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Suagee-Bedore, J. K., Linden, D. R., & Bennett-Wimbush, K. (2021). Effect of Pen Size on Stress Responses of Stall-Housed Horses Receiving One Hour of Daily Turnout. J. Equine Vet. Sci., 98, 103366.
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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Paramastri, Y., Royo, F., Eberova, J., Carlsson, H. - E., Sajuthi, D., Fernstrom, A. - L., et al. (2007). Urinary and fecal immunoglobulin A, cortisol and 11-17 dioxoandrostanes, and serum cortisol in metabolic cage housed female cynomolgus monkeys (Macaca fascicularis). Journal of Medical Primatology, 36(6), 355–364.
Abstract: Background and methods Quantitative enzyme-immunoassays of urinary and fecal immunoglobulin A (IgA), cortisol and 11-17-dioxoandrostanes (11,17-DOA), and serum cortisol in eight metabolic-cage-housed female cynomolgus monkeys were performed. The monkeys were divided into two groups, B and NB. Group B animals were blood sampled every 6 hours, whereas Group NB animals were not handled/blood sampled. Results No differences were recorded between the amounts of feces and urine excreted by the two groups. Group B animals excreted more urinary cortisol than did Group NB animals indicating that restraint-blood sampling resulted in a stress response. Excreted amounts of IgA and 11,17-DOA (urine and feces) did not differ between the groups. Conclusions Urinary cortisol was a reliable marker of the stress associated with repeated blood sampling. Declining amounts of excreted urinary cortisol indicated that cynomolgus monkeys acclimated quickly to repeated blood sampling in metabolism cages. Within and between animal variation in amounts of feces voided demonstrated the importance of expressing fecal markers as ‘amounts excreted per time unit per kg body weight’ rather than just measuring the concentrations in fecal samples.
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Batt, L. S., Batt, M. S., Baguley, J. A., & McGreevy, P. D. (2009). The relationships between motor lateralization, salivary cortisol concentrations and behavior in dogs. Journal of Veterinary Behaviour, 4(6), 216–222.
Abstract: The degree of lateralization (LI) indicates both the direction and strength of a paw preference. Here, a positive value is indicative of a right paw bias, and a negative value of a left paw bias. Higher numbers on the positive side of the scale and lower numbers on the negative side of the scale indicate a greater strength of that lateralization. The strength of motor lateralization (|LI|) is the absolute value of the LI. The use of absolute value removes directionality (i.e., does not indicate left or right paw bias) and instead indicates only the strength of the paw preference. Both LI and |LI| have been associated with behavioral differences in a range of species. The assessment of motor lateralization in the dog can be conducted by observing the paw used to perform motor tasks. Elevated cortisol concentrations have been associated with fearfulness in many species. Additionally, fearfulness and boldness can be assessed in response to so-called temperament tests. Consequently, in this study we examine the relationship between lateralization, temperament test results, and cortisol concentrations in 43 potential guide dogs, of which 38 were Labrador retrievers and 5 were golden retrievers. Over a 14-month period, the current study assessed motor lateralization and salivary cortisol concentrations 3 times (approximately 6 months of age, 14 months of age, and after the dogs' performance in the guide dog program had been determined) and behavior twice (approximately 6 and 14 months of age). This study is the first to examine the relationship between behavior, lateralization, and cortisol concentrations in dogs. It implemented an objective and quantifiable assessment of behavior that may be of use to a variety of dog-focused stakeholders. Findings show that during the Juvenile testing period (6 months of age), dogs with higher cortisol concentrations were typically less able to rest when exposed to the unfamiliar testing room. Results from both Juvenile and Adult Test (14 months of age) periods showed that a greater |LI| and LI were associated with more confident and relaxed behavior when dogs were exposed to novel stimuli and unfamiliar environments. Significant elevations of cortisol concentrations were found at the completion of guide dog training when compared with results from the 2 prior test periods. This finding may reflect maturation or the effect of the prolonged kenneling which occurred during this period.
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Dalmau, A., Ferret, A., Chacon, G., & Manteca, X. (2007). Seasonal Changes in Fecal Cortisol Metabolites in Pyrenean Chamois. J Wildl Manag, 71(1), 190–194.
