© Springer International Publishing Switzerland 2017Walter K.H. Krause and Rajesh K. Naz (eds.)Immune Infertility10.1007/978-3-319-40788-3_18
18. Immune Contraception in Wildlife Animals
Leibniz Institute for Zoo and Wildlife Research, PF 700430, Berlin, D-10324, Germany
Fertility control by immune contraception is suggested to offer a long-term, effective and humane approach for reproduction control in captive animals as well as to reduce free-ranging wildlife populations. The chapter presents a summary of target species for reproduction control by immunization, the available vaccines for wildlife animals and prospective delivery methods. Based on published literature, health issues as well as behavioural changes and population level effects of contraception in wildlife animals are discussed.
18.1 Immune Contraception in Wildlife Species
18.1.1 Captive Population
Not only is there a need of reproduction control in captive animals, but also exotic animals in captivity serve as models for the establishment of contraceptive approaches for wildlife.
Because improved animal husbandry and veterinary care has led to a low adult mortality and an increase in longevity, especially in large ungulates and carnivores and even species managed in breeding programs, high quality enclosures are consequently overcrowded. This results in the demand for population control. Prevention of offspring (sterilization or contraception) is an alternative to the elimination of captive animals (euthanasia or transfer to other institutions) .
In addition, zoos usually do not share the same limitations as wildlife managers, thus making it possible to apply a wider variety of contraceptive techniques, or even allow research on contraceptive approaches which would not be possible on wildlife animals. Thus, not surprisingly the first anti-fertility vaccinations were applied to captive wildlife animals [2, 3].
Twenty-five years after first attempts on immunocontraception in captive wildlife animals , a huge number of animals and species has been treated with a variety of contraception measures including contraceptive vaccines. Nevertheless, knowledge on efficacy and treatment protocols remains patchy. To meet this challenge, in North America, and also in Europe, a centralized database has been compiled to produce guidelines for a variety of species (www.egzac.org). The recommendation for captive animals (www.stlzoo.org/animals/scienceresearch/contraceptioncenter) lists two options of anti-fertility immunocontraception: vaccines against porcine zona pellucida proteins (PZP) and against the GnRH . Both vaccines still require repeated injections and are limited to captive or small populations of free-ranging wild animals.
18.1.2 Wildlife Population
Reproductive control is important for the management of certain wildlife, and ironically also endangered species, particularly in the increasingly common situation in which the size and the nature of habitat is restricted by man’s encroachment . Additionally, some wildlife species have adapted successfully to changing environmental conditions and concentrated in large populations significantly impacting on their habitats or their prey species and served as reservoirs for infectious diseases, making them “pest species”.
The challenge is to develop acceptable and sustainable methods to reduce and maintain populations of these animals and maintain them at levels that minimize their impact on environment. There is also a major need to control population sizes in non-indigenous animals and/or imported species such as the raccoon and American mink in Europe, fox in Australia. At the population management level with free-ranging species, the primary focus has been on wild horses, urban deer, bison, and African elephants .
18.104.22.168 Pest Species
Pest mammals have severe economic, environmental and social impacts throughout the world . Methods of immune contraception are being investigated for small rodents, in particular for rabbits and house mice, and for overabundant marsupial species, like brushtail possum in New Zealand . Most small rodent species are characterized by high reproductive rate. Therefore, fertility control need to be adapted to their reproduction strategy (r-strategy) which is characterized by a high rate of reproduction, high juvenile mortality, and strongly fluctuating population sizes within fluctuating habitats. Population models which were developed to predict possible outcomes of fertility control show that at least 80 % of females will need to be infertile and that this infertility will need to be permanent .
Virally vectored immunocontraception (VVIC) has been proposed as an economic way to achieve this [10, 11]. Although VVIC may have the advantage of self-regulating depending on the density of the target species , biological safety and regulatory concerns must be overcome in future before VVIC can be applied for field testing [7, 13, 14]. Therefore, specific bait-delivered fertility control is more likely to be used in the near future .
22.214.171.124 Alien Species
Alien species are any species that have spread beyond their natural range into new locations as a result of human activity. Invasive alien species are species that have some advantage over native species. These advantages are often enhanced when aliens move into ecological niches and thrive because outside their natural environments they are not held back by natural predators, parasites, disease, or competition in the way that native species are. Therefore, there is a major need to control population sizes in non-indigenous animals and/or imported species such as the grey squirrel  and several deer species in Europe , and fox in Australia . In case of some alien species, a full eradication from their new habitats might be considered. The IUCN Invasive Species Specialist Group (www.issg.org) provides support for the control of invasive species through the Global Invasive Species Programme (GISP) (www.gisp.org). The expertise about eradication and control varies widely, with a wealth of experience available in a few areas, such as on islands (for review .
