Effect of weaning and feed provision times on the performance and several behavioural traits of post-weaning lambs.

The aim of the present study was to assess if weaning time (morning or evening) and feed provision (immediate or 4 h delayed) can influence growth performance and expression of juvenile lamb behaviour immediately after and at the age of 50-125 days old. Forty lambs were randomly assigned to 4 groups: E1 (evening + immediate), E2 (evening + delayed), M1 (morning + immediate) and M2 (morning + delayed). All lambs were especially active during the first hour after weaning. However, their activity within the first 24 h after weaning was mainly affected by their circadian rhythms, since lambs were more active during the day compared to night. On the other hand, no effect of alfalfa hay provision on lamb activity during these first 24 h was observed. At the age of 50-125 days old, body weight was measured, and an isolation and flight distance test was implemented. There were no significant differences among lambs in body weight, number of jumps and heart rates assessed during isolation test and flight distance. However, the number of vocalizations was lower (65-110th day, P < 0.05) for the E2 compared to E1 lambs. Breed also slightly affected (95th day, P < 0.05) the average heart rate of lambs, with greater values for Karagouniko compared to Chios lambs. It is concluded that no discernible effects of weaning or feeding time were observed on growth performance and the display of stress-induced behavioural components of juvenile lambs till the age of 125 days old.

Spatial distance and reactivity traits alter the positive perception of brushing by ewes.

Gentle handling seems to elicit positive states in sheep. The study investigated whether spatial distance alters sheep responses to brushing and whether spatial distance is influenced by reactivity. Twenty Romane ewes were assessed in three sessions: in Sessions 1 and 3, one grid separated the test animal from pen mates, with no distance between them, and in Session 2 two grids separated the test animal from pen mates by a distance of about 1.7 m. Ewes had been genetically selected for low (R-) or high (R+) behavioural reactivity to social isolation. Body postures, head orientation, ear postures, closed and half-closed eyes, tail wagging and feeding behaviour, in addition to heart rate (HR) and HR variability, as the root mean square of successive differences (RMSSD), standard deviation of all normal-to-normal (NN) intervals (SDNN), RMSSD/SDNN ratio and ratio between low-frequency (LF) and high-frequency (HF) powers (LF/HF) were assessed. Data were analysed using generalized linear models and linear mixed models. Session, genetic line and phase (pre-, brushing and post-brushing) were considered fixed effects. Increased distance in Session 2 might not have influenced ewes' responses. Fewer changes in ear postures were noted in Session 3 than 1 (P<0.01), suggesting that ewes were more relaxed in Session 3. The RMSSD/SDNN ratio was higher mainly during brushing in Sessions 1 and 3 (P<0.05), indicating that ewes were more relaxed during brushing, and at no distance between pen mates. However, spatial distance influenced R- and R+ ewes' responses; R+ ewes performed more asymmetric ear postures in Session 2 than 1 and 3 (P<0.01), and in Session 3 than 1 (P<0.01), indicating that spatial distance had a negative effect on R+ ewes. Low reactive ewes spent less time on horizontal ear postures in Session 2 than 1 and 3 (P<0.01), and R+ ewes spent more time on horizontal postures in Session 1 than 3 (P<0.01). Curiously, R- ewes spent more time eating and ruminating in Session 3 than 1 (P<0.01), and in Session 2 than 1 and 3 (P<0.01), whereas R+ ewes ate and ruminated more in Session 1 than 3 (P<0.05). Higher HR was found among R- ewes in Session 2 than 1 and 3, and in Session 3 than 1 (P<0.01). High reactive ewes showed higher HR in Session 1 than 3 (P<0.01). The findings suggest that the social context might influence sheep responses to gentle handling, and the effects depend on their reactivity traits.

Effects of ammoniated pearl millet silage on intake, feeding behavior, and blood metabolites in feedlot lambs.

