Appeasing pheromones for the management of stress and aggression during conservation of wild canids: could the solution be right under our nose?

Thirty-six species of canid exist globally, two are classified as critically endangered, three as endangered, and five as near threatened. Human expansion and the coinciding habitat fragmentation necessitate conservation interventions to mitigate concurrent population deterioration. The current conservation management of wild canids includes animal translocation and artificial pack formation. These actions often cause chronic stress, leading to increased aggression and the suppression of the immune and reproductive systems. Castration and pharmaceutical treatments are currently used to reduce stress and aggression in domestic and captive canids. The undesirable side effects make such treatments inadvisable during conservation management of wild canids. Pheromones are naturally occurring chemical messages that modulate behaviour between conspecifics; as such, they offer a natural alternative for behaviour modification. Animals are able to distinguish between pheromones of closely related species through small compositional differences but are more likely to have greater responses to pheromones from individuals of the same species. Appeasing pheromones have been found to reduce stress- and aggression-related behaviours in domestic species, including dogs. Preliminary evidence suggests that dog appeasing pheromones (DAP) may be effective in wild canids. However, the identification and testing of species-specific derivatives could produce more pronounced and beneficial behavioural and physiological changes in target species. In turn, this could provide a valuable tool to improve the conservation management of many endangered wild canids

Improved sperm freezing in the endangered African wild dog (Lycaon pictus) using a two-step dilution TRIS-egg yolk extender containing Equex STM

Development of assisted breeding techniques can aid conservation and management of the endangered African wild dog (Lycaon pictus). Previous attempts to freeze sperm from this species has proven unsuccessful with sperm motility dropping to nearly 0% within 2 h of thawing. The aim of this study was to improve the freezing success of African wild dog sperm by testing two routinely used canine cryopreservation protocols. Sperm was frozen from n=3 captive African wild dog males housed at Albuquerque BioPark (Albuquerque, NM, USA) and Binder Park Zoo (Battle Creek, MI, USA) during the breeding season (Aug-Sept 2014). Freshly collected semen samples were evaluated for volume, colour, pH, motility, viability, morphology, sperm number, acrosome status and DNA integrity. Each sample was split and frozen using two different protocols. Protocol 1: semen was diluted with a Tris-egg yolk extender containing 8% glycerol and 20% egg yolk, and slowly cooled from 37°C to 4°C over 2.5 h. The sample was then loaded into 0.25 mL straws, suspended 4 cm over liquid nitrogen vapour for 10 min, then plunged in liquid nitrogen. Protocol 2: semen was first diluted with a Tris-egg yolk extender containing only 3% glycerol and 20% egg yolk, followed by a second extender (same composition) now containing 7% glycerol and 1% Equex STM, added after the 2.5 h refrigeration period. The freezing procedure was the same as Protocol 1. Straws from both protocols were thawed in a 37ᵒC water bath, but Protocol 2 straws were further diluted by with a thawing solution which that consisted of the initial extender solution without glycerol and egg yolk. Sperm were incubated at 37 ᵒC and motility evaluated at 5 min and every 2 h for 8 h after thawing. Viability, morphology and acrosome integrity was evaluated over 6 h and DNA integrity was evaluated immediately post-thaw. Sperm motility declined significantly for both protocols immediately after thawing (fresh 78.9 ± 2.6%; Protocol 1 24.4 ± 5.0%; Protocol 2 36.7 ± 4.2%; P ≤ 0.05). Motility was significantly higher for Protocol 2 from 2 h after thawing (Protocol 1 1.0 ± 0.8%; Protocol 2 30.8 ± 1.9%; P ≤ 0.05) and sperm remained motile for up to 8 h. Sperm frozen with Protocol 2 also had significantly higher viability (Protocol 1 37.0 ± 5.7%; Protocol 2 65.3 ± 9.9%; P ≤ 0.05) and acrosome integrity (Protocol 1 22.8 ± 8.2%; Protocol 2 69.3 ± 8.8%; P ≤ 0.05) immediately after thawing. There was no difference in the proportion of normal morphology or DNA fragmentation between both protocols. Our results demonstrate that using a two-step dilution with TRIS-egg yolk extender containing Equex STM yields greatly improved post-thaw quality and longevity in African wild dog sperm; making it suitable for use in artificial insemination

