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JoGayle Howard, Theriogenologist, Co- Investigator, Smithsonian Conservation Biology
Institute, National Zoological Park, Washington, DC.
Rebecca Hobbs, Post-doctoral Fellow, Co-investigator, Smithsonian Conservation Biology
Institute, National Zoological Park, Washington, DC.
The North American clouded leopard population managed by the Species Survival Plan is not a genetically healthy,
self-sustaining population. The major problem affecting captive breeding of clouded leopards is the high incidence
of male aggression. This has severely limited the number of naturally breeding pairs, and consequently, most animals
are housed individually. It is integral that all animals within the SSP contribute to the population ensuring greater
genetic diversity.
The Smithsonian Conservation Biology Institute at the National Zoo has studied the reproductive biology of clouded leopards
for over 20 years. We have used this information to develop ovarian control and assisted reproductive techniques to enhance
reproduction in this species. Unfortunately, attempts at artificial insemination (AI; placing sperm directly into the uterus)
have not been successful in clouded leopards as they have in other cat species.
AI failure may be due to multiple factors, including: changes in oocyte (egg) quality, reduced sperm function, poor
fertilization rates, compromised embryo development, corpora lutea (ovulation site) insufficiency, or uterine abnormalities.
To analyze these problems, we are developing in vitro fertilization (IVF) for clouded leopards. IVF is a tool that allows direct
evaluation of oocyte and embryo quality through observational assessment of fertilization, embryo development and molecular
analysis of mRNA (the message read from DNA and translated into protein) in follicle cells. IVF involves hormonal stimulation to
induce ovarian activity and oocyte
maturation, collection of oocytes by laparoscopy and placement of these oocytes in culture
medium with sperm for fertilization in vitro. Not only can IVF be used to diagnose problems with AI, there also is potential
to cryopreserve and store the IVF embryos for future use.
The primary objectives are to:
a) characterize clouded leopard oocyte fertilization in vitro;
b) assess the quality of oocytes following the AI hormone stimulation protocol; and
c) assess the quality of embryos generated from young (under 8 years) versus aged (over 9 years old) clouded leopards.
The overall goal of this project is to develop an IVF protocol for clouded leopards to identify any potential problems with
oocyte quality leading to AI failure and ultimately to improve reproductive and genetic health in the North American Clouded
Leopard SSP program.
Investigators: Heather DeCaluwe, PhD Candidate in Animal and Avian Sciences at the University
of Maryland, College Park, MD and Smithsonian Conservation Biology Institute, National
Zoological Park, Washington, DC; JoGayle Howard, DVM, PhD, Smithsonian Conservation
Biology Institute, National Zoological Park, Washington, DC
Clouded leopards are a striking and elusive wild cat species whose secretive
nature has made it difficult to gather information on population statistics and behavior in
nature. However, it is clear that the clouded leopard is in decline throughout its historic
range in Southeast Asia due to habitat loss and poaching. The captive population of
clouded leopards serves as a hedge against extinction, but is one of the most challenging
felid species to breed in captivity. In the current North American SSP population of ~ 65
animals, there have been only 2 successful breeding pairs in the past 6 years. Although
many animals are still of reproductive age, the scarcity of successful breeding pairs
is primarily due to male aggression which can be so extreme that attacks sometimes
result in death. For this reason, zoos have become extremely cautious about initiating
introductions, and males with a history of violent behavior are especially challenging for
pairing.
In other species, male aggression has been linked both to circulating testosterone
and to personality. In many cases, pharmaceuticals have been instrumental in reducing
aggression. Particularly effective have been hormonal therapies that reduce testosterone,
such as gonadotropin releasing hormone (GnRH) agonists. Psychotropic drugs that target
stress-associated aggression through changes in serotonin metabolism in the brain also
have been effective. Through the administration of deslorelin (a GnRH agonist) and
clomipramine (a psychotropic drug), a study has been initiated to understand and mitigate
aggression in captive male clouded leopards. Behavioral, endocrine, and reproductive
data are being collected and analyzed to find the most effective treatment for decreasing
male aggression. Ultimately, this information may be useful in forming compatible male-
female pairs with the assistance of drug therapy.
Investigator: Jilian Fazio, Principal Investigator, Master’s Student in Environmental Science
and Public Policy at George Mason University and Animal Keeper, Smithsonian’s National Zoo.
