PENILAIAN NEMATOD PANAGRELLUS REDIVIVUS SEBAGAI MAKANAN LARVA LAMPAM JAWA, BARBODES GONIONOTUS

UNIVERSITI PUTRA MALAYSIA EVALUATION OF FREE-LIVING NEMATODE PANAGRELLUS REDIVIVUS AS A LIVE FOOD ORGANISM FOR SILVER BARB BARBODES GONIONOTUS LARVAE ABDOLSAMAD JAHANGARD FP 2003 7 EVALUATION OF FREE-LIVING N EMATODE PANAGRELLUS REDIVIVUS ASA LIVE FOOD ORGANISM FOR SILVER BARB BARBODES GONIONOTUS LARVAE By ABDOLSAMADJAHANGARD Thesis Su�mitted to the School of Graduate Studies, Universiti Putra Malaysia, in Fulfilment of the Requirements for the Degree of Doctor of Philosophy March 2003 DEDICATION To my most beloved wife, Ladan, for all her understanding, patience and support during all difficulties and for her technical help during my study To my father and mother, for their support, principle guide and encouragement since my childhood To all scientists and researchers, who have contributed directly and indirectly in the quest of knowledge ii iii Abstract of the thesis presented to the Senate of Universiti Putra Malaysia in fulfilment of the requirement for the degree of Doctor of Philosophy EVALUATION OF FREE-LIVING N EMATODE PANAGRELLUS REDIVIVUS AS A LIVE FOOD ORGANISM FOR SILVER BARB BARBODES GONIONOTUS LARVAE By ABDOLSAMADJAHANGARD March 2003 Chairman: Associate Professor Dr. Mohd. Salleh Kamarudin Faculty: Agriculture A series of experiments were conducted to develop mass production and improve nutritional quality of free-living nematode Panagrellus redivivus. The performance of nematodes P. redivivus produced on different culture media on growth and survival of silver barb Barbodes gonionotus larvae were also studied. Prior to evaluation of P. redivivus as a larval live food organism, a study was conducted on the optimal stocking density of B. gonionotus larvae. Best larval growth and survival were obtained at 10 larvae L-1 for a 1 6-days rearing period. However, a range of 34-65 larvae L-1 was recommended for its commercial hatchery production. A comparative study on performance of nematode, rotifer, Moina and Artemia was carried out. B. gonionotus larvae fed with Artemia and iv nematode exhibited significantly (PO.05) effect on the biochemical composition and nutritional value of P. redivivus for silver barb P. gonionotus larvae. Another study was conducted to determine the effect of lipid enrichment on the production of P. redivivus. The results showed that the best lipid enrichment level for the maximum production of P. redivivus was 3.43%. The extremely low production of P. redivivus in unenriched medium suggested that lipid has an important role in reproduction, metabolism and as the main energy source. The results of a feeding trial showed that nematodes grown in media enriched with> 2.6% oil level were unsuitable for silver barb B. gonionotus larvae. Finally, this study demonstrated that lipid source has an enormous effect on production and fatty acid composition of P. redivivus. The highest production v was achieved when nematodes were grown on sunflower oil enriched medium, followed by those grown in corn, linseed, fish, bleached palm kernel and bleached palm oil enriched respectively. The fish feeding trial, however, demonstrated that silver barb B. gonionotus larvae had a high capability to utilize a wide range of lipid source and dietary fatty acids without any negative effect on its growth and survival. Abstrak tesis yang dikemukakan kepada Senat Universiti Putra Malaysia sebagai memenuhi keperluan untuk ijazah Doktor Falsafah PENILAIAN NEMATOD PANAGRELLUS REDIVIVUS SEBAGAI MAKANAN LARVA LAMPAM JAWA, BARBODES GONIONOTUS Oleh ABDOLSAMADJAHANGARD Mac 2003 Pengerusi: Prof. Madya Dr. Mohd. Salleh Kamarudin Fakulti: Pertanian vi Beberapa eksperimen telah dilakukan bagi membangunkan pengeluaran dan peningkatan mutu pemakanan nematod Panagrellus redivivus secara besarbesaran. Prestasi pengeluaran nematod P. redivivus menggunakan media kultur yang berlainan terhadap pertumbuhan dan kemandirian larva lampam Jawa Barbodes gonionotus juga telah dikaji. Sebelum kajian penilaian P. redivivus dijalankan, satu kajian telah dilakukan untuk menentukan kadar perlepasan optimum larva B. gonionotus. Kadar perlepasan 10 larva L-1 telah memberi pertumbuhan dan kemandirian tertinggi dalam tempoh 16 hari pengkulturan. Kadar perlepasan 34-65 larva L-1 bagaimanapun telah disarankan untuk pengeluaran benih lampam Jawa secara komersial. vii Satu kajian perbandingan prestasi nematod, rotifer, Moina dan Artemia. telah dilakukan. Larva lampam Jawa yang diberi Artemia dan nematod menunjukkan pertumbuhan tertinggi (P<0.05), diikuti oleh ratifer dan Moina. Nematod didapati sesuai sebagai makanan larva lampam Jawa. Kajian selanjutnya menunjukkan 20 nematod mL- 1 adalah kadar pemberian makanan optimum bagi larva lampam Jawa pada kadar perlepasan 10 larva L-1 dalam tempoh 16 hari pengkulturan. Kajian ini juga menunjukkan 8% adalah paras optimum kanji dalam media bagi pengeluaran maksimum P. redivivus. Larva lampam Jawa yang diberi nematod yang dikultur menggunakan media 8% kanji juga menunjukkan pertumbuhan dan kemandirian yang tertinggi. Kajian selanjutnya menunjukkan sumber kanji dalam media mempengaruhi pengeluaran keseluruhan nematod P. redivivus. Kanji kentang merupakan sumber yang terbaik pagi pengeluaran nematod. 