Development of Genetic Improvement in the African Catfish (Clarias gariepinus, Burchell, 1822) A Thesis Submitted for the Degree of Doctor of Philosophy By Suleiman Ihiabe Isa Institute of Aquaculture, Faculty of Natural Sciences, University of Stirling, Scotland, UK January 2019 Suleiman I. Isa Institute of Aquaculture 1 Declaration I declare that this thesis has been compiled entirely by me based on research I carried out. It has not been submitted for award of another degree or certificate elsewhere. All information and other sources of assistance have been duly acknowledged. Suleiman Ihiabe Isa January, 2019 Signature of Candidate ---------------------------------------------------------- Signature of First Supervisor ---------------------------------------------------------- Signature of Second Supervisor -------------------------------------------------------- 2 Suleiman I. Isa Institute of Aquaculture Acknowledgements I give praises and gratitude to God Almighty, for preserving my life and giving the ability to complete this journey. What a journey! This journey would not be complete and memorable without acknowledging the people that made the journey possible and rewarding. I am indeed very indebted to my Principal Supervisor, Dr David James Penman, who to me is more than supervisor, but also considered guardian, a mentor and a father. Without your patience, generous support, guidance, scholastic supervision, empathy and encouragement, this experience would never have been the same. I sincerely acknowledge and appreciate your of depth of knowledge, enthusiasm, quick, constructive and valuable feedbacks, and contribution to this research. Words cannot express my appreciation for your kindness. Thank you for always being there for me. I will like to thank my supervisor Dr John Taggart. Sir, permit me to call you “Stirling’s father of molecular biology”. You thought me a lot in the lab, including how to think – yes, how to think about causes of problems and possible solutions in molecular biology. I am indeed very grateful for your intellectual stimulation, time and involvement in my studies. Thank you both for the assistance during illness and bereavement. My sincere appreciation goes to my supervisor –retired, Prof. Brendan McAndrew for the support during the PhD and for the books. Thank you for your kindness. I will also like to also thank Prof. Graham Mair (External Supervisor) and Dr Andrew Davie (Internal Supervisor) for doing a very thorough job of enriching this research work with very constructive and valuable inputs. Thanks vey much for the details. Many thanks to Dr Michael Bekaert, for all the bioinformatics and timely support. I really appreciate your assistance. My special gratitude goes to Dr Kerry Bartie. You taught me how to take the first step in a molecular biology lab (DNA extraction), and assisted me all through to the end of my PhD. I owe a lot to you, thank you. I will like to express my profound gratitude to Dr Sarah-Louise Counter Selly. Your commitment, patience, and ability to simplify stuff for a beginner is exceptional, and I am indeed very grateful. I will also like to thank all the staff and students of the reproduction and genetics group Prof. Harve Migaud, Dr Andrew Davie, Dr John Taylor, Dr Stefano Carboni, Jacquie Ireland, Dr Robyn Harris, Mikey Clarkson and Yewah Jin for the good time and for the valuable inputs at one stage or the other in my PhD. A special thanks to Melanie Suleiman I. Isa Institute of Aquaculture 3 Cruikshan, Prof. David Little, Prof. Simon Mackenzie, Prof. Brett Glencross, Prof. T. C. Telfer, Prof. Sandra Adams, Prof. James Bron, Mr William Leschen, Dr John Bostock, Dr Francis Murray, Dr Armin, Dr Monica Betancor, James Dick, Fiona, Keith, Greme, Anda Kilpatrick, Sandra Tulloch and all the other staff of the institute who have made my stay here a memorable one. Thank you. Having spent such a long time here, my list of people to acknowledge is equally very long. I will like to thank the following friends and families with whom this experience was shared. My profound gratitude goes to Mr Merajuddin and his family for such a wonderful time, Mr Ian Hogg and family, Dr Ma’amun Abdullah and family, Dr Taslima Khanam and family, Dr Aisha Ambu Ali and family, Dr Aliya Al-Gapshi and family, Dr Geuseppe Paldini, Dr Olga R. and their lovely kid, Dr Khalid Shahin and family, Dr Ahmed El-Keish and family, Dr Munnever Oral, Dr janielle Wallace, Dr Amjad Ullah, Dr Kamran and family, Dr Sayfudden, Dr Julliet Nattabi, Khalfan, Atif Al-Balusi, Aqila Junaid, Claudia, Carolina, Jadwiga and others, whose name I have unintentionally omitted. Thank you all for the amazing experience. My Nigerian folks, Dr Angela Oboh, Isah Lawal, Elizabeth Buba and family, Suleiman Yakubu