i BINOMICS, CULTURE TRIAL AND DNA BARCODING OF CARIDEAN SPECIES FROM THREE LAGOONS, SOUTH-WEST NIGERIA BY AKINWUNMI, MOSUNMOLA FLORENCE 119077072 B. Agric. Tech. (FEDERAL UNIVERSITY OF TECH. AKURE), 2007 A THESIS SUBMITTED FOR THE DEGREE OF DOCTOR OF PHILOSOPHY (Ph.D.) IN FISHERIES (CRUSTACEAN BIOLOGY) DEPARTMENT OF MARINE SCIENCES UNIVERSITY OF LAGOS, LAGOS NIGERIA DECEMBER, 2015. ii SCHOOL OF POSTGRADUATE STUDIES UNIVERSITY OF LAGOS CERTIFICATION This is to certify that the Thesis: BINOMICS, CULTURE TRIAL AND DNA BARCODING OF CARIDEAN SPECIES FROM THREE LAGOONS, SOUTH-WEST NIGERIA Submitted to the School of Postgraduate Studies University of Lagos For the award of the degree of DOCTOR OF PHILOSOPHY (Ph.D.) is a record of original research carried out By AKINWUNMI, MOSUNMOLA FLORENCE in the Department of Marine Sciences AUTHOR’S NAME SIGNATURE DATE 1 ST SUPERVISOR’S NAME SIGNATURE DATE 2 nd SUPERVISOR’S NAME SIGNATURE DATE 1 st INTERNAL EXAMINER SIGNATURE DATE 2 nd INTERNAL EXAMINER SIGNATURE DATE EXTERNAL EXAMINER SIGNATURE DATE SPGS REPRESENTATIVE SIGNATURE DATE iii CERTIFICATION We certify that the work embodied in this Thesis for the award of the degree of Doctor of Philosophy (Fisheries) is a record of original research carried out by AKINWUNMI Mosunmola Florence under our supervision. ___________________________ ___________________________ Dr. (Mrs) A. O. Lawal-Are Date (Associate Professor) ___________________________ ___________________________ Dr. S. O. Ayoola Date (Senior Lecturer) iv DEDICATION This thesis is deservedly dedicated to God Almighty, my parents, husband and children. v ACKNOWLEDGEMENT My profound gratitude goes to my major supervisor, Dr. (Mrs) Aderonke Omolara Lawal-Are who not only fine-tuned the topic, but supervised, encouraged and scrutinised the various stages of the drafts of the Thesis. Her corrective measures and academic motherhood since my M.Phil. studies till this level cannot be fathomed and will forever be remembered. My sincere appreciation goes to my co-supervisor Dr. Simeon Oluwatoyin Ayoola for his drive, concerted efforts and encouragement with prayers to see to the successful completion of this work. I must express my appreciation to my own academic grandfather: Emeritus Professor Kola Kusemiju who nurtured, prayed and guided me all through this study. I remain eternally grateful to Professor Lucian Obinna Chukwu for his kind heart and positive attitude to my progress and at completing this work and also to my academic father: Professor Oluayo Anthony Bello-Olusoji, who sowed the seed in me from the beginning and keeps on watering it till date. I must acknowledge Professor Dike Nwankwo, Dr. Olufemi Soyinka, Dr. (Mrs) Adesola Osibona and Professor Clement Edokpayi for their moral support. Dr. Ikenna Onyema for his assistance in identifying planktonic food items in fish stomach. Dr. Olugbenga Olude, Dr. Julius Agboola and Dr. Olusegun Samuel for their assistance in correcting the statistical analysis. Dr. Ademola Aderolu and Miss Bola Seriki for their assistance during the feed formulation and culture period. Dr. Minasu Kuton, Dr. Muyideen Lawal and Dr. Babatunde Emmanuel for their prayers, concern, encouragement and moral support. I remain eternally grateful to Professor Olusoji Ilori for his encouragement and support during the study, Professor Oluwatoyin Ogundipe for giving me the opportunity to make use of the Molecular laboratory in Department of Botany and also Dr. (Mrs) Temitope Onuminya for her support during the DNA analysis. I appreciate the effort of Dr. (Mrs) Temitope Samuel, Dr. Gbenga Adeogun for their moral support and encouragement at every stage of this work. Mr. A. Ezekiel for the provision of the microscope that was used in the laboratory throughout the study period. vi I also thank Mr. Tijani Lawal and Mrs. Remi Babalola for their laboratory assistance. Mr. Jubril Olaniyi for his help in computer work and during the culture period. Mr. Tunde Lamidi for his assistance during collection of specimens. I thank Mr. M.A. Adelabu who took the pictures of eggs developmental stages, Mr. C. James for the proximate analysis of the carcass and experimental feeds, Professor Chris Oke for his advice on statistical analysis. I will not do without appreciating my friends and colleagues; Elegbeleye Yemi, Usese Amii, Bassey Bassey, Ogundele Adurayemi, Mrs. Olumide-Ale Abiola, Akinnigbagbe Toyin, Mr. Ibanga Luke, Ogunbambo Mogbonjubola, Sandra Akagha, Oshingboye Dolapo, Nodza George, Ajikah Linus and Ngonadi Ifeanyi. I appreciate all for their competitive support. My profound appreciation goes to my parents Chief Martins Ogunwo and Deaconess Afolashade Ogunwo who brought me up in a family