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Schmidt, A., Biau, S., Möstl, E., Becker-Birck, M., Morillon, B., Aurich, J., et al. (2010). Changes in cortisol release and heart rate variability in sport horses during long-distance road transport. Domest Anim Endocrinol, 38(3), 179–189.
Abstract: It is widely accepted that transport is stressful for horses, but only a few studies are available involving horses that are transported regularly and are accustomed to transport. We determined salivary cortisol immunoreactivity (IR), fecal cortisol metabolites, beat-to-beat (RR) interval, and heart rate variability (HRV) in transport-experienced horses (N = 7) in response to a 2-d outbound road transport over 1370 km and 2-d return transport 8 d later. Salivary cortisol IR was low until 60 min before transport but had increased (P < 0.05) 30 min before loading. Transport caused a further marked increase (P < 0.001), but the response tended to decrease with each day of transport. Concentrations of fecal cortisol metabolites increased on the second day of both outbound and return transports and reached a maximum the following day (P < 0.001). During the first 90 min on Day 1 of outbound transport, mean RR interval decreased (P < 0.001). Standard deviations of RR interval (SDRR) decreased transiently (P < 0.01). The root mean square of successive RR differences (RMSSD) decreased at the beginning of the outbound and return transports (P < 0.01), reflecting reduced parasympathetic tone. On the first day of both outbound and return transports, a transient rise in geometric HRV variable standard deviation 2 (SD2) occurred (P < 0.01), indicating increased sympathetic activity. In conclusion, transport of experienced horses leads to increased cortisol release and changes in heart rate and HRV, which is indicative of stress. The degree of these changes tended to be most pronounced on the first day of both outbound and return transport.
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Stull, C. L., Spier, S. J., Aldridge, B. M., Blanchard, M., & Stott, J. L. (2004). Immunological response to long-term transport stress in mature horses and effects of adaptogenic dietary supplementation as an immunomodulator. Equine Veterinary Journal, 36(7), 583–589.
Abstract: Reasons for performing study: Little information exists on the immunological effects of transport or the use of supplements to minimise transport stress. Objectives: To establish baseline ranges and evaluate immunophenotypic and functional changes associated with transport and a nutritional ‘adaptogen’ supplement. Methods: Horses received either supplement (n = 10) or placebos (n = 9) during the 30 day study. After 28 days in stalls, 12 horses (6 supplement; 6 placebo) were transported for 24 h, then unloaded and recovered. Venous blood samples were collected on Days 1, 14 and 28 to establish baselines, and on Days 28, 29 and 30 to examine changes during transport and recovery. Results: Transport prompted elevations (P<0.05) in cortisol concentration, neutrophil count and white blood cell counts, while lymphocyte subpopulation counts (CD3+, CD4+, CD8+, CD21+) decreased (P<0.05). Normal phenotypic lymphocyte profiles returned within 24 h of recovery. Supplement effects on immunophenotype (CD21+ and CD8+) were observed in stabled horses (P<0.05), but not in transported horses. Conclusions: These results provide insights into the immunological mechanisms associated with long-term transport. Potential relevance: The existence of a small window of immunological uncertainty follows long-term transportation, enhancing the potential risk of infectious disease in susceptible individuals.
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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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Flauger, B., Krueger, K., Gerhards, H., & Möstl, E. (2010). Simplified method to measure glucocorticoid metabolites in faeces of horses. Vet Res Comm, 34(2), 185–195.
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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Kedzierski, W., Wilk, I., & Janczarek, I. (2014). Physiological response to the first saddling and first mounting of horses: comparison of two sympathetic training methods. Animal Science Papers and Reports, 32(3), 219–228.
Abstract: There is not much research done on the influence of sympathetic training on the emotional reaction
of horses. The aim of the present study was to evaluate the emotional response and the stress level
in horses to two sympathetic training methods: (1) with the use of the “round pen technique” (RP),
and (2) in which the RP was not applied (SH). Twenty two half-bred Anglo-Arab horses (2.5
years ±3 months of age) were subject to an initial training. Eleven horses were randomly included
to the RP method and the other 11 horses for the SH method. Heart rate (HR) and saliva cortisol
concentration were measured as indicators of horse emotional arousal and stress level, respectively.
