eppley Foundation for Research, the NOAA Marine Mammal Health and Stranding Program, the Woods Hole Oceanographic Institution Ocean Life Institute, and the New England Aquarium

Fecal glucocorticoids and anthropogenic injury and mortality in North Atlantic right whales Eubalaena glacialis

© The Author(s), 2017. This article is distributed under the terms of the Creative Commons Attribution License. The definitive version was published in Endangered Species Research 34 (2017): 417-429, doi:10.3354/esr00866.As human impacts on marine ecosystems escalate, there is increasing interest in quantifying sub-lethal physiological and pathological responses of marine mammals. Glucocorticoid hormones are commonly used to assess stress responses to anthropogenic factors in wildlife. While obtaining blood samples to measure circulating hormones is not currently feasible for free-swimming large whales, immunoassay of fecal glucocorticoid metabolites (fGCs) has been validated for North Atlantic right whales Eubalaena glacialis (NARW). Using a general linear model, we compared fGC concentrations in right whales chronically entangled in fishing gear (n = 6) or live-stranded (n = 1), with right whales quickly killed by vessels (n = 5) and healthy right whales (n = 113) to characterize fGC responses to acute vs. chronic stressors. fGCs in entangled whales (mean ± SE: 1856.4 ± 1644.9 ng g-1) and the stranded whale (5740.7 ng g-1) were significantly higher than in whales killed by vessels (46.2 ± 19.2 ng g-1) and healthy whales (51.7 ± 8.7 ng g-1). Paired feces and serum collected from the live-stranded right whale provided comparison of fGCs in 2 matrices in a chronically stressed whale. Serum cortisol and corticosterone in this whale (50.0 and 29.0 ng ml-1, respectively) were much higher than values reported in other cetaceans, in concordance with extremely elevated fGCs. Meaningful patterns in fGC concentration related to acute vs. chronic impacts persisted despite potential for bacterial degradation of hormone metabolites in dead whales. These results provide biological validation for using fGCs as a biomarker of chronic stress in NARWs.This research was funded by the NOAA/NMFS, Office of Naval Research Marine Mammals and Biology Program, Northeast Consortium, Island Foundation, Irving Oil, NEAq Internal Research Fund, Prescott Grant NA08NMF4390590, and NOAA CINAR Cooperative Agreement NA09OAR4320129

Overcoming the challenges of studying conservation physiology in large whales : a review of available methods

Large whales are subjected to a variety of conservation pressures that could be better monitored and managed if physiological information could be gathered readily from free-swimming whales. However, traditional approaches to studying physiology have been impractical for large whales, because there is no routine method for capture of the largest species and there is presently no practical method of obtaining blood samples from free-swimming whales. We review the currently available techniques for gathering physiological information on large whales using a variety of non-lethal and minimally invasive (or non-invasive) sample matrices. We focus on methods that should produce information relevant to conservation physiology, e.g. measures relevant to stress physiology, reproductive status, nutritional status, immune response, health, and disease. The following four types of samples are discussed: faecal samples, respiratory samples (‘blow’), skin/blubber samples, and photographs. Faecal samples have historically been used for diet analysis but increasingly are also used for hormonal analyses, as well as for assessment of exposure to toxins, pollutants, and parasites. Blow samples contain many hormones as well as respiratory microbes, a diverse array of metabolites, and a variety of immune-related substances. Biopsy dart samples are widely used for genetic, contaminant, and fatty-acid analyses and are now being used for endocrine studies along with proteomic and transcriptomic approaches. Photographic analyses have benefited from recently developed quantitative techniques allowing assessment of skin condition, ectoparasite load, and nutritional status, along with wounds and scars from ship strikes and fishing gear entanglement. Field application of these techniques has the potential to improve our understanding of the physiology of large whales greatly, better enabling assessment of the relative impacts of many anthropogenic and ecological pressures.Publisher PDFPeer reviewe

Estimating the effects of stressors on the health, survival and reproduction of a critically endangered, long-lived species

Funding: Office of Naval Research (Grant Number(s): N000142012697, N000142112096); Strategic Environmental Research and Development Program (Grant Number(s): RC20-1097, RC20-7188, RC21-3091).Quantifying the cumulative effects of stressors on individuals and populations can inform the development of effective management and conservation strategies. We developed a Bayesian state–space model to assess the effects of multiple stressors on individual survival and reproduction. In the model, stressor effects on vital rates are mediated by changes in underlying health, allowing for the comparison of effect sizes while accounting for intrinsic factors that might affect an individual's vulnerability and resilience. We applied the model to a 50-year dataset of sightings, calving events and stressor exposure of critically endangered North Atlantic right whales Eubalaena glacialis. The viability of this population is threatened by a complex set of stressors, including vessel strikes, entanglement in fishing gear and fluctuating prey availability. We estimated that blunt and deep vessel strike injuries and severe entanglement injuries had the largest effect on the health of exposed individuals, reinforcing the urgent need for mitigation measures. Prey abundance had a smaller but protracted effect on health across individuals, and estimated long-term trends in survival and reproduction followed the trend of the prey index, highlighting that long-term ecosystem-based management strategies are also required. Our approach can be applied to quantify the effects of multiple stressors on any long-lived species where suitable indicators of health and long-term monitoring data are available.Publisher PDFPeer reviewe

