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Introduction and General Information

"Successful reproduction wherein offspring are parentally reared to independence is a keynote to a biologically adequate animal facility and husbandry program." (B438.24.w24)

Reproductive management includes deciding that a particular species should be bred, choosing which individual animals to pair with one another, monitoring the reproductive health of individuals, providing appropriate social structures for breeding and parent rearing, artificial assistance with reproduction in some cases, and preventing reproduction when appropriate.

Decisions on whether animals of a particular species should be bred will depend on factors such as the conservation status of the species, its educational value, and/or its value as a model for a related, rarer species, as well as the space available for keeping animals of that species and related species with which it may "compete" for enclosure space.

Major basic factors which need to be considered if breeding is to occur successfully include appropriate enclosure design, including sufficient space, avoidance of overcrowding, providing suitable social groups, provision of psychological security and meeting behavioural needs; adequate nutrition; and prevention and control of disease. (B105.19.w6, B438.24.w24, J51.18,w1, P1.1970.w1). Further information on these aspects is provided in:

Note: Control and prevention of reproduction is as important in management as is encouragement of reproduction. Available accommodation in appropriate facilities is limited, and populations of animals in zoos need to be carefully managed to maximise the number of species which can be maintained in self-sustaining, genetically viable, populations. (B429.40.w40) 

Bear Consideration
Conservation status

When deciding whether or not bears should be bred in captivity, it should be remembered that five of the seven Ursus, Melursus, Helarctos and Tremarctos bear species are red-listed as Vulnerable (Melursus ursinus - Sloth bear, Tremarctos ornatus - Spectacled bear, Ursus maritimus - Polar bear and Ursus thibetanus - Asiatic black bear) or Data deficient (Helarctos malayanus - Sun bear). (W2.Jun06.w8)

Management

Reproductive management of bears needs to include consideration of space requirements both for breeding these large, solitary, species and for holding the cubs, including the likelihood that there will be appropriate enclosure space available for grown offspring either at the breeding institution or at another suitable location. (B407.w6, B432.w3)

Lagomorph Consideration
Conservation status
Management
  • Reproductive management of lagomorphs needs to consider space requirements for breeding and holding adequate numbers of animals which may be territorial or may be aggressive to one another in confined spaces. (J23.14.6, J51.19.w1, J332.10.w1)
  • If breeding an endangered species or subspecies for reintroduction, adequate holding spaces are required to maintain a large enough captive population to provide sufficient animals for release as well as enough individuals remaining in the captive population. For short-lived species such as Brachylagus idahoensis - Pygmy rabbit, this may be problematic. (J471.11.w1)
Ferret Consideration
Conservation status
Management
Bonobo Consideration Conservation status
  • Bonobos are endangered and on the Red List as endangered under criteria A4cd (version 3.1). (W2.Sept09.w1)

Management

  • Breeding management of bonobos should aim for genetically desirable offspring, while maintaining good social structures.
Published Guidelines linked in Wildpro

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Genetic Considerations and Breeding Programmes

  • For all species with a managed breeding programme (EEP, SSP etc.) the species manager/studbook keeper should be contacted for recommendations for breeding.
Breeding programme aims and priorities

Alongside efforts to preserve endangered species as self-sustaining wild populations, captive populations may be vital to ensure survival of some species and for preservation of genetic diversity. (B115.9.w13) Breeding programmes in zoos may be carried out to develop and maintain self-sustaining captive populations in order to allow continued exhibit of the species, or to conserve species or gene pools whose continued existence in the wild is doubtful. (P97.1.w34) Priorities of breeding programmes vary depending on the starting population and the aim of the programme.

  • Reproduction should start with a pair or group (as appropriate) of genetically unrelated animals. (B438.24.w24)
  • In species with very few founders, breeding programmes should aim to maximise reproduction, expanding the size of the population as rapidly as possible. (J23.27.w3)
    • This reduces loss of genetic diversity and minimises the risk of genetic loss through founders not breeding, or through loss of the population due to disease, accident, a distorted sex ratio, a small number of non-breeding females etc. (J23.27.w3)
      • Populations numbering less than 50 (or even 100) are particularly vulnerable to crashes due to disease, distorted sex ratio, natural disasters etc. (J23.27.w3)
      • Genetic diversity is important; genetic variation within a population is the basis for change within a population, allowing adaptation, and in an individual, the degree of heterozygosity and avoiding homozygosity of deleterious genes is important to maintain fitness (survival and fertility). (J23.17.w8, J23.27.w3, J54.5.w1)
  • For short-term captive breeding prior to reintroduction. (J54.5.w2)
    • Rapid breeding of as many offspring as possible is required. The captive population ideally should be held in an environment similar to the natural environment (or similar to the environment into which the animals will be released). (J54.5.w2)
    • Selection should be used only to select against outliers, reducing the genetic load [of deleterious alleles]. (J54.5.w2)
  • For long-term management of endangered species in captivity. (J54.5.w2)
    • The aims are to maintain a viable population, preserve as much as possible of the genetic variation, and preserve the option for future reintroduction. (J54.5.w2)
    • Management should aim to equalise founder representation and maximise effective population size. (J54.5.w2)
    • Selection may be required to control the genetic load. Culling to achieve this may be in conflict with maintaining equal representation of founders; culling of outliers may be delayed until the population size has grown, to ensure that the genetic loss from the culling is not greater than the genetic gain from removal of outliers. (J54.5.w2)
    • It may be necessary to allow some adaptation to captivity (e.g. improved survival and breeding) but this should be minimised, limited only to what is essential for the captive population to survive. Some inadvertent selection for adaptation to captive conditions may be inevitable. (J54.5.w2) 
    • Selection for genetic diversity should not be carried out; genetic diversity should be maintained by maximising the effective population size and maintaining equal representation of founders. (J54.5.w2)
    • It has been suggested that captive breeding programmes should aim to retain 90% of the average heterozygosity of the founders for 200 years. (J54.5.w1)
    • Maintenance of as much as possible of the genetic variability present in the wild gene pool requires both deliberate avoidance of inbreeding and demographic management of the population to establish and maintain a stable population. (P97.1.w34)
    • Variables to be considered include: the number of generations over which it may be necessary to maintain the captive population; the age and sex composition of the breeding population; age-specific and sex-specific fecundity and survivorship; the size of stable population required (based on known life history features); the carrying capacity of zoos for the species; requirements for surplus individuals; and how practical it is to develop controlled breeding or mating. (P97.1.w34)
    • Accurate record keeping is required. (P97.1.w34)
    • A breeding policy needs to be explicitly formulated and agreed by all parties concerned. (P97.1.w34)
    • Animals need to be of known origin, with pedigrees preferably traceable back to the original wild population. (P97.1.w34)
    • The age and sex structure should be defined, and the number of animals preferably should be chosen based on the breeding strategy.
    • "Explicit definitions of the need for introduction of new wild stock should be made in terms of breeding strategy and breeding management programs with a view to maintain some defined level of genetic variability in the population." (P97.1.w34)
    • Data collection and data sharing are vital; ISIS can provide census and vital statistic data, demographic projections, pedigrees, studbooks, and data for analysis of breeding relationships including calculation of inbreeding coefficients. (P97.1.w34)
    • For a maximally efficient genetic management programme, it is important to remember that the effective population size will determine the rate of loss of genetic diversity per generation. Given the use of a maximum avoidance of inbreeding scheme, the size of the effective, rather than the actual, population, depends on equal contribution by all members of the population to the next generation, i.e. equal family sizes, and an equal number of males and females breeding - for species where social organisation does not match this, management (e.g. rotation of breeding males) is required. (P97.1.w34)
      • The tendency to favour individuals which do well and breed well in captivity must be avoided to avoid selection of captive-adapted, inbred strains. (P97.1.w34)
      • From a given pairing, selection of offspring to contribute to the next generation must be randomized, avoiding selection based on unconscious preferences. (P97.1.w34)
  • Where the aim is to maintain populations purely for education/exhibition, selection should be used to control the genetic load. Selection can be used also to select for classic phenotypes, ensuring animals on display are representative of the typical wild form. Moderate selection for easy management and maintenance (e.g. tameness, willingness to breed in captivity, adaptation to inexpensive readily available diets) may be useful. (J54.5.w2)
    • However, species considered common may change in conservation status, so that captive populations may become of conservation importance in the future. (J54.5.w2)
    • Selection of zoo-domesticated stock probably would result in unrepresentative animals not suitable for life history studies or for development of data suitable for use in managing the species in the wild. (P97.1.w34)
    • Zoo-domesticated, inbred strains would be at high risk of loss of vigour, viability, growth rate and fertility, and could therefore become extinct. (P97.1.w34)
Preserving genetic diversity and avoiding inbreeding
  • As a general rule, inbreeding is to be avoided. Inbreeding occurs inevitably when the population is very limited in size, as may be the case for endangered species. In these circumstances, studbooks may be kept and pairings made to minimise inbreeding. Some species are known to have fallen to very low numbers and recovered without any indications of problems arising from inbreeding. In others, infertility, poor hatchability (in birds) and poor survival have been noted and thought to be related to inbreeding. 
  • Inbreeding is used in domestic animal breeding in a controlled manner when breeding to "fix" a particular characteristic, such as an unusual colouration of pelage. When used for this purpose (which is not appropriate for conservation purposes) the risks of also enhancing undesirable characteristics should always be considered.
  • In a few species, populations have descended from only a few individuals without any apparent deleterious effects (e.g. Anas laysanensis - Laysan teal, Elaphurus davidianus - Pčre David's deer), however these should be taken to be the exception rather than the norm. (J23.17.w7) In general, inbreeding results in inbreeding depression with reduced fertility, fecundity, lactation, viability, growth rate etc. (J23.17.w8)
    • Inbreeding depression leading to reduced fertility can quickly cause a small population to become extinct. (J23.17.w8)
    • Inbreeding depression may be more likely to arise with animals which were previously outbred rather than those with high natural inbreeding. (B482.3.w3)
    • In captivity, the effects of inbreeding may be compounded by management of breeding to ensure that different founders provide equal genetic contributions, and by the lack of the reproductive competition and associated social stimulation which generally occurs in the wild. (B482.3.w3)
  • "Maximum avoidance of inbreeding" can be carried out by, in each generation, mating the least-related individuals with each other. For a captive population of about 50-100 individuals, this should preserve about 50% of the original genetic diversity over 100 generations. (J23.17.w8)
  • The population should be expanded to carrying capacity as fast as possible, while breeding all available animals and ensuring equal family sizes. (J23.17.w8)
  • As well being advantageous to reduce inbreeding, managing for equal family sizes slows the rate of unintentional domestication which may occur from greater breeding of less aggressive, easier to handle, more readily breeding individuals. (J23.17.w8)
  • Genetic diversity is greatest when a 1:1 male: female ratio of breeding individuals is maintained. Where the actual ratio of breeding animals is far removed from this, e.g. if social criteria require one male with several females, the males should be rotated to equalise their breeding potential. (J23.17.w8)
  • Genetic strategies and demographic management should be coordinated. (J23.17.w8)
  • For maximum preservation of genetic diversity over time, generation time should be maximised, since there is the opportunity for loss of genetic variation at each generation. (J54.5.w1)
  • To minimise loss of genetic diversity in a captive population, breeding programmes should:
    • Involve an adequate number of wild-caught founders. (J54.3.w2)
      • For both conservation and reintroduction purposes, the founder population should be as large as possible with an approximately even sex ratio. (J23.17.w8)
      • To maintain 90% of genetic diversity over a period of 200 years, a minimum of 20 effective individuals, and preferably at least 25-30 is generally required. (J54.5.w1)
        • If there are 20-30 effective founders, average heterozygosity will be preserved. To preserve rare alleles from the wild population, 30-50 effective founders are required. (J23.27.w3)
      • If even one effective founder from the wild population enters the captive population each generation, this can be sufficient to keep the captive population representative of the wild gene pool. (J23.27.w3)
    • Equalise, as much as possible, the genetic contribution of the founders to the population. (J23.17.w8, J54.3.w2)
      • It is preferable for genetic purposes to maintain an even sex ratio of animals contributing to each generation, and for all the members of one generation to contribute equally to the next breeding generation (i.e. for an equal number of offspring to be produced and used for breeding). This minimises inbreeding and decreases the risk of domestication which may arise from the individuals which breed most easily in a captive situation becoming over-represented in the breeding stock. (J23.17.w8)
    • Specify matings to minimise inbreeding. (J54.3.w2)
      • Ideally, animals who are least related to one another are mated - a process known as "maximum avoidance of inbreeding." If this is used in each generation, then with a starting population of 50-100 animals it would be possible to preserve over half of the genetic diversity of the original population over 100 generations. (J23.17.w8)
    • Maximise the effective population size, by manipulating the number of offspring per parent, and the sex ratio of the population. (J54.3.w2)
      • The effective population size is largest when each individual parent produces the same number of progeny. (J23.27.w3, J407.17.w1)
    • Maintain a demographically stable population. (J54.3.w2)
  • To maximise genetic diversity: (B115.9.w13)
    • The population should be started with an adequate number of founders (at least 20 effective, i.e. reproducing, founders).
    • The population should be expanded to carrying capacity as fast as possible; this capacity must be larger than the minimum viable population size.
    • The sex ratio should be equalised.
    • Family size should be equalised (in each generation, breeding individuals should produce equal numbers of offspring to contribute to the next breeding generation).
    • Once carrying capacity is reached, the size and growth of the population should be stabilised, avoiding population size fluctuations.
    • Once carrying capacity is reached, the population should be managed for longer generation times.
    • Inbreeding should always be minimised.
    • Always manage for a stable age structure.
    (B115.9.w13)
    • To properly use a computer-based demographic program, a model for each species must be developed based on: (P97.1.w34)
      • Estimate of the carrying capacity of the captive breeding groups.
      • The need for surplus individuals (for exhibit in non-breeding institutions, use in research programmes etc.).
      • Determined or estimated fecundity and survivorship by age and sex classes. In particular, for relatively long-lived species, adequate first-year survivorship and average annual adult survivorship data are required, in order to determine the average number of offspring required per parent to maintain a stable population size.
        • It is essential that mortality data are recorded accurately, for survivorship data and for detection of detrimental inbreeding effects.
      • The known age and sex structure of the existing population. This is used with survivorship and fertility data to predict the demographic structure of the population.
      • An explicit plan for moving from the current to the desired stable population must be devised, tested and agreed on. This should be examined and stochastic models used to predict possible fluctuations.
      • Note: similar modelling and projections can be used to formulate a strategy for planned reintroduction projects. (P97.1.w34)
    • N.B. A deliberate breeding plan can more effectively preserve the maximum heterozygosity than can random breeding. (P97.1.w34)
  • Note: Following the recommendations for breeding which have been made based on genetic management may not always be practical or possible, due to considerations such as behavioural incompatibilities or psychological problems; differences in disease status between animals; and logistical problems in moving animals between collections.
    • Use of assisted reproductive techniques (see below: Assisted Reproduction) may be appropriate in some cases to allow appropriate genetic matching. However, the applicability of such techniques in non-domestic animals is very limited. 
Use of reproductive technology in maintaining genetic diversity
  • Semen collection and cryopreservation can help in the maintenance of genetic diversity by increasing the number of potential founders (collecting semen from wild or captive individuals that, for whatever reason, have not contributed genetically to the captive population, or are underrepresented genetically in the population), and, via cryopreservation, extending the reproductive life of those founders and their closest descendents. (J54.3.w2)
    • Collection and preservation of semen from founders does not allow preservation of genetic material from female founders. However, collection of semen from male offspring of female founders (particularly those genetically underrepresented in the population) does allow some preservation of their genetic material. (J54.3.w2)
    • Ideally, F1 offspring should be produced from different pairings (male A with female B and male C with female D, but also e.g. male A with female D and male B with female D), to allow separate management of the genetic contributions from different founder individuals. (J54.3.w2)
    • Since each F1 individual can have inherited only part of the allelic diversity of each parent, for maximum probability of representation of all the allelic diversity of the founders, semen should be collected from several (ideally six to nine) F1 offspring of given founders. (J54.3.w2)
    • In the absence of founders or their F1 descendents, semen should be collected and stored from individuals with genomes least exposed to genetic drift and inbreeding, and within these individuals, particularly from descendents of genetically underrepresented founders. (J54.3.w2)
      • Careful analysis of pedigrees and calculation of inbreeding coefficients and the relative genetic contribution of each founder is required. (J54.3.w2)
    • For further information on semen collection and cryopreservation see: Assisted Reproduction.
Demographic management

Demographic management is an important part of the management of captive populations, alongside genetic and behavioural management. (J23.17.w9)

  • A demographic model needs to consider the required population size, as indicated by carrying capacity plus any need to provide surplus individuals for other purposes. (J23.17.w9)
  • The demographic part of a management plan should specify the number, ages and (in part) sexes of individuals to be retained in the breeding population and will indicate how many progeny need to be produced within a given time interval, and specify the number and ages of animals to be bred.
    • Exactly which individuals within a given sex and age class should breed will depend on genetic management.
    • Behavioural management will indicate the grouping and movement of animals (e.g. rotation of males where behavioural grouping requires uneven sex ratios of breeding groups).

    (J23.17.w9)

  • Usually, demographic management aims to produce a stationary (stable) population, with a constant population size and stable age distribution. (J23.17.w9)
    • A stable population is advantageous because it minimises the risk of demographic disaster due to random fluctuations or unpredicted developments in population.
    • Production and maintenance of a stable population involves manipulation of potential survivorships (by removing individuals from the breeding population) and potential fertilities (by limiting breeding).
      • Animals removed from the breeding population would be available for other uses. 

