NOTE TO USERS This reproduction is the best copy available. UMI* EXPLORING THE EFFICACY, UTILITY, AND LIMITATIONS OF DNA BARCODING WITHIN THE CLASS AVES A Thesis Presented to The Faculty of Graduate Studies of The University of Guelph by KEVIN CHARLES ROBERT KERR In partial fulfilment of requirements for the degree of Doctor of Philosophy April, 2010 © Kevin C. R. 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The author retains copyright L'auteur conserve la propriete du droit d'auteur ownership and moral rights in this et des droits moraux qui protege cette these. Ni thesis. Neither the thesis nor la these ni des extraits substantiels de celle-ci substantial extracts from it may be ne doivent etre imprimes ou autrement printed or otherwise reproduced reproduits sans son autorisation. without the author's permission. In compliance with the Canadian Conformement a la loi canadienne sur la Privacy Act some supporting forms protection de la vie privee, quelques may have been removed from this formulaires secondaires ont ete enleves de thesis. cette these. While these forms may be included Bien que ces formulaires aient inclus dans in the document page count, their la pagination, il n'y aura aucun contenu removal does not represent any loss manquant. of content from the thesis. •+• Canada ABSTRACT EXPLORING THE EFFICACY, UTILITY AND LIMITATIONS OF DNA BARCODING WITHIN THE CLASS AVES Kevin C. R. Kerr Advisors: University of Guelph, 2010 Professor P. D. N. Hebert Professor A. J. Baker This thesis investigates the efficacy of a recently proposed molecular bioidentifcation system known as "DNA barcoding". This system employs a short, standardized gene region (648bp of the mitochondrial gene cytochrome c oxidase I, in the case of animals) as a unique species identifier. To test species-level resolution, I constructed a library of DNA barcode sequences for birds from three regions: the Nearctic (North America), the southern Neotropics (Argentina), and the eastern Palearctic (Russia, Mongolia, and Kazakhstan). The accuracy of barcode-based species identification was assessed using the currently accepted avian taxonomy, which is the most robust of any taxonomic group. I also tested the use of DNA barcodes for species discovery via detection of large intraspecific divergences. Common intraspecific and interspecific trends in phylogeography were compared within and between biogeographical realms. Using the avian barcode library, I also compared the performance of several different methods for species delimitation, including distance-based thresholds, tree-based methods, and character-based methods (wherein each nucleotide of the sequence is treated as a unique character). Finally, I used the abundance of sequence data to test for signs of selection in cytochrome c oxidase I. Whole mitochondrial genomes available from GenBank were used to review the consistency of selective pressure throughout the genome. This largely confirmed the role of purifying selection in the evolution of the mitochondrial genome in birds. Overall, this study substantiates the utility of DNA barcoding as a reliable tool for the purposes of species identification and for highlighting taxa in need of further taxonomic review. ACKNOWLEDGEMENTS This research was funded by an Ontario Graduate Scholarship and an Elgin Card terrestrial zoology scholarship to myself, and, to a greater extent, by formidable grants to Paul Hebert from Genome Canada, the Natural Sciences and Engineering Research Council of Canada, and the Gordon and Betty Moore Foundation. In its entirety, the research presented in this thesis is truly a culmination of the efforts of many, to all of whom I owe a debt of gratitude. First and foremost, I must thank Paul Hebert for seeing my potential and for providing me with so many tremendous opportunities, some beyond which I could have ever imagined. Without him, none of this would have been possible. I have been incredibly fortunate to have an extensive support network and the opportunity to collaborate with so many wonderful people. My co-advisor, Allan Baker, and Mark Peck of the Royal Ontario Museum provided samples, insightful discussion, and behind-the-scenes tours. Dr. Mark Stoeckle of the Rockefeller University was always willing