Abstract

Pathology Review

A review of Siberian tiger (Panthera tigris altaica) pathology records from 1915–2000 has been completed. The mean age at death was 12.1 (+/- 5.9) years, with females living longer than males (12.8 vs.11.3 years). Neoplasm was the most common diagnosis on necropsy, occurring in 25.5% of the reports. Degenerative musculoskeletal problems were noted in over 20% of the cats and renal disease was seen in over 25%. A complete review of the data is in preparation for presentation at the American Association of Zoo Veterinarians annual meeting in 2005.¹⁰ Records are incomplete and any zoos that have not submitted information should contact Albert Lewandowski, DVM.

Vaccination

The veterinary advisor for the Tiger SSP recommends the vaccination of normal healthy tigers for feline rhinotracheitis, calici virus, panleukopenia, feline leukemia, rabies, and canine distemper with killed vaccine. Only killed vaccines should be used in tigers, not modified live vaccines due to the potential risk of inducing disease. The following vaccination protocol is suggested for use in all Tiger SSP managed animals:

• 1 ml given intramuscularly of Purevax Ferret Distemper Vaccine, Merial, Inc., Athens, GA, USA. (Commercial killed vaccines for canine distemper are not available, this recombinant canarypox vectored vaccine has been used extensively in tigers with no observed problems.)
• 1 ml given intramuscularly of Fel-O-Vax LV-K, Fort Dodge Laboratories Inc., Fort Dodge, IA, USA.
• 1 ml given intramuscularly of Purevax Feline Rabies Vaccine, Merial, Inc., Athens, GA, USA, or Imrab 3 from Merial or other killed rabies vaccines.

Some institutions use 2 ml doses due to the greater body weight of tigers. Although there is no direct evidence that this is more effective, it also does not do any harm. Animals never before vaccinated should receive at least two and preferably three booster vaccinations approximately 3 weeks apart after 6 weeks of age. Previously vaccinated animals should receive an annual booster. There are no vaccines, including those listed above, that are legally approved for use in non-domestic felids. This is particularly relevant with rabies vaccines where human exposure through bites may occur, especially in privately owned animals. We do not know how protective these vaccines actually are or statistically how effective they might be.

It is known that most species of large felids are susceptible to canine distemper virus and the virus should be regarded as a significant potential threat to zoo populations.^{1,2,7,9,11,13}

Training Programs

During the past 3 years, professionals from the Wildlife Conservation Society (WCS) Siberian Tiger Project, the Henry Doorly Zoo, and WCS zoo and field program professional staff, as well as other groups have been active in building capacity in the Russian Far East (RFE) by training professional Russian tiger protection personnel who are required to respond to problem tiger situations. This is a group of trained Russian professionals that has the responsibility and capacity to respond with a variety of tools to problem tiger situations involving tiger conflict with humans. Through a series of workshops with their North American counterparts, Russian personnel responsible for dealing with problem tigers received intensive training in techniques dealing with this problem including wildlife health and disease management (including necropsy technique), immobilization and safe animal handling practices, and methods of animal-human conflict resolution. The two groups worked together sharing ideas and procedures to develop methods for coping with various aspects of the problem tiger issue. This training was made up of four workshops, carried out in Russia and the United States.¹² An advanced program is planned for the fall of 2004. This project has been supported by the Trust for Mutual Understanding.

Contraception (This Section is Taken From the 2002 Tiger SSP Report)

Tiger Contraception Recommendations

Genevieve Dumonceaux (Busch Gardens, Tampa Bay) and Douglas Armstrong (Henry Doorly Zoo and Tiger SSP Veterinarian Advisor)

While there are several methods of contraception in felids being investigated currently, none are currently useable. The recommended methods for contraception of large cats including tigers are as follows:

Physical Separation

Typically, the safest, most effective, reversible method with least risk of side effects. Available space, resources, and facilities tend to be the limiting factors with this management method.

Male Vasectomy

This involves the surgical removal of a section of each vas deferens to prevent transmission of semen to the female. Testicles are left intact. This method is very effective when performed properly. It does not inhibit testosterone-related behavior. It is not reversible and incurs an anesthesia and surgical risk.

Female Tubal Ligation

This is tying the fallopian tubes to prevent transmission of the sperm to oocytes at ovulation. The entire reproductive tract is left intact. This may or may not be reversible depending on technique, animal involved, and the skill of the surgeon. This technique does not inhibit female estrogen-related behavior. This method incurs anesthetic risk and laparoscopic or surgical risk depending on the method used.

Neutering

This involves castration of the male or spaying the female for complete and permanent sterilization. This method is recommended primarily for animals of little or no future genetic value. It is 100% effective for contraception. There is anesthetic and surgical risk with this procedure.

Melengestrol Acetate Implants

Currently still the method of choice for temporary, reversible, minimally invasive contraception. This has proven to be successful as a primary means of contraception in felids and primates. This method is recommended for use for a maximum of 2 consecutive years at a time to minimize the risk of development of reproductive pathology. Repeated, continuous use of implants significantly increases the pathology risk. Pregnancy following or between implants may reduce the risk. This method does incur brief anesthetic and surgical risk.

Methods Under Investigation

Leuprolide Acetate Injections

An option for cat contraception but still very expensive at about $600 per injection per Dr. Asa. Details on the use of this agent are still pending discussion with Dr. Briggs who has used it in various carnivores.

Deslorelin Implants

As GnRH analogs this drug has potential for contraceptive action in the big cats. However, currently it is no longer available due to changes with companies manufacturing the product and FDA issues. Undetermined if it will be available in the future. Investigators including Dr. Cheri Asa of St. Louis Zoo are looking into other GnRH analogs but no available agents yet.

