As part of the program to develop contraceptive tools to control populations of over-abundant wildlife species, the NWRC has developed a single-injection gonadotropin-releasing hormone (GnRH) immunocontraceptive vaccine, GonaCon™. GonaCon™ has been tested and shown to provide contraceptive effects lasting 1-3 years in many pest species including white-tailed deer, domestic and feral pigs, bison, wild horses, cats, dogs, and California ground squirrels. GonaCon™ contains a GnRH peptide conjugated to keyhole limpet hemocyanin combined with AdjuVac™, an adjuvant also developed at the USDA National Wildlife Research Center. Immunization against GnRH prevents the circulating GnRH from stimulating the release of pituitary luteinizing hormone and follicle-stimulating hormone. This process of immuno‑neutralization of GnRH effects a temporary non‑surgical castration in both males and females. Contraceptive and behavioral effects of GonaCon™ are discussed for a variety of species.

The term “vaccine” has traditionally been associated with establishing immunity (antibodies) to a disease. This immunity is usually developed following administration of killed microorganisms. Disease vaccines typically require 1 to 3 injections, depending on the antigen design and efficacy of the vaccine. The effectiveness of the disease vaccine depends on the immune response developed by the host following exposure to the disease organism. The immunocontraceptive vaccine GonaCon™ is designed to produce immunity to the “self” hormone (GnRH), which is essential to reproductive activity in the mammal. Antibodies to GnRH reduce its biological activity resulting in infertility of both sexes. GonaCon™’s effectiveness as a single-injection immunocontraceptive wildlife vaccine depends on 4 factors. The first is the use of a large foreign mollusk protein in the GnRH conjugate. Second is the design of mollusk/GnRH protein conjugate that presents the GnRH antigen in a repetitious fashion. This design mimics the “danger signal” found in bacterial pathogens to which the animal has been previously exposed. Third is the addition to the vaccine of micrograms of Mycobacterium avium, which is ubiquitous in the environment and activates memory cells. The fourth factor is use of a water-in-oil emulsion, which provides a depot at the injection site, allowing a slow release of the vaccine. With this formulation, the vaccine is presented to the body as a “chronic infection”, even though it is not infectious. The granuloma that normally develops at the injection site plays a prime role in the host’s defense against this “chronic infection”. A WHO report on the use of the alum adjuvant in human vaccines states that “development of a small granuloma is inevitable with vaccines adjuvanted with aluminum, and is to be considered necessary to the efficacy of the adjuvant.” Researching GonaCon™ for use in companion animals, NWRC has looked at many different adjuvants intended to reduce the injection site reaction while at the same time retaining an effective vaccine. This paper reports on the role of the adjuvant and the injection site on the effectiveness of the vaccine.

GonaCon™ is a gonadotropin releasing hormone (GnRH) contraceptive vaccine developed at the USDA National Wildlife Research Center (NWRC) that contains keyhole limpet hemocyanin (KLH) coupled to GnRH. The vaccine is administered intramuscularly in an emulsion with the NWRC adjuvant, AdjuVac™. This first-generation vaccine, developed in 1998, has been very effective in contracepting every species tested. However, because of the high cost of KLH, NWRC has developed a second generation GnRH vaccine, replacing KLH with a less expensive mollusk protein. This second generation GnRH vaccine, called GonaCon-Blue™, is currently being tested in several species and has proven to be as, or more effective than the original GonaCon™. Scientists with the Reproductive Control Methods Project at the NWRC are also testing a recombinant GnRH antigen developed by Dr. Talwar of the Talwar Research Institute, India. The recombinant antigen was produced from plasmid gene expression grown in and extracted from E. coli. The antigen is combined with the NWRC AdjuVac™ to form an emulsion similar to the GonaCon™ preparation. The recombinant vaccine has been shown to be effective as a single or dual injection for contraception of adult female pigs in a short-term study at the Swine Research Facility at Penn State University. Although the long term effectiveness of the recombinant vaccine remains to be established, this small molecule of approximately 14,000 MW is of considerable interest because it may be effective in oral or transcutaneous applications. It could also be scaled-up for large field applications.