8agaimanapun sumber kanji didapati tidak mempengaruhi komposisi kimia dan nilai pemakanan P. redivivus terhadap larva lampam Jawa, B. gonionotus. Kajian selanjutnya telah dilakukan bagi menentukan kesan paras pengkayaan lipid dalam media terhadap pengeluaran P. redivivus. Keputusan menunjukkan paras pengkayaan lipid maksimum bagi pengeluaran P. redivivus adalah 3.43%. Pengeluaran yang amat rendah dalam media tanpa lipid menunjukkkan lipid berperanan penting dalam pembiakan, metabolisme dan sebagai sumber utama tenaga bagi nematod. Kajian juga menunjukkan nematod yang dikultur dalam media yang viii diperkaya dengan lipid melebihi >2.6% didapati tidak sesuai untuk larva B. gonionotus. Kajian juga menunjukkan sumber lipid mempunyai kesan penting terhadap pengeluaran dan komposisi asid lemak P. redivivus. Pengeluaran nematod tertinggi didapati apabila ia dikultur dalam media diperkaya dengan minyak bunga matahari, disusuli oleh minyak jagung, minyak bijan, ikan, isirong kelapa sawit diluntur dan kelapa sawit yang diluntur. Keputusan kajian pemberian makanan menunjukkan larva B. gonionotus memiliki keupayaan untuk menggunakan ranj sumber lipid dan asid lemak dietari yang luas tanpa sebarang kesan pada kemandirian dan pertumbuhannya. ix ACKNOWLEDGEMENTS First and foremost, I want to thank Associate Professor Dr. Mohd. Salleh Kamarudin, chairman of my advisory committee, for providing me with a wonderful opportunity to complete my doctoral studies under his exceptional guidance. Other than the provision of the necessary funding, this work would not have been possible without his patience, constant encouragement, guidance and knowledge. Through frequent meetings and his open door policy, Dr. Kamarudin made an immense contribution to this dissertation and my academic growth, as well as my professional and personal life. My sincerest appreciation is also extended to Dr. Che Roos Sa ad and Associate Professor Dr. Kamaruzaman Sijam, who are members of my supervisory committee, for their constructive suggestions and guidance during the study period. I am also grateful for their willingness to serve on my committee, provide me assistance whenever required, involvement in my oral qual ifying examination, and for reviewing this dissertation. I would like also to extend my most sincere gratitude and thanks to Assoc. Prof. Dr. Razak Alimon who provided me some laboratory facilities and cooperation during the course of the study. I would also like to extend my- appreciation and thanks to: • The European Union for providing the research grant (ERB1C1 8-CT 98- 0333) for this project; x • Dr. Manuele Ricci of BioTecnologie B.T. s.r.l. Pantalla di Todi, 06050 PG, Italy who provided me a training course relevant to my study and for his warm hospitality and continuous technical advices; • Dr. Paymon Roustaian, Dr. Hamid Rezai and Dr. Ehsan Kamrani for their sincere and deep friendship, scientific guidance and technical advices; • My laboratory mates in Aquatic Biotechnology Laboratory, Dr. Annie Christianus, Mannuel, Emil, Reza and Carina for their technical assistance during the conduct of this study; • Professor Abdul Rahman Abd. Razak for his advices and constructive criticism during the major part of my study period; • Mr. Kambiz Shamsi for his critical review and editorial assistance, Alireza for his valuable computer assistance and Yasmin for her support, encouragement, moral, spiritual, faith and concern during my study; • The staff of Aquatic Resources Technology Laboratory, Faculty of Agriculture, UPM especially Mr. Jasni for their valuable assistance; • Mr. Kavoos Kasbi for his financial support in the beginning of my study and Dr. Farshad Shishechian for his technical assistance; • My family and most of all, my brother Ali and sister Mahvash, for their love, understanding and encouragement, and my parents in-law for their moral support towards this achievement; Above all, to the GOD almighty for making this study poss·ible. xi I certify that an Examination Committee met on 19th March 2003 to conduct the final examination of Abdolsamad Jahangard on his Doctor of Philosophy thesis entitled "Evaluation of Free-living Nematode Panagrellus redivivus as a Live Food Organism for Silver Barb 8arbodes gonionofus Larvae" in accordance with Universiti Pertanian Malaysia (Higher Degree) Act 1980 and Universiti Pertanian Malaysia (Higher Degree) Regulations 1981. The Committee recommends that the candidate be awarded the relevant degree. Members of the Examination Committee are as follows: Abdul Razak Alimon, Ph.D. Associate Professor, Faculty of Agriculture, Universiti Putra Malaysia, (Chairman) Mohd. Salleh Kamarudin, Ph.D. Associate Professor, Faculty of Agriculture, Universiti Putra Malaysia, (Member) Che Roos Saad, Ph.D. Lecturer, Faculty of Agriculture, Universiti Putra Malaysia, (Member) Kamaruzaman Sijam, Ph.D. Associate Professor, Faculty of Agriculture, Universiti Putra Malaysia, (Member) Alexendra James Anderson, Ph.D. School of Life Sciences, Queensland University of Technology, Brisbane, Australia, (Independent Examiner) GULAM RUSU Professor I Deputy School of Graduat Studies, Universiti Putra Malaysia Date: 2 1 MAY 2003 