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Author Flauger, B.; Krueger, K.; Gerhards, H.; Möstl, E.
Title Simplified method to measure glucocorticoid metabolites in faeces of horses Type Journal Article
Year 2010 Publication Veterinary Research Communications Abbreviated Journal (down) Vet Res Comm
Volume 34 Issue 2 Pages 185-195
Keywords ACTH challenge; enzyme immunoassay; stress behaviour; cortisol
Abstract Glucocorticoids or their metabolites can be measured in several body fluids or excreta, including plasma, saliva, urine and faeces. In recent years the measurement of glucocorticoid metabolites (GCMs) in faeces has gained increasing attention, because of its suitability for wild populations. In horses, however, the group-specific enzyme immunoassay described so far has a limited racticability due to its complex extraction procedure. Therefore, we tested the applicability of

other enzyme immunoassays for glucocorticoid metabolites. The present study clearly proved that an enzyme immunoassay (EIA) for 11-oxoetiocholanolone using 11-oxoetiocholanolone-17-CMO: BSA (3α,11-oxo-A EIA) as antigen showed high amounts of immunoreactive substances. Therefore it was possible to use just a small amount of the supernatant of a methanolic suspension of faeces. The results

correlated well with the already described method for measuring GCMs in horse faeces, i.e. analysing the samples with an EIA after a two step clean up procedure of the samples (Merl et al. 2000). In addition, the 3α,11-oxo-A EIA has the advantage of providing a bigger difference between baseline values and peak values after ACTH stimulation. The new assay increased the accuracy of the test,

lowered the expenses per sample, and storing samples at room temperature after collection was less critical than with other assays investigated in our study. This is a big advantage both in the field of wildlife management of equids and in the field of equestrian sports and it shows the importance of choosing an assay which is in good accordance with the metabolites excreted in a given species.
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Call Number Equine Behaviour @ team @ Serial 5073
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Author Wyss, C.
Title Does housing in a „social box“ change faecal cortisol metabolites concentration in stallions? Type Conference Article
Year 2015 Publication Proceedings of the 3. International Equine Science Meeting Abbreviated Journal (down) Proc. 3. Int. Equine. Sci. Mtg
Volume Issue Pages
Keywords housing system, stallions, social interaction, stress, faecal cortisol metabolites
Abstract In order to improve the housing conditions of stallions in individual boxes by offering a possibility to have more social contact, the Swiss national stud farm tested a new box system for horses, allowing increased physical contact with the neighbouring stallion. The aim of this part of the study was to investigate whether this type of housing system (named “social box”) potentially induces a change in stress reactions in stallions compared to conventional boxes. Therefore faecal cortisol metabolite (FCM) concentration was measured as a non-invasive parameter to assess endocrine responses related to this new environment.

Four groups each consisting of eight adult Freiberger breeding stallions were included in the test design. Every stallion spent three weeks in a conventional box and in a social box respectively (cross-over design). The conventional box consisted of a separation wall with a lower opaque part and an upper part with vertical barriers (5 cm between barriers), allowing visual and olfactory contact but strongly limiting tactile contact. The separating wall of the social box consisted of two lateral sections, one part being opaque to the ceiling and the second part consisting of vertical barriers (30 cm between barriers), allowing the horse to have physical contact with its neighbour or to avoid it.

In horses, FCM concentration reflects an average level of circulating cortisol over a period of approximatively 24h. Faecal samples were collected the day following integration in social / conventional boxes, reflecting the potential stress induced by increased social interactions during the integration. In order to asses potential chronical stress, faeces samples were also collected in week one, two and three after the integration into the social / conventional box (in total: 4 samples per horse and housing system). The samples were immediately stored at -20°C until they were analysed. The samples were not analysed in the laboratory until the end of the experiment, therefore the duration of conservation in the freezer varied from 40 to 429 days.

A considerable percentage of data from groups 1 and 2 was below the detection limit (<0.8 ng/g) (Tab. 1). Thus the statistical analysis was conducted with the FCM concentration from groups 3 and 4 (n horses = 16) which contained no values below the detection limit.