GonaConTM is a single-shot immunocontraceptive vaccine that targets the reproductive hormone gonadotropin releasing hormone (GnRH). The GnRH peptide used in the vaccine is secreted by the hypothalamus of the brain and is specifically known as luteinizing hormone releasing hormone. This peptide stimulates the synthesis and secretion of luteinizing hormone and follicle stimulating hormone by the anterior pituitary gland. These hormones in turn stimulate hormone and gamete production by the ovary. In earlier studies with deer, we showed that antibodies produced in response to a single shot of GonaCon inactivated endogenous GnRH and greatly reduced fertility, reproductive steroids, and associated behaviors for at least 2 years. In the present study, we report on contraceptive efficacy and behavioral observations up to 6 years after female white-tailed deer were given a single shot of one of several formulations of GonaCon. We compared the standard GonaCon made by conjugating the GNRH peptide to keyhole limpet hemocyanin (GonaCon-KLH) to one with the GnRH peptide conjugated to a blue mollusk protein (GonaCon-Blue) and to a 2-shot regimen with GonaCon-KLH. All GonaCon preparations were made into an emulsion with AdjuVacTM and administered to 5 mature female deer per treatment. The results showed that the GonaCon-Blue preparation significantly out-performed the other single-shot vaccines, as well as the 2-shot regimen of GonaCon-KLH, with efficacy of 80-100% for each of the 5 years in the study. Interestingly, expression of estrous behavior was minimal in Years 1-2 following treatment for all groups, but expression of estrus increased in later years of the study, even though does remained infertile. This expression of estrus and fertility was reversed with follicle stimulating hormone releasing hormone (FSH-RH) peptide conjugated to a mollusk protein and mixed as an emulsion with AdjuVac. This suggests that does possess an FSH-RH peptide, secreted by the hypothalamus, which modulates follicle development and resulting estrogen production and reproductive behavior.

Overpopulation of feral horses in several western states is an unquestioned problem. Current management strategies of removal and adoption are expensive, logistically challenging, and ineffective as a means of population control. We are testing three long-acting contraceptive approaches on feral Nevada mares. Modified reversible immunocontraceptive vaccines for gonadotropin releasing hormone (GnRH) and porcine zona pellucida (PZP) (SpayVac), and intrauterine contraceptive devices (IUDs), are being evaluated to determine: 1) their safety and efficacy for preventing pregnancy for multiple years, 2) whether the effects are reversible, and 3) whether there are notable contraindications. Preliminary data after 1 year suggest that IUD- and PZP-treated mares continue to exhibit breeding and estrus, while GnRH-treated mares are less likely to cycle. All mares in the GnRH and PZP treatments were infertile for the breeding season. Eighty percent of the IUD-treated mares were infertile; those mares that became pregnant likely failed to retain the IUD. A notable contraindication was that uterine edema normally observed in mares in the follicular phase of the estrous cycle was commonly observed in PZP-treated mares. Because administration of each contraceptive approach is different, and each has different effects and expected duration, one approach or a combination of approaches may be best suited for specific field applications. Subsequent years of this study should establish the efficacy and safety of one or more long-acting contraceptive approaches for feral horse population control.

The physiological mechanisms controlling ovarian follicular growth and ovulation involve a complex exchange of systemic signals and a localized exchange of molecules between the oocyte and surrounding somatic cells. It has been demonstrated that the oocyte itself plays an essential role in regulating these processes by secreting two key regulatory proteins: bone morphogenetic protein-15 (BMP15) and growth and differentiation factor-9 (GDF9). Natural mutations in their expression and vaccination against these growth factors have been shown to cause sterility in sheep. The aim of this 3-year study was to determine the effect of vaccination against GDF9 and BMP15 on fertility in female white-tailed deer. Does were randomly assigned to two groups (n = 10/group). Each doe received a primary vaccination followed by a booster 47 days later. After the first year, a subset of animals (n = 4-5/group) received an additional booster vaccination. Blood samples were collected at regular intervals to determine antibody titers and progesterone concentrations. Early pregnancy was assessed by ultrasound, and fawning was subsequently monitored. For the BMP15 group, over the 3 years 80%, 100%, and 75% of does fawned with fawning rates of 2.4, 3.5, and 3.3 fawns/doe, respectively. For the GDF9 group, 80%, 25%, and 25% of does fawned with fawning rates of 2.6, 1, and 2 fawns/doe, respectively. The fawning rate of untreated animals in the herd was 1.8 fawns/doe. Although all animals in the BMP15 group had high antibody titers, they were not made infertile, and they became more fecund. Most animals in the GDF9 group had high antibody titers, but fecundity was not affected the first year; however, they were made infertile in Years 2 and 3. Results from this study demonstrate that vaccination against GDF9 has potential to control fertility in deer. Further research will be required to determine the appropriate timing for administering the vaccine and the longevity of effect.