structure that encouraged hard work, honesty, humility, integrity, sharing and above all love. Your life has been a guiding light and beacon of hope and I am eternally grateful to you. I thank all my siblings; Mrs. Olabisi Afolabi, Miss Jumoke Ogunwo, Mrs. Damilola Omotoso and Master Damilare Ogunwo and their husbands for their immeasurable role in my life. I also appreciate my parent-in-law: Apostle Israel Oladokun Akinwunmi (of blessed memory) and Mrs. J. Akinwunmi for their spiritual and moral support. I deeply express my love and gratitude to my wonderful husband Mr. Oluwaseun Olusoji Akinwunmi who stood by me and assisted in keeping the home front peaceful and congenial especially during my long stay in the laboratory and to my precious children Toluwanimi and Toluwalase for their excellent comradeship, encouragement, total support, perseverance and confidence they had in me throughout the study. vii TABLE OF CONTENT Page Title page i Certification ii Dedication iv Acknowledgement v Table of Content vii List of Tables x List of Figures xii List of Plates xvi List of Appendices xvii List of Abbreviations xix Definition of Operational Terms xx Abstract xxi Chapter One 1 1.0 INTRODUCTION 1 1.1 Statement of problem 3 1.2 Aim and objectives 4 1.3 Significance of the study 5 Chapter Two 6 2.0 LITERATURE REVIEW 6 2.1 Classification of prawns 7 2.2 Distribution of prawns 8 2.3 Behaviour of M. vollenhovenii 9 2.3.1 Resting posture 9 2.3.2 Locomotion 9 2.3.3 Digging 9 2.3.4 Escape 10 2.3.5 Aggression 10 2.3.6 Learning and social behavior 11 2.4 Age and growth 11 2.4.1 Condition factor 12 2.5 Colour changes 13 2.6 Life history of prawns 13 2.7 Reproductive biology 14 2.7.1 Breeding season 15 2.7.2 Sex differentiation 16 2.7.3 Fecundity 17 2.8 Food and feeding habits 18 2.9 Culture of shrimps/prawns 20 2.10 DNA barcode 21 Chapter Three 24 3.0 MATERIALS AND METHODS 24 3.1 Description of study sites 24 3.1.1 Badagry Lagoon 24 3.1.2 Lagos Lagoon 24 3.1.3 Epe Lagoon 26 3.2 Field studies 26 3.2.1 Physico-chemical parameters of the water samples 27 3.2.1.1 Temperature 27 viii 3.2.1.2 Salinity 27 3.2.1.3 Dissolved oxygen (DO) 27 3.2.1.4 Hydrogen ion concentration 27 3.2.1.5 Water transparency 28 3.2.1.6 Conductivity 28 3.2.2 Collection of specimens 28 3.3 Laboratory procedures 30 3.3.1 Length-weight measurements of the Macrobrachium species 30 3.3.2 Growth pattern 30 3.3.2.1 Length-frequency distribution 30 3.3.2.2 Length-weight relationship 30 3.3.2.3 Condition factor (K) in Macrobrachium species 31 3.3.3 Food and feeding habits in Macrobrachium species 31 3.3.4 Reproductive biology 31 3.3.4.1 Sex ratio in Macrobrachium species 31 3.3.4.2 Fecundity 32 3.3.4.3 Gonadosomatic Index (GSI) 32 3.3.4.4 Egg diameter 33 3.3.5 Racial study 33 3.3.6 Flesh/Body mass ratio 34 3.3.7 Culture potentials of M. vollenhovenii using varied Crude Protein (CP) percentages 34 3.3.7.1 Composition and preparation of experimental diets 34 3.3.7.2 Experimental system 37 3.3.7.2.1 Experimental animal (M. vollenhovenii) 37 3.3.7.3 Water quality analysis 39 3.3.7.4 Feeding procedure 39 3.3.7.5 Determination of growth and nutrient utilisation parameters 39 3.3.7.5.1 Weight Gain 39 3.3.7.5.2 Percentage Weight Gain 40 3.3.7.5.3 Specific Growth Rate 40 3.3.7.5.4 Food Conversion Ratio 40 3.3.7.5.5 Food Efficiency Ratio 40 3.3.7.5.6 Protein Efficiency Ratio 40 3.3.8 Proximate Analysis of experimental diets and prawn carcasses 41 3.3.8.1 Determination of crude protein 41 3.3.8.2 Determination of crude fat 41 3.3.8.3 Determination of crude fibre 42 3.3.8.4 Determination of moisture content 42 3.3.8.5 Determination of total ash 42 3.3.8.6 Determination of nitrogen free extract 42 3.3.9 DNA barcoding 43 3.3.9.1 DNA extraction 43 3.3.9.2 Gel electrophoresis of the DNA (0.5 X TBE) 45 3.3.9.3 Gel dye preparation 45 3.3.9.4 Agarose preparation 46 3.3.9.5 Spectrophotometer analysis 46 3.3.9.6 DNA Amplification and Sequencing 47 3.4 Statistical Analysis 48 Chapter Four 49 ix 4.0 RESULTS 49 4.1 Physico-chemical parameters of the lagoons 49 4.1.1 Air and water temperatures 49 4.1.2 Dissolved oxygen (DO) 52 4.1.3 Hydrogen ion concentration (pH) 52 4.1.4 Salinity 52 4.1.5 Transparency 53 4.1.6 Conductivity 55 4.2 Size composition of M. macrobrachion and M. vollenhovenii 55 4.3 Growth pattern M. macrobrachion and M. vollenhovenii 58 4.3.1 Total-length frequency distribution of M. macrobrachion and M. vollenhovenii 58 4.3.2 Carapace length frequency distribution M. macrobrachion and M. vollenhovenii 61 4.3.3 Length-Weight relationship in M. macrobrachion and M. vollenhovenii 61 4.3.3.1 Log total-length/Log weight distribution in M. macrobrachion and M. vollenhovenii 61 4.3.3.2 Log