    (J23.17.w9)

  • Application of demographic models to assist long-term captive management and possible reintroduction would require: (J23.17.w9)
    • Determination of the existing and, if possible, the potential age- and sex-specific survivorships and fertilities of the population; calculation of the existing age/sex structure.
    • Calculation of near-future changes in the population size and age structure, subject to the survivorships and fertilities determined above.
    • Determining the finite rates of change and stable age distribution associated with the determined sets of existing and potential survivorships and fertilities.
    • Determining, as closely as possible (a) the carrying capacity for the captive population; (b) the number of surplus animals needed for other purposes (e.g. reintroduction programmes).
    • Within the upper limits set by the potential or maximum existing values of survivorships and fertilities, construction of (computer-aided) the one or more sets of managed survivorships and fertilities which, by varying amounts of culling (removal of individuals from the managed population) and/or breeding restriction, will produce a stationary captive population. 
    • Determining the sets of stationary age distributions associated with each of the calculated stationary sets of managed survivorships and fertilities. The total number of animals in one or more of these sets should be approximately equal to the desired carrying capacity for the population.
    • With computer-assistance, preparation of a plan for moving from the existing population to a stationary population which best fits the carrying capacity and surplus population desired.
    • Selecting individual animals for demographic manipulation in accordance with genetic models.
    • N.B. It would also be necessary to consider stochastic (random or chance) population fluctuations that might occur in the future.

    (J23.17.w9)

Hybrids
  • Breeding of hybrids should be avoided by not housing closely related species with one another. Sub-specific hybridisation also should be avoided. This may be more difficult if subspecies are not readily identifiable from one another on the basis of external features, and if records do not accurately indicate the origin of the individuals.
Bear Consideration Bears in zoos are managed to differing extents depending on the species and the priorities of the regional zoo associations. For example:-

In Europe (EAZA)

In North America (AZA - Bear TAG):

  • Tremarctos ornatus - Spectacled bear, Melursus ursinus - Sloth bear and Helarctos malayanus - Sun bear are managed under SSPs. (W642.Jun07.w1)
    • The SSP for Tremarctos ornatus - Spectacled bear works closely with the European EEP as well as with zoos in range state countries (Venezuela and Peru). (W642.Jun07.w1)
    • Melursus ursinus - Sloth bear is highly endangered and the CBSG has recommended that North American zoos take the lead in managing these bears in an international studbook. Additionally, the Bear TAG "will place high priority on initiating efforts to develop an in situ conservation program". (W642.Jun07.w1)
    • Helarctos malayanus - Sun bear. New founders are needed for the North American population. Importation of founders "needs to be done with an active conservation linkage program in the range country where the animals will be obtained in order to protect and preserve as much remaining habitat as possible while at the same time carrying out intensive field studies on the sun bear itself. This is vitally important as so very little is known about these animals in the wild." (W642.Jun07.w1)
  • Ursus maritimus - Polar bears are managed under a regional studbook, and zoos are encouraged "to emphasize the importance of the polar bear's delicately balanced ecosystem in their conservation message." (W642.Jun07.w1)
  • The Bear TAG does not recommend that zoos in North America breed Ursus americanus - American black bear (which have a stable population), Ursus arctos - Brown bear (which is stable in its large population groups) or Ursus thibetanus - Asiatic black bear (the North American population "is an unknown, with few animals, many of varying subspecies, and is considered unsuitable to manage for genetic success"). (W642.Jun07.w1)
Genetic considerations
Lagomorph Consideration There are few breeding programmes for wild lagomorph species.

Brachylagus idahoensis - Pygmy rabbit

  • In the breeding program for the Columbia Basin (Washington State, USA) Brachylagus idahoensis - Pygmy rabbit population, it has been found that this distinct population of the pygmy rabbit is genetically distinct from other populations. It has also shown that over the past 50 years it appears to have suffered a considerable loss of genetic diversity. (D372)
  • Compared with the Idaho pygmy rabbit, the Columbia Basin pygmy rabbits have shown lower pregnancy rates, fewer litters per breeding male, fewer litters per breeding female and fewer young produced per female. There were also behavioural differences: longer times to show reproductive behaviour (964 versus 124 minutes) and longer periods chasing, although similar numbers of mountings. Additionally, Idaho pygmy rabbits continued breeding through May and into early June, while Columbia Basin pygmy rabbits did not breed successfully after 12th April. (D372)
  • Interbreeding Columbia Basin and Idaho rabbits has produced young, with successful backcrossing to Columbia Basin pygmy rabbits. Intercross rabbits have shown markedly increased reproductive success compared with pure Columbia Basin individuals. (D372)
  • Genetic management priorities of the Columbia Basin Pygmy Rabbit Captive Breeding and Genetic Management Plan (2006) included: (D372)
    • "Maintain genetic diversity of Columbia Basin pygmy rabbits by matching the most appropriate Columbia Basin males and females (minimize loss of genetic diversity)";
    • "Produce offspring of pure-bred Columbia Basin pygmy rabbits";
    • "Conduct intercrosses to produce 87.5% Columbia Basin pygmy rabbits";
    • "De-emphasize breeding of rabbits with a high relatedness to the Idaho rabbit Alder;
    • "Conduct intercrosses to produce 75% Columbia Basin pygmy rabbits";
    • "Create no new F1 intercrosses (Columbia Basin/Idaho intercrosses) and avoid pairings to produce 62.5% Columbia Basin pygmy rabbits".

(D372 - The Columbia Basin Pygmy Rabbit Captive Breeding and Genetic Management Plan - full text included)

  • It has been noted that there are significant challenges for the captive breeding and recovery of short-lived lagomorphs. Rapid expansion of the captive population is important to take the Columbia Basin pygmy rabbit population out of the demographic and genetic bottleneck, minimise genetic drift, and to enable harvest of the captive population, i.e. taking of individuals for the reintroduction programme, without increasing to unacceptable levels the risk of extinction of the captive population. (J471.11.w1)

Sylvilagus bachmani - Brush rabbit 

  • In a propagation and reintroduction plan for riparian brush rabbits Sylvilagus bachmani riparius, each year new founder animals are taken from a population in the South Delta on the San Joaquin Sacramento rivers' delta, west of Lathrop, California, USA and kept in large, predator-free propagation pens. Surviving founders are then repatriated to their original capture site, as well as rabbits bred in the propagation pens being released. (D339, V.w137)
Ferret Consideration Breeding programmes for domestic ferrets vary depending on the goal of the programme, such as particular coat colours, ferret size or shape. (B651.6.w6, B652.2.w2)
Bonobo Consideration Captive bonobo populations in Europe and North America are managed in an EEP and SSP respectively, with studbooks maintained and breeding recommendations made, aiming to minimise inbreeding. (P86.9.w1)
  • Captive bonobo breeding programmes avoid inbreeding as much as possible, but there is still some evidence of inbreeding and inbreeding depression. A study found higher mortality in infant bonobos which were inbred (inbreeding coefficient greater than zero) than in infants which were not inbred (31.25% versus 12.1%; difference not statistically significant). (P86.9.w1)
Associated techniques linked from Wildpro
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Pair / Group Formation and Maintenance

  • Mammalian mating systems show considerable variation from monogamous to promiscuous. In some species temporary pairs are formed or males and females come together only for mating; males may mate with one or several females; females may mate with one or several males. In other species, pairings are long term or permanent. In many hoofstock species, dominant males defend a herd of several females.
  • Captive animals frequently are not allowed to chose their own mates. Rather, the choice is made for them on the basis of genetics and/or expediency (available individuals). While mate choice may not be relevant in many species, it may play a role in the breeding of some species, with two individuals not forming a pair and not breeding, despite the studbook keeper's recommendations.
  • Care is required when introducing animals to one another and animals should be introduced to one another gradually. (B438.24.w24)
  •  An animal being introduced to conspecifics is at a disadvantage if also being introduced to a new enclosure. If possible, the animal should be given a chance to familiarise itself with the enclosure, then to meet its new mate or group through a barrier before the animals have full access to one another. Periods of contact usually should be brief initially and increased as it becomes clear that the animals are behaviourally compatible. (B438.24.w24)
  • Consideration should be given to the need, in species which are normally predominantly solitary, to separate the male and female for much of the time. This may improve reduce stress from e.g. food competition, and may improve both breeding and rearing of young. (B105.19.w6)
  • It is important to remember that the introduction of unfamiliar individuals into a social group, resulting in significant alterations in social interactions, may produce immediate physiological effects which may interfere with breeding (e.g. abortion at certain stages of pregnancy in many small mammals if an unfamiliar male is introduced to the group). (B105.19.w6)
Bear Consideration Bears are polygamous/promiscuous in the wild. In zoos, bears which are intended to breed are commonly maintained either as a pair or in a group with several females and one male.

In Helarctos malayanus - Sun bears in Europe, breeding has been more successful when bears were kept as pairs rather than in groups. (P6.2.w5)

Recommendations (EEP Ursid Husbandry Guidelines) are as follows:

  • If a breeding group is to be kept containing several females, preferably these females should be sisters or mother and female offspring. (D247.4.w4)
    • If female offspring are to be kept with their mother and be bred, the male needs to be exchanged for an unrelated male before his daughters become sexually mature. This reflects the natural, wild situation more closely than does keeping the same male and introducing unrelated females to one another. (D247.4.w4)
    • With Helarctos malayanus - Sun bear, female offspring may be kept with their parents for several years. However, if unrelated females are introduced to one another there may be (a) aggression, fighting and injuries; (b) reduced reproductive success. (D247.4.w4)
Lagomorph Consideration
Domestic rabbits
  • The doe should be brought to the buck, not the buck brought into the doe's territory, to avoid fighting: does tend to be more territorial than bucks. (B339.8.w8, B550.16.w16, J35.151.w2)
  • Usually, copulation occurs within a few minutes. (B550.16.w16)
  • If a receptive doe does not allow copulation, the doe should be placed with a different buck; it has been observed that a doe may reject one buck and accept another buck. (B550.16.w16, J35.151.w2)
Wild lagomorphs
  • With Romerolagus diazi - Volcano rabbit at Jersey Zoo, it was found that housing a pair in a small cage was not successful. Breeding was achieved by development of a 4 x 4 m area, dividable into two sections each with a tunnel system connecting underground nestboxes. Male and female would be given one side of the area each, for a week, then the partition was removed and the pair left together for 35 days (allowing re-isolation of the female before the earliest possible littering date , given a gestation of 38 - 40 days. (J51.19.w1)
  • Brachylagus idahoensis - Pygmy rabbit can be maintained with a female and two males in large pens (e.g. 75 - 100 m˛) (J332.87.w1, V.w134). When kept in smaller pens, only a single adult can be kept per pen. For breeding, a door between the male's and the female's pen is opened; this can then be left open for two or three days to allow mating. Animals are not given access to each other in this manner until the male is known to be in breeding condition (testes are fully descended, purple and firm). (J332.87.w1)
    • Following parturition, either the male (or another male) is given access to the female again while the young are in the natal tunnel, or the male is introduced after the young have been removed from the pen. (J332.87.w1)
    • Note: Higher reproductive success appear to occur if immediate post partum mating is allowed, rather than waiting and mating the female again later. (V.w134)
    • Breeding has been more successful for rabbits kept in a larger, semi-naturalistic pen rather than in small individual pens. (V.w134)
Ferret Consideration
  • Normally either the female and male are placed together in neutral territory or the female is placed into the male's territory for mating. (B651.6.w6)
  • The male soon bite's the female's neck and drags her around before mating. Mating is violent and noisy, with the female screaming. Afterwrads, both male and female clean themselves, and they may eat a little. If left together, mating thn starts again. They may be left together for up to 24 hours. Leaving them longer is not advised as repeated mating can result in excessive damage to the female's neck. (B651.6.w6) Once mating is completed, the female's neck should be examined and the wounds cleaned and treated with antiseptic powder. (B651.6.w6)
Bonobo Consideration
  • In the wild, it is the females that move between communities, and the status of males may be affected by the presence of their mother. Therefore in zoo breeding programmes, generally females rather than males are moved between collections (D386.2.1.w2a, D386.3.2.w3b, D386.App1.w6)
    • An adolescent female preferably should be transferred into a group containing an older, established female with whom she can bond, as occurs in the wild. (D386.3.2.w3b)
  • If males must be transferred, it is suggested that transfer occurs when they are older juveniles or young adolescents, 5 - 7 years of age, and that they be transferred into a group with other bonobos of the same age, with whom they can play. (D386.3.2.w3b)
  • Males which are genetically overrepresented or genetically undesirable may be housed in an all-male group. (D386.App1.w6)
Associated techniques linked from Wildpro

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Hybridisation

Hybridisation may occur in captive animals, if closely-related species, or subspecies, normally separated (e.g. geographically) are brought together. In most cases, hybrids are infertile, although hybrids between subspecies or between closely related species may be fertile. 
  • Hybridisation is sometimes a useful tool in indicating ancestral relationships, but should usually be avoided.
  • Hybrids take up valuable resources (living space, food costs, keeper time etc.), which might otherwise be available for the maintenance of pure-bred individuals of endangered species. (B444.w1)
  • Fertile hybrids may be a threat to the maintenance of the pure form of either or both parent species or subspecies.
Bear Consideration

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Bears should not be kept in an enclosure with other bears in circumstances allowing hybridisation (i.e. there should not be a reproductively capable male of one bear species sharing an enclosure with a reproductively capable female of another bear species). 
Lagomorph Consideration Hybridisation is probably most likely to occur among Lepus spp.; all Lepus spp. karyotyped have been found to have 48 chromosomes and very similar karyotypes. (J336.12.w2)
Ferret Consideration

Hybridisation has been reported with other species in the genus:

Bonobo Consideration
  • Hybrids between bonobos and Pan troglodytes - Chimpanzee has occurred in captivity between a male Pan paniscus and two female Pan troglodytes. (P119.2003.w13) with two aborted fetuses and three healthy offspring reported. (J187.56.w1)
  • Given the close genetic relationship between Pan paniscus and Pan troglodytes - Chimpanzee, it is not surprising that hybridisation should be possible. Given the conservation importance of bonobos, it is unlikely that hybridisation is likely to be permitted in modern zoos. (V.w5)
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Monitoring Reproductive Health and Reproductive Status in the Female 

Many factors may influence reproductive function, including gross nutritional status, micronutrients, behavioural and social factors, season etc. 

Assessment of reproductive status is required for day-to-day management of captive breeding programmes, including decision-making regarding animal housing and pairing; selection of individuals for intensive breeding management; evaluating the outcome of breeding attempts; group/herd management; monitoring the health of the fetus and predicting birth dates to allow preparations for birth; regulation of fertility; and establishing the causes of breeding failure and monitoring treatment. Monitoring the timing of ovulation further allows the scheduling of timed matings, and the application of reproductive technologies such as artificial insemination, and embryo recovery and transfer. (B445.w4)

A variety of methods have been developed to monitor the reproductive status and reproductive health of animals.

Behavioural observation
  • An important part of reproductive monitoring is behavioural monitoring. (B445.w4, J23.38.w2)
  • This is non-invasive and does not require any animal contact or restraint. (B445.w4)
Ultrasonography
  • The use of real-time ultrasound examination is of increasing use for monitoring reproductive health and reproductive status. (B429.38.w38, B445.w4, P17.60.w1)
  • This method allows the organs to be observed without the need for any surgical invasion. (B429.38.w38, B445.w4, P17.60.w1)
  • The main advantages of ultrasonography are that it is non-invasive and atraumatic; gives real-time, visual, reproducible information about the organs; gives cross-sectional images of the tissues/organs, including images of organs which are moving; allows organs or other objects to be measured; can be integrated easily with other methods of examination; and can be stored. (B445.w6, P17.60.w1)
  • The main disadvantage is the need for physical restraint and often for sedation or anaesthesia. (B429.38.w38, B445.w4, B445.w6)
    • Note: Some animals can be trained to allow ultrasonography without either physical restraint or sedation. (V.w5)
  • Other potential problems include the need for a good acoustic coupling with the animal, which may be disrupted for example by fur; insufficient penetration of sound waves through tissues; the need for modification of some components to allow for different species; and the need to develop experience with different species, as well as cost issues. (B445.w6, P17.60.w1)
  • Ultrasound can be used for sex determination of monomorphic species (species in which male and female cannot be distinguished by external characteristics). (B429.38.w38, B445.w6, P17.60.w1)
  • Ultrasound can be used to monitor the female reproductive cycle, but this involves repeated examination, and physical or chemical restraint may be needed on each occasion. (B445.w6)
  • Use of ultrasonography may allow a diagnosis (e.g. of pregnancy) from a single observation. (B429.38.w38, B445.w6)
  • Ultrasonography may allow monitoring of early pregnancy and, by measuring fetal structures and comparing these to reference data, may allow calculation of the expected parturition date. (B445.w6)
  • Ultrasound is able to detect pathological changes in the genital tract, and may detect pathological changes in other maternal organs that may affect general health and reproductive function. (B445.w6)
  • Ultrasound may detect abnormalities in the embryo, and sometimes in the fetus up to as late as the end of the second trimester. (B445.w6)
Vaginal smear
  • Vaginal smears can provide useful information about the reproductive cycle in the female animal. (B445.w4)
  • This is a non-invasive technique but requires capture and restraint of the animal. (B445.w4)
  • Some animals can be trained to allow collection of vaginal smears without either physical restraint or sedation. (V.w5)
Endocrinology (measurement of hormone concentrations)
  • Analysis of hormones in the blood, urine, faeces or saliva can be a precise way of indirectly monitoring reproductive function. However, before these can be used for monitoring it is necessary to develop a knowledge of the reproductive physiology of the species, including hormone metabolism and patterns of hormone secretion and excretion. (B429.38.w38, B445.w4, B445.w5, J372.42.w1)
  • Hormones may be detected and their concentrations measured using radioimmunoassays or enzyme immunoassays. (B429.38.w38)
  • With the right tests it is possible to monitor the ovarian cycle, and to detect and monitor pregnancy. (B445.w5)
  • Analysis of blood may provide the most information, but generally requires collection of serial blood samples, which is a practical disadvantage. (B429.38.w38)
  • Non-invasive monitoring of reproductive status may be carried our by tests on voided urine or faeces. These may allow study of gonadal function including cyclicity, seasonality, and pregnancy, as well as adrenal (i.e. stress) status. (B445.w4, J23.38.w2)
    • Detection of excreted hormone metabolites in urine has been carried out for some years; measurement of metabolites in faeces was developed later. (B445.w5)
    • For each species, it is necessary to determine the main hormone metabolites produced and the main route of excretion of hormone metabolites; some species, such as many ungulates, excrete oestrogens mainly in the urine, while others, such as carnivores (Carnivora - Carnivores (Order)), excrete oestrogens mainly in the faeces. (B445.w5, J372.42.w1)
    • It is important to validate tests used in each species; measurements of metabolites which give excellent information on reproductive status in one species may not be useful in another species. (B445.w5)
    • Urinalysis is advantageous compared with analysis of blood, since collection of urine is a non-invasive process. There are practical problems associated with urine collection, and correction factors, usually relating values to creatinine concentrations, may be needed to allow for differences in concentration and volume. (B429.38.w38)
      • In equids (Equidae - Horses (Family)), pregnancy can be confirmed by detection of the pregnancy-specific equine chorionic gonadotrophin in urine. (B429.38.w38)
    • Analysis of faeces can be used since fecal excretion is the predominant route of hormone excretion in many species. (B429.38.w38)
      • It is necessary to determine, for each species, which metabolites should be assayed. (B429.38.w38, B445.w5)
      • Analysis of oestrogens in faeces has been used reliably for indication of pregnancy in some ungulates and primates. (J372.42.w1)
      • Collection of faeces may be easier than collection of urine, particularly for free-living animals but also for some captive animals. (B429.38.w38, B445.w5)
      • Complex, laborious laboratory techniques are required to process the faeces before the assays can be carried out. (B429.38.w38, B445.w5)
      • For analysis of progesterone metabolites in faeces, group-specific antibodies against 5-alpha-pregnanes or 5-beta-pregnanes are recommended. (J372.42.w1)
  • Saliva contains minute quantities of hormones, which can be measured by sensitive assays. In most situations, use of this method is limited by practical considerations regarding collection of saliva. (B429.38.w38)
Laparoscopy/laparotomy

