Studying vocal correlates of emotions is important to provide a better understanding of the evolution of emotion expression through cross-species comparisons. Emotions are composed of two main dimensions: emotional arousal (calm versus excited) and valence (negative versus positive). These two dimensions could be encoded in different vocal parameters (segregation of information) or in the same parameters, inducing a trade-off between cues indicating emotional arousal and valence. We investigated these two hypotheses in horses. We placed horses in five situations eliciting several arousal levels and positive as well as negative valence. Physiological and behavioral measures collected during the tests suggested the presence of different underlying emotions. First, using detailed vocal analyses, we discovered that all whinnies contained two fundamental frequencies (“F0” and “G0”), which were not harmonically related, suggesting biphonation. Second, we found that F0 and the energy spectrum encoded arousal, while G0 and whinny duration encoded valence. Our results show that cues to emotional arousal and valence are segregated in different, relatively independent parameters of horse whinnies. Most of the emotion-related changes to vocalizations that we observed are similar to those observed in humans and other species, suggesting that vocal expression of emotions has been conserved throughout evolution.

Summary Reasons for performing study: Studies on the prevalence of behavioural disorders in horses and on associated risk factors have revealed inconsistent results. There are many studies on the neuropharmacological, surgical ormechanical therapy of stereotypies, but little is known about their causation. Objectives: To explore risk factors associated with the occurrence of behavioural disorders in horses. Methods: A sample of horse owners, selected randomly and representative for Switzerland, was contacted in a postal survey. Answers were provided for 622 stables (response rate 35.2%). Individual data of 2341 horses were examined with path analysis (multivariable linear and logistic regression), and adjustment made for possible confounding effects due to age and breed. Results: Out of 60 possible risk factors, 11 were associated with the outcome at the univariable level (null‐hypothesis path model) and 3 factors remained after the backward logistic regression procedure. Mature Warmbloods and Thoroughbreds, assessed by the owners to be reactive, fed 4 times a day and without daily pasture, had increased odds of displaying crib‐biting, weaving and box‐walking. Furthermore, indirect associations of 5 factors with the outcome were identified. Conclusions: The final logistic regression model of risk factors leads to the hypotheses that causal prevention of stereotypic behaviours should be based upon housing and management conditions which allow tactile contact with other horses (e.g. mutual grooming), daily free movement (paddock or pasture), as well as the provision of high amounts of roughage but of little or no concentrates. Potential clinical relevance: It is one of the aims of population medicine to prevent the development of behavioural disorders. Further research is needed to test the concluding hypotheses in experimental studies or to verify them in the context of similar observational studies.

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Finding valid indicators of emotional states is one of the biggest challenges in animal welfare science. Here, we investigated in horses whether variation in the expression of eye wrinkles caused by contraction of the inner eyebrow raiser reflects emotional valence. By confronting horses with positive and negative conditions, we aimed to induce positive and negative emotional states, hypothesising that positive emotions would reduce whereas negative emotions would increase eye wrinkle expression. Sixteen horses were individually exposed in a balanced order to two positive (grooming, food anticipation) and two negative conditions (food competition, waving a plastic bag). Each condition lasted for 60 seconds and was preceded by a 60 second control phase. Throughout both phases, pictures of the eyes were taken, and for each horse four pictures per condition and phase were randomly selected. Pictures were scored in random order and by two experimenters blind to condition and phase for six outcome measures: qualitative impression, eyelid shape, markedness of the wrinkles, presence of eye white, number of wrinkles, and the angle between the line through the eyeball and the highest wrinkle. The angle decreased during grooming and increased during food competition compared to control phases, whereas the two phases did not differ during food anticipation and the plastic bag condition. No effects on the other outcome measures were detected. Taken together, we have defined a set of measures to assess eye wrinkle expression reliably, of which one measure was affected by the conditions the horses were exposed to. Variation in eye wrinkle expression might provide valuable information on horse welfare but further validation of specific measures across different conditions is needed.

The identification of quantitative trait loci (QTL) such as height and their underlying causative variants is still challenging and often requires large sample sizes. In humans hundreds of loci with small effects control the heritable portion of height variability. In domestic animals, typically only a few loci with comparatively large effects explain a major fraction of the heritability. We investigated height at withers in Shetland ponies and mapped a QTL to ECA 6 by genome-wide association (GWAS) using a small cohort of only 48 animals and the Illumina equine SNP70 BeadChip. Fine-mapping revealed a shared haplotype block of 793 kb in small Shetland ponies. The HMGA2 gene, known to be associated with height in horses and many other species, was located in the associated haplotype. After closing a gap in the equine reference genome we identified a non-synonymous variant in the first exon of HMGA2 in small Shetland ponies. The variant was predicted to affect the functionally important first AT-hook DNA binding domain of the HMGA2 protein (c.83G>A; p.G28E). We assessed the functional impact and found impaired DNA binding of a peptide with the mutant sequence in an electrophoretic mobility shift assay. This suggests that the HMGA2 variant also affects DNA binding in vivo and thus leads to reduced growth and a smaller stature in Shetland ponies. The identified HMGA2 variant also segregates in several other pony breeds but was not found in regular-sized horse breeds. We therefore conclude that we identified a quantitative trait nucleotide for height in horses.