carapace-length/Log weight distribution in M. macrobrachion and M. vollenhovenii 67 4.3.3.3 Flesh weight/Body weight relationship in M. macrobrachion and M. vollenhovenii 72 4.4 Condition factor (K) 74 4.5 Food and feeding habits 81 4.5.1 Empty stomach analysis 81 4.5.2 Food items in M. macrobrachion and M. vollenhovenii 81 4.6 Reproductive biology in M. macrobrachion and M. vollenhovenii 93 4.6.1 Sex ratio 93 4.6.2 Fecundity 103 4.6.3 Stages of egg development in M. macrobrachion and M. vollenhovenii 109 4.6.4 Egg diameter of M. macrobrachion and M. vollenhovenii 109 4.6.5 Gonadosomatic Index (GSI) 125 4.7 Racial study in M. macrobrachion and M. vollenhovenii 127 4.8 Culture trial of M. vollenhovenii 127 4.8.1 Physico-chemical parameters of the culture media 127 4.8.2 Growth and nutrient utilization parameters of M. vollenhovenii 132 4.8.3 Molting in M. vollenhovenii 136 4.8.4 Cannibalism in M. vollenhovenii 136 4.8.5 Mating in M. vollenhovenii 136 4.9 DNA barcoding 136 Chapter Five 145 5.0 DISCUSSION AND CONCLUSION 145 5.1 Discussion 145 5.2 Conclusion 158 5.3 Summary of findings 159 5.4 Contributions to knowledge 160 References 161 Appendices 180 x LIST OF TABLES Table Page 1. Purified diets formulation at varied CP levels 35 2. Local diets formulation at 40 % CP level 36 3. Proximate composition (%) of experimental diets 38 4. Ranges and means of physico-chemical parameters of Badagry, Lagos and Epe Lagoons (June 2013 – May 2015) 50 5. Size composition of M. macrobrachium and M. vollenhovenii from Badagry, Lagos and Epe Lagoons (June 2013 – May 2015) 57 6. Size groupings of M. macrobrachium and M. vollenhovenii from Badagry, Lagos and Epe Lagoons (June 2013 – May 2015) 59 7. Condition factor (K) by sex and size group of M. macrobrachion from Badagry Lagoon (June 2013 – May 2015) 75 8. Condition factor (K) by sex and size group of M. macrobrachion from Lagos Lagoon (June 2013 – May 2015) 75 9. Condition factor (K) by sex and size group of M. macrobrachion from Epe Lagoon (June 2013 – May 2015) 76 10. Condition factor (K) by sex and size group of M. vollenhovenii from Badagry Lagoon (June 2013 – May 2015) 76 11. Condition factor (K) by sex and size group of M. vollenhovenii from Lagos Lagoon (June 2013 – May 2015) 77 12. Condition factor (K) by sex and size group of M. vollenhovenii from Epe Lagoon (June 2013 – May 2015) 78 13. Empty stomachs in M. macrobrachion from Badagry, Lagos and Epe Lagoons (June 2013 – May 2015) 82 14. Empty stomachs in M. vollenhovenii from Badagry, Lagos and Epe Lagoons (June 2013 – May 2015) 82 15. Monthly variation in empty stomach of M. macrobrachion from Badagry Lagoon (June 2013 – May 2015) 83 16. Monthly variation in empty stomach of M. macrobrachion from Lagos Lagoon (June 2013 – May 2015) 84 17. Monthly variation in empty stomach of M. macrobrachion from Epe Lagoon (June 2013 – May 2015) 85 18. Monthly variation in empty stomach of M. vollenhovenii from Badagry Lagoon (June 2013 – May 2015) 86 19. Monthly variation in empty stomach of M. vollenhovenii from Lagos Lagoon (June 2013 – May 2015) 87 20. Monthly variation in empty stomach of M. vollenhovenii from Epe Lagoon (June 2013 – May 2015) 88 21. Food items in M. macrobrachion from Badagry, Lagos and Epe Lagoons (June 2013 – May 2015) 89 22. Food items in M. vollenhovenii from Badagry, Lagos and Epe Lagoons (June 2013 – May 2015) 90 23. Egg stages of M. macrobrachion from Badagry, Lagos and Epe Lagoons (June 2013 – May 2015) 116 24. Egg stages of M. vollenhovenii from Badagry, Lagos and Epe Lagoons (June 2013 – May 2015) 116 25. Monthly egg stages of M. macrobrachion from Badagry Lagoon (June 2013 – May 2015) 118 26. Monthly egg stages of M. macrobrachion from Lagos Lagoon xi (June 2013 – May 2015) 119 27. Monthly egg stages of M. macrobrachion from Epe Lagoon (June 2013 – May 2015) 120 28. Monthly egg stages of M. vollenhovenii from Badagry Lagoon (June 2013 – May 2015) 121 29. Monthly egg stages of M. vollenhovenii from Lagos Lagoon (June 2013 – May 2015) 122 30. Monthly egg stages of M. vollenhovenii from Epe Lagoon (June 2013 – May 2015) 123 31. Diameter of eggs in mature female M. macrobrachion and M. vollenhovenii from Badagry, Lagos and Epe Lagoons 124 32. Gonadosomatic index (GSI) of M. macrobrachion and M. vollenhovenii from Badagry, Lagos and Epe Lagoons 126 33. Mean meristic characters of M. macrobrachion and M. vollenhovenii from Badagry, Lagos and Epe Lagoons 128 34. Mean morphometric characters of M. macrobrachion and M. vollenhovenii from Badagry, Lagos and Epe Lagoons 128 35. Water quality within the rearing systems during the experimental culture phase 131 36. Growth and nutrient utilisation parameters of M. vollenhovenii for the 60 