Both laparotomy and laparoscopy may be used to examine the female reproductive organs and determine reproductive status. (B429.38.w38, J23.18.w3)

  • Both methods are surgical, invasive procedures and require anaesthesia. (B429.38.w3, J23.18.w38)
  • Laparoscopy causes less surgical trauma than does laparotomy and is now generally preferred. (B429.38.w38, J23.18.w3)
  • Laparoscopy can be valuable for visualising the internal reproductive organs and monitoring reproductive cycles in a variety of species. (B429.38.w3, J23.18.w3, P1.1976.w1)
  • Laparoscopy can be particularly valuable for determining the precise timing of ovulation, for identifying abnormalities and pathological conditions of the internal reproductive tract, and in some cases for surgical treatment of pathology (e.g. adhesions). (B429.38.w3, J23.18.w3)
Post mortem examination
  • As part of the necropsy, the ovaries, uterus and other parts of the reproductive tract should be examined.
  • Thorough examination of the reproductive tract at necropsy allows correlation with, and confirmation of, findings from ante-mortem examinations such as ultrasonography. (P6.4.w3)
Bear Consideration The reproductive health and reproductive status of bears have been monitored by a variety of methods including behavioural observation, physical examination, laparoscopic examination, ultrasonography and measurement of hormones in blood or faeces.

In bears, it is not possible to distinguish between failure to conceive and failure to implant. Either condition could lead to no cubs being born.

Behavioural observation
  • Oestrous behaviour, such as presenting to the male and standing to be mounted by the male, and the occurrence of matings, can be observed. (J27.65.w1, J296.66.w1)
Ultrasonography
  • Transrectal ultrasonography can be used to assess the reproductive system in female bears and to confirm pregnancy. (J296.66.w1, J370.51.w1, P1.1997.w7)
  • Special adaptors, 15, 25 and 50 cm long, allowed scanning of the elongate genital tract in Ursus arctos - Brown bear, Tremarctos ornatus - Spectacled bear and Melursus ursinus - Sloth bear, using a 7.5 MHz curved linear intraoperative transducer and a real-time B-mode ultrasound scanning system. It was found possible to examine the urinary bladder, urethra, kidneys (but not the ureters), vagina, cervix, uterus and ovaries, to detect pregnancy and to detect abnormalities such as a leiomyoma and purulent vaginitis. (J370.51.w1, P1.1997.w7)
  • There are no differences in the uterus detectable by ultrasonography between non-pregnant bears and pregnant bears pre-implantation. Ovulation can be confirmed by the presence of active corpora lutea. Pregnancy can be detected only following implantation of the embryo. The pregnant uterus shows local uterine enlargement, anechoic fetal fluid and endometrial proliferation at sites of implantation. The placentae appear as hypoechoic discoid structures. (J370.51.w1, P1.1997.w7)
  • Transabdominal (transcutaneous) ultrasonography may be used for detection of pregnancy in bears, but not for visualisation of the ovaries, non-gravid uterus or early pregnancy. (J370.51.w1, P1.1997.w7)
Endocrinology 
Blood steroid analysis
  • Levels of progesterone in blood can be determined using an enzyme-immunoassay (EIA) (J370.51.w1 P1.1997.w7) or radioimmuneassay (RIA). (J27.62.w2, J206.68.w1, J371.65.w1, J372.53.w1)
    • A study of Ursus arctos - Brown bear found very low concentrations of plasma progesterone (less than 0.1 ng/mL) during February to April (non-breeding season) and higher levels, indicating active corpora lutea, both in September-October (before implantation) (3.3 +/- 0.9 ng/mL) and in November-December (after implantation) (3.1 +/- 0.1 ng/mL). (J370.51.w1)
    • A study of Ursus americanus - American black bears found that non-pregnant bears, whether in lactational anoestrus or non-lactational anoestrus, had very low levels of plasma progesterone (0.6 - 2.5 ng/mL). Levels were increased in pregnant bears to 5.0 - 12.5 ng/mL during the mid- to late preimplantation period, and at about the time of implantation in late November/early December increased a further two- to three-fold. It was noted that both non-pregnant and pregnant levels of plasma progesterone showed variation between individual bears and that levels during pregnancy varied between years for a given bear. (J206.68.w1)
    • A study of captive and free-living Ursus americanus - American black bears found that serum progesterone levels rose, from a very low level, about one month after oestrus, and increased dramatically around the time of implantation in November/December. (J372.53.w1)
    • A study of Japanese black bears Ursus thibetanus japonicus (Ursus thibetanus - Asiatic black bear) found that serum progesterone levels (measured by radioimmuneassay) were low in April to July, tending to increase in August, rising much higher November and December then falling to low levels by March. In captive females known to be pregnant (cubs produced the following spring), there was a gradual increased from 0.5-2.4 ng/mL in August to 0.9-3.6 ng/mL in October and reaching significantly higher maximum levels (7.2-18.0 ng/mL) in December. Serum levels of estradiol-17β were low in November and December, higher in January and variable in April to October. For eight pregnant females, estradiol-17β was high in May (95.6-191.4 pg/mL, varied in August to October (35.6-143.3 pg/mL), dropped to significantly lower minimum levels (5.3-11.9 pg/mL) in December, then increased to 67.6-153.1 pg/mL in January. (J27.62.w2)
    • A study of Japanese black bears Ursus thibetanus japonicus (Ursus thibetanus - Asiatic black bear) found that serum progesterone levels (measured by radioimmuneassay) were low in non-pregnant bears, while both pregnant and pseudo-pregnant bears showed a significant increase in progesterone in December (time of implantation of the embryo in pregnant females). Progesterone is produced by the corpora lutea, not the placenta, in carnivores. (J371.65.w1)
    • A study of Ursus thibetanus formosanus in Taiwan, found that in a female which became pregnant, serum progesterone was 3 ng/mlL in April, lower (about 1.5 to less than 1 ng/mL) in May to August, rising to about 7 ng/mL in September. (J467.5.w1)
Fecal steroid analysis
  • Analysis of faecal steroid levels may allow non-invasive monitoring of the reproductive status. (J370.51.w1)
  • Levels of progesterone in faeces can be determined using an enzyme-immunoassay (EIA). (J370.51.w1)
  • Fecal progesterone concentrations can be used to monitor luteal activity in Ursus arctos - Brown bear: (J296.66.w1)
    • A study of Ursus arctos - Brown bear (Ursus arctos yesoensis, Hokkaido brown bears) found that mean fecal estradiol-17β concentrations were high during the oestrus period then decreased, while fecal progesterone concentrations increased after the oestrus period. (J27.65.w1)
    • A study of Ursus arctos - Brown bear found very low concentrations of fecal progesterone (9.6 +/- 6.7 ng/g faeces) during February to April (non-breeding season) and higher levels (50 - 800 ng/g) in bears with active corpora lutea: both in September-October (before implantation) (151 +/- 133 ng/g) and in November-December (after implantation) (379 +/- 238 ng/g); in the first two pregnant bears examined, an increased excretion of progesterone was recorded. (J370.51.w1)
    • In one Ursus arctos - Brown bear, fecal progesterone concentrations were found to be very low (0.98 +/1 2.04 ng/g faeces) prior to the mating period, and tended to be higher (mean 7.36 +/- 7.94 ng/g) in the post mating period, with a dramatic increase (4,650 ng/g) in December, at the time of embryo implantation (implantation was confirmed by subsequent ultrasonographic examination). (J296.66.w1)
  • A study of Helarctos malayanus - Sun bear found that the level of 17-oxo-androgen in faeces (epiandrosterone antibody) generally peaked at the time of behavioural oestrus (no such peak was detected at the time of behavioural oestrus in one bear on one occasion). Levels of 20alpha-OH-pregnanes (pregnanediol antibody) increased during the luteal phase of the cycle. The length of the luteal phase was similar in pregnant and non-pregnant bears. (P6.2.w5)
  • The Australasian Husbandry Manual for the Malayan Sun Bear indicates that measurement of progesterone metabolites in faeces, using an ELISA, is useful for monitoring pregnancy in Helarctos malayanus - Sun bear, while measurement of pregnanediol-glucuronide is not useful in this species. It was noted that there were wide variations in excretion of progesterone over the course of pregnancy and that it was important to run controls, including a non-pregnant control, in each assay. (D255.6.w6)
  • In a Tremarctos ornatus - Spectacled bear, low concentrations of progesterone were measured in faeces, except for alternating high and low concentrations during the second half of pregnancy. (J296.66.w1)
  • A study of Ursus thibetanus formosanus in Taiwan, found that in a non-pregnant female, fecal progesterone remained low (below 25 ng/g faeces) except for two higher readings in late May and in early August. A female which became pregnant showed rising fecal progesterone during October (parturition in early November); fecal progesterone levels also rose in a "pseudopregnant" female. (J467.5.w1)
Urinary steroid analysis
  • Enzyme immunoassays (EIA) can be used to detect total oestrogens, progesterone, pregnanediol and epiandrosterone. However, these do not appear to be useful for monitoring reproduction in bears. (J296.66.w1)
    • In an Ursus arctos - Brown bear, urinary oestrogen concentrations fluctuated at about 0.5 ng/mL; there was an increase to 3.3 ng/mL in June, but this occurred after matings, which were observed 27 May to 4 June. Measurement of urinary pregnanediol and progesterone did not reveal increased luteal activity. (J296.66.w1)
    • In a Tremarctos ornatus - Spectacled bear, urinary oestrogen concentrations fluctuated (0.1 to 0.8 ng/mL urine), with no peak related to observed mating dates. Urinary progesterone concentrations were generally below 1.0 ng/mL, with two higher values measured in July and around the time of implantation, with an increase to 3 ng/mL about three weeks post-implantation, and a decrease seven days before parturition. (J296.66.w1)
Laparoscopy
  • Laparoscopy has been used for monitoring ovarian function in bears. (B429.38.w38, J23.18.w3)
Post mortem examination
  • Thorough examination of the reproductive tract at necropsy allows correlation with, and confirmation of, findings from ante-mortem examinations such as ultrasonography. (J370.51.w1)
Lagomorph Consideration
Behavioural observation
  • Experienced personnel may detect when a female rabbit is receptive by her general behaviour, such as increased restlessness. (J35.151.w2)
  • Increased restlessness, trying to reach rabbits in nearby hutches, and scent marking by rubbing the chin on surfaces are signs of sexual receptivity. (B550.16.w16)
  • A receptive female should allow the male to mount her. However, even when the female is receptive, mating may not occur immediately when a male and female rabbit are placed together. (B618.21.w1)
  • If receptive, the female will lie down and raise her hindquarters to allow the male to mate. (B550.16.w16)
Physical Examination
Receptivity
  • Receptivity in Oryctolagus cuniculus domesticus - Domestic rabbits can be detected by the colour of the vulva - swollen, moist and dark red to purple at the right time for mating, rather than narrow and pale pink. (B612.8.w8, B618.21.w1, B550.16.w16)
    • However, some females with a purple vulva are not receptive, and some females have a purple vulva while pseudopregnant or pregnant. (J35.151.w2)
Pregnancy detection
  • In Oryctolagus cuniculus domesticus - Domestic rabbits, pregnancy may be detected by gentle palpation of the caudal abdomen at 10 - 14 days after mating. (B618.7.w7); 12 to 14 days of gestation. (B550.16.w16, B612.8.w8)
    • Fetuses are felt as "marbles" slipping between the thumb and fingers. (B550.16.w16)
    • This is not always effective when only one or two fetuses are present. (J35.151.w2)
    • There is a risk of damaging the fetuses if too much pressure is used. (B550.16.w16, J35.151.w2)
    • Distinguishing between pregnancy and digesta is more difficult after the 14th day. (B550.16.w16)
    • Note: "Rabbits are particularly prone to fetal loss at day 13, when placentation changes from yolk sacs to hemochorial, and at 23 days when the fetuses are susceptible to dislodgement by rough handling." (B612.8.w8)
  • By day 24, the mammary glands are notably thicker than in a non-pregnant doe, if the two are compared directly. (B550.16.w16)
  • Examination and palpation can be used to determine mammary gland development and estimate the number of fetuses in anaesthetised hares. (J148.67.w1)
  • In wild Lepus europaeus - Brown hares examined under general anaesthesia, it was noted that palpation did not allow detection of pregnancy before 24 days of gestation: prior to this it was not possible to distinguish between fetuses and ingesta. (J148.68.w1)
Ultrasonography
  • Ultrasound can be used for pregnancy diagnosis and to diagnose and distinguish between Uterine Neoplasia in Rabbits, pyometra (Uterine Infection in Lagomorphs) and haemorrhage due to Endometrial Venous Aneurisms in Rabbits. (B601.5.w5, B543.15.w15, J29.10.w1)
    • Pyometra: large, fluid-filled uterus. (B601.9.w9, B602.18.w18, J213.5.w1)
    • Uterine neoplasia: uterine mass or masses; often multiple and affecting both horns. (B601.9.w9, B602.18.w18, J27.64.w4, J34.24.w3, J213.5.w1, J213.7.w1)
  • Fetuses can be assessed to see if they are alive or dead. (B543.15.w15, J29.10.w1)
  • The ovaries can be assessed for size and presence of cysts. (B543.15.w15)
  • In Lepus europaeus - Brown hares, 7.5 MHz and 10.0 MHz transducers have been used percutanously to scan the female reproductive tract in anaesthetised hares wild hares and in hares held for reproductive research. (J148.68.w1, J503.35.w1, J503.35.w4) Good contact was promoted by wetting the fur with 70% alcohol and use of ultrasonic transmitter gel. (J148.68.w1) It has been possible to evaluate:
    • Ovarian activity (e.g. presence of follicles, corpora lutea);
    • Reproductive status (pregnant/non-pregnant);
      • Detectable with percutaneous ultrasonography by 14 days of gestation. Amniotic pouch about 23 mm diameter, fetal heart rate 180 bpm. (J148.68.w1)
    • The number of embryos or fetuses and their viability (e.g. fetal integrity and heartbeat). 
    • The age of the fetuses.
      • Head and body visible by day 23 (of the 42-day gestation).
      • By day 25, lung and liver visible (lung more echogenic than liver).
      • By day 31, vertebrae, long bones, stomach and bladder detected. (J148.68.w1)
      • By day 33, cardiac septum detected.
      • By day 38, kidneys imaged.

