Abstract

Introduction

Regular manipulation of large, nondomestic hoofstock species is a necessary component of effective preventative health programs at zoological facilities and is often restricted by the zoo’s limited handling capabilities. In the majority of cases, large ruminants must be chemically immobilized for basic physical examination, research procedures, disease testing and certain types of medical therapy. As an alternative to chemical immobilization, appropriate handling facilities permit the physical restraint of hoofstock species in a manner that is safe for both animal and handler.^1,2 The Wilds (Cumberland, OH) has developed an effective handling system which has been designed to reduce adverse visual stimuli, ensure the smooth advancement of animals through the system without the need for human contact, and takes advantage of the animal’s ‘natural’ behavior to accomplish our specific goals. The system incorporates a series of corridors of varying widths, strategically placed transfer gates and doors that may be remotely operated, and a modified version of a commercially available hydraulic restraint device (The Tamer™, Fauna Products Inc., Red Hook, NY). This system has been regularly used for physiologic sample collection, testing and medical treatment of such species as scimitar horned oryx (Oryx dammah), sable (Hippotragus niger), eland (Taurotragus oryx), Jackson’s hartebeest (Alcelaphus buselaphus jacksoni), Pere David’s deer (Cervus elaphurus), Bactrian deer (Cervus elaphus bactrianus) and Urial sheep (Ovis vignei). In addition to being used in preventative health programs, this handling system has also proven valuable in several reproductive studies carried out at this facility.

Physical and psychologic stress associated with handling can result in physical injury, capture myopathy and, on occasion, may even result in the animal’s death. The handling system developed at the Wilds has been designed to move animals in a step-wise but smooth fashion, from a holding pen into a transfer corridor and finally into the Tamer™. A series of sliding gates allows animals to be moved through the system without direct human manipulation and allows the easy separation of individuals within the transfer corridors. The walls of the chute are designed to minimize disturbing visual stimuli to the animals and are designed to prevent animals from attempting to jump or climb walls and gates and are spaced to prevent turning while in the transfer corridor. When animals enter the final corridor section, movement into the restraint device is affected by means of a sliding wall pushed up behind the animal. After the subject has been restrained, the system allows for the animal to either return to the animal holding pens or be moved into a padded induction/recovery room connected to a medical treatment suite. When in the restraint device, adjustable hydraulic pressure allows the animal to be squeezed laterally between padded walls and then lifted so that its feet cannot gain purchase on the ground, similar to the action of a drop floor restraint system. At this stage, movement of the animal is limited to dorsoventral flexion of the back and neck. Movement is further reduced by restraining the head. The Tamer™ itself allows access to the head and neck, the muscles of the thoracic and pelvic limbs, and provides access to the perineal area for rectal or vaginal examination. In an attempt to reduce the adverse effects of any restraint episode, animals are fed preferred feed (alfalfa) following return to their holding stalls whenever possible.

Methods

In order to evaluate the stress associated with the use of this handling system, blood samples were collected to determine the plasma concentration of cortisol during a series of reproduction research studies in eland antelope. Initially, 10 adult, female elands underwent estrous synchronization and superovulation in preparation for non-surgical embryo collection. Exogenous hormone administration and sample collection occurred nine times during the 17-day trial. In addition to per-rectum fecal collection carried out at these times, jugular venipuncture was performed on each animal, and serum and plasma were collected and subsequently evaluated for progesterone and cortisol, respectively. Plasma and serum were stored at -20°C until the content of cortisol and progesterone could be determined by radioimmunoassay.^3,4,7 For specific techniques, see Note.

Despite the fact that corticoid levels on their own may not accurately measure stress,⁵ corticoids are still considered reliable indicators of stress. In this study, the difficulty of interpreting single cortisol values was obviated by the collection of multiple samples and the evaluation of trends. For the collection of further research data and to rule out the possibility that any significant cortisol reduction was a result of adrenal exhaustion, we initiated a follow-up project which involved the regular collection of blood and fecal samples from a subset of the group of elands (n=5) handled in the same way, for approximately 8 months, to evaluate endocrine profiles and estrous cyclicity.

Stress associated with multiple physical restraint sessions was evaluated in the following ways:

1.  Subjective evaluation of animal’s adaptation to regular handling during the trial period

2.  Stress evaluation—plasma cortisol evaluated over 17-day trial period

3.  Endocrine profile evaluation over 8-month period from subset (n=5) of original group

Results

1.  Subjective evaluation: During the initial 17-day trial period, all animals appeared to tolerate regular manipulation, medical treatment and blood and fecal collection with no untoward effects. It became apparent that the more experienced and skilled the operators, the easier and quicker the operation proceeded. As animals became familiar with the procedures of the trial, speed and efficiency of restraint, sample collection and drug administration improved significantly.
Following a restraint episode, animals were observed for a short period to ensure normal behavior and appetite. It was apparent that animals rapidly became accustomed to the physical requirements of the trial.

