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Testing the Utility of DNA Barcoding for the Rapid Assessment of Formicidae Biodiversity in the Ethekwini Region

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Testing the Utility of DNA Barcoding for the Rapid Assessment of Formicidae Biodiversity in the Ethekwini Region Testing the utility of DNA barcoding for the rapid assessment of Formicidae biodiversity in the eThekwini region by Sohana Singh BSc. ( Hons ) Genetics Submitted in fulfilment of the academic requirements for the degree of Master of Science in The School of Life Sciences University of KwaZulu-Natal Pietermaritzburg As the candidate’s supervisor I have approved this dissertation for submission. Signed: _______________________ Name: Dr S. Willows-Munro Date: ________________ Preface The experimental work described in this dissertation was carried out in the Discipline of Genetics, School of Life Sciences, University of KwaZulu-Natal, Pietermaritzburg, from May 2011 to April 2013 under the supervision of Dr S. Willows-Munro. These studies represent original work by the author and have not otherwise been submitted in any form to another University. Where use has been made of the work by other authors it has been duly acknowledged in the text. Name: ___________________________ Signed: ___________________________ Date: ___________________________ ii College of Science and Agriculture Declaration of Plagiarism I, Sohana Singh declare that: 1. The research reported in this thesis, except where otherwise indicated, is my original research. 2. This thesis has not been submitted for any degree or examination at any other university. 3. This thesis does not contain other persons’ data, pictures, graphs or other information, unless specifically acknowledged as being sourced from other persons. 4. This thesis does not contain other persons' writing, unless specifically acknowledged as being sourced from other researchers. Where other written sources have been quoted, then: a. Their words have been re-written but the general information attributed to them has been referenced b. Where their exact words have been used, then their writing has been placed in italics and inside quotation marks, and referenced. 5. This thesis does not contain text, graphics or tables copied and pasted from the Internet, unless specifically acknowledged, and the source being detailed in the thesis and in the References sections. Signed: _______________ Date: _______________ Declaration Plagiarism 22/05/08 FHDR Approved iii Abstract Testing the utility of DNA barcoding for the rapid assessment of Formicidae biodiversity in the eThekwini region. Student: Sohana Singh Supervisor: Dr S. Willows-Munro The biodiversity of Durban (eThekwini municipality) in KwaZulu Natal is primarily threatened by urbanization although other factors such as climate change and the spread of invasive species also pose a significant threat. Knowledge of what species exist within the city is important for biodiversity surveillance, detecting invasive taxa and uncovering cryptic species. Conducting a comprehensive biodiversity inventory is a daunting task, especially for hyperdiverse groups such as terrestrial arthropods, where closely related species can often only be separated by subtle morphological characters. This study investigated whether the barcoding marker, Cytochrome Oxidase C Subunit 1 (COI) can be used to efficiently and accurately delineate species of ants (family Formicidae) in comparison to traditional taxonomic approaches. The feasibility of DNA barcoding for assembling biodiversity inventories for urban areas which could be useful in conservation planning was also evaluated. A total of 619 individuals were sequenced from 23 geographic localities within the eThekwini region and surrounding regions. DNA barcoding revealed 80 provisional species/ “barcode clusters” or monophyletic lineages which could represent distinct species, while morphology revealed 51 different morphospecies. Extrapolation measures of species richness indicated that as many as 153 species of ants could occur in the city. Phylogenetic and phylogeographic analyses were performed on co-distributed species belonging to the genera Lepisiota , Camponotus , Pheidole and Pachycondyla to better understand the spatial distribution of genetic variability in the eThekwini region. Nuclear markers 18S rDNA and 28S rDNA were also sequenced and compared for a subsample of individuals from Camponotus and Pachycondyla. There was genetic variation at COI and the nuclear markers for each of the species examined. In order to fully elucidate the population genetic patterns which could be expected in eThekwini and surrounding regions, further sampling across more localities is essential. The use of more nuclear markers could also assist in uncovering these unique patterns of genetic variation in an urban setting. In this study, the utility of COI as a species diagnostic tool in