Tab. 1: Details about FCM values and storage time for the 4 groups of stallions

Group Storage duration [d] Proportion of data below the detection limit (<0.8 ng/g) Mean [ng/g] Median [ng/g]

Group 1 384-429 55.6 % 2.2 0

Group 2 315-360 25.5 % 5.8 6.3

Group 3 41-79 0.0 % 8.7 8.0

Group 4 40-85 0.0 % 5.8 5.4

Despite the impressive social interactions observed between the stallions directly after being introduced into the social boxes, we did not find any differences in FCM concentration between the stallions being introduced into the conventional box and the social box on the day of integration (social box: n samples = 16, mean±SD: 6.9±4.7 ng/g; conventional box: n samples = 16, mean±SD: 9.0±11.2 ng/g; Wilcoxon signed rank test V = 70, p = 0.94).

Overall the samples taken during integration and in week one, two and three did not show evidence of changes in FCM concentration in either housing system over a longer period of time (social box: n samples = 64, mean±SD: 7.9±6.2 ng/g; conventional box: n samples = 64, mean±SD: 6.6±3.4 ng/g; Linear mixed model (LMM), p = 0.56).

Our results suggest that the possibility of having physical contact with a conspecific does not induce changes in FCM concentration in breeding stallions. The considerable percentage of values below the detection limit in groups 1 and 2 seemed to correlate with the increasing duration of storage before analysis. During the IESM Network Meeting 2015, we would like to discuss possible methodological issues and the possibilities to correctly integrate these low values in the statistical analysis.
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Corporate Author Wyss, C. Thesis
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Call Number Equine Behaviour @ team @ Serial 5869
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Author Erber, R.; Wulf, M.; Aurich, J.; Becker-Birck, M.; Rose-Meierhöfer, S.; Möstl, E.; Hoffmann, G.; Aurich, C.
Title Physiological stress parameters in sport horse mares transferred from group housing to individual stabling Type Conference Article
Year 2012 Publication Proceedings of the 2. International Equine Science Meeting Abbreviated Journal (down) Proc. 2. Int. Equine. Sci. Mtg
Volume in press Issue Pages
Keywords horse, isolation, cortisol, heart rate, locomotion
Abstract Initial equestrian training and especially first mounting of a rider are stressful challenges for young horses (1). Most young horses are raised in groups but, in association with equestrian training, they are commonly transferred to individual stabling in loose boxes. Although, in most stables, visual contact with horses in adjacent boxes is possible, separation from the herd might be an additional stressor. We have studied physiological stress parameters, in 3-year-old sport horse mares (n=8), transferred from a group stable with access to a paddock to individual boxes without paddock. Once stabled in the individual boxes, mares underwent a standard training for young horses. Horses had been accustomed to lunging and tolerating a rider on their back several weeks before the study. Mares were studied from 5 days before to 5 days after changing the stable. Cortisol concentration in saliva, locomotion activity (ALT pedometers), heart rate (HR) and HR variability (RMSSD: root mean square of successive beat-to-beat intervals) were determined. We hypothesized that the change of the stable increases cortisol release and is associated with changes in HR and RMSSD and reduced locomotion. Before mares were moved to individual boxes, cortisol concentration showed a pronounced diurnal rhythm with values around 0.6 ng/ ml in the morning and a continuous decrease throughout the day. When the mares were moved to individual boxes, cortisol concentration increased to 1.8±0.2 ng/ml and did not return to baseline values within 6 h (p<0.05 over time). On subsequent days, a diurnal rhythm was re-established but shifted to a higher level than before. Locomotion activity determined by ALT pedometers was increased for some minutes only after mares has been placed in individual boxes but was only slightly higher than during the time mares spent with the group in a paddock. On days 2-5 in individual boxes, locomotion activity was reduced compared to the group stable. HR increased and the HRV variable RMSSD decreased when mares were separated. In conclusion, separating horses during initial training from their group is an additional stressor, although the stress is less pronounced than induced by other social challenges, e.g. weaning of foals (2). When stabled in individual boxes, mares move less than when kept as a group. Horses kept in a group thus appear to exercise themselves freely, such an effect is absent when the animals are kept individually.