days culture trial 134 37. Carcass composition (%) of prawns fed varied crude protein for 60 days 135 38. Percentage closeness of the identified species in relation to pre-identified strains of M. macrobrachion, M. vollenhovenii and Penaeus monodon 140 39. The absorbance ratio and concentration of the DNA samples 143 xii LIST OF FIGURES Figure Page 1. Ventral and cross section of the first abdominal segments of female and male M. vollenhovenii 17 2. Second pleopod of a female and a male M. vollenhovenii 17 3. Map of Badagry, Lagos and Epe Lagoons showing the sampling stations 25 4. Air temperature variation in Badagry, Lagos and Epe Lagoons (June 2013 – May 2015) 51 5. Water temperature variation in Badagry, Lagos and Epe Lagoons (June 2013 – May 2015) 51 6. Dissolved oxygen (DO) variation in Badagry, Lagos and Epe Lagoons (June 2013 – May 2015) 51 7. Hydrogen ion concentration (pH) variation in Badagry, Lagos and Epe Lagoons (June 2013 – May 2015) 54 8. Salinity variation in Badagry, Lagos and Epe Lagoons (June 2013 – May 2015) 54 9. Transparency variation in Badagry, Lagos and Epe Lagoons (June 2013 – May 2015) 54 10. Conductivity variation in Badagry, Lagos and Epe Lagoons (June 2013 – May 2015) 56 11. Total length frequency distribution of M. macrobrachion from Badagry, Lagos and Epe Lagoons (June 2013 – May 2015) 60 12. Total length frequency distribution of M. vollenhovenii from Badagry, Lagos and Epe Lagoons (June 2013 – May 2015) 60 13. Carapace length frequency distribution of M. macrobrachion from Badagry, Lagos and Epe Lagoons (June 2013 – May 2015) 62 14. Carapace length frequency distribution of M. vollenhovenii from Badagry, Lagos and Epe Lagoons (June 2013 – May 2015) 62 15. Log total length – Log weight relationship of male M. macrobrachion from Badagry Lagoon (June 2013 – May 2015) 63 16. Log total length – Log weight relationship of male M. macrobrachion from Lagos Lagoon (June 2013 – May 2015) 63 17. Log total length – Log weight relationship of male M. macrobrachion from Epe Lagoon (June 2013 – May 2015) 63 18. Log total length – Log weight relationship of male M. vollenhovenii from Badagry Lagoon (June 2013 – May 2015) 63 19. Log total length – Log weight relationship of male M. vollenhovenii from Lagos Lagoon (June 2013 – May 2015) 63 20. Log total length – Log weight relationship of male M. vollenhovenii from Epe Lagoon (June 2013 – May 2015) 63 21. Log total length – Log weight relationship of female M. macrobrachion from Badagry Lagoon (June 2013 – May 2015) 64 22. Log total length – Log weight relationship of female M. macrobrachion from Lagos Lagoon (June 2013 – May 2015) 64 23. Log total length – Log weight relationship of female M. macrobrachion from Epe Lagoon (June 2013 – May 2015) 64 24. Log total length – Log weight relationship of female M. vollenhovenii from Badagry Lagoon (June 2013 – May 2015) 64 25. Log total length – Log weight relationship of female M. vollenhovenii from Lagos Lagoon (June 2013 – May 2015) 64 xiii 26. Log total length – Log weight relationship of female M. vollenhovenii from Epe Lagoon (June 2013 – May 2015) 64 27. Log total length – Log weight relationship of M. macrobrachion (combined sexes) from Badagry Lagoon (June 2013 – May 2015) 65 28. Log total length – Log weight relationship of M. macrobrachion (combined sexes) from Lagos Lagoon (June 2013 – May 2015) 65 29. Log total length – Log weight relationship of M. macrobrachion (combined sexes) from Epe Lagoon (June 2013 – May 2015) 65 30. Log total length – Log weight relationship of M. vollenhovenii (combined sexes) from Badagry Lagoon (June 2013 – May 2015) 65 31. Log total length – Log weight relationship of M. vollenhovenii (combined sexes) from Lagos Lagoon (June 2013 – May 2015) 65 32. Log total length – Log weight relationship of M. vollenhovenii (combined sexes) from Epe Lagoon (June 2013 – May 2015) 65 33. Log carapace length – Log weight relationship of male M. macrobrachion from Badagry Lagoon (June 2013 – May 2015) 68 34. Log carapace length – Log weight relationship of male M. macrobrachion from Lagos Lagoon (June 2013 – May 2015) 68 35. Log carapace length – Log weight relationship of male M. macrobrachion from Epe Lagoon (June 2013 – May 2015) 68 36. Log carapace length – Log weight relationship of male M. vollenhovenii from Badagry Lagoon (June 2013 – May 2015) 68 37. Log carapace length – Log weight relationship of male M. vollenhovenii from Lagos Lagoon (June 2013 – May 2015) 68 38. Log carapace length – Log weight relationship of male M. vollenhovenii from Epe Lagoon (June 2013 – May 2015) 68 39. Log carapace length – Log weight relationship of female M. macrobrachion from Badagry Lagoon (June 2013 – May 