    (J148.68.w1, J503.35.w1, J503.35.w4)

Radiography
Endocrinology 
Blood steroid analysis
  • Plasma progesterone and 17B-estradiol can be assayed using radioimmunoassay (J35.151.w2, J206.92.w1) or enzyme immuonassay (EIA). (J35.151.w2, J372.X2008.w1) 
    • These have been used in Lepus europaeus - Brown hare. (J206.92.w1, J372.X2008.w1)
    • In domestic rabbits, measuring plasma or serum progesterone taken two hours after the ovulatory stimulus can be used to confirm ovulation (progesterone concentration peaks about 10 hours before ovulation, then declines sharply). (J35.151.w2)
    • In domestic rabbits, the Ocuckeck ELISA kit (Cambridge Life Sciences) can be used to measure rabbit progesterone and reliably distinguishes between pregnant and pseudopregnant rabbits at about day 17 - 18 following insemination; if the doe is not pregnant, day 18 is an optimal time for repeat mating. J35.151.w2
  • In wild Lepus europaeus - Brown hares, serum progesterone levels were measured and correlated with pregnancy detection by ultrasonography. Mean serum progesterone concentrations were 97.9 ng/mL in pregnant hares and 14.5 ng/mL in non-pregnant hares. (J503.35.w4)
  • In Lepus europaeus syriacus (Lepus europaeus - Brown hares) stimulated to ovulate by injection of human chorionic gonadotrophin, and artificially inseminated, plasma progesterone levels rose in pregnant hares (from a baseline of < 1.0 ng/mL) starting on day 2 - 3 and reached a high of 41.4 ng/mL at about day 14, after which levels remained at about this level for a further week, then increased again rapidly in the fourth week to a peak of 67.7 ng/mL about day 28, followed by a decline to a mean of 37.5 ng/mL about a week before parturition, another peak of 55 ng/mL a few days before parturition, then falling, although still relatively high at 37.5 ng/mL 24 hours before parturition, with a rapid fall to baseline by the day after parturition. In pseudopregnant hares, progesterone levels also rose from day 2, peaking at days 11 - 18, before declining rapidly back to baseline at about day 22. (J206.54.w2)
  • In Lepus europaeus - Brown hares, LH was low (basal) during pregnancy, but in females left with a male, peaked (31.24 +/- 15.41 ng/mL) on the day of prepartum mating, 1 - 3 days before parturition, when circulating progesterone was at about 60 mg/mL. An acute rise in progesterone also occurred on the day of prepartum mating, reaching 142 +/- 15 ng/mL. FSH were high at the start and end of pregnancy, increasing rapidly to peak at day 5, remaining high to day 16, then decreasing, with a significant  rise on the same day as the LH peak, then remaining hgh for three days before declining around the time of parturition. (J206.92.w1)
Endoscopy
  • Vaginoscopy is useful in the diagnosis of diseases of the caudal reproductive tract. (J513.7.w2)
  • Laparoscopy can be used for assessment of the female reproductive organs in situ, although the ovaries may be obscured by fat. (B602.36.w36, P6.3.w1)
  • See: Imaging in Lagomorph Diagnosis and Treatment - Endoscopy
Post mortem examination
  • A wide variety of uterine pathologies, as well as the presence or absence of fetuses either within or outside the uterus, can be detected at necropsy. (B600.17.w17, J72.49.w4) 
  • Examination of the reproductive tract at necropsy can be used to investigate aspects of female reproduction on a population level in wild populations. This necessitates the use of dead females, either killed for other purposes (e.g. hunting) or for the purpose of the study. (J148.67.w1, J148.68.w1)
    • Note: recently, methods applicable to live hares (e.g. ultrasonography) have been developed). (J148.68.w1)
  • In wild Lepus europaeus - Brown hares, placental scars have been used as an indicator of previous pregnancy. (J503.35.w3, J503.35.w5) In one study, in 21 hares more than five years old (as determined by eye lens weight) in which there were no signs of pregnancies, pathological alterations of the genital tract were detected. (J503.35.w3)
Ferret Consideration The female ferret (jill) comes into oestrus in the spring (seasonal breeder). While in oestrus, the vulva is swollen to about the size of a hazelnut. The jill remains in season until mated or until the end of the breeding season. If unmated, the vulval swelling may decrease after several weeks but this does not mean the female has come out of oestrus. Prolonged oestrus causes Hyperoestrogenism in Ferrets, with resultant anaemia. (W755.Sept11.w1)
  • Except during the breeding season, the vulva is a simple slit just ventral to the anus. (B602.1.w1)
  • During the breeding season the vulva swells and protrudes (B602.1.w1) as much as 2-3 cm. Full swelling takes about two weeks to develop and the jill should not be mated until this time. The female also develops a vaginal discharge. (B651.6.w6)
  • After the jill has been mated, the vulva becomes dry and reduces in size, and she moults. (B652.7.w7)
  • Parturition should occur after 41-42 days. (B602.5.w5) 
Monitoring of Reproductive condition
  • The onset of oestrus can be detected by examination of vaginal smears. A cotton-tipped applicator is moistened with sterile saline and inserted into the vagina; the sample is then rolled across the surface of a clean glass slide, air-dried  and stained with Wright's stain. (P1.1977.w2)
  • Cells for cytological evaluation can be obtained by gently inserting a 1 mL syringe or plastic pipette 1.0  -1.5 cm into the vagina until slight resistance is felt, then flushing in 0.05 - 0.1 mL sterile physiological saline and aspirating it out again, several times. The contents of the syringe are then expelled onto a clean slide, sprayed with cytological fixative, air-dried and stained with Papanicoulaou stain or Wright's stain. During anoestrus, most vaginal epithelial cells are intermediate or superficial intermediate cells, with a few parabasal cells; superficial cells are basophilic or acidophilic with Papanicolaou stain, rarely orangeophilic (which indicats keratinisation). During oestrus, usually more than 90% of cells are superficial cells. In the first 3-5 days of estrus, superficial cells show mixed staining properties (basophilic, acidophilic or orangeophilic with degenerated nuclei and by 4-6 days, cells are highly keratinised and mainly anucleate. Neutrophils may be present at any stage of the cycle. Bacteria are rarely present and are found in association with superficial cells. If oestrus is prolonged, numerous bacteria may be present together with neutrophils, cellular debris and erythrocytes. During proestrus (lasting 2-3 weeks) the size of the vulva increases. (J212.4.w4)
  • After mating, a vaginal smear can be examined to confirm the presence of sperm. (P1.1977.w2)
Detection of pregnancy

The fetuses can be detected by gentle palpation by about 14 days after mating. (B631.17.w17, B602.1.w1); by 21 days. (B652.7.w7)

Pregnancy can be diagnosed using ultrasonography from about 14 days. (B631.17.w17, B602.1.w1)

  • Ultrasonography assists with determining gestational age, number of fetuses and viability of fetuses, as well as confirming pregnancy. (B631.27.w27)

In the last days of pregnancy the teats become visibly enlarged. (B631.17.w17)

Inducing Parturition
  • Litters of only one or two kits may not produce sufficient hormonal stimulus to initiate parturition. Prolonged pregnancy (43 days) may result in oversized kits and therefore dystocia, and if parturition does not occur by this date the kits may die in utero. If palpation indicates only one kit, induce parturition at 41 days by giving 0. 5 mg prostaglandin F, then 1-4 hours later 6 USP units of oxytocin. Expect parturition 2-12 hours later. If the jill does not deliver the kit(s) within 24 hours, repeat the treatment of perform a Caesarean section. (B602.5.w5) 
Bonobo Consideration Observation

The sexual swellings of female bonobos should be observed and the state of the swelling recorded, as well as the occurence of menstuation.

  • A regular pattern of penineal swelling is generally taken as an indicator of normal ovarian function.
  • If the pattern of swelling is altered or even absent, this may indicate:
    • Pregnancy.
    • Ovarian neoplasia
    • Ovarian failure
    • Altered pituitary function leading to reduced stimulation of the ovaries to produce the normal hormones (oestrogen and progesterone).
    • Old age.
  • Lack of both swelling and menstruation for a long period, in a female which is not raising an infant, probably indicates a pathological condition
  • Prolonged swelling with erratic reductions in swelling probably indicates infertile cycles; these may be seen in adolescents and lactating females, also in females reaching a post-reproductive phase.
  • Intra-menstrual intervals with little or no pre-swollen or post-swollen phase suggests pathology and may be associated with  a pre-menopausal stage.

(D386.2.3.w2c)

Detection of ovulation and pregnancy using human urinary test kits

  • Bonobos can be trained to urinate in response to a cue (P129.1.w5), or can be kept in an overhead chute (between living areas), with a wire floor, until they urinate. The urine can then be collected from the floor using a syringe. This allows collection of urine from a specific bonobo. (J54.30.w1)
    • Human ovulation test kits can be used to detect ovulation and pregnancy in bonobos. The kit OvuQuick One Step Ovulation, Nanorepro AG, Marburg, GE, which is based on qualitative detection of luteinising hormone (LH) in urine was used to monitor ovulation in bononbos at Milwaukee Zoo, Wisconsin, USA. (J54.30.w1)
  • Human pregnancy test kits can be used to confirm pregnancy in bonobos. At Milwaukee Zoo, Wisconsin, USA, Accustrip hCG Pregnancy Test (JANT Pharmacal Corporation, Encino, California, USA) was used; the test is based on qualitative detection of human chorionic gonodaotropin (hCG). The test was carried out if a female did not show menstrual bleeding at the expected time. (J54.30.w1)

Monitoring pregnancy using ultrasonography

  • Ultrasonography can be used to monitor the progression of pregnancy in bonobos. With appropriate positive reinforcement training, bonobos can be scanned without either physical or chemical restraint. this allows repeated scans and monitoring of the fetus during the whole gestation. (J54.30.w1, P129.1.w2) See Mammal Handling & Movement (Mammal Husbandry and Management)
    • Ultrasonography allowed monitoring of fetal heart rate, body movements and swallowing, and thereby of fetal well-being. (J54.30.w1)
    • The examinations allowed size and position to be monitored for the fetal heart, lungs, kidneys, urinary bladder, gastric vesicle, limbs, umbilical cord insertion, features of the face, and the cerebellum. 
    • Measurements of the size of the embryonic vesicle and the embryonic crown-rump length were recorded during the first trimester, with head circumference, biparietal diameter, abdominal circumference and femur length measured during the second and third trimesters. These measurements allowed the construction of fetal growth curves. (J54.30.w1)
    • The formation of the placenta and a qualitative assessment of the amniotic fluid were possible also. (J54.30.w1)

Radiography

  • Radiography with positive contrast medium introduced via the cervix has been used in  investigation of infertility. (J339.47.w1)

Laparoscopy

  • Laparoscopy has been used to visualise the internal reproductive organs in the investigation of infertility. (J339.47.w1)

Biopsy

  • Endometrial biopsy has been used in investigation of infertility. (J339.47.w1)
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Monitoring Reproductive Health in the Male 

Failure of reproduction may be associated with male infertility. Evaluating the male for reproductive health is as important as examining the female.

Behavioural observation
  • An important part of reproductive monitoring is behavioural monitoring. (J23.38.w2)
  • Males housed in bare (non-enriched) environments may fail to show breeding behaviour; provision of a more complex environment may improve breeding behaviour. (B429.39.w39)
History
  • Information should be gathered on the animal's age; any previous disease or injury; whether the individual has previously been fertile (produced pregnancies and offspring); an estimate of libido; and whether the animal has been exposed to any pesticides, heavy metals or other potential toxins. Additionally, consider the captive environment and how well close relatives have reproduced. (B429.39.w39)
    • It is important to remember that the effect of an "insult" to the testes may take several weeks to become apparent, due to the length of the spermatogenic cycle. (B429.39.w39)
Physical examination
  • The genitalia should be examined in detail. Note the size, tonicity and integrity of each testis. Note whether there are any adhesions within the scrotum (indicated by the testes not being normally mobile within the scrotal sac). Observe and palpate the penis and sheath; check for any persistent frenulum, phimosis (abnormal contraction of the sheath) or adhesion preventing proper protrusion of the penis (and therefore preventing vaginal penetration), as well as for abnormalities such as penile fibropapillomas. (B429.39.w39)
  • Cryptorchidism (undescended testes) is highly heritable, therefore males which are unilateral cryptorchids (i.e. one testis has descended) and fertile should not be used for breeding. (B429.39.w39)
  • Persistent frenulum and congenital (rather than traumatic) phimosis are generally genetic in origin, therefore males with these conditions should not be used for breeding. (B429.39.w39)
  • The accessory glands may be assessed by palpation per rectum; chronic inflammatory conditions of the accessory glands often reduce fertility. (B429.39.w39)
  • Note: The state of the testes (e.g. size, tonicity) must be assessed in relation to season in seasonally-breeding species. (B429.39.w39)
Ultrasonography
  • The use of real-time ultrasound examination is of increasing use for monitoring reproductive health and reproductive status. (B429.38.w38, B445.w4, P6.4.w6)
  • This method allows the organs to be observed without the need for any surgical invasion. (B429.38.w38, B445.w4, P6.4.w6)
  • The main advantages of ultrasonography are that it is non-invasive; gives real-time, visual, reproducible information about the organs; gives cross-sectional images of the tissues/organs, including images of organs which are moving; allows organs or other objects to be measured; can be integrated easily with other methods of examination; and can be stored. (B445.w6, P6.4.w6)
  • The main disadvantage is the need for physical restraint and often for sedation or anaesthesia. (B429.38.w38, B445.w4, B445.w6)
  • Other potential problems include the need for a good acoustic coupling with the animal, which may be disrupted for example by fur; insufficient penetration of sound waves through tissues; the need for modification of some components to allow for different species and the need to develop experience with different species, as well as cost issues. (B445.w6)
  • Ultrasonography can be used for sex determination of monomorphic species. (B429.38.w38, B445.w6)
  • Ultrasound is able to detect pathological changes in the genitalia, such as calcification, fibrosis, cysts or tumours in the testes, and also may allow detection of pathological changes in other organs, which may affect general health and reproductive function. (B445.w6, P6.4.w6)
Endocrinology
  • This is less used in assessing male reproductive function than in assessing female reproductive function. (B429.39.w39)
  • Levels of hormones may be assessed in blood, or concentrations of hormone metabolites in excreta (urine, faeces) may be measured. (B429.39.w39)
  • Generally, serial samples are required, with little data being gained from a single sample. (B429.39.w39)
  • Serum testosterone levels may be assayed to confirm puberty (sexual maturity), and to indicate problems such as deficiency in pituitary gonadotrophins, and profound Leydig cell dysfunction. (B429.39.w39)
  • Testosterone levels show marked seasonal variations in many species, and also diurnal variation, therefore samples which are to be compared with one another should be taken at about the same time of day.
    • There may be marked variations in testosterone level between species. 
    • Abnormally low testosterone levels are associated with low libido and with infertility. 
    • Very high testosterone levels may be seen in association with an interstitial cell tumour. 

    (B429.39.w39)

  • The response of the male to an injection of GnRH (gonadotropin-releasing hormone) can be tested: 25-100 µg GnRH is injected intravenously or intramuscularly, and samples are collected at 15-30 minute intervals from before injection to two or three hours after injection. GnRH should stimulate release of LH (luteinizing hormone) from the pituitary and result in a detectable rise in serum testosterone within 15 to 90 minutes; failure of this response indicates an abnormality in the pituitary or Leydig cells. (B429.39.w39)
    • Increased LH in response to GnRH can also be detected; failure of such increase indicates pituitary dysfunction. (B429.39.w39)
  • Interpretation of baseline LH, rather than response to stimulation, is complicated by the pulsatile release of this hormone, so that even relatively frequent sampling such as every 15 minutes may fail to detect an LH pulse, and by possible interference with FSH (follicle-stimulating hormone)/LH patterns by anaesthesia. (B429.39.w39)
  • Measurement of oestrogen levels in blood or urine may indicate the presence of Sertoli cell tumours or seminomas, although not all Sertoli cell tumours secrete oestrogen. (B429.39.w39)
  • High levels of stress hormones (glucocorticoids) may be associated with the presence of large numbers of pleomorphic (various-shaped) sperm. (B429.39.w39)
Semen evaluation
Collection of semen for examination
  • Semen for evaluation generally is collected by electroejaculation carried out on an anaesthetised individual. (B429.39.w39)
    • Electroejaculation requires the animal to be in a surgical plane of anaesthesia. The anaesthetised animal is placed in a position allowing access to the anus and the glans penis. Copper or stainless steel electrodes are arranged longitudinally, or less commonly as rings, on a plastic or Teflon rectal probe with a diameter about the same as that of a normal fecal bolus. Longitudinal electrodes may be arranged on one side of the probe only, to provide stimulation ventrally, towards the accessory glands. The ejaculate should be collected into a pre-warmed (to prevent cold-shock) sterile plastic vial. (B429.39.w39)
    • Stimuli, either direct current or alternating current, are applied generally in groups and with increasing voltage, generally starting at 2.0 to 5.0 volts and rarely exceeding 10 volts. (B429.39.w39)
    • An ejaculate containing sperm is generally produced in response to electrical stimulation. (B429.39.w39)
    • Contamination with urine may occur; this is more common with the use of some sedatives including xylazine, diazepam and acepromazine, all of which relax the urethral musculature. (B429.39.w39)
    • Electroejaculation using a rectal probe is safe when used correctly; males show normal sexual behaviour and may mate and fertilise females within days of electroejaculation. (B429.39.w39)
  • Individuals of some species have been trained to allow semen collection into an artificial vagina (various species), or by manual stimulation of the penis (some canids (Canidae - Dogs, foxes (Family))). (B429.39.w39)
  • It is possible to collect mature spermatozoa post mortem, by flushing the ductus deferentia and caudal epididymes with warmed saline or air. If there is a delay in collection, the tissues should be maintained at 5 °C prior to collection. (B429.39.w39)

Evaluation

  • The volume, colour and pH of the ejaculate should be tested immediately. A yellow colour indicates urine contamination; this also will lower (in carnivores) or raise (in ungulates) the pH. (B429.39.w39)
  • Other common evaluations are for sperm concentration (this can be greatly reduced with urine contamination), percentage motile sperm and forward motility, both assessed at 37°C, percentage abnormal sperm, with records of types of abnormalities (e.g. abnormal head shape or size, missing midpiece, coiled flagellum, biflagellate, bicephalic or a defective acrosome). (B429.39.w39)
    • For samples with very high sperm concentrations (greater than one billion sperm per millilitre) it is necessary to dilute the semen (in saline or an appropriate culture medium) before motility can be evaluated. (B429.39.w39)
    • Motility estimates should be repeated for several hours at intervals of 30 to 60 minutes, since infertility has been reported associated with rapid loss of motility. (B429.39.w39)
    • Reduced longevity in vitro is common with samples which are urine contaminated; seminal fluid constituents also can reduce motility. (B429.39.w39)
    • More complex evaluations of sperm viability can be carried out, such as the ability of sperm cells to move through cervical mucus (assessed using a polyacrylamide gel) and sperm penetration assays looking at penetration of heterologous ova. (B429.39.w39)
  • Examinations should be made on several (three to four) ejaculates collected at intervals of three to four weeks before a judgment is made on the male being infertile. (B429.39.w39)
Testicular biopsy
  • Biopsies can be used to assess the presence of cellular elements within the testis, the presence or absence of sperm production, and the severity of degeneration of the testis in acquired diseases. (B429.39.w39)
    • Anaesthetise the animal. Surgically prepare the scrotum. Stabilise the testis within the scrotum. Using an ultrafine surgical blade, make an incision through the scrotum, tunica vaginalis and tunica albuginea, and though into the testis to a depth of 2 - 3 mm. Using moderate pressure at the neck of the scrotum, press the surface of the testis though the incision. Shave the testicular sample off the testis with a sharp scalpel. Place the sample in glutaraldehyde (not formalin; formalin distorts chromatin patterns and the walls of the seminiferous tubules). Suture all layers of the biopsy site. Give prophylactic systemic antibiotics. (B429.39.w39)
  • Alternatively a needle biopsy may be taken but this often does not provide sufficient tissue for histological evaluation. (B429.39.w39)
Bear Consideration The size and weight of the sexual organs in bears varies with body size. The size of the testes of Ursus arctos - Brown bear, Helarctos malayanus - Sun bear and Melursus ursinus - Sloth bear were found to be similar to one another, but the size of the accessory glands was related to body size. (P9.2004.w5)
  • Ursus arctos - Brown bear (200 - 250 kg bodyweight); testis diameter 26.1 +/- 7.1 mm (breeding season, n=4); 17.9 +/- 2.2 mm (non-breeding season, n=2).
  • Helarctos malayanus - Sun bear (70 - 85 kg bodyweight) testis diameter 24.5 +/-3.8 mm (breeding season, n=4); this species is a non-seasonal breeder.
  • Melursus ursinus - Sloth bear (74 - 156 kg bodyweight) no breeding season data; testis diameter 17.8 +/- 8.4 mm (non-breeding season, n=4).
  • Tremarctos ornatus - Spectacled bear (150 - 160 kg bodyweight; testis diameter 26.6 +/1 3.1 mm (breeding season, n=8); 25.7 +/- 2.1 (non-breeding season, n=3).

