Tackling Evolutionary Questions in Fishes with Genome-Wide Data from Recent Speciation to Ancient Divergences by Lily Claire Hughes B.A. in Biology, May 2011, Clark University M.A. in Biology, October 2012, Clark University A Dissertation submitted to The Faculty of The Columbian College of Arts and Sciences of The George Washington University in partial fulfillment of the requirements for the degree of Doctor of Philosophy January 19, 2018 Dissertation directed by Guillermo Ortí Louis Weintraub Professor of Biology The Columbian College of Arts and Sciences of The George Washington University certifies that Lily Claire Hughes has passed the Final Examination for the degree of Doctor of Philosophy as of August 30, 2017. This is the final and approved form of the dissertation. Tackling Evolutionary Questions in Fishes with Genome-Wide Data from Recent Speciation to Ancient Divergences Lily Claire Hughes Dissertation Research Committee: Guillermo Ortí, Louis Weintraub Professor of Biology, Dissertation Director Robert Alexander Pyron, Robert F. Griggs Assistant Professor of Biology, Committee Member Vanessa Liz González, Computational Genomics Scientist, Global Genome Initiative, Smithsonian Institution, National Museum of National History, Committee Member ii © Copyright 2018 by Lily Claire Hughes All rights reserved iii Acknowledgements The author wishes to thank her advisor, Guillermo Ortí, and her wonderful committee, Alex Pyron, Vanessa González, Keith Crandall, and Liz Alter, for their comments and valuable insights on this dissertation. Additionally, I would like to acknowledge the support from the Biological Sciences department I received, and funding from the Weintraub and Harlan families that supported me over the course of my studies at the George Washington University. I want to thank the many collaborators I worked with during the course of my research, from all over the world. These individuals include: Gustavo M. Somoza and Yamila Cardoso (Instituto de Investigaciones Biotecnológicas- Instituto Tecnológico de Chascomús (CONICET-UNSAM), Chascomús, Argentina), Bryan B. Nguyen and James P. Bernot (Computational Biology Institute, George Washington University), Mariano González-Castro and Juan Martín Díaz de Astarloa (Grupo de Biotaxonomía Morfológica y molecular de peces, IIMyC- CONICET, Universidad Nacional de Mar del Plata, Mar del Plata, Argentina), Julie Sommer (School of Biological Sciences, University of Nebraska-Lincoln), Roberto Cifuentes and Evelyn Habit (Departmento de Sistemas Acuáticos, Facultad de Ciencias Ambientales y Centro EULA, Universidade Concepción y Centro de Ivestigaciones en Ecosistemas Patagónicos, Concepción, Chile), Mariela Cuello (Facultad de Ciencias Naturales y Museo, Universidad Nacional de La Plata, Buenos Aires, Argentina), Victor Cussac (Instituto de Investigaciones en Biodiversidad y Medioambiente (INIBIOMA), Universidad Nacional del Comahue, Consejo Nacional de Investigaciones Cieníficas y Técnicas (CONICET), Bariloche, Rio Negro, Argentina), Luiz Malabarba (Departamento de Zoologia, Intituto de Biociências, Universidade iv Federal do Rio Grande do Sul, Porto Alegre, Brazil), Yu Huang, Ying Sun, Xiaomeng Zhao, Xiafeng Li, Min Wang, Chao Fang, Bing Xie, Qiong Shi (BGI-Shenzhen, Shenzhen, China), Zhuocheng Zhou (China Fisheries Association, Beijing, China), Soling Chen (Yellow Sea Fisheries Research Institute, Chinese Academy of Fishery Sciences, Quingdao, China), Carole Baldwin (Smithsonian Institution, National Museum of Natural History), Chenhong Li (Key Laboratory of Exploration and Utilization of Aquatic Genetic Resources, Shanghai Ocean University, China), Leandro Becker and Nicolás Bellora (Universidad Nacional de Comahue (CONICET), Laboratorio de Ictiología y Acuicultura Experimental (IPATEC), Bariloche, Argentina. I especially want to thank all the members of the GO Lab who supported and inspired me along the way: Andrew W. Thompson, Daniela Campanella, Dahiana Arcila and Ricardo Betancur-R (now at University of Puerto Rico- Rio Piedras). My fellow graduate students in Biological Sciences including Robert Kallal, Jesus Ballesteros, Mariana Abarca, Joao Tonini, Andrew Moore, Thiago Moreira, Amy Milo, David Stern, Meredith Fontana, Karen Poole, and Joshua Storch were an invaluable source of knowledge and support. Hartmut Doebel and Catriona Hendry made me a better teacher during my time as a T.A. The incredible faculty of the department also supported me, including Gustavo Hormiga, Jim Clark, Diana Lipscomb, Kathy Forester, Leon Grayfer, Keryn Gedan, Arnaud Martin, and Amy Zanne. I would also like to thank my Master’s advisors Susan Foster and John Baker at Clark University, for encouraging me to pursue my PhD in the first place. Finally, on a personal note, I want to thank my