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Berger, A. (2015). Evaluation of living conditions in free running animals by chronobiological analysis of continuously recorded behavioural data. In Proceedings of the 3. International Equine Science Meeting.
Abstract: We developed a biorhythmical method to assess behaviour patterns and to evaluate living conditions of animals. All kinds of continuous and equidistant long-term recordings of behaviour are suitable for this method. As simple behavioural parameters, such as motor activity, can be conveniently recorded by telemetry from wild animals now, it is possible to investigate stressors by analysing its biorhythmic structure. It is the purpose of this report to describe the basic idea, and the procedure, and to give some examples of application measured on Przewalski horses in an Semireserve.
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Gorgasser I., Tichy A., & Palme R. (2007). Faecal cortisol metabolites in Quarter Horses during initial training under field conditions[Messung der Kortisolmetaboliten im Pferdekot während der Grundausbildung von 2jährigen Quarter Horses]. Wien. Tierärztl. Mschr. – Vet. Med. Austria, 94, 226–230.
Abstract: The first month of training of a young horse is suspected to be stressful, but the endocrine responses to initial training are unknown. Therefore in our study a total of 40 Quarter Horses (QH), all at the age of almost 2 years, were followed during the first 30 days of their training. During this time faecal samples were collected twice daily and faecal cortisol metabolites (FCM) were measured. Baseline values of FCM ranged between 1.3 and 20.1 (median: 6.7) ng/g faeces. No differences in FCM values between days of training were found. Mares showed the highest values. Significant diurnal variations were observed in mares (p=0.035) and stallions (p=0.003), but not in geldings (p=0.282). As in this study adrenocortical activity was not increased during initial training, horses seem to cope very well with this new situation. The results of our large-scale study provide basic physiological data about initial training. This gives additional input in an emotional debate about animal welfare aspects of first time handling and training of horses.
Abbreviations: 11,17-DOA = 11,17-dioxoandrostanes; EIA = Enyzme Immunoassay; FCM = faecal cortisol metabolites; GC = glucocorticoids; HPA-axis = hypothalamic-pituitary-adrenocortical-axis; QH = Quarter Horses
[Das Einreiten eines jungen Pferdes steht unter Verdacht belastend zu sein. Bisher gibt es aber keine Veröffentlichungen über endokrine Vorgänge während dieser Phase. Mit der vorliegenden Studie wurde überprüft, ob Pferde aufgrund physischer und psychischer Belastungen während des Trainings höhere Konzentrationen an Kortisolmetaboliten im Kot (FCM) aufweisen. Es wurden dazu 40 Quarter Horses im Alter von 2 Jahren während der ersten 30 Tage der Grundausbildung des Westernreitens beobachtet und ihre FCM Werte gemessen. Während dieser Zeitspanne wurden täglich morgens und abends Kotproben der Pferde genommen. Die Basalwerte der FCM Konzentration variierten zwischen 1,3 und 20,1 (Median: 6,7) ng/g Kot, wobei Stuten die höchsten Werte hatten. Signifikante Unterschiede während der einzelnen Trainingstage konnten nicht festgestellt werden. In der Tagesrhythmik wurden signifikante Unterschiede bei Stuten (p=0,035) und bei Hengsten (p=0,003), jedoch nicht bei Wallachen (p=0,282) ermittelt. In dieser Studie konnte keine erhöhte Aktivität der Nebennierenrinde im Verlauf der Grundausbildung eines Pferdes im Westernreitstil festgestellt werden. Das legt nahe, dass Pferde mit dieser neuen, zeitlich kurz andauernden Situationen gut zurechtkommen. Unsere Studie wurde an einer großen Anzahl von Tieren unter Feldbedingungen durchgeführt. Sie bietet daher eine gute Datenbasis über Belastungen während des Einreitens. Damit liefert sie einen zusätzlichen Beitrag zu einer mitunter emotional geführten Debatte über tierschutzrelevante Aspekte bei der Grundausbildung von Pferden.]
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Palme, R. (2019). Non-invasive measurement of glucocorticoids: Advances and problems. Physiol. Behav., 199, 229–243.
