University of Texas at El Paso DigitalCommons@UTEP Open Access Theses & Dissertations 2015-01-01 Systematics Of The African River Frog Genus Amietia (anura: Pyxicephalidae) From Eastern Democratic Republic Of The onC go Thornton Robert Larson University of Texas at El Paso, [email protected] Follow this and additional works at: https://digitalcommons.utep.edu/open_etd Part of the Biodiversity Commons, Biology Commons, Developmental Biology Commons, and the Evolution Commons Recommended Citation Larson, Thornton Robert, "Systematics Of The African River Frog Genus Amietia (anura: Pyxicephalidae) From Eastern Democratic Republic Of The onC go" (2015). Open Access Theses & Dissertations. 1085. https://digitalcommons.utep.edu/open_etd/1085 This is brought to you for free and open access by DigitalCommons@UTEP. It has been accepted for inclusion in Open Access Theses & Dissertations by an authorized administrator of DigitalCommons@UTEP. For more information, please contact [email protected]. SYSTEMATICS OF THE AFRICAN RIVER FROG GENUS AMIETIA (ANURA: PYXICEPHALIDAE) FROM EASTERN DEMOCRATIC REPUBLIC OF THE CONGO THORNTON LARSON Department of Biological Sciences APPROVED: Eli Greenbaum, Ph.D., Chair Jerry D. Johnson, Ph.D. Shizue Mito, Ph.D. Charles Amber, Ph.D. Dean of the Graduate School Copyright © by Thornton R. Larson 2015 SYSTEMATICS OF THE AFRICAN RIVER FROG GENUS AMIETIA (ANURA: PYXICEPHALIDAE) FROM EASTERN DEMOCRATIC REPUBLIC OF THE CONGO by THORNTON R. LARSON, B.S. THESIS Presented to the Faculty of the Graduate School of The University of Texas at El Paso in Partial Fulfillment of the Requirements for the Degree of MASTER OF SCIENCE Department of Biological Sciences THE UNIVERSITY OF TEXAS AT EL PASO May 2015 ACKNOWLEDGMENTS I would like thank my family for their support through the whole graduate process and encouraging me to have a fascination with nature and the outdoors. Thanks especially to my parents, for putting up with the many frogs and muddy clothes, allowing me to pursue my interest in reptiles and amphibians. I am grateful to my Uncle Mark for exposing me to many of my first experiences with reptiles and amphibians. I thank my siblings for the adventures in various degrees of suburban wilderness that brought more than one scrape and bruise. Great appreciation is due to my grandparents for the immense support provided through the years of work. I would like to thank my graduate committee, Dr. Eli Greenbaum, Dr. Jerry Johnson and Dr. Shizue Mito. Special thanks to my thesis advisor Dr. Eli Greenbaum for allowing me into his lab at the last minute, and his excellent guidance which produced a fantastic project. I also thank other faculty members including, Dr. Craig Tweedie and Dr. Carl Lieb for their helpful opinions and advice on future directions. I thank Jens V. Vindum (California Academy of Sciences) and Alan Resetar (Field Museum of Natural History) for access to specimens. I thank former undergraduate student Delilah Castro for her initial work on Amietia, which provided the basic foundation for the project. DNA samples were sequenced by Ana Betancourt at the UTEP Border Biomedical Research Center Genomic Analysis Core Facility for services and facilities provided, supported by grants from the National Center for Q4 Research Resources (5G12RR008124-12) and the National Institute Q5 on Minority Health and Health Disparities (8G12MD007592-12) from the National Institutes of Health. I am also thankful to lab members, both past and present, for all of their patience and support. Thanks to Frank Portillo and Chris Anderson for helping me get started in the research process. Thanks to Fernie Medina, Nancy Conkey, Danny Hughes, Adan Lara, Samantha Stewart and Waleeja Rashid for all the added help and support that comes with working together in a lab. iv ABSTRACT The African river frog genus Amietia is found near rivers and other lentic water sources throughout central, eastern and southern Africa. Because the genus includes multiple morphologically conservative species, taxonomic studies of river frogs have been limited. We sampled 49 individuals of Amietia from multiple localities in and near the Albertine Rift (AR) of Burundi, Democratic Republic of the Congo and Uganda. We utilized single-gene (16S) and concatenated (12S, 16S, cytochrome b and RAG1) gene-tree analyses and coalescent species-tree analyses to construct phylogenetic trees. Two divergence dating approaches were used in BEAST, including secondary calibration points with 12S, 16S and RAG1 and a molecular clock with the 12S, 16S and cyt b gene. All analyses recovered Amietia as monophyletic with strong support, and revealed several well-supported cryptic lineages, which is consistent with other recent phylogeography studies of AR amphibians. Dating estimates were similar, and Amietia diversification is consistent with global cooling and aridification events in the Miocene and Pliocene, respectively. Our results suggest additional taxonomic work is needed to describe multiple new species of AR Amietia, which have limited geographic distributions that are likely to be of conservation concern. v TABLE OF CONTENTS ACKNOWLEDGMENTS ........................................................................................................ iv ABSTRACT .............................................................................................................................. v TABLE OF CONTENTS ......................................................................................................... vi LIST OF TABLES ................................................................................................................. viii LIST OF FIGURES .................................................................................................................. ix CHAPTER 1: INTRODUCTION ............................................................................................. 1 1.1 Background ................................................................................................................ 1 1.2 Natural history ............................................................................................................ 3 1.3 Taxonomy and systematics......................................................................................... 4 1.4 Study site .................................................................................................................... 5 1.5 Research objectives .................................................................................................... 8 CHAPTER 2: MATERIALS AND METHODS ....................................................................... 9 2.1 Specimen acquisition and taxon sampling ................................................................. 9 2.2 Laboratory protocols ................................................................................................ 10 2.3 Sequencing alignment and phylogenetic analysis .................................................... 10 2.4 Species trees and species delimitation ..................................................................... 11 2.5 Divergence dating .................................................................................................... 12 CHAPTER 3: RESULTS ........................................................................................................ 15 3.1 Gene-tree analyses.................................................................................................... 15 3.2 Species tree and species delimitation ....................................................................... 20 3.3 Divergence dating .................................................................................................... 21 CHAPTER 4: DISCUSSION .................................................................................................. 26 4.1 Biogeography of Albertine Rift Amietia .................................................................. 26 4.2 Species limits and taxonomy .................................................................................... 28 vi 4.3 Conservation concerns ............................................................................................. 31 LITERATURE CITED ........................................................................................................... 33 APPENDIX I ........................................................................................................................... 49 VITA ....................................................................................................................................... 53 vii LIST OF TABLES Table 2.1.1. Primers used for sequencing mitochondrial and nuclear genes .......................... 9 Table 3.3.1. Calibrated time estimates for most relevant splits within Ranoidea from BEAST analyses. Calibration points are provided in the methods. ..................................................... 22 Table 3.3.2. Molecular-clock estimates for most relevant splits within Ranoidea from BEAST analyses. Molecular clock rates are provided in the methods. ............................................... 23 viii LIST OF FIGURES Fig. 1.4.1 Elevation map for the Albertine Rift showing sampling localities for Amietia in this study. Colored shapes correspond to localities of clades within the phylogenies of Figs. 3.1.1.–3.1.2. ...................................................................................................................... 7 Fig. 3.1.1 Maximum-likelihood phylogeny with