ants was confirmed. The barcoding library constructed showed promise in highlighting reserves that should be preserved and possible cryptic speciation for further investigation. iv Acknowledgements This dissertation is dedicated in memory of my late grandfather, Mr. J.L. Singh, who always encouraged me to further my education and to strive for the best. A heartfelt thanks goes out to my dad, P.J. Singh, and grandmother Mrs L. Singh, who sacrificed a lot while providing unconditional love and support so that I could study towards my dreams. I would also like to thank my siblings, Pooja and Nikhil Singh who also showed love, support and encouragement. I would like to thank my supervisor, Dr. Sandi Willows-Munro for her guidance, motivation and inspiration and for instilling in me the ethics of hard work. Thank you for sharing your knowledge and passion with me and I really appreciate the quick feedback, the ideas and encouragement. I would like to acknowledge the National Research Foundation (NRF) and the University of KwaZulu-Natal for providing the funding for this project. I would also like to thank the University of Guelph in Canada for assistance with this project. Few endeavours would be bearable without the support of good friends. Thanks goes out to my housemates, Sandipa Bhikraj, Ashnie Moolchand, Serisheena Nunnan, Kireshnie Naicker, Reansha Bissin, Kerisha Ahir and Dane Seunanden, for the late night chats, laughter and help with cooking. Thanks to my awesome friends for keeping me sane, for always being there for the amazing fun times we’ve had and for the down times… Laurencia Harris, Chavonne Chetty, Bilaal Ismail, Kerwin Moonsamy. All of you guys were basically my family away from home. Life at campus would have been boring without these epic ladies – Letrisha Padayachee, Mary Ranketse, Melissa (Milly) Govender, Lefentse Mashamaite, Mampolelo Photolo. Thanks for the fun, crazy and sometimes enlightening chats at ‘lunch time’, and for supporting me during the conference in Cape Town where we had tons of fun and adventure, and for supporting me at the campus research day. Thank you to the past and present members of the Conservation Genetics Lab – Ayanda Shelembe, Connor Stobie, Thina Duze, Qiniso and Sihle Mthetwa, Joro, Riel Coetzee and Tara Vorster, for making the lab a fun environment to be in and for thoughts and help that were needed. Collection of data for this project would not have been easy without the help of the SANBI interns, Zuki, Kirby, Matabara, Patrick, Lorika and Vince, and their assistance with field work and sorting of the insects. I would also like to thank Pooja, Ashnie, Kerisha, Letrisha, Mary, Ayanda, Shravan Ramdial, Leeth Singh and Kapil Ramisur for assistance with field work. v Finally, I want to express my gratitude for the support and assistance of the staff of the Department of Genetics at the University of KwaZulu-Natal – Dr. Pillay, Mr. Dzomba, Dr. Meusel, Megan Brunkhorst and Goodman Zondi. vi Table of Contents Preface........................................................................................................................................ ii Declaration of Plagiarism ...................................................................................................... iii Abstract ..................................................................................................................................... iv Acknowledgements .................................................................................................................... v List of tables ............................................................................................................................... x List of figures ........................................................................................................................... xii Chapter One: DNA Barcoding of Invertebrates in Africa and its Prospects for Landscape Genetics and Phylogeography – A Case Study Using the Ants of Madagascar .............................................................................................................................. 1 Abstract ...................................................................................................................................... 1 1.1 Introduction .......................................................................................................................... 2 1.1.1 Protecting our biodiversity ............................................................................................ 2 1.1.2 DNA barcoding ............................................................................................................. 3 1.1.2.2 DNA barcoding methodology, advantages and criticisms ..................................... 6 1.1.3 Importance of applying DNA barcoding to arthropods .............................................. 10 1.1.4 Below the species level – landscape
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