2015) 69 40. Log carapace length – Log weight relationship of female M. macrobrachion from Lagos Lagoon (June 2013 – May 2015) 69 41. Log carapace length – Log weight relationship of female M. macrobrachion from Epe Lagoon (June 2013 – May 2015) 69 42. Log carapace length – Log weight relationship of female M. vollenhovenii from Badagry Lagoon (June 2013 – May 2015) 69 43. Log carapace length – Log weight relationship of female M. vollenhovenii from Lagos Lagoon (June 2013 – May 2015) 69 44. Log carapace length – Log weight relationship of female M. vollenhovenii from Epe Lagoon (June 2013 – May 2015) 69 45. Log carapace length – Log weight relationship of M. macrobrachion (combined sexes) from Badagry Lagoon (June 2013 – May 2015) 70 46. Log carapace length – Log weight relationship of M. macrobrachion (combined sexes) from Lagos Lagoon (June 2013 – May 2015) 70 47. Log carapace length – Log weight relationship of M. macrobrachion (combined sexes) from Epe Lagoon (June 2013 – May 2015) 70 48. Log carapace length – Log weight relationship of M. vollenhovenii (combined sexes) from Badagry Lagoon (June 2013 – May 2015) 70 49. Log carapace length – Log weight relationship of M. vollenhovenii (combined sexes) from Lagos Lagoon (June 2013 – May 2015) 70 50. Log carapace length – Log weight relationship of M. vollenhovenii (combined sexes) from Epe Lagoon (June 2013 – May 2015) 70 xiv 51. Log flesh weight/Log body weight relationship of M. macrobrachion from Badagry Lagoon (June 2013 – May 2015) 73 52. Log flesh weight/Log body weight relationship of M. macrobrachion from Lagos Lagoon (June 2013 – May 2015) 73 53. Log flesh weight/Log body weight relationship of M. macrobrachion from Epe Lagoon (June 2013 – May 2015) 73 54. Log flesh weight/Log body weight relationship of M. vollenhovenii from Badagry Lagoon (June 2013 – May 2015) 73 55. Log flesh weight/Log body weight relationship of M. vollenhovenii from Lagos Lagoon (June 2013 – May 2015) 73 56. Log flesh weight/Log body weight relationship of M. vollenhovenii from Epe Lagoon (June 2013 – May 2015) 73 57. Condition factor (K) by sex of M. macrobrachion from Badagry Lagoon (June 2013 – May 2015) 79 58. Condition factor (K) by sex of M. macrobrachion from Lagos Lagoon (June 2013 – May 2015) 79 59. Condition factor (K) by sex of M. macrobrachion from Epe Lagoon (June 2013 – May 2015) 79 60. Condition factor (K) by sex of M. vollenhovenii from Badagry Lagoon (June 2013 – May 2015) 80 61. Condition factor (K) by sex of M. vollenhovenii from Lagos Lagoon (June 2013 – May 2015) 80 62. Condition factor (K) by sex of M. vollenhovenii from Epe Lagoon (June 2013 – May 2015) 80 63. Stomach contents of M. macrobrachion from Badagry Lagoon (June 2013 – May 2015) 94 64. Stomach contents of M. macrobrachion from Lagos Lagoon (June 2013 – May 2015) 94 65. Stomach contents of M. macrobrachion from Epe Lagoon (June 2013 – May 2015) 95 66. Stomach contents of M. vollenhovenii from Badagry Lagoon (June 2013 – May 2015) 95 67. Stomach contents of M. vollenhovenii from Lagos Lagoon (June 2013 – May 2015) 96 68. Stomach contents of M. vollenhovenii from Epe Lagoon (June 2013 – May 2015) 96 69. Monthly variation in sex ratio in M. macrobrachion from Badagry Lagoon (June 2013 – May 2015) 102 70. Monthly variation in sex ratio in M. vollenhovenii from Badagry Lagoon (June 2013 – May 2015) 102 71. Monthly variation in sex ratio in M. macrobrachion from Lagos Lagoon (June 2013 – May 2015) 102 72. Monthly variation in sex ratio in M. vollenhovenii from Lagos Lagoon (June 2013 – May 2015) 104 73. Monthly variation in sex ratio in M. macrobrachion from Epe Lagoon (June 2013 – May 2015) 104 74. Monthly variation in sex ratio in M. vollenhovenii from Epe Lagoon (June 2013 – May 2015) 104 75. Log total length/Log fecundity relationship of M. macrobrachion from Badagry Lagoon 106 xv 76. Log carapace length/Log fecundity relationship of M. macrobrachion from Badagry Lagoon 106 77. Log weight/Log fecundity relationship of M. macrobrachion from Badagry Lagoon 106 78. Log total length/Log fecundity relationship of M. vollenhovenii from Badagry Lagoon 106 79. Log carapace length/Log fecundity relationship of M. vollenhovenii from Badagry Lagoon 106 80. Log weight/Log fecundity relationship of M. vollenhovenii from Badagry Lagoon 106 81. Log total length/Log fecundity relationship of M. macrobrachion from Lagos Lagoon 107 82. Log carapace length/Log fecundity relationship of M. macrobrachion from Lagos Lagoon 107 83. Log weight/Log fecundity relationship of M. macrobrachion from Lagos Lagoon 107 84. Log total length/Log fecundity relationship of M. vollenhovenii from Lagos Lagoon 107 85. Log carapace length/Log fecundity