Citation
Sustainable Feed Production to Support Nile Tilapia (Oreochromis Niloticus) Aquaculture in Haiti

Material Information

Title:
Sustainable Feed Production to Support Nile Tilapia (Oreochromis Niloticus) Aquaculture in Haiti
Creator:
St Martin Francois, Marie Pascale G.
Place of Publication:
[Gainesville, Fla.]
Publisher:
University of Florida
Publication Date:
Language:
english
Physical Description:
1 online resource (113 p.)

Thesis/Dissertation Information

Degree:
Master's ( M.S.)
Degree Grantor:
University of Florida
Degree Disciplines:
Interdisciplinary Ecology
Committee Chair:
Kane, Andrew
Committee Members:
Pine, William
Adesogan, Adegbola T
Graduation Date:
12/15/2012

Subjects

Subjects / Keywords:
Amino acids ( jstor )
Aquaculture ( jstor )
Breadfruits ( jstor )
Cassava ( jstor )
Diet ( jstor )
Experimental diets ( jstor )
Fertilizers ( jstor )
Fish ( jstor )
Food ( jstor )
Tilapia ( jstor )
Interdisciplinary Ecology -- Dissertations, Academic -- UF
aquaculture -- diet -- feeding-trial -- formulation -- haiti -- isonitrogenous -- nutrition -- palatability -- sustainable -- tilapia
Genre:
Electronic Thesis or Dissertation
bibliography ( marcgt )
theses ( marcgt )
government publication (state, provincial, terriorial, dependent) ( marcgt )
Interdisciplinary Ecology thesis, M.S.

Notes

Abstract:
In an effort to address under- and malnourishment in Haiti, feed formulations for Nile tilapia (Oreochromis niloticus) were developed using regionally available and sustainable foodstuffs that are not commonly eaten by people, to support the development of aquaculture-based protein in Haiti. Moringa leaves (Moringa oleifera), spent brewer’s grain (Hordeum vulgare), wet and dried spent brewer’s yeast (Saccharomy cescerevisiae), cassava root (Manihot esculenta), breadfruit (Arthocarpus altilis) and blood meal were examined as ingredients in six feed formulations developed using linear programming. Two experiments with six experimental isonitrogenous diets were conducted under controlled laboratory conditions to examine fish growth, and feed palatability, digestibility and pellet stability. Results from Experiment 1 indicated that tilapia fingerlings had better growth (120% weight gain), feed conversion, efficiency and palatability fed with Diet2 compared with Diet 1 (58% weight gain) after 69 days. A second experiment conducted with the samecohort of fish discerned formulation variables associated with differences observedbetween diets in Experiment 1. New diet formulations in Experiment 2, however,failed to provide positive growth, improved palatability or feed efficiencyafter 24 days. Results indicated that breadfruit and cassava are acceptablebinders for other proteinaceous ingredients, but >5% brewer’s yeast in dietformulations negatively impacts feed palatability.(5% brewer’s yeast in diet formulations negatively impacts feed palatability. Further, feed palatability may vary for fish of different sizes. Ingredients examined in this study provided important utility in formulating sustainable feeds that, along with phytoplankton stimulation, should augment community-based tilapia culture in Haiti. ( en )
General Note:
In the series University of Florida Digital Collections.
General Note:
Includes vita.
Bibliography:
Includes bibliographical references.
Source of Description:
Description based on online resource; title from PDF title page.
Source of Description:
This bibliographic record is available under the Creative Commons CC0 public domain dedication. The University of Florida Libraries, as creator of this bibliographic record, has waived all rights to it worldwide under copyright law, including all related and neighboring rights, to the extent allowed by law.
Thesis:
Thesis (M.S.)--University of Florida, 2012.
Local:
Adviser: Kane, Andrew.
Electronic Access:
RESTRICTED TO UF STUDENTS, STAFF, FACULTY, AND ON-CAMPUS USE UNTIL 2014-12-31
Statement of Responsibility:
by Marie Pascale G. St Martin Francois.

Record Information

Source Institution:
UFRGP
Rights Management:
Applicable rights reserved.
Embargo Date:
12/31/2014
Resource Identifier:
870309390 ( OCLC )
Classification:
LD1780 2012 ( lcc )

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1 SUSTAINABLE FEED PRODUCTION TO SUPPORT NILE TILA PIA ( Oreochromis n iloticus ) AQUACULTURE IN HAITI By MARIE PASCALE G. ST MARTIN FRANOIS A THESIS PRESENTED TO THE GRADUATE SCHOOL OF THE UNIVERSITY OF FLORIDA IN PARTIAL FU LFILLMENT OF THE REQUIREMENTS FOR THE DEGREE OF MASTER OF SCIENCE UNIVERSITY OF FLORIDA 2012

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2 2012 Marie Pascale G. St Martin Franois

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3 To my husband, my daughter and my parents, thank you for all your lov e, support and encouragement

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4 ACKNOWLEDGMENTS First and foremost I want to thank God almighty for his immense love, grace, and fidelity. I give him the honor and glory of this journey. I would l like to thank my supervisory committee members, Dr. Andr ew Kane, Dr. Adegbola Adesogan and Dr. William Pine, for giving me the opportunity to work with them, and for their great contributions to my personal and professional development. I am especially grateful for their guidance, perseverance, and patience. Wi thout Dr commitment to work in Haiti this wor k would have not been possible. I also want to express my gratitude to the USAID/WINNER Program for enabling me to pursue my graduate studies. I extend my thanks to all the IFAS internatio nal program staff for their help; especially Florence Serg ile and Melissa Wokasch O'Hern. I extend my deepest gratitude to the WINNER program staff in Haiti, especially Parnell Dimanche and Marie Claude Vorbe. I also would like to thank ECHO Foundation an d Swamphead Brewery for their support of this project and willingness to supply experimental ingredients. The support and insights of Mr. Mike Picchietti, AquaSafra, is also very much appreciated in supplying healthy tilapia fingerlings for this project. E fforts provided by Ross Brooks are greatly appreciated, and were instrumental in assisting with the care and maintenance of the fish, and collection of the data. I would like to thank my parents, Guy and Mervela Francois, and my husband, Henriot Saint Mart in, sisters and friends for all of their love and support from the distance, for personal support and encouragement during my graduate experience. Finally, I thank all of my friends for their friendship and fun and happy memories.

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5 TABLE OF CONTENTS page ACKNOWLEDGMENTS ................................ ................................ ................................ .. 4 LIST OF TABLES ................................ ................................ ................................ ............ 8 LIST OF FIGURES ................................ ................................ ................................ ........ 11 ABSTRACT ................................ ................................ ................................ ................... 12 CHAPTER 1 OVERVIEW ................................ ................................ ................................ ............ 14 Problem Statement ................................ ................................ ................................ 14 Thesis Rational ................................ ................................ ................................ ....... 15 Objectives ................................ ................................ ................................ ......... 16 Hypothesis ................................ ................................ ................................ ........ 17 2 LITERATURE REVIEW ................................ ................................ .......................... 18 State of t he Country ................................ ................................ ................................ 18 Poverty ................................ ................................ ................................ ............. 18 Hunger Statistic ................................ ................................ ................................ 19 Protein Sources Scarci ty ................................ ................................ .................. 21 Fish as Animal Based Protein ................................ ................................ .......... 23 Fish Consumption ................................ ................................ ............................ 24 Aquacultu re ................................ ................................ ................................ ............. 25 Tilapia Biology and Ecology ................................ ................................ ............. 26 Asia ................................ ................................ ................................ ............ 28 Africa ................................ ................................ ................................ .......... 28 North America and Caribbean ................................ ................................ .... 29 Tilapia Culture in Haiti ................................ ................................ ...................... 29 Successes and Failur es ................................ ................................ ................... 30 Natural Aquatic Resources of Haiti with Potential to Support Aquaculture ....... 33 Nutrient Requirements of Tilapia ................................ ................................ ............ 35 Proteins and Amino Acids ................................ ................................ ................ 35 Energy ................................ ................................ ................................ .............. 36 Lipids and Fatty Acids ................................ ................................ ...................... 37 Carbohydrates ................................ ................................ ................................ .. 37 Vitamins and Minerals ................................ ................................ ...................... 38 Non Conventional Feed Resources Availabl e for Feed Production in Haiti ............ 39 Moringa Leaves ................................ ................................ ................................ 40 Leucena Leaves ................................ ................................ ............................... 41 ................................ ....................... 41 Blood Meal ................................ ................................ ................................ ....... 42

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6 Cassava Root ................................ ................................ ................................ ... 42 Jatropha Meal ................................ ................................ ................................ ... 42 Duckweeds ................................ ................................ ................................ ....... 43 Earthworms ................................ ................................ ................................ ...... 43 Coconut Meat ................................ ................................ ................................ ... 44 Breadfruit ................................ ................................ ................................ .......... 44 Distillery Waste ................................ ................................ ................................ 45 3 DEVELOPMENT OF EXPERIMENTAL F EEDS FOR TILAPIA .............................. 46 Introduction ................................ ................................ ................................ ............. 46 Materials and Methods ................................ ................................ ............................ 47 Identi fication and Availability of Nonconventional Feed Resources in Haiti ...... 47 Proximate and Amin o Acid Composition ................................ .......................... 48 Fee d Formulation ................................ ................................ ............................. 49 Experimental Diet Preparation ................................ ................................ .......... 50 Fee ding Trial Studies ................................ ................................ ....................... 54 Experim ental Design ................................ ................................ ........................ 54 Experimental Parameters ................................ ................................ ................. 56 Growth ................................ ................................ ................................ ....... 56 Palatability ................................ ................................ ................................ .. 57 Digestibility ................................ ................................ ................................ 57 Pellet stability ................................ ................................ ............................. 58 Water Quality ................................ ................................ ................................ .... 58 Dissolved oxygen ................................ ................................ ....................... 58 Ammonia ................................ ................................ ................................ .... 59 Nitrite ................................ ................................ ................................ ......... 59 pH ................................ ................................ ................................ .............. 59 Data Analysis ................................ ................................ ................................ ... 59 Results ................................ ................................ ................................ .................... 60 Identi fication and Availability of Nonconventional Feed Resources in Haiti ...... 60 Proximate and Amino Acid Compos ition ................................ .......................... 60 Feed Formulation ................................ ................................ ............................. 67 Diets Manufactured ................................ ................................ .................... 67 Commercial Fish Feed ................................ ................................ ............... 69 Composition of Zeig ler Bronze Fish Food ................................ .................. 69 Fee ding Trial Studies ................................ ................................ ....................... 69 Experimental parameters (Experiment 1) ................................ ......................... 69 Growth performance an d feed efficiency ................................ ................... 70 Palatability ................................ ................................ ................................ .. 72 Digestibility ................................ ................................ ................................ 72 Pellet stability ................................ ................................ ............................. 73 Water Quality ................................ ................................ ............................. 73 Experimental parameters (Experiment 2) ................................ ......................... 73 Growth performance an d feed efficiency ................................ ................... 74 Palatability ................................ ................................ ................................ .. 76 Pellet stability ................................ ................................ ............................. 77

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7 Water Quality ................................ ................................ ............................. 77 Discussion ................................ ................................ ................................ .............. 77 Experiment 1 ................................ ................................ ................................ .... 78 Experiment 2 ................................ ................................ ................................ .... 81 Alternative NCFRS ................................ ................................ ........................... 82 Antinutritional and anti quality factors in NCFRs ................................ .............. 83 Additional notes on palatability ................................ ................................ ......... 84 Conclusion ................................ ................................ ................................ .............. 85 4 CONCLUSION AND APPLI CATIONS ................................ ................................ .... 86 Feedstuffs Sustainability in Haiti ................................ ................................ ............. 87 Moringa (Benzolive) ................................ ................................ ......................... 87 Breadfruit ................................ ................................ ................................ .......... 88 Cassava ................................ ................................ ................................ ........... 89 Animal Blood ................................ ................................ ................................ .... 90 Brewery and Distiller y Waste ................................ ................................ ............ 90 Why Fish? ................................ ................................ ................................ ............... 92 Nutritional Benefits of Fish ................................ ................................ ................ 92 Characteris tics of Consumption in the Region ................................ .................. 92 Where Do We Go From Here? ................................ ................................ ............... 93 APPENDIX A ADDITIONAL RESOURCES ................................ ................................ ................... 97 B ANALYSIS OF VARIANCE ................................ ................................ ................... 100 LIST OF REFERENCES ................................ ................................ ............................. 103 BIOGRAPHICAL SKETCH ................................ ................................ .......................... 113

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8 LIST OF TABLES Table page 2 1 Comparison of health statistics among under 5 years olds in six countries, based on national surveys ................................ ................................ .................. 20 2 2 Safe level of protein intake for infants, children and adolescent boys and girls .. 22 2 3 Comparison of common human feedstuffs in Haiti. Data presented in gram s per cup ................................ ................................ ................................ ............... 22 2 4 Annual per capita fish consumption in varying countries and regions in 2007 .... 25 2 5 Ongoing aquaculture developm ent projects in Haiti ................................ ........... 30 2 6 Protein requirements of cultured Nile tilapia ................................ ....................... 36 2 7 Essential amino acid requirements of tilapia a s a percentage of dietary protein and of total diet (in parentheses) ................................ ............................ 3 6 2 8 Vitamin requirements (dry matter basis) of Nile tilapia ................................ ....... 39 2 9 Mineral requirements (dry matter basis) of Nile tilapia ................................ ........ 39 2 10 Crude protein contents of some nonconventional feedstuffs .............................. 40 3 1 Methods used for proximate analysis ................................ ................................ 48 3 2 Proximate analysis of ingredients used in the formulation of the diets (a is basis) ................................ ................................ ................................ .................. 49 3 3 Constraints for formulation of protein optimization feed for O. niloticus ............. 50 3 4 Experimental design for used for Experiment 1 and Experiment 2 ..................... 55 3 5 Palatability ranking scheme for evaluating palatability of experimental diets in Experiments 1 and 2. ................................ ................................ .......................... 57 3 6 Availability of nonconventional feedstuff in Haiti ................................ ................. 61 3 7 Proximate and amino acid analysis of experimental diet ingredients from Haiti. ................................ ................................ ................................ ................... 62 3 8 Proximate and amino acid analysis of formulated diets manufactured using sample ingredients from Florida versus Haiti (as fed basis) ............................... 64 3 9 Chemical analyses of diets used in Experiment 1 ................................ .............. 65

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9 3 10 Proximate and amino acid composition of diets in Experiment 2.. ...................... 66 3 11 Proximate analysis of Zeigler fish food (control Diet) ................................ .......... 66 3 12 Composition of formulated diets on dry matter basis. ................................ ......... 67 3 13 Growth performance of tilapia in Experiment 1. ................................ .................. 70 3 14 Stability of extruded feeds in Experiment 1 ................................ ........................ 73 3 15 Feed intake for tilapia in Experiment 2 ................................ ............................... 74 3 16 Pellet stability after immersion in water for 10 minutes. ................................ ...... 77 4 1 Feed and fertilization strategies suggested for optimum yield of tilapia in semi intensive culture systems ................................ ................................ ........... 95 A 1 Theoretical composition of diets using composition of sample ingredients from Haiti and Value from literature ................................ ................................ .... 97 A 2 Water quality parameter throughout the duration of Experiment 1. .................... 98 A 3 Weekly water quality parameters throughout duration Experiment 2 .................. 98 B 1 Analysis of variance of protein intake (PI) of fish fed experimental Diet 1, Diet 2 and Control diet (Diet 3) ................................ ................................ ................ 100 B 2 Analysis of variance of feed conversion ratio (FCR) of fish fed experim ental Diet 1, Diet 2 and Diet 3 ................................ ................................ ................... 100 B 3 Analysis of variance of protein efficiency ratio (PER) of fish fed experimental Diet 1, Diet 2 and Diet 3 ................................ ................................ ................... 100 B 4 Analysis of variance of body weight gain (BWG) of fish fed experimental Diet 1, Diet 2 and Diet 3 ................................ ................................ ........................... 101 B 5 Analysis of variance of growth of fish fed experimental Diet 1, Diet 2 and Diet 3 ................................ ................................ ................................ ....................... 101 B 6 Analysis of variance of pellet stability of experimental Diet 1, Diet 2 and Diet 3 ................................ ................................ ................................ ....................... 101 B 7 Analysi s of variance of palatability of experimental Diet 1, Diet 2 and Diet 3 ... 101 B 8 Analysis of variance of digestibility of experimental Diet 1, Diet 2 and Diet 3 ... 102 B 9 Analysis of variance of palatability of experimental Diet 1, Diet 4, Diet 5, Diet 6 and Diet 7 ................................ ................................ ................................ ...... 102

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10 B 10 Analysis of variance of final weight of fish fed experimental Diet 1, Diet 4, Diet 5, Diet 6 and Diet 7 ................................ ................................ ................... 102

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11 LIST OF FIGURES Figure page 2 1 Prevalence of stunting, underweight, and wasti ng by age in Haiti. .................... 21 2 2 Contribution of fish to animal protein supply worldwide and fish proteins pe r capita per day ................................ ................................ ................................ .... 23 2 3 Value of farmed tilapia (US $ 1000) ................................ ................................ .... 27 2 4 Natural and manmade water bodies that could potentially support freshwater aquaculture in Haiti. ................................ ................................ ............................ 34 3 1 Digestibility ranking scheme based on observations. ................................ ......... 58 3 2 Essential amino acids analyzed fro m Florida d erived ingredients. .................... 62 3 3 Comparison of ingredients derived from Florida versus ingredients derived from Haiti based on crude protein. ................................ ................................ ...... 63 3 4 Samples of feeds used in Experiments 1 and 2. ................................ ................ 68 3 5 Processed ingredients used in the study. ................................ ........................... 68 3 6 Growth dyna mics of tilapia in Experiment 1 ................................ ........................ 71 3 7 Crude protein and weig ht gain changes for tilapia observed in Experiment 1. ... 72 3 8 Palatability and digestibility diets in Experiment 1. ................................ ............. 73 3 9 Change in tilapia body weight observed in Experiment 2. ................................ .. 75 3 10 Percent weight change in medium and large fish after 24 days in Experiment 2. ................................ ................................ ................................ ........................ 75 3 11 Palatability of diets in Experiment 2. ................................ ................................ ... 76 4 1 Moringa seedlings. ................................ ................................ ............................. 88 4 2 Breadfruit growing in Haiti. ................................ ................................ ................. 89 4 3 Cassava plant showing leaves and roots. ................................ ......................... 90 A 1 Marketing channel of fish in Haiti ................................ ................................ ........ 98 A 2 Block diagram of the scheme used to manufacture fish feed in this study. ........ 99

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12 Abstract of Thesis Presented to the Graduate School of the University of Florida in Partial Fulfillment of the Requirements for the Degree of Master of Science SUSTAINABLE FEED PRODUCTION TO SUPPORT NILE TILAPIA ( Oreochromis nilotic us ) AQUACULTURE IN HAITI By Marie Pascale G. St Martin Franois December 2012 Chair: Andrew S. Kane Major: Interdisciplinary Ecology In an effort to address under and malnourishment in Haiti, feed formulations for Nile tilapia ( Oreochromis niloticus ) w ere developed using regionally available and sustainable foodstuffs that are not commonly eaten by people, to support the development of aquac ulture based protein in Haiti. Moringa leaves ( Moringa oleifera ), Hordeum vulgare ), wet and dried ( Saccharomyces cerevisiae ), cassava root ( Manihot esculenta ), breadfruit ( Arthocarpus altilis ) and blood meal were examined as ingredients in six feed formulations developed using linear programming. Two experiments with six expe rimental isonitrogen ous diets were conducted under controlled laboratory conditions to examine fish growth, and feed palatability, digestibility and pellet stability. Results from Experiment 1 indicated that tilapia fingerlings had better growth (120% weig ht gain), feed conversion, efficiency and palatability fed with Diet 2 compared with Diet 1 (58% weight gain) after 69 days (P<0.05) A second experiment conducted with the same cohort of fish discerned formulation variables associated with differences obs erved between diets in Experiment 1. New diet formulations in Experiment 2, however, failed to provide positive growth, improved palatability or feed efficiency after 24 days. Results indicated that breadfruit

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13 and cassava are acceptable binders for other p roteinaceous ingredients, but >5% palatability may vary for fish of different sizes. Ingredients examined in this study provided important utility in formulating sustai nable feeds that, along with phytoplankton stimulation, should augment community based tilapia culture in Haiti.