Abstract: Glucocorticoids (GCs; i.e. cortisol/corticosterone) are a central component of the stress response and thus their measurement is frequently used to evaluate the impact of stressful situations. Their metabolites from faeces of various animal species are more and more taken as a non-invasive aid to assess GC release and thus adrenocortical activity. The current literature review includes an extensive collection (1327 papers) and evaluation (see also Supplementary Tables) of the literature on faecal cortisol/corticosterone metabolite (FCM) analysis published to date. It aims at giving reference for researchers interested in implementing FCM analysis into their study or seeking to improve such methods by providing background knowledge on GC metabolism and excretion, conveying insights into methodological issues and stating caveats of FCM analysis and by highlighting prerequisites for and some examples of a successful application of such methods. Collecting faecal samples and analysing FCMs may appear simple and straightforward, but researchers have to select and apply methods correctly. They also need to be aware of the many pitfalls and potentially confounding factors and, last but not least, have to carefully interpret results. Applied properly, measurement of FCMs is a powerful non-invasive tool in a variety of research areas, such as (stress) biology, ethology, ecology, animal conservation and welfare, but also biomedicine.
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Palme, R., Touma, C., Arias, N., Dominchin, M.N., & Lepschy, M. (2013). Steroid extraction: Get the best out of faecal samples. Wien Tierärztl Monat – Vet Med Austria, 100, 238–246.
Abstract: Faecal steroid hormone metabolites are becoming increasingly popular as parameters for reproductive functions and stress. The extraction of the steroids from the faecal matrix represents the initial step before quantification can be performed. The steroid metabolites present in the faecal matrix are of varying polarity and composition, so selection of a proper extraction procedure is essential. There have been some studies to address this complex but often neglected point. Radiolabelled steroids (e.g. cortisol or progesterone) have frequently been added to faecal samples to estimate the efficiency of the extraction procedures used. However, native, unmetabolized steroids are normally not present in the faeces and therefore the results are artifi- cial and do not accurately reflect the actual recoveries of the substances of interest. In this respect, recovery experiments based on faecal samples from radiometabolism studies are more informative. In these samples, the metabolite content accurately reflects the mixture of metabolites present in the given species. As a result, it is possible to evaluate different extraction methods for use with faecal samples. We present studies on sheep, horses, pigs, hares and dogs that utilized samples containing naturally metabolized, 14C-labelled steroids. We recommend extracting faecal steroids by simply suspending the faeces in a high percentage of a primary alcohol (for glucocorticoid metabolites 80% aqueous methanol proved best suited for virtually all mammalian species tested so far). Not only does the procedure significantly increase the total amount of recovered radioactivity, it also increases the percentage of unconjugated metabolites, which are more likely to be recognized by the antibodies used in various immunoassays. The advantages of this extraction procedure are clear: it is very easy to use (no evaporation step is needed), it yields high recoveries and variation based on the extraction procedure is reduced to a minimum.
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Ramos, D., Reche-Junior, A., Fragoso, P. L., Palme, R., Yanasse, N. K., Gouvêa, V. R., et al. (2013). Are cats (Felis catus) from multi-cat households more stressed? Evidence from assessment of fecal glucocorticoid metabolite analysis. Physiol. Behav., 122, 72–75.
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.
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Schwarzenberger, F., Mostl, E., Palme, R., & Bamberg, E. (1996). Faecal steroid analysis for non-invasive monitoring of reproductive status in farm, wild and zoo animals. Animal Reproduction: Research and Practice, 42(1-4), 515–526.
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.
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Young, T., Creighton, E., Smith, T., & Hosie, C. (2012). A novel scale of behavioural indicators of stress for use with domestic horses. Applied Animal Behaviour Science, 140(1–2), 33–43.
Abstract: Behaviour scores (BS) offer non-invasive, objective and easy to use ways of assessing welfare in animals. Their development has, however, largely focused on behavioural reactions to stressful events (often induced), and little use of physiological measures has been made to underpin and validate the behavioural measures. This study aimed to develop a physiologically validated scale of behavioural indicators of stress for the purpose of welfare assessment in stabled domestic horses. To achieve this, behavioural and physiological data were collected from 32 horses that underwent routine husbandry procedures. Principal component analysis (PCA) of the behavioural and physiological data revealed three meaningful components that were used as the basis of the scale. Analysis of video clips of the horses’ responses to the husbandry procedures was undertaken by a panel of equestrian industry professionals using a free choice profiling (FCP) methodology. These results were added to the scale along with key definitions from relevant literature. Salivary cortisol levels were significantly correlated with the BS confirming the scale was meaningful and reflected physiological stress. The scale offers an easy to use ‘tool’ for rapid, reliable non-invasive welfare assessment in horses, and reduces the need for potentially invasive physiological measures.
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