Abstract: After 14C-labelled cortisol infusion in ponies and pigs, faecal samples were collected. Extraction of 0.5 g faeces with 5 ml 80–90 % methanol yielded the highest radioactivity in the supernatant. Most of the metabolites were ether soluble. After high performance liquid chromatography (HPLC), the presence of immunoreactive metabolites was demonstrated by measuring each HPLC fraction using enzyme immunoassays for cortisol, corticosterone and 11-oxoaetiocholanolone. Only the assay for 11-oxoaetiocholanolone revealed peaks with co-eluting radioactivity. For biological validation of the test system, adrenocorticotrophic hormone (ACTH) and dexamethasone were injected intravenously successively in both species (n = 6). Cortisol concentration in blood and the 11-oxoaetiocholanolone immunoreactive substances in faeces were determined. In horse faeces, basal values of 2.3–35.2 nmol/kg were measured. After ACTH administration, an increase (more than 200 % above basal values) of these metabolites was seen about 1 day after ACTH administration. After dexamethasone injection the levels decreased, reaching minimum concentrations 2 days after administration. In pigs, an increase in these metabolites was measured in only three animals after ACTH; dexamethasone did not cause a decrease. The stability of the samples after defecation was tested by storing samples from cows, horses and pigs at room temperature. It was shown that there was a significant increase in the concentration of measured cortisol metabolites in bovine, equine and porcine faeces after storage for 1 h, 4 h and 24 h, respectively. In frozen samples this effect was diminished after thawing samples at 40°C; thawing the samples at 95°C prevented an increase in immunoreactive substances.

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: Abstract Given the social and territorial features described in feral cats, it is commonly assumed that life in multi-cat households is stressful for domestic cats and suggested that cats kept as single pets are likely to have better welfare. On the other hand, it has been hypothesized that under high densities cats can organize themselves socially thus preventing stress when spatial dispersion is unavailable. This study was aimed at comparing the general arousal underpinning emotional distress in single housed cats and in cats from multi-cat households (2 and 3–4 cats) on the basis of fecal glucocorticoid metabolites (GCM) measured via enzyme immunoassay (EIA). GCM did not significantly vary as a function of living style (single, double or group-housing); highly stressed individuals were equally likely in the three groups. Young cats in multi-cat households had lower GCM, and overall cats that tolerate (as opposed to dislike) petting by the owners tended to have higher GCM levels. Other environmental aspects within cat houses (e.g. relationship with humans, resource availability) may play a more important role in day to day feline arousal levels than the number of cats per se.

Abstract: Animal welfare is of increasing importance and absence of chronic stress is one of its prerequisites. During stress, various endocrine responses are involved to improve the fitness of the individual. The front-line hormones to overcome stressful situations are the glucocorticoids and catecholamines. These hormones are determined as a parameter of adrenal activity and thus of disturbance. The concentration of glucocorticoids (or their metabolites) can be measured in various body fluids or excreta. Above all, fecal samples offer the advantage that they can be easily collected and this procedure is feedback free. Recently, enzyme immunoassays (EIA) have been developed and successfully tested, to enable the measurement of groups of cortisol metabolites in animal feces. The determination of these metabolites in fecal samples is a practical method to monitor glucocorticoid production.

Abstract: Non-invasive methods enable stress evaluation through measuring fecal glucocorticoid metabolites (FGMs), and immunoglobulin A (IgA) in the feces avoiding stressful blood drawing or stressful restraining of animals in the field. However, FGMs and IgA are mostly analysed in freshly frozen samples, which is difficult when fresh samples cannot be frozen immediately or frozen samples cannot be stored or transported. Good results were also derived from air-dried fecal samples, which are hampered by unstable air humidity in the field. These difficulties may be overcome, when drying of samples could be induced with colorless silica gel (SiO2) granules in a secure set-up, such as an air tight tube. We determined the speed of drying 1.5 g of a fresh fecal sample from six horses on air and on silica gel. Furthermore, FGMs and IgA were analysed in differently stored subsamples from 12 horses: in frozen fecal samples, in air- or silica gel-dried samples stored for 1 day and for 7 days, and in wet fecal samples kept in a tube at room temperature for 7 days. FGM levels remained stable in feces dried on air or on silica gel for 7 days, whereas IgA quantities showed a significant loss. Under field conditions, when freezing or transporting the frozen samples is not possible and humidity hampers air drying, drying samples on silica gel in air tight tubes appears to be very helpful and reliable for analysing FGMs.