GENETIC DIFFERENTIATION AMONG POPULATIONS OF BALD EAGLES, HALIAEETUS LEUCOCEPHALUS by Ericka Elizabeth Helmick A Thesis Submitted to the Faculty of The Charles E. Schmidt College of Science in Partial Fulfillment of the Requirements for the Degree of Master of Science Florida Atlantic University Boca Raton, Florida May 2011 Copyright by Ericka Elizabeth Helmick 2011 ii GENETIC DIFFERE TIAnON AMO G POPULATIO S OF BALD EAGLES, HALIAEETUS LEUCOCEPHALUS by Ericka Elizabeth Helmick This thesis was prepared under the direction of the candidate's thesis advisor, Dr. Colin R. Hughes, Department of Biological Sciences, and has been approved by the members of her supervisory committee. It was submitted to the faculty of the Charles E. Schmidt College of Science and was accepted in partial fulfillment of the requirements for the degree ofMaster of Science. SUPE:~---==M=I_TT_E_E_: _ Colin Hughes, Ph.D. Thesis Advisor ale GawlIk, Ph.D. 4co~ Gary . erry, P .D. Dean, The Charles E. Sc dt College of Science 13~7. ~~ Barry T. Rosson,'--Ph.D. Dean, Graduate College III ACKNOWLEDGEMENTS Special thanks go to my committee members who without their patience I would not have been able to accomplish this study. I am extremely grateful to those who were willing to share their bald eagle samples for this project, without their cooperation, this study definitely would not have been possible: Brian Mealey, Phil Schempf, and Dr. Daniel Wolf, I owe each of you quite a bit. Thanks to the DIS students that I had for helping me in the lab: M. Creamer and R. Debernardi (special thanks for working into the wee hours of the morning and keeping me company). For my parents who always knew that I would finish ―schooling‖ someday and supported me with lots of love; hours of political conversations, that not only succeeded in getting my mind off my work but also increased my blood pressure exponentially. Thanks to all of my fellow students at FAU and their support in conversations and diversions. I would also like to thank my colleagues and friends at the University of Florida, especially Dr. N. Harrison who gave me a job, even though I was pursuing a graduate degree. To all of my friends, thanks for the support and the ―you can do it‖ attitudes. Finally, thanks to Dr. T. Chouvenc for keeping me motivated, fed, and for always letting me know when I was just too tired to keep going. iv ABSTRACT Author: Ericka Elizabeth Helmick Title: Genetic Differentiation Among Populations of Bald Eagles, Haliaeetus leucocephalus Institution: Florida Atlantic University Thesis Advisor: Dr. Colin Hughes Degree: Master of Science Year: 2011 The bald eagle, Haliaeetus leucocephalus, population declined dramatically in the early 20th century reducing the population from tens of thousands of birds within the lower 48 states, to <450 pairs of birds, effectively inducing a population bottleneck. The overall population has recovered and was removed from the endangered species list in 2007. This study investigates whether such overall population statistics are appropriate descriptors for this widespread species. I investigated the genetic differentiation between three populations of bald eagles from Alaska, North Florida and Florida Bay using both mitochondrial and nuclear DNA loci to determine whether discrete subpopulations comprise the broad range. Significant FST values, for both mtDNA and microsatellites, were found between both Florida populations and Alaska, but not within Florida populations. Results indicate that there is strong population structure, rejecting the null hypothesis of a panmictic population. Future conservation efforts should focus on subpopulations rather than the overall population. v GENETIC DIFFERENTIATION AMONG POPULATIONS OF BALD EAGLES, HALIAEETUS LEUCOCEPHALUS TABLES .......................................................................................................................... viii FIGURES ............................................................................................................................ x INTRODUCTION .............................................................................................................. 1 Overview ................................................................................................................. 1 Objectives ............................................................................................................... 2 Molecular Markers .................................................................................................. 2 Theoretical Background .......................................................................................... 6 Study Populations ................................................................................................... 7 Habitat and Nesting Preferences ................................................................. 8 Dispersal and Migration .............................................................................. 9 Breeding and Reproduction ...................................................................... 12 Population Structure.................................................................................. 14 Current Genetic Data of Bald Eagles ........................................................ 15 MATERIALS AND METHODS ...................................................................................... 17 Sample Collection ................................................................................................. 17 DNA Extraction .................................................................................................... 18 PCR Amplification and Sequencing ..................................................................... 18 Mitochondrial Control Region .................................................................. 18 Microsatellite Loci .................................................................................... 19 vi DATA ANALYSES ......................................................................................................... 21 Mitochondrial Control Region .............................................................................. 21 Microsatellite Loci ................................................................................................ 22 RESULTS ......................................................................................................................... 23 Mitochondrial Control Region Variability – Domain I ........................................ 23 Haplotypes and polymorphic sites ............................................................ 23 Overall population differentiation............................................................. 23 Differentiation among populations ........................................................... 24 Mitochondrial Control Region Variability – Domain I and II .............................. 24 Haplotypes and polymorphic sites ............................................................ 24 Overall population differentiation............................................................. 25 Differentiation among populations ........................................................... 26 Other Outcomes – Assessment of Female Turnover in Florida Bay .................... 26 Microsatellite gene diversity ................................................................................. 27 Gene diversity among populations............................................................ 28 Gene diversity within populations ............................................................ 28 Assignment of Individuals to Populations ................................................ 29 DISCUSSION ................................................................................................................... 31 Genetic diversity ................................................................................................... 31 Implications for conservation of Florida populations ........................................... 34 CONCLUSION ................................................................................................................. 36 REFERENCES ................................................................................................................. 50 vii TABLES Table 1. Variable sites, numbers and frequency of 22 mtDNA haplotypes (H) based upon Domain I of mtDNA ...............................................................................38 Table 2. Overall population variation for 22 haplotypes from Domain I ......................38 Table 3. Matrix of within population FST values (below diagonal) and corresponding P-values (above diagonal) for Domain I of the mtDNA control region ...................................................................................................39 Table 4. Exact test of differentiation P-values for Domain I .........................................39 Table 5. Molecular diversity estimates for Domain I haplotypes. Sample size (n), number of haplotypes (H), nucleotide diversity (π), haplotype diversity (h) and average number of nucleotide differences (k). Standard error is in parenthesis........................................................................................................40 Table 6. Variable sites, numbers and frequency of 24 mtDNA haplotypes (H) based upon Domains I and II of mtDNA control region............................................41 Table 7. Overall population variation for 24 haplotypes of Domains I and II ...............42 Table 8. Molecular