Abstract
Successful captive elephant management is a priority among zoo and wildlife organizations worldwide. Captive populations have been traditionally maintained by collecting from the wild, transport of captive females on long-term breeding loans, and to a lesser extent, by management of males on-site, which can be unrealistic for many zoos. Conservation and safety concerns, along with the growing acknowledgment among elephant caretakers that removing females from their familiar social groupings for breeding loans can cause distress, have all contributed to the need for development of assisted reproductive techniques. Demonstration of a successful artificial insemination (AI) would open up many possibilities for captive elephant management, including the collection of genetic material from the wild for integration into captive populations once semen cryopreservation techniques have been perfected. A new technique involving the application of ultrasonography for reproductive assessment and AI has been implemented in the elephant management program of several zoos. The technology for this project was developed at the Institute for Zoo Biology and Wildlife Research in Berlin in cooperation with the company A. Schnorrenberg.1 The AI component of this project involved simultaneous imaging by ultrasonography and endoscopy for verifiable semen placement. Both these components have never been accomplished together in an elephant AI. The insemination technique is noninvasive and has resulted in verifiable sperm deposition directly into the cervix.
Ultrasound-guided AI has been attempted in four nulliparous African cows at the Indianapolis Zoo: “Ivory” (16-year-old), “Tombi” (22-year-old), “Kubwa” (22-year-old), and the Vienna Tiergarten Schönbrunn “Sabi” (13-year-old). Additionally, the uniparous Asian cow “Shanthi” (23-year-old) was artificially inseminated at the National Zoo. All five females are wild-born.
The reproductive hormone levels (progesterone [P4]; luteinizing hormone [LH]) have been monitored from blood samples taken from their ear veins on a routine basis. All five females were excellent candidates for this project for several reasons:
1. They are of prime breeding age.
2. They are extremely calm-mannered, tractable and well-trained.
3. They are in very good general and reproductive health.
4. They have been palpated both vestibularly and rectally on a routine basis and have shown no signs of distress or injury during these procedures.
The technique for transrectal ultrasonography in elephants is performed in standing or laying position without the use of tranquilizers, anesthetics, or restrictive devices. A real-time B-mode ultrasound scanning system was used. For visualizing the caudal component of the urogenital tract (vestibule, urethra, vagina, urinary bladder, cervix, caudal corpus uteri), a 3.5-MHz transducer was manually introduced into the rectum with ultrasound gel for coupling. To visualize the cranial component of the genital tract (cranial corpus uteri, uterine horns, ovaries, surrounding tissues), a 5.0–7.5-MHz transducer is attached to a specially adapted extension and guided manually into the rectum. Ultrasonography can provide valuable information on ovarian activity, uterine integrity, and reproductive disease or dysfunction.2,3 It can be used to visualize structures of the entire reproductive tract.
Ultrasonographic examinations of the five AI candidates revealed that there were no indications of reproductive pathology in the urogenital tract or in the ovaries. Both endocrine data and ultrasonographic images were used to determine the timing of AI trials. In preparation for the AI attempts, the females were monitored daily for circulating levels of P4 and LH. Two LH peaks,4 separated by 19–21 days, were detectable in their estrous cycle, with the second peak being the ovulatory LH surge. Detection of the first peak provided us with a 3-week window to prepare for the inseminations. Transrectal ultrasonography was employed daily in this time period to identify morphologic changes in the vagina and endometrium and to characterize developing ovarian structures during the follicular phase. Ultrasonography allowed the visualization of follicle growth and maturation and the development of the Graafian follicle. The ruptured ovulatory follicle and corpus hemorrhagicum could also be visualized. The visualization of these ovulatory events has never been possible before.
The semen donor for the AIs in “Tombi” and “Kubwa” was a 20-year-old African bull named “Dale”. He is owned by Jo-Don Farms currently being housed at the Kansas City Zoo. The ejaculate for the insemination in “Ivory” was collected from a 16-year-old African bull with the name “McLean” at Disney’s Animal Kingdom. The selected breeding partner for the African cow “Sabi” at the Vienna Tiergarten was a 16-year-old bull named “Tembo” housed at the Colchester Zoo. The insemination in the Asian female “Shanthi” was performed with semen collected from the 13-year-old bull “Calvin” (African Lion Safari) and the 35-year-old bull “Onyx” (Dickerson Zoo). In general, the semen was collected by rectal palpation of the accessory glands sometimes combined with manual penile stimulation.6 Extended semen was transported by air at 4°C in a refrigerated vessel. The samples were warmed to 37°C prior to insemination. Samples used for AI trials ranged in motility status from 5–95%.
