Abstract: Non-invasive faecal oestrogen and progesterone metabolite evaluations are well established approaches for monitoring reproductive function in a variety of mammalian species. The route of excretion of steroid hormone metabolites varies considerably among species, and also between steroids within the same species. Steroid concentrations in faeces exhibit a similar pattern to those in plasma, but have a lag time, which depending upon the species, can be from 12 h to more than 2 days. Faecal steroid metabolites in mammals are mainly unconjugated compounds. Faecal oestrogens consist predominantly of oestrone and/or oestradiol-17α or -17β. Therefore, specific oestrogen antibodies or antibodies against total oestrogens can be used for their determination. Progesterone is metabolised to several 5α- or 5β-reduced pregnanediones and hydroxylated pregnanes prior to its faecal excretion. Therefore, relevant antibodies for their determination show considerable cross-reactivities with several pregnane metabolites, whereas specific progesterone antibodies are less suitable. Faecal oestrogen evaluations have been used as reliable indicators of pregnancy in several ungulate and some primate species. They have also been used to determine the preovulatory period in carnivores, corpus luteum activity in New World primates, and to diagnose cryptorchidism in horses. Faecal progesterone metabolite analysis has been successfully used for monitoring corpus luteum function and pregnancy, abortion, seasonality and treatment therapies in an ever expanding list of species.

Abstract: The ability of horses to learn and remember new tasks is fundamentally important for their use by humans. Fearfulness may, however, interfere with learning, because stimuli in the environment can overshadow signals from the rider or handler. In addition, prolonged high levels of stress hormones can affect neurons within the hippocampus; a brain region central to learning and memory. In a series of experiments, we aimed to investigate the link between performance in two learning tests, the baseline level of stress hormones, measured as faecal cortisol metabolites (FCM), fearfulness, and social rank. Twenty-five geldings (2 or 3 years old) pastured in one group were included in the study. The learning tests were performed by professional trainers and included a number of predefined stages during which the horses were gradually trained to perform exercises, using either negative (NR) or positive reinforcement (PR). Each of the learning tests lasted 3 days; 7min/horse/day. The NR test was repeated in a novel environment. Performance, measured as final stage in the training programme, and heart rate (HR) were recorded. Faeces were collected on four separate days where the horses had been undisturbed at pasture for 48h. Social rank was determined through observations of social interactions during feeding. The fear test was a novel object test during which behaviour and HR were recorded. Performance in the NR and PR learning tests did not correlate. In the NR test, there was a significant, negative correlation between performance and HR in the novel environment (rS=-0.66, P<0.001, i.e. nervous horses had reduced performance), whereas there was no such correlation in the home environment (both NR and PR). Behavioural reactions in the fear test correlated significantly with performance in the NR test in the novel environment (e.g. object alertness and final stage: rS=-0.43, P=0.04), suggesting that performance under unfamiliar, stressful conditions may be predicted by behavioural responses in a fear test. There was a negative correlation between social rank and baseline stress hormones (rS=-0.43, P=0.04), i.e. high rank corresponded to low FCM concentrations, whereas neither rank nor FCM correlated with fearfulness or learning performance. We conclude that performance under stressful conditions is affected by activation of the sympathetic nervous system during training and related to behavioural responses in a standardised fear test. Learning performance in the home environment, however, appears unrelated to fearfulness, social rank and baseline FCM levels.

Abstract: The aim of this comparative study was to gain more information about the excretion of steroid hormones in farm animals. This should help to establish or improve non-invasive steroid monitoring procedures, especially in zoo and wildlife animals. Over a period of 4 h the 14C-steroid hormones (3.7 MBq) progesterone (three females), testosterone (three males), cortisol and oestrone (two males, two females) were infused intravenously in sheep, ponies and pigs. Faeces were collected immediately after defecation. Urine was sampled via a permanent catheter in females and after spontaneous urination in males. A total of 88 +/- 10% (mean +/- SD) of the administered radioactivity was recovered. Considerable interspecies differences were measured both in the amounts of steroid metabolites excreted via faeces or urine and the time course of excretion. Progesterone and oestrone in ewes, and progesterone in mares were excreted mainly in the faeces (over 75%). The primary route of excretion of all other 14C-steroids was via the urine but to a different extent. In general, sheep showed the highest degree of faecal excretion and pigs the least. The highest radioactivity in urine (per mmol creatinine) was observed during the infusion or in one of the next two samples thereafter, whereas in faeces it was measured about 12 h (sheep), 24 h (ponies) or 48 h (pigs) after the end of the infusion. Thereafter the radioactivity declined and reached background levels within 2-3 weeks. In faeces, steroid metabolites were present mainly in an unconjugated form, but in blood and urine as conjugates. Mean retention time of faecal radioactivity suggested that the passage rate of digesta (duodenum to rectum) played an important role in the time course of the excretion of steroids. The information derived from this investigation could improve the precision of sampling as well as the extraction of steroids from the faeces. Furthermore, the study demonstrates that it should be possible to establish methods for measuring faecal androgen and cortisol metabolites for assessing male reproductive endocrinology and stress in animals.

Abstract: Stress generated by humans on wildlife by continuous development of outdoor recreational activities is of increasing concern for biodiversity conservation. Human disturbance often adds to other negative impact factors affecting the dynamics of vulnerable populations. It is not known to which extent the rapidly spreading free-riding snow sports actually elicit detrimental stress (allostatic overload) upon wildlife, nor what the potential associated fitness and survival costs are. Using a non-invasive technique, we evaluated the physiological stress response induced by free-riding snow sports on a declining bird species of Alpine ecosystems. The results of a field experiment in which radiomonitored black grouse (Tetrao tetrix) were actively flushed from their snow burrows once a day during four consecutive days showed an increase in the concentration of faecal stress hormone (corticosterone) metabolites after disturbance. A large-scale comparative analysis across the southwestern Swiss Alps indicated that birds had higher levels of these metabolites in human-disturbed versus undisturbed habitats. Disturbance by snow sport free-riders appears to elevate stress, which potentially represents a new serious threat for wildlife. The fitness and survival costs of allostatic adjustments have yet to be estimated.

Abstract: On Days 26, 28, and 30 after AI, ultrasonographic pregnancy diagnoses were performed on 207 gilts and sows by using a 3.5 MHz linear-array transducer. Fecal samples were taken from the rectum after each ultrasonographic examination, and the concentrations of unconjugated estrogens in selected samples (n = 73) were measured by RIA. Fecal unconjugated estrogen concentration of 11.7 ng/g feces or higher was indicative of pregnancy. The overall sensitivity and specificity of the ultrasonographic test was 99% for farrowing sows and 73.1% for nonfarrowing sows. With one exception, sows with a false negative diagnosis by ultrasonography on Day 26 were correctly diagnosed pregnant by elevated fecal unconjugated estrogens or repeated ultrasonographic examinations on Days 28 or 30. Return to estrus around the sampling period may cause false positive results in the unconjugated estrogen assay, while early embryonic mortality can result in false positive diagnoses in both the ultrasonographic test and estrogen assay. Although there was a positive correlation between the concentrations of unconjugated estrogens in the feces and litter size at farrowing in the selected sows, it seems very unlikely that fecal estrogens can provide an accurate tool for predicting litter size.