Abstract: Since the non-invasive field endocrinology techniques were developed, several fecal preservation and extraction methods have been established for a variety of species. However, direct adaptation of methods from previous studies for use in crested macaques should be taken with caution. We conducted an experiment to assess the accuracy and stability of fecal estrogen metabolite (E1C) and glucocorticoid metabolite (GCM) concentrations in response to several preservation parameters: (1) time lag between sample collection and fecal preservation; (2) long-term storage of fecal samples in 80% methanol (MeOH) at ambient temperature; (3) different degrees of feces drying temperature using a conventional oven; and (4) different fecal preservation techniques (i.e., freeze-drying, oven-drying, and field-friendly extraction method) and extraction solvents (methanol, ethanol, and commercial alcohol). The study used fecal samples collected from crested macaques (Macaca nigra) living in the Tangkoko Reserve, North Sulawesi, Indonesia. Samples were assayed using validated E1C and GCM enzyme immunoassays. Concentrations of E1C and GCM in unprocessed feces stored at ambient temperature remained stable for up to 8 h of storage after which concentrations of both E1C and GCM changed significantly compared to controls extracted at time 0. Long-term storage in 80% MeOH at ambient temperature affected hormone concentrations significantly with concentrations of both E1C and GCM increasing after 6 and 4 months of storage, respectively. Drying fecal samples using a conventional oven at 50, 70, and 90 °C did not affect the E1C concentrations, but led to a significant decline for GCM concentrations in samples dried at 90 °C. Different fecal preservation techniques and extraction solvents provided similar results for both E1C and GCM concentrations. Our results confirm previous studies that prior to application of fecal hormone analysis in a new species, several preservation parameters should be evaluated for their effects on hormone metabolite stability. The results also provide several options for fecal preservation, extraction, and storage methods that can be selected depending on the condition of the field site and laboratory.

Abstract: Relatively few large-scale faecal DNA studieshave been initiated due to difficulties inamplifying low quality and quantity DNAtemplate. To improve brown bear faecal DNA PCRamplification success rates and to determinepost collection sample longevity, fivepreservation methods were evaluated: 90%ethanol, DETs buffer, silica-dried, oven-driedstored at room temperature, and oven-driedstored at -20 °C. Preservationeffectiveness was evaluated for 50 faecalsamples by PCR amplification of a mitochondrialDNA (mtDNA) locus (~146 bp) and a nuclear DNA(nDNA) locus (~200 bp) at time points of oneweek, one month, three months and six months. Preservation method and storage timesignificantly impacted mtDNA and nDNAamplification success rates. For mtDNA, allpreservation methods had >= 75% success atone week, but storage time had a significantimpact on the effectiveness of the silicapreservation method. Ethanol preserved sampleshad the highest success rates for both mtDNA(86.5%) and nDNA (84%). Nuclear DNAamplification success rates ranged from 26-88%, and storage time had a significant impacton all methods but ethanol. Preservationmethod and storage time should be importantconsiderations for researchers planningprojects utilizing faecal DNA. We recommendpreservation of faecal samples in 90% ethanolwhen feasible, although when collecting inremote field conditions or for both DNA andhormone assays a dry collection method may beadvantageous.

Abstract: Sensory lateralization in dogs (n = 74) was investigated in this study using our innovation, the Sensory Jump Test. This required the modification of head halters to create three different ocular treatments (binocular, right, and left monocular vision) for eye preference assessment in a jumping task. Ten jumps were recorded as a jump set for each treatment. Measurements recorded included (i) launch and landing paws, (ii) type of jump, (iii) approach distance, (iv) clearance height of the forepaw, hindpaw, and the lowest part of the body to clear the jump, and (v) whether the jump was successful. Factors significantly associated with these jump outcomes included ocular treatment, jump set number, and replication number. Most notably, in the first jump set, findings indicated a left hemispheric dominance for the initial navigation of the Sensory Jump Test, as left monocular vision (LMV) compromised of jumping more than right monocular (RMV) and binocular vision, with a significantly reduced approach distance and forepaw clearance observed in dogs with LMV. However, by the third jump set, dogs undergoing LMV launched from a greater approach distance and with a higher clearance height, corresponding to an increase in success rate of the jump, in comparison with RMV and binocular vision dogs. A marginally non-significant RMV bias was observed for eye preference based on the laterality indices for approach distance (P = 0.060) and lowest body part clearance height (P = 0.067). A comparison between eye preference and launching or landing paws showed no association between these measures of sensory and motor laterality. To our knowledge, this is the first study to report on sensory lateralization in the dog, and furthermore, to compare both motor and sensory laterality in dogs.