xii This thesis submitted to the Senate of Universiti Putra Malaysia has been accepted as fulfilment of the requirement for the degree of Doctor of Philosophy. The members of the Supervisory Committee are as follows: Mohd. Salleh Kamarudin, Ph.D. Associate Professor, Faculty of Agriculture, Universiti Putra Malaysia, (Chairman) Che Roos Saad, Ph.D. Lecturer, Faculty of Agriculture, U niversiti Putra Malaysia, (Member) Kamaruzaman Sijam, Ph.D. Associate Professor, Faculty of Agriculture, Universiti Putra Malaysia, (Member) -e -. i' � AINIIDERIS, Ph.D. Professor I Dean, School of Graduate Studies, Universiti Putra Malaysia, Date: 1 0 JUL 2003 xiii OECLARA TION I hereby declare that this thesis is based on my original work except for quotations and citations, which have been duly acknowledged. I also declare that it has not been previously or concurrently submitted for any other degree at UPM or other institutions. """..,nan Jahangard Date: 5 MAY 2003 xiv TABLE OF CONTENTS DEDiCATION ...... ......... .................................. ................................................... ii ABSTRACT ............. ........... .......................... , ....... ............ ................ ................ iii ABSTRAK ............................. ............ ........ ..... ................... ....... .................. . ..... vi ACKNOWLEDGEMENTS ......... , ......... ... ..................... ........ ............................. ix APPROVAL SHEETS .... .................................................................................. xi DECLARATION .... .......... ............................................................................... xiii TABLE OF CONTENTS ................................................................................ xiv LIST OF TABLES ..... ......................... .......................................................... xviii LIST OF FIGURES ......................................................................................... xx LIST OF ABBREVIATIONS .......................................................................... xxii CHAPTER INTRODUCTION .................. . . ..................... . ................ ............................ 1 Background of the Study ...... . ... .... ... ... ......... ... .......... . ........ ..... .. ..... ....... . ... 1 Statement of Problem . ..................... . . ..................... . .............. ................... 3 The Significance of Study . . ................................... . .......... . .................. . ..... 5 Objectives . . ..... ..... ............. . . ..................................... .............. . ............... . .. 6 II LITERATURE REViEW ...... .................................................. . ................... 8 Free Living Nematode Panagrellus redivivus: Its Biology, Culture and Application ............... . ................................ . ....... ...... . ..... . ........................... 8 Biology ................. . ................................... ................ .................. 8 Growth and Development.. ..... . ..................................... . ............ 9 Population Growth .......... . ....... . ................................................ 10 Temperature ....... . ............. . .......... . . . ............ . ....... . ....... ... . . . . ...... 11 pH ..................... .............................. .............. . .... .. . .......... .. ....... 12 Osmotic Regulations .......................................... ............ . ......... 13 Light .. . .......... . . ...... .................. . . ........... . .............. ...... .............. . . 13 Oxygen .... . ..... . .......................... . ............... . .............................. . 13 Biochemical Composition and Nutritional Value ......... . ...... . .... 14 P. redivivus Culturel Production . . ................... . .... . . . ..... . ........... 15 Application of Nematode P. redivivus in Fish Larviculture ..... 18 Application of P. redivivus in Shrimp Larviculture ..... . . ............ 18 Enrichment with Different Lipid Sources . . .. . ........... . ........ ........ 19 Bioencapsulation of Drugs Using Nematode P. redivivus ....... 20 Silver Barb Barbodes gonionotus .......................................................... 21 Biology, Ecology and Culture .... ................ .. ...... . ..................... 21 Feed and Feeding .................................................................... 22 Larval N utrition ..... .. ............. .. .... ................. . . . . ............. .......... . ................ 23 Common Live Food Organisms for Larviculture ............. . ................... . .. 26 Rotifer ... . ....... ....... . ............. ..... . ..... ........................................... 27 Artemia .... . ................................................... ............................. 28 Moina ................................... . ................. . ........... ....... . .......... . ... 30 "I GENERAL METHODOLOGy ....................... ............... . .......................... 32 Location of the Study ........................ . ...................................... .. . ............ 