The Pennsylvania State University The Graduate School School of Forest Resources KEY AND ATLAS TO THE HAIR OF TERRESTRIAL PENNSYLVANIA MAMMALS A Thesis in Wildlife and Fisheries Science by Andrea Nickoloff Ó 2013 Andrea Lee Nickoloff Submitted in Partial Fulfillment of the Requirements for the Degree of Master of Science December 2013 The thesis of Andrea Nickoloff was reviewed and approved* by the following: Jay R. Stauffer Distinguished Professor of Ichthyology Thesis Advisor Michael G. Messina Head and Professor, Department of Ecosystem Science and Management Matthew D. Hurteau Assistant Professor of Forest Resources *Signatures are on file in the Graduate School iii ABSTRACT Hair is considered one of the synapomorphies (shared derived characters) of extant mammals. Keys and atlases of mammal hairs can be useful for many purposes. Although rare, such keys and atlases can be used for studies of food habits of predators, species identification of material recovered in the illegal trade of wildlife parts and products, determining diet changes, taxonomic and phylogenetic studies, archaeology, research on the contamination of mercury and other metals in mammals, and behavioral studies. These keys and atlases can also be used as a non- invasive method for censusing. In Pennsylvania, there are approximately 70 extant or extinct species of mammals. I determined if: 1) the families of Pennsylvania mammals could be diagnosed based on hair; and 2) whether species within the families Sciuridae (squirrels) and Soricidae (shrews) could be diagnosed by their hair. For this investigation, I examined guard hairs from mammalian species within the Commonwealth of Pennsylvania to 1) determine the synapomorphies of guard hairs from each family; 2) create a taxonomic tree and table to the mammals of Pennsylvania and; 3) create a tool for identifying mammal species that can be utilized with non-lethal sampling approaches. iv TABLE OF CONTENTS List of Figures..................................................................................................................... v List of Tables ...................................................................................................................... vi Acknowledgements ............................................................................................................. vii Chapter 1: Introduction ....................................................................................................... 1 Mammal Hair...................................................................................................................... 1 Characteristics to Identify Hair ............................................................................................ 3 How Hairs are Obtained ...................................................................................................... 4 Previous Studies ................................................................................................................. 4 Chapter 2: Purpose .............................................................................................................. 5 Objectives................................................................................................................................ .6 Hypothesis ......................................................................................................................... 30 Chapter 3: Procedure ........................................................................................................... 30 Gross Features of Dorsal Guard Hairs and Making Slides......................................................30 Microscopic Features of Dorsal Guard Hairs I: Cuticular Scale Patterns............................... 31 Microscopic Features of Dorsal Guard Hairs II: Medullar Patterns........................................31 Part III: MALDI-TOF Mass Spectrometry..............................................................................32 Results and Discussion............................................................................................................36 Conclusion............................................................................................................................. .40 Effectiveness of Each Method by Order..................................................................................41 References................................................................................................................................207 v LIST OF FIGURES Figure 1: Cuticular and Medullar Patterns......................................................................................34 Figure 2: Dichotomous Key............................................................................................................45 Appendix A: Pennsylvania Mammal Species Cuticular and Medullar Slides with Accompanying Measurement and Counts Data......................................................................................................53 Appendix B: Data Spreadsheet.....................................................................................................107 Appendix C: Pennsylvania Mammal Species Mass Spectrometry of Guard Hair Protein Composition................................................................................................................................147 Appendix D: Supplies for MALDI-TOF Mass Spectrometry.....................................................204 Appendix E: Glossary................................................................................................................. 206 vi LIST OF TABLES Table 1-1. Family Accounts of Pennsylvania Mammals.....................................................7 Table 2-1. List of Pennsylvania Mammals ........................................................................... 14 Table 3-1. Semi-Specific Peaks (SEMPs)...……………………………………….........................................................28 vii ACKNOWLEDGEMENTS I would, first of all, like to thank my original advisor, Dr. Jacqualine Grant, for designing such a unique and fascinating thesis project for me. I would also like to thank my current advisor, Dr. Jay Stauffer Jr. for taking me in as one of his students, being such a great mentor to me with his support and guidance, and sharing his interesting stories of his fish research and travels around the world. I would like to also thank my other committee members, Dr. Michael Messina and Dr. Matthew Hurteau, for their feedback, and for our weekly conversations during Coffee Hour. I would like to thank Dr. Bruce Stanley, of the Penn State College of Medicine in Hershey, who conducted the mass spectrometry experiments, Dr. Tatiana Laremore, Director of the Proteomics and Mass Spectrometry Core Facility of the Huck Institutes of the Life Sciences, who supplied the chemicals and other equipment, and Denny Coleman, an undergraduate student who coordinated this endeavor for me. I am also grateful for the suggestions and useful information provided to me by Dr Reena Roy, from the Department of Forensic Science. I am thankful for my undergraduate professors at Delaware Valley College, especially Drs. Benjamin Rusiloski and Christopher Tipping, and Mr. Reginald Hoyt, for their support, and for their patience in writing countless letters of recommendation for my graduate school applications. Without all of these people, my thesis would not be possible. I am fortunate to have had several helpful friends, both in the Stauffer Lab and other labs. These include Rich Taylor, who helped me find and purchase equipment, Haskell Sie (Department of Statistics), and Bill Hanson, who were always willing to help me with the challenges of statistics, and Keith Price (Hunter Carrick Lab), who helped with microscope issues. The labmate I am most thankful for is Casey Weathers, who was always ready to lend a hand and assisted me with many things, including formatting my thesis, and develop techniques viii for hair measurement and photography with the microscope. I enjoyed spending time with them, learning about, sharing our enthusiasm for, and discussing our research, and any other topic we felt like talking about. Most of all, I am very thankful for my parents, Diane Nickoloff, and Dr. Edward Nickoloff, D.Sc., Professor of Radiology at the Columbia University College of Physicians and Surgeons, for being very supportive of me and my goals and ambitions. I especially want to thank my dad for teaching me to never give up, for inspiring and encouraging my interest in science and all things related to the outdoors, and for his unending love and guidance. 1 Chapter 1: Introduction Mammals (Class Mammalia) include 26 orders, and over 5000 species. Hair is one of the synapomorphies (shared characteristics) that are unique to mammals, and all mammals have hair at some point in their development (University of Michigan Museum of Zoology 2013). There are two layers to the skin of a mammal: the epidermis and the dermis (Teerink 1991). The dermis is the layer that has pain and touch receptors, and gives strength and flexibility to the skin (The Ohio State University Wexner Medical Center). The epidermis, or upper layer, contains both dead and living cells. Part of the epidermis is a basal layer, which has cells that constantly divide to maintain the epidermis. The dermis, or lower layer, has cells that form a connective-tissue sheath around the hair follicle, which has small blood capillaries that supply nutrients to the tissue. A wax gland is formed from outgrowths