126.96.36.199 Overabundant Wildlife
Overabundance of wildlife often has nothing to do with biological carrying capacity, when population reduces the growth rate at which food resources are replenished. Wildlife is considered to be overabundant (ecological overabundance) when a population is so dense that it threatens the persistence of other species or when a population is so dense that it becomes unacceptable to humans (societal overabundance). In this respect, a number of wildlife species have become overabundant on a local or regional scale throughout the world. Traditionally overabundant wildlife is controlled by hunting and trapping, but this may be restricted or infeasible in parks and suburban areas. Application of wildlife fertility control is suggested for use in urban or suburban areas (e.g. deer species, wild boars) and in situations where immigration is limited and lethal control is restricted (e.g. elephants in national parks).
188.8.131.52 Human Wildlife Conflicts
Population control of wildlife is also considered in case of “human wildlife conflicts”, which reflect any conflict in interaction between wild animals and people. It occurs when growing human populations overlap with established wildlife territory, creating reduction of resources or life to people and/or animals. The conflict takes many forms ranging from loss of life or injury to humans, and animals both wild and domesticated, to competition for scarce resources to loss and degradation of habitat. Ethical considerations regarding humane treatment of animals are shaping public attitudes toward acceptable methods of mitigating human-wildlife conflicts . Conflict management strategies usually compromise lethal control and translocation . Recent management approaches also consider fertility control to reduce human-wildlife interactions .
184.108.40.206 Transmitters of Zoonoses
Zoonoses are infectious diseases that can be transmitted between humans and animals, both wild and domestic. Approximately 75 % of recently emerging infectious diseases that affect humans are diseases of animal origin, and approximately 60 % of all human pathogens are zoonotic. During the last 40 years, new epidemiological patterns have emerged as free-ranging wildlife have become progressively more involved in the epidemiology of both common and emerging infectious diseases of humans and domestic animals (for review . This has been seen in rabies, bovine tuberculosis and more recently in wild-boar classical swine fever  or brucellosis .
Offensive lethal control, however, failed to control animals, like badgers, wild boar and foxes for tuberculosis, classical swine fever and rabies, respectively. Culling these species reduced their populations to a logarithmic part of their growth curve, such that any losses due to culling were very quickly replaced by individuals that normally would have died due to population density pressures. Consequently, immune contraception in combination with disease vaccination is being discussed as an alternative strategy to lethal control.
18.2 Development of IC Vaccines for Wildlife Species
The challenges in the development and application of vaccine-based wildlife contraceptives are diverse and include differences in efficacy across species. Specific requirements for wildlife, both free-ranging and captive animal populations are defined by side-effects. For instance, a reduction of sexual/aggressive behaviour might be advantageous in zoo settings but not for free-ranging animals when the specific sexual behaviour, like herd hierarchy, should be retained. For wildlife animals, remote delivery is important to avoid stress of capturing and restraining animals for hand injection . In addition, the uncertain reproductive status at treatment should also guarantee safety of vaccines during pregnancy. Beyond the constraints imposed by the public and a host of regulatory concerns, there exists a real limitation for funding of well-designed programs that apply this type of fertility control .
The most widely tested immune contraceptive vaccine for wildlife species is based on developing antibodies to zona pellucida (ZP), which surrounds the mammalian egg. The PZP antigens are isolated from porcine ovaries obtained from slaughter house material. When PZP is injected into females others than pigs, the target species will produce antibodies against the antigen. These antibodies attach to the zona pellucida of ovulated eggs and cause steric hindrance which then blocks fertilization. Long-term application or hyperactive immune response, however, can cause ovarian failure and permanent sterilization.
The initial PZP vaccine was based with multiple-shot boostering . Since then several technologies have been developed to achieve efficiency with a single immunization [29, 30].
PZP vaccines and species-specific ZP (native/recombinant, whole/individual ZP) components have been investigated in various animal species (wild, zoo, farm, and domestic) (for review: ). PZP has been shown to be effective in a wide variety of ungulates [4, 31–33], equines [34, 35], elephants [6, 36–38] and some carnivores [32, 39, 40]. Recently it was also investigated in marsupials [41, 42]. It was shown that that the fertility of grey kangaroo, brushtail possum and koalas can be compromised by immunization against ZP antigens, but immunization with bacterial recombinant brushtail possum ZP3 did not reduce fertility in the koala.