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14 CHAPTER 1 OVERVIEW Problem S tatement The state of food i nsecuri ty in the w orld presents a challenge in the recent decades Under nourishment o r chronic hunger : status of persons, whose food intake regularly provides less than their minimum energy requirements is related to food insecurity. T he percentage of the world's population experiencing hunger is approximately 868 million and the prevalen ce of hunger aro und the globe remains unacceptably high at close to one billion (FAO, 2010) Food insecurity in Haiti is severe and widespread The number of undernourished people is 5.0 million and prevalence of u ndernourishment is 45% (FAO, 2010) Maln utrition is broad term for a range of conditions that hinder good health, caused by inadequate or unbalanced food intake or from poor absorption of food consumed (FAO, 2010) It affects 50% of the Haitian population (WFP, 2008) Under and malnourishment i n Haiti is a pervasive problem plaguing many per i urban and rural communities. Poverty is the underlying cause of these nutritional problems, which is unavailability of protein sources inadequate purchasing powe r, inappropriate utilization of the resources, increases in global food prices, and the recent frequency of natural disasters Lack of animal based protein, particularly for young children is a primary challenge. Haiti has a need to develop sustainable, lo w cost sources of protein for its people, and tilapia aquaculture is a viable option to help meet this need. One of the main constraints to tilapia aquaculture development in Haiti is the lack of adequate feed and fertilizers. Low cost, regionally availabl e, sustainable feeds to promote fish growth are lacking. Ideally ingredients for

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15 such feeds would need to be developed from unconventional sources, i.e., not items that could be used to feed people directly (such as corn, soybean and fish meal). Sustainabl e feed formulations to support tilapia culture in Haiti, and develop ment of tilapia c ulture programs in rural areas might reduce hunger targets. Thesis Rational Much attention has been paid to aquaculture nutrition in recent years. Nevertheless, the challe nge that faces developing countries like Haiti is to figure out the efficient and sustainable ways to support rural aquaculture, including development of sustainable feeds. The most feasible solution is the development of cost effective tilapia feeds using locally available, cheap and nonconventional resources. Non conventional feed resources (NCFRs) are feed resources that are not usually common in the markets and are not the traditional ingredients used for commercial fish feed production (Devendra, 1988; Becker and Makkar, 2001; Madu et al., 2003). NCFRs are credited for being noncompetitive in terms of human consumption, inexpensive to purchase, incorporating by products or waste products from agriculture, farm made feeds and processing industries, servi ng as a form of waste management and enhancing sanitation. Ingredients can be recycled to improve their value if there are economically justifiable and technological means for converting them into useable products. More information has recently become avai lable about how to formulate tilapia feeds such as nutrient requirements, nutrient composition, and digestibility of feed ingredients (Lim and Webster, 2006). Literature suggests that diet formulation and manufacture are fundamentally a compromise between the ideal situations and practical considerations. The perfect feed that meets the nutritional requirements of an animal or

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16 feed may need to be modified so that it can be prepared and be cost effective For example, practical matters such as ingredient pri ces and availability, the ability to pelletize the formulation, and levels of antinutritional factors in certain ingredients are all very important (Hardy and Barrows, 2002). Therefore, specific feed applications are often considered carefully when formula ting feeds for differe nt animals in different regions Considering these issues, the aim of this study is to identify and analyze chemical composition o f some NCFRs available in Haiti use NCFRs such as moringa ( Moringa olifera ) leaves, cassava ( Manihot es culenta ) grain yeast, breadfruit ( Arthocarpus alti lis ) and blood meal as nonconventional protein sources to formulate and manufacture utilitarian fish feed. Results for this study will help incorporating any of this non conventiona l feedstock into fish feed as on feed ingredients in Haiti and therefore help improve current harvest and provide more protein for children. As in initial effort, this project focused on discerning appropriate sustainable NCFRs in Haiti, and conducting con trolled laboratory experiments to learn more about the feasibility of these ingredients in formulations, feed acceptance and performance, and ramifications relative to future, follow up field studies in Haiti. As such, the following objectives have been id entified: Objectives 1. Identify potential fish feed ingredients in Haiti that have the following characteristics: regionally available, sustainable in Haiti, relatively high in crude protein, and not typically consumed by people. 2. Determine proximate and ami no acid composition of ingredients and experimental diet formulations.

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17 3. Formulate experimental diets using linear programming based on proximate and amino acid composition, and produce experimental diets for laboratory testing with fish. 4. Conduct controlled laboratory feed trials to determine effects of formulat ed diets on tilapia. Effects of interest include growth, palatability, feed eff iciency, and feed stability in water With regard to Objective 4, the following specific aims will be examined: 1. Conduct feed trials with experimental isonitrogenous d iets, based on sustainable NCFR s ingredients that are available in Haiti; and 2. Determine outcomes of feed trials with experimental diets based, and discern fish growth, and feed palatability, efficiency and stab ility Hypothesis With regard to Objective 4, the following hypotheses will be examined: 1. Different isonitrogenous feed formulation s do not affect growth and/weight for tilapia fed experimental diets. 2. Differences in diet formulation s do not affect palatabil ity, digestibility, feed performance and stability of experimental diets in the water 3. Addition of breadfruit to experimental diet formulations does not alter palatability.

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18 CHAPTER 2 LITERATURE REVIEW State o f t he Country Haiti is a country of about 28, 000 square kilometers. It takes up the western third of the Caribbean island of Hispaniola; the Dominican Republic occupies the eastern two thirds. Haiti is shaped like a horseshoe. The most recent statistical survey estimates the population of Haiti of ab out 9,801,664 people (Ju ly 2012 est.) (CIA, 2012). The country annually imports 10,000 tons of fish per year valued $10 million while exports are estimated at 500 tons per year (Damais et al., 2007). One of the major obstacles facing Haitian communities is the desperate need for novel animal based protein Fishery and aquaculture is under developed. Problems that aggravate the state of sustainable aquaculture in Haiti include a lack of baseline data describing the state of aquaculture and aquaculture resour ces portraying the range of successes and failures, weak institutional capacity, rudimentary gear, small size and outdated vessels that prevent the exploitation of deep lakes, lack of fingerling production, lack of refrigeration that increases the risk o f losses at all levels of the value chain, and a paucity of agriculture products where production by products could support alternate animal feed production. In addition to lack of adequate nutrition, natural disasters, poverty, and poor access to educatio n and healthcare for much of the population are among Haiti's most serious problems (CIA, 2012). Poverty The rate of the poverty continues to rise in Haiti (CIA, 2012). U nemployment and underemployment are widespread; more than two thirds of the labor for ce does not have formal jobs Haiti's economy suffered a severe setback in January 2010 when a

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19 7.0 magnitude earthquake destroyed much of its capital city, Port au Prince, and neighboring areas. Already the poorest country in the Western Hemisphere with 80 % of the population living under the poverty line, 54% in abject poverty, and 40.6% (2010 est.) unemployed, the earthquake inflicted $7.8 billion in damage and caused the country's GDP to contract form 2.9% in 2009 to 5.4% in 2010 (CIA, 2012). Two years a fter the January 12, 2010 earthquake struck Haiti, an estimated 500,000 people are still living in about 800 camp sites in earthquake affected areas of Haiti, according to the International Organization for Migration. Two fifths of all Haitians depend on t he agricultural sector, which is ancient without modern means of production, mainly small scale subsistence farming, and remain vulnerable to damage from frequent natural disasters, exacerbated by the country's widespread deforestation. Hunger S tatistic La ck of adequate nu trition for child development is another major problem in rural Haitian communities Dietary protein is notably limiting for children and families in many communities throughout Haiti. In fact, chronic hunger and food insecurity affects a significant portion of the population. Haitian farms produce less than 40% of the malnutrition (FAO, 2010). E dible protein is scarce Despite Haiti's geographic locati on, Haitians consume only four pounds of fish per person per year, seven times less than the Caribbean norm, making fish protein rare An estimated 46% of the population is undernourished, resulting in underweight, stunting, and micronutrient deficiencies (WFP, 2008). Poverty is the underlying cause of these global food prices, and the recent frequency of natural disasters. Haiti holds one of the

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20 lowest life expectancies, and the third highest hunger rate in the world, trailing only Somalia and Afghanistan (UNICEF, 2010) The rural population is most affected by food insecurity because of their dependence on low income generating a griculture. Food insecurity is defined as consuming less than 1900 Kcal per person per day (WFP, 2008). suffer from chronic malnutrition, a direct result of chronic hunger. Ma lnutrition is a major threat to child health in Haiti. As many as 300,000 Haitian children suffer from malnutrition, and up to half of child deaths in the country are caused by malnutrition. One third of 1 year old children have severe growth retardation, 29.7% of children under 5 are stunted, and 18.9% are underweight for age (UNICEF, 2010). Approximately 9% of children under 5 suffer from mod erate to severe wasting (Table 2 1 ). Table 2 1 Comparison of health statistics among under 5 years olds in six cou ntries, based on national surveys (UNICEF, 2010) Country Population size (thousands) Under 5 mortality rate % Under weight % S tunting % Low Birth weight Haiti 9 993 165 18 29 25 Dominican Republic 9 927 27 7 18 11 Jamaica 2 741 24 2 4 12 South Africa 50 133 57 9 24 China 1 341 335 18 4 10 3 United S tates 310 384 8 8 Stunting levels for children under 5 remained relatively constant from 2005 to 2008, at about 24% (Figure 2 1 ) (Cayemittes 2007 ). In 2009, malnutrition contributed to 60% of al l deaths in children in Haiti (UNICEF, 2010).

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21 Figur e 2 1 Prevalence of stunting, underweight, and wasting by age in Haiti. Data represent 5 month moving averages based on 2006 data from Cayemittes 2007 Protein Sources S carcity Haiti currently imports animal feed that is too expensive for small scale rural farmers, effectively creating a large barrier of entry for animal husbandry, such as chicken, beef, goat, tilapia, and dairy farming. Lack of animal based protein is common in the Latin America and C aribbean region. Studies have shown that animal protein supply in this region is low around 40g/day/per capita ( FAO, 2010). Typical diet for rural Haitians consists primarily of rice and beans, and meat is only eaten on Sunday or for big celebrations (e.g ., weddings, Christmas, Easter, funerals). The amount of protein in this meal is clearly under the protein requirement for children or adult (Table 2 2). Protein requirements for human health, however, are much higher than that and more specific (Table 2 3 ).

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22 Table 2 2. Safe level of protein intake for infants, children and adolescent boys and girls Source: WHO, 2002 Boys Girls Age Weight (Kg) (g/Kg/day) (g/day) Weight (kg) (g/kg/day) (g/day) 6 mo 7.8 1.31 10.2 7.2 1.31 9.4 12 mo 10.2 1.14 11.6 9.5 1.14 10.8 18 mo 11.5 1.03 11.8 10.8 1.03 11.1 2 yrs 12.3 0.97 11.9 11.8 0.97 11.4 3 yrs 14.6 0.90 13.1 14.1 0.90 12.7 4 6 yrs 19.7 0.87 17.1 18.6 0.87 16.2 7 10 yrs 28.1 0.92 25.9 28.5 0.92 26.2 11 14 yrs 45.0 0.90 40.5 46.1 0.89 41.0 15 18 y rs 66.5 0.87 57.9 56.4 0.84 47.4 Table 2 3. Comparison of com mon human feedstuffs in Haiti. Data presented in grams per cup. Source: Nicole, 2011 Food items Calories Carbohydrates Protein Fat Rice and b eans 282.0 52.0 8.0 4.0 Black beans sauce 257.0 2 8.2 6.9 12.2 Cook ed r ice 170.0 38.0 4.0 0.0 Cook ed b eef 42.0 0.0 5.0 2.3 In addition, crops such as red and black beans and legumes provide the main source of protein for millions of Haitians. The cull chicken, that has replaced pork as countr y's main source of animal protein, is very expensive. Prices for staple goods, such as rice, corn, beans, and cooking oil, have also increa sed dramatically, 30 40% over one year period (2007 2008) whereas 80% of Haitians live on less than $2 per day; some survive on as little as 44 cents per day ( WFP, 2008)

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23 Fish as Animal Based P rotein Fish is an excellent source of healthy protein and an important source of food for people. quality protein, providing 16 % of animal protein con sumed by the world's population (FAO, 1997). It is particularly important in regions where livestock are relatively scarce (fish supply <10% of animal protein consumed in North America and Europe, but 17% in Africa, 26% in Asia and 22 % in China) (FAO, 2000). About one billion people worldwide rely on fish as their primary source of animal protein (FAO, 2000). The fishery sector plays a key role in food security, not only for subsistence and small scale fishers who rely directly on fish ery for food, incomes and services, but also for consumers who profit from an excellent source of affordable high quality animal protein. Figure 2 2 Contribution of fish to animal protein supply worldwide and fish proteins per capita per day (average 2005 2007) (FAO, 2010)

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24 A 150 g portion of fish provides about 50 60 % of the daily protein requirements for an adult (FAO, 2010). Fish is also a source of essential micronutrients, including vitamins and minerals. In 2007, fish accounted for 15.7 % animal protein and 6.1 % o f all protein consumed (Figure 2 2 ). Globally, fish provide more than 1.5 billion people with almost 20 % of their average per capita intake of animal protein, and 3.0 billion people with 15 % o f such protein (Figure 2 2 ). In terms of a world average, the contribution of fish to calories is rather low at 30.5 calories per capita per day (FAO, 2010). However, it can reach 170 calories per capita per day in countries where there is a lack of altern ative protein food and where a preference for fish has been developed and maintained (e.g., Iceland, Japan (FAO, 2010). Fish C onsumption The fishery sector in Haiti, has more than 50,000 fishermen and fish farmers. Together, they produce approximately 16 ,000 ton metrics of fish per year, only one quarter of which comes from aquaculture (Damais et al., 2007). Despite annual imports of 10,000 metrics tons of fish valued at $10 million, fish consumption is still very low about 2.5 kg / person/year. By compar ison, fish consumption in Jamaica is about 17 kg/person/year (Damais et al. 2007). Fish consumption per capita in Haiti has remained static while global annual per capita fish consumption has grown from an average of 9.9 kg in the 1960s,12.6 kg in the 198 0s, an d up to 17.1 kg in 2009 (Table 2 4 ). The most substantial increases in annual per capita fish consumption occurred in East Asia (from 10.8 kg in 1961 to 30.1 kg in 2007) and North Africa (from 2.8 kg in 1961 to 10.1 kg in 2007).