The aim of this study was to evaluate the effects of urea as an additive in the ensiling of pearl millet on the intake, feeding behavior, and metabolic parameters of feedlot-finished lambs. Thirty-two uncastrated, mixed-breed male lambs were used in the experiment. Diets were composed of pearl millet silage enriched with 0, 2, 4, or 6% urea plus a concentrate containing ground corn, soybean meal, and a mineral mixture. The treatments did not affect feed intake (P > 0.05) but influenced (P < 0.05) eating time (in min/day, in min/kg of dry matter (DM), and in min/kg of neutral detergent fiber (NDF)) and chewing time in min/kg of DM. Eating efficiency (in g DM/h and in g NDFap/h) responded linearly (P < 0.05) to the increasing urea levels in the silages. By contrast, there was no effect (P > 0.05) of diets on the blood protein profile (total proteins and albumin), although the serum urea levels responded quadratically (P < 0.05). Increasing urea levels in the silage did not change the blood energy profile (cholesterol and triglycerides) or blood enzyme activity (alanine aminotransferase (ALT), aspartate aminotransferase (AST), and gamma-glutamyl transferase (GGT); P > 0.05). In conclusion, the treatment of pearl millet silage with urea does not influence the DM intake or metabolic parameters of lambs, but leads to increased eating time and decreased eating efficiency.

Dynamics of social behaviour at parturition in a gregarious ungulate.

Group living is the behavioural response that results when individuals assess the costs vs benefits of sociality, and these trade-offs vary across an animal's life. Here we quantitatively assess how periparturient condition (mother/non-mother) and births affect the dynamics of social interactions of a gregarious ungulate, and how such can help to explain evolutionary hypotheses of the mother-offspring bond. To achieve this we used data of the individual movement of a group of Scottish blackface sheep (Ovis aries) marked with GPS collars and properties of mathematical graphs (networks). Euclidean pair-wise distance between sheep were threshold at different percentiles to determine network links, and these thresholds have a profound effect on the connectivity of the resulting network. Births increased the average pair-wise distance between mothers, and between mothers and non-mothers, with less effect on the distance between non-mothers. Mothers occupied peripheral positions within the flock, more evident following births. Associations between individuals (i.e. network community change) were highly dynamic, though mothers were less likely to change community than non-mothers, especially after births. Births hampered individual communication within the flock (assessed via network closeness centrality), especially in mothers. Overall leadership (lead positioning relative to flock movement) was not associated to reproductive condition, and individual leadership rank was not affected by births. A ten minute GPS acquisition time was adequate to capture complex social dynamics in sheep movement. The results on mother's isolation behaviour support the hypotheses of selection for maternal imprint facilitation, reducing risks to nursing alien offspring, and group/multilevel selection on group formation.

The ability of ewes with lambs to learn a virtual fencing system.

The Nofence technology is a GPS-based virtual fencing system designed to keep sheep within predefined borders, without using physical fences. Sheep wearing a Nofence collar receive a sound signal when crossing the virtual border and a weak electric shock if continuing to walk out from the virtual enclosure. Two experiments testing the functionality of the Nofence system and a new learning protocol is described. In Experiment 1, nine ewes with their lambs were divided into groups of three and placed in an experimental enclosure with one Nofence border. During 2 days, there was a physical fence outside the border, during Day 3 the physical fence was removed and on Day 4, the border was moved to the other end of the enclosure. The sheep received between 6 and 20 shocks with an average of 10.9±2.0 (mean±SE) per ewe during all 4 days. The number of shocks decreased from 4.38±0.63 on Day 3 (when the physical fence was removed) to 1.5±0.71 on Day 4 (when the border was moved). The ewes spent on average 3%, 6%, 46% and 9% of their time outside the border on Days 1, 2, 3 and 4, respectively. In Experiment 2, 32 ewes, with and without lambs, were divided into groups of eight and placed in an experimental enclosure. On Day 1, the enclosure was fenced with three physical fences and one virtual border, which was then increased to two virtual borders on Day 2. To continue to Day 3, when there was supposed to be three virtual borders on the enclosure, at least 50% of the ewes in a group should have received a maximum of four shocks on Day 2. None of the groups reached this learning criterion and the experiment ended after Day 2. The sheep received 4.1±0.32 shocks on Day 1 and 4.7±0.28 shocks on Day 2. In total, 71% of the ewes received the maximum number of five shocks on Day 1 and 77% on Day 2. The individual ewes spent between 0% and 69.5% of Day 1 in the exclusion zone and between 0% and 64% on Day 2. In conclusion, it is too challenging to ensure an efficient learning and hence, animal welfare cannot be secured. There were technical challenges with the collars that may have affected the results. The Nofence prototype was unable to keep the sheep within the intended borders, and thus cannot replace physical fencing for sheep.