Despite a high individual variability in immune response observed , the biological efficacy of PZP depends on species-specific antigenicity and immunogenicity of porcine zona pellucida [44–46], and therefore must be validated before it can be applied for contraceptive population control in a particular species.
18.2.2 GnRH Vaccine
Vaccination against gonadotropin-releasing hormone (GnRH) entails the administration of a modified form of the GnRH hormone in order to stimulate the production of anti-GnRH antibodies, which bind and inactivate the endogenous GnRH. The hormone (GnRH) stimulates the pituitary to secret the gonadotropins LH and FSH. Thus, immunization against GnRH disrupts the reproductive axis by depression of gonadotropin production in order to inhibit follicle growth, ovulation or spermatogenesis. In addition, vaccination against GnRH is the most promising alternative to castration for reducing male aggressive behaviour in captive adult animals . The molecular structure of GnRH is conserved between mammalian species. Therefore anti-GnRH vaccines are applied in a wide range of animals.
Studies on the use of GnRH vaccination for suppression of fertility, aggression and sexual behaviour show promising results in several domestic species, such as sheep , pigs , cattle  and horses . In particular, for population control of feral cats  and street dogs , GnRH is discussed as an ideal contraceptive target because it regulates pituitary and gonadal hormone responses in both males and females, thus suppressing nuisance behaviours associated with sex hormones in addition to preventing pregnancy.
GnRH vaccination has also been suggested to be applied in fertility control of overabundant wildlife, such as wild boar , white-tailed  or black-tailed deer , white Alpine sheep , prairie dog , and ground , fox  and tree  squirrels. Also for captive wildlife animals it is recommended and applied [62–64], in particular in connection to aggression and sexual behaviour suppression in males.
Suppression of steroid production by GnRH vaccines, however, is accompanied by effects which are characteristically for castration: changes in secondary sex characteristics, like antler growth in deer or the mane in lions, and therefore should be taken into account. In addition, safety trials should be performed to assure that non-reproductive function of the hypothalamus and pituitary gland are retained in treated animals.
18.2.3 Other Antigens for Immune Contraception
Most currently available targets for immune contraception interact at some point in the sequence of hormone synthesis or are involved in essential reproductive events, like ovulation, spermatogenesis, sperm or egg transport, or implantation . New molecular technologies are applied to generate vast peptides libraries that are screened for their potential impact on fertility.
A variety of proteins derived from sperm and oocytes have been experimentally assessed for their contraceptive potential in wildlife species [18, 66]. In particular, sperm antigens are suggested to act as species-specific immunogens, an essential prerequisite for oral vaccines applied in free-ranging wildlife. In this respect, a promising strategy appears to be the construction of immunogens that include repeated peptides from proteins involved in fertilization [67–69]. Multi-epitope contraceptive vaccines and preformed engineered antibodies of defined specificity may eliminate concern related to inter-individual variability of the immune response .
Fertilization-related antigens were isolated from germ cell plasma membranes  or are identified from cDNA libraries of oocytes or sperm cells . Also the identification epitope peptide by phage display was introduced to contraceptive vaccine development [73, 74].
Some of the sperm proteins already proved to be promising antigens for contraceptive vaccine include lactate dehydrogenase, protein hyaluronidase, Eppin or Catsper . After identification of the specific sperm fusion protein Izumo  it is discussed as a potential contraceptive antigen as well .
18.3 Delivery Methods
Fertility management is not yet a practical reality in wildlife management. Before it can be implemented it must meet the demands on risk assessment for health and behavioural effects in the target species, as well as any potential effects on non-target species . Immune contraception in wildlife requires the acceptable and safe application (e.g. humane use, environmental safety, target specificity) and knowledge of how to apply it strategically (e.g. where, when, how often, how intensively) .
The species-specific application can either be achieved through vaccine compound itself or the delivery method. Vaccine-developing strategy includes the identification of an anti-fertile antigen and the development of an effective vaccine composition including acceptable carriers, adjuvants and delivering systems . In free-ranging wildlife population, a large-enough proportion of the population must be reached to suppress growth  still ensuring a safe, effective and efficient delivery.
Delivery can be oral, by implant or hand injection after capture, or by dart in unrestrained populations. The following delivery methods had been used or suggested for wildlife species.
18.3.1 Parenteral Immunization
Attempts to use anti-fertile vaccines to stop breeding in wildlife animals date back to late 1980. Initial studies showed that pregnancies could be prevented by multiple-shot vaccines  containing PZP and Freund’s adjuvants. These were followed by studies showing that these vaccines could be remotely delivered effectively to free-ranging animals (horses, deer and elephants), but the requirement for repeated initial shots and annual boosters limited management application. During the following years the formulation of vaccines was improved steadily. Now vaccines are effective for several years with a single treatment [84–87].