Zona Pellucida Vaccine

Has been investigated in domestic cats and shows no immunocontraception in them per one study from University of Georgia. More information forthcoming.

Summary

There are still no 100% effective, safe, and reversible methods for contraception in large cats. Still the method of choice seems to be MGA implants as a temporary means of contraception but with significant side effects if used beyond 2 years. Investigations continue in better, longer term safer means of contraception.

Assisted Reproduction

In 1990 and 1991 single successes were achieved using fresh semen with each of the assisted reproduction techniques of in vitro fertilization⁴ and by laparocopic artificial insemination^5,6 at the Henry Doorly Zoo in a cooperative project with the National Zoo and the Minnesota Zoo. The birth of Siberian tiger cubs following transvaginal artificial insemination with fresh semen was reported in 2000 by a team in Portugal.² There were no subsequent successes with any of these techniques in this species, in spite of multiple attempts with each until 2003 when a single cub was born at the Henry Doorly Zoo. The pregnancy had been produced by intrauterine insemination with fresh semen introduced to the uterus by laparoscopy. The cub was born after a 107-day gestation and had to be hand raised due to maternal neglect. It subsequently died at approximately 3 weeks of age due to pneumonia.

At the present time, we believe that the primary problems preventing repeatable success with these techniques are attributable to the use in tigers of hormones derived from other species (human, equine, bovine) to manipulate the reproductive cycles. In essence the hormones used historically in these projects seemed to decline in effectiveness after initial use in any individual animal, possibly due to immunologic responses although this is not confirmed. In order to resolve this issue, we collected the pituitary gland from both an Amur tiger (Panthera tigris altaica) and from a snow leopard (Panthera uncia), extracted follicle stimulating hormone (FSH) and luteinizing hormone (LH) from the pituitaries and then determined the amino acid sequence for the FSH and LH for each of these species. These sequences were then compared to the known FSH and LH sequences for other species including canine, bovine, porcine, equine, ovine, and human. The protein structures for pig most closely matched the sequence for tiger hormones of all of those that were commercially available. Porcine FSH and LH were evaluated for efficacy in tigers during trials at the Henry Doorly Zoo. Although improved over hormone regimens utilized previously, the regimen of porcine hormones still proved less than ideal. The cats were inconsistent in the formation of corpora lutea and abnormalities in ultrastructure were found during electron microscopic evaluation of the oocytes.⁸

Concurrently during the above trial, a second project was undertaken to produce tiger hormones via transfection of cell cultures with the DNA sequence to produce tiger FSH and tiger LH. Plasmids containing the DNA sequences for the FSH and LH hormone subunits of tigers were produced and were used to transfect Chinese hamster ovary cell lines and cat kidney cell lines with these sequences. These cells were cultured, lines were selected for maximum hormone production by Western Blot analysis and these were propagated. Currently the effluent from these selected cell lines is in the process of being assessed for biologic activity in vitro in rat Leydig and rat granulosa cell bioassays. Large-scale production is being investigated by the National Cell Culture Center in Minnesota. Additional work remains, particularly determining the steps needed to produce purified hormone for injection. Initial hormone purification was done using a FLAG Tag system. It is not clear at this time whether additional purification steps will be needed or not. This project will have wide application to improve the success of assisted reproduction techniques in a variety of endangered cat species besides tiger. Initial support for this hormone production project came from the Morris Animal Foundation.

Literature Cited

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4.  Donoghue A, Johnston L, Seal U, Armstrong D, Tilson R, Wolf P, et al. In vitro fertilization and embryo development in vitro and in vivo in the tiger (Panthera tigris). Biol Reprod. 1990;43:773–744.

5.  Donoghue A, Byers A, Johnston L, Armstrong D, Wildt D. Timing of ovulation after gonadotrophin induction and its importance to successful intrauterine insemination in the tiger (Panthera tigris). J Reprod Fertil. 1996;107:53–58.

6.  Donoghue AM, Johnston LA, Armstrong DL, Simmons LG, Wildt DE. Birth of a Siberian Tiger cub (Panthera tigris altaica) following laparoscopic intrauterine artificial insemination. J Zoo Wildl Med. 1993;24:185–189.

7.  Fix AS, Riordan DP, Hill HT, Gill MA, Evans MB. Feline panleukopenia virus and subsequent canine distemper virus infection in two snow leopards (Panthera uncia). J Zoo Wildl Med. 1989;20(3):273–281.

8.  Gjorret JO, Crichton EG, Loskutoff NM, Armstrong DL, Hyttel P. Ultrastructure of oocyte maturation, fertilization and early embryo development in vitro in the Siberian tiger (Panthera tigris altaica). Mol Reprod Dev. 2002;63:79–88.

9.  Gould DH, Fenner WR. Paramyxovirus-like nucleocapsids associated with encephalitis in a captive Siberian tiger. J Am Vet Med Assoc. 1983;183:1319–1322.

10.  Lewandowski AH. Preliminary studies on the morbidity and mortality of Siberian tigers (Panthera tigris altaica). In Preparation.

11.  Montali RJ, Bartz CR, Teare JA, et al. Clinical trials with canine distemper vaccines in exotic carnivores. J Am Vet Med Assoc. 1983;183:1163.

12.  Quigley KS, Armstrong DL, Miquelle DG, Goodrich JM, Quigley HB. Health Evaluation of wild Siberian tigers (Panthera tigris altaica) and Amur leopards (Panthera pardus orientalis) in the Russian Far East. In: The American Association of Zoo Veterinarians Annual Conference, Orlando, FL, Sept. 18–23, 2001:179–182.

13.  Roelke-Parker ME, Munson L, Packer C, et al. A canine distemper virus epidemic in Serengeti lions (Panthera leo). Nature. 1996;379:441–445.