32 Experimental Fish ................................. . ...................... . ................ . ......... 32 Mass Production of Live Food ........ . ........... ............................................ 33 xv Green Water ......................... ......................... . ........... .............. 34 Rotifer Culture ....................................... ................................... 34 Moina Culture ...... ......... . ........................................................... . 34 Artemia Preparation ......................................................... . ....... 35 Small Scale Nematode Culture (Petri dish Culture) ........... . .... 36 Mass Scale Nematode Culture (Axenic Culture Method) ........ 37 Experimental Set-up ......... ............ ......... ........ ........ ... ............. ..... ............ 39 Water Management ............. ....... . . .......................................................... 39 Samplings ............. ..... ............................................................................. 39 Proximate Analyses of Live Food and Fish Samples ............................ 40 Total Protein ................. ...... ....... ............................................... 40 Total Lipid ......... .......................... . . ....... . ....... . ........................... 40 Fatty Acids ............ . .............................. ............ ........................ 40 Carbohydrate ............... . . ........ .................................................. 41 Statistical Analyses ............ ........ ...... ............................. . ........ ...... .......... 42 IV EFFECTS OF STOCKING DENSITY ON GROWTH, SURVIVAL AND YIELD PRODUCTION OF SILVER BARB BARBODES GONIONOTUS LARVAE ...................... ........................................................... ................ 43 Introduction ..... ..................... ................................................................... 43 Materials and methods .................................................. ......................... 44 Experimental Design ....................................................................... 44 Larval Rearing ....... .......................................................................... 45 Survival, Growth and Yield ............................................................. 45 Water Quality ................... ............................................................... 46 Data Analysis ........................... ............. ............ .............................. 46 Results ................................................................................................... 47 Discussion .............................................................................................. 51 Conclusion .................................................... .......................................... 56 V EFFECTS OF FREE LIVING NEMATODE PANAGRELLUS REDIVIVUS AND SELECTED LIVE FOOD ORGANISMS ON GROWTH AND SURVIVAL OF SILVER BARB BARBODES GONIONOTUS (BLEEKER) LARVAE ................................................................................................. 57 Introduction ........... .................................. ................................................ 57 Materials and Methods ........ ...................................... , ............................ 59 Larval Rearing . . ............................... ................................................ 59 Survival, Growth and Yield ................................................... .......... 60 Biochemical Analysis ...................................................................... 61 Statistical Analysis ....................................... ................................... 61 Results ......................... ...................................... .................................... 62 Water Quality ........................................................... ....................... 62 Growth and Survival ............................. ........................................... 62 Biochemical Composition ................................................................ 65 Discussion .............................................................................................. 66 Conclusion .............................................................................................. 68 VI AN OPTIMAL FEEDING DENSITY OF FREE-LIVING NEMATODE PANAGRELLUS REDIVIVUS FOR SILVER BARB BARBODES GONIONOTUS (BLEEKER) LARVAE .......................... .......... ............... 69 Introduction .......... . .................................................................................. 69 xvi Materials and Methods ....... .................................... .......... ...................... 71 Spawning and I ncubation ............................ .................................... 71 Larval Rearing ................... ............ ............................... ................... 71 Feeding Trial ........ : .......................................................................... 71 Water Quality .............................................................. . . ............. . .... 72 Fish Growth and SurvivaL ........................................ . ...................... 72 Statistical Analysis ....................................................... ...... ............. 73 Results ..................... ....... .............................................. ........ ................. 73 Discussion ...................................... ....................... ............... .................. 77 Conclusion .............................................................................................. 82 VI I EFFECTS OF VARYING STARCH LEVEL ON THE PRODUCTION AND BIOCHEMICAL COMPOSITION OF FREE- LIVING NEMATODE PANAGRELLUS REDIVIVUS AND ITS EVALUATION USING SILVER BARB BARBODES GONIONOTUS (BLEEKER) LARVAE ................... 83 Introduction .................. ........................................................................... 83 Materials and Methods ........................................................................... 84 Nematode Culture ........................................................................... 84 Larval Rearing ............................................. .................................... 85 Water Qual ity ...................................................................... . ........... 86 Survival and Growth ............... ..................... ................... ................. 86 Biochemical Analysis ................ . ..... ... ....... ...... . ........ ........ ............... 87 Statistical Analysis ....... ............................................................ ....... 87 Results .................................................... . .............................................. 88 Nematode Production ..... . ............................................................... 88 Water Quality ....... : .......................................................................... 88 Survival and Growth of Larvae ........ ................. . ...... ................ ....... 89 Biochemical Composition ................................................................ 93 Discussion . . ....................................................... . ...... . . ............................ 97 Conclusion ................................................. ........................................... 100 VI II EFFECTS OF STARCH SOURCES ON THE PRODUCTION OF FREELIVING NEMATODE PANAGRELLUS REDIVI VUS AND ITS NUTRITIONAL VALUE FOR SILVER BARB BARBODES GONIONOTUS LARVAE .................... ................................................. 101 Introduction ................................................................ ; .......................... 101 Materials and Methods ......................................................................... 102 Nematode Culture ....................................................... .................. 102 Larval Rearing ........................................... .................................... 103 Water Quality ....................................... ...................... . ............ ...... 103 Survival and Growth ............................ .............................. ............ 104 Viscosity Measurement. ...................................................... . ......... 104 Biochemical Analysis ........................................ . ..... . ..................... 105 Statistical Analysis . .-...................................................................... 105 Results ........................................................ ......................................... 106 Physical and biochemical properties of starches .......................... 106 Nematode Production and Biochemical Composition .................. 106 Survival and Growth of Fish Larvae ................. ................... .......... 109 Water Quality ...... .......................................................................... 1 12 Biochemical Composition ................................................. . ............ 112 Discussion .... . ..................................................... .................................. 1 13 xvi i Conclusion ............................................................................................ 115 IX EFFECTS OF LIPID ENRICHMENT LEVEL ON PRODUCTI ON AND B IOCHEMICAL COMPISITON OF FREEE-LiVING NEMATODES PANAGRELLUS REDIVIVUS AND ITS EVALUATION USING SILVER BARB BARBODES GONIONOTUS (BLEEKER) LARVAE ..... . ........ ... 1 1 6 Introduction ................... . .... . ...... .............. ......... ... . . .............. . ................. 1 1 6 Materials and Methods ................... . . ..... . . ..... ........ ...... . . ......... .. . . ......... .. 1 1 7 Nematode Culture ............... ............. ........ ..................................... 1 1 7 Larval Rearing .... ............................... .................... ........................ 118 Water Quality . . ........................................... . .................... . ............. 1 1 9 Fish Survival and Growth ............ .......... .................. ...... . ........ ....... 1 1 9 Biochemical Analysis .............. . ..... ... ............................................. 1 20 Statistical Analysis .... .................... . ............................................... 