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25 Table 2 4 Annual per capita fish consumption in varying countries and regions in 2007 (FAO, 2010) Region Fish consumption (kg/person/year) East Asia 30.0 Southeast Asia 29.8 North Africa 10.0 Oceania 25.2 North America 4.0 Europe 22.2 Central America 9.4 Caribbean and South America 9.1 Haiti 2.5 Fish consumption is low in Haiti, primarily because of the high cost of fish products not because of cultural reasons (personal observation). If a low cost cultured fish were available, it is reasonable that the m arket for this product would grow and per capita consumption would increase, particularly for poor people who have limited access to other protein sources. It is therefore important to explore avenues for improving aquaculture in Haiti to help increase the availability of fish as a high quality low cost food source for Haitian people. Aquaculture is becoming very successful all over the world. The following paragraphs present global aquaculture worldwide. Aquaculture Aquaculture, the farming of freshwater, brackish water and marine plants and animals including fish, mollusk, crustaceans, is one of the fastest growing segments of agriculture. It provided 46% of the total worldwide food fish supply in 2008 (FAO, 2010). Over e xploitation of marine and freshwate r fisheries stocks has put escalating pressure on aquaculture production (FAO, 2000). With increase in per capita consumption and the growing human population there is a steadily growing demand for food fish. This has intensified the pressure on the harves ters that, in turn has translated into increased pressure on and overfishing of many commercial fisheries. Fish consumption

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26 increased by 31% from 1990 to 1999 although the supply from marine capture fisheries increased by only 9% (FAO, 1999). Nearly h alf of the known ocean fisheries are completely exploited (FAO, 1999), and 70% are in need of urgent management (MacLennan, 1995). To meet the ever increasing demand for fish, aquaculture has expanded very rapidly and is now the fastest growing food produc tion industry on the world (Tidwell et al 2001). Compared to only 1.3% for capture fisheries, the average annual growth rate of aquaculture was 9% per year from 1970 2000 (Tacon, 2003). The use of aquaculture as a production method has the potential to r elieve some of the overwhelming pressures on natural fish species and can lead to improve d production in developing countries and promote economic and environmental Sustainability. Fifty percent of the total global aquaculture production in 2002 was finfis h (25,728,611 Mt) ( El Sayed 2006) Tilapia B iology and Ecology The genus name Tilapia is a latiniz ation of the word thiape, which means fish in Tswana language (Chapman, 1992) Tilapia includes over 70 species of freshwater fishes within the genera Oreoch romis and Seratherondon (family Cichlidae). Among cultured fish of the world, tilapia rank third in terms of production behind carps and salmonids (first and second, respectively) Nile tilapia ( Oreochromis niloticus ) is, by far, the most important farmed tilapia species in the world. It represents more than 80% of total tilapia production during 1970 2002. Mozambique tilapia ( O. mossambicus ), comes second, with a production of 54,146 Mt in 2002, representing 3.6% of the production of total farmed tilapia. In 2002, production for t hree spotted tilapia ( O andersonii ) blue tilapia ( O. aureus ) redbreast tilapia ( Tilapia rendalii ) and longfin tilapia ( O macrochir )

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27 was respectively 2,700, 1 350, 860 and 210 Mt ( El Sayed 2006) These species are also gainin g some popularity in certain parts of the world. Tilapia are the most adaptable and successful tropic al species worldwide. Tilapia are native to Africa. They were introduced into many tropical, subtropical and temperate regions of the world (Pillar, 1990) Because of their biological and environmental attributes, tilapia is an ideal candidate for aquaculture, especially in developing countries. Those attributes include fast growth, high resistance to stress and disease, tolerance to high stocking densities tolerance to a wide range of environmental factors including poor water quality conditions, easy reproduction in captivity and short generation time, acceptance of artificial feeds immediately after yolk sac absorption and high economic value ( El Sayed 2006) Figure 2 3 Value of farmed tilapia (US $ 1000), 1984 2002 (Fitzsimmons, 2006)

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28 Tilapia farming started as an industry based on fish introduction around the world by development agencies to reduce hunger but has quickly become a highly domestic ated livestock product with sales now exceed ing $2 billion in 2000 (Figure 2 3 ) (Fitzsimmons, 2006). Tilapia is described as the aquatic chicken because its farming can be successful on any scale, from subsistence farming to culture by multinational corpor ations with farms or with processing plants in multiple countries. World production of farmed tilapia was estimated to be around 2,002,087 metric tons (Mt) in 2004 (Fitzsimmons, 2006). Asia Asia produce d 79% of the global farmed tilapia in 2002 which make s it the largest tilapia producer in the world. The growth rate in the production of farmed tilapia during 1950 2002 was among the fastest in the world with an overall average of 20.5% annually ( El Sayed 2006). Over twenty countries in Asia practice tilapi a culture; however China is the largest producer. Without China, Asia would only account for 32.2% of the production of global farmed tilapia. In 2004, China produce d 897,300 M t (Fitzsimmons, 2006). Africa In 2001, t hirty two African countries reported pro duction of farmed tilapia but despite the fact that tilapia is an African fish, farmed tilapia in Africa is relatively new; contribution to world tilapia production is only 12.8%. Most of farmed tilapia production in Africa comes from the brackish water en vironment and Egypt is the major producer. In 2004 tilapia p roduction in Egypt was 220,000 M t ( El Sayed 2006).

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29 North America and Caribbean The first FAO record of tilapia produc tion appeared in 1970 with 200 M t from Mexico. Twenty countries from North Ame rica and the Caribbean reported tilapia production in 2002 but no single co untry dominates the production. Costa Rica, the USA, Mexico and Jamaica accounted respectively for 29.3%, 20%, 16.1%, and 13.3 % and together for 78.7% of tilapia production in 2002. In 2004 Mexico had the largest national produ ction in the western hemisphere, a pproximately 110,000 M t, which represented about 3% of global tilapia production (Fitzsimmons, 200 6 ) Count r ies such as Guatemala, Cuba, Honduras, Panama, Dominican Republic a nd El Salvador start to pay considerable attention to tilapia culture. The production of the rest of North America countries is not significant ( El Sayed 2006). Tilapia Culture in Haiti which is characterized by low input and cost and very low output; and (ii) weak semi intensive aquaculture, characterized by medium input and medium output ( Martinez, 1995) The second category has already received assistance from governments a nd international organization such as FAO, for the formulation of general plans for the development of rural aquaculture in region (Damais et al., 2007). Fish culture was introduced to Haiti in the early 1950s by the FAO technical assistance p lopment of Fish Farming in Haiti For the first two years work concentrated on the construction of nursery and experimental ponds at the Damien Fish Culture Station and at Mariani. Fry production of common carp and Tilapia mossambica started in 1951 and fingerlings of these two species were stocked in rivers, lakes and irrigation canals where they reproduced naturally (FAO, 1984). Over the years several

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30 projects on small scale aquaculture and poultry farming for food secu rity have been executed (Table 2 5 ). Aquaculture production in Haiti is limited, even with perfectly justified objectives such as extensive fish production in small lakes; grow out tilapia in floating cages, aquaculture remain unsuccessful. Since lack of fingerlings and feed represents the major bottlenecks for aquaculture development in Haiti, more attention should be focused on farm made feed and fingerlings production in pond to allow farmers to cover their means of production (FAO, 2010). Table 2 5 Ongoing aquaculture development proje cts in Haiti Aquaculture Development Projects Location Promotion and Development of Freshwater aquaculture Artibonite, North East Petit Freres de l'Incarnation Petite place Cazeau Petit Freres de l'Incarnation Pandiassou Programme National de Lac Collinaires Petite place Cazeau, French Development Agency Hinche Caribbean Harvest Croix des bouquets Double H arvest Croix des bouquets Rural Center for Sustainable Development Croix des bouquets Ponsonde Hatchery Artibonite Fish ministries Chr istianville, Gressier Damien Fish Farm Croix des missions Heartline M inistries Village Theodate Sustainable Aquaculture Program Love a Child Tilapi a project Fond Parisien Morgan Fish farm Ile a vache Lil lavois Groupe d'Appui pour la Reconstruction Effective d'Haiti Grand'Anse Success es and Failure s A program of fingerli ng production of T. mossambica was carried out at the Damien Fish Culture Station during the period 1958 65. During these years the tot al fingerling production, according to the Fisheries Service, was 799, 000 T. mossambica

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31 The average annual productio n was 108 000 carp/year and 114, 000 T. mossambica per year. From 1966 to 1977 production declined and only 616 500 fingerlings were produce d The average production was 37, 0 00 carp fingerlings and only 14, 000 T. mossambica fingerlings per year (FAO, 1984). In 1984, the national production decreased to 110 tons of fish This program was a failure without any follow up and assessment (FAO, 1984 ). According to Randolph ( 1978), 5, 207 ponds were constructed during the years 1958 77 b ut a lack of adequate management and trained personnel in the Fisheries Service brought fish farming more or less to a standstill in 1966. The Fisheries Service estima ted that in 1980 about 500 ponds remained in production in the country. Given this general situation, the aquaculture sub sector of the Ministry of Agriculture Natural Resources and Rural Development (MARNDR), with the support of United Nations Development Program / Food and Agriculture Organization ( PNUD/FAO ) established several aquaculture projects to intro duce T ilapia nilotica in rural communities as the main species and launch fish farming for families (FAO, 1986). The fishing industry in Haiti has disap peared over the past 20 years as a result of poor Because of this, fish farmers have lost their source of income and f ishing villages have declined to a level of abject poverty. Basically the PNUD/FAO project endeavored to create viable aquaculture conditions and to provide new sources of cash income for rural famers. Haiti has a number of successful fish farming initiatives already designed and built by Haitians A Haitian Christian miss etit has excavated small ponds at Pandiassou to raise tilapia to help feed school children and

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32 families at the nutrition centers. Only a few kilometers from Hinche in the Central Plateau of Haiti, Pandiassou, was uninha bited, devastated by land erosion and despair 20 years ago Today, with tilapia culture, the ent ire area has become a natural bounty that feeds thousands of people ( personal observations ). P FI received $25 million from the Haitian government in 2008 to build about 150 catchment ponds throughout the country, and since then, aquaculture production from natural lakes and catchment ponds registered an increase Another successful tilapia culture project is the Caribbean Harvest foundation This project directed by Dr. Val entin Abe is a cooperative project for tilapia f ish f arming established in 2006 to cr eate 500 new jobs in 7 villages around Lake Azuei. Projects are underway to complete this facility in Lake Azeui in 2011 12 and expand to Lake Peligre in the central plateau. For the past 13 years Dr. Valentin Abe, a world renowned agronomist, has worked t In late 2005 he created jobs and provided economic benefits to destitute fishing villages by constructing a modern fish hatchery. This operation and its hatchery, Caribbean Harvest, are located in Croix des Bouquets, a f arming area about 10 miles north of Port au Prince. Caribbean Harvest maintains a hatchery and grow 5 10 grams each), and conducts adult grow out production in the brackish water of Etang Saumtre (Lake Az uei ). Thi s system has been highly successful using hybrid tilapia (name the two strains something x something) that flourishes in brackish water. Another successful tilapia farming operation is FISH Ministries, an American Christian mission based at Christianville a village located in Logne Haiti. They raise

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33 tilapia to feed school children. Protein supplements in the form of eggs or fish provide essential nutrition to over 1,500 school children daily. Because of this high quality nutrition, these students tend t o be healthier than many of their peers and children in other schools, have high attendance records and academ ic scores (Kane unpublished observations ). Natural Aquatic Resources of Haiti with Potential to Support Aquaculture Haiti, a country with a rich h istorical heritage, has considerable potential for marine resources. It has a large potential for aquaculture development. After Cuba, Haiti is the Caribbean country with the most inland water resources in the Caribbean region. Half of these resources are composed of freshwater lakes covering a total area of 11,000 ha. The other half is brackish water lakes, beyond the sea, for a total equivalent. Furthermore, numerous bodies of water including lakes, rivers, reservoirs, catchment ponds exist (Celestin, 200 4). The Republic of Haiti is counted among the territories with renewable resources of marine and inland diverse and varied. These constitute significant potential that can contribute to the overall development of the country (Celestin, 2003). Haiti has a wide variety of ecosystem s with strong potential for developing aquaculture activities. The inland and c oastal waters are very promising environment s for fish culture. Many short rivers with high gradients flow from the mountains to the sea. The largest ri ver, the Artibonite, flows into the Gulf of Gonave. In 1957, a hydro electric dam was built across the dam and it Lake Peligre (3 200 ha). The total area of natural lakes and lagoons is approximately 23 000 ha of which two ar e large lakes; Miraogaone lake, one of the largest freshwater lakes in the Caribbean. Its surface area and maximum depth are respectively 7.06 km 2 and 41 m (Damais et al. 2007)

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34 Figure 2 4. Natural and manmade water bodies that could pote ntially support freshwater aquaculture in Haiti Etang Saumtre (Brackish lake) which is a natural lake of about 16 000 ha was isolated from the Caribbean Sea by tectonic uplift. Because it is fed by springs arising from calcareous rocks, the western part of the lake is slightly saline but the water in the eastern part is fresh (Florvil, 1992). Haiti has about 140 catchment ponds built nationwide over a 3 year period from 2008 to 2011. Those catchment lakes have been built on community land all over the Cen tral Plateau; n ortheast and northwest (Figure 2 4 ).Their areas varied between 1.2 and 15 ha with a capacity of 10,000 to 2,000,000 cubic meters. The country has 1,771 km of coastline and numerous water bodies yet the capture volume was almost unchanged ove r the decades from 1970 to 1980, during

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35 which the annual marine catch fluctuated around 5,000 to 8,000 tons (Celestin, 2004). Finally there are numerous ponds and fish farms in Haiti. Fish farms are operated by stakeholders involved in aquaculture producti missions, researchers, government, scientists, small enterprise owners, farm managers and freshwater fishermen and women. In addition of water bodies Haiti has numerous renewed biological resources. Nutrient Requireme nts of Tilapia Research to improve tilapia feeds and feeding practices has increased over the past two decades. Studies have focused on formulating and manufacturing high quality and nutritionally complete fish feeds due to increased tilapia demand in both domestic and international markets. Research has provided more information on the nutritional requirements of tilapia with respect to protein, energy, lipids, carbohydrates, vitamins, and minerals. Tilapia nutrition deals with the requirement for five cla sses of dietary nutrients, namely, protein, lipids, carbohydrates, vitamins and minerals. Data on nutrient requirements, feed composition and costs are needed for least cost feed formulation. Proteins and Amino Acids Proteins are essentials for the structu re and fu nction of all living organisms. They are the principal constituents of animal tissues and their principal components are amino acids. In fish diets, protein is the most expensive dietary ingredient. The protein requirement of tilapia depends, amon g other things, on fish size or age, proteins sources and the energy content of the diet ( Lim and Webster, 2006 ). Generally, protein requirements decrease with increasing fish size (Table 2 6 ). V alues in Table 2 6 are similar to those from NRC (1993).

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36 Tabl e 2 6 Protein requirements of cultured Nile tilapia Life stage Weight (g) Requirement (%) Reference Fry 0.012 45 El Sayed and Teshima (1992) 0.51 40 Siddiqui et al. (1988) 0.56 35 Teshima et al. (1985) Fingerlings 1.29 40 Teshima et al. (1982) 2.4 35 Toledo et al. (1983) 3.5 30 Wang et al. (1985) Adults 24 28 Wee and Tuan (1988) 40 30 Siddiqui et al. (1988) 45 264 30 Siddiqui et al. (1988) 19 29 Wannigama et al. (1985) Table 2 7 Essential amino acid (EAA) requirements of tilapia as a pe rcentage of dietary protein and of total diet (in parentheses) EAA Requirements O. niloticus a O. mossambicus b Arginine 4.20 (1.18) 2.82 (1.13) Histidine 1.72 (0.48) 1.05 (0.42) Isoleucine 3.11 (0.87) 2.01 (0.80) Leucine 3.39 (0.95) 3.40 (1.35) Lysine 5.12 (1.43) 3.78 (1.51) Methionine 2.68 (0.75) 0.99 (0.40) Phenylalanine 3.75 (1.05) 2.50 (1.00) Threonine 3.75 (1.05) 2.93 (1.17) Tryptophan 1.00 (0.28) 0.43 (0.17) Valine 2.80 (0.78) 2.20 (0.88) Notes: a Jauncey et al. 1983. b Santiago and Lovell 1 988. Amino acids are divided into two groups: Non essential amino acids (NEAA), which can be synthesized by the organism, in the presence of the right precursors and essential amino acids (EAA) which cannot be synthesized by living organisms therefore must be provided in the diets. Tilapia requires the same essential amino acids (arginine, histidine, isoleucine, leucine, lysine, methionine, phenylalanine, threonine, tryptophan and valine) as 7). Energy Energy intake is a basi c nutritional requirement because maintenance of life processes takes priority over growth and other functions (NRC, 1993). Diets should be

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37 balanced to maximize the use of protein for growth by providing optimal amounts of energy as carbohydrates and lipid s. The ratio of protein to energy (P:E; mg / Kcal) varies with fish age and size. For tilapia the optimum ratio for growth varies between 68 and 125, depending on species and size (Shiau and Huang, 1990). Lipids and Fatty A cids Studies have shown that tilapi a utilize dietary lipids very efficiently. Tilapia require s diets of about 10 15% lipids for maximum growth performance (Teshima et al. 1985). Dietary lipids provide highly digestible energy to fish and are the only sources of essential fatty acids for the ir normal growth and development (Lim and Webster, 2006). Apart from satisfying the requirement of a fish for essential fatty acids, dietary lipid acts as a source of energy. According to Li et al. (1991), dietary lipids have sparing effect on the utilizat ion of dietary proteins. The level of protein in the diet of Nile tilapia can be reduced from 33.2 to 27.5 % by increasing dietary lipids from 5.7 to 9.4 % and carbohydrate from 31.9 to 36.9 % The sparing effect of dietary protein by increasing dietary lipid s has been also reported by Jauncey (2000). Carbohydrates Tilapia can very efficiently utilize 30 to 40% digestible carbohydrates as a source of starch (Anderson et al., 1984; El Sayed and Garling, 1988). However, tilapia does not have a specific requireme nt for carbohydrate. It is included in their diet because it is cheap and is the most available food source in the world ( El Sayed 2006). Fiber is usually considered indigestible, as tilapia does not possess the required enzymes for fiber digestion. For t his reason, and to attain maximum growth, crude fiber levels in tilapia diets should probably not exceed 5% (Anderson et al., 1984).

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38 Vitamins and Minerals Vitamins and minerals are essential for normal fish metabolism. Vitamin and mineral supplementation in the form of premixes may be beneficial in intensive systems, although most of these requirements are usually met naturally in extensive and semi intensive pond cultures. Vitamins are generally required in small quant ities in animal diets. Table 2 8 give s ranges of vitamin requirements that have been determined for O. niloticu s Because of the limited knowledge and the uncertainty regarding vitamin requirements, it is difficult to make general recommendations as to what the optimal concentrations should b e, but general minimum levels are commonly applied to feeds. However, tilapia is very sensitive to pyridoxin e (vitamin B6) deficiency. Red h ybrid tilapia ( O. mossambicus O. niloticus ) fed a diet deficient in pyridoxine developed abnormal neurological sig ns, mouth lesions, poor growth and high mortality within 2 to 3 weeks (Lim and Webster, 2006). For Mozambique tilapia, the requirement of this vitamin is reported to range from 5.0 to 11.7 mg.kg 1 diet (Oyetano et al. 1985). Vitamin B 12 (Cyanocobalamin) i s synthesized in tilapia gastrointestinal tract by bacteria to meet their metabolic requirement. General mineral requirements for various tilapia species are presented in Table 2 9 Basic knowledge of mineral toxicity and interactions among minerals is ne cessary when supplementation is made. Fish fed magnesium deficien t diets present poor growth, low tissue magnesium concentrations and abnormal tissue mineralization. Watanabe et al. (1988) recommended a dietary level of 12 mg manganese per kg for O. niloti cus Deficiency of this mineral resulted in anorexia, poor growth, loss of equilibrium and high mortality.

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39 Table 2 8 Vitamin requirements (dry matter basis) of Nile tilapia Vitamin Size (g) Requirement (mg/kg of diet) Reference Vitamin B1 (thiamine) 4 Li m et al. (2000) Folic acid 0.5 Lim and Klesius (2001) Vitamin C (ascorbic acid) 0.56 4.5 50 Abdelghany (1996) 1.0 18.0 420 Soliman et al. (1994) Vitamn A (retinol), IU/kg 11.4 33.1 5000 Saleh et al. (1995) Vitamin E (tocopherol) 0.49 7.8 10 Satoh et al. (1987) Table 2 9 Mineral requirements (dry matter basis) of Nile tilapia Minerals Size (g) Requirement Reference Major (g/kg of diet) Phosphorus 6.1 32.0 < 9 Watanabe et al. (1980) Potassium 0.77 3.5 2 3 Shiau and Hsieh (2001) Magnesium 20.0 54.4 0.59 Dabrowska et al. (1989) Trace (mg/kg of diet) Iron 85 (Fe sulfate) Kleemann et al. (2003) 60 (available Fe) Manganese 12 Watanabe et al. (1988) Zinc 3.1 22.1 30.0 Eid and Ghomin (1994) Copper 2.0 3.0 Watanabe et al. (1988) No n Conventional Feed Resources Available for Feed Production in Haiti Developing an animal feed using unconventi on al ingredients requires finding ingredients that meet the nutritional requirements of the animal, but al so are palatable and consumed. These in gredients should also be available in some form year round feedstuff or can be processed to create a stable feed that can be preserved and used when the ingredients are not available. Ideally these ingr e dients also will not represent human f ood items such as corn or grain that could be consumed by people. Finally these ingredients should be extremely low cost to procure, readily available, and sustainable.