2.  Stress evaluation: During the initial trial period, each animal was investigated by assessing physiologic stress response through the evaluation of plasma cortisol trends. Of the 10 animals evaluated, seven (70%) showed a decreasing trend in plasma cortisol values over the 17-day study period, two animals (20%) showed no significant change, with one animal (10%) showing a slight increase. First sample cortisol values varied from 1.3831 ng/ml to 8.4810 ng/ml (x=3.6337 ng/ml), whereas cortisol values from final samples varied from 1.4039 ng/ml to 6.8447 ng/ml (x=2.6877 ng/ml). Morton, et al. investigated capture-associated plasma cortisol levels in Zimbabwe and found a mean of 3.52 ng/ml for 20 wild-caught, physically restrained elands.⁶ In this investigation, the trend of decreasing plasma concentration of cortisol in 70% of the animals is an indication that animals became familiar with the requirements of the program and were not unduly stressed by regular physical manipulation.

3.  Endocrine profile evaluation: Following the 17-day trial period, a subset (n=5) of the group of elands being investigated was selected to undergo further evaluation over a longer time period. Five animals were manipulated through the handling system three times per week for a period of 30 weeks. At each restraint episode, blood and fecal samples were collected and stored for later evaluation. After 20 weeks, a mature male eland was placed with the group; effect on female endocrine profiles was recorded, and gestation periods were determined following birth of offspring. Despite being intensively manipulated through the Wilds’ handling system for a period of more than 7 months, all five study elands showed evidence of regular estrous cyclicity with a mean cycle length of 23.46 days, and all five animals conceived and became pregnant with a mean gestation length of 272 days. It is well recognized that ‘management’ stressors can affect both cycle length and time of ovulation in domestic hoofstock.⁵ These results indicate that the management stressors associated with handling animals in the manner described above appeared not to be significant. Plasma cortisol values associated with restraint during this follow-up trial are currently being determined and were not available at the time of writing.

Conclusion

We recommend the use of an appropriate and safe handling system that incorporates a means of physical restraint as described above, as a suitable method for the regular manipulation of large nondomestic hoofstock species such as eland. When carried out in an efficient and professional manner, regular manipulations are not unduly stressful nor detrimental to the health of the animals, as evidenced by decreasing trends in plasma cortisol values, the presence of regular estrous cycles, and the ability to conceive and produce healthy offspring.

Note

Titrated hydrocortisone and progesterone were purchased from New England Nuclear, Inc. (Boston, MA, USA). The cross-reactivity of the cortisol antisera (Pantex, Inc., Santa Monica, CA, USA) with corticosterone, deoxycorticosterone, progesterone, androstenedione and estradiol was 60%, 48%, 0.01%. 0.01% and 0.01%, respectively. The sensitivity of the cortisol RIA was 31 pg/tube. The intra- and inter-assay coefficients of variation were 4.7% and 8%, respectively. The cross-reactivity of the progesterone antisera (GDN 337; obtained from Dr. G.D. Niswender, Colorado State University, Ft. Collins, CO, USA) was 1.1%, 0.1%, 0.3%, 0.3%, 0.2%, 0.2% with pregnenolone, cortisol, estrone, estradiol, testosterone, and androstenedione. The intra- and inter-assay coefficients of variation were 7.3% and 8.9%, respectively. The sensitivity of the progesterone RIA was 31 pg/tube. Radioimmunoassay data were processed by the AssayZap program (Biosoft, Ferguson, MO).

Literature Cited

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4.  Kemper Green, C.N., D.A. Hawkins, A. Rocha, J.W. Tanner, P.G. Harms, D.W. Forrest, and T.H. Welsh, Jr. 1996. Temporal aspects of ovarian follicular growth and steroidogenesis following exogenous follicle-stimulating hormone in Angus heifers. Anim. Repro. Sci. 45. Pp 157–176.

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7.  Willard, S.T., J.A. Carroll, R.D. Randel, and T.H. Welsh, Jr. 1995. In vitro cell culture and adrenocorticotropin secretion by Indian blackbuck antelope (Antilope cervicapra) anterior pituitary glands collected under field conditions. J. Zoo Wildl. Med. 26(2). Pp. 252–259.