1 20 Results .................................... . .............. . . ..... . .............. ........................ 121 Nematode Production ..... ....... ..................... ............ ... ................... 1 21 Survival and Growth of Fish Larvae ..................................... . ........ 1 22 Water Quality ............ .............. ...................................................... 1 24 Biochemical Composition . ............... ............ . .......................... . ...... 1 26 Discussion ........................................... ..................... ............. . . ............. 131 Conclusion .................... . ................................. . . ................... . . ................ 1 35 X EFFECTS OF LIPID ENRICHMENT USING DIFFERENT OIL SOU RCES ON PRODUCTION AND BIOCHEMICAL COMPOSITION OF FREE-LIVING NEMATODE PANARELLUS REDIVIVUS AND ITS NUTRITIONAL EVALUATION USING SILVER BARB BARBODES GONIONOTUS, (BLEEKER) LARVAE ................................................ 1 36 Introduction ................................................................ ............ ............... 1 36 Materials and Methods ................... ...................................................... 1 38 Nematode Culture ....... .................................................................. 1 38 Larval Rearing ............................... ................................................ 1 39 Chemical analysis .................................................................... . .... 1 40 Statistical Analysis .................................................................. ...... 1 40 Results ................................................................................................. 1 42 Nematode Production ........... ................................................. ....... 1 42 Survival and Growth of Larvae .... ...... ........................................... 1 44 Proximate Composition of Nematodes ......................................... 147 Proximate Composition of Fish ................ ................. .................... 1 47 Fatty Acid Composition of Nematode ..... ...................................... 1 50 Fatty Acid Composition of Fish ........................... . ......................... 1 50 Discussion ................................. ........................................................... 1 53 Conclusion ......................... . ....... ........................................................... 1 58 XI GENERAL DiSCUSSiON ......... ............................................................ 1 59 REFERENCES ............................................................................................ 1 65 APPENDiCES ........................................ . ........... ....... ......... .......................... 1 89 ViTA ..... ............................... ......................................................................... 1 93 xviii LIST OF TABLES Table Page 1. Physical-chemical parameters in rearing tanks of B. gonionotus larvae grown at different stocking density, during 16 days of rearing period . ..................................................................................................... 47 2. Survival, final body weight, final total length, body weight gain, total length gain, specific growth rate and total wet weight yield of B. gonionotus . ............. . ........ .. ..................... ...... ............... ... . ....................... 48 3. Mean total length, total width and biochemical composition of different live food organisms . ............................. .................... ......... ....... 60 4. Survival and growth parameters of silver barb B. gonionotus larvae fed with different live foods for a period of 21-days . .............................. 63 5. Protein and lipid composition of fish larvae B. gonionotus fed on different live food organisms . .................. .......................... ..................... 66 6. Survival, final body weight, final total length and final yield of silver barb B. gonionotus fed on different nematode density ................... ... ... . 74 7. Summary of multiple regression analysis of survival and growth of fish (dependent variables) versus feed concentration, a mmonia and pH in culture tanks. ......... ............. ..... ...... .................. ... ............ ... ... .... ... 77 8. Optimal feeding densities of different live foods used by d ifferent authors for larval rearing of fish ........................... ................................... 81 9. Mean production, multiplication factor and medium cost of nematode P. redivivus grown on varying levels of starch . .......................... ............ 88 1 0. Survival, specific growth rate, total length, body weight, total length gain and weight gain of B. gonionotus larvae fed on nematodes grown on different starch level for a period of 1 6-days . . ............ ..... .. ..... 91 1 1 . Proximate composition (% OW) of P. redivivus cultured on d ifferent levels of starch media . ................. : ............................. : ........ ...... ..... ......... 93 1 2. Proximate composition (% OW) of B. gonionotus larvae fed with nematodes grown on different starch levels . ......................................... 