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 Cadí 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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Hoffmann, G., Bockisch, F. - J., & Kreimeier, P. (2009). Einfluss des Haltungssystems auf die Bewegungsaktivität und Stressbelastung bei Pferden in Auslaufhaltungssystemen. Landbauforschung – vTI Agriculture and Forestry Research, 2(59), 105–112.
Abstract: Frühere Untersuchungen haben gezeigt, dass die tägliche Bewegung für die Gesunderhaltung der Pferde notwendig ist. Inwieweit sich jedoch unterschiedliche Bewegungsangebote auf das Stress-und Bewegungsverhalten von Pferden in einer Gruppen-Auslaufhaltung auswirken und ob der Bewegungsbedarf der Pferde durch eine Auslaufhaltung ohne zusätzliche Bewegung gedeckt werden kann, ist der Literatur bisher nicht zu entnehmen. Daher sollte in der nachfolgend beschriebenen Untersuchung der Frage nachgegangen werden, welche Auswirkungen verschiedene Bewegungsangebote auf die Bewegungsaktivität von Pferden in Gruppen-Auslaufhaltungen haben und ob diese das Wohlbefinden der Tiere beeinflussen. Letzteres wurde durch Messung der Herzfrequenzvariabilität und Bestimmung von Cortisolmetaboliten im Pferdekot erfasst und die Bewegungsaktivität der Pferde wurde mit ALT-Pedometern bestimmt.
Verglichen wurden eine Einzel-und Gruppenhaltung mit jeweils angrenzendem Auslauf, aber ohne eine zusätzliche Bewegung der Pferde außerhalb des Stalls. In drei weiteren Varianten der Gruppenhaltung bekamen die Pferde täglichen Auslauf auf einer unbegrünten Koppel, auf einer Weide oder durch gezielte Bewegung in einer Führanlage. Die Bewegungsaktivität konnte durch die zusätzliche Bewegung in Form von Weide oder Führanlage signifikant gesteigert werden.
Ein zusätzliches Bewegungsangebot führte bei den Pferden zu einer Abnahme der Stressbelastung und sollte auch den Pferden ermöglicht werden, die in einer Gruppenhaltung gehalten werden, um ihre physische und psychische Gesundheit zu erhalten.
[Former studies confirm the necessity of daily movement for the health of a horse. But so far no description could be found in the literature how different movement offerings impact the stress and movement behaviour of horses in group husbandries with close-by discharge. The same holds true for the question whether a discharge husbandry system can meet the movement requirements of horses if there isn�t any additional movement possibility. The aim of the present study was to examine different movement offerings, their effects on the movement activities of horses in a group horse husbandry with close-by discharge and the impact of the movement on the wellbeing of the animals.
The heart rate variability and the concentration of the cortisol metabolites in the horse excrement were analyzed for detecting the wellbeing of the horses. Additionally ALT-Pedometers were used for determining the movement activity.
A single and a group husbandry system, each with closeby discharge, were compared when horses had no additional movement outside the stable. In three further variants the group husbandry was supplemented with daily time on a non-grassy pasture land, a pasture or in a horse walker. Pasture or horse walker increased movement activity significantly. Nevertheless an additional movement offering resulted in a lower stress load of the horses and should also be allowed to horses in group husbandry systems to ensure the horse�s physical and mental health.]
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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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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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Yarnell, K., Hall, C., & Billett, E. (2013). An assessment of the aversive nature of an animal management procedure (clipping) using behavioral and physiological measures. Physiol. Behav., 118, 32–39.
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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Erber, R., Wulf, M., Aurich, J., Becker-Birck, M., Rose-Meierhöfer, S., Möstl, E., et al. (2012). Physiological stress parameters in sport horse mares transferred from group housing to individual stabling. In K. Krueger (Ed.), Proceedings of the 2. International Equine Science Meeting (Vol. in press). Wald: Xenophon Publishing.
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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Wyss, C. (2015). Does housing in a „social box“ change faecal cortisol metabolites concentration in stallions? In Proceedings of the 3. International Equine Science Meeting.
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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