    (P9.2004.w5)

Physical examination
  • The presence of the testes can be confirmed, and their size measured, in anaesthetised males. It may also be possible to detect abnormalities such as the presence of a tumour. (J296.55.w1, P6.4.w3)
Ultrasonography
  • Ultrasonography can be used to assess the reproductive system in male bears. The testes can be scanned and measured transcutaneously, while the internal organs can be scanned transrectally. The size of the testes and accessory glands can be determined and abnormalities (e.g. tumours, fibrosis, calcifications) detected. Ultrasonographic examination allows detection of abnormalities in the male reproductive tract before they can be detected by physical examination and before any apparent effect on sperm quality. (P6.4.w3)
    • In a study, a B-mode ultrasound scanning system was used, with a 10 MHz linear array for transcutaneous imaging of the testes and a 7.5 MHz curved linear transducer fitted into a specially designed adapter used transrectally for imaging internal structures such as the urinary bladder, urethra, prostate gland and ampulla. Testis, prostate and ampulla volume were calculated from measurements of their length, width and height. Abnormalities such as testicular fibrosis, a tumour in the testis of one Tremarctos ornatus - Spectacled bear and a hyperechoic structure in the ampulla were detected. (P6.4.w3)
Endocrinology
  • A study of Ursus americanus - American black bears found that serum prolactin, measured with a radioimmuneassay, varied seasonally, being lowest in December (1.1 +/- 0.1 ng/mL (mean +/- SD), and highest in May (17.6 +/- 4.7 ng/mL), preceding the June testosterone peak. Prolactin concentrations were considered to be a useful indicator of testicular steroidogenic activity. (P1.1993.w3)
Electroejaculation
  • Electroejaculation can be used to assess semen characteristics such as ejaculate volume, sperm concentration, total sperm count, sperm motility, viability and abnormal morphology. (J296.55.w1, P6.4.w3)
Post mortem examination
  • Thorough examination of the reproductive tract at necropsy allows correlation with, and confirmation of, findings from ante-mortem examinations such as ultrasonography. (P6.4.w3)
Lagomorph Consideration Infertility in the male may be congenital (can occur in a significant proportion of domestic rabbit bucks). or associated with infectious diseases: 

(B618.21.w1)

Physical examination
  • The genitalia can be assessed by physical examination. In males older than about 10 - 12 weeks, the testes should be seen in the scrotal sacs on either side just cranial to the penis; they should be symmetrical, smooth and non-painful. (B601.2.w2)
  • The position (intra- or extra-abdominal) of the testes can be determined during examination of live wild hares; this can be useful in assessing the timing of the breeding season. (J148.67.w1)
  • Testicular swelling may indicate: (B601.App2.w20) 
Ultrasonography
  • Ultrasound can be used for assessing the reproductive tract in male lagomorphs. (J503.35.w1, J503.X.w1)
  • In wild Lepus europaeus - Brown hare, a 7.5 MHz transducer was used rectally to image the pelvic portions of the male genital tract. (J503.35.w1)
  • The contents of the scrotum of 28 New Zealand white rabbits was evaluated using a 5 MHz linear array transducer, with the rabbits in dorsal recumbency. For the examination, a rubber pad, 0.8 cm thick and with a central hole 0.5 cm diameter was placed between the testis and the rabbit's body, ultrasound coupling gel was applied to the scrotum, and a rubber sac containing 250-300 mL tap water was placed over the scrotum as a stand-off. (J503.X.w1)
    • The dimensions of the testes were measured (right testis average 3.2 +/- 0.08 cm (mean +/- SEM) long, 1.1 +/- 0.03 cm wide; left testis mean 3.3 +/- 0.09 cm long, 1.1 +/- 0.03 cm wide). (J503.X.w1)
    • The testicular parenchyma was described as "homogenous and moderately echoic" while the cauda of the epididymis seen in sagittal plane images was "a homogenous, less echoic structure localised next to the testicular parenchyma" and the caput epididymis was "smaller than the cauda, homogenous and less echoic compared with the testicular parenchyma" but could only be visualised in eight rabbits (28.6%). (J503.X.w1)
    • Abnormalities which could be detected included biopsy needle scars (hyperechoic, from 3-4 days to day 14 after biopsy), while in a testis undergoing necrosis, reduction in testicular volume and variation in echogenicity were noted. (J503.X.w1)
Electroejaculation
  • Semen can be collected from rabbits and hares by electroejaculation. (J372.93.w1, J372.X2008.w1)
  • In wild Lepus europaeus - Brown hare, semen samples were collected by electroejaculation using specially designed equipment. (J503.35.w1)
Endocrinology
Semen evaluation
  • Average ejaculate volume in a medium-sized domestic rabbit is 0.8 mL. (B550.16.w16)
  • Samples can be assessed by light microscopy. (J372.93.w1)
  • The concentration of sperm can be assessed in a Thom-Zeiss chamber, using the cytometric method. (J372.93.w1)
  • Staining using a Live/Dead Sperm Viability Kit (L-7011, Molecular Probes) can be used to assess the percentage of live sperm. (J372.93.w1)
  • In wild Lepus europaeus - Brown hare semen samples were evaluated for volume, colour, concentration of sperm, sperm motility and sperm morphology. (J503.35.w1)
  • Note: there is variation is semen composition between individuals, also in an individual there are variations such as cyclic changes in sperm count. (B550.16.w16)
Post mortem examination
  • The position (intra- or extra-abdominal) of the testes can be determined; testicular mass can also be measured. These can be useful in assessing the timing of the breeding season. (J148.67.w1)
  • Histological examination of testicular parenchyma can be used to assess seasonal regression and reactivation of testes. (J503.35.w2)
  • Orchitis and Epididymitis in Rabbits may be detected at necropsy. (B600.17.w17, J72.49.w4)
Ferret Consideration
  • On the male, the preputial opening is easily seen on the ventral surface of the abdomen, caudal to the umbilicus, and the os penis can be palpated. (B602.1.w1)
  • Outside the breeding season the male's testes are small and intra-abdominal. In early spring, they enlarge and descend into the scrotum. (B651.6.w6)
  • Enlargement and descent of the testes starts once day length increases. As testosterone production increases, the hob puts on weight and the activity of the skin glands increases, with a resultant increase in odour as well as stickiness of the fur coat. (B631.17.w17)
  • Spermatogenesis starts in December and continues to July. (B602.1.w1)
  • Electroejaculation with a rectal electrode can be used in an anaesthetised ferret to obtain a sperm sample. The semen is mixed with an extender then can be examined for sperm concentration and mobility (and can be used for artificial insemination or cryopreserved for later use). (J206.86.w1, P1.1977.w2)
    • Anaesthesia with medetomidine or xylazine plus ketamine is less likely to rasult in urine contamination than diazepam, acepromazine or inhalational anaesthetics.
    • Semen needs to be collected into a warm collection vessel (glass or plastic) to avoid cold shock of the sperm.
    • Three series of stimulations can be given, totalling 80 stimulations, In each, the voltage is increased from zero to the maximum required voltage over a period of a second, held for 2-3 seconds then stopped, with a three second break between stimulations and a 2-3 minute rest between sets of stimulations. Suggested:
    • Series one: ten stimulations at 4V, ten at 5V, ten at 6V;
    • Series two: ten stimulations at 5V, ten at 6V, ten at 7V;
    • Series three: ten stimulations at 6V, ten at 7V.
    • Usually 0.05 +/- 0.01 mL semen can be collected by this method, with about 706 x 106 spermatozoa/mL, with 67% normal sperm and 80% motility. (B232.17.w17)
Bonobo Consideration
  • The external genitalia of primates should be examined as part of a full physical examination e.g. when individuals are moved between collections. (B10.44.w44f)
  • Male bonobos can be trained to allow masturbation and semen collection by keepers; collected semen can be used for artificial insemination [as well as for evaluation of male fertility]. (P129.1.w5)
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Assisted Reproduction

Assisted reproduction includes "any technique that circumvents natural procreation to produce young." The most commonly used assisted breeding techniques are artificial insemination and embryo transfer. (J23.38.w2)

Note: 

  • Assisted reproduction can be a part of conservation, but is not sufficient in itself for conservation. (J3.143.w10, J370.57.w3) Assisted reproduction should only be used as part of an overall breeding programme, to counteract the effects of human interference which may make breeding difficult. (P1.1976.w1) While assisted breeding has been responsible for some landmarks in breeding of wild carnivores, these techniques are only a very small part of conservation. (J370.57.w2)
  • Assisted reproduction techniques have only limited applicability for use in non-domestic species. Techniques developed for assisted reproduction of humans and domestic animals cannot easily be applied to other species; much needs to be known about the reproductive physiology of the species before even the simplest techniques, such as artificial insemination, can be used successfully. (B23.61.w19, J23.38.w2, J370.57.w3)
  • Effective use of assisted reproductive technologies requires considerable research for each species: to characterise normal sperm traits and develop methods for cryopreserving and thawing sperm; to characterise the reproductive cyclicity of the female and understand the timing of ovulation; to understand seasonal aspects of reproduction; to learn the in vivo and in vitro requirements of gametes; to characterize ovulation timing, oestrus synchronization, induction of ovulation, and maturation of oocytes; to develop appropriate in vitro sperm-oocyte interaction and culture conditions for gametes and embryos; to find the appropriate site for deposition of sperm or embryos in the female; and to understand the effect of stress on reproduction. (J23.38.w2)
Benefits of assisted reproductive techniques

In non-domestic animals, reproductive technologies including assisted reproduction and germ plasm banks may be useful to:

  • Improve fertility. (B445.w4)
  • Stimulate reproduction in individuals which are genetically important and increase founder representation in small populations. (B23.61.w19, B445.w4)
  • Allow production of offspring while the male and female are kept at different locations, reducing the need for shipping individual animals between collections, with associated reduction in risks, permits and expense. (B23.61.w19, J3.143.w10, J23.38.w2, J51.18.w1, P1.1976.w1)
    • This also removes the possibility that, after the transfer has taken place, the two animals turn out not to be a compatible pair. (J23.38.w2, P1.1976.w1)
  • Reduce the risk of transfer of microparasites and macroparasites (which may be transferred between locations with their host). (B23.61.w19, J51.18.w1, P1.1976.w1)
  • Allow insemination when a pair of animals are behaviourally incompatible, or there is a risk to the female during normal mating. (B23.61.w19, J51.18.w1)
  • Introduce new genetic material into captive populations, either from other captive populations or from the wild. (B23.61.w19, B445.w4, J51.18.w1, P1.1976.w1)
    • This could reduce the need to bring new wild animals into captivity to maintain the genetic diversity of the captive animals. (B23.61.w19, J23.38.w2)
  • Introduce new genetic material into isolated wild populations, either from other isolated wild populations or from captive animals (i.e. genetic transfer between in situ populations or from ex situ to in situ populations). (B23.61.w19, B445.w4, J23.38.w2)
  • Allow preservation of genetic material (frozen semen) for future use, including from individuals in (presently) healthy wild populations (i.e. develop genome resource banks), preserving genetic diversity and providing insurance against genetic loss, disease and unnatural selection. (B445.w4, J51.18.w1,J23.38.w2)
  • Possibly improve bloodlines, reducing or eliminating undesirable genetic traits. (P1.1976.w1)
  • Allow breeding of individuals which are behaviourally unsuitable for mating, (B445.w4) for example due to imprinting. Note: it is preferable to avoid imprinting or development of other behavioural impediments to normal mating. 
  • Allow breeding despite anatomical problems, (B445.w4)  e.g. when a male is fertile but has suffered a penile injury reducing his ability to mate naturally, or if a genetically valuable female has an injury such as a broken pelvis making normal parturition impossible.
  • Increase population growth. (B445.w4)
Artificial insemination

Artificial insemination is the assisted reproduction technique most likely to be of practical use for wildlife propagation. (J23.38.w2)

  • Artificial insemination involves collection of semen from the male and deposition of the semen into the female. The semen may or may not be stored before use, either short term (days, in a proper medium) or longer term (frozen). (J23.38.w2)
  • In domestic mammals, artificial insemination is commonly used in farm species, particularly cattle, for convenience (the farmer does not have to maintain a bull), to improve desired characteristics (by using semen from a bull proven for such characteristics) and to allow a given bull to sire greater numbers of offspring.
  • Collection of semen from most mammal species requires electroejaculation of the male, under deep anaesthesia. A rectal probe of an appropriate size and shape for the species is used, with either AC or DC power. Semen collected is evaluated microscopically as well as macroscopically. Semen can be used for immediate insemination, or placed in an appropriate extender and held for up to two to three days at 5°C, or frozen for storage (equilibrated at 5°C, pelleted on dry ice then deposited in liquid nitrogen). It may be possible to use frozen semen successfully after several years of correct storage. (P1.1976.w1)
  • Note: Collection of semen from conscious mammals by electroejaculation is painful. (J296.64.w1)
  • Sperm from different species, and even from different individuals within the same species, may show different susceptibilities to freezing and thawing, i.e. different percentages of sperm which are mobile and able to fertilise an egg following freezing and thawing. (B445.w8)
  • In non-domestic mammals, deposition of sperm into the female requires an anaesthetic, which has deleterious effects on sperm transport. Fertility is much higher if laparoscopic intrauterine deposition is used rather than intravaginal deposition. (B23.61.w19)
  • Note: While artificial insemination may have an important role to play in breeding programmes for some endangered species, it should be seen only as one tool in such a programme, not as an end in itself. (J51.18.w1, P1.1976.w1)
  • Note: In some countries there may be legislation restricting the use of electroejaculation for semen collection to licensed veterinarians.
Oocyte collection (gamete rescue)
  • Oocytes can be "rescued" from the ovaries at necropsy or at the time of ovariohysterectomy. Generally, immature eggs are collected from the ovary and matured in vitro. This technology has been proved by use in domestic cats (oocyte retrieval, maturation, fertilisation, embryo transfer and subsequent birth of live young). Immature eggs from a variety of wild felids have been collected and matured in vitro. (B23.61.w19)
In vitro fertilisation
  • This has been used primarily in felids. (B23.61.w19)
  • Development of ovarian follicles is stimulated by injecting equine chorionic gonadotrophin (eCG) or using follicle-stimulating hormone (FSH) followed by human chorionic gonadotrophin (hCG). (B23.61.w19)
  • Oocytes can be collected by transabdominal laparoscopy; this is comparatively easy in felids but more difficult in some other taxa (e.g. in mustelids (Mustelidae - Weasels (Family)) the ovaries are embedded in adipose tissue and in canids (Canidae - Dogs, foxes (Family)) they are enclosed in a bursa, therefore it is not possible to observe ovarian activity or retrieve oocytes). (B23.61.w19)
  • Collected oocytes are examined by stereomicroscopy and oocyte-cumulus cell complexes classified as mature (with loose and expanded oophorus cells) are co-cultured with sperm for about 20 hours then assessed for fertilisation and embryo cleavage at about 30 hours post insemination.
  • Confirmed embryos can be transferred into an appropriate female. (B23.61.w19)
Embryo transfer
  • Embryo transfer involves the collection of fertilised ova or embryos from one female and deposition into the uterus of another female. Either fresh or frozen embryos may be used. (J23.38.w2)
  • Embryo transfer has been highly developed for use in domestic cattle, to increase reproduction from highly desirable cows. It has been used in non-domestic animals, but its use is likely to remain minor.
Bear Consideration Wildlife Information Network again thanks WSPA for its invaluable sponsorship without which the Wildpro "Bears: Health and Management" volume would not have been researched and published. A message from WSPA follows:
LOGO-WSPA OrangeBlack.jpg (31514 bytes) WSPA does not condone the captive breeding of bears unless there is a programme to rehabilitate and release the cubs back to the wild as part of a valid conservation or animal welfare programme.