Monitoring stress in captive and free-ranging African wild dogs (Lycaon pictus) using faecal glucocorticoid metabolites

An understanding of stress physiology is important for species management because high levels of stress can hamper reproduction and affect an individual’s ability to cope with threats to their survival, such as disease and human–wildlife conflict. A commonly used indicator of stress, faecal concentrations of cortisol metabolites (FCM), can be used to assess the impact of social, biological and environmental factors. Measurements of FCM are particularly valuable for endangered species that are logistically challenging to study and where non-invasive techniques are preferred. As the second most endangered canid in Africa, the African wild dog (Lycaon pictus) has been the focus of considerable conservation research, yet there is still little understanding of factors associated with stress, in either captive or free-ranging populations. The present study therefore aimed to determine whether stress levels differ between captive and freeranging populations, and to detect social, biological and environmental factors that are stressful in these populations. Faecal samples were collected from 20 captive and 62 free-ranging animals. Within freeranging populations, the sexes differed significantly, but there was no effect of social status, age or breeding period for either sex. Captive females had higher FCM concentrations than free-ranging females. In captive populations, FCM concentrations differed among zoos and with reproductive status in females, but were not related to age class or group-housing structure. In conclusion, FCM is a useful indicator of stress and should be considered an integrative aspect of management, for both in situ and ex situ African wild dog populations.Australian Postgraduate Award and funding provided by the School of Animal Biology (University of Western Australia) and the Institute for Breeding Rare and Endangered African Mammals (IBREAM).http://www.elsevier.com/locate/ygcen2017-01-31hb201

Innovative parental care in a myrmecophageous mammal

Male bat-eared foxes, Otocyon megalotis, are known to contribute extensively to parental care. Yet, the exact roles that males and females play in raising offspring remain relatively unexplored. Here, we describe interactions between adult foxes and their presumed offspring based on a pilot study on three family groups of a wild population in South Africa. We report the first recorded instance of dung provisioning observed in canids. A male bat-eared fox provided dung to his offspring during a foraging trip, presumably to give them access to the ensconced insects. Further,this male provisioned the young foxes with large, live insects. Similar to other researchers, we never observed provisioning by females, but the females in this population did interact socially with their young in addition to suckling.We emphasize the importance of anecdotal reports of novel behavioural responses in wild canids, as an accumulation of such evidence may reveal patterns of innovative behaviour presently unrecognized in this family.NRF focal area grant awarded to Professor Elissa Cameron, an NRF SARCHi Chair in Mammal Behavioural Ecology and Physiology awarded to Professor Nigel Bennett and NRF incentive funds as well as a research fellowship from the University of Pretoria awarded to F. Dalerum.http://link.springer.com/journal/10211hb201

Novel appeasing pheromones to minimize stress during metapopulation management of African wild dogs

In the last century, human persecution and increased habitat fragmentation has reduced African wild dog populations from around 500,000 to 6,600 individuals, with the population continuing to decline. In South Africa, metapopulation management has been instrumental to the survival of populations across fragmented habitats, and entails the translocation and artificial new pack formation of animals to reinforce populations, maximize genetic dispersal and reintroduce animals into their former range. However, temporary captivity during such conservation interventions regularly cause chronic stress in African wild dogs, and can result in increased aggression, disrupted pack hierarchy, injury and occasionally mortality. Pheromones are naturally occurring chemicals that can moderate behaviours and physiology in conspecifics. Appeasing pheromones identified in domestic dogs (DAP) are known to reduce stress and aggression. When applied to African wild dog packs, we showed DAP treatment decreased faecal androgen metabolite concentrations and shifted dominance behaviour from contact to non-contact compared to controls (Van den Berghe et al. PLoS ONE 14(3): e0212551). However, because pheromones are largely species-specific, an African wild dog-specific appeasing pheromone (AWDAP) should elicit a stronger beneficial physiological and behavioural effect during conservation interventions. In this project, AWDAP will be isolated and applied to packs undergoing translocation in South Africa. Using a combination of behaviour, faecal hormone metabolites and antibodies, animals will be monitored for reduced stress and aggression, improved immune function and greater social cohesion, all of which should ultimately confer a survival advantage to the pack upon release into the wild