94 13. Proximate composition (% dry matter) and viscosity (x1 03 CPS) of different starch sources . .............................. .......................... ............... 105 14. Mean production, multiplication factor and medium cost of nematodes P. redivivus grown on different starch sources for a period of 1 4-days ................................. ................................................. 1 06 1 5. Proximate composition (% dry matter) of P. redivivus cultured on media of different starch sources . ...................... .................................. 1 08 1 6. Mean survival, total length, body weight, length gain, weight gain and specific growth rate of B. gonionotus larvae fed on nematodes grown on different starch sources for a period of 1 6-days ... ........................... 1 09 xix 17. Proximate composition (% dry matter) of B. gonionotus larvae fed with nematodes grown in different starch sources .. .... . .............. . ...... . .. 111 18. Mean production, multiplication factor and medium cost of nematode P. redivivus grow on various lipid levels for a period of 14-days . ........ 120 19. Survival, growth and yield production of B. gonionotus larvae fed on different level of lipid-enriched nematodes reared for a period of 16 days . ..................................................................................................... 122 20. Biochemical composition of nematode P. redivivus cultured on media of different levels of lipid enrichment.. .................................................. 126 21. Biochemical composition of B. gonionotus larvae fed on nematodes P. redivivus grown on media of different levels of lipid enrichment. .... 127 22. Results of multiple regression analysis between survival and growth of fish (Dependent variables) and dietary nutritional factors ................ 130 23. Iodine value and fatty acid compositions of different lipid sources used in the experiment ......................................................................... 140 24. Mean production, multiplication factor and medium cost for nematode P. redivivus grow in enriched media of various oil sources for a period of 14-days . ............................................................................... 142 25. Multiple regression analysis between production of free-living nematodes Panagrelfus redivivus (dependent variable) and characteristics of lipid sources . ............................................................ 142 26. Survival, mean body length, mean body weight, length gain, weight gain, specific growth rate production of B. gonionotus larvae, fed on nematodes grown on enriched media of different oil sources for a period of 16-days .......... , ....................................................................... 145 28. Proximate composition of B. gonionotus larvae fed on nematodes P. redivivus grown on media enriched with different oil sources .............. 148 29. Fatty acid compositions (percent of total FAME) of nematodes grown in media enriched with different oil sources ......................................... 150 30. Fatty acid composition (percent of total FAME) of fish fed with nematodes grown in enriched media of different oil sources ............... 151 xx LIST OF FIGURES Figure Page 1. Survival of silver barb B. gonionotus larvae reared at different stocking density over the 16-days experimental period . . ....................... 49 2. Changes in a) mean total length and b) mean body weight of B. gonionotus larvae reared in six different stocking densities for 16- days . ....................... .............. ....................... ...................... ...... ............. 50 3. Optimum stocking density of B. gonionotus larvae based on polynomial (second order) regression model (Zeitoun et al., 1976) of larval yield versus stocking density . .... .. ....................... ......... ................. 51 4. Optimum stocking density for hatchery production based on a) acceptable body weight and b) acceptable total length of B. gonionotus larvae . .................. ....................... .. ............... ..... ................... 55 5. Effects of nematodes and selected live food organisms on larval B. gonionotus yield. Means with different alphabetical letter are significantly different (P<0.05) . . ........................................ ...................... 64 6. Total length size frequency histograms of fish larvae fed on different live food organisms . .................................................. .......... .... ............... 65 7. Changes in a) total length b) body weight of silver barb B. gonionotus larvae fed on different nematode densities (nematodes mL·1) over a 16-days feeding trial ............................................. ............. ..... ................ 75 8. Survival of silver barb B. gonionotus larvae fed on different nematode densities (nematodes mL-1) over the 16-days experimental period . . ..... 76 9. Survival of B. gonionotus larvae fed on nematodes from various levels of starch during a 16-days trial... ............... .............. ... . . ................ 