Major
Animal Science and Technology
Advisor
Hoffman, Maria L
Advisor Department
Animal and Veterinary Science
Date
4-2019
Keywords
Fecal glucocorticoid metabolites; ELISA; rabbit
Creative Commons License
This work is licensed under a Creative Commons Attribution-Noncommercial-Share Alike 4.0 License.
Abstract
The topic for this research project was based on the problem of the diminishing population of the wild New England Cottontail rabbits. Over the last 50 years, the population of these once very common rabbits has decreased to only inhabit New England and parts of New York (1). This is less than one-fifth of the geographic range they once inhabited (1). Roger Williams Park Zoo currently has a New England Cottontail breeding program that has experienced limited success, which may be caused by the domestic environment into which this wild species is being introduced. The central objective of this study was to determine the glucocorticoid metabolite concentrations in the fecal matter of established and newly obtained New England Cottontails using an enzyme linked immunosorbent assay. This would allow us to determine if the rabbits experience periods of increased stress during captivity. In order to determine the glucocorticoid metabolite concentrations, we started by using a protocol and antibodies purchased from the Smithsonian Institute (Front Royal, VA) which had been successfully used in species of pygmy rabbits. We also performed isolations with the fecal samples using ethanol, methanol, and water. Isolates were dried using fish tank aerators and fans to remove it. In optimizing this assay we determined that different buffer formulations were needed and amounts of reagents used in order to successfully coat the plate. Specifically, we determined that a protocol from Bethyl (Montgomery, Texas), along with new plates (Nunc-maxisorp plates; Thermofisher, Waltham, MA) coated much more efficiently. To date we have been able to successfully coat the plate and run hydrocortisone standards. However, we have found that the binding of the antibody appears to be non-specific. Therefore, we are going to attempt using a different block for the assay in an effort to determine if the issue is related to the blocking step or the primary antibody/secondary antibody concentrations. References: “New England Cottontail.” New England Cottontail | New England Cottontail Management, newenglandcottontail.org/.v
Comments
For this project, 12 New England Cottontail rabbits in the Roger Williams Park Zoo conservation and breeding program had fecal samples taken, labeled in separate tubes and stored at the University of Rhode Island in a -80 degrees celsius laboratory freezer. These samples were utilized in order to work to develop a fecal glucocorticoid metabolite assay that could be used to assess stress levels in these rabbits. Different protocols, methodologies and materials were used in order to work on this research project, each with their own strengths and weaknesses. The troubleshooting we have done throughout this research project has helped to put future work on this project in the right direction in order to properly assess stress levels in these rabbits. Ultimately, I would like this research to lead to management changes that can cause an overall increase in reproductive success and population size through further conservation efforts. After working on this research project, I believe that I have expanded my knowledge in laboratory and research work, which I would not have had the opportunity to do otherwise. When I begin veterinary school this fall, I know that I will have the necessary skills to work in a research laboratory if the opportunity is presented to me. I also believe that this research project has taught me the importance of research, how much hard work is required and how difficult projects can be to complete, all of which are essential lessons to learn in the field of animal and veterinary science.