mom, Linda Ziemer, and Mary Peterson, for all the love, support, and care packages a woman could ask for. I want to v thank Pepper, for reminding me to take breaks when I was writing this dissertation by standing on my keyboard. Last, but never least, I thank Joey Stiegler, who told me I could do it, every step of the way. vi Abstract of Dissertation Tackling Evolutionary Questions in Fishes with Genome-Wide Data from Recent Speciation to Ancient Divergences Ray-finned fishes (Actinopterygii) form the largest clade of vertebrates with more than 34,000 valid species currently described. Here, I use a variety of sequencing approaches to tackle evolutionary questions in fishes, from recent habitat transitions in a group of silverside fishes, to disentangling over 400 million years of evolutionary change to infer the phylogeny of all ray-finned fishes. Chapter two of my dissertation focuses on silverside fishes in the genus Odontesthes (Atherinopsidae), which have recently and rapidly transitioned from marine to freshwater. Two closely related species, marine O. argentinensis and freshwater O. bonariensis, span this ecological divide, and yet show little genetic divergence in mitochondrial markers. We sequenced gill transcriptomes from wild-caught O. argentinensis and O. bonariensis to look for candidate genes that might be of ecological importance in the adaptation of these fish to freshwater habitats. Chapter three focues on resolving relationships in the genus Odontesthes using a large ddRADseq dataset with more than 150 samples, and cytochrome b haplotype dataset with more than 400 individuals, spanning the geographic range of Odontesthes. This chapter addresses the evolutionary relationships and some species boundaries in this group, which have been difficult to disentangle. Marine O. argentinensis and freshwater O. argentinensis, (discussed in chapter 2), despite having no differentiation in cytochrome b, clearly separate into distinct clades with ddRAD data. Chapter four produced the largest phylogenomic matrix (in basepairs, and backbone taxonomic sampling) to date for Actinopterygiians, using a database of 305 genomes and transcriptomes to address paralogy for an exon-capture set for fish phylogenomics. We developed a heuristic using vii topology tests to detect paralogs in our marker set based on known genome duplications affecting fishes, and inferred a phylogenetic tree to test our marker set of 1105 exons. We also addressed topological conflict in the literature using the ‘gene genealogy interrogation’ technique. viii Table of Contents Acknowledgments ......................................................................................................... iv Abstract of Dissertation ................................................................................................. vi List of Figures ................................................................................................................. x List of Tables ................................................................................................................ xii Chapter 1: Introduction ................................................................................................... 1 Chapter 2: Transcriptomic differentiation underlying marine-to-freshwater transitions in South American silversides Odontesthes argentinensis and O. bonariensis (Atheriniformes) ......................................................................................... 6 Chapter 3: Broad survey of mitochondrial DNA and genomic RAD data: species boundaries, introgression, and phylogeny of Odontesthes silverside fishes (Teleostei: Atheriniformes) ......................................................................................... 32 Chapter 4: Comprehensive phylogeny of ray-finned fishes (Actinopterygii) enhanced by transcriptomic and genomic data ............................................................. 62 References ..................................................................................................................... 91 ix List of Figures Figure 1…………………………………………………………………………………...20 Figure 2…………………………………………………………………………………...21 Figure 3…………………………………………………………………………………...22 Figure 4…………………………………………………………………………………...23 Figure 5…………………………………………………………………………………...51 Figure 6…………………………………………………………………………………...52 Figure 7…………………………………………………………………………………...53 Figure 8…………………………………………………………………………………...54 Figure 9…………………………………………………………………………………...55 Figure 10………………………………………………………………………………….56 Figure 11………………………………………………………………………………….57 Figure 12………………………………………………………………………………….58 Figure 13………………………………………………………………………………….59 Figure 14………………………………………………………………………………….82 Figure 15………………………………………………………………………………….83 Figure 16……………………………………………………………………………….....84