skip to main content

This content will become publicly available on January 27, 2023

Title: Validation and effects of drying and prey hair on fecal hormone concentrations in spotted hyenas
Abstract As fecal steroid methods increasingly are used by researchers to monitor the physiology of captive and wild populations, we need to expand our validation protocols to test the effects of procedural variation and to identify contamination by exogenous sources of steroid hormones. Mammalian carnivore feces often contain large amounts of hair from the prey they consume, which itself may contain high concentrations of hormones. In this study, we report first a validation of two steroid hormone antibodies, corticosterone and progesterone, to determine fecal concentrations of these hormones in wild spotted hyenas (Crocuta crocuta). Next, we expand on these standard validation protocols to test two additional metrics: (i) whether hair from consumed prey or (ii) the specific drying method (oven incubation vs. lyophilization) affect steroid hormone concentrations in feces. In the first biological validation for the progesterone antibody in this species, progesterone concentrations met our expectations: (i) concentrations of plasma and fecal progesterone were lowest in immature females, higher in lactating females, and highest in pregnant females; (ii) across pregnant females, fecal progesterone concentrations were highest during late pregnancy; and (iii) among lactating females, fecal progesterone concentrations were highest after parturition. Our additional validation experiments indicated that contamination with prey more » hair and drying method are hormone-specific. Although prey hair did not release hormones into samples during storage or extraction for either hormone, its presence appeared to “dilute” progesterone (but not corticosterone) measures indirectly by increasing the dry weight of samples. In addition, fecal progesterone, but not corticosterone, values were lower for lyophilized than for incubated samples. Therefore, in addition to the standard analytical and biological validation steps, additional methodological variables need to be tested whenever we measure fecal hormone concentrations, particularly from predatory mammals. « less
Authors:
; ; ;
Editors:
Hopkins, Jack
Award ID(s):
1755089 1853934
Publication Date:
NSF-PAR ID:
10317314
Journal Name:
Journal of Mammalogy
ISSN:
0022-2372
Sponsoring Org:
National Science Foundation
More Like this
  1. Abstract Understanding reproductive physiology in mysticetes has been slowed by the lack of repeated samples from individuals. Analysis of humpback whale baleen enables retrospective hormone analysis within individuals dating back 3–5 years before death. Using this method, we investigated differences in four steroid hormones involved in reproduction and mating during confirmed pregnant and non-pregnant periods in two female humpback whales (Megaptera novaeangliae) with known reproductive histories based on sightings and necropsy data. Cortisol, corticosterone, testosterone, and estradiol concentrations were determined via enzyme immunoassay using subsamples of each baleen plate at 2 cm intervals. There were no significant differences in cortisol or corticosterone during pregnancy when compared to non-pregnancy (inter-calving interval), but there were significant differences between the two whales in average glucocorticoid concentrations, with the younger whale showing higher values overall. For testosterone, levels for the younger female peaked at parturition in one pregnancy, but also had spikes during non-pregnancy. The older female had three large spikes in testosterone, one of which was associated with parturition. Estradiol had large fluctuations in both whales but had generally lower concentrations during non-pregnancy than during pregnancy. There were peaks in estradiol before each pregnancy, possibly coinciding with ovulation, and peaks coinciding with themore »month of parturition. Both estradiol and testosterone could be useful for determining ovulation or impending birth. Using baleen to investigate retrospective steroid hormone profiles can be used for elucidating long-term patterns of physiological change during gestation. Lay summary Case studies of two pregnant humpback whales whose hormones were analyzed in baleen may illuminate when humpback whales ovulate, gestate, and give birth. These physiological metrics could assist in accurate population growth assessments and conservation of the species. This study shows that baleen hormone analysis can be a useful tool for understanding whale reproductive physiology.« less
  2. Hormones mediate physiological and behavioral changes in adults as they transition into reproduction. In this study, we characterize the circulating levels of five key hormones involved in reproduction in rock doves ( Columba livia ): corticosterone, progesterone, estradiol, testosterone, and prolactin using univariate and multivariate approaches. We show similar patterns as previous studies in the overall patterns in circulating levels of these hormones, i.e., testosterone (males) and estradiol (females) high during nest-building or egg-laying, prolactin increasing at mid-incubation and peaking at hatching (both sexes), and elevated corticosterone levels in later incubation and early nestling development. In our investigation of hormone co-variation, we find a strong correlation between prolactin and corticosterone across sampling stages and similarities in earlier (early to mid-incubation) compared to later (late incubation to nestling d9) sampling stages in males and females. Finally, we utilized experimental manipulations to simulate nest loss or altered caregiving lengths to test whether external cues, internal timing, or a combination of these factors contributed most to hormone variation. Following nest loss, we found that both males and females responded to the external cue. Males generally responded quickly following nest loss by increasing circulating testosterone, but this response was muted when nest loss occurredmore »early in reproduction. Similar treatment type, e.g., removal of eggs, clustered similarly in hormone space. These results suggest internal drivers limited male response early in reproduction to nest loss. In contrast, circulating levels of these hormones in females either did not change or decreased following nest manipulation suggesting responsiveness to external drivers, but unlike males, this result suggests that reproductive processes were decreasing.« less
  3. Abstract