Multiple booster treatment were replaced by microsphere particles or polymer-based controlled-released pellets [30, 88, 89] containing the antigen and the adjuvant. Immunogenicity of antigens was increased by tagging them to another protein, and implementation of different adjuvants [30, 90, 91]. The replacement of Freund’s adjuvant was aimed to prevent local and systemic side effects described in several wildlife animals .
Beside the progress made in delivery procedure, the parenteral immunization requires an individual approach to the animal and is therefore limited to captive or small populations of free-ranging wild animals. In this respect, an individual-based rotational vaccination was suggested in long-lived, social species (e.g. wild elephant) to stretch the inter-calving interval for each individual, preventing exposing females to unlimited long-term immune contraception use . It is also discussed to combine parenteral vaccination campaigns (e.g. for elimination of canine rabies) with immune contraception [94, 95] for population control.
18.3.2 Oral Delivery via Baits
Alternative contraceptive vaccines are actively being developed to enable large numbers of wild animals to be targeted by oral delivery of baits, when single animal’s treatment will be not effective or possible (pest or alien species). The distribution of baits and collecting remaining bait could be handled similarly to rodendicide baits . It is proposed that vaccines which will be delivered orally by baits will stimulate a mucosal immune response to the foreign antigen(s). Such a vaccine requires a detailed understanding of reproductive-tract mucosal immunity in target species. Oral contraceptive vaccines under consideration include viral or bacterial vectors and microencapsulated antigens . Although baits are increasingly used in wildlife management to deliver vaccines for disease control, a contraceptive oral vaccine is not available yet. In addition, safety requirements concern the transmissibility of the antigen in case of viral or bacterial vectors, the reversibility of the intervention within an individual animal and in animal populations, as well as the species specificity of the antigen used .
18.3.3 Nasal Inoculation
Vaccination based nasal inoculation resulted in a high antibody response in mucosal tissues, including genital tracts , and were suggested to avoid a pathogenic T cell activation , a side effect causing autoimmune destruction of gonads. Thus, the intranasal co-delivery may present a safe strategy for the development of contraceptive vaccine. Vehicles for potential nasal delivery of vaccines include bacterial ghost, virus-like-particles [101, 102] or nanoparticles .
18.3.4 Plant Based Contraceptives
An alternative approach to protein production using bacteria or virus presents the plant-based immune contraception especially for herbivore animals. Female possums vaccinated with immunocontraceptive antigens showed reduced fertility, and possums fed with potato-expressed heat labile toxin-B (LT-B) expressed mucosal and systemic immune responses to the antigen. This demonstrated that immune contraception was effective in possums and that oral delivery in edible plant material might be possible. Prior to attempts at large scale production, more effective antigen-adjuvant formulations are probably required before plant-based immunocontraception can become a major tool for population control of overabundant vertebrate pests .
18.3.5 Virally-Vectored Immunocontraception (VVIC)
One approach to deliver immunocontraceptive vaccines are self-disseminating agents as a viruses. This approach employs live genetically modified viruses to deliver immunocontraceptives and has proved successful under laboratory conditions. Under field condition, a virus may have the advantage of self-regulating depending on density of the target species  and can be species specific if the viral vector is species specific [66, 105]. However, despite a large number of studies on VVIC [14, 66], so far no product has been developed for field testing. The ability of an immunocontraceptive virus to control populations is not only compromised by several factors like sufficient transmission rate, competition with field strains of virus or its ability to induce infertility in the presence of field strains , there are also safety and regulatory concerns about maintaining the species specificity of the viral vector and other potential unexpected changes in such genetically modified virus. Once released, a vector cannot be recalled and may spread to regions where the original target species is not a pest . These constraints indicate that it is very unlikely that VVIC will be applied in near future .
18.4 Problems Connected with IC
18.4.1 Animal Welfare and Health Issues
Potential adverse effects of contraception may include harmful effects on pregnant animals, inhibition of parturition or dystocia, changes in ovarian structure or function, changes in sex ratio, changes in lactation or mammary glands, impact on fertility of young, changes in testicular structure or function, changes in secondary sex characteristics, changes in bodyweight, changes in behaviour, changes in annual breeding season, and other physiologic and pathologic changes . Reactions at injection sites and in lymph nodes are typical responses to injection of vaccines formulated as water-in-oil emulsions, especially those that contain mycobacteria [60, 108, 109].