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40 Moringa L eaves Moringa ( Moringa oleifera ) leaves are a potential alternative protein source for Nile t ilapia feed Moringa is a member of the family Moringaceae. This fast growing plant is widely available in the tropics and subtropics with great economic importance for the food and medical industry (Becker and Makkar, 199 9; Foidl et al., 2001). The leaves are rich in carotenoids, ascorbic acid and iron (Siddhuraju and Becker, unpublished data). It contains approximately 25% crude protein (Table 2 10 ). Table 2 10 Crude protein contents of some nonconventional feedstuff s Ingredients Crude Protein Conten t (%) References Blood meal 80 Lovell (1998) Moringa leaf 25 Richter et al. (2002) Brewer's yeast 48 El Sayed (2006) Distillery waste 27 Lovell (1998) Leucena leaf 1 8 Wee and Wang (1987) Jatropha seed 60 63 Akinleye (2011) Breadfruit 15 Oladunjoye e t al.(2010) Cassava root 12 Wee and Ng (1986) Duckweed 45 Essa (1997) Earthworm meal 56 Sogbesan and Ugwumba (2008) Coconut cake 19 24 Hardy and Barrows (2002) Brewer s grain 20 30 El Sayed (2006) Up to 10% inclusion of moringa is recommended for N ile tilapia diet (Ritcher et al., 2002). In view of the favorable amino acid profile of moringa leaves and their wide and ready availability throughout Haiti, Moringa called Benzolive in Haiti can be considered as a potential feed component with high nutri tive value for fish. Moringa has the advantage of growing readily in almost any tropical environment lower than 400 season and as a protein source for tilapia feed would per sist nearly year round If

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41 Moringa can provide the key protein needed as part of developing a feed for tilapia, this perennial drought resistant plant may be ideal. This plan can be grown on non agronomic land, and is already a recognizable tre e though out Haiti. Further culture of this tree could reduce the labor costs associated with obtaining protein for the feed develop ment and helping to provide a sustainable protein source for feed development in Haiti. Leucena L eaves Leucena ( Leucena leuc ocephala ) is a member of the family Fabaceae. Leucena leaves are grown in tropical and subtropical regions in the world. Leaves from this plant are widely used in Haiti as a supplemental protein source for ruminant livestock. Leucena leaves have excellent palatability, digestibility and chemical composition. It contains 17.7% crude protein but has high fiber content approximately 15% (Wee, 1987) The nutritive value of suitably processed Leucaena leaf meal as an alternative protein source has been demonstra ted in study. It was possible to include soaked leaf meal (submerged in water for 48 h and sundried) up to 25% of the total protein with no adverse effects on the growth of the fish (Wee, 1987). S pent B Grain and B Y east S ( Hordeum vulgare east ( Saccharomyces cerevisiae ) are potential protein by product s for fish diet s It is promising especially because they are an alternative protein source, that are by products and that are not good for human consumption grain 32% and 48% crude protein (Desale et al. 2008) They are the solids and liquids left over after fermentation in the brewery process. Desale et al. (2008) showed that 50% of

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42 the fish meal protein in grain Blood M eal Blood meal derived from collecting and drying blood from livestock slaughter is a dark chocolate colored powder with characteristic smell. Its protein content varies from 65 8 5% (Hardy and Borrow, 2002). It is rich in lysine, arginine, methionine, cystine, and leucine but is very poor in isoleucine (NRC, 1993). Amino acid profile revealed that all essential amino acids were present in blood meal in sufficient quantity for amino acid requirement of tilapia Isoleucine was the first limiting amino acid and methionine was the second limiting amino acid for tilapia Blood meal can be used as a supplemental source of protein and can be used to increase the crude protein content of di ets containing cereal grain and plant by products (Khawaja et al. 2007). Previous research (McDonald et al., 1992, and Oyenuga 1968 ; Onwudike 1981) reported similar findings. Cassava R oot Cassava ( Manihot esculenta ) of the Euphorbiaceae family is widely cultivated in the tropics for its starchy storage root that is an excellent source of food and energy. The whole cassava plant may have potential for incorporation into fish feed in this regard, with the roots and leaves used as energy and protein sources, respectively. Ng and Wee (1989 ) reported good growth of Nile tilapia fed cassava based diets without supplementary micronutrients such as vitamin and mineral premixes. Jatropha M eal Jatropha curcas kernel is a multipurpose drought resistant shrub, availab le in Haiti belonging to the family Euphorbiaceae. The species available in Haiti is not edible by people. I t contains phorbol esters a powerful toxin and some others anti nutrients

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43 such as tannins, saponins, phytates, trypsin inhibitor and lectins. Jatro pha contains 60 63% crude protein (CP) (Akinleye et al., 2012). Studies show that j atropha k ernel meal can be used as feed ingredient for Nile tilapia. It can replace up to 62.5% of fishmeal protein in the diet of Nile tilapia without any unfavorable effec ts on the growth performance, nutrient utilization and biochemical activities in the fish and it can be utilized in tilapia diet as a good quality protein source (Akinleye et a l., 2012). However, the use of j atropha in Haiti can be chall enging because deto xication of j atropha meal or cake may be difficult and unsustainable economically. Duckweeds Duckweed (family Araceae) is highly productive with high protein content when cultivated in nutrient rich water (Hillman and Culley, 1978; Culley et al., 1981). D uckweed is readily consumed by a variety of herbivorous fish such as grass carp ( Ctenopharyngodoni della ), silver barb ( Barbus choloensis ) and tilapias ( Oreochromis spp (Zhu, 1999; Fasakin e t al., 2001; Singh et al., 2003 ). It may have potential as fish fo od in the development of low cost aquaculture systems in the tropics. Its crude protein content is high and ranged from 21 to 33% ( Culley et al., 1981) Earthworms Cultured earthworms ( Lumbricus terresstris ) from Lumbricidae family contain low fat and low fibe r concentrations and approximately 63% crude protein (Sogbesan and Ugwumba, 2008). They populate rapidly Earth worms can produce between 3 and 80 colonies per year. Worms take 3 4 months to double the colony. They live 4 to 10 years. Because of that e a rthworms can be excellent food for cultured fish species. In U.S. at the Fish and Wildlife Service's Nati onal Fisheries Research Center, c ultures of the West African nightcrawler and brandl ing worm have been used to feed Gulf sturgeon

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44 ( Acipenser oxyrhynchu s desotoi ). In addition, earthworms have been used alone and in combination with other foods, such as commercial feeds, in diets of other fish species (Willian et al., 1992) Coconut M eat Coconut, Cocos nucifera is a member of the family Arecaceae (palm f amily). The chief constituent of mature coconut meat (the edible white meat of coconut) was carbohydrate, followed by lipid, in contrast to that of the normal mature meat which is an oil source (Mepba, 2003). Dietary fiber of the normal mature meat is comp osed of cellulose, hemicellulose and lignin in proportional amounts. Coconut is rich in vitamins and minerals and its protein content is 21.2 % and can vary with several factors (Hardy and Barrows, 2002). Coconut proteins contain a high percentage of lysine cystine, histidine, arginine, methionine and other essential amino acids. Breadfruit B readfruit ( Arthocarpus altilis ), native to Polynesia, is a large, round, starchy fruit. The plant belongs to the Moraceae family and was introduced to Haiti as a food f or slaves. There are two varieties with either seeded or seedless fruits. The seedless fruits are considered a non conventional food product consumed and mostly cultivated only in the south and southwest of Haiti, where they are Seedless fruits harvested before complete maturity are consumed boiled or deep fried as chips. Some varieties have been studied and are appreciated for their nutritional properties because they are rich in carbohydrates, lipids and proteins. C arbohydrate composition s (starch content and free sugar) of b readfruit have been reported as a good energy source for animals. Studies on the chemical composition for the seedless and seeded varieties have shown a protein content of 15.10 and 1.70 g / 100g, fat 29.0 and 0.30

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45 g / 100 g and moisture 20.20 and 70.80 g/100g respectively (Rincn and Padilla, 2005). The b readfruit tree is widely grown and used in Haiti and its fruit is an important staple crop during the fruiting season, but only one or two varieties are cultiva ted. Breadfruit has tremendous potential for the country as starch and energy sources for fish food Distillery W aste Distillery waste is the primary residue, after removal of the alcohol by distillation, from the yeast or bacterial fermentation of cereals grain or sugar cane. The product contains approximately 27% protein and is highly palatable to fish; however it is relatively low in l ysine (Tom Lovell, 1998).

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46 CHAPTER 3 DEVELOPMENT OF EXPERIMENTAL FEEDS FOR TILAPIA Introduction Aquaculture is the fastes t growing sector of world human food production and has an annual increase of about 10% (FAO, 1997). Worldwide, considerable emphasis has been focused on the use of nonconventional protein sources in order to attain a more economically, environmentally fri endly and viable aquaculture production. Research interest has been directed towards the evaluation and use of nonconventional protein sources, particular from plant products such as seeds, leaves, and agricultural by products (Becker and Makkar, 2001). A quaculture in Haiti is scarce despite available water resources that could support aquaculture development. Water resources for aquaculture in the country are high and are believed to have an excellent potential for rural development and protein supply. Ho wever, the high cost of standard commercial feed imported from United States or Dominican Republic led to the need to identify alternative protein sources for fish feeds and to manufacture the feed locally. In Haiti, some of those feedstuffs are available for increasing animal production. There are large quantities of unutilized agriculture and brewery by products such as crops, vegetables, fruit residues, and need of protein for malnourished children in Haiti a matching increase in non conventional fish feed production is imperative. This project focused on discerning appropriate sustainable NCFRs in Haiti, and conducting controlled laboratory experiments with Nile t ilapia to learn more about the feasibility of these ingredients in formulations, feed acceptance and performance, and

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47 ramifications relative to future, follow up field studies in Haiti As such, the following broad objectives have been identified: 1. Objectiv e 1. Identify potential fish feed ingredients in Haiti that have the following characteristics: regionally available, sustainable in Haiti, relatively high in crude protein, and not typically consumed by people. 2. Objective 2. Determine proximate and amino acid composition of ingredients and experimental diet formulations. 3. Objective 3. Formulate experimental diets using linear programming based on proximate and amino acid composition, and produce experimental diets for laboratory testing with fish. 4. Objectiv e 4. Conduct controlled, laboratory feed trials to determine effects of formulated diets on tilapia. Effects of interest include growth, palatability, feed eff iciency and feed stability in water. With regard to Objective 4, the following specific aims were examined: 1. Conduct feed trials with experimental isonitrogenous diets, based on sustainable NCFRs ingredients that are available in Haiti ; and 2. Determine outcomes of feed trials with experimental diets based, and discern fish growth, palatability, feed effi ciency, and feed stability in water. With regard to Objective 4, the f ollowing hypotheses were tested: 1. Ho 1 : Different isonitrogenous feed formulations do not affect growth and/weight for tilapia fed experimental diets. 2. Ho 2 : Differences in diet formulations do not affect palatability, digestibility, feed performance and stability of experimental diets in the water. 3. Ho 3 : Addition of breadfruit to experimental diet formulations does not alter palatability. Materials and Methods Identification and Availability of Nonconventional Feed Resources in Haiti (Objective 1) Professional networking, including interactions at the World Aquaculture Society Meeting Special Session on Tilapia Cu lture in Haiti; literature review s; and internet search es, were used to id entif y nonconventional feed resource s in Haiti that provide

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48 excellent source s of protein or/and starch. A qualitative survey in the field in Haiti and dialog with academic professionals at the State University of Haiti, business persons and farmers were used t o identify availability of potentially sustainable NCFRs that could be used for fish feed Proximate a nd Amino Acid Composition (Objective 2) Analyses were conducted with potential NCFRs to determine nutritional value Proximate analyses of diet ingredien ts dry matter, crude protein, fat, fiber and ash contents of the NCFR s ingredients and experimental diets formulated with these ingredients, were analyzed prior to conducting feed trials, according to the methods of Association of Official Analytical Chem ists ( AOAC 2005) Amino acid analys e s and proximate analys e s were performed by the University of Missouri Agricultural Experiment Station Chemical Laboratories (ESCL). Methods used are presented in Table 3 1 Table 3 1 Methods used for proximate analysi s Analysis Methods Crude protein Kjeldahl, AOAC Of ficial Method 984.13 (A D) (2005 ) Ash AOAC Official Method 942.05 Crude Fat Ether Extraction, AOAC Official Method 920.39 (A), 2005 Crude Fiber A OAC Official Method 978.10, 2005 Moisture A OAC Official Method 934.01, 2005 vacuum oven Amino acid profile AOAC Official Method 982.3 0 E (a,b,c), chapter. 45.3.05, (2005 ) Crude Protein (CP) is the t otal protein equivalent including nitrogen from both protein and non protein sources. Since proteins contain 1 6% nitrogen on average, the nitrogen value is multiplied by a factor of 6.25 to calculate the crude protein content of the feed. Complete amino acid profile (AAP) was used to determine the amino acid composition of feed ingredients and diets. The samples w ere hydrolyzed for 24 hours

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49 with 6M HCl at 110C and the sulfur containing amino acids were oxidized using perchloric acid before acid hydrolysis. Proximate analyses of feed ingredients used in the experimental diets are presented in Table 3 2 Table 3 2 Proximate analysis of ingredients used in the formulation of the diets (a is basis) Ingredients Crude Protein* % Dry Matter % Crude Fat % Crude Fiber % Ash % NFE % GE Kcal/100g Moringa leaf 27.06 91.79 7.50 6.04 12.19 47.21 445.22 16.13 92.49 3.96 20.09 7.95 51.87 435.15 32.56 96.26 2.85 0.89 7.56 56.14 443.83 Cassava root 1.67 91.39 0.84 1.85 2.17 93.47 403.08 Blood meal 84.52 91.15 0.47 0.31 7.48 7.22 518.50 Breadfruit flour 4.05 89.57 3.04 4.7 7.17 81 .04 401.15 Percentage N X 6.25. Nitrogen free extract (NFE) = 100 (CP + EE + CF + Ash) and Gross energy (GE) were calculated by multiplication of the factors 5.72, 9.5, 4.79, 4.03 kcal GE/100g by percentage of CP, Fat, CF and NFE respectively (Jobling, 1983). Feed Formulation (Objective 3) Feed formulation is required to meet the minimum essential nutrient requirements of the fish. The formulation process demands combination of several suitable feed ingredients to obtain a mixture that is palatable and can be pelletized (Li et al., 2006). Fish feed formulations for this study was based on protein optimization with a fixed formula using linear programing. Linear programming was used as a mathematical model to simultaneously give the solution of a series o f equations. Protein optimizations of the feeds were calculated using Non linear Generalized Reduced Gradient (GRG) algorithm of Solver from Excel add in (Ragsdale, 2011). To formulate the feeds the following information was needed: nutrient concentrations in feedstuffs, table nutrient requirement and chemical composition of ingredients, nutritional and non nutritional restrictions.

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50 The solver application in Microsoft Excel 2010 was utilized to solve for Xi percentage ingredients by maximizing the crude protein of the diet. The formula used for diets in the first feeding trial is the following: The objective function is to Maximize Crude Protein Diet MAX: Z= Z= Crude protein Diets Xi (with i=1) =% Moringa grain Xi (with i=4) =% Cassava roots, Xi (with i=5) = % Blood meal For the second feeding trial, the formula was the same but different ingredient s were added Xi (with i=6) =% Breadfruit M AX: Z= The objective function was subjected to the constraints presented in Table 3 3 Table 3 3 Constraint s for formulation of protein optimization feed for O. niloticus Item Restriction Amount a Moringa (perc ent) Maximum 10 Crude Fat (percent) Maximum 5 Blood meal (percent) Maximum 5 Blood meal (percent) b Maximum 3 Brewer's grain (percent) Minimum 50 Brewer's yeast (percent) Maximum 20 Breadfruit (percent) b Maximum 10 Total % ingredient 100 100 Percenta ge each ingredient >0 Available lysine (g/100g protein) Minimum 3.00 Available methionine (g/100g protein) Minimum 1.46 a Amounts are expressed on an as fed basis. b Amount used in second exper iment. Adapted from Li et al., 2006 Experimental Diet Prepa ration For Experiment 1, t wo diets (Diet 1 and Diet 2) containing different percentages

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51 u sed to make a mash out of the mixed ingredients. Starchy cassava meal was gelatinized, added to the mixture and mixed well. Gloves were used as the mixture was hot. Finally, the feed was extruded through a g arlic press and then cut into pellets by hand. Fe eds were dried in a drying oven at low temperature and collected in plastic bags, sealed and stored in a cool place. The Ziegler Bronze standard commercial fish food was used as a positive control diet (denoted as Diet 3) in Experiment 1. This complex and high quality floating fish food containing 37% crude protein was used to compare diets made with nonconventional proteins sources. Ziegler Bronze is a mixture of feedstuffs, including fish meal, and vitamin and mineral premixes, that provide optimal essent ial nutrients for digestion, growth and energetics. For Experiment 2, Diet 1 from the first experiment was repeated, and was modified slightly to formulate four additional, novel diets (Diets 4, 5, 6 and 7) to attempt to discern why Diet 1 was less palata ble and provided poorer growth than Diet 2 (despite both diets being approximately isonitrogenous ) in Experiment 1 The and to try breadfruit to enhance palatability. Breadfruit was added as a new ingredient to Diet 6 and Diet 7 to replace 50% of the cassava. Breadfruit and cassava were used to make a starchy paste and all other ingredients were added and mixed to form a firm temperature under fans for one day.