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Corporate Author Erber, R. Thesis
Publisher Xenophon Publishing Place of Publication Wald Editor Krueger, K.
Language Summary Language Original Title
Series Editor Series Title Abbreviated Series Title
Series Volume Series Issue Edition
ISSN 978-3-9808134-26 ISBN Medium
Area Expedition Conference
Notes Approved no
Call Number Equine Behaviour @ team @ Serial 5542
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Author Ramos, D.; Reche-Junior, A.; Fragoso, P.L.; Palme, R.; Yanasse, N.K.; Gouvêa, V.R.; Beck, A.; Mills, D.S.
Title Are cats (Felis catus) from multi-cat households more stressed? Evidence from assessment of fecal glucocorticoid metabolite analysis Type Journal Article
Year 2013 Publication Physiology & Behavior Abbreviated Journal (down) Physiol. Behav.
Volume 122 Issue Pages 72-75
Keywords Feline; Housing; Non-invasive; Enzyme immunoassay; Cortisol
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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Language Summary Language Original Title
Series Editor Series Title Abbreviated Series Title
Series Volume Series Issue Edition
ISSN 0031-9384 ISBN Medium
Area Expedition Conference
Notes Approved no
Call Number Equine Behaviour @ team @ Serial 5997
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Author Palme, R.
Title Non-invasive measurement of glucocorticoids: Advances and problems Type Journal Article
Year 2019 Publication Physiology & Behavior Abbreviated Journal (down) Physiol. Behav.
Volume 199 Issue Pages 229-243
Keywords Glucocorticoids; Non-invasive; Faecal cortisol/corticosterone metabolites; Immunoassays; Physiological/biological validation
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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Language Summary Language Original Title
Series Editor Series Title Abbreviated Series Title
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ISSN 0031-9384 ISBN Medium
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Notes Palme2019_attachment.pdf Approved no
Call Number Equine Behaviour @ team @ Serial 6517
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Author Yarnell, K.; Hall, C.; Billett, E.
Title An assessment of the aversive nature of an animal management procedure (clipping) using behavioral and physiological measures Type Journal Article
Year 2013 Publication Physiology & Behavior Abbreviated Journal (down) Physiol. Behav.
Volume 118 Issue Pages 32-39
Keywords Thermography; Eye temperature; Cortisol; Horse; Welfare; Clipping
Abstract Animal management often involves procedures that, while unlikely to cause physical pain, still cause aversive responses. The domestic horse (Equus caballus) regularly has excessive hair clipped off to facilitate its use as a riding/driving animal and this procedure causes adverse behavioral responses in some animals. The aim of this study was to compare behavioral and physiological measures to assess the aversive effect of this procedure. Ten horses were selected on the basis of being either compliant (C: n=5) or non-compliant (NC: n=5) during this procedure. The horses were subjected to a sham clipping procedure (SC: where the blades had been removed from the clippers) for a period of ten minutes. Measures were taken pre, during and post SC (-10min to +30min) and mean values calculated for ALL horses and for C and NC separately. Behavioral activity was scored (scale 1-5) by twenty students from video footage in (phase/group-blind scoring). Heart rate (HR), salivary cortisol and eye temperature were monitored throughout the procedure. The NC horses were found to be significantly more behaviorally active/less relaxed throughout the trial than C horses (p<0.05) with the greatest difference occurring during the SC procedure (p<0.01). NC horses were more active/less relaxed during, compared with pre or post SC (p<0.05), but showed no behavioral difference pre and post SC. HR of the NC horses was higher than that of the C horses throughout the trial but only significantly so after 10min of SC (p<0.01). ALL horses showed a significant increase in HR between +5 and +10min into the procedure (p<0.05). There was a significant increase in salivary cortisol concentration in ALL horses post procedure (p<0.01) with levels peaking at 20minute post SC. No significant differences in salivary cortisol concentration between C and NC were found at any stage of the trial. Eye temperature increased significantly in ALL horses during SC, peaking at +10min into the procedure (p<0.05) and then decreased substantially when SC had ceased (p<0.01). Although no significant differences were found between