View metadata, citation and similar papers at core.ac.uk brought to you by CORE provided by South East Academic Libraries System (SEALS) Genetic and morphometric variation of Octopus vulgaris in the Benguela Current Region A thesis submitted in fulfilment of the requirements for the degree of MASTERS OF SCIENCE of RHODES UNIVERSITY By Chénelle Lesley de Beer February 2014 Abstract The Benguela Current is a cold eastern boundary current located on the south-western coast of the African continent. The establishment of its present day features approximately two million years ago has triggered allopatric events which have driven genetic and/or phenotypic differentiation in many of the warm-temperate organisms that previously had continuous distributions along the south and west coast of southern Africa. However, since many of these species have responded differently, despite similar isolation times, research in this region provides a unique opportunity to increase our understanding of evolutionary processes. The common octopus (Octopus vulgaris, Cuvier 1797) is a coastal, sedentary species, inhabiting coral reefs or rocky environments at depths of up to a 100m. It is considered to be one of the most extensively studied cephalopod species due to its worldwide distribution. However, very little research has been conducted on O. vulgaris in southern Africa. In order to gain a holistic understanding of the effects of the Benguela Current on population connectivity, genetic and phenotypic diversity, and evolutionary history of O. vulgaris, a comparative genetic and morphological study was conducted across the Benguela region. A total of 168 specimens of O. vulgaris were collected from four different regions across the Benguela system. A small tissue sample was preserved in ethanol for molecular analysis, and the specimen was frozen whole for morphometric analysis in the laboratory. Octopus vulgaris genetic population structure and evolutionary history was investigated using a 580bp fragment of the mitochondrial cytochrome b (cytb) gene for 76 individuals located within the Benguela region, yielding 10 different haplotypes. AMOVA and pairwise FST analyses revealed significant genetic differentiation suggesting a northern-southern Benguela divergence. Estimates of time since most recent common ancestor, based on biogeographical calibrators and coalescent analyses, indicated that isolation between the Angolan and South African population occurred between ~231 Ka and 1 Ma. Mismatch distribution analyses revealed a past population expansion within the South African O. vulgaris roughly 129.31 Ka, whilst Bayesian skyline plots were indicative of gradual demographic growth within the Angolan population in the last ~100 Ka. Observed O. vulgaris population structure and demographic history was likely the result of historical climate-induced change within the system. Reconstruction of phylogenetic relationships within the Octopus genus, using cytb and COI suggest that O. vulgaris is not a monophyletic group and a major systematic revision is required. Furthermore, unidentified individuals from South Africa were found to group with ii species from Indo-West Pacific Oceans and were therefore considered to have been translocated through ballast water from Asia. While the molecular analysis indicated a significant northern-southern Benguela structure results from the principle component analysis (PCA) and discriminate function analysis (DFA) were unable to distinguish between O. vulgaris from different sampling localities throughout the Benguela Current region based on soft-parts, hard-parts and meristic characters. The lack of phenotypic variation, despite significant genetic divergence, highlights the importance of multi-method approaches in gaining a holistic understanding of the taxonomy and biogeography of species. iii Acknowledgements I would like to start by expressing my sincere gratitude to my supervisor, Warren Potts. Your unwavering enthusiasm and dedication over the past few years has really inspired diligence and passion within me. Thank you not only for the opportunity to travel and experience one of the most unique environments to which I have had the privilege to visit but for continually providing me with the opportunity to learn and progress. To Romina Henriques, my co-supervisor, your guidance, patience, support and friendship was indispensable and essential to the completion of this thesis. Furthermore, thank you for opening my eyes to the world of genetics and igniting my enthusiasm within this field. I would like to gratefully acknowledge the support of the NRF-KFD-South African Biosystematics Initiative without which this project would not have been possible Furthermore, I would like to extend my greatest gratitude to Genevieve Maharaj, from the Department of Agriculture, Forestry and Fisheries, Jan Strugnell from La Trobe University, Peter Teske and everyone else who provided and assisted with sample collection. I would also like to express my profound gratitude to the owners and staff of Flamingo Lodge, your warm hospitality will not be forgotten A special thank you to Roy Bealey for all your assistance and support throughout our postgraduate years; they definitely would have been rather dull without you. To all of my colleagues and friends, thank you for all your support, input and motivation throughout the past two years. Completion of this thesis would not have been possible without all the unconditional love and encouragement from my dad, mom and brother. Thank you for all that you have and for always providing clarity and reassurance during moments of frustration and confusion. Finally, to Carin Eaton, words cannot even begin to describe how grateful I am for all your love and support over the years. Thank you for putting up with all the craziness and tears and for always keeping me focused on what really matters. I couldn’t have done this without you! iv Contents CHAPTER 1 ......................................................................................................................................... 1 General Introduction ............................................................................................................................ 1 Study species .......................................................................................................................... 4 Commercial importance of octopus .................................................................................... 7 Project Aim ............................................................................................................................. 8 CHAPTER 2 ....................................................................................................................................... 10 General Methods and Materials........................................................................................................ 10 Sample collection, fixation and preservation ....................................................................... 10 Description of the study site ................................................................................................... 3 Major oceanographic features of the Benguela Current upwelling system ....................... 3 Changing State of the Benguela ....................................................................................... 10 CHAPTER 3 ....................................................................................................................................... 15 Population structure, evolutionary history and phylogenetic relationship of Octopus vulgaris across the Benguela Current system ................................................................................................ 15 Introduction .......................................................................................................................... 15 Method and Materials .......................................................................................................... 21 Sample collection and DNA extraction ............................................................................ 21 Cytochrome b ................................................................................................................ 22 Cytochrome Oxidase subunit I...................................................................................... 22 1st Intron of the nuclear S7 ribosomal protein .............................................................. 23 Elongation factor-1 alpha .............................................................................................. 23 Statistical analysis ............................................................................................................ 23 Population structure and phylogeographic patterns ...................................................... 24 Evolutionary History ..................................................................................................... 25 Phylogenetic Analyses .................................................................................................. 28 Results .................................................................................................................................. 30 Preliminary testing ........................................................................................................... 30 Population structure and phylogeography ....................................................................... 30 Phylogenetic analysis ......................................................................................................