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52 I ngredients used for feeds in the two separate experiments were derived from the same Florida sources but different batches, and stored at room temperature until used d yeast which were stored at refrigerated at 4C until used. Next, they were ground using a Willey mill (grinding plate mesh size 1 and 2 inches diameter). After manufacture, dried feed s were crumbled to the appropriate size with a hand grinder. The feeds were stored in sealed bags at room temperature until used in the experiments (Refer to Appendix A 2 for diagram of the scheme used to man ufacture fish feed in this study). Ingredient p reparation Ingredients used to formulate experimental diets were prepared separately before mixing together to manufacture the feed. The pr ep aration process was as follow. Since all feed formulations were dev eloped on a dry weight basis, pilot data was conducted to discern the amount of wet ingredients needed to generate dr y ingredients, as appropriate. Results from these pilot observations indicated the following wet: dry rations: b rewer's grain= 0.1 (1: 10) ; c assava= 0.40 (2: 5) ; b lood= 0.11 (1: 9) ; b rewer's yeast= 0.1 (1: 10) ; and m oringa= 0.1 (1: 10) Blood meal. Fresh blood was collected from a local chicken slaughter house, Pilgrim's Pride Corporation in Live Oak, FL. Four grams of trisodium citrate in 10 ml o f water per liter of blood were added as anticoagulant. Blood was stirred well in order to facilitate evaporation of water and mixed with the citrate. At the lab, all debris were removed, liquid blood with the remaining small amount of the coagulated fract ion were poured into a shallow pan. The blood was weighed and sun dri ed for 3 to 4 days. Finally a W iley mill with a screen of 2 mm mesh size was used to reduce the material to a

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53 uniform particle size. The sun dried blood meal was then stored in tightly cl osed nylon bags at room temperature until used in the experiment. Moringa. Fresh oven dried Moringa oleifera leaves were obtained from Educational Concerns for Hunger Organization (ECHO) Foundation, Fort Pierce, FL. A W iley mill of 2 mm mesh size was used to finely ground the leaves Ground moringa leaf powder was stored in Ziploc bags at room temperature until incorporation into feed formulations. Spent grain and spent yeast were supplied by a local micro brewing company, Swamphead Br ewery, Gainesville, FL. Spent grain used in this study had a protein content of 16.13% and the spent yeast had 32.56% crude protein. They were obtained from the following process. Barley grain and hot water were mixed in a large tank for mashing. Mashing b roke down starch into simple sugars ready for fermentation. The resulting sugars were transferred into a fermentation tank and yeast was added and left for about 2 week to ferment. Once the beer was drained, the solids used wet and fresh for both expe riments. Cassava and breadfruit meal. After washing and peeling manually in bowl, raw cassava roots were soaked in water for 1 hour. The soaking process is done to remove HCN (Amsalu and Esubalew, 2011). Cassava roots were shopped by hand into small sizes, put into boiling water, and simmered for 6 hours to facilitate cooking of starch. Breadfruit followed the same process but was washed instead of soaked in water.

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54 Feed ing Trial Studies (Objective 4) T wo controlled experimental feeding trial s were conducte d at the UF Aquatic Pathobiology Laborator ies to test experimental feeds derived from NCFRs, and discern the feasibility of producing pilot feeds that may be subsequently used for sustainable, community scale tilapia culture in Haiti. The primary purpose f or these experiments was to evaluate tilapia growth, gain experience with formulating experimental diets by studying feed floatation and palatability, and to discern possible interactions with water quality. Experiment 1 provided initial insights into two formulated diets, and the relative growth that may be expected from these diets compared with a commercial fish feed under controlled laboratory conditions. A follow up Experiment 2 was designed to potentially improve feed trial outcomes, based on results from Experiment 1 Seven diets were formulated with varying percentage s of moringa leaves, cassava root, dry and wet for both Experiments 1 and 2 Experimental D esign Three hundred sex reversed male Nile tilapia fingerlings were obtained from a local source, AquaSafra). Fish were laboratory acclimated and observed for three weeks to provide a healthy stable population for the treatments. Three fish diets were used as experimental variables in Experim ent 1: Diet 1 and Diet 2 formulated using feed supplements that are locally available by product ingredients found in Haiti (moringa grain a third diet (Diet 3) of commercial ly (Ziegler Bros, Inc ., Gardners, PA). Diet 3 served as a positive control. For Experiment 2

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55 five diets were used Breadfruit was added as a new ingredient to potentially improve palatability of Diet 1. One hundred and twenty six, laboratory acclimated juvenile tilapia (3.9 0.04 g, Mean SE) were used in Experiment 1. All treatments were conducted in triplicate. Therefore, 14 fish x triplicate tanks x three treatments give a total of 126 fish. Acclimated f ish were randomly sorted into the 9 experimental holding tanks (3 treatments in triplicate) for the feeding trials (Table 3 4 ). Table 3 4 Experimental design for used for Experiment 1 and Experiment 2 # Fish/Replicate Grow O ut Tank Experiment 1 Rep 1 Re p 2 Rep 3 Diet 1 n=14 n=14 n=14 Diet 2 n=14 n=14 n=14 Diet 3 (Control) n=14 n=14 n=14 Experiment 2 Diet 1 n=7 ** n=7 ** n=6 *** Diet 4 n=7 ** n=7 ** n=6 *** Diet 5 n=7** n=7** n=6*** Diet 6 n=7** n=7** n=6*** Diet 7 n=7* n=7** n=6*** denotes small (fingerling) size fish (3g). ** denotes medium size fish (20g). *** denotes large size fish (43 g). Ninety three Nile tilapia were assigned to two blocks based on size and distributed into fifteen tanks for Experiment 2. E ach of the five treatments had 3 replicate tanks each containing 6 or 7 fish. Fish were blocked by size such that one replicate tank for each treatment contained medium fish weighing on average 20.1 0.68 g (mean SE) and two replicate tanks per treatmen t contained large fish weighing 42.5 1. 76 g ( mean SE). Fish within each block were randomly assigned to the five treatments.

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56 photoperiod, under flow through conditions 240 ml/h. Lengths and weights were obtained for each fish during the distribution process at the beginning of the experiments. Fish were fed twice a day at 5% body weight per day during Experiment 1 over 69 days, and once a day at 1% of their body weight per day in Experiment 2 over 24 days. F eed quantities in Experiment 1 were re adjusted according to the change in live body weight every two w eeks. The amount of feed provided during the feeding trials was such that no feed remained at the bottom of the tanks after 3 minutes. Fish were observed daily for appetite and general health. Water quality parameters including temperature, pH, ammonia and nitrite were monitored throughout the acclimation period and feed trials. Fish were weighed after one month, and again at the conclusion of the experiment. At the end of the experiment fish were euthanized by immersion in ice slurry to instantly render th em insensate, followed by cervical transection. Experimental P arameters Growth Average total gain (ATG), relative growth (RG % ), condition factor (K), protein intake (PI %), feed conversion ratio (FCR), and protein efficiency ratio (PER %), were calculate d a ccording to Fawzy et al. (2008). 1. ATG (g/fish) = [average final weight (g) average initial weight (g) 2. RG (%) = [final weight (g) initial weight (g)] x100 / initial weight (g) 3. K= 100x weight (g)/( length) 3 (cm) 4. PI= food supply crude protein (%) 5. FCR= feed intake, dry weight (g) / live weight gain. 6. PER = live weight gain (g) / protein intake (g)

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57 Palatability A ranked scale from 0 to 3 was used to evaluate the relative palatability of the different experimental feeds. A ranked palatability scale for consump tion of the different experimental diets by fish in grow out tanks was developed for Experiment 1 based on initial observations. In Experiment 2, however, with more grow out tanks to observe, the ranking scale was modified to allow a longer o bservation tim e between tanks. Each tank was observed at feeding times, and a rank score was provided on a scale of 0 3). Experiment 2) were averaged to discern possi ble difference s between diets. Table 3 5 illustrates the ranking schemes used in both experiments Table 3 5. Palatability ranking scheme for evaluating palatability of experimental diets in Experiments 1 a nd 2. Experiment 1: Experiment 2: 0: Food was not palatable 0: Food was not palatable 1: Some food remained after 3 min 1: Some food remained after 3 0 min 2: All food consumed within 1 min 2: All food consumed within 1 0 min 3: All food consumed within 10 sec 3: All food consumed within 1 min Digestibility A rank ed digestibility scale was developed to see if there were differences in the relative amount of feed passed to feces from fish in the different treatment tanks. A scale from 0 to 4 was established based on initial observations, where 0 represented the most amount of feces observed (low amount digested), and 4 represented no feces observed (high amount digested and no feces). The digestibilit y scale is presented in Figure 3 1

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58 Ranked digestibility observation scheme for daily digestibility observations are : 0: Lots of feces at tanks bottom after one day of feeding 1: More or less feces after one day of feeding 2: Small amount of feces after one day of feeding 3: Almost no feces after one day of feeding 4: No feces after one day of feeding Figure 3 1. Digest ibility ranking scheme based on observations Pellet stability Experimental diets were weighed; dried for 1 hour in a drying oven at 100 C and reweighed. Sample of 1g of dried feeds were then immersed in 10 ml of tap water for 10 minutes. Water was decante d and the remaining were put on pre weighed aluminu m foil, dried in a drying oven and reweighed. Percent dry matter recovered after immersion in water was measure as stability of feed in the water. Water Quality Dissolved o xygen A YSI model 57 dissolved o xygen meter was used to measure dissolved oxygen (DO) and water temperature in the middle of the water column of the tanks. DO was

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59 measured in mg/L, and data were also expressed in percent saturation. The DO meter was calibrated prior to use each day as p e r manufacturer's instructions. Ammonia Ammonia nitrogen was measured using a Lamotte test kit, model SL NH (3351 01, LaMotte Corporation, 1997). This kit uses an Octa Slide Bar to compare sample color to a color standard after a chemical reaction. Data wa s rec orded as mg/L ammonia nitrogen as per instructions. Toxic unionized ammonia in the sample was calculated using the equation: (mg/L Total NH 3 N kit table value)/100 1.2 = mg/L NH 3 Nitrite Lamotte test kit model SL LNR code 3352 was used to measure nitrite nitrogen in the water. This kit uses an Octa Slide Bar to compare sample color to a color standard after a chemical reaction Data was recorded as mg/L nitrite nitrogen. To convert to nitrite, nitrite nitrogen was multiplied by 3.3, as per instructions. pH An expandable ion analyzer pH meter (Thermo Fisher Scientific, Waltham, MA) using a Ross Sure Flow combination electrode was used for measuring pH. Water samples from fish tanks were obtained and were gently stirred with a magnetic stir bar during pH measurement. Data Analysis Feed performance for each of the 6 experimental diets was assessed by weight gain, FCR PER, feed intake palatability, and dige stibility and pellet stability These variables were analyzed usi ng one way ANOVA using repeated measures. In

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60 Experiment 2, data were blocked by fish size (medium and large). Block effects and interactions effects were considered. Analyses were facilitated using JMP computer software from Statistical Analysis System (SA S Institute, Cary, NC), Proc Mixed Procedure. Standard error of the mean of each parameter was estimated by the sample standard deviation divided by the square root of the sample size. When F test result was significant (P<0.05), the least significant dif ference was calculated to compare tiple range test (Duncan, 1955). Results Identification and Availabil ity of Nonconventional Feed Resources in Haiti (Objective 1) The result of our investigation indicated that nonconvent ional feed resources are readily available in Haiti. M breadfruit and blood meal as nonconventional protein sources in feed formulations are sustainable. Those feedstuffs are underutilized and som e of them are being wasted. Most of them are available at no cost in rural areas in Haiti ( personnel observations) ( Table 3 6 ). Proximate a nd Amino Acid Composition (Objective 2) Non conventional feed resources ye ast, cassava, animal blood, breadfruit were considered because of their promising crude protein content from the literature values. Feedstuff sample from Haiti were shipped to Florida for laboratory analysis. When compared to amino acid requirements for Ni le tilapia, the laboratory results confirmed that they were nutritionally adequate to formulate tilapia feed (Table 3 7 ). Results from laboratory analysis of sample ingredients from Haiti indicated that when compared against amino acid requirement for

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61 Nile tilapia their essential amino acids profiles are excellent and proved they are good n lysine and methionine (Table 3 7 ). Table 3 6 Availability of nonconventional feedstuff in Haiti Ingredients Availability Price (US$) Moringa leaf Environment < 400 m altitude in Haiti, cut back annually to 1 2 meters to regrow pods and leaves Free grain By product of largest brewery in Haiti, BRANA (ton per day) Free yea st By product of largest brewery in Haiti, BRANA (ton per day) Free Cassava roots Produce as feedstock for animal and human food throughout Haiti $0.10/root Animal blood Slaughter house throughout the country, meat provider for market and supermarket Fre e Breadfruit flour Millions of breadfruit tree throughout Haiti, year round production $0.05/fruit However, to be able to conduct this study in the expected timeframe, ingredients from Florida were used. Shipping the amount of ingredients needed from Ha iti would have too expensive. Results from laboratory analysis show ingredients from Florida are great protein sources compared to Amino acid requirements for Nile tilapia except for hich are low in lysine ( Figure 3 2 ). Nonconve ntional feedstuffs sampled from Haiti were analyzed and compared to feedstuffs from Florida. The protein concentrations differed with source as did the feeds formulated with those feedstuffs u sing linear programming ( Figure 3 3 Table 3 8 ). Diets formulate d with sample ingredients from Haiti had higher percentage of crude protein. Those results are theoretical because no diets were manufactured using ingredients from Haiti.

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62 Table 3 7. Proximate and amino acid analysis of experimental diet ingredients from Haiti Diets are compared to optimal growth requirements for Nile tilapia as described by Santiago and Lovell (1988) Nutrients (%) Moringa l eaf waste yeast Distillery waste O. niloticus Crude Protein 28.42 22.53 46.97 24.26 30.00 Dry Matter 8.15 8.66 6.44 8.96 Crude Fat 8.74 7.76 0.66 11.57 10 15 Crude Fiber 5.68 15.86 0.21 3.07 <5.00 Ash 10.19 4.18 5.51 11.11 EAA (g/100g protein) Arginine 4.86 5.06 5.22 3.79 4.20 Histidine 1.79 2.09 2.13 1.81 1.72 Isoleucine 3.94 3.95 4.22 4.78 3.11 Leucine 7.11 7.32 6.26 6.97 3.39 Lysine 3.73 3.99 6.75 6.88 5.12 Methionine 1.41 1.78 1.47 1.53 2.68 Phenylalanine 5.38 5.19 3.64 4.08 3.75 Threonine 3.91 3.46 4.17 4.82 3.75 Tryptophan 1.37 0.93 1.21 0.66 1.00 Valine 4.86 5.24 5.09 5.65 2.80 Figure 3 2 Essential amino acids analyzed from Florida derived ingredients Ingredients are compared with required levels of EAAs for Nile tilapia (red bars) (Santiago and Lovell, 1988)

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63 Figure 3 3 Comparison of ingredients derived from Florida versus ingredients derived from Haiti based on crude protein. Diets were formulated to be approximately isonitrogenous (1 9.10 to 20.07% crude protein and isocaloric (5810.82 to 5881.41) Kcal/kg dry matter (DM). The crude protein content and the gross energy calculated were higher in the control diet respectively 37.05% and 7488.19 Kcal/kg DM. Nitrogen free extract (NFE) was calculated by this formula. NFE= 100 (moisture + crude protein + crude fat + crude fiber + a sh). G ross energy (GE ) was calculated using the values 4.1 5.6 and 9.44 Kcal GE/g DM of carbohydrate protein and fat, res pectively (Jobling, 1983). M etabolizabl e energy (ME ) was calculated using the value of 3.49, 8.1, and 4.5, Kcal/g for carbohydrate, fat, and protein, respectively, according to Pantha (1982).

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64 Table 3 8 Proximate and amino acid analysis of formulated diets manufactured using sample ingredien ts from Florida versus Haiti (as fed basis) Ingredients % Diet 1 Diet 2 Diet 1* Diet 2* O. niloticus 2 Moringa Leaf 10 10 10 10 50 60 50 60 18 3 18 3 2 2 2 2 Cassava roots 20 20 20 20 Dr ied Blood -5 -5 Total 100 100 100 100 Proximate analysis (%) Crude Protein 1 20.07 19.00 23.84 23.27 Moisture 6.64 6.54 8.16 8.50 Crude Fat 2.84 3.31 5.05 5.75 Crude Fiber 12.67 10.53 8.91 10.48 Ash 7. 05 6.80 4.65 4.61 EAA(g/100g protein) Arginine 4.17 4.27 4.42 4.40 4.20 Histidine 2.08 2.27 2.20 2.09 1.72 Isoleucine 3.66 3.13 3.69 3.64 3.11 Leucine 6.92 7.27 6.34 6.64 3.39 Lysine 3.00 3 .49 4.20 3.99 5.12 Methionine 1.46 1.47 1.44 1.46 2.68 Phenylalanine 5.04 5.22 4.94 5.20 3.75 Threonine 3.41 3.36 3.43 3.38 3.75 Tryptophan 1.16 1.24 1.32 1.31 1.00 Valine 4.64 4.33 4.60 4.69 2.80 S ample ingredients c ollected in Haiti 1 Percentage N X 6.25. 2 Essential Amino acids (EAA) requirement for O. niloticus (Santiago and Lovell, 1988) Proximate and amino acid composition of feed ingredients, and diets in Experiments 1 and 2, are presented i n Tables 3 8 and 3 9, respectively. Only 10% of moringa leaves were used in diets because of its anti nutrient content. According to Richter et al., 2002, moringa leaves contain about 0.53% tannins, a substantial

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65 concentration of total phenolics (2.74%), n onhaemolytic saponins (6.38%) and phytic acid (2.25%). Thus, if increased moringa leaf content in feeds, total phenolics, saponin and phytic acid would increase in the diets. Table 3 9 Chemical analyses of diets used in Experiment 1 Nutrient % (as fed bas is) Diet 1 Diet 2 Control Crude p rotein* 19.00 20.07 37.05 Moisture 6.54 6.64 8.13 Crude f at 3.31 2.84 4.00 Crude f iber 10.53 12.67 2.84 Ash 6.80 7.05 7.98 NFE 60.36 57.37 48.13 Calculated energy value GE kcal/kg 5810.82 5881.41 7488. 19 ME Kcal/kg 4384.86 4391.45 5197.51 Percentage N X 6.25. Proximate and amino acid composition of diets for Experiment 2 are pres ented in Table 3 10 Crude fiber content of all five diets are notably high and exceed the requirement of Nile tilapia fo r fiber (< 5%) whereas the levels of crude protein (17.86% All the diets are deficient lysine and methionine based on Nile tilapia requirements described by Lovell (1988). B readfruit replaced half of the cassava in the formulations of Diet 6 and Diet 7. The formulated diets in Experiment 2 were approximately isonitrogenous (17.86% to 19.00% crude protein) and isocaloric (5,668 to 5,743 Kcal/kg dry mat ter). Proximate and amino acid analyses were also conducted for the commercial fish food. Crude protein, moisture, crude fiber, crude fat a nd ash were measured and reported in Table 3 11

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66 Table 3 10 Proximate and amino acid composition of diets in Experiment 2 A mino acid require ment s of Nile tilapia (from Santiago and Lovell, 19 8 8) are provided for comparison. Proximate analysis (as fed basis) Diet 1 Diet 4 Diet 5 Diet 6 Diet 7 O. niloticus Crude Protein % 19.00 19.00 18.03 17.86 18.26 -Moisture % 6.54 6.54 7.46 6.19 6.60 Crude Fat % 3.31 3.31 3.47 3.38 3.39 -Crude Fiber % 10.53 10.53 12.52 11.01 12.15 -Ash % 6.80 6.80 7.16 6.92 6.95 -NFE % 60.36 60.36 58.81 60.83 59.24 -GE kcal/kg 5810.82 5810.82 5690.20 5721.43 5743.26 -ME Kcal/kg 4384.86 4384.86 4322.11 4351.72 4356.06 -EAA (g/100g protein) Arginine 4.06 4.06 4.24 3.99 4.06 4.20 Histidine 1.67 1.67 2.20 1.90 2.02 1.72 Isoleucine 3.59 3.59 3.63 3.42 3.39 3.11 Leucine 5.48 5.48 7.15 6.56 6.79 3.39 Lysine 3.00 3.00 3.83 3.47 3.59 5.12 Met hionine 1.28 1.28 1.47 1.40 1.41 2.68 Phenylalanine 4.19 4.19 5.16 4.50 4.62 3.75 Threonine 2.79 2.79 3.37 3.17 3.13 3.75 Tryptophan 0.34 0.34 1.21 0.92 0.97 1.00 Valine 3.78 3.78 4.67 4.40 4.43 2.80 Table 3 11 Proximate a nalysis of Zeigler fi sh food (control Diet) Proximate and AA analysis Control Diet (g/100 g of Protein) Crude Protein* 37.05 Moisture 8.13 Crude Fat 4.00 Crude Fiber 2.84 Ash 7.98 Arginine 6.48 Histidine 2.40 Isoleucine 3.97 Le ucine 7.07 Lysine 5.29 Methionine 1.65 Phenylalanine 4.24 Threonine 3.59 Tryptophan 1.03 Valine 5.16 Percentage N X 6.25.