C and NC per se, there was a significant interaction between group and phase of trial (p<0.05) with the NC group showing a greater decrease in eye temperature post SC. There was a significant positive correlation between changes in salivary cortisol concentration and eye temperature (p<0.01) but no correlation between any of the other measures. Although the behavioral response of C and NC to this procedure was significantly different the physiological responses indicated that ALL horses found the procedure aversive. Eye temperature could be used as an objective and immediate measure of how an animal is responding to a specific situation in order to evaluate management procedures and adapt them where appropriate to reduce the negative impact on animal health and welfare.
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Language Summary Language Original Title
Series Editor Series Title Abbreviated Series Title
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ISSN 0031-9384 ISBN Medium
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Notes Approved no
Call Number Equine Behaviour @ team @ Serial 6611
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Author Suagee-Bedore, J.K.; Linden, D.R.; Bennett-Wimbush, K.
Title Effect of Pen Size on Stress Responses of Stall-Housed Horses Receiving One Hour of Daily Turnout Type Journal Article
Year 2021 Publication Journal of Equine Veterinary Science Abbreviated Journal (down) J. Equine Vet. Sci.
Volume 98 Issue Pages 103366
Keywords Agonistic behaviors; Cortisol; Group turnout; Paddock sizes
Abstract Group turnout provides important socializing opportunities for horses, particularly those that are primarily stalled. A high percentage of equine injuries occur during group turnout, which could partly be due to the physical constraints of fencing. To investigate appropriate paddock sizes for group turnouts, horses (n = 12) from a single herd were divided into groups of 4, stalled for 24 hours, and then turned out for 1 hour into one of three differently sized pens: 342, 263, and 184 m2 per horse. Groups rotated through pens across 3 days, receiving one treatment per day. Blood was sampled for cortisol concentrations at 08:00 hours each morning, and then at 15 and 60 minutes into the turn out sessions, and 60 minutes after return to individual stalls. Groups rotated through three turnout times: 09:00, 12:00, 14:00 hours. Counts of agonistic behaviors (chasing, contact biting, and kicking) and low-level threats (pinned ears, tail swishing, bite and kick threats) were recorded. When turned out in pens that provided 342 m2 per horse, horses exhibited reduced plasma cortisol concentrations by 15 minutes after turnout and at 1 hour after return to their stalls (P < .05). Horses in pens providing 184 m2 per horse exhibited greater agonistic (P < .001) and low-level threat (P < .01) behaviors than horses in larger pens. These data provide insight into appropriate pen sizes for horses from established herds. Providing at least 342 m2 per horse may reduce the chance of injury in horses accustomed to group turnout.
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Language Summary Language Original Title
Series Editor Series Title Abbreviated Series Title
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ISSN 0737-0806 ISBN Medium
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Notes Approved no
Call Number Equine Behaviour @ team @ Serial 6694
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Author Dalmau, A.; Ferret, A.; Chacon, G.; Manteca, X.
Title Seasonal Changes in Fecal Cortisol Metabolites in Pyrenean Chamois Type Journal Article
Year 2007 Publication Journal of Wildlife Management Abbreviated Journal (down) J Wildl Manag
Volume 71 Issue 1 Pages 190-194
Keywords Cadí-Moixeró, Nature Reserve, chamois, cortisol metabolites, feces, hunting reserve, Pyrenees, Rupicapra pyrenaica pyrenaica, seasonal rhythm, stress
Abstract We studied seasonal changes in fecal cortisol metabolites (FCM), which have been widely used as indicators of stress, in a population of Pyrenean chamois (Rupicapra pyrenaica pyrenaica) in the Cad&#237; Range of northeastern Spain. We collected fecal samples from 2001 to 2003 in 3 particular locations with different altitudes and male or female presence, and we analyzed them for FCM and fecal nitrogen as an indicator of diet quality. We observed a clear seasonal pattern, with the highest FCM in winter, and we obtained correlations between FCM and monthly mean minimum temperatures and fecal nitrogen. We observed no effects of tourism presence, trophy hunting, or rut season on FCM. Analysis of cortisol metabolites in feces can be a good measure of winter stress in Pyrenean chamois.