Intake and ingestive behavior of lambs fed diets containing ammoniated buffel grass hay.

This study aimed to evaluate the effect of diets with ammoniated buffel grass hay on the ingestive behavior of feedlot lambs. Thirty-two sheep of no defined breed with an average body weight of 17.7 ± 1.8 kg were used. A completely randomized design with four treatments (0, 18, 36, and 54 g/kg dry matter (DM) basis) and eight repetitions was used. Ingestive behavior, rumination, and idle time were similar (P > 0.05) among the diets containing ammoniated buffel grass hay, with mean values of 294.5, 554.44, and 594.25 min per day, respectively. Regarding the chews, all of the variables resulted in similar behavior (P > 0.05). The quadratic effect (P < 0.05) observed for daily intake can be explained based on the amount of DM and neutral detergent fiber (NDF) per meal per rumination. There was no effect of urea levels from the ammoniation (P > 0.05) on the efficiency of DM and NDF consumption. However, the rumination efficiency of DM and NDF showed a quadratic effect (P < 0.05). Thus, the use of ammoniated buffel grass hay with urea in lamb diet affects the ingestive behavior by increasing the rumination efficiency, increased intake, and feed utilization.

Behavioral evidence of heterospecific bonding between the lamb and the human caregiver and mapping of associated brain network.

While behavioral mechanisms of bonding between young mammals and humans have been explored, brain structures involved in the establishment of such processes are still unknown. The aim of the study was to identify brain regions activated by the presence of the caregiver. Since human positive interaction plays an important role in the bonding process, activation of specific brain structures by stroking was also examined. Twenty-four female lambs reared in groups of three were fed and stroked daily by a female caregiver between birth and 5-7 weeks of age. At 4 weeks, an isolation-reunion-separation test and a choice test revealed that lambs developed a strong bond with their caregiver. At 5-7 weeks of age, lambs were socially isolated for 90min. They either remained isolated or met their caregiver who stroked them, or not, at regular intervals over a 90-min period. Neuronal activation was investigated at the end of the period for maximum c-Fos expression. Reunion with the caregiver appeased similarly the lambs whether stroking was provided or not. Stroking did not activate a specific brain network compared to no stroking. In both cases, brain regions associated with olfactory, visual and tactile cue processing were activated in the presence of the caregiver, suggesting a multisensory process involved. In addition, activation of the oxytocinergic system in the hypothalamic paraventricular nucleus induced by the presence of the caregiver suggests similar neuroendocrine mechanisms involved in inter-conspecific and animal-human bonding.

Attention bias to threat indicates anxiety differences in sheep.

Humans and animals show increased attention towards threatening stimuli when they are in increased states of anxiety. The few animal studies that have examined this phenomenon, known as attention bias, have applied environmental manipulations to induce anxiety but the effects of drug-induced anxiety levels on attention bias have not been demonstrated. Here, we present an attention bias test to identify high and low anxiety states in sheep using pharmacological manipulation. Increased anxiety was induced using 1-methyl-chlorophenylpiperazine (m-CPP) and decreased anxiety with diazepam, and then we examined the behaviour of sheep in response to the presence of a dog as a threat. Increased attention towards the threat and increased vigilance were shown in sheep that received the m-CPP and reduced in sheep receiving the diazepam. The modulated attention towards a threat displayed by the m-CPP and diazepam animals suggests that attention bias can assess different levels of anxiety in sheep. Measuring attention bias has the potential to improve animal welfare assessment protocols.