Abstract: We used an opportunistic review of photographs of different adult and juvenile horses walking, trotting, and cantering (n = 828) to compare the angle of the nasal plane relative to vertical in feral and domestic horses at liberty (n = 450) with ridden horses advertised in a popular Australian horse magazine (n = 378). We assumed that horses in advertisements were shown at, what was perceived by the vendors to be, their best. Of the ridden horses, 68% had their nasal plane behind the vertical. The mean angle of the unridden horses at walk, trot, and canter (30.7 ± 11.5; 27.3 ± 12.0; 25.5 ± 11.0) was significantly greater than those of the ridden horses (1.4 ± 14.1; ?5.1 ± ?11.1; 3.1 ± 15.4, P < 0.001). Surprisingly, unridden domestic horses showed greater angles than feral horses or domestic horses at liberty. We compared adult and juvenile horses in all 3 gaits and found no significant difference. Taken together, these findings demonstrate that the longitudinal neck flexion of the degree desirable by popular opinion in ridden horses is not a common feature of unridden horses moving naturally. Moreover, they suggest that advertised horses in our series are generally being ridden at odds with their natural carriage and contrary to the international rules of dressage (as published by the International Equestrian Federation). These findings are discussed against the backdrop of the established doctrine, which states that carrying a rider necessitates changes in longitudinal flexion, and in the context of the current debate around hyperflexion.

Abstract: The traditional way to train horses is by the application of negative reinforcement (NR). In the past few years, however, the use of positive reinforcement (PR) has become more common. To evaluate the effectiveness and the possible stressor effect of the 2 training methods, 12 horses showing severe trailer-loading problems were selected and exposed to trailer-loading. They were randomly assigned to one of the 2 methods. NR consisted of various degrees of pressure (lead rope pulling, whip tapping). Pressure was removed as soon as the horse complied. PR horses were exposed to clicker training and taught to follow a target into the trailer. Heart rate (HR) was recorded every 5 seconds and behavior denoting discomfort was observed using one-zero sampling with 10 seconds sampling intervals. Training was completed when the horse could enter the trailer upon a signal, or was terminated after a maximum of 15 sessions. Of the 12 horses, 10 reached the criterion within the 15 sessions. One horse was eliminated from the study because of illness and 1 PR horse failed to enter the trailer. A Mann-Whitney U-test indicated that the horses trained with NR displayed significantly more discomfort behavior per training session than horses trained with PR (NR: 13.26 ± 3.25; PR: 3.17 ± 8.93, P < 0.0001) and that horses in the PR group spent less time (second) per session to complete the training criterion (NR: 672.9 ± 247.12; PR: 539.81 ± 166.37, P < 0.01). A Mann-Whitney U-test showed that no difference existed in mean HR (bpm) between the 2 groups (NR: 53.06 ± 11.73 bpm; PR: 55.54 ± 6.7 bpm, P > 0.05), but a Wilcoxon test showed a difference in the PR group between the baseline of HR and mean HR obtained during training sessions (baseline PR: 43 ± 8.83 bpm; PR: 55.54 ± 6.7 bpm, P < 0.05). In conclusion, the PR group provided the fastest training solution and expressed less stress response. Thus, the PR procedure could provide a preferable training solution when training horses in potentially stressing situations.