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Call Number Equine Behaviour @ team @ Serial 4254
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Author Schmidt, A.; Möstl, E.; Wehnert, C.; Aurich, J.; Müller, J.; Aurich, C.
Title Cortisol release and heart rate variability in horses during road transport Type Journal Article
Year 2010 Publication Hormones and Behavior Abbreviated Journal (down) Horm. Behav.
Volume 57 Issue 2 Pages 209-215
Keywords Horse; Transport; Cortisol; Heart rate variability
Abstract Based on plasma cortisol concentrations it is widely accepted that transport is stressful to horses. So far, cortisol release during transport has not been evaluated in depth by non-invasive techniques such as analysis of salivary cortisol and faecal cortisol metabolites. Transport also causes changes in heart rate and heart rate variability (HRV). In this study, salivary cortisol, faecal cortisol metabolites, heart rate and HRV in horses transported by road for short (one and 3.5 h) and medium duration (8 h) were determined. With the onset of transport, salivary cortisol increased immediately but highest concentrations were measured towards the end of transport (4.1 ± 1.6, 4.5 ± 2.6, 6.5 ± 1.8 ng/ml in horses transported for one, 3.5 and 8 h, respectively). Faecal cortisol metabolite concentrations did not change during transport, but 1 day after transport for 3.5 and 8 h had increased significantly (p < 0.01), reflecting intestinal passage time. Compared to salivary cortisol, changes in faecal cortisol metabolites were less pronounced. Heart rate increased and beat-to-beat (RR) interval decreased (p < 0.05) with the onset of transport. Standard deviation of heart rate increased while root mean square of successive RR differences (RMSSD) decreased in horses transported for 3.5 (from 74 ± 5 to 45 ± 6 ms) and 8 h (from 89.7 ± 7 to 59 ± 7 ms), indicating a reduction in vagal tone. In conclusion, transport of horses over short and medium distances leads to increased cortisol release and changes in heart rate and HRV indicative of stress. The degree of these changes is related to the duration of transport. Salivary cortisol is a sensitive parameter to detect transient changes in cortisol release.
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Publisher Place of Publication Editor
Language Summary Language Original Title
Series Editor Series Title Abbreviated Series Title
Series Volume Series Issue Edition
ISSN 0018-506x ISBN Medium
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Notes Approved no
Call Number Equine Behaviour @ team @ Serial 5387
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Author Dreschel, N.A.; Granger, D.A.
Title Methods of collection for salivary cortisol measurement in dogs Type Journal Article
Year 2009 Publication Hormones and Behavior Abbreviated Journal (down) Horm. Behav.
Volume 55 Issue 1 Pages 163-168
Keywords Dog; Canine; Salivary cortisol; Methods; Measurement; Stress
Abstract Salivary cortisol has been increasingly used as a measure of stress response in studies of welfare, reaction to stress and human–animal interactions in dogs and other species. While it can be a very useful measure, there are a number of saliva collection issues made evident through studies in the human and animal fields which have not been investigated in the canine species. Collection materials and the volume of saliva that is collected; the use of salivary stimulants; and the effect of food contamination can all dramatically impact cortisol measurement, leading to spurious results. In order to further examine the limitations of the collection method and the effects of collection material and salivary stimulant on salivary cortisol levels, a series of clinical, in vitro and in vivo studies were performed. It was found that there is a large amount of inter- and intra-individual variation in salivary cortisol measurement. Beef flavoring of collection materials leads to unpredictable variability in salivary cortisol concentration. Using salivary stimulants such as citric acid also has the potential to affect cortisol concentration measurement in saliva. Hydrocellulose appears to be a useful collection material for salivary cortisol determination. Recommendations for collection materials and use of salivary stimulants are presented.
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Publisher Place of Publication Editor
Language Summary Language Original Title
Series Editor Series Title Abbreviated Series Title
Series Volume Series Issue Edition
ISSN 0018-506x ISBN Medium
Area Expedition Conference
Notes Approved no
Call Number Equine Behaviour @ team @ Serial 5560
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