to offer thoughtful commentary and, when in a pinch, sound medical advice. Carla Dove and her myriad of technicians at the Smithsonian Institution helped tremendously in wrangling up specimens and by contributing sequences. Sharon Birks and staff at the Burke Museum of Natural History (including, but not limited to, Rob Faucett, Chris Wood, and Sievert Rohwer) were kind enough to host me at their museum and provide extensive access to their remarkable tissue collection. Charles Francis and Michel Gendron of the Canadian Wildlife Service also offered valuable collaboration. I owe unfuerte abrazo to Pablo Tubaro for having the foresight to get involved with DNA barcoding. Our ongoing student exchange has been one of the best parts of my i graduate experience. Accordingly, I thank Dario Lijtmaer for being such an excellent collaborator and for occasionally serving as my translator, I thank Ana Barreira and Pilar Benites for getting me home from the jungle in one piece, and I thank Cecilia Kopuchian, Leonardo Campagna, and everyone else at the museum for all of our interactions. The Hebert Lab has been a great academic home. I must thank all lab mates, past and present: Tyler Zemlak, Elizabeth Clare, John Wilson, Taika von Konigslow, Vazrick Nazari, Christina Carr, and Erin Corstorphine. I thank Sujeevan Ratnasingham, who on account of his genius has been the backbone of this whole endeavour (and occasionally stopped to have fun too). I also thank all of the other brains that have helped me out along the way: Jeremey deWaard, Jonathan Witt, Alex Smith, Mehrdad Hajibabaei, Nataly Ivanova, Alex Borisenko, Dirk Steinke, Rob Dooh, and Justin Schonfeld. Lab support came from Chris Grainger, Janet Topan, Constantine Christopoulos, Isabelle Meusnier and Liuqiong Lu, as well as notably from Angela Hollis of the Genomics Facility. Jinzhong Fu and Teri Crease have both served very important roles on my committee and I have relied on them both for support. The Department of Integrative Biology is generally filled with terrific faculty, of whom I would like to especially thank Jim Bogart, Elizabeth Boulding, Bob Hanner, Brian Husband, Ryan Gregory, Denis Lynn, Steve Newmaster, Beren Robinson, and Don Stevens. The students in this department (Darren Sleep, Marc Freeman, Nathalie Newby, Mark Sherrard, Kate Crosby, John Urquhuart, Joe Crowley, Martin Brummell, and Sarah Alderman, to name just a few) have been great friends and I could not imagine the experience without them. This experience has taught me the essential value of museum collections, without which a study of this magnitude would never be possible, as should be evidenced by this ii laundry list of collaborating institutions. For providing tissue samples I thank the Royal Ontario Museum (including volunteers of Toronto's Fatal Light Awareness Program), Burke Museum of Natural History and Culture, Zoological Museum of Moscow Lomonosov University, Museo Argentino de Ciencias Naturales, and the Canadian Wildlife Service. For additional samples processed via the Smithsonian I would like to thank the Academy of Natural Sciences Philadelphia, American Museum of Natural History, Field Museum, Museum of Comparative Zoology, Louisiana State University, Museum of Southwestern Biology, Museum of Vertebrate Zoology, National Museum of Natural History, University of Alaska-Fairbanks, and the University of Kansas. Bob Montgomerie (Queen's University) and Chris Earley (The Arboretum, University of Guelph) provided additional specimens and Irby Lovette (Cornell University) shared unpublished sequences. Feather samples were generously provided by the following stations: Albert Creek Bird Observatory, Appalachian Highlands Science Learning Center of the US National Park Service, Brier Island Bird Migration Research Station, Gros Morne National Park Migration Monitoring Station, Haldimand Bird Observatory, Innis Point Bird Observatory, Inglewood Bird Observatory, Long Point Bird Observatory, Mackenzie Nature Observatory, McGill Bird Observatory, Rock Point Bird Banding Station, Rocky Point Bird Observatory, St. Andrews Banding Station, Tommy Thompson Park Bird Research Station, and Vaseux Lake Migration Monitoring Station. Lastly, but most importantly, I thank my parents, Gary and Sandy,