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67 Feed Formulation (Objective 3) Two diets (Diet 1 and Diet 2) containing different percentages of moringa, experiment 1. Compositions of the experimenta l d iets are presented in Table 3 12 The primary protein source and the other used less yeast but added blood meal. Ingredients used to manufacture diets in Experiment 2 were from the same sources as ingredients used for diet formulations in Experiment 1, but they were from different batches. Diet 1(uses as a referenc breadfruit) ( Table 3 12 ) Table 3 1 2 Composition of formulated d iets on dry matter basis Diets 1 and 2 were used in Experiment 1 and Diets 1, 4, 5, 6 and 7 were used in Experiment 2 Ingredients (% ) Diet 1 Diet 2 Diet 4 Diet 5 Diet 6 Diet 7 Moringa leaf 10 10 10 10 10 10 Br ewers grain 50 60 50 57 50 57 s yeast 18 3 0 0 0 0 2 2 20 10 20 10 Cassava root 20 20 20 20 10 10 Dried blood 0 5 0 3 0 3 Breadfruit flour 0 0 0 0 10 10 Total 100 100 100 100 100 100 Diets M anufactured Investigated Ingredients were formulated and processed to manufacture experimental diets usin g in this experiments. Figures 3 4 show s A) extruded non floating garlic press extruded feed used in Experiment 1 and B) small crumbl es of meat

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68 grinder ext ruded feed used in Experiment 2 and Figure 3 5 show s processed ingredients used as com ponent of the diets A B Figure 3 4 Sample s o f fe ed s u sed in Experiments 1 and 2 A ) F eed extruded using a garlic press used in Experiment 1. B ) Feed e xtruded using a meat grinder used i n Experiment 2 Photos courtesy Pascale St. Martin and Andrew Kane. A B C D E F Figure 3 5 Processed ingredients us ed in the study. A) Ground B ) Ground st C) Ground moringa leaves D) D ) E ) Wet yeast. F ) Dried blood meal. Photos courtesy Pascale St. Martin and Andrew Kane.

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69 Commercial Fish Feed Empirical evidence and studies have indicated that tilapia farming is expanding and production methods are focused on intensive culture systems with fish stocked at higher densities, fed with nutritionally balanced and high quality feeds ( El Sayed 2006). Commercial fish food containing 37% crude protein was used as a control diet to com pare the formulated diets. A high quality diet was used because the goal of this study is not to make high performance feed capable to compete with the commercial diet. Control diet was used to show maximum growth in fish under laboratory condition when mo ney is not a limiting factor. Composition of Zeigler Bronze Fish Food The commercial fish food used in the experiments consisted of these components: dehulled soybean meal, wheat, wheat middlings, poultry by product meal, hydrolized feather meal, fish meal dehydrated alfalfa meal, soy lecithin, salt manganese proteinate, zinc proteinate, copper proteinate, calcium iodate, iron proteinate, cobalt proteinate, calcium carbonate, sodium selenite, yeast culture, calcium propionate (preservative), vitamin a acet ate, vitamin D3 supplement, di alpha tocopheryl acetate (vitamin E supplement), vitamin B12 supplement, riboflavin, niacin, calcium pantothenate, menadione sodium bisulfite complex (sources of vitamin K activity), folic acid, thiamine mononitrate, pyridoxi ne hydrochloride, biotin, choline, chloride, di methionine, l ascorbyl 2 polyphosphate (sources of vitamin C). Feed ing Trial Studies (Objective 4) Experimental parameters (Experiment 1) This section presents results of use of moringa leaf cassava root, sp grain and yeast, breadfruit and blood meal as nonconventional protein sources

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70 available in H aiti in feed formulations and growth performance o f Nile tilapia ( O. niloticus ). Growth p erformance and f eed e fficiency (Experiment 1) The growth re sponse and feed utilization data of tilapia fed thre e di ets are presented in Table 3 13 The effects of Diet 1, Diet 2 and Diet 3 on fish weight increa sed with time (P<0.001, Figure 3 6 ). There was no feed related mortality observed during the ent ire perio d of the experiment (69 days). There were no differences between the average initial weights of the fish used in the experiment. There were differences between the average final weights of the fish fed on each of the diets, Diet 1, Diet 2 and the c o ntrol d iet (P<0.001) (Table 3 13 ). Fish fed Diet 1 had less weight gain over time compared to fi sh fed Diet 2 (P<0.001, Figure 3 6 ). Table 3 13 Growth performance of tilapia in Experiment 1 Datas are means SE (n=3). Means in a row followed by different lette rs are different ( P< 0.0 01 ) Parameters Diet 1 Diet 2 Control diet (Diet 3) Initial body wt (g) 3.9 0.2 3.9 0.3 3.9 0.1 Final body wt (g) 6.2 0.3a 8.7 0.6b 19.3 0.9c Mean wt gain (g) 2.3 0.2a 4.7 0.4b 15.5 1.0c Rel ative growth (%) 58.6 4.4a 120.3 b 400.7 25.2c Condition factor 1.6 0.9 1.7 1.0 1.7 0.7 Feed intake/fish (g) 15.6 0.0a 15.6 0.0a 15.6 0.0a Protein intake/fish(g) 3.0 0.0a 3.1 0.0a 5.5 0.0b Feed conversion ratio 6.9 0.5a 3.3 0.3b 1.0 0.1c Protein efficiency ratio 0.8 0.1a 1.5 0.1b 2.8 0.2c In turn, fish fed either Diet 1 or Diet 2) had less growth compared to fish fed Diet 3, the control diet. Diet 1 and Diet 2 have simila r crude protein percentage (19.00, 20.7% respectively) but fish fed Diet 2 has higher relative growth (P<0.001) (Figure 3 7). There were no differences between feed intake of the fish fed Diet 1, Diet 2 or Diet 3

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71 (P>0.05). Protein intake for Diet 3, howeve r, was greater than Diet 1 and Diet 2 (P<0.001), while protein intake of fish fed Diets 1 and 2 did not differ significantly (P>0.05). However fish fed Diet 2 used protein more ef ficiently than Diet 1(Table 3 13 ). Figure 3 6 Growth dynamics of tilapia in Experiment 1 Fish were fed two experimental diets Diet 1 and 2, and the control diet, Diet 3, for 69 days Y axis is broken to data from fish fed the positive control diet, but to also maintain focus on data from fish fed experimental Diets 1 and 2. Data are means SE. The feed conversion ratio (FCR) of Diet 1 are lower than FCR of Diet 2 and Diet 3 (P<0.001) (Table 3 13 ). Protein efficiency ratio (PER) was higher for the control diet R was noticeably intake was similar. Condition factor is similar for all fish whi ch means fish were adequately fed and stay in good shape with no regard to the different diets

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72 Figure 3 7 C rude protein and weight gain changes for tilapia observed in Experiment 1 Palatability (Experiment 1) Palatability of Diets 1, 2 and 3 in Experi ment 1 we re different (P< 0.001, Figure 3 8 ). Palatability between diets was greatest in Diet 3 > Diet 2 >> Diet 1. There is no additional effect of time on palatability (P>0.05), and palatability scores were consistent within treatments throughout the exp eriment. Digestibility (Experiment 1) Digestibility ranks for the diets was significantly different (P< 0.0 01 ) between treatments in Experiment 1 Diet 3 (control ) was the most digestible and Diet 1 was the least digestible. Results for dige stibility of d iets in Experiment 1 are shown in Figure 3 8

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73 Figure 3 8 Palatability and digestibility diets in Experiment 1. Data are means SE Different letters above bars indic ate differences between diets (P <0.001) Pellet stability (Experiment 1) Experimental d iets were not different for pellet stability. Percentage recovery of the feeding trial diets after immersion for 10 minutes in unperturbe d w ater are presented in Table 3 14 Table 3 14 Stability of extruded feeds in Experiment 1 Pellet stability Diet 1 Di et 2 Diet 3 % dry matter recovery after immersion for 10 min 84.30 3.44 81.30 0.78 94.25 1.04 Water Quality (Experiment 1) Water quality was monitored throughout feed trials in both Experiment 1 and Experiment 2 For all treatments and replicat 0.1; unionized ammonia was < 0.02 mg/L; nitrite was < 0.1 mg/L; and dissolved oxygen remained above 60% saturation Weekly water quality data are presented in Appendix A 2 Experimental parameters ( Experiment 2 ) Experiment 2 was conducted to revisit feed performance and palatability issues that were observed in Experiment 1. In Experiment 1, Diet 1 had poorer feed performance and palatability compared with Diet 2. In this second experiment, we repeat ed a feed trial with Diet 1, and added 4 modified formulations of that diet to

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74 discern ways to improve diet performance. Results from Experiment 2 show the effects wet versus dried with or without breadfrui t, on growth, palatability, feed efficiency, feed stability of experimental diets. Specific aim 2 was examined for Experiment 2. Growth performance and f e ed e fficiency (Experiment 2) There was no feed related mortality observed during the entire perio d of the experiment (23 days). The feed intake and protein intake of tilapia fed with different diets for the entire experim ent al period is shown in Table 3 15 Table 3 15 Feed intake for tilapia in Experiment 2 Feed intake Diet 1 Diet 4 Diet 5 Diet 6 Diet 7 FI / medium fish (g) 3.18 0.05 3.57 0.10 3.44 0.05 3.17 0.05 3.44 0.05 FI / large fish (g) 7.65 0.00 7.37 0.00 6.80 0.00 7.37 0.00 7.04 0.00 PI / medium fish (g) 0.60 0.07 0.67 0.14 0.62 0.07 0.57 0.07 0.63 0.07 PI / large fish (g) 1.45 0.00 1.40 0.00 1.23 0.00 1.32 0.00 1.29 0.00 All of fish from both medium and large size classes lost weight at the end of Experiment 2 (P<0.001, Figures 3 10 ). Weight loss for medium and large fish was negative but was lower for mediu m compared to large fish (P<0.001) Figure 3 10 ). All diets have the same effects on fish rather they are medium or large, but P = 0.076 is weakly significant. Change in weight blood meal) was observed to be hi gher than change in weight of fish fed the other diets However, there was no statistical difference (P=0.319). Small fish (3.9 0.04 g) fed Diet 1 grew by 58% of their initial weight at the end of 69 days in Experiment 1 while medium (20.1 0.68 g) and large fish (42.5 1.76 g) fed Diet 1 show negative growth. When compare fish fed Diet 1 during 24 days for both experiment, results show that small fish have positive change in weight and medium fish

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75 A B has lower percentage of change and large fish have the lowest percent change in weight. Resu lts are presented in Figure 3 9 Figure 3 9 Change in tilapia body weight observed in Experiment 2. Data for A) medium fish and B) large fish after 24 days are means SE. Figure 3 10 Percent weight change i n medium and large fish after 24 days in Experiment 2. Time adjusted data for small fish from Experiment 1, that were fed Diet 1, are added for comparison between experiments. Data are means SE.

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76 Palatability (Experiment 2) Palatability of diets in Experi ment 2 is presented in F igure 3 11 Both m edium and large size fish fed on the experimental diets were reluctant to consume the feed throughout the experiment Observations included repeated "spitting out" of food. Although all diets were not obviously em braced by fish in any diet treatment, p alatability varied between diets (P<0.001). Further, p alatability of all diets in Experiment 2 was higher for medium fish than for large fish (P<0.001). Diets with reduced (10%) yeast Diet s 5 and 7 were more palatable than diets with higher levels (20%) brewer's yeast ( Diet s 1, 4 and 6). Figure 3 11 Palatability of diets in Experiment 2. Data are means SE. Different letters above bars indicate dif ferences between diets (P<0.001).

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77 All feeds were less pal atable for large fish as compared with medium fish in Experiment 2 (P<0.001). Appendix B 7 provides Analysis of Variance output for palatability. Pellet stability (Experiment 2) The stability of pelleted experimental diets in Experiment 2 was all similar ( P> 0.05). Pelle t stability of Diets 1, 4, 5, 6 and 7 after immersion in water demonstrated 79 91% re covery after 10 minutes ( Table 3 16 ). Table 3 1 6 Pellet stability after immersion in water for 10 minutes Data are means SE (n = 3) Diet1 Diet 4 Diet 5 Diet 6 Diet 7 % dry matter recovery 86.86 3.96 79.31 1.52 81.16 0.41 86.66 6.74 91.29 0.36 Water Quality (Experiment 2) Water quality data is presented in Appendix A 3 Water quality was measured weekly. Results throughout the 4 weeks o f maintenance on the 5 experimental diets indicated that water quality remained within limits for tilapia health, and did not vary between treatments. Temperature was 28 0.3 mg/L, unionized ammonia <0.01 mg/L (below detectable limits), and nitrite <0.05 mg/L (below detectable limits). Discussion Expansion of aquaculture in Haiti has been limited due to the high cost and inconsist ent supply of commercial fish feed s, and the lack of regionally available conventional protein sources such as fishmeal and soybean meal to facilitate local manufacture. Consequently, the need for finding alternative protein sources, especially

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78 those that are by products and not typically not used for human consumption, is needed (Hoffman et al. 1997). The search for nonconventional feed resources (NCFRs) to support tilapia production in Haiti was pursued in this project by conducting feeding experiments with experimental diets formulated using regionally available, sustainable protein resources in Haiti. Diets were formulated using linear programing and controlled feeding studies were conducted in the laboratory t o evaluate experimental diets. Specific ai ms of this project are listed below. 1. Conduct feed trials with experimental isonitrogenous diets, based on sustainable NCFRs ingredients that are available in Haiti; and 2. Determine outcomes of feed trials with experimental diets based, and discern fish growt h, and feed palatability, efficiency and stability. Results from this project indicate that NCFRs such as moringa lea ves cassava may serve as functional feed ingredients in tilapia diet for mu lations. Diets formulated with these ingredients should offer notable aquaculture growth performance, when supplemented with algal production, in fish culture ponds (versus algal stimulation alone) Several observations and notes from data obtained during the feeding trials in Experiments 1 and 2 are worth discussion. Experiment 1 Growth data from Experiment 1 demonstrated that fish fed Diet 2 and Diet 3 (positive control commercial diet) provided better growth than fish fed Diet 1 (P<0.0001). Feed performa nce (P<0.0001) and fish weight gain (P<0.0001) was better for Diet 2 compared with Diet 1, that was likely associated with lower amounts of brewer's yeast and addition of blood meal These data suggest that Diet 2 had better protein utilization

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79 and the fis methionine, which might be the limiting amino acid in a diet with 20% versus 5% complement the lower amount of all growth parameters. Differences between diet composition, in addition to pellet texture and palatability, play an important role in differences seen in fish growth and feed utilization Diet 1 cont ained approximately 19% crude protein and Diet 2 contains 20% crude protein. Although this 1% difference in crude protein may not be insignificant when trying to use perfectly isonitrogenous diets for experimental comparison, this difference likely does no t play a critical role in overall differences obser ved in growth and performance. This is based, in part, on the disproportionate relationship between crude protein in the diets and weight gain (Figure 3 7), as well as differences between protein and weigh t change observed in Experiment 2. Amino acid constituents in the diets are also important. Diet 2 had a more optimal amino acid profile for tilapia, compared with Diet 1, although both Diets 1 and 2 were deficient in lysine and methionine. Generally, lys ine and methionine tend to be lower in plant based sources; and indeed the formulated diets are mostly plant protein based. These effects are underscored by the difference in growth for fish fed Diet 2 versus Diet 3 (commercially formulated control diet). The additional 17% crude protein in Diet 3, and the supplementation with amino acids, vitamins and minerals, should certainly be associated with the better feed performance. The control diet (Diet 3) with 37% crude protein is a n expensive, high performance diet that contains conventional protein

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80 source s including fish meal and soybean meal. It is supplemented with lysine, methionine and vitami ns. The objective of this study was not to achieve the same level of performance as the control diet ; i ts high effic iency in Experiment 1 is normal. In contrast the formulated diets are least cost feed deficient in lysine and methionine and using very cost free or very inexpensive ingredient s A further possible reason for better growth for fish fed Diet 2 (versus Die t 1) is its content of 5% blood meal. Blood meal used in this study contains 84.52% crude protein and is a good source of amino acid except for methionine. It is likely that the more blood meal that can be added to a diet, the better will perform This wou ld be associated with the relatively high amount of protein and excellent digestibility that blood meal provides. Blood meal, however, is the least available NCFR in Haiti because animal protein is scarce and there are not too many slaughterhouses from whi ch to collect raw ingredient. Further, a relatively large volume of blood is needed to generate blood meal; the ratio fresh animal blood to solar dried blood is approximately 1:9 (0.11%). There was no difference in pellet stability between any of the diets used in Experiment 1 and Experiment 2, including the commercially formulated Di et 3 (p>0.05). Even th ough methods used to manufacture the experimental feeds were low technology (feed was extruded with meat grinder or garlic press instead of extruder), res ults indicated that p ercentage recovery after immersion for 10 m inutes in water was acceptable. However, the formulated diets did not float very well compared to the commercial Diet 3 that was formulated as a floating pellet This issue is important becaus e floating feeds are preferred from an aquaculture management perspective The reason for this preference is because it is important to be able to visualize fish while

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81 feeding in order to observe behavioral responses to feeding, appetite and amount of food consumed. Experiment 2 Growth of fish fed experimental Diets 1, 4, 5, 6 or 7 was not different between diets in Experiment 2 (P>0.05). In fact, fish in all diet treatments lost weight Weight loss observed in "medium" fish was less than weight loss observ ed in "large" fish (~43g) had negative weight change in Experiment 2. Lack of gr owth and poor feed intake in Experiment 2 was, at least in part, associated with low palatability of the diets The amount of brewers yeast in the diet formulations in Experiment 2 likely played a role in reduced palatability. Drying the yeast, versus usi ng it wet (directly from the brewery; that would have been easier to formulate into diets) did not change the (Diet 1; 18% dry + 2% wet yeast) versus a diet with wet brewe yeast) and did not observe palatability differences between those formulations in medium or large fish tested in Experiment 2 (P>0.05). Interestingly, the addition of breadfruit to diets with wet yeast appeared to negate the neg ative impact of brewer's yeast on palatability Palatability of Diet 6 (20% wet Diet 1 (18% dry yeast + 2% wet yeast) (P<0.0001). Further, the addition of blood meal to the (Diet 7 and Diet 5, respectively), appeared to improve palatability compared with diets without blood meal (Diets 1, 4 and 6) (P<0.0001).