Understanding the combined effects of multiple stressors : a new perspective on a longstanding challenge

This work was supported by the Office of Naval Research [grant numbers N000142012697, N000142112096]; and the Strategic Environmental Research and Development Program [grant numbers RC20-1097, RC20-7188, RC21-3091].Wildlife populations and their habitats are exposed to an expanding diversity and intensity of stressors caused by human activities, within the broader context of natural processes and increasing pressure from climate change. Estimating how these multiple stressors affect individuals, populations, and ecosystems is thus of growing importance. However, their combined effects often cannot be predicted reliably from the individual effects of each stressor, and we lack the mechanistic understanding and analytical tools to predict their joint outcomes. We review the science of multiple stressors and present a conceptual framework that captures and reconciles the variety of existing approaches for assessing combined effects. Specifically, we show that all approaches lie along a spectrum, reflecting increasing assumptions about the mechanisms that regulate the action of single stressors and their combined effects. An emphasis on mechanisms improves analytical precision and predictive power but could introduce bias if the underlying assumptions are incorrect. A purely empirical approach has less risk of bias but requires adequate data on the effects of the full range of anticipated combinations of stressor types and magnitudes. We illustrate how this spectrum can be formalised into specific analytical methods, using an example of North Atlantic right whales feeding on limited prey resources while simultaneously being affected by entanglement in fishing gear. In practice, case-specific management needs and data availability will guide the exploration of the stressor combinations of interest and the selection of a suitable trade-off between precision and bias. We argue that the primary goal for adaptive management should be to identify the most practical and effective ways to remove or reduce specific combinations of stressors, bringing the risk of adverse impacts on populations and ecosystems below acceptable thresholds.Publisher PDFPeer reviewe

Decreasing body size is associated with reduced calving probability in critically endangered North Atlantic right whales

Funding: This work was supported by the Office of Naval Research (grant nos. N000142012697 and N000142112096) and the Strategic Environmental Research and Development Program (grant nos. RC20-1097, RC20-7188 and RC21-3091). Photogrammetry was supported by NOAA grant no. NA14OAR4320158 to Woods Hole Oceanographic Institution, and by NOAA's Southwest Fisheries Science Center.Body size is key to many life-history processes, including reproduction. Across species, climate change and other stressors have caused reductions in the body size to which animals can grow, called asymptotic size, with consequences for demography. A reduction in mean asymptotic length was documented for critically endangered North Atlantic right whales, in parallel with declines in health and vital rates resulting from human activities and environmental changes. Here, we tested whether smaller body size was associated with lower reproductive output, using a state-space model for individual health, survival and reproduction that quantifies the mechanistic links between these processes. Body size (as represented by the cube of length) was strongly associated with a female's calving probability at each reproductive opportunity. This relationship explained 62% of the variation in calving among reproductive females, along with their decreasing health (20%). The effects of decreasing mean body size on reproductive performance are another concerning indication of the worsening prospects for this species and many others affected by environmental change, requiring a focus of conservation and management interventions on improving conditions that affect reproduction as well as reducing mortality.Peer reviewe

A tripartite paternally methylated region within the Gpr1-Zdbf2 imprinted domain on mouse chromosome 1 identified by meDIP-on-chip

The parent-of-origin specific expression of imprinted genes relies on DNA methylation of CpG-dinucleotides at differentially methylated regions (DMRs) during gametogenesis. To date, four paternally methylated DMRs have been identified in screens based on conventional approaches. These DMRs are linked to the imprinted genes H19, Gtl2 (IG-DMR), Rasgrf1 and, most recently, Zdbf2 which encodes zinc finger, DBF-type containing 2. In this study, we applied a novel methylated-DNA immunoprecipitation-on-chip (meDIP-on-chip) method to genomic DNA from mouse parthenogenetic- and androgenetic-derived stem cells and sperm and identified 458 putative DMRs. This included the majority of known DMRs. We further characterized the paternally methylated Zdbf2/ZDBF2 DMR. In mice, this extensive germ line DMR spanned 16 kb and possessed an unusual tripartite structure. Methylation was dependent on DNA methyltransferase 3a (Dnmt3a), similar to H19 DMR and IG-DMR. In both humans and mice, the adjacent gene, Gpr1/GPR1, which encodes a G-protein-coupled receptor 1 protein with transmembrane domain, was also imprinted and paternally expressed. The Gpr1-Zdbf2 domain was most similar to the Rasgrf1 domain as both DNA methylation and the actively expressed allele were in cis on the paternal chromosome. This work demonstrates the effectiveness of meDIP-on-chip as a technique for identifying DMRs