No abstract

One of the most significant challenges in the recovery of threatened species is the ability to maintain genetic diversity, avoid inbreeding and sustain population health and reproduction. Assisted reproductive techniques, including artificial insemination (AI), have been touted for decades as approaches that could contribute to the demographic and genetic management of rare species. Here, we report the first successful integration of AI with frozen semen into a formal recovery program and the positive impact on genetic diversity for the critically endangered black-footed ferret Mustela nigripes. Techniques developed in the taxonomically related domestic ferret Mustela putorius furo and Siberian polecat Mustela eversmannii were applied over time to selected black-footed ferrets, including semen banking from six of the last 18 survivors. After evaluation, processing and storage in liquid nitrogen (−196°C/−321°F), for as long as 20 years, sperm samples were thawed and transabdominally inseminated into the uterine horns of female conspecifics. Eight black-footed ferret offspring were produced using thawed sperm samples (including after two decades of cryopreservation) with inseminates containing as few as 3.4 × 10 6 motile spermatozoa. The incorporation of these offspring and/or their descendants into the ex situ breeding program prevented heterozygosity loss in the population and actually enhanced gene diversity (GD) significantly by 0.2% and lowered measures of inbreeding by 5.8%. This study demonstrates the utility and genetic diversity benefits of applying AI with cryopreserved spermatozoa 20 generations removed from the contemporary population for a wild animal revival program.

Hormonal analysis provides information about wildlife populations, but is difficult to conduct in the field. Our goal was to develop a rapid and effective field method for fecal steroid analysis by comparing: (1) three extraction methods (laboratory (LAB), homogenize (HO) and handshake (HS)) and (2) two storage methods (solid-phase extraction (SPE) tubes vs. plastic tubes (PT)). Samples (n=23) from captive African wild dogs (Lycaon pictus) were thoroughly mixed, three aliquots of each were weighed ( approximately 0.5 g) and 5 ml of 90% ethanol was added. For LAB, samples were agitated (mixer setting 60; 30 min), centrifuged (1,500 rpm; 20 min) and poured into glass tubes. Or aliquots were HO (1 min) or HS (1 min) and poured through filter paper into glass tubes. Samples were split, analyzed for corticosterone (C) and testosterone (T) metabolites using enzyme immunoassays or stored in SPE or PT. Samples were stored (room temperature) for 30, 60 or 180 days, reconstituted in buffer and analyzed. Mean C and T recoveries of HO were greater (P=0.03) than HS compared with LAB, which was similar to HO (P>0.05). After 30 days <21% of C and T was recovered from SPE, but approximately 100% of each was recovered from HO-PT and HS-PT. Similarly, after 60 and 180 days, approximately 100% of C and T was recovered from HO-PT and HS-PT. Results demonstrated that, for C and T, HO was more comparable (P<0.001) to LAB than HS and PT storage was more efficient than SPE (P<0.001).

We are developing a novel method using non-invasive skin swabs to measure stress in several amphibians from terrestrial, semi-aquatic and fully aquatic species both at the zoo and in the wild. Upon further testing, this technique will allow future studies to understand how the environment is impacting amphibian health and susceptibility to disease.

Monitoring adrenocortical activity in wild primate populations is critical, given the well-documented relationship between stress, health and reproduction. Although many primate studies have quantified fecal glucocorticoid metabolite (FGM) concentrations, it is imperative that researchers validate their method for each species. Here, we describe and validate a technique for field extraction and storage of FGMs in wild chimpanzees (Pan troglodytes). Our method circumvents many of the logistical challenges associated with field studies while yielding similar results to a commonly-used laboratory method. We further validate that our method accurately reflects stress physiology using an ACTH challenge in a captive chimpanzee and a FGM peak at parturition in a wild subject. Finally, we quantify circadian patterns for FGMs for the first time in this species. Understanding these patterns may allow researchers to directly link specific events with the stress response.

Population augmentation with translocated individuals has been shown to alleviate the effects of bottlenecks and drift. The first step to determine whether restoration for genetic considerations is warranted is to genetically monitor reintroduced populations and compare results to those from the source. To assess the need for genetic restoration, we evaluated genetic diversity and structure of reintroduced (n = 3) and captive populations of the endangered black-footed ferret (Mustela nigripes). We measured genotypic changes among populations using seven microsatellite markers and compared phenotypic changes with eight morphometric characters. Results indicated that for the population which rapidly grew postreintroduction, genetic diversity was equivalent to the captive, source population. When growth languished, only the population that was augmented yearly maintained diversity. Without augmentation, allelic diversity declined precipitously and phenotypic changes were apparent. Ferrets from the genetically depaupertate population had smaller limbs and smaller overall body size than ferrets from the two populations with greater diversity. Population divergence (F ST = 0.10 ± 0.01) was surprisingly high given the common source of populations. Thus, it appears that 5-10 years of isolation resulted in both genotypic divergence and phenotypic changes to populations. We recommend translocation of 30-40 captive individuals per annum to reintroduction sites which have not become established quickly. This approach will maximize the retention of genetic diversity, yet maintain the beneficial effects of local adaptation without being swamped by immigration.

Free-roaming dogs (Canis lupus familiaris) are of public health and conservation concern because of their potential to transmit diseases, such as rabies, to both people and wildlife. Understanding domestic dog population dynamics and how they could potentially be impacted by interventions, such as rabies vaccination, is vital for such disease control efforts. For four years, we measured demographic data on 2,649 free-roaming domestic dogs in four rural villages in Tanzania: two villages with and two without a rabies vaccination campaign. We examined the effects of body condition, sex, age and village on survivorship and reproduction. Furthermore, we compared sources of mortality among villages. We found that adult dogs (>12mos) had higher survival than puppies in all villages. We observed a male-biased sex ratio across all age classes. Overall survival in one non-vaccination village was lower than in the other three villages, all of which had similar survival probabilities. In all villages, dogs in poor body condition had lower survival than dogs in ideal body condition. Sickness and spotted hyena (Crocuta crocuta) predation were the two main causes of dog death. Within vaccination villages, vaccinated dogs had higher survivorship than unvaccinated dogs. Dog population growth, however, was similar in all the villages suggesting village characteristics and ownership practices likely have a greater impact on overall dog population dynamics than vaccination. Free-roaming domestic dogs in rural communities exist in the context of their human owners as well as the surrounding wildlife. Our results did not reveal a clear effect of vaccination programs on domestic dog population dynamics. An investigation of the role of dogs and their care within these communities could provide additional insight for planning and implementing rabies control measures such as mass dog vaccination.

Introductions of sable antelope (Hippotragus niger) can be difficult due to the potential ensuing aggression compounded by their large horns. The goal was to use hormonal assays and behavioral analyses to evaluate the success of an introduction of 2 adult females at Lincoln Park Zoo. The objectives were to (a) document behavioral and hormonal changes in 2 female sable antelope during the introduction, (b) compare fecal glucocorticoid metabolites (FGM) in each individual during the introduction stages, (c) measure fecal androgen metabolites (FAM) during introduction and compare with dominance rank and observed aggression, and (d) monitor estrous cycle synchronization. Results demonstrate that FGM were higher before than during and after the introduction. Behavioral observations indicated limited aggression between females, although the keeper survey results revealed that the new female was more dominant and had higher mean FGM and FAM than the resident. Both sable antelope were reproductively active throughout the year. Results indicate that fecal hormone analysis can provide zoo management with valuable information to minimize the risk of aggression, injury, and stress during introductions of nonhuman animals.