relationship of M. vollenhovenii from Lagos Lagoon 107 86. Log weight/Log fecundity relationship of M. vollenhovenii from Lagos Lagoon 107 87. Log total length/Log fecundity relationship of M. macrobrachion from Epe Lagoon 108 88. Log carapace length/Log fecundity relationship of M. macrobrachion from Epe Lagoon 108 89. Log weight/Log fecundity relationship of M. macrobrachion from Epe Lagoon 108 90. Log total length/Log fecundity relationship of M. vollenhovenii from Epe Lagoon 108 91. Log carapace length/Log fecundity relationship of M. vollenhovenii from Epe Lagoon 108 92. Log weight/Log fecundity relationship of M. vollenhovenii from Epe Lagoon 108 93. Stages of egg development of M. macrobrachion from Badagry, Lagos and Epe Lagoons (June 2013 – May 2015) 117 94. Stages of egg development of M. vollenhovenii from Badagry, Lagos and Epe Lagoons (June 2013 – May 2015) 117 95. Level of significance of the morphometric/meristic characters of M. macrobrachion from Badagry, Lagos and Epe Lagoons 129 96. Level of significance of the morphometric/meristic characters of M. vollenhovenii from Badagry, Lagos and Epe Lagoons 129 97. Level of significance of the egg diameters of M. macrobrachion and M. vollenhovenii from Badagry, Lagos and Epe Lagoons 130 98. Mean ± S.E. of the physico-chemical parameters of the culture media and the natural environment 133 99. Concentration of extracted DNA samples 138 100. Relative absorbance ratio of extracted DNA samples 138 101. Phylogenetic tree of the sequenced data 144 xvi LIST OF PLATES Plate Page 1. Trap (igun) used in the collection of the prawn species 29 2. The dorsal view of male M. macrobrachion 97 3. The dorsal view of female M. macrobrachion 98 4. The dorsal view of male M. vollenhovenii 99 5. The dorsal view of female M. vollenhovenii 100 6A. The ventral view of female M. vollenhovenii with bright colour eggs still attached to the abdomen (immature stage) 110 6B. The ventral view of female M. vollenhovenii with yellow colour eggs still attached to the abdomen (developing stage) 111 6C. The ventral view of female M. vollenhovenii with light brown colour eggs still attached to the abdomen (ripening stage) 112 6D. The ventral view of female M. vollenhovenii with dark brown colour eggs still attached to the abdomen (ripe stage, yet to spawn) 113 6E. The ventral view of female M. vollenhovenii with greyish black colour eggs still attached to the abdomen (ripe stage, ready to spawn) 114 7. Various stages of development of ovary of M. vollenhovenii detached from the abdomen of the female 115 8. Molted female M. vollenhovenii during the culture phase 137 9. Cannibalism observed during the molting stage in M. vollenhovenii 137 10. Mating in M. vollenhovenii 137 11. Electropherogram of extracted DNA samples using phenol-chloroform protocol 139 12. Electropherogram of extracted DNA samples using Norgen kit 139 13. Identified prawns from the DNA barcode 142 xvii LIST OF APPENDICES Appendix Page 1. Preparation of solutions I, II and III for the phenol-chloroform extraction 180 2. Monthly physico-chemical parameters of Badagry Lagoon (June 2013 – May 2015) 181 3. Monthly physico-chemical parameters of Lagos Lagoon (June 2013 – May 2015) 182 4. Monthly physico-chemical parameters of Epe Lagoon (June 2013 – May 2015) 183 5. Air temperature variation in Badagry, Lagos and Epe Lagoons (June 2013 – May 2015) 184 6. Water temperature variation in Badagry, Lagos and Epe Lagoons (June 2013 – May 2015) 185 7. Dissolved oxygen (DO) variation in Badagry, Lagos and Epe Lagoons (June 2013 – May 2015) 186 8. Hydrogen ion concentration (pH) variation in Badagry, Lagos and Epe Lagoons (June 2013 – May 2015) 187 9. Salinity variation in Badagry, Lagos and Epe Lagoons (June 2013 – May 2015) 188 10. Transparency variation in Badagry, Lagos and Epe Lagoons (June 2013 – May 2015) 189 11. Conductivity variation in Badagry, Lagos and Epe Lagoons (June 2013 – May 2015) 190 12. Total length frequency distribution of M. macrobrachion from Badagry, Lagos and Epe Lagoons (June 2013 – May 2015) 191 13. Total length frequency distribution of M. vollenhovenii from Badagry, Lagos and Epe Lagoons (June 2013 – May 2015) 192 14. Carapace length frequency distribution of M. macrobrachion from Badagry, Lagos and Epe Lagoons (June 2013 – May 2015) 193 15. Carapace length frequency distribution of M. vollenhovenii from Badagry, Lagos and Epe Lagoons (June 2013 – May 2015) 193 16. Mean monthly condition factor (K) by sex and size of M. macrobrachion from Badagry Lagoon (June 2013 – May 2015) 194 17. Mean monthly condition factor (K) by sex and size of M. macrobrachion from Lagos Lagoon (June 2013 – May 2015) 195 18. Mean monthly condition