No sedation or physical restraint was used for the insemination protocol. Sterile technique was employed throughout the procedures. A balloon catheter was inserted in the vestibule (Canalis urogenitalis) to slightly distend the reproductive tract for optimal visualization and placement of an endoscope and insemination catheter. Both endoscopic and ultrasonographic visualizations were used to guide the placement of semen deep into the cervix if possible. The actual introduction of semen was monitored ultrasonographically to verify its placement. The time needed for an AI procedure varied from 10 minutes to 2 hours at maximum. Olfactory cues, such as semen, urine, feces, and temporal gland secretions were presented to the AI candidates before and after the AI series. They showed strong reactions to these stimuli, often rumbling and pelvic thrusting. They have shown no adverse behavioral, physical, or physiologic effects of this AI program.
Following the AI procedures, the females were monitored endocrinologically for P4 on a weekly basis. The first ultrasonographic examinations3 were performed about 8 weeks after the insemination. The preliminary results of this project are two successful AIs. The 16-year-old female African elephant “Ivory” and the 22-year-old “Kubwa” are pregnant. The AI in the 22-year-old African female “Tombi” was not successful due to poor semen quality (low motility and concentration). There were no results regarding the AIs in the cows “Sabi” (13-year-old) and “Shanthi” (23-year-old) before the abstract was submitted.
Artificial insemination (AI) is one of the most effective methods for improving the breeding success of domestic species. But for 35 years, different AI methods have never produced a confirmed elephant pregnancy until the year 1998. The introduction of AI in captive elephant breeding programs is enormously important in terms of the ultimate viability of assisted reproduction for use in the entire captive population of elephants—most of which may never have the opportunity for natural conception. The Dickerson Park Zoo, one of the collaborators in this project, announced the first successful insemination of a primiparous female Asian elephant in June of 1998.5 Last year, a total of three elephants became pregnant by AI in North America. “Kubwa” and “Ivory” are the first virgin elephant cows to be successfully impregnated by artificial insemination. The combination of a reproductive assessment program with a newly developed insemination technology resulted in this groundbreaking success. This project has been a team effort, requiring the cooperation and expertise of many individuals: curators, keepers, researchers, pathologists, volunteers, educators, and public relations specialists.
Acknowledgments
The authors are grateful for the assistance from the elephant staff of the Indianapolis Zoo, Smithsonian, National Zoological Park, and Vienna Tiergarten Schönbrunn for training the female elephants to stand unrestrained for the AI procedure. The authors thank the elephant staff of the Kansas City Zoo, Disney’s Animal Kingdom, Dickerson Zoo, Colchester Zoo, and the African Lion Safari for the semen collections.
Literature Cited
1. Hildebrandt, T.B. and A. Schnorrenberg. 1996. Besteck zur künstlichen Besamung von Elefanten. Deutsches Patentamt, Offenlegungsschrift, DE 196 06 925 A1.
2. Hildebrandt, T.B., F. Göritz, N.C. Pratt, D. Schmitt, J.L. Lehnhardt, R. Hermes, S. Quandt, J. Raath, G. West, and R.J. Montali. 1997. Assessment of health and reproductive status in African and Asian elephants by transrectal ultrasonography. Proc. Annu. Meet. Am. Assoc. Zoo. Vet. 207–213.
3. Hildebrandt, T.B., and F. Göritz. 1998. Use of ultrasonography in zoo animals. In: M.E. Fowler, and R.E. Miller (eds.): Zoo and Wild Animal Medicine. Current Therapy 4. W.B. Saunders Company, Philadelphia, 41–54.
4. Kapustin, N., J.K. Critser, D. Olson, and P.V. Malven. 1996. Nonluteal estrous cycles of 3-week duration are initiated by anovulatory luteinizing hormone peaks in African elephants. Biol. Reprod., 55: 1147–1154.
5. Schmitt, D.L. 1998. Report of a successful artificial insemination in an Asian elephant. Proc. 3rd Intern. Elephant Res. Symp., p. 7.
6. Schmitt, D.L., and T.B. Hildebrandt. 1998. Manual collection and characterization of semen from the Asian elephant (Elephas maximus). Anim. Reprod. Sci. 53: 309–314.