Molecular Systematics and Biogeographic History of Oniscidean Isopod Troglofauna in Groundwater Calcretes of Central Western Australia Seyedmohammad Javidkar Australian Centre for Evolutionary Biology and Biodiversity School of Earth and Environmental Sciences The University of Adelaide, South Australia A Thesis Presented for the Degree of Doctor of Philosophy March 2014 CONTENTS Declaration ................................................................................................................................................................................................ iv Abstract ....................................................................................................................................................................................................... vi Acknowledgements .............................................................................................................................................................................. viii CHAPTER I: GENERAL INTRODUCTION ............................................................................................................................................. 1 Isopoda classification and phylogeny ......................................................................................................................................... 2 Subterranean ecosystems, calcrete aquifers of central Western Australia and the associated diversity ....... 6 Troglofauna associated with the Western Australian calcretes ..................................................................................... 11 Colonisation and evolution of subterranean fauna ............................................................................................................. 13 Aims of project ................................................................................................................................................................................... 16 References ........................................................................................................................................................................................... 17 CHAPTER II: MOLECULAR PHYLOGENY OF ISOPODS ATTRIBUTED TO THE PLATYARTHRIDAE (CRUSTACEA, ISOPODA, ONISCIDEA) REVEALS A NEW SOUTHERN HEMISPHERE ONISCIDEAN FAMILY ........................................ 28 Statement of Authorship ............................................................................................................................................................... 29 Abstract ................................................................................................................................................................................................. 31 Introduction......................................................................................................................................................................................... 32 Evolution of the water conducting system and lungs (pseudotracheae) ............................................................... 33 Material and Methods .................................................................................................................................................................... 35 collecting methods ...................................................................................................................................................................... 35 DNA extraction and sequencing ............................................................................................................................................. 36 Alignment, taxon sampling and phylogenetic analyses ................................................................................................ 38 Scanning Electron Microscopy (SEM) ................................................................................................................................... 42 Results ................................................................................................................................................................................................... 45 Individual and combined phylogenetic analyses ............................................................................................................. 45 V1/V2 length variation, GC content and molecular signatures ................................................................................. 49 Structure of the water conducting system on the second antennae ...................................................................... 50 Systematics ..................................................................................................................................................................................... 54 Discussion ............................................................................................................................................................................................. 66 Molecular phylogeny and relationships .............................................................................................................................. 66 Morphological comparisons .................................................................................................................................................... 69 Conclusion ............................................................................................................................................................................................ 71 Acknowledgements .......................................................................................................................................................................... 72 References ........................................................................................................................................................................................... 72 CHAPTER III: MOLECULAR SYSTEMATICS AND DIVERSIFICATION PATTERNS OF ONISCIDEAN ISOPOD TROGLOFAUNA IN GROUNDWATER CALCRETES OF CENTRAL WESTERN AUSTRALIA ................................................ 81 i Statement of Authorship ............................................................................................................................................................... 82 Abstract ................................................................................................................................................................................................. 85 Introduction ........................................................................................................................................................................................ 86 Material and Methods .................................................................................................................................................................... 88 field sampling/Sorting ................................................................................................................................................................ 88 DNA extraction and sequencing ............................................................................................................................................. 89 Identifying a new candidate nuclear gene for phylogenetics using next generation sequencing (NGS) analyses. .......................................................................................................................................................................................... 91 Phylogenetic analyses ................................................................................................................................................................ 93 Species delimitation .................................................................................................................................................................... 95 Results ................................................................................................................................................................................................... 99 Single mitochondrial “COI” and nuclear “Lysyl-tRNA Synthetase” analyses; GC content and Transition/Transversion estimation .................................................................................................................................. 100 Species delimitation ................................................................................................................................................................. 101 Combined Phylogenetic Analyses ....................................................................................................................................... 102 Discussion ......................................................................................................................................................................................... 110 Diversity, divergences and species delineation ............................................................................................................ 110 Distribution patterns ............................................................................................................................................................... 113 Nuclear marker candidates and relationships ............................................................................................................... 114 The role of historical events in the formation of the oniscidean troglofauna