factor (K) by sex and size of M. macrobrachion from Epe Lagoon (June 2013 – May 2015) 196 19. Mean monthly condition factor (K) by sex and size of M. vollenhovenii from Badagry Lagoon (June 2013 – May 2015) 197 20. Mean monthly condition factor (K) by sex and size of M. vollenhovenii from Lagos Lagoon (June 2013 – May 2015) 198 21. Mean monthly condition factor (K) by sex and size of M. vollenhovenii from Epe Lagoon (June 2013 – May 2015) 199 22. Stomach contents of M. macrobrachion from Badagry Lagoon (June 2013 – May 2015) 200 23. Stomach contents of M. macrobrachion from Lagos Lagoon (June 2013 – May 2015) 200 24. Stomach contents of M. macrobrachion from Epe Lagoon (June 2013 – May 2015) 201 25. Stomach contents of M. vollenhovenii from Badagry Lagoon (June 2013 – May 2015) 201 26. Stomach contents of M. vollenhovenii from Lagos Lagoon xviii (June 2013 – May 2015) 202 27. Stomach contents of M. vollenhovenii from Epe Lagoon (June 2013 – May 2015) 202 28. Monthly variation in stomach contents of M. macrobrachion and M. vollenhovenii from Badagry Lagoon (June 2013 – May 2015) 203 29. Monthly variation in stomach contents of M. macrobrachion and M. vollenhovenii from Lagos Lagoon (June 2013 – May 2015) 208 30. Monthly variation in stomach contents of M. macrobrachion and M. vollenhovenii from Epe Lagoon (June 2013 – May 2015) 213 31. Monthly variation in sex ratio in M. macrobrachion from Badagry Lagoon (June 2013 – May 2015) 221 32. Monthly variation in sex ratio in M. vollenhovenii from Badagry Lagoon (June 2013 – May 2015) 222 33. Monthly variation in sex ratio in M. macrobrachion from Lagos Lagoon (June 2013 – May 2015) 223 34. Monthly variation in sex ratio in M. vollenhovenii from Lagos Lagoon (June 2013 – May 2015) 224 35. Monthly variation in sex ratio in M. macrobrachion from Epe Lagoon (June 2013 – May 2015) 225 36. Monthly variation in sex ratio in M. vollenhovenii from Epe Lagoon (June 2013 – May 2015) 226 37. Summary of fecundity, egg diameter and GSI estimate for M. macrobrachion from Badagry Lagoon 227 38. Summary of fecundity, egg diameter and GSI estimate for M. macrobrachion from Lagos Lagoon 230 39. Summary of fecundity, egg diameter and GSI estimate for M. macrobrachion from Epe Lagoon 236 40. Summary of fecundity, egg diameter and GSI estimate for M. vollenhovenii from Badagry Lagoon 237 41. Summary of fecundity, egg diameter and GSI estimate for M. vollenhovenii from Lagos Lagoon 238 42. Summary of fecundity, egg diameter and GSI estimate for M. vollenhovenii from Epe Lagoon 239 43. Morphomerics characters of M. macrobrachion from Badagry Lagoon 240 44. Morphomerics characters of M. macrobrachion from Lagos Lagoon 243 45. Morphomerics characters of M. macrobrachion from Epe Lagoon 249 46. Morphomerics characters of M. vollenhovenii from Badagry Lagoon 250 47. Morphomerics characters of M. vollenhovenii from Lagos Lagoon 251 48. Morphomerics characters of M. vollenhovenii from Epe Lagoon 252 xix LIST OF ABBREVIATIONS ANOVA: Analysis of Variance AOAC: Association of Official Analytical Chemists AWG: Average Weight Gain BLAST: Basic Local Alignment Search Tool Bmm: Macrobrachium macrobrachion from Badagry Lagoon Bmv: Macrobrachium vollenhovenii from Badagry Lagoon Bpm: Penaeus monodon from Badagry Lagoon CO1: Cytochrome c Oxidase Sub unit 1 CP: Crude Protein DMRT: Duncan Multiple Range Test EDTA: Ethylene Di-amine Tetra Acetate Emm: Macrobrachium macrobrachion from Epe Lagoon Emv: Macrobrachium vollenhovenii from Epe Lagoon Estr: Strain of Macrobrachium vollenhovenii from Epe Lagoon FCR: Food Conversion Ratio FER: Food Efficiency Ratio Lmm: Macrobrachium macrobrachion from Lagos Lagoon Lmv: Macrobrachium vollenhovenii from Lagos Lagoon Lpa: Parapenaeopsis atlantica from Lagos Lagoon Lpm: Penaeus monodon from Lagos Lagoon NFE: Nitrogen Free Extract PER: Protein Efficiency Ratio SGR: Specific Growth Rate WG: Weight Gain xx DEFINITION OF OPERATIONAL TERMS AWG: The mean weight gain by the experimental prawns in the course of the experimental trial. Binomics: The branch of biology concerned with the relations between organisms and their environment (distribution, ecology and occurrence). Culture: The cultivation of prawn in captivity under defined environmental conditions. DNA barcoding: DNA barcoding, use of short segment of DNA sequence in identifying a species. FCR: The ratio of total dry feed fed to total wet weight gain in prawns. FER: The reciprocal of FCR; ratio of total wet weight gain to total dry feed fed. Lagoon: A body of water with little or no direct connection to the sea, thereby having far less saline conditions. Meristic: The counting of body features such as rostrum teeth. Morphometric: Measurement