WSPA believes that bears should not be kept in captivity in enclosures (including zoo enclosures) where their living environment lacks sufficient space and natural habitat to enable the bears to exhibit their natural behaviour.
Artificial Insemination

Artificial insemination could be used as a means of transferring genetic material between isolated bear populations in the wild, and between captive collections.

Embryo transfer
Embryo transfer could be used as a way to protect existing genetic diversity in bears (J296.51.w1) and to transfer genetic material between isolated bear populations.
  • Bears show seasonal breeding (except for Helarctos malayanus - Sun bear) and delayed implantation. These features increase the potential usefulness of embryo transfer, since transplantation can be carried out without the donor and recipient having their reproductive cycles synchronised. (J296.51.w1)
  • It has been proven possible to collect a blastocyst nonsurgically from an Ursus americanus - American black bear, and to laparoscopically transfer the blastocyst to a recipient Ursus americanus - American black bear, resulting in the birth of a live, apparently normal cub (the cub, together with a cub genetically descended from the recipient, died at several weeks old when the maternal den was flooded). (J296.51.w1)
    • Further work is required to improve the technique for passing the stylet through the cervix, which is tortuous, for embryo collection and to allow non-surgical implantation into the recipient. Use of oestradiol cypionate (156 µg/head) has improved passage of a stylet through the cervix. (J296.51.w1)
Oocyte recovery and maturation
  • Oocytes have been recovered from antral follicles of the excised ovaries of bears (hunter-killed Ursus americanus - American black bear, and euthanased elderly Helarctos malayanus - Sun bear and Melursus ursinus - Sloth bear), and incubated to produce nuclear maturation (indicated by development to metaphase II chromosome configuration). Unsurprisingly, efficiency of recovery of oocytes capable of maturation appeared to be affected by the age, reproductive status and general health status of the bears involved. It was noted that considerable research would be required before "oocyte rescue" could be used for ursid conservation. (J369.269.w1)
Lagomorph Consideration
Artificial insemination
  • Artificial insemination can be carried out in rabbits. (B550.16.w16, B614.2.w2)
  • Artificial insemination has been carried out in hares (Lepus spp.) and Sylvilagus spp. (J206.10.w1, J296.4.w1)
Semen collection
  • In domestic rabbits, semen can be collected from the buck into an artificial vagina of appropriate size. (B550.16.w16, B614.2.w2)
    • Note: This is unlikely to be useful for wild lagomorphs. (J206.43.w1)
  • An artificial vagina for rabbits generally consists of a plastic or thick-walled rubber casing and a thinner rubber liner. (B550.16.w16)
  • For example, 1.25 inch diameter radiator hose, 5 inches long, lined with surgical drainage tubing and leading to a 5 mL test tube, attached to the tubing via a rubber stopper with a hole bored through), maintained at about 40 °C (not over 45 °C) by use of a water jacket. (B614.2.w2)
    • If the temperature is too low, the buck may not ejaculate; if it is too warm, the sample may be contaminated by urine. (B550.16.w16, J35.151.w2)
  • Either a female teaser rabbit is used or a rabbit skin is worn over the arm of the person collecting the sample and is held such that it is easy for the buck to mount it. (B614.2.w2, B550.16.w16, J35.151.w2)
    • A young male can easily be trained to mount a rabbit skin. The male should be allowed to approach the skin and sniff it when it is first introduced, not have the skin moved towards the rabbit, which may be intimidating. Usually the male will soon approach and mount, ejaculating as soon as the penis has entered the artificial vagina (held in the operator's hand under the skin). Initially, during training, the skin should be introduced every day or every second day. (J35.151.w2)
  • Usually collections are made twice a week. Daily collections have been made for a year. (B614.2.w2)
    • Semen collected from daily collections contain more (two to four times) spermatozoa than those from weekly collections, but excessive collection can cause decreased semen production and decreased sex drive. (B614.2.w2)
    • With semen collection very frequently (two or three times a day for nine months, seminal fluid was found to contain antibodies to sperm antigens. (J35.151.w2)
  • There are both breed and individual differences in semen production. (B614.2.w2)
  • In hares (Lepus europaeus - Brown hare), collection of semen by artificial vagina apparently is not possible. (J372.93.w1)
  • Semen can be collected from Lepus europaeus - Brown hare by electroejaculation under anaesthesia. (J372.93.w1, J372.X2008.w1)
    • If no ejaculate was obtained after four or five impulses, the collection attempt was cancelled. (J372.93.w1)
    • An interval of at least one week between collection of samples. (J372.93.w1)
  • Sperm mass has also been collected from lagomorphs post mortem or at castration, by slicing or cutting the epididymis or the vas deferens and epididymis in 0.9% physiological saline or another appropriate diluent. (J346.46.w1, J369.144.w1, J495.28.w4)
  • Semen has been collected directly from the epididymis of hares (Lepus americanus - Snowshoe hare, Lepus europaeus - Brown hare) and Sylvilagus floridanus - Eastern cottontail and used successfully for artificial insemination. (J206.10.w1, J206.54.w1, J296.4.w1)
    • Semen was collected from the epididymis following castration. (J495.28.w4)
    • Semen was collected from the epididymis of anaesthetised male Lepus europaeus - Brown hare. (J206.54.w1)
Semen dilution and storage
  • Semen can be stored for up to 12 hours undiluted and used for insemination, or diluted in glucose-yolk-citrate and used within four day. (B614.2.w2)
  • Successful insemination can be made after diluting the semen up to 10 times in physiological saline or in Ringer's solution or Krebs-Ringer solution. (B614.2.w2, J35.151.w2)
    • Antibiotic should be added. (J35.151.w2)
  • Semen from rabbits can be stored for longer periods in Tris buffer-based diluents with glucose and citrate, plus a cryoprotectant for frozen storage, commonly dimethylsulphoxide (DMSO). (J372.64.w1, J372.93.w1)
    • Semen is diluted in Tris-citrate-glucose-DMSO extender at 33 °C then cooled to 5 °C over 90 minutes, then cooled in straws (0.25 mL) above liquid nitrogen for 10 minutes then plunged into the liquid nitrogen. To thaw, straws are thawed in a water bath at 37 °C for 30 seconds. (J372.93.w1)
  • The rates of freezing and thawing can greatly affect the quality of sperm following thawing. In one study, using semen in Tris-citric-acid-glucose extender with 1.75M DMSO and 0.05 M sucrose as cryoprotectants, freezing in liquid nitrogen vapour for ten minutes gave significantly (P < 0.01) higher post-thaw fertility compared with freezing in a freezer at minus 30 °C. Thawing at 50 °C for 10-12 s produced higher numbers of young born and numbers of live young than thawing at 70 °C for 10 - 12 s. (J296.60.w1)
  • Semen from Lepus europaeus - Brown hare has been frozen diluted in extender and gave acceptable motility post-thaw. (J372.93.w1)
    • Extender used (rabbit semen extender): Tris 3.028g, citric acid 1.675 g, glucose 1.25 g, dimethylsulphoxide (DMSO) 4.5% v/v, egg yolk 17% v/v and distilled water to 100 mL). Dilution of ejaculates of at least 0.4 mL, greater than 75% motile spermatozoa and sperm concentration over 250 x 106 per mL to give 5 x 106 spermatozoa per 0.25 mL. Post thaw, motility 40.50 +/- 4.21%, live spermatozoa 35.05 +/- 4.21%, percentage with normal acrosomes 76.10 +/- 3.69%. (J372.93.w1)
Insemination of the female
  • For successful fertilisation, 0.3 - 0.7 mL of sperm suspension, containing 20 - 50 x 106 spermatozoa should be inseminated into the female. (B614.2.w2)
    • As low as 1 x 106 spermatozoa may produce acceptable fertility. (B614.2.w2)
    • Semen should be examined at least for apparent viability and motility, before being used. Place a drop on a warmed slide for microscopic. examination. (J35.151.w2)
    • In fertile males, at least 70% of the sperm usually show progressive forward motility. (J35.151.w2)
  • A 15 - 20 cm length of tubing, rubber, soft plastic or glass can be used, with an external diameter of 6 - 7 mm. (B614.2.w2)
  • Inseminate when the doe is in oestrus. (B614.2.w2)
    • Receptivity can be determined by the red colouration and turgidity of the lips of the vulva (versus white or pale pink vulval lips in non-receptive does). (J296.68.w1, J296.66.w3)
  • Note: Conception rates with artificial insemination can be similar to those from natural mating if carried out when the female is receptive. (B614.2.w2, J296.66.w3)
    • Conception when the female is not receptive to natural mating results in a lower conception rate. (B614.2.w2, J296.66.w3)
    • Both the colour of the vulva and general behaviour can be used to determine when the doe is receptive. In general, the vulva of a receptive doe is purple, rather than pale pink, the the doe may appear restless. (J35.151.w2)
  • Hold the doe on her back and deposit the sperm suspension deep into the vagina, in the cervical area. (B614.2.w2, J35.151.w2)
  • OR place the doe in a box with a non-slip surface and lift the tail gently. A receptive doe then shows lordosis, lifting her hindquarters, while a non-receptive doe fails to lift her hindquarters, and clamps down her tail. (J35.151.w2)
    • Insert the pipette at an angle of 45 degrees. (J35.151.w2)
Stimulation of ovulation
It is necessary to stimulate the doe to induce ovulation. This can be carried out by:
  • Mating by a vasectomised buck (not always effective). (B614.2.w2, J35.151.w2)
    • Several vasectomised bucks must be kept, since some females will reject some males. (J35.151.w2)
    • Note: vasectomised males can develop anti-sperm antibodies in their seminal plasma, therefore the vasectomised buck should not be allowed to mate the female until 1 - 4 hours after insemination. (J35.151.w2)
    • Mounting and ejaculation by the buck does not always result in ovulation. (J35.151.w2)
  • Injection of the GnRH analogue buserelin (Receptal (Intervet Schering-Plough) - data sheet included) intramuscularly or subcutaneously. (B600.4.w4, B601.15.w15, B603.5.w5, J35.151.w2, J296.68.w1)
    • This method is simple and reliable, causing ovulation about 10 - 12 hours after injetion. It can be used repeatedly. (J35.151.w2)
    • A recent alternative has been addition of a GnRH analogue (buserelin 5 µg or triptorelin 5 µg) to the semen suspension, for mucosal vaginal absorption. (J296.68.w1)
    • In Lepus europaeus - Brown hare, intramuscular injection with buserelin (0.4 mL Receptal, i.e. 0.00168 mg buserelin) during the breeding season resulted in an increase in blood progesterone on the fourth day after injection, and successful pregnancies following artificial insemination with diluted semen (in Tris buffer extender). (J372.X2008.w1)
      • Injection with buserelin outside the breeding season did not cause an increase in plasma progesterone concentration. (J372.X2008.w1)
  • Alternatives which have been used include:
    • Injection with human chorionic gonadotropin (hCG). (B550.16.w16, J35.151.w2)
      • Intravenous injection is required. (B550.16.w16, J35.151.w2)
      • 20 - 25 IU in 0.25 mL sterile water. (B550.16.w16)
      • This is highly antigenic, therefore with repeated use antibodies are produced and further use does not cause ovulation. (J35.151.w2)
      • Injection with human chorionic gonadotropin (100 units intravenously) in Lepus europaeus syriacus Lepus europaeus - Brown hare. (J495.28.w4)
        • This was effective for successful induction of ovulation throughout the year. (J495.28.w4)
    • Injection of 20 - 25 IU luteinising hormone (LH), intravenously or intramuscularly. (B614.2.w2, B550.16.w16)
    • Injection with 20 UI of eCG (Folligon, Intervet) to synchronise oestrus and induction of ovulation using 10 µg of GnRH (Fertagyl, Janssen Animal Health). (J296.66.w3)
    • Injection with FSH twice daily for three days then LH at 50 mg/kg on the fourth day in Sylvilagus floridanus - Eastern cottontail. (J296.4.w1)
  • Note: Feeding of artificially inseminated domestic rabbit does with sprouted wheat improved sexual receptivity and litter size. Sprouted wheat contains high levels of 6-MBOA (6-methoxybenzoxazolinone), a non-oestrogenic phenol which has been shown to stimulate reproductive functions. (J372.99.w1)
Ferret Consideration
  • Ferrets are photoperiod controlled seasonal breeders. Oestrus occurs when daylength increases after a period of short daylength - about three weeks after an abrupt change from eight hours light/16 hours darkness to 16 hours light/eight hours darkness, under artificial lighting conditions. Changing the light conditions periodically can be used to bring the female into oestrus two or three times a year (by changing lighting hours every four months or every two months, respectively), and at whatever time of the year is desired. (J503.43.w1)
  • Ovulation occurs in response to the physical stimuli of mating: copulation leads to release of LH (about 3-4 fold elevation) which leads to ovulation of about 12 oocytes 30-40 hours after copulation. Fertilisation is most effective up to 12 hours after ovulation, therefore at 42-52 hours after copulation. (J503.43.w1)
  • Treatment with Human chorionic gonadotropin (hCG) also induces release of LH and thereby ovulation. (J503.43.w1)
    • The optimum dose of hCG for ovulation is 100 IU per ferret. (J54.7.w2)
Artificial Insemination
  • Artificial insemination has been used successfully for breeding ferrets. It is necessary to induce ovulation, e.g. by injection of 100 ug LH prior to insemination. (P1.1977.w2)
  • Sperm for insemination can be obtained by electroejaculation. (J206.86.w1, J206.92.w1, P1.1977.w2)
    • Sperm has also been obtained from the epididymis of surplus male ferrets immediately following euthanasia. (J54.17.w2, J422.11.w1)
    • Semen can be cryopreserved and used later for fertilisation. (J206.86.w1)
  • Inoculation of semen into the jill's vagina does not result in fertilisation, even if the jill ovulates. (J54.17.w2, J422.11.w1)
  • Artificial insemination can be carried out laparascopically or surgically to deposit semen into the vagina. (J54.17.w2, J206.92.w1, P1.1977.w2)
  • Non-surgical artificial insemination is successful if sperm are deposited transcervically, into the uterine body or anterior horns. (J54.17.w2)
    • Improved results were obtained by insemination 24 hours after injection of hCG, rather than at the same time (79% pregnancies versus 17%). (J54.17.w2)
  • Artificial insemination has been used to improve breeding and improve founder representation of the endangered Mustela nigripes - Black-footed ferret including (via cryopreservation) allowed breeding by males several years after the death of the ferret. (J503.44.w1)
Bonobo Consideration

Assisted reproduction may be required when genetically it is preferred that two animals should breed, but the two individuals show an inhibition against mating. This may happen in bonobos which have lived with each other for prolonged periods and it is possible that this represents a natural barrier to inbreeding. (P1.2002.w10)

Artificial insemination
  • Non-invasive artificial insemination (AI) has been carried out in bonobos with the help of positive reinforcement training. A female was trained to provide urine samples (to enable accurate timing of ovulation) and to permit a speculum to be inserted, allowing insemination deep into the vagina. Training also permitted collection of ejaculate from a male. After the artificial insemination, the female was allowed to mate with males who had been fitted with vas deferens clips to render them infertile. Two months later the female tested pregnant (urinary test). The offspring was proven to be the result of the AI. (P1.2002.w10)
Associated techniques linked from Wildpro

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Control of Reproduction

Every animal born within a zoo ought to have a place within the zoo's overall collection plan. Control of reproduction may be used as one method of population control in a zoo. Factors which should be considered before the decision to use any form of contraception is made include possible negative impacts on behaviour, as well as side-effects which may occur following use of either surgical or pharmacological contraception methods. (J23.38.w3, D271, D273)

Captive breeding programmes for many species have become more efficient and control of breeding is an integral part of responsible zoo animal management. (B445.w9)

There are many circumstances in which it is desirable to prevent or delay reproduction, including:

  • Prevention of overcrowding;
  • Avoiding inbreeding when related animals are maintained in the same enclosure;
  • Genetic management, such as reducing breeding of animals which are already genetically over-represented in a population;
  • Avoiding producing hybrids or sub-specific hybrids;
  • Preventing hybrid animals from breeding.
  • Increase the time between generations.

Various methods may be used for preventing reproduction including: (P76.1990.w1)

Physical separation of males and females. (D271.w2) This may be appropriate for:
  • Species which are solitary except during the breeding season;
  • Social species which may be maintained in all-female and all-male groups;
  • Some social species with a short, well-defined breeding season.

(B429.40.w40)

  • Note: Reproductive failure may occur if animals are prevented from breeding for several years, even when only separation has been used. (B429.28.w28)
Holding single-sex groups of animals at each collection. (D271.App1.w3)

Surgical castration of males, or ovariectomy or ovariohysterectomy of females

  • These procedures require anaesthesia and a surgical operation.
  • These methods are permanent.
  • Sexual behaviour is generally eliminated, which may be advantageous in some situations but may be undesirable in other situations.
  • Social hierarchies may be disrupted. 
  • Desirable secondary sexual characteristics may be lost (e.g. the mane in lions), or seasonal cycles disrupted (e.g. deer antlers remaining in velvet). (B23.40.w18)
  • Note: Viable sperm may remain in the vas deferens after castration; the male should not be considered sterile immediately (six weeks is recommended as a conservative estimate). (B23.40.w18)

(B23.40.w18, B429.40.w40, B445.w9, D271..App1.w3)

Vasectomy in males/tubal ligation in females;
  • These methods require anaesthesia and a surgical operation.
  • Although theoretically reversible, these operations should be considered permanent.
    • The technique required for reversing vasectomy requires highly skilled microsurgery and has been developed only for humans.
  • injection of a sclerosing agent into the cauda epididymis or vas deferens is a possible non-surgical method in males.
  • Hormone production and sexual behaviour typically are unaffected.
  • In females, generally ovariectomy is preferable to tubal ligation on health grounds, although in very long-lived species the possibility of long-term bone density loss due to lack of oestrogen (as in humans) might make tubal ligation preferable.