Moods can influence our judgment of ambiguous stimuli as positive or negative. Measuring judgment bias in animals is a promising method to objectively assess their emotional states. Our study aimed to develop a cognitive bias test in horses, in order to assess the effect of training using positive reinforcement (PR) or negative reinforcement (NR) on their emotional states. We trained 12 mares to discriminate between a rewarded and a non-rewarded location situated on each side of a paddock. The mares were then trained during five days to perform several exercises using PR (n = 6) for one group, and NR (n = 6) for the other (treatment). Finally, we compared the responses of the two groups to three ambiguous locations situated between the rewarded and non-rewarded locations (judgment bias test). During the training exercises, according to our predictions, behavioural measures suggested that NR mares experienced more negative emotions than PR mares. Surprisingly, the results of the judgment bias test suggest that NR mares were in a more optimistic mood compared to PR mares, despite previously experiencing more negative emotions during the treatment. NR mares could have been more motivated to obtain a food reward than PR mares, which had been rewarded throughout the treatment phase. Alternatively, NR mares could have developed optimistic bias triggered by release from the negative state experienced during treatment. This first attempt to test judgment bias in horses suggests that this is a promising method to measure horse mood. Knowledge about the effect of training methods on the mental health of domesticated animals can add a new dimension to animal welfare, in order to promote better ways to work with animals. 2.3. Judgment bias We used a judgment bias test, which uses spatial location as a stimulus (Burman et al., 2008a, 2009), to assess horse mood following treatment.

BackgroundNon-human animals often produce different types of vocalisations in negative and positive contexts (i.e. different valence), similar to humans, in which crying is associated with negative emotions and laughter is associated with positive ones. However, some types of vocalisations (e.g. contact calls, human speech) can be produced in both negative and positive contexts, and changes in valence are only accompanied by slight structural differences. Although such acoustically graded signals associated with opposite valence have been highlighted in some species, it is not known if conspecifics discriminate them, and if contagion of emotional valence occurs as a result. We tested whether domestic horses perceive, and are affected by, the emotional valence of whinnies produced by both familiar and unfamiliar conspecifics. We measured physiological and behavioural reactions to whinnies recorded during emotionally negative (social separation) and positive (social reunion) situations.ResultsWe show that horses perceive acoustic cues to both valence and familiarity present in whinnies. They reacted differently (respiration rate, head movements, height of the head and latency to respond) to separation and reunion whinnies when produced by familiar, but not unfamiliar individuals. They were also more emotionally aroused (shorter inter-pulse intervals and higher locomotion) when hearing unfamiliar compared to familiar whinnies. In addition, the acoustic parameters of separation and reunion whinnies affected the physiology and behaviour of conspecifics in a continuous way. However, we did not find clear evidence for contagion of emotional valence.ConclusionsHorses are thus able to perceive changes linked to emotional valence within a given vocalisation type, similar to perception of affective prosody in humans. Whinnies produced in either separation or reunion situations seem to constitute acoustically graded variants with distinct functions, enabling horses to increase their apparent vocal repertoire size.Electronic supplementary materialThe online version of this article (doi:10.1186/s12983-017-0193-1) contains supplementary material, which is available to authorized users.

Horses can sleep while standing; however, recumbency is required for rapid eye movement (REM) sleep and therefore essential. Previous research indicated a minimal duration of recumbency of 30 min per 24 h to perform a minimal duration of REM sleep. For group-housed horses, suitable lying area represents a potentially limited resource. In Switzerland, minimal dimensions for the space allowance of the littered area are therefore legally required. To assess the effect of different space allowances of the littered area on lying behavior, 38 horses in 8 groups were exposed to 4 treatments for 11 days each; T0: no litter provided, T0.5: 0.5× minimal dimensions, T1: minimal dimensions, and T1.5: 1.5× minimal dimensions. Non-littered areas were covered with hard rubber mats. Lying behavior was observed during the last 72 h of each treatment. The total number of lying bouts per 24 h was similar in treatments providing litter, whereas in treatment T0, recumbency occurred only rarely (F1,93 = 14.74, p = 0.0002) with the majority of horses lying down for less than 30 min per 24 h (χ12=11.82, p = 0.0006). Overall, the total duration of recumbency per 24 h increased with increasing dimensions of the littered area, whereby the effect attenuated between treatment T1 and T1.5 in high-ranking horses but continued in low-ranking horses (F1,91 = 3.22, p = 0.076). Furthermore, low-ranking horses showed considerably more forcedly terminated lying bouts in treatments T0.5 and T1, but were similar to high-ranking horses in T1.5 (F1,76 = 8.43, p = 0.005). Nonetheless, a number of individuals showed durations of recumbency of less than 30 min per 24 h even in treatment T1.5. The lying behavior was dependent on the availability of a soft and deformable surface for recumbency. A beneficial effect of enlarged dimensions of the littered area was shown by increased durations of recumbency and decreased proportion of forcedly terminated lying bouts in low-ranking horses. Taking this into account, it became evident that the minimal dimensions for the littered area as implemented in the Swiss animal welfare legislation do not ensure undisturbed lying behavior for all members of a given group.