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82 Alternative NCFRS In addition to the N CFRs examined in the present study, other NCFR like resources have been described that have utility in this discussion For example, good tilapia growth in extensive culture has been achieved with a diet composed of yellow maize, ground nut cake, fish meal as described by Oribhabor and Ansa (2006) B road bean leaf meal has been shown to effectively r eplace 25% of the protein provided by soybeans without loss of growth performance in Nile tilapia ( Fawzy et al., 2008) Combina tions of Lactobaccilus and brewer's yeast (15% total by dry weight) have successfully been used in experimental diets, already containing 29% fishmeal, to supplement total protein in experimental tilapia diets (Schneider et al., 2004). Duckweeds, earthwor ms, coconut meat and jatropha meal have value as contributions for future consideration in formulating diets for tilapia in Haiti (refer to NCFRs as discussed in the Literature Review chapter) Jatropha, however, is a highly palatable and sought after food stuff for humans, and is likely too expensive to consider as a viable NCFR at this time. Brewer's grain has been important as an animal feed supplement around the world Primarily because of the lack of sufficient plant protein sources like soybean in deve of beer production (Westendorf and Wohlt 2002). Its widespread availability makes it a good candidate in the search for alternative protein sources for aquafeeds. The resu lts Nile tilapia O. niloticus with no likely adverse effect on growth. Desale et al. (2008) in a study of O niloticus waste can effectively substitute up to 50%

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83 of the fishmeal protein of a typical commercial feed with no adverse effect on growth of tilapia. Antinutritional and anti quality factors in NCFRs Antinutritional factors are natural or synthetic substances in the diet that have the p otential to adversely affect health and growth by preventing the absorption of nutrients from food These factors can include protease, lipase or amylase inhibitors, phytic acid, oxalic acid, glucosinolates saponins and phenols, among other compounds (Fra ncis et al., 2001) Diets in the present study were formulated with 10% moringa leaves. Richter et al. (2002), in his studies of nutritional quality of raw moringa leaves as partial feed for fish, indicated that up to 10% inclusion of this material can be recommended for Nile tilapia. Based on the favorable amino acid profile of moringa leaves, and their wide availability throughout Haiti, moringa leaves can be considered as a potential feed component with high nutritive value for fish. Moringa leaves have been reported to have some saponins, tannins, phenols and phytate that may serve to act as antinutritional factors (Makkear and Becker, 1997; Francis et al., 200 1 ; Madalla 2008). Incorporation of moringa leaf, mallow and water hyacinth leaf in Nile tilapi a diets at a 10% level supported positive growth, while 20% had no significant effect on growth and 30% depressed growth (Richter et al., 2003; Abdelhamid et al., 2004, 2006). (32 48% crude protein), it is extremely bitter as described in the literature and based on observations palatability at levels higher than 5% in diets.

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84 In addition, elevated levels of plant pr otein in general tend to negatively affect texture and palatability, and may lower feed intake (Al Hafedh and Siddiqui, 1998). High intake (Oliva Teles and Goncalves 2 001). These data suggest that high level of growth could likely be attributed to amino acid deficiency and/or the bitterness of According to Jauncey (20 00) EAA deficiency in the diet leads to poor protein utilization, growth and food conversion efficiency as well as loss of appetite. Additional notes on palatability Data suggest that palatability of feeds may be different for tilapia at different life sta ges as seen in Experiment 2 This is concept is corroborated by additional data from Experiment 1, where palatability for Diet 1 was empirically observed to be greater than palatability observed for fishes fed Diet 1 in Experiment 1 Although palatability data from Experiment 1 cannot be directly compared with data from Experiment 2 with modified methods, it appears that smaller fish may be less sensitive to sub palatable feeds than relatively larger fish. Florida versus Haiti NCFR sources It is important to notice that experimental diets were manufactured using ingredients from Florida. Laboratory analysis of sample ingredients form Haiti and simulation using linear programming suggested that feed manufactured using Haitian ingredients will be higher in cr ude protein than feed manufactured using ingredient from Florida. The difference in crude protein might be caused by a difference in the process of making beer in case of Brewery waste or in the process of drying moringa leaves (solar dried versus drying o ven). Further studies

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85 should manufacture feed with ingredient from Haiti and conduct field trial in Haiti for a better evaluation of growth in Nile tilapia. Conclusion The aquaculture industry in Haiti can use sustainable resources to decrease cost of feed improve needs for animal protein, and minimize the widespread under and malnutrition of children. Tilapia aquaculture needs requires alternative protein sources that are locally available for continued expansion. The results of this study on sustainable feed production to support Nile tilapia aquaculture in Haiti indicate that a 10% moringa and 5% blood meal can be recommended for Nile tilapia feed. In view of the favorab le amino acid profile of moringa leaves and blood meal, the availability of components with high nutritive value for fish in Haiti. However, more investigations must be done to yeast for its inclusion in practical diets for tilapia at higher levels. Ingredients examined in this study may provide excellent utility in formulating sustainable feeds that, along with phytoplankton stimulation, should augment community based tilapia culture in Haiti.

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86 CHAPTER 4 CONCLUSION AND APPLICATIONS Nile tilapia is the main tilapia species cultured in the Caribbean. It represents about 54% of tilapia production in the region in 2002. The rapid growth rate of cultured tilapia provides an opportunity for this species to play a substantial role as a n animal protein source and to meet the needs of the poor in Haiti It also holds promise of becoming an important cash crop in the future As with more traditional forms of animal production, nutrition plays a critical role in intensive aquaculture because it influences not only production costs but also fish growth, health and waste production (Gatlin, 2002). However, the chal lenge that faces Haiti is that feed represent the major expense in fish farming. Commercial feed s are expensive a nd farmer s cannot afford to use them for tilapia production. Therefore, an efficient and sustainable way to develop rural aquaculture in Haiti is to develop least cost, highly nutritive diets using regionally available and sustainable feedstuffs. To develop nutritious, cost effective diets for Nile tilapia we must know its nutritional requirements and meet those requirements with balanced diet fo rmulations and appropriate feeding practices. Research over the last two decades has expanded our knowledge of the nutritional requirements of cultured fishes. Therefore, the current study entailed controlled laboratory experiments to discern the feasibili ty of making feeds with locally available feedstuffs in Haiti for sustainable, community scale tilapia culture in Haiti. The three primary objectives of this study were to investigate availability of ingredients that are sustainable, can be locally harvest ed in Haiti, and that are both high in crude protein and not consumed by people; formulate and manufacture the tilapia feeds ; and conduct lab based controlled feed ing trial s to evaluate the effect of the feeds on the growth of t ilapia

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87 Results for Experime nt 1 disprove hypothesis 1 and 2 : 1. Different isonitrogen ous feed formulations do not affect growth and/weight for tilapia fed experimental diets. 2. Differences in diet formulations do not affect palatability, digestibility, feed performance and stability of experimental diets in the water. Th e result s indicated that the formulated and manufactured Diets 1 and Diet 2 provided positive growth. Diet 2 provided better growth, better Feed Conversion Ratio (FCR), higher Protein Efficiency ratio (PER) and better pal atability than Diet 1 Results for Experiment 2 are in partial agreement with hypothes is 2 and disprove hypothesis 3 : 2. Differences in diet formulations do not affect palatability, digestibility, feed performance and stability of experimental diets in the w ater. 3. Addition of breadfruit to experimental diet formulations does not alter palatability. The results show that diet s had significantly lower palatability than diet with 10% b affecte d is study was less palatable to Nile tilapia at more than 5% level in diets. Data suggest that / or blood meal improves palatability of diets. Addition of breadfruit to experimental diet form ulations improves palatability. Feedstuffs S ustainability in Haiti Moringa (Benzolive) The Moringa oleifera than one season, and is a year round source for nutrition, and potential job creation. It woul d be a huge mista ke not to consider the availability of moringa in Haiti as a harvestable and sustainable crop The idea cannot be to go to Haiti and collect moringa. We are

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88 talking about sustainability. Deforestation in Haiti is a severe environmental pro blem. In 1923, over 60% of Haiti's land was forested; by 2006, less than 2% was. The use of moringa should be part of a strategy for reforestation. Farmers in Haiti do not take full advantage of the unique opportunity of having this year round tree because they do not consider it as a source of income. Moringa can tolerate the intense heat or dr o ught conditions. It is a perennial based crops that is reliable and easy to manage, can be agronomic land and is highly dr o ught resistant. T he strategy is to culture moringa, extend the growing season, reduce labor costs, and provide sustained, long term food security by using it in fish food. Moringa can also be process ed to make the leaf powder as a human nutritional supplement; to make oil; and utilize the seed within water treatment plants (Signa Haiti 2012) Figure 4 1. Moring a seedling s Photos courtesy of SignaHaiti. Breadfruit Reforestation, food security and incom e generation are dire ly needed in Haiti. The b readfruit tree is widely grown and used in Haiti and its fruit is an important staple crop during the fruiting season, but only one or two varieties are cultivated. Breadfruit

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89 has tremendous potential for the c ountry. Breadfruit planting project can be established at areas near fish farm s Breadfruit trees can be interplanted with fruit t rees such as mango, bananas, avocados, m oringa and other plants as part of an integrated, sustainable, managed agro forestry plantation. Our sustainable goal can be achieved if we set up a model management system incorporating interplanting, mulching, composting, ground covers, animal husbandry, and pruning methods to keep breadfruit trees low for ease of harvest. Breadfruit wil l provide a nutritious carbohydrate food for home consumption and fish food and generate income from sale of fresh fruit for processing into flour or meal. A B Figure 4 2. Breadfruit growing in Haiti A) Breadfruit orchard. B) Seedless breadfruit Photos courtesy of SignaHaiti. Cassava Cass ava ( Manihot esculenta manioc in French) is a popular tropical crop cultivated i and as cassava bread It has to be properly cooked for detoxification purpose before being eaten. Cassava is capable of growing in poor soil s. Cassava roots are a source of energy and starch; th e leaves are a so urce of protein

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90 Figure 4 3 Cassava plant showing leaves and roots Photo courtesy of SignaHaiti. Animal Blood Animal bloo d is the least available product as discussed in this thesis, in Haiti because of the deplorable condition of our existent s laughterhouses. Although, within the Caribbean community ( CARICOM ) Haiti is the largest beef producer followed by Jamaica, Guyana, Belize and Suriname (FAO, 2002) ; and Haiti has by far the highest goat population in the region (more than twice the goat po pulations from Cuba, Dominican Republic and Jamaica); slaughterhouses are somehow scarce in Haiti and collecting animal blood will be c hallenging However, the national agricultural i nvestment p lan (2010 2016) h as a budget of 4,896,000 USD to develop the c omponent slaughter, meat preparation and packaging. Construction or rehabilitation of six slaughterhouses from 2010 to 2013 is part of this plan. Brewery and Distillery W aste Haiti has two well known brewery and r um companies In addition, there are numero us small er producer of Haitian alcohol drinks Brasserie Nationale D'Haiti also called BRANA, is the leading beverage company in Haiti, producing, marketing and distributing various beer and malt brands, most notably

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91 P re stige, Malta and Guinness, as well as various P epsico licensed soft drink brand s. It is a top Caribbean beer producer as well. The brewery manufactures the popular P restige beer, one of the premium American s tyle lagers produced in the Caribbean whose popu larity is gradually crossing national boundaries in terms of consumption and marketing. Barbancourt is a well known rum produced and bottled by Socit du Rhum Barbancourt, T. Gardre & Cie. in Haiti. The company is one of Haiti's oldest and its rum is the country's most famous export, widely regarded as among the finest rums in the world. Those industries produce hundreds of tons of solids and liquid waste per day. Not only those waste can be used as source of protein in fish diets to improve aquaculture i n Haiti as it is presented in this study but also can be part of an urban and environmental management. The idea of using those wastes is a model for environmental management and may reduce potential hazards on aquatic ecosystem. This is important because industrial activities are source of excessive nitrogen and phosphate loads in aquatic environments, generating a high pollution in Haiti (Personal observation) Most of these pollutants are poured off into the sea without prior treatment and can cause phen omenon of eutrophica tion and destroy or weaken our c oastal ecosystem. Excess of nutrients from those industries can cause turbidity and odor, toxic algal blooms, r ed tides, loss of biodiversity, damaging coastal environment, sedimentation, loss of habitat, decrease coastal population and collapse of fishing in Haitian coastal ecosystems.

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92 Why F ish? Nutritional Benefits of F ish First and foremost, increasing the per caput consumption of fish in any country benefits health. Aquatic animals contain a high leve l of protein (17 20%), with an amino acid profile similar to that of the meat of land animals. The flesh of fish is therefore readily digestible and immediately utilizable by the human body. Compared with land animals (with some exceptions, such as shellfi sh), aquatic animals have a far higher percentage of edible flesh, and there is little wastage. Fish are a source of minerals, such as calcium, iron, and phosphorus, as well as trace elements and vitamins. Marine species are particularly rich in iodine. Th e poly unsaturated fatty acid content is high and particularly those which are attributed to reduce blood cholesterol. Characteristics of Consumption in the R egion Fish have played a central role in the food economy of man from early times. The developmen t of civilizations in the Caribbean islands was undoubtedly influenced by the availability of fish and shellfish found in the surrounding waters. Fish constitutes a traditional element of the diet of most Caribbean people and cannot be easily substituted b y other foods. Fish is preferred in a fresh whole form, although salted dried fish is well accepted in some islands. Processed forms are bought in larger quantities due to greater availability and sometimes cheaper prices. Fish imports into the region comp rise a wide range of species and product forms, mostly dried, smoked, salt dried, gutted, iced, and canned. In the Greater Antilles most fish are acceptable without preference for those of demersal or pelagic origin. Some freshwa ter fishes, namely the tila pias and carps, have recently become popular. In fact, in countries like Cuba, Dominican Republic, Jamaica, and Trinidad and Tobago, the growth and transformation

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93 of the productive structure of fish farming have had a strong impact on the marketing and con sumption of fish. In Haiti, fishing yield is very low 16,000 MT per year for fisheries and 400 MT per year for aquaculture. Sedimentation and damage of costal environment affect fishing and poor fish farmers cannot afford the mean of production of fish far ming. Haitians consume only 2.5 Kg of fish per year per person compared to other areas in the Caribbean where fish is a staple for protein. Caribbean fish consumption is 7 times greater than in Haiti. pounds of fish c onsumption are imported (WFP, 2008) Where Do We Go From Here ? According to the CIA World Fact book, Haiti imported more than $325 million in rice, wheat, meat, poultry and other food items from the United States in 2008. Meat and poultry represented $33.8 million, up 26.4% of U.S. exports to Haiti in 2008. This situation is critical and more than ever we need to have a plan to improve fish production in Haiti. The objectives of a sustainable project cannot be to feed Haitian s im for a day. Teach a person to fish feed him for a live Non conventional protein sources need to be investigated further for the development of low cost pelleted feeds, high in crude protein and suitable for use by small scale farmers in Haiti. Stu dy must focus on how to make floating pellet since Haiti is a mountainous land and all of the water bodies are deep. However the formulated diets in this study can be part of a sustainable project to re establish fishing as a reliable occupat ion in Haiti. The most efficient system is to grow fish to 100 150 g with fertilizers alone, then add feed. El S ayed (2006) reviewed strategies for optimum yield of tilapia production. Research experiments indicated that supplemental feeding in fertilized ponds can resu lt in significantly higher growth rates and greater yields than fertilization

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94 alon e (Green 1992; Diana et al.1994; Re fer to Table 4 1 ) Raising tilapia in green ponds supplemented with Diet 2 (or an improved derivation thereof) can help build a fish farmin g industry for the benefit of thousands of poor Haitians. Different strategies can be developed: 1. Micro enterprise: with proper funding several thousand good jobs can be created in Haiti for fishermen and other employees in the process of manufacturing the diet cultivate and collect the feedstuff. For local market, the establishment of micro credit systems (type community bank) to best suited their need for mean of production such as make or buy feed. 2. Location: Because of lack of infrastructure and lack of p ower energy in Haiti, the proper location for this project is important. One should take into consideration, access to roads, the marketing channel, etc. 3. Human dimension: A project like this can be a whole new way to make women participate and get involv ed in the fishing industry. 4. Job creation: There is a lack of job in Haiti. Rural farmers live to subsist every not be sustainable. For instance they need to eat a small fish but most important ly they need to be able to sell a big fish to buy clothes soap or pay tuition They cannot have an improvement in their standard of living without earning cash.