(B23.40.w18, B429.40.w40, P1.1991.w7, B445.w9, D271.App1.w3)

Occlusion of the vas deferens, either by a material inside the vas deferens to plug it, or by extravasal devices such as clips. (B23.40.w18,, B429.40.w40, B445.w9 P1.1991.w7)
  • These procedures require anaesthesia and a surgical operation.
  • Theoretically reversible, but in practice should be regarded as irreversible since preventing sperm passage without either perforation of the vas or development of fibrosis is not reliable by any method developed to date. (B23.40.w18, B429.40.w40)
  • Fertility may not be restored after removal of a vas deferens plug. (P9.1995.w9)
  • The animal may develop antisperm antibodies, compromising subsequent fertility even if the occlusive device is removed and the vas is patent. (B429.40.w40)
  • Note: Viable sperm may remain in the vas deferens; the male should not be considered sterile immediately (six weeks is recommended as a conservative estimate). (B23.40.w18)
Use of IUDs
  • These may be appropriate for some species, such as the great apes. (B23.40.w18)
Use of steroid hormones in females - synthetic oestrogen plus synthetic progestin, or synthetic progestin alone - orally, by intramuscular injection or in a surgical implant. (B23.40.w18, B429.40.w40, D271.App1.w3, P76.1990.w1)
  • Commercially available contraceptives include: megestrol acetate for oral use (Ovaban®, Schering-Plough; Megace®, Bistol-Meyers Squibb); medroxyprogesterone acetate for oral use (Provera®, Pfizer Inc.) or intramuscular injection (Depo-Provera®, Pfizer Inc.); melengesterol acetate in an implant (ZooPharm); and levonorgesterol implant (Norplant®, Wyeth-Ayerst). (J296.66.w2)
  • These must be given at appropriate intervals depending on the hormone and formulation used: daily (oral drugs); every two to three months (e.g. Depo-Provera (Upjohn) - medroxyprogesterone acetate in a slow-release formulation); replaced at intervals of two years (melenegestrol acetate (MGA) in a silicone matrix); or left for up to five years (Norplant (levonorgestrel)). (B429.40.w40, P76.1990.w1)
    • Implants must be placed in the animal, which requires an anaesthetic. (B429.40.w40)
    • Implants may be lost, particularly in primates which may remove them. (B429.40.w40)
    • It may be difficult to tell whether an implant is still in place or has been lost. (B445.w9)
    • Loss of implants may be reduced by using implants such as Norplant, injected through a trochar (the small entry wound reduces the risk of loss during grooming); by gas sterilisation then thorough degassing and use of sterile instruments for implantation; and by separating the implanted individual from her social group while the implant site heals. (B23.40.w18)
    • With oral contraceptives, it is vital to ensure daily ingestion, and this can be difficult. (B23.40.w18)
  • For seasonal breeders, oral or injectable steroids may be used during the breeding season only. (P76.1990.w1)
  • Note: when a progestin is first given, it may not block ovulation if follicles are already present, therefore the female should still be considered fertile for at least the first two weeks. (B23.40.w18)
  • If given to animals which are pregnant, there may be adverse effects on pregnancy (e.g. abortion in golden lion tamarins), parturition (prevented in contracepted Odocoileus virginianus - White-tailed deer), or the offspring of the implanted animal. (B429.40.w40)
  • If oestrogens are given to a lactating female, lactation may be suppressed. (B429.40.w40)
  • There may be considerable variations in time to ovulation following removal of implants. (B23.40.w18)
  • Prolonged use in felids, canids (Canis lupus - Wolf) and other carnivores has been associated with development of a variety of proliferative diseases (including aggressive tumours) and inflammatory diseases of the reproductive tract. (B23.40.w18, P1.1991.w7, P9.1995.w9)
  • Mibolerone (Cheque, Upjohn Co.) is a synthetic androgen used in domestic female dogs, but may cause increased aggression. (B23.40.w18)
Immunocontraception in males. (B429.40.w40)
  • Male animals can be immunised against LH or GnRH.
  • Spermatogenesis is blocked.
  • This method also interferes with production of testosterone; typical male behaviour is suppressed, therefore this method is functionally similar to castration. 

(B429.40.w40)

Immunocontraception in females, using porcine zona pellucida (PZP) vaccines. (B23.40.w18, B429.40.w40, B445.w9, D271.App1.w3, J296.66.w2, P1.1991.w7, P1.1993.w4)
  • Commercially-available PZP vaccines include SpayVac® (TerraMar Environmental Research Ltd.) and a vaccine available from Zoo Montana. (J296.66.w2)
  • The "standard" dose of PZP vaccine is about 65 microgrammes of antigen (equivalent to about 5000 zonae pellucida)
  • In theory, immunocontraception prevents fertilisation without affecting either ovulation or the expression of oestrous behaviour. In practice there may be wider effects on the ovary, although these may be reduced by increased purification of the antigen. (P1.1991.w7)
  • There may be reactions, sometimes severe, if Freund's adjuvants are used, while other adjuvants may have a lesser immunostimulatory effect therefore result in less successful contraception. (P9.1995.w9)
    • Freund's Complete Adjuvant may lead to tuberculosis-positive test reactions, and produces small abscesses in some animals. (B23.40.w18, B445.w9, P1.1993.w4)
    • Alternative adjuvants which result in adequate, contraceptive, antibody titres include Freund's incomplete adjuvant (FIA = Quil-A, Superfos Specialty Chemicals, Vedback, Denmark), Carbopol (Goodrich Co., Cleveland, Ohio, USA) and DEAE-dextran (Sigma Chemical Co., St Louis, MO, USA). (P1.1993.w4)
  • The antigen/adjuvant emulsion has high viscosity, which may cause problems for delivery by dart, although delivery by this method can be used. (P1.1993.w4)
  • Usually three injections are given, at intervals of about a month. (B23.40.w18)
  • The female should not be considered protected from contraception until after administration of the last booster vaccination. (B23.40.w18)
  • Immunocontraception may last for only 6-12 months, which is adequate for seasonal breeders with a breeding season of under six months. (B23.40.w18)
    • However, some seasonal breeders, if not pregnant (due to contraception), may continue to cycle for longer periods than is usual. (B23.40.w18)
  • PZP immunocontraception may fail because: (J54.24.w3)
    • The vaccine fails to elicit sufficient antibody titres; this is most likely to occur due to relative homology between the PZP and the zona proteins of the target animal; (J54.24.w3)
    • The antibodies produced in response to the PZP fail to cross-react with the sperm receptor proteins on the zona pellucida of the target animal;
    • The antibodies formed do not remain at sufficiently high levels over the breeding season. (J54.24.w3)
  • Long term use of PZP for contraception may be irreversible. (B23.40.w18)
  • Note: PZP vaccines should not be given to pregnant females until more data is available; early abortion and premature births have been recorded in treated individuals. (B23.40.w18)
Hormone agonists or antagonists in males
  • Hormone agonists act by negative feedback. Because pituitary hormones respond to pulsatile rather than chronic GnRH stimulation, continuous administration of a GnRH agonist, after an initial stimulatory phase, causes suppression of production of LH and FSH.
  • In males, suppression of LH and FSH results in suppression of spermatogenesis.
  • These drugs should function as "equivalent to reversible gonadectomy." (B23.40.w18)
  • Because testosterone production is suppressed, typical male behaviours and secondary sexual characteristics are suppressed.
    • In some species, reduced aggression of the male may be desirable.
    • If maintenance of male-typical behaviour and/or characteristics is desirable, the GnRH agonist may be given together with testosterone.
  • Leuprolide (Leupron, TAP) is a GnRH agonist available in injectable form which has had some success in males of various species. (B23.40.w18)
  • Use of GnRH agonists may become more applicable if subcutaneous implants are developed to allow sustained release of the GnRH-analogue peptide hormones.
  • Use of antagonists (which block hormone actions by binding the bind hormone receptors) would be practical only if they became sufficiently inexpensive for chronic use.
  • Use of the GnRH analogue deslorelin in wild carnivores has had variable results. (J370.57.w2)

(B23.40.w18, B429.40.w40, J370.57.w2)

  • Commercially available GnRH analogs include deslorelin (Suprelorin®, Peptech Animal Health), and leuprolide acetate (Lupron Depot®, TAP Pharmaceuticals; and a formulation from ZooPharm). These are given intramuscularly. (J296.66.w2)
Antispermatogenics in males - bisdiamine.
  • Antispermatogenics inhibit spermatogenesis in a reversible manner and do not affect hormone levels.
  • The main side-effect in humans (inhibition of alcohol dehydrogenase) is not a problem in animals (since they do not usually consume alcohol). (P1.1991.w7)
  • In wolves (Canis lupus - Wolf), a bisdiamine (WIN 18,446, Stirling Winthrop, Rensselaer, New York, USA) was found to effectively and reversibly block spermatogenesis in males when given orally at a dose of at least 190 g/kg (in meat) prior to the beginning of spermatogenesis and continuing through the breeding season. It was not effective in wolves treated after the start of the breeding season at a dose of 250 mg/kg. (P9.1995.w8)
  • Bisdiamine is commercially available only in a laboratory-grade, very expensive formulation (Fertilysin, Aldrich). (B23.40.w18)
    • Bisdiamine WIN 18,446 is commercially available as Fertilysin® (SAF Bulk Chemicals, Milwaukee, Illinois, USA). (J296.66.w2)
  • Bisdiamine is teratogenic to developing embryos; it is important, during administration of the drug, to separate males from females which might be pregnant. (B23.40.w18)
  • Indenopyridine analogues have been successfully used in rodents, but in cats they caused unacceptable side effects (e.g. lethargy, occasionally bloody diarrhoea). (B23.40.w18)
  • Busulfan suppresses spermatogenesis but also bone marrow, therefore is unsuitable. (B23.40.w18)
Disadvantages of reproductive control

All methods of reproduction control have disadvantages. 

  • One obvious potential problem is failure of the chosen method. With implanted contraceptives, this may be caused by failure of the contraceptive, or by loss of the implant from the animal. (P76.1990.w1)
  • A potential problem when any form of contraception has been used on a temporary basis is failure of the animal to return to normal reproductions after the period of contraception. (B23.40.w18, P76.1990.w1) This may occur even when the only form of contraception used is physical separation. (B429.28.w28)
  • Effective breeding control may result in an aged population with reduced fecundity and reduced breeding potential if the population suffers from a catastrophe such as severe disease. (B429.28.w28)
Reproductive control of free-ranging wild animals
  • There has been considerable research into the possibilities of contraceptive products useable in free-living wild animals, and in particular the development of immunocontraceptive vaccines. (P69.22.w1)
  • It is probable that the use of wildlife contraceptives will be limited; they may be useful in urban/suburban areas when traditional methods such as trapping and hunting cannot be used. (P69.22.w1)
  • In general, except with very short-lived species, contraception would not cause rapid reductions in populations. (P69.22.w1)
  • The following criteria would be required for contraception to be useful for control of pest populations: (P115.1993.w1)
    • Existence of appropriate technology to reduce fertility;
    • Development of an effective delivery mechanism for wild animals;
    • Use of the contraception results in the required goal of reduced damage due to animals;
    • The effects are humane and non-toxic;
    • The product being target-specific, cost effective and environmentally acceptable. 

    (P115.1993.w1)

  • The American Association of Wildlife Veterinarians has suggested (1993 resolution) the following criteria to be met in order for fertility control to be an acceptable method of regulating populations of free-ranging wild animals: (D367)
    1. "The compound doe not affect the health of target species and humans.
    2. A risk assessment is completed delineating potential effects on nontarget species.
    3. The application is limited to site-specific, well-defined subpopulations or populations.
    4. The application does not alter the gene pool of the species.
    5. Sort- and long-term effects on population dynamics, including age structure and behaviorral effects, are evaluated through modeling and monitoring.
    6. The program is evaluated by regulatory and wildlife management agencies before use, with full public participation.
    7. Costs of the fertility control program are borne by the organizations or public that benefit from the program." 

    (D367)

Bear Consideration Wildlife Information Network again thanks WSPA for its invaluable sponsorship without which the Wildpro "Bears: Health and Management" volume would not have been researched and published. A message from WSPA follows:
LOGO-WSPA OrangeBlack.jpg (31514 bytes) WSPA does not condone the captive breeding of bears unless there is a programme to rehabilitate and release the cubs back to the wild as part of a valid conservation or animal welfare programme.

WSPA believes that bears should not be kept in captivity in enclosures (including zoo enclosures) where their living environment lacks sufficient space and natural habitat to enable the bears to exhibit their natural behaviour.

Note: Collections holding animals within a breeding programme (e.g. EEP animals in Europe) should consult with the species coordinator and follow their recommendations regarding control of reproduction. (D247.6.w6)

Reproduction in bears may be controlled by a variety of methods, each with advantages and disadvantages.

