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 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.

中文翻译：

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干燥和捕食毛发对斑点鬣狗粪便激素浓度的验证和影响

随着研究人员越来越多地使用粪便类固醇方法来监测圈养和野生种群的生理机能，我们需要扩展我们的验证方案以测试程序变化的影响并确定外源类固醇激素的污染。哺乳动物食肉动物的粪便通常含有大量来自它们所吃猎物的毛发，这些毛发本身可能含有高浓度的激素。在这项研究中，我们首先报告了对两种类固醇激素抗体皮质酮和黄体酮的验证，以确定野生斑点鬣狗 (Crocuta crocuta) 中这些激素的粪便浓度。接下来，我们扩展了这些标准验证协议以测试两个额外的指标：（i）是否来自消耗的猎物或（ii）特定的干燥方法（烤箱孵化与干燥）冻干）影响粪便中的类固醇激素浓度。在该物种中孕酮抗体的首次生物学验证中，孕酮浓度符合我们的预期：（i）血浆和粪便中的孕酮浓度在未成熟雌性中最低，在哺乳期雌性中较高，在怀孕雌性中最高；(ii) 在怀孕女性中，孕晚期的粪便孕酮浓度最高；(iii) 在哺乳期女性中，分娩后粪便中的孕酮浓度最高。我们的额外验证实验表明，猎物毛发的污染和干燥方法是激素特异性的。尽管猎物毛发在储存或提取任何一种激素的过程中都不会向样品中释放激素，它的存在似乎通过增加样品的干重间接“稀释”孕酮（但不是皮质酮）测量值。此外，冻干样品的粪便黄体酮值（而不是皮质酮）低于孵育样品。因此，除了标准的分析和生物学验证步骤之外，每当我们测量粪便激素浓度时，都需要测试其他方法变量，尤其是来自捕食性哺乳动物的粪便激素浓度。