    Research in captive birds and mammals has demonstrated that circadian (i.e., daily) behavioral rhythms are altered in response to increases in sex-steroid hormones. Recently, we and others have demonstrated a high degree of individual repeatability in peak (gonadotropin-releasing hormone [GnRH]-induced sex) steroid levels, and we have found that these GnRH-induced levels are highly correlated with their daily (night-time) endogenous peak. Whether or not individual variation in organization and activity of the reproductive endocrine axis is related to daily timing in wild animals is not well known. To begin to explore these possible links, we tested the hypothesis that maximal levels of the sex steroid hormone estradiol (E2) and onset of daily activity are related in a female songbird, the dark-eyed junco (Junco hyemalis). We found that females with higher levels of GnRH-induced E2 departed from their nest in the morning significantly earlier than females with lower stimulated levels. We did not observe a relationship between testosterone and this measure of onset of activity. Our findings suggest an interaction between an individual’s reproductive endocrine axis and the circadian system and variation observed in an individuals’ daily activity onset. We suggest future studies examine the relationship between maximal sex-steroid hormones andmore »timing of daily activity onset.

    « less
  4. Monitoring wildlife stress levels is essential to ensure their quality of life in captivity or in the wild. One promising method to assess the stress response is the comeasurement of glucocorticoids (GC) and dehydroepiandrosterone sulfate (DHEAS), adrenal hormones involved in the modulation of the stress response. Although noninvasive methods to measure GCs have been validated in several species, only a few studies have validated DHEAS assays. The aims of this study were (1) to describe an enzyme immunoassay (EIA) to measure DHEAS levels, (2) to validate this assay for fecal samples in gibbons and siamangs, and (3) to test hormonal stability after one freeze-thaw cycle and over time at two freezer temperatures (-20°C and -80°C). Subjects included 32 gibbons and siamangs from U.S. zoological parks. The EIA was validated analytically by parallelism and accuracy tests, and biologically by confirming a DHEAS response 1-2 days after a stressful event (accident, vaccination, or transportation) in three individuals. In addition, fecal DHEAS levels in a pregnant female were above nonpregnant/nonlactating levels and declined progressively the following parturition. The hormonal stability experiments revealed no significant changes in fecal DHEAS levels after one freeze-thaw cycle. Hormonal levels in fecal extracts were stable for 2 months,more »regardless of the storage temperature, with no significant differences between -20°C and -80°C conditions. The EIA described has high sensitivity and it is suitable for fecal DHEAS measurement in gibbons and siamangs, with a potential to be applied to other species.« less
  5. Abstract

    Glucocorticoids and glucocorticoid metabolites are increasingly used to index physiological stress in wildlife. Although feces is often abundant and can be collected noninvasively, exposure to biotic and abiotic elements may influence fecal glucocorticoid metabolite (FGM) concentrations, leading to inaccurate conclusions regarding wildlife physiological stress. Using captive snowshoe hares (Lepus americanus) and simulated environmental conditions, we evaluated how different realistic field conditions and temporal sampling constraints might influence FGM concentrations using an 11-oxoetiocholanolone-enzyme immunoassay. We quantified how fecal pellet age (i.e., 0–6 days), variable summer temperatures, and precipitation affected FGM concentrations. Fecal pellet age had a strong effect on FGM concentrations (βAge = 0.395, s.d. = 0.085; β2Age = −0.061, s.d. = 0.012), which were lowest at the beginning and end of our exposure period (e.g., meanday6 = 37.7 ng/mg) and typically highest in the middle (meanday3 = 51.8 ng/mg). The effect of fecal pellet age on FGM concentrations varied across treatments with warm-dry and cool-wet conditions resulting in more variable FGM concentrations relative to control samples. Given the confounding effects of exposure and environmental conditions, if fresh fecal pellet collection is not an option, we encourage researchers to develop a temporally consistent sampling protocol to ensure all samples are exposed to similar environmental conditions.