  • Note: "The limited data available for pubertal animals suggest that contraception probably does not affect future fertility, but this is not certain." (D315)
Physical separation
  • Separating males and females during the breeding season may be considered as a method of reproductive control for northern bear species with a defined breeding season. (D247.6.w6)
  • Separation may be required for as long as five months. (D247.6.w6)
  • Separated individuals should be kept in enclosures well away from one another; if they remain within range of olfactory communication this could act as a stressor and promote stereotypic behaviours. (D247.6.w6)
Hormonal contraception
  • Hormonal implants have been used in various bear species including Helarctos malayanus - Sun bear, Ursus americanus - American black bear, Melursus ursinus - Sloth bear, Ursus arctos - Brown bear and Ursus maritimus - Polar bear. (P76.1990.w1)
  • Implants of melengestrol acetate (MGA) have been used in bears. (B445.w9)
  • Implants containing 500 mg melengestrol are effective for at least two years. (D247.6.w6)
    • The bear has to be immobilised (anaesthetised) and the implant placed surgically. (D247.6.w6)
    • An Ursus arctos - Brown bear conceived 87 months after implantation of an MGA implant. (P1.1991.w)
  • Note: After use of an implant, the bear should then be given at least one season without contraception, to reduce the risk of adverse health effects. (D247.6.w6)
  • Oral contraceptives can be used. (D247.6.w6)
    • Note: Care is required to ensure that the dose is swallowed and has not been spat out. (D247.6.w6)
    • In a female Ursus maritimus - Polar bear at Cologne Zoo, 400 mg medroxyprogesteronacetate (two pills of Clinovir 200, Upjohn) were given weekly, crushed and mixed with honey which was then presented to the bear on the end of a spoon. (D247.6.w6)
      • No side effects were noted with this contraception. (D247.6.w6)
    • In a female Kodiak bear (Ursus arctos - Brown bear), oral megestrol acetate was given at 200 mg daily (0.25 mg/lb bodyweight) for 64 days, in fish, starting 1st April - about two weeks before oestrus was expected, and appeared effective: the female, which had cubbed for the previous seven years, did not produce cubs. (P1.1978.w1)
  • N.B. Long-term progestin treatment of felids (Felidae - Cats (Family)) has been associated with the development of uterine and mammary pathology, including neoplasia. (J296.66.w2, D315). Additionally, progestins may enhance the onset of signs of diabetes mellitus. Therefore synthetic progestins, with or without oestrogen, are not recommended for long-term use in carnivores. (D315)
  • Progestins should not be used in pregnant females as they may inhibit uterine contractions and therefore interfere with the normal progression of parturition. (D315)
  • Use of progestins during the period of embryonic diapause could stimulate the onset of active gestation, resulting in an earlier than normal birth date. (D315)
  • Progestins may or may not interfere with the development of normal oestrous behaviour. (D315)
Immunocontraception using Porcine Zonae Pellucidae (PZP)
  • This involves two vaccinations in the first year, one at one to two months before the start of the breeding season and the second one to two weeks before the breeding season. For seasonally breeding species, a single vaccination is then required yearly, four weeks before the breeding season. In Helarctos malayanus - Sun bear, which does not have a defined breeding season, a booster would be required every eight months. (D247.6.w6)
  • The vaccine can be given by darting (remote injection). However, until more is known about the effects, it is preferable to immobilise the bear so that a blood sample can be taken to check antibody titres. (D247.6.w6)
  • The vaccination should not affect the reproductive behaviour of the bears. (D247.6.w6)
  • This has been used in Ursus arctos - Brown bear and Ursus thibetanus - Asiatic black bear. (No data was available on its efficacy in these species). (P1.1993.w4)
  • In Helarctos malayanus - Sun bear, two females were treated with PZP. In one, treatment was followed by absence of ovarian activity; a normal oestrus cycle occurred four years after PZP treatment and when the bear died one year after the end of the endocrine study the ovaries were found to be grossly and microscopically normal. In the other treated bear, persistent follicular and luteal activity occurred. However, similar activity/lack of activity were also seen in other individuals not treated with PZP. (J296.62.w1, P6.2.w5)
  • PZP contraception has been used in 25 bears with 100% effectiveness (no pregnancies); boosters are recommended at 12-month intervals. (J54.24.w3)
  • More information is required:
    • At present this should not be used in bears which may be needed to reproduce later, since it is not known whether the effects are reversible. (D247.6.w6)
    • At necropsy, ovaries of both treated and untreated bears should be collected, preserved in 4% formalin (make small incisions to increase formalin penetration into the tissue) and sent to the Institute for Zoo Biology and Wildlife Research (IZW), Berlin, Germany. (D247.6.w6, D247.7.w7) See: Necropsy of Mammals
  • Note: use of a PZP vaccine is not recommended in carnivores since it has been associated with damage, potentially permanent, to ovarian tissue, which may result in infertility. (D315)
Castration, ovariectomy/ovariohysterectomy and sterilization (vasectomy/tubal ligation)
  • These methods are surgical and require anaesthesia. (D247.6.w6)
  • These methods are irreversible. (D247.6.w6)
  • Castration affects reproductive behaviour, and castrated males tend to gain weight. (D247.6.w6)
    • Some castrated males continue to show breeding behaviour. (D315)
  • Vasectomy is not recommended because vasectomised males are likely to continue normal sexual behaviour and copulation with females, which can induce ovulation followed by pseudopregnancy. (D247.6.w6) During pseudopregnancy, high endogenous progesterone levels develop (J467.5.w1); repeated exposure to endogenous progesterone not associated with pregnancy may carry risks similar to those recognised with repeated exposure to exogenous progestins (see above). (D315)
  • Castration or ovariohysterectomy are preferred for permanent sterilisation. (D315)
Antiprogestin to prevent implantation
  • Antiprogestins competitively displace progesterone at the cellular receptor and thereby prevent implantation or induce abortion. (J296.56.w1, J370.57.w1, P1.2002.w2)
  • Because bear reproduction involves a period of embryonic diapause prior to implantation, and progesterone levels increase at the end of the embryonic diapause, displacement of progesterone using an antigestagen (AG) has been considered as a means of preventing the blastocyst from implanting. (P5.39.w9) In seasonally breeding bears, antiprogestins can be given parenterally during diapause (October) or in the early post-implantation period. (P1.2002.w2)
  • Note: If the antiprogestin is given too late after implantation (e.g. one month after implantation) there may be incomplete resorption of embryos, or fetal abortion. It is therefore important to give the treatment in the late diapause/early implantation period. (J370.57.w1, P1.2002.w2)
    • J956 intramuscularly at 10 mg/kg together with an oestrogen (ethinyloestrodiol at 10 µg/kg) given after implantation (mid-November or early December), resulted in abortion, but in the two bears (Ursus arctos - Brown bear) treated in December (late pregnancy), abortion was incomplete. Also, two of the bears treated post-implantation developed serosanguinous vaginal discharge and discomfort. (J370.57.w1)
  • Antiprogestins given orally were ineffective. (J370.57.w1, P1.2002.w2)
    • Oral (four doses each 10 mg/kg) or low-dose (1 mg) intramuscular administration of antiprogestin J956 did not prevent implantation and pregnancy. (J296.56.w1)
  • Antiprogestin J956 can be delivered intramuscularly by remote injection (darting). (J296.56.w1)
  • At a dose rate of 10 mg/kg by intramuscular injection, plasma levels of antiprogestin J956 were sustained (6.4 +/- 1.3 ng/mL) for nearly two months in Ursus americanus - American black bear. (J296.56.w1)
  • Full information on fertility following antiprogestin treatment has not yet been gathered. However, one bear treated with J956 and ethinyloestadiol became pregnant a year after treatment. (J370.57.w1)
GnRH agonist in males
  • An initial trial in a bear (Tremarctos ornatus - Spectacled bear) indicated that this may be a useful method for contraception in bears in the future. (P36.1994.w5)
    • A GnRH agonist, leuprolide acetate (Leupron® Depot, TAP Pharmaceutical, Chicago, Illinois, USA), injected subcutaneously at 0.075 mg/kg/28 days in a male Tremarctos ornatus - Spectacled bear produced a reduction in testosterone levels in blood from 1.75 ng/mL to less than 0.156 ng/mL, i.e. a 91% reduction. Within eight weeks of cessation of treatment, levels returned to pre-treatment values. The male was held adjacent to a female bear. (P36.1994.w5)
  • If a GnRH agonist is used in seasonally-breeding bears, it needs to be implanted about two months before the onset of the breeding season, certainly at least six weeks before, so that any sperm produced have been eliminated from the male before the female will ovulate. (D315)
  • This treatment may reduce male aggression. (D315)
GnRH agonist in females
  • This is particularly suitable for use in seasonal breeders. Because of the initial stimulatory effect, the agonist needs to be implanted about two months before the earliest possible onset of the breeding season. Initial ovulation may be prevented by treating with a progestin short term (e.g. oral contraceptives for about ten days) at the time of the insertion of the GnRH agonist implant. (D315)
  • Oestrous behaviour should be eliminated by use of a GnRH agonist. (D315)
  • Note: 
    • GnRH agonists may cause abortion if used in pregnant females. (D315)
    • The effects of GnRH agonist use on lactation have not been properly tested. (D315)
Lagomorph Consideration
Physical separation
  • Physical separation can be used to prevent breeding in lagomorphs. (B600.3.w3, B601.1.w1, P113.2005.w6)
Castration, ovariectomy/ovariohysterectomy and sterilization (vasectomy/tubal ligation)
  • Castration is effective at preventing breeding. Note: following castration, the buck may remain fertile for at least four weeks and sometimes longer than this; the buck should be kept apart from any does until six weeks after castration. (B600.3.w3, B601.1.w1, P113.2005.w6)
  • Ovariohysterectomy is effective at preventing breeding. In pet rabbits this is highly recommended because it also prevents development of conditions such as Endometrial Adenocarcinoma and other Uterine Neoplasia in Lagomorphs
  • Vasectomy can be carried out in rabbits. Vasectomised males are used sometimes to induce ovulation in female rabbits being artificially inseminated. (B614.2.w2, J35.151.w2)
  • Tubal ligation can be carried out in rabbits. Experimentally, surgical sterilisation, by tubal ligation, of 60 - 80% of wild Oryctolagus cuniculus - European rabbit populations in Australia, was shown to reduce seasonal peaks of rabbit abundance, but did not reduce the average adult population size. Sterilised females showed a much better survival than unsterilised females (demonstrating the high costs associated with reproduction), while unsterilised adults (males and females) and immature rabbits also showed higher survival in the sterilised population than in unsterilised populations. There was a density-dependent increase in juvenile survival. (J182.37.w1, J182.44.w1)
Immunocontraception using Zonae Pellucidae (ZP) Proteins
  • Immunization with intact or heat-solubilized porcine zonae pellucidae together with various adjuvants has been shown to be immunogenic, inducing a strong autoimmune response, and to cause infertility, with failure to ovulate even after treatment with hCG, and with development of ovarian pathology. (J371.25.w1, J371.74.w1, J531.21.w1)
  • Immunization with whole rabbit zonae pellucidae failed to induce infertility, although antibodies to zonae were found in the serum. (J371.25.w1)
  • Recombinant rabbit zona pellucida glycoprotein B (ZPB), expressed in eukaryotic cells in vitro was found to be an effective antigen when injected with Freunds complete adjuvant. With the use of two booster doses, sustained infertility was produced in 70% of female rabbits; this required production of high antibody titres (at least 12,800). In male, but not female, rabbits, high antibody titres developed after a single immunization. When ZPB was expressed in a recombinant myxoma virus, serum antibody levels were similar in male and female rabbits, but did not reach the 12,800 level; 25% of females were infertile. Booster injections of recombinant ZPB protein resulted in a higher antibody response and 80% infertility of female rabbits. The primary mechanisms of infertility appeared to be degeneration of oocytes and follicles. (J371.61.w2)
  • Immunization with recombinant myxoma virus expressing rabbit zona pellucida 3 (rZP3) glycoproteins resulted in production of serum antibodies, binding of antibodies to zona pellucida from day 10, transient inflammatory response in the ovaries, and infertility in 70% of rabbits when first bred. Fertility later returned, as indicated by pregnancy in 92% of rabbits at later matings (return to fertility within two months). (J371.74.w1)
  • Infection of rabbits with a recombinant myxoma virus expressing rabbit zona pellucida protein B elicited a specific IgG response to ZPB with the antibody binding strongly to the zonae pellucidae of oocysts in both secondary and tertiary follicles. There were significantly reduced preovulatory and tertiary follicles in the ovaries at 30 days post infection. Histologically, in the medulla of the ovary there were no follicles but a large number of probably luteinized cells, indicating severe ovary pathology. (J20.318.w1)
  • "In more recent experiments, recombinant myxoma viruses constructed with rZPC under the control of a synthetic combined early/late promoter induced infertility in 90-100% of rabbits mated 35 days after immunization and in 50% of rabbits at subsequent matings (Kerr, Perkins and van Leeuwen, unpublished results)." (J532.71.w1)
Ferret Consideration
Castration, Ovariohysterectomy and Vasectomy
Breeding in ferrets can be prevented by surgical neutering: castration of males, ovariohysterectomy (spaying) of females, or by vasectomy of males. 
  • Note: There is an increased risk of the development of adrenal gland disease (Adrenocortical Neoplasia in Ferrets) in early-neutered ferrets. It is recommended that ferrets (male or female) be neutered only at puberty (6-8 months of age), not sooner and should be implanted with a GnRH implant until fully mature. (W755.Sept11.w1)
  • A study in the Netherlands found a significant correlation between age of neutering and age of onset of adrenal disease. Within the studied 1,274 ferrets, seven (0.55%) developed hyperadrenocorticism confirmed by histological examination of excised adrenal glands while a further seven were suspected cases based on clinical signs, but the owners did not allow further studies to confirm the diagnosis. (J4.216.w2)
  • Hyperadrenocorticism has been seen in entire (unneutered) ferrets, but only rarely. (J4.216.w2)
  • Vasectomy of males prevents fertilisation but does not affect behaviour. 
Preventing reproduction in entire females

Entire (fertile) jills which are not mated remain in oestrus throughout the breeding season (until daylength decreases) and develop anaemia due to bone marrow suppression caused by Hyperoestrogenism in Ferrets. (B627.8.w8, W755.Sept11.w1) If breeding is not required, it is important to end oestrus, not simply prevent pregnancy. Options include:

  • Mating with a vasectomised male (hoblet). (W755.Sept11.w1)
    • Ferrets are induced ovulators; the physical stimulus of mating results in ovulation. (J503.43.w1)
    • It is recommended that each ferret owner keep their own hoblet, rather than sharing a vasectomised male with others, in order to reduce the risks of disease transmission. (W755.Sept11.w1)
  • Proligestone (the "jill jab")
    • Proligestone (0.5 mL Delvosterone, Intervet, subcutaneously) will elicit ovulation. (B232.9.w9)
      • This should be given in late March [northern hemisphere]. (J3.126.w7)
      • Signs of oestrus start to reduce after three to four days. (J3.126.w7)
      • If the initial 50 mg injection (0.5 mL of 100 mg/mL drug per jill, subcutaneously) has no effect, give 25 mg seven days later, (B626.App.w22, B631.21.w21) and again 25 g seven days thereafter if necessary (should not be needed). (B626.App.w22)
      • Some jills return to oestrus later in the summer and require a repeat injection. (J3.126.w7)
        • Following injection with 40 mg proligestone in mid-February [northern hemisphere], ferrets returned to oestrus in 99 +/-40 days (in April to July). Treatment later in the year (e.g. late March) may prevent return to oestrus for the whole year if ferrets are kept under natural light conditions. (J3.166.w1)
        • Ferrets mated after returning to oestrus became pregnant (3/4) or pseudopregnant (1/4). (J3.166.w1)
    • Very occasionally a jill may return to oestrus a third or even a fourth time in the same year. (J3.126.w7)
    • Hormone-related alopecia develops in some treated ferrets. (J3.166.w1)
    • Jills treated with proligestone can be bred as normal in following years. (J3.126.w7)
    • Note: Jills accidentally mated in May and June after being treated with proligestone produced small litters (three and four kits); a jill accidentally mated in July developed pseudopregnancy. (J3.126.w7)
  • Medroxyprogesterone acetate (MPA)
    • Following injection with 15 mg MPA in mid-February [northern hemisphere], ferrets returned to oestrus in 94 +/-18 days (in May to June). (J3.166.w1)
    • Ferrets mated after returning to oestrus became pregnant (3/4) or pseudopregnant (1/4). (J3.166.w1)
    • Hormone-related alopecia develops in some treated ferrets. (J3.166.w1)
  • Human chorionic gonadotropin (hCG) (Chorulon, Intervet), 20 - 100 units intramuscularly will induce ovulation. (B232.9.w9)
    • Human chorionic gonadotropin hCG 100 IU can be given intramuscularly after the 10th day of the vulva being swollen and will induce ovulation. (B627.8.w8)
    • Following injection with hCG in mid-February [northern hemisphere], ferrets returned to oestrus after 53 days +/- 9 days i.e. May to June. (J3.166.w1)
      • Ferrets mated after returning to oestrus became pregnant (3/4) or pseudopregnant (1/4). (J3.166.w1)
  • Gonadotropin releasing hormone (GnRH) 20 µg can be given intramuscularly after the 10th day of the vulva being swollen and will induce ovulation. (B627.8.w8)
  • Deslorelin acetate (gonadotropin-releasing hormone agonist) in a slow-release formulation. 
    • Following implantation of deslorelin acetate in a slow-release formulation in mid-February [northern hemisphere], jills showed intensive signs of oestrus within four days, but signs stopped within two weeks. They did not return to oestrus that year; first oestrus occurred after 698 +/- 122 days (July of the year after implantation to May of the year after that). (J3.166.w1)
    • A new implant is needed about every 18 months or when the jill shows signs of sexual activity. (W755.Sept11.w1)
Preventing reproduction in entire males
Vasectomy prevents males from fertilising females, but does not reduce odour, nor does it change behaviours such as aggression towards other males.
  • Deslorelin acetate (gonadotrophin-releasing hormone agonist) in a slow-release formulation may be given to males as a form of "chemical castration". (J296.70.w1) Experimentally, males with such an implant (9.4 mg deslorelin per ferret) were shown to have reduced plasma FSH and testosterone concentrations, reduced testis size, and reduced musky odour (lower odour even than in surgically castrated males). Spermatogenesis was suppressed (no germ cells present). This appeared to be a useful alternative to surgical castration. (J296.70.w1)
    • Treatment of entire male ferrets with deslorelin (GnRH agonist) implant greatly reduces sexual behaviours. Treated ferrets also showed reduced intermale aggressive behaviour (more so than surgically castrated ferrets), and increased play behaviours. (J288.115.w1)
    • If two entire (uncastrated) males are to be kept together, then implanting both deslorelin is advisable to reduce breeding season aggression and territoriality. (W755.Sept11.w1)
Bonobo Considerations Given the small size of the captive bonobo population, and the associated potential need for breeding by any bonobo, contraception should be used only as a last resort; separation of individuals by moving one to another group is preferable if particular pairings need to be avoided. Any method of contraception used should ideally be reversible. (D386.5.w3e)
  • In general, methods used in humans should be considered. It is likely that contraception methods usedd in Pan troglodytes would be appropriate for use in bonobos. (D386.5.w3e)
Males 
  • At present, there are no reliable methods available for reversible contraception of male bonobos. (D386.5.w3e)
  • Vasectomy can be used for male contraception, but is not reliably reversible and therefore this is not recommended. (D386.5.w3e)
  • Ligation of the vas deferens by use of surgical clips would be a possible method and preferable to vasectomy. Risks include that this is a surgical procedure, requiring anaesthesia; the risk of infection; and the possibility of spontaneous reversal by recanalisation of the vas deferens, such that frequent follow-up analysis of ejaculate obtained by electrostimulation or via an artificial vagina is required. (D386.5.w3e)
    • This method has been used in bonobos. (P1.2002.w10)
Females
  • Melengestrol acetate (MGA) implants have been used in Pan troglodytes - Chimpanzee and in Pongo pygmaeus - Orang-utan, generally with a high success rate although with some failures in Pongo pygmaeus - Orang-utan due to imlant loss associated with allogrooming. Tagging the implant with a microchip transponder would allow earlier detection of implant loss. Unwanted effects include definite weight gain and a possible link with development of diabetes. (D386.5.w3e)
  • Levonorgestrel implant (Norplant) has been used successfully in Pan troglodytes - Chimpanzee and in Pongo pygmaeus - Orang-utan. If placed deeply and with complete sterility, retention is good. Reintroduction of the animal to the group should be delayed until the site has completely healed, to minimise the risk of implant loss during allogrooming. This method is more likely to fail in obese individuals. There is an unknown risk of long-term impairment of fertility, but this method could be used. (D386.5.w3e)
  • Medroxyprogesterone acetate (DepoProvera) has been used in Pan troglodytes - Chimpanzee successfully. It is administered by intramuscular injection, either by dart or with the animal anaesthetised. It is important that the whole dose is given. The effects of long-term use are unknown and this would be recommended only for short term (less than six months) use. (D386.5.w3e)
  • Birth control pills have been used in great apes, but there is always a risk of incomplete compliance with taking the pills, therefore their routine use is not recommended. (D386.5.w3e)
  • Intrauterine devices (IUD) have been used in Pan troglodytes - Chimpanzee; the smallest human size (suitable for a nulliparous woman) should be fitted. To reduce the risk of the female removing the device, the removal strand can be shortened; obviously this makes removing it, when this is wanted, more difficult. A failure rate of about 5% should be expected and there is a risk of complications if the individual becomes pregnant while the device is in place. (D386.5.w3e)
  • Low-dose oestrogen oral contraceptive pills have been used in bonobos; problems of breakthrough bleeding have been reported. (P20.2010.w1)
Associated techniques linked from Wildpro

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Authors & Referees

Authors Dr Debra Bourne MA VetMB PhD MRCVS (V.w5)
Referees Liz Carter BSc MSC (V.w144); Neil Dorman (V.w104); Mike Jordan (V.w30); Chris Lasher (V.w110)

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