Profiling patterns of fecal 20-oxopregnane concentrations during ovarian cycles in free-ranging southern white rhinoceros (Ceratotherium simum simum)

Unlike their wild counterparts, many white rhinoceros females in captivity fail to reproduce successfully such that current captive populations are not self-sustaining. The causes of the problem are poorly understood. Variation in cycle length and long periods of acyclicity are characteristics of the majority of these non-reproducing females in captivity but it is unknown whether these characteristics are a feature of reproductively successful free-ranging females. This study therefore aimed to monitor cyclic activity in a wild population of southern white rhinoceros at Lapalala Wilderness, South Africa, by measuring the concentrations of immunoreactive fecal progestagen metabolites (fPM). Five adult females were tracked twice per week for 20 months and if located a fresh fecal sample was collected. Reproductive events and group structural dynamics were also recorded and subsequently correlated with the fPM data. The baseline concentration of fPM was 0.69 ± 0.20 μg/g DW while concentrations during pregnancy were 30- to 400- fold higher. The females exhibited estrous cycle lengths of 30.6 ± 7.7 days and, based on fPM data, gestation length in one female was 502 ± 3 days. Year-round monitoring showed no clear evidence of seasonality in ovarian activity. During cyclic luteal activity females were often seen in the presence of a dominant bull. One female stopped cycling after removal of the local dominant bull and luteal activity only returned after a new bull was introduced. This suggests that white rhinoceros females in the wild might need external stimuli from a male to ovulate. These findings indicate that the irregular cyclicity reported for white rhinoceros housed in zoos and animal parks may result from conditions in captivity and account for reduced fertility.The University of Western Australia, the Mammal Research Institute and Endocrine Research Laboratory, University of Pretoria, the Institute for Breeding Rare and Endangered African Mammals, the AAZK‘s Bowling For Rhinos, and the SAVE African Rhino Foundation.http://www.elsevier.com/locate/anireprosci2016-10-31hb201

Faecal progestagen profiles in wild southern white rhinoceros (Ceratotherium simum simum)

Knowledge of the reproductive biology of wild animals can provide valuable information for the development of appropriate in situ and ex situ management plans. The present study aimed to establish a non-invasive protocol for monitoring faecal progestagen metabolite (FPM) patterns in wild female southern white rhinoceroses (Ceratotherium simum simum). Six adult females at Lapalala Wilderness, South Africa, were tracked and accurately identified at least once every week. Three animals gave birth during the study period. Fresh faecal samples were collected for 12 months and stored frozen at –20°C until analysis with an enzyme immunoassay utilizing an antiserum raised against 5 -pregnan-3 -ol-20-one which cross-reacts with a number of progestagens. Mean FPM concentrations were 35 to 64-fold higher during pregnancy (55–145 days before parturition) compared to postpartum (120–140 days after parturition) (P < 0.001). Also, the non-pregnant animals had mean FPM concentrations significantly higher than postpartum values (P = 0.006). Our results show that non-invasive FPM measurements provide information on the pregnancy status of wild female white rhinoceroses, and may be used for the detection of pregnancy in free-living individuals, without the necessity of immobilization and/or relocation of the animal. This information has potential value for optimizing breeding management of wild and captive populations.We wish to thank the Institute for Breeding Rare and Endangered African Mammals (IBREAM) for making this research possible.We are indebted to the following students from the faculty of Veterinary Medicine at Utrecht University for their contribution in collecting faecal samples: Yvette Charbon, Belinda Bitter, Meike Meijers and Julie Swinkels. We are especially thankful to Thomas Litshani, whose exceptional rhinoceros knowledge and tracking skills greatly contributed to the number of samples collected.We are most grateful to the management of the Lapalala Wilderness reserve: Duncan Parker, Mike Gregor, Anton Walker, Roger Collinson and Anthony and Erin Roberts for permission to conduct research in the reserve as well as for their help and support. We also thank John Hanks and Henk Bertschinger for their encouragement and helpful advice, and Stefanie Ganswindt for expert help in laboratory techniques.http://africanzoology.journals.ac.za/am2013ab201