Omale and family, Ekpeki Akpo and family, thank you so much for the wonderful time. “I dey hail una”. To my beloved parents Late Alhaji Isah Ihiabe and Maryam Isa Ihiabe, I am very grateful for the sacrifice, upbringing, wisdom, patience discipline, care, unconditional love, support and prayers. You made me who I am today. Words cannot express my gratitude. Thanks for the grit Mom and I miss you Dad. To my siblings, thank you so much for the peace, love, sacrifice and wonderful kinship. To my beloved wife, my friend and my companion. Thank you so much for the patience, love, understanding and prayers. You stood by me during the highs and lows of this journey. I am really grateful. My daughter Maryam I. I. Suleiman and son Isa I. I. Suleiman, thanks for making me smile on a rough day. I will like to thank the UK Commonwealth Scholarship commission for funding mu PhD and taking care of me during my illness. To the doctors and nurses at Airthery Park Medical Centre and Forth-Valley Royal hospital, thank you for diagnosing and treating my illness. I am cured of cancer. Thank you. I will also like to thank the immediate past DGs of NABDA, Prof. Lucy Jummai Ogbadu, for your support and for believing in me. To the 4 Suleiman I. Isa Institute of Aquaculture present DG, Prof. Alex Akpa and the management of NABDA, thank you for the study leave and for the funding received. To Abubakar Isah, Salihu Isa, Umar Isa, Abdulqadir Adam, Tahir Nuhu, Abubakar Alhassan Muhammad, Emmanuel Oje, Jibrin and Nnnake Agbam, thanks for the support. Finally, I will like to thank Prof. B.M.B. Ladu, Prof. A. Y. Ribadu, Prof. B. O. Solomon, Mal. Sadiq, Mr Segun Adeyemi, Dr Ilyasu Jauro, Prof. A. B. Haruna and once again Prof. Lucy Jummai Ogbadu for the tutorship during my undergraduate studies and guidance in my carrier. Thank you. Suleiman Ihiabe Isa January, 2019. Suleiman I. Isa Institute of Aquaculture 5 Dedication Dedicated To The Departed Soul of My Beloved Father (Alh. Isah Ihiabe) 6 Suleiman I. Isa Institute of Aquaculture Abstract The African catfish, Clarias gariepinus, is the most important fish species for aquaculture in sub-Saharan Africa. Despite its long-standing history in aquaculture (since the 1950’s) and current rapid expansion, little work has been done on its genetics and the genetic management/improvement of different populations globally. The industry, currently worth over USD 720 million in Nigeria, and with so much more growth potential, is faced with numerous challenges. To understand the extent of these challenges and possible areas/types of intervention, the Nigerian catfish aquaculture industry was reviewed. Inadequate supply of good quality fingerlings/broodstock and feeds were notably the most significant challenges. As a step towards addressing the former, a survey of the current practise in catfish hatcheries was conducted, to identify problems and prospects therein. Over 90% of the hatcheries surveyed use shooters (fast growers) as broodstock, use only farmed broodstock and have no broodstock management/replacement programmes. Findings from these studies informed research on the development of genetic improvement for C. gariepinus. Just as in salmon, tilapia, carp, etc., the use of molecular markers as tools for genetic management and improvement of C. gariepinus was explored. Problem-solving markers, separating C. gariepinus from its closest relative, C. anguillaris, were developed. A total of 24 diagnostic SNP markers were identified from double-digest restriction-site associated DNA sequencing (ddRADseq). Following validation using KASP assay, 8 of the 24 SNPs were tested on a total of 291 Clarias catfishes and 7 Heterobranchus longifilis (out groups). The Clarias samples were separated into 259 putative C. gariepinus and 32 putative C. anguillaris. These are the first diagnostic markers for separating these species, for which morphological features perform poorly (effectively cryptic species). A set of eight new microsatellite markers was developed from the ddRADseq data and microsatellite enrichment. These microsatellite markers, together with four others sourced from the literature were optimised, multiplexed and used to genotype populations of C. gariepinus being evaluated for suitability for aquaculture. Although incomplete (due to problems with parental DNA quality), preliminary assessment of the assignment power by simulation shows that over 90% of the offspring could be assigned to a pair of parents. The high parentage assignment power and polymorphic information content (>0.5), suggest the usability and reliability of these markers in genetic management and improvement in the Clarias catfish industry, enabling parental assignment and kinship studies, and for evaluation of practices such as the use