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PAIRING CLOUDED LEOPARDS (NEOFELIS NEBULOSA) IN A CAPTIVE BREEDING PROGRAM By KATHERINE MARGARET MACKINNON A thesis submitted in partial fulfillment of the requirements for the degree of MASTER OF SCIENCE IN ANIMAL SCIENCES WASHINGTON STATE UNIVERSITY Department of Animal Sciences MAY 2008 To the Faculty of Washington State University: The members of the Committee appointed to examine the thesis of KATHERINE MARGARET MACKINNON find it satisfactory and recommend that it be accepted. ___________________________________ Chair ___________________________________ ___________________________________ ___________________________________ ii ACKNOWLEDGEMENTS After many tears, smells, laughter, stress, much time, and a bit of heartache I am completing this project with great appreciation for the opportunity and the many people who helped guide and support me through it. Multiple individuals, both in Thailand and the U.S., were involved in this project, and it would not have been completed without their assistance and contributions. First, I’ll express intense gratitude for the guidance and support by my advisors, Ruth Newberry, Ph.D. and Katey Pelican, D.V.M., Ph.D. They are outstanding mentors, providing a blend of leadership and friendship that is priceless and eternally appreciated. Ruth’s attention to maintaining the scientific integrity of the research and analysis has been an invaluable usher for this thesis. Katey’s indispensable understanding of the challenges involved with wildlife research kept me grounded in the understanding that results can be revealing and still very beneficial in this field of study, even if they are not statistically significant. They both lead by tremendous example with quiet encouragement and steadfast dedication. I’d also like to thank Dr. Paul Verrell and Dr. Jerry Reeves for serving on my committee, Dr. Rich Alldredge for statistical advice, and Dr. Jaak Panksepp for providing me with a Research Assistantship during my time at WSU. Second, I cannot fully express how crucial the support of my family has been. They are my foundation and strength, and they have made my difficult choice of living so far from them for this career a lesser burden. My parents have always pushed me to follow my dreams. Without the encouragement, inspiration and support of my mother I would not have followed iii this path, much less dared to reach for this goal. The love and pride all my parents share with me has allowed me to keep my dreams alive and even bring them within reach. Before I started on this project there were several people that have been involved in establishment and continuation of the Thailand clouded leopard breeding program, so my third thank you goes out to them. Much appreciation goes to JoGayle Howard, D.V.M., Ph.D. and Nadja Wielebnowski, Ph.D. for this amazing opportunity. The hard work and dedication (blood, sweat and tears) of Ken Lang, Rick Passaro, and the interim project managers, assistant Nonglak (Nok) Sumlanthin and keepers Tanat (Tong) Gadsomboon, Rungarun (Arun) Prasongkij, and Tossapol (Ping) Puttaraksa handling fecal collection and pairing observations made the breeding program possible and has kept it going. Appreciation also goes to the Association of Zoos and Aquariums Conservation Endowment Fund for the financial support of this project. A final thank you goes to my friends, fellow students, and coworkers, in Virginia and Washington for the support and kindness through this process. A big thank you also goes to endocrine technician Bernardo Mesa Cruz, D.V.M., and the students and interns who have helped with processing and running samples and entering data. You all helped make working in a fecal lab fun, and you made getting through this project much easier. iv PAIRING CLOUDED LEOPARDS (NEOFELIS NEBULOSA) IN A CAPTIVE BREEDING PROGRAM Abstract By Katherine Margaret MacKinnon, MS Washington State University May 2008 Chair: Ruth C. Newberry Pairing the endangered clouded leopard (Neofelis nebulosa) for reproduction in captivity is risky due to unpredictable male aggression toward females, which can result in lethal attacks or a need to intervene to prevent injury. To assess potential early indicators for successful pairings (i.e., pairings producing cubs), behavioral observations and fecal hormones were evaluated on four juvenile-male (< 1 y) and nine adult-male (> 1 y) pairs of clouded leopards in a breeding program in the Thailand Clouded Leopard Consortium at the Khao Kheow Open Zoo. Fecal cortisol concentrations, stereotyped pacing behavior and fighting were predicted to be lower in successful than failed pairs, and animals in successful pairs were predicted to display more affiliative behaviors than those in failed pairs. It also was predicted that pairs are more likely to be successful when females are paired with a juvenile male, rather than an adult male. Behavioral observations were conducted on paired males and females 2-5 times weekly. Daily fecal samples were collected for hormonal analysis of cortisol by enzyme immunoassay. Data were analyzed to identify predictors of successful (n = 5) versus failed (n = 8) pairings using generalized linear models. As predicted, animals in successful pairs exhibited higher (p < 0.05) v levels of affiliative behaviors (grooming, rubbing on other cat) than animals in failed pairs. Neither pacing nor exploratory behaviors were significantly different between animals in successful versus failed pairs. Male fecal cortisol baseline, mean and peak mean were higher in failed than successful pairs (p < 0.01). No differences were observed in female fecal cortisol mean, baseline or peak mean concentrations (p > 0.05). However, fecal cortisol coefficient of variation around the mean was higher (p < 0.05) in successful than failed females. Although pairing females with juvenile versus adult males did not directly predict success (p > 0.05), specific behaviors (e.g., groom other cat, female rub on other cat) did predict success (p < 0.05) when the juvenile/adult-male variable was included in the model. These results suggest that behavior and hormone levels could be used to predict whether a pair of clouded leopards will reproduce successfully. vi TABLE OF CONTENTS Page ACKNOWLEDGEMENTS .............................................................................................iii ABSTRACT .....................................................................................................................v LIST OF TABLES ...........................................................................................................viii LISTOF FIGURES ..........................................................................................................ix INTRODUCTION ...........................................................................................................1 Aggression: Overview .........................................................................................1 Sexual Aggression ..........................................................................................3 Aggression in Felids ......................................................................................4 Neural Pathways of Aggression .....................................................................4 Steroids and Aggression ................................................................................5 The Stress Response ............................................................................................7 The Negative Impact of Chronic Stress .........................................................8 Chronic Stress in the Captive Context ...........................................................9 How are Aggression and Stress Managed in Captive Animals? ..........................11 Hormone Monitoring of Wildlife .........................................................................13 Behavioral Observation: How are Behavior Data Collected? .............................14 Monitoring Behavior in Captive Wildlife ......................................................16 Felid Social Systems ............................................................................................16 Captive Breeding: Why is it Needed? Why Difficult for Some Species? ..........18 Clouded Leopards ................................................................................................19 Natural History and Biology ..........................................................................20 Radio-tracking Studies ...................................................................................21 Stress in Captive Clouded Leopards ....................................................................22 Challenges to Breeding Clouded Leopards in Captivity ......................................22 The Thailand Breeding Program ..........................................................................25 OBJECTIVE AND HYPOTHESES ................................................................................26 RESEARCH DESIGN AND METHODOLOGY ...........................................................28 Animals, Housing, and Husbandry ......................................................................28 Introduction Protocol ...........................................................................................30 Fecal Sample Collection, Steroid Extraction, and Enzyme Immunoassay ..........31 Behavior Observations .........................................................................................33 Statistical Analysis ...............................................................................................34
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