Controlling fertility of feral horses through the use of long-acting contraceptives or sterilization approaches has been championed as a reasonable and humane solution for addressing overpopulation problems in several western states. However, methods to accomplish long-term contraceptive efficacy of horses following a single treatment have been lacking. In fall 2002 and spring 2003, we initiated a study to compare the long-term efficacy of a single-shot contraceptive vaccine directed at gonadotropin releasing hormone (GonaCon™) to that of a single-shot vaccine directed at the zona pellucida (SpayVac) and to the use of intrauterine contraceptive devices (IUD). Both vaccines were administered with AdjuVac™, an adjuvant developed at the National Wildlife Research Center. The objectives of the study were to determine: 1) 3-year efficacy for preventing pregnancy, 2) whether the contraceptive effects are reversible, and 3) whether there are contraindications. The Nevada Department of Agriculture provided the feral mares, which were maintained at the Nevada State Penitentiary, Carson City, facility. Mares were dewormed and given health vaccinations annually. Eight untreated control mares were compared to 12 mares treated with SpayVac, 16 mares treated with GonaCon™, and 15 mares treated with copper-containing IUDs. Only 25% (2/8) of the control mares were not pregnant or infertile in the first year. All mares in the SpayVac group were infertile, and 94% (15/16) were infertile in the GonaCon™ group during the first breeding season. In Year 2, 80% (10/12) of the SpayVac-treated mares and 60% (9/15) of the GonaCon™-treated mares were infertile. In Year 3, 80% of the SpayVac mares and 53% (8/15) of the GonaCon™-treated mares were infertile. For IUD-treated mares, 80% (12/15) were infertile after Year 1, but only 29% (4/14) and 14% (2/14) were infertile after Years 2 and 3, respectively. For IUD mares that were infertile, it was possible to visualize the IUD by ultrasonography, leading us to conclude that mares that became pregnant had lost their IUDs. For mares given SpayVac, uterine edema was commonly observed. In Years 2 and 3, antibody titers for SpayVac were progressively lower compared to titers observed in Year 1.

GonaCon™ Immunocontraceptive Vaccine targets the reproductive hormone gonadotropin releasing hormone (GnRH) secreted by the hypothalamus of the brain. Antibodies produced in response to the vaccine inactivate endogenous GnRH, which in turn eliminates stimulation of the pituitary gland and gonads in males or females. The resulting “immunocastration” renders animals unable to produce reproductive steroids or gametes. In previous studies, we demonstrated that GonaCon™ was effective in impairing fertility for up to 3 years in female deer without apparent side effects. However, detailed post mortem evaluations were not done. To better understand the entire physiological response of deer to GonCon™ and to establish if contraindications were associated with its use, we undertook the present study. To evaluate toxicity and safety, 7 does were given the standard single injection GnRH-KLH vaccine dose (1000 µg) delivered IM in 1 ml using AdjuVac™ adjuvant and compared to 6 does given a single control saline IM injection and 6 does given 3 injections of GonaCon™ at 2-week intervals per dose. The study was conducted for 20 weeks. Does were blocked by weight and randomly assigned within blocks to treatment groups. Blood samples were drawn immediately prior to vaccination and at 5, 10, 15, and 20 weeks post immunization. Blood was assayed for LH, testosterone and progesterone, and anti-GnRH titers, as well as hematology and blood chemistry. At each sampling period, the general health of the doe was observed and the injection site was inspected for the formation of abscesses or other tissue reactions. At Week 20, all deer were euthanized and evaluated at necropsy by veterinary pathologists and samples of lymph nodes, reproductive organs, lung, liver, heart, kidney, spleen, and brain were taken for histology. Aside from granulomata formation at the injection site, there were no significant contraindications or toxic effects associated with GonaCon™.