of body characteristics or features such as length and width. Numerical method: The number of food items of a given type that were found in all the samples examined which is expressed as a % of all food items. Frequency of Occurrence method: The number of stomach sample in which one or more of a given food item is found which is expressed as a % of all excluding empty stomachs examined. PER: The relationship between wet weight gain of prawn and the protein content of feed. Purified diet: A single nutrient diet which has no anti-nutritional factors. Racial: The comparison of the morphometric and meristic features. SGR: The daily percentage increase in weight. xxi ABSTRACT Macrobrachium species are crustaceans found in most inland water and low brackish areas and are of economic importance and possible recruitment into aquaculture. The binomics, culture trial and DNA barcoding of caridean species Brackish water prawn (Macrobrachium macrobrachion) and African River prawn (Macrobrachium vollenhovenii) from Badagry, Lagos and Epe Lagoons, Nigeria were carried out between June 2013 and May 2015. The physico-chemical parameters of the study sites (N 6º 29′ 24.9856″ latitude, E 2º 45′ 52.5347″ longitude – Badagry Lagoon, N 6º 31′ 31.7846″ latitude, E 3º 48′ 23.9768″ longitude – Lagos Lagoon, N 6º 34′ 26.0″ latitude, E 3º 58′ 35.3″ longitude – Epe Lagoon) were measured in- situ. M. macrobrachion and M. vollenhovenii collected for 24 months were analyzed to determine their morphometric characteristics. M. vollenhovenii was reared in triplicate glass tanks with different Crude Protein (CP) levels of purified feed (25%, 30%, 35%, 40% and 45%) and 40% local diet as a control. Molecular technique (DNA barcoding) was used to identify Macrobrachium species found in the study areas. The DNA of the species was extracted using the phenol-chloroform protocol and Norgen tissue kit. Amplification and sequencing of M. macrobrachion and M. vollenhovenii were carried out and further identification of the species was done by comparing with sequenced data in the Genbank. The water temperature for the three lagoons ranged between 21.00 and 32.30 ºC; Dissolved Oxygen ranged from 1.50 to 14.00 mg/L; pH ranged between 5.00 and 9.39; salinity values ranged from 0.00 to 25.00 ‰; water transparency ranged between 17.78 and 175.00 cm and the conductivity ranged between 65.40 and 39600.00 µS/cm. Out of the 4729 specimens collected during the study period, 1770 specimens were from Badagry Lagoon, 924 from Lagos Lagoon and 2035 from Epe Lagoon. The total length (total weight) of M. macrobrachion from the three lagoons ranged from 4.8 cm to 12.8 cm (1.1 – 40.1 g), while M. vollenhovenii ranged from 4.3 cm to 22.6 cm (1.1 – 140.0 g). The condition factor (K) for the combined sexes of M. macrobrachion ranged from 0.3 – 1.4, 0.3 – 1.4 and 0.3 – 1.9 while M. vollenhovenii ranged from 0.4 - 2.7, 0.4 – 3.1 and 0.1 – 2.1 for Badagry, Lagos and Epe Lagoons respectively. The edible parts of M. macrobrachion constituted 44.93 % - 92.04 % of the body weight while that of M. vollenhovenii constituted 40.32 % - 89.80 % in the three lagoons. The stomach contents of M. macrobrachion and M. vollenhovenii from the three lagoons were made up of Bacillariophyta, Chlorophyta, Cyanophyta, Bivalves, Crustaceans, Copepods and detrital materials. The results of the food and feeding habit showed that M. macrobrachion and M. vollenhovenii had preference for detritus. The sex ratio (male: female) for M. macrobrachion and M. vollenhovenii were 1:1.03 and 1:0.21 respectively from Badagry Lagoon, 1:2.60 and 1:1.65 from Lagos Lagoon and 1:0.67 and 1:0.3 from Epe Lagoon. There were significant difference (P < 0.05) in the sex ratio recorded in the three lagoons except for M. macrobrachion from Badagry Lagoon, which showed no significant difference (P > 0.05). At the end of the culture trial, the tank that received 40% CP, compounded from local ingredients performed significantly (P < 0.05) better than the other treatments in terms of average Weight Gain (1.18g), Specific Growth Rate (0.61 %/day), Food Conversion Ratio (5.50), Protein Efficiency Ratio (0.60), Food Efficiency Ratio (0.21) and the survival rate (70.83%). New species of M. asperulum and M. nipponense were observed in Badagry and Epe Lagoons respectively. This study established the occurrence and distribution of M. macrobrachion and M. vollenhovenii from three interconnecting lagoons with the culture potentials of M. vollenhovenii in the laboratory. The phenol-chloroform protocol gave better gel-images than the Norgen tissue kit. The DNA barcode identified that some other species of Macrobrachium were present in Badagry and Epe Lagoons.