The HR values were analysed: at rest, during the habituation period, just after the first saddling
and tightening of the girth, during the first time a human leaned over the horse’s back, and during
the mounting of the horse. Saliva samples were taken before and 15 min after each training session
studied. After saddling, the HR occurred significantly higher when the RP technique was used. The
significant increase in saliva cortisol concentration was observed only after the first mounting of
the horse. Generally, the use of the RP technique did not involve more important physiological
reactions in the trained horses than did the SH method.
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Palme, R. (2012). Monitoring stress hormone metabolites as a useful, non-invasive tool for welfare assessment in farm animals. Animal Welfare, 21(3), 331–337.
Abstract: A multitude of endocrine mechanisms are involved in coping with challenges. Glucocorticoids, secreted by the adrenal glands, are in the front line of the battle to overcome stressful situations. They are usually measured in plasma samples as parameters of adrenal activity and thus of disturbance. Unfortunately, collecting blood samples itself can disturb an animal. Thus, non-invasive methods for the determination of glucocorticoids or their metabolites have become increasingly popular. The pros and cons of various non-invasive sample materials (saliva, excreta, milk, hair/feathers and eggs) for glucocorticoid determination are given. Above all, faecal samples offer the
advantage that they can be collected easily. In faecal samples, circulating hormone levels are integrated over a certain period of time and represent the cumulative secretion of hormones. Thus, the levels are less affected by short fluctuations or the pulse-like nature of hormone secretion. However, using this technique to assess an animal’s adrenocortical activity is not especially simple. Whether frequent sampling is necessary or single samples will suffice depends upon the study’s aim (whether one is examining the impact of acute or chronic stressors). Background knowledge of the metabolism and excretion of cortisol/corticosterone metabolites is required and a careful validation for each species and sex investigated is obligatory. The present review also addresses analytical issues regarding sample storage, extraction procedures and immunoassays and includes a comprehensive list of published studies (up to 2011) describing the use of such methods in farmed animals. Applied properly, non-invasive techniques to monitor glucocorticoid metabolites in faecal samples of various species are a useful tool for welfare assessment, especially as they are easily applied at farm or group level.
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Cerasoli, F., Podaliri Vulpiani, M., Saluti, G., Conte, A., Ricci, M., Savini, G., et al. (2022). Assessment of Welfare in Groups of Horses with Different Management, Environments and Activities by Measuring Cortisol in Horsehair, Using Liquid Chromatography Coupled to Hybrid Orbitrap High-Resolution Mass Spectrometry (Vol. 12). Animals, 12(14).
Abstract: Horses have always been animals used for companionship, work, transportation, and performance purposes over the history of humanity; there are different ways of managing horses, but studies on how horse welfare is influenced by different activities and managements are scanty. Understanding how the management, the environment, and the different uses of horses can affect the level of stress and well-being is important not only for people associated with horses. Three groups of horses with different management, environments, and activities were selected: (1) stabled horses ridden frequently, (2) horses that perform public order service under the Italian state police, and (3) free-ranging horses. Cortisol analysis was carried out on horsehair samples using liquid chromatography coupled to hybrid orbitrap high-resolution mass spectrometry (LC-HRMS/MS), a laboratory technique used for the first time to quantify horsehair cortisol. The selection of horses to be included in the three groups was carried out by including only subjects with positive welfare assessment in accordance with the horse welfare assessment protocol (AWIN). These analyses demonstrated that the cortisol levels detected in the horsehair of free-ranging animals were significantly higher compared to those detected in stabled and working horses. These results may have been a consequence of complex environmental, managerial, and behavioral factors, which should be worth further investigation
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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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Skandakumar, S., Stodulski, G., & Hau, J. (1995). Salivary IgA: a Possible Stress Marker In Dogs. In Animal Welfare (Vol. 4, pp. 339–350).
Abstract: Stress in humans has been reported to be associated with a decrease in the salivary immunoglobulin A (s-IgA) levels enabling the possible use of s-IgA to assess stress. Prolonged stress, if reliably assessed in a non-invasive manner, may be used to assess animal welfare. This study analysed groups of dogs undergoing physical and temperamental training and s-IgA levels were measured by rocket immunoelectrophoresis in prospective samples. Behavioural assessment was carried out and cortisol levels in saliva were measured by ELISA. A significant negative correlation (P < 0.007) between the logarithmic cortisol concentrations and s-IgA levels in saliva was recorded. The behavioural assessment of the dogs agreed well with the biochemical markers. It is concluded that IgA levels in saliva may be a useful marker of dog well-being and that stress results in decreased s-IgA levels.
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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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