Effect of pasture size on behavioural synchronization and spacing in German Blackface ewes (Ovis aries).

This study aims to assess plot size related changes in spacing and behavioural synchronization in a herd of 14 German Blackface ewes kept on three different pasture sizes: S (126m(2)), M (1100m(2)), and L (11,200m(2)). In direct field observations, behaviour and nearest neighbour distance were recorded individually. Additionally, interindividual and nearest neighbour distances were derived from aerial photographs of the herd taken on plot sizes S and M. Nearest neighbour distances <1m accounted for more than 60% of observations, and were more frequent on plot size L than on plot sizes S (Z=3.3; p<0.01) and M (Z=3.2; p<0.01). Average interindividual distances were significantly smaller on S (4.89±2.62m) than on M plots (5.99±3.06m; t=7.3; p<0.01). Synchronization tended to increase with plot size (K(S)=0.42; K(M)=0.52; K(L)=0.66), but was not accompanied by a concomitant increase in dispersion. Aerial photography proved a valuable tool in the analysis of spacing behaviour as intraindividual repeatability of the derived distances was highly significant (Kendall's W between 0.32 and 0.58; p<0.01). The sheep kept small distances on all plot sizes, thus the high degree of behavioural synchronization might be mainly attributed to the motivation for close proximity to any conspecific.

Laterality as an indicator of emotional stress in ewes and lambs during a separation test.

We assessed motor laterality in sheep to explore species-specific brain hemi-field dominance and how this could be affected by genetic or developmental factors. Further, we investigated whether directionality and strength of laterality could be linked to emotional stress in ewes and their lambs during partial separation. Forty-three ewes and their singleton lambs were scored on the (left/right) direction of turn in a y-maze to rejoin a conspecific (laterality test). Further, their behavioural response (i.e. time spent near the fence, vocalisations, and activity level) during forced separation by an open-mesh fence was assessed (separation test). Individual laterality was recorded for 44.2% ewes (significant right bias) and 81.4% lambs (equally biased to the left and the right). There was no significant association in side bias between dams and offspring. The Chi-squared test revealed a significant population bias for both groups (p < 0.05). Evolutionary adaptive strategies or stimuli-related visual laterality may provide explanation for this decision-making process. Absolute strength of laterality (irrespective of side) was high (Kolmogorov-Smirnov test, dams: D = 0.2; p < 0.001; lambs: D = 0.36, p < 0.0001). The Wilcoxon test showed that lateralised lambs and dams spent significantly more time near each other during separation than non-lateralised animals (p < 0.05), and that lateralised dams were also more active than non-lateralised ones. Arguably, the lateralised animals showed a greater attraction to their pair because they were more disturbed and thus required greater reassurance. The data show that measures of laterality offer a potential novel non-invasive indicator of separation stress.

Effect of a screen with flaps and straw on behaviour, stress response, productive performance and meat quality in indoor feedlot lambs.

We analysed the effect of a modified pen using a wooden screen with flaps and cereal straw as forage and bedding, on behaviour, stress response, performance and meat quality variables of lambs housed in feedlots. Sixty male lambs were placed in enriched (ESF) or conventional (CO) pens (3 pens/treatment, 10 lambs/pen). The CO environment was barren. The ESF lambs showed a great preference for the provided items, which encouraged more natural and richer behaviour, reducing stereotypies and lamb aggressions, and increasing affiliations (P ≤ 0.05), which improves group cohesion. However, ESF lambs also developed a more natural coping style to the handling, evidenced by the higher cortisol levels (65.4 vs. 43.8 nmol/L) and a higher eye temperature as response to the reactivity test (38.1 vs. 37.8 °C). The ESF lambs had a higher (P ≤ 0.05) slaughter weight (27.2 vs. 26.3 kg), conformation score (7.38 vs. 6.07) and pH 24 (5.63 vs. 5.56) but lower cooking losses (12.9 vs. 14.9%) than CO lambs.