89 10. Changes in a) total length and b) total body weight of silver barb B. gonionotus fed on nematodes grown on different levels (%) of starch during a 16-days of feeding . . ... ............................................ ................... 90 11. Relationship between medium starch level and a) nematode carbohydrate content, b) nematode lipid content and c) nematodes energy content. ............ : .................................................. ..... .... .. .... . . ....... 95 12. Relationship between a) nematode lipid content and fish lipid content b) nematode energy content and fish energy content. .............. 96 13. Relationship between carbohydrate content (% DW) of starch sources and nematode production . . ......................... ............................ 107 14. Relationship between starch viscosity of starch sources and nematode production .. .................................... ...................................... 107 15. Changes in a) total body length b) body weight of silver barb B. gonionotus fed on nematodes grown on different starch sources for a 16-days trial. ................. ........ ..... ..... ...................... ..... ........................... 110 1 6. Polynomial (second order) regression model: lipid enrichment level vs P. redivivus multiplication factor for a period of 1 4-days culture xxi period ... ........ . . ...... . ..... ... ........ . ..... . . ... . . .... .... . .. .... ... ....... ... ................. ...... 1 21 1 7. Survival of B. gonionotus larvae fed with different lipid enrichment level of nematodes during a 1 6-days feeding trial. .............................. 1 23 1 8. Changes in a) total length b) body weight of silver barb B. gonionotus fed nematodes grown on different lipid enrichment level during a 1 6- days feeding trial. ................. ................................ ................................ 1 24 1 9. Relationships between medium lipid enrichment and a) nematode lipid content (% DW) b) nematode energy content c) nematode carbohydrate content (% DW) . . ................................................. ........... 1 28 20. Relationships between nematode lipid content and a) fish lipid contents b) fish energy content. ..................... ...................................... 1 29 21 . Optimal lipid enrichment level of P. redivivus for P. gonionotus larvae . .................................. .................... .... ......... ......... ............ ........... 1 33 22. Survival of B. gonionotus larvae fed with nematodes grown in enriched media of different oil sources during a 16 d feeding trial. ..... 1 43 23. Changes in growth a) total length b) total body weight of silver barb B. gonionotus fed on nematode grown in enriched media of different oil sources during a 1 6-days feeding trial . ........................................... 1 44 xxii LIST OF ABBREVIATIONS ANOVA Analysis of Variance AOAC Association of Official Analytical Chemist BW Body weight BSA Bovine Serum Albumin CHCI3 Chloroform CH30H Methanol CRD Complete Randomised Design DHA Decosahexanoeic acid DMRT Duncan Multiple Range Test OW Dry weight DO Dissolved oxygen EFA Essential fatty acid Expt. Experiment FAME Fatty acid methyl esters HCL Hydrochloric acid H2SO4 Sulphuric acid HUFA Highly unsaturated fatty acid M.F. Multiplication factor MSA Methanol sulphuric acid MUFA Mono unsaturated fatty acid N Normality NaCI Sodium chloride NaOH Sodium hydroxide NPK Nitrogen Potassium Phosphorous NPU Net protein utilization RGR Relative growth rate RPM Revolution per minute SAS Statistical analysis system SEAFDEC Southeast Asian Fisheries Development Centre SFA Saturated fatty acid SGR Specific growth rate SEM Standard error of mean TRT Treatment tJm Microgram UPM Universiti Putra Malaysia UV Ultra Violet YSI Yellow spring instrument xxiii 1 CHAPTER I INTRODUCTION Background of the Study Asia has the world's highest production and consumption of aquaculture products (Liao, 1 991 ). Similar to other Asian developing nations, seafood is also one of the major sources of dietary protein for Malaysians. Fish culture was introduced to Malaysia with the immigration of Chinese workers to Malaya. However, the industrial farming of several species of fish and shellfish in this country has recently and rapidly expanded due to technology advancement, government support, tropical condition and sufficient water sources (Endinkeau and Kiew, 1 993). Despite of rapid development in culture techniques, inadequate fry supply coupled with relatively h igh feed prices limits Malaysian aquaculture production (Ang, 1 993). Most of fish and shellfish hatcheries in Malaysia are very dependent on imported expensive Artemia cysts (USD 50-80kg- 1 ) as a main larval food source, which could cost up to 80% of the total production cost per se (Sorgeloos, 1 980). However, Artemia may not always be the best larval food for all species (Jones et a/. , 1 993). Therefore, cheaper alternative live food should be explored. Free-living nematode Panagrellus redivivus with its suitable wide size range, ease of culture, h igh n utritional value and low cost production (Fontaine et al. , 1 982; Kahan and Appel, 1 975; Kahan et al. , 1 980) has been found to be 

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