 
There are many challenges to the captive management of the clouded leopard, however the issue of
highest concern is male aggression during introduction to a female for breeding. According to the
Clouded Leopard Species Survival Plan, mate incompatibility is the most pressing challenge facing this
species survival (Fletchall 2007). There is an urgent need to pinpoint behavioral differences among
individuals that are reproductively successful.
For this study, the investigator will conduct a complete behavioral assessment of clouded leopards to
determine if certain behavioral responses may act as indicators of reproductive success. It is hypothesized
that there will be quantifiable behavioral differences between clouded leopards that can be used to
categorize different temperaments. This study will utilize three separate methods to determine temperament
in each individual, including behavioral observations, a keeper temperament assessment, and non-invasive
fecal hormone analysis. The results of this research will also establish if certain behavioral responses
can act as indicators of breeding success in clouded leopards. The results of this study can be used to
avoid unnecessary animal transfer, injury and possible death. Such findings would assist in future
management and conservation of the species both in North America and in range countries. The successful
pairing of the limited captive population is integral to the survival of the clouded leopard and behavioral
research is a powerful tool in achieving this goal.
Investigator: Dr. Nadja Wielebnowski of the Brookfield Zoo and Dr. Janine Brown of
the National Zoological Park's Conservation and Research Center (CRC)
In the last few years, clouded leopards in zoos have been studied in an attempt
to help zoologists better manage them by mitigating behavioral problems and enhancing
reproduction. The following is a summary of this research.
Since 1998, Dr. Nadja Wielebnowski of the Brookfield Zoo and Dr. Janine Brown of
the National Zoological Park's Conservation and Research Center (CRC) have been
studying the link between clouded leopard behavior and their production of stress
hormones (i.e. glucocorticoids: cortisol and corticosterone) in order to identify
factors that lead to problems such as fur-plucking, tail-biting, and excessive pacing
and hiding that are often demonstrated by clouded leopards in zoos. Many of these
aberrant behaviors are thought to be the result of high levels of stress faced by
clouded leopards living in captivity. Finding ways to alleviate this stress is a
top priority for zoos housing clouded leopards as well as the Clouded Leopard Species
Survival Plan management team.
Their study consisted of monitoring 74 clouded leopards in North American zoos (65%
of the population) to gather data on their hormone production. In order to evaluate
the level of hormones produced by the cats, the keepers collected fecal samples
every other day for six weeks and sent them to the researchers who analyzed them
for the hormones in question.
At the same time, the keepers made behavioral assessments evaluating the cats' temperaments,
indicating if the cats appeared "stressed" or "calm" and if they exhibited any abnormal
behavior. The keepers also noted if there were any changes in the cats' husbandry
routines or o
The results indicated that clouded leopards evaluated by keepers as being "stressed"
did produce higher levels of stress hormones than those evaluated as being "calm."
Additionally, stress hormone levels were found to be higher in cats exhibiting behaviors
such as self-mutilation and excessive hiding. Furthermore, when the researchers
analyzed a variety of husbandry factors, such as exhibit height, their ability to
see other large cats (i.e. potential predators), and the amount of keeper time spent
around the animals, they were found to be linked to the levels of stress hormones
produced. For example, high concentrations of stress hormones were measured when
potential predators were visible and when cats were on public display. Lower levels
of stress hormones were measured when cats had enclosures with high climbing space
and also when keepers were able to spend more time with them.
These results indicate that zoo staff may be able to control some of the factors
that lead to stress in clouded leopards to more effectively manage this species.
In order to identify the best ways to improve clouded leopard management, the researchers
are now undertaking a new study to vary several of the husbandry factors that are
known to be associated with stress hormone levels. They will be looking at how providing
cats with additional hiding and climbing opportunities affects the production of
stress hormones in the hopes of being able to make specific management recommendations
for improving the behavior and breeding success of clouded leopards in zoos.
The following references provide more information on these studies:
Wielebnowski, N., Busso, J.M., Brown, J.L. 1999.
Adrenal activity in relation to subjective temperament assessment in clouded leopards (Neofelis
nebulosa). In: Felid Taxon Advisory Group Action Plan. Annual report. Eds.
D.E. Wildt, J.D. Mellen, J. Brown. American Zoo and Aquarium Association,
Disney's Animal Kingdom Publ., Lake Buena Vista, FL: p. 74-75.
Wielebnowski, N., Fletchall, N., Carlstead, K., Busso, J.M., Brown, J.L 2002.
Non-invasive assessment of adrenal activity associated of adrenal
activity associated with husbandry and behavioral factors in the North
American clouded leopard population. Zoo Biol. 21 (In Press).
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