Development of affinity to the stockperson in lambs from two breeds.

The interactions between genetic and environmental factors on the development of lamb affinity to humans was assessed on 48 animals from two breeds (Gentile di Puglia and Comisana) and two treatments (Gentled and Not Gentled) producing 4 groups (GPg, GPng, Cg, Cng, respectively). The Gentile di Puglia and Comisana gentled animals were subjected to a period of training consisting of gently handling each lamb for 5 min three times a day for the first week and then twice a week for three additional weeks. The gentling procedure included both tactile and visual/auditory interactions. At 30-32 days of age lambs were subjected to three arena tests conducted in a novel environment: i) isolation test (each animal was exposed to a novel environment, and isolated from tactile and visual contact with conspecifics for 5 min), ii) stationary human test (as previously but a human sitting in a corner of the pen), and iii) pen-mates' test (each animal was tested in the presence of 2 pen-mates whose behaviour was not recorded). During the 13 training sessions lambs exhibited an increasing number of contacts with the human and a decreasing number of bleats (P<0.001), although Gentile di Puglia lambs interacted more with the human (P<0.001) and tended to bleat less (P<0.10) than Comisana lambs. Lambs vocalised more and climbed more when tested in isolation as compared with lambs tested either with the human or with the pen-mates (P<0.01) and they vocalised more in the presence of the human than with the pen-mates (P<0.001). A higher number of contacts with the human stimulus was observed in gentled animals (P<0.05). However, differences between gentled and not gentled animals were only significant in Gentile di Puglia subjects (P<0.01). GPng lambs displayed the longest ambulatory activity during the isolation test (P<0.05) and exhibited more climbing attempts during the isolation test as compared with the human or the pen-mates' tests (P<0.001). A higher cortisol level was shown by GPng lambs in comparison with Cng subjects during the stationary human test (P<0.01), whereas no differences were detected between the two gentled groups. Gentling determined an improvement of the quality of human animal relationship in more reactive breeds such as Gentile di Puglia sheep.

Social rank affects reproductive development in male lambs.

Under field conditions, dominant rams may prevent mating by other rams, thereby depressing flock fertility. However, little is known about the effects of social rank on prepuberal reproductive development in male lambs. The objective of this experiment was to determine if social rank of lambs is related to body weight, scrotal circumference, testosterone concentrations, semen production and courtship behavior until 38 weeks of age. An additional aim was to characterize male-male sexual behavior and to determine if this behavior was related to social rank. Body weight, scrotal circumference and testosterone serum concentrations were measured weekly in 13 male lambs. In addition, semen was obtained by electroejaculation, and sexual behavior towards non-estrous ewes was registered every 2 weeks. Male-male mounts were also recorded weekly. Individual social rank success indexes were determined using the food competition test when lambs were 3 and 6 months old. The five rams with greater (GR) and the five with lesser (LR) success indexes were used in the experiment. Body weight and scrotal circumference increases were greater in GR than in LR lambs. Testosterone concentrations increased with age, but were not related to social rank. Semen characteristics matured earlier in GR than in LR lambs. Most courtship and mating behaviors (lateral approaches, flehmen, ejaculations and ejaculations/total mounts) increased with age, but no effect of social rank was observed. The number of ano-genital sniffing and mount attempts increased with age and was greater in GR than LR. The total number of male-male mounts in which each lamb took part was positively related with the success index (r=0.48; P=0.05), and tended to be related with the number of times each lamb was mounted (r=0.42; P=0.06). No relation was observed between the number of times each lamb mounted and the social rank success index. We observed that GR lambs mature earlier than LR lambs, which is reflected by a greater increase of their body weight and scrotal circumference. Semen production and male-female sexual behavior also occurred earlier in high-ranked than in low-ranked lambs. It remains to be determined if low-ranked lambs can achieve similar development than high-ranked lambs over longer time intervals, or if their maximum developmental capacity is lower than the maximum obtained by high-ranked lambs.