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95 Table 4 1. F eed and fertilization strategies suggested for optimum yield of tilapia in semi intensive culture systems ( as compiled by El Sayed 2006) Cultured fish Species (Country) Size (g) Density / ha Fertilization regime Feeding Culture period (days) Yield (Mt/ha) Remarks Reference Nile tilapia (Honduras) 10 000 Chicken manure, 1000 kg/ha/week 152 1.76 During rainy season Green et al. (1990) Nile tilapia (Panama) 10 000 Chicken manure, 1000 kg/ha/week 1.49 2.07 During rainy season Green et al. ( 1990) Nile tilapia (Philippines 20 000 Chicken manure, 500 kg/ha/week 23% cp diet 1.5% bw/day 151 4.35 Based on cost/benefit analysis Green et al. (1992) Nile tilapia (Cameroon) 202 222 7 600 Cattle manure, daily, 226 kg dry wt/ha/day CSC; 3% bw/day 6% bw/day 100 4.80/yea r Polyculture with C larias gariepinus (1100 fish/ha) as police fish Middendorp (1995) (Thailand) 15 20 000 280 kg chicken manure + 56.3 kg urea + 17.5 TSP/ha/week 234 4.0 4.2 2.5 m deep ponds, no water addition. Wee kly fertilization produced best growth Diana and Lin (1998)

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96 bw, body weight; CSC, cottonseed cake; TSP, triple superphosphate Table 4 1. Continued Cultured Fish Species (Country) Size (g) Density / ha Fertilization regime Feeding Culture period (days) Yield (Mt/ha) Remarks Reference Red til apia (Thailand) 33 62.5/m 3 Urea + TSP; 4kg N + 2.18 kg P/ha/day Com merci al diet, 0 100% satiation 90 13.5 kg/m3 C ages in fertilized ponds, 50% satiation was economically best Y i et al (2004) (Kenya) 90 1000 Diammonium phosphate + urea, 20kg N/ha/week 133 1.72 During warm season (23.5 28.1 c) Veverica et al. (2001) (Egypt) 1 3 20 000 Chi c ken ma n ure 1000 kg/ha/week, for 60 days. 54.4 kg urea + 92.4 kg Superphosphate / ha/week 30% cp diet, 3% bw/day starts day 60 145 7.40/yea r African catfish (59g) were used for seed control Green et al. (2002) No feeding 145 3.20/yea r Same regime Nile tilapia (Egypt) 13.8 30 000 750 kg Chicken litter/ha, biweekly application, 1 00 TSP and 20 kg urea/ha 25% cp diet, 3% bw/day, 6 week after stocking 133 4.75 Feeding at 6 week delay was comparable to no delay Abdelghany et al. (2002) Silver carp 1.9 Common carp 10.7

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97 APPENDIX A ADDITIONAL RESOURCES A 1 Theoretical composition of diets using composition of sample ingredients from Haiti and Value from literature Sample ingredients from Haiti Literature values Ingredients (%) Diet 1 Diet 2 Diet1 Diet 2 Moringa leaf 10 10 10 10 Brewer s grain 50 60 50 60 20 5 20 5 ----Cassava roots 20 20 20 20 Dried blood meal -5 -5 Distiller's waste Haiti ----Breadfruit flour ----Total 100 100 100 100 EAA's (g/100g protein) Arginine 4.42 4.40 4.06 4.33 Histidine 2.20 2.09 1.67 2.06 Isoleucine 3.69 3.64 3.59 3.18 Leucine 6.34 6.64 5.48 6.34 Lysine 4.20 3.99 3.00 3.49 Methionine 1.44 1.46 1.28 1.46 Phenylalanine 4.94 5.20 4.19 4.76 Threonine 3.43 3.38 2.79 2.93 Tryptophan 1.32 1.31 0.34 0.31 Valine 4.60 4.69 3.78 4.33 Crude Protein (%) 23.84 23.27 29.01 29.58 Moisture (%) 8.16 8.50 --Crude Fat (%) 5.05 5.75 4.02 4.47 Crude Fiber (%) 8.91 10.48 10.64 11.45 Ash (%) 4.65 4.61 --

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98 A 2 Water quality parameter throughout the duration of Experiment 1. Values are means SE (n=3) A 3 Weekly water quality parameters thr oughout duration E xperiment 2 Date Temperature pH DO Unionized ammonia Nitrite Week 1 28.4 0.3 7.78 0.06 6. 3 0.3 <0.02 <0.05 Week 2 28.2 0.2 7.83 0.04 6.8 0.1 <0.02 <0.05 Week 3 29.1 0.3 7.76 0.03 6.5 0.2 <0.02 <0.05 Week 4 27.7 0.3 7.33 0.26 6.9 0,2 <0.02 <0.05 A 1 Marketing channel of fish in Haiti Temperature (C) pH Unionized NH3 (mg/L) Nitrite (mg/L) DO saturation % Diet 1 26.6 0.2 7.21 0.03 0.00 0.00 0.05 0.00 81% 1 Diet 2 27.0 0.1 7.40 0.09 0.01 0.01 0.05 0.00 75% 2 Diet 3 26.9 0.1 7.33 0.05 0.01 0.00 0.08 0.00 63% 5

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99 A 2 Block diagram of the schem e used to manufacture fish feed in this study. Adapted from Robinson et al. (2004)

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100 APPENDIX B ANALYSIS OF VARIANCE Analysis of Variance of parameters for Experiment 1 B 1. Analysis of variance of protein i ntake (PI) of fish fe d experimental Diet 1, Diet 2 and Control diet ( Diet 3 ) ANOVA Source of Variation SS df MS F P value F crit Between Groups 11.64 2 5.82 335298.7 <0.0001 5.14 Within Groups 0.00 6 <0.0001 Total 11.64 8 B 2. Analysis of variance of feed c onversion ratio (FCR) of fish fed experimental Diet 1, Diet 2 and Diet 3 ANOVA Source of Variation SS df MS F P value F crit Between Groups 52.67 2 26.33 209.08 <0.00 0 1 5.14 Within Groups 0.76 6 0.13 Total 53.42 8 B 3. Analysis of var iance of protein efficiency r atio (PER) of fish fed experimental Diet 1, Diet 2 and Diet 3 ANOVA Source of Variation SS df MS F P value F crit Between Groups 6.56 2 3.28 183.80 <0.0001 5.14 Within Groups 0.11 6 0.02 Total 6.67 8

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101 B 4. Analysis of variance of b ody weight gain (BWG) of fish fed experimental Diet 1, Diet 2 and Diet 3 ANOVA Source of Variation SS df MS F P value F crit Between Groups 199536.70 2 99768.34 388.21 <0.0001 5.14 Within Groups 1541.99 6 256.99 Total 201078.70 8 B 5. Analysis of variance of growth of fish fed experimental Diet 1, Diet 2 and Diet 3 Source Nparm DF Sum of Squares F Ratio Prob > F Diet 2 2 166.86519 358.7070 <.0001* Time 2 2 255.95630 550.2245 <.0001* Diet*Time 4 4 154.23481 165.7779 <.0001* B 6. Analysis of variance of p ellet stability of experimental Diet 1, Diet 2 and Diet 3 ANOVA Source of Variation SS df MS F P value F crit Between Groups 183.74 2 91.87 10.18 0.0460 9.55 Within Groups 27.08 3 9.03 Total 210.82 5 B 7 Analysis of variance of palatability of experimental Diet 1, Diet 2 and Diet 3 ANOVA Source of Variation SS df MS F P value F crit Between Groups 13.39 2 6.69 8868.47 <0.0001 5.14 Within Group s 0.005 6 0.0008 Total 13.39 8

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102 B 8 Analysis of variance of digestibility of experimental Diet 1 Diet 2 and Diet 3 ANOVA Source of Variation SS df MS F P value F crit Between Groups 2.53 2 1.27 348.34 <0.0001 5.14 Within Gro ups 0.02 6 0.003 Total 2.55 8 Analysis Of V ariance of parameters for Experiment 2 B 9 Analysis of variance of p alatability of experimental Diet 1, Diet 4, Diet 5, Diet 6 and Diet 7 Source Nparm DF Sum of Squares F Ratio Prob > F Time (day) 1 1 2.7834320 24.8588 <.0001* Diet 4 4 6.4558824 14.4143 <.0001* Time (day)*Diet 4 4 0.9648494 2.1543 0.0769 Block 1 1 8.4941176 75.8608 <.0001* Time (day)*Block 1 1 0.0264108 0.2359 0.6279 Diet*Block 4 4 0.2558824 0.5713 0.6838 Time (day)*Diet*Block 4 4 0.9004185 2.0104 0.0959 B 10 Analysis of variance of final weight of fish fed experimental Diet 1, Diet 4, Diet 5, Diet 6 and Diet 7 Source Nparm DF Sum of Squares F Ratio Prob > F Diet 4 4 10.2333 1.3465 0.3190 Block 1 1 31 39.2667 1652.246 < 0 .0001* Diet*Block 4 4 22.2333 2.9254 0.0768 Time 1 1 6.6667 3.5088 0.0905 Diet*Time 4 4 2.1667 0.2851 0.8811 Block*Time 1 1 4.2667 2.2456 0.1649 Diet*Block*Time 4 4 0.5667 0.0746 0.9884

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103 LIST OF REFERENCES AOAC (Association of Official Analytical Chemists) 2005. Official Methods of Analysis of AOAC INTERNATIONAL. 18th Ed., AOAC INTERNATIONAL, Gaithersburg, MD, USA Abdelghany, A. E. 1996 Growth response of Nile tilapia ( Oreochro mis niloticus ) to dietary L ascorbic acid, L ascorbyl 2 sulfate and L ascorbyl 2 polyphosphate. World Aquaculture Society 27(4) : 449 455. Abdelghany, A.E. and Ahmad, M. H. 2002 Effects of feeding rates on growth and production of Nile tilapia, common carp e and Silver carp polycultured in fertilized ponds. Aquaculture Research 33 : 415 423. Abdelhamid, A. M., M. F. I. S. and A. E. T. 2004 Evaluation of sesame meal as a dietary protein source for Nile tilapia ( Oreochromis niloticus ) fingerlings J. Agric. S ci. Mansoura Univ. ( 29 ): 6887 6897. Abdelhamid, A. M., M. F. I. S. and M. M. K. 2006 Substitution of soybean meal by water hyacinth hay in diets of Nile tilapia. The 2nd International Scientific Congress for Environment, recent environmental problems and sharement.28 30 March, South Valley University (pp. 114 125). Akinleye, A. O., Kumar, V., Makkar, H. P. S., Angulo Escalante, M. a, & Becker, K. 2012 J atropha platyphylla kernel meal as feed ingredient for Nile tilapia ( Oreochromis niloticus ): growth, nut rient utilization and blood parameters. Journal of animal physiology and animal nutrition 96 (1): 119 29. Retrieved September 19, 2012, from http://www.ncbi.nlm.nih.gov/pubmed/21410563 Al Hafedh, Y. S. and A. Q. S. 1998 Evaluation of guar seed as a protein source in Nile tilapia ( Oreochromis niloticus ) practical diets. Aquaculture Research ( 29 ): 703 708. Amsalu, N. and Esubalew G. 2011 Soaking and drying of cassava roots reduced cyanogenic potential of three cassava varieties at Jimma, South west Ethiopia. World Journal of Agricultural Sciences, 7 (4): 439 443. IDOSI Publications. Anderson, J., Jackson, A.J., Matty, A.J. and capper,B.S. 1 984 Effects of dietary carbohydrate and fiber on the tilapia Oreochromis niloticus Aquaculture ( 37 ): 303 314. Becker, K and H. P. M. 2001 Effect of dietary tannic acid on growth performance and metabolic rates of common carp. Aquaculture ( 175 ): 327 332. Beve ridge, M. C. and McAndrew, B. 2000 Tilapias: biology and expl oitation (M. C Beveridge & B. McAndrew, editors .) (Kluwer Aca. Press p. 508).

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104 Cayemittes, M. 2007 Enqute mortalit, morbidit et utilisation d es services IV Hati 2005 2006. Accessed on September 9, 2011. Chapman, F. A. 1992 Culture of hybrid tilapia: a reference profile". University of Florida, In stitute of Food and Agricultural Sciences. CIA. 2010 The world factbook Source: https://www.cia.gov/library/publications/the world factbook/geos/ha.html Culley, D. D., Rejmankova, E., Kvet, J. and Frye, J.B. 1981 Production, chemical quality, and use of duckweeds (Lemnaceae) in aquaculture, was te management and animal feed. Journal of World Mariculture Society ( 12 ): 27 49. DAMAIS, G., P. V. of A. S. and D. S. C. 2007 Study of the fishing industry in Haiti. IRAM / inesa Interim p. 116. Dabrowska, H., Me yer Brugdoff, K. and Gunther, K.D. 1989 Interaction between dietary protein and magnesium level in Tilapia ( Oreochromis niloticus ). Aquaculture ( 76 ): 277 291. Desale, Z. B., Fitzsimmons, K. M., Collier, R. J. AND Duff, C. G. 2008 Evaluation of ste as partial replacement of fish meal protein in Nile t ilapia Oreochromis niloticus d iets. Journal of the World Aquaculture Society 39 (4): 556 564. Devendra, C. 1988. General approaches to animal n utrition research and their relevance to fish product ion in the Asian region. Pages 7 24 i n S.S. DeSilva, editors Finfish Nutrition Research in Asia ). Heinemannn Asia Singapore, Singapore. Diana, J.S., and Lin, C. K. 1998 The effect of fertilization and water management on g rowth and production of Nile ti l apia in deep ponds during the dry season. Journal of the World Aquaculture Society ( 29 ): 405 413. Duncan, D. B. 1 955 Multiple Ranges and Multiple F test. Biometrics (11th ed., pp. 1 42). Eid, A. L. and G., S. 1994. Dietary z inc requirem e n t of fingerlings Oreochromis niloticus Aquaculture ( 119 ): 259 264. El Sayed A. M. and S. T. 1991 Tilapia nutrition in aquaculture. Reviews in Aquatic Sciences 5(3 4) : 247 265. El Sayed A. F.M. and Garling, D.L., Jr. 1988 Carbohydrates to lipids ratios in diets for T il apia z illii fingerlings. Aquaculture 73(1 4) : 157 163. El Sayed A.F 2006 Tilapia c ulture. CABI publishing, p. 277. Cambrige USA

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105 El Sayed A.F. and Teshima, S. 1992 Protein and energy requirements of Nile tilapia, Oreochromis niloticus fry. Aquacultu re ( 103 ): 55 63. Essa, M. A. 1997 Utilization of some aquatic plants in diets for Nile tilapia, Oreochromis niloticus fingerlings. Egyptian Journal of Aquatic Biology and Fisheries ( 1 ): 19 34. FAO. 1984 Dam design and operation to optimize fish productio n in impounded river basins. CIFA Technical paper, FAO, Rome 11 pp FAO. 1986 Water resources legislation and administration in selected Caribbean countries. New york: un: FAO descriptors: water resources; laws and regulations; water management ; Antigua and Barbuda; Cayman Islands; Dominican Republic; Haiti; Jamaica; Montserrat; S (p. 171). New York: Department of Technical Cooperation for Development. FAO. 1997 Review of the state of world aquaculture. FAO Fisheries Circular (No. 886, Rev. 1. FAO, Rom e), 163 pp. FAO. 1999 The state of world fisheries and aquacult ure 1998 A nnual report. Rome. 176 pp FAO. 2000 The state of world fisheries and aquaculture 2000 Annual report. Rome, FAO. 176 p p. FAO 2010. The state of world fisheries and aquaculture 2 010. Rome, FAO. 197 p p. FAO. 2010 Document du en Hati Proje ct d ocument. Aquaculture development in rur al comm unities in Haiti, (PNUD/FAO/HAI/10/010), 21p. FAO. 2011 Food security data and definitions food g roups. 9, 2011. http://www.fao.org/economic / ess/ess fs/fs data/ess fadata/en/. (See Food consumption: Quantities Fawzy, I. M., EL Gendi, M. O. Salem, M. F. I. A., & Elazab, A. A. 2008 Use of cucumber, squash and broad bean leaves as non conventional plant pro tein sources in N ile tilapia ( O reochromis niloticus ) diet. Eigh th International Symposium on Tilapia in Aquaculture 2008 (pp. 847 859). Fitzsimmons, K. 2006 Prospect and potential for global productio n. Pages 51 72 i n C. E. L im & C. D. Webster, e d itors. Tilapia: biology, culture, and nutrition The Hawort Food Products Press. Fitzsimmons, K. & C. F., J. 2000 Tilapia aquaculture in the 21st century: Pages 682 in p roceedings of the Fifth International Symposium on Tilapia in A quaculture Brasilia, Brazil: Rio de Janeiro, Brazil,Ministrio de Agricultura, Dep artmento de Pesca e Aqicultura

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106 Florvil, A. 1992 Etude compare diffrents reprise des propagules du manglier rouge ( Rhizophora m angle ) et la croissance State University of Haiti. Foidl, N., Makkar, H.P.S., Becker, K. 2001 The potential of Moringa oleifera for agricultural and industrial uses. Pages 45 76 i n L. J. Fuglie, e d itors. The Mi racle Tree CTA, Wageningen, Net herlands and CWS, New York, USA Francis, G., Makkar, H.P.S., and Becker, K. 2001. Antinutritional factors present in plant derived alternate fish feed ingredients and their effects in fish. Aquaculture 199:197 227. Gatlin, D. M. I. 2002 Nutrition and fish health. Pages 671 702 i n R. W. Halver, J. E. and Hardy, editors. Fish Nutrition (3rd ed.). London: Academic Pres. Green, B. W. 1992 Substitution of organic manure for pellet feed in tilapia production. Aquaculture ( 101 ): 2 13 222. Green, B. W., El Nagdy, Z. and Hebicha, H. 2002 Evaluation of Nile tilapia pond management strategies in Egypt. Aquaculture Research ( 33 ): 1037 1048. Green, B. W., Teichert Coddington, D. R. and Phelps, R. P. 1990 Response of tilapia yield and econ omics to varying rates of organics f ertilization and season in two Central A merica countries. Aquaculture ( 90 ): 279 290. Hardy, W. R., Barrows, F. T. 2002 Diet Formulation and Manufacture. Pages 506 596 i n John E. Halver and Ronaldo W. Hardy e d itors. Fish Nutrition (3th ed). Academic press. Hillman W.S. and Culley D.D. 1978 The uses of duckweed American Scientist ( 66 ): 442 451. Hoffman, L. C., J. F. Prinsloo, and G. R. 1997 Partial replacement of fish meal with either soybean meal, yeast or tom ato meal in the diets of African sharptooth catfish, Clarias gariepinus. Water SA 23(2) : 181 186. Jauncey, K. 2000 Nutritional requirements. Pages 327 375 i n M C. Beveridge & B. McAndrew, e d itors. Tilapias: Biology and Exploitation London,UK: Kluwer Aca demic Publishers. Jauncey, K., Tacon, A.G.J. and Jackson, A. J. 1983 The quantitative essential amino acid s requirements of Sarotherodon mossambicus Pages 328 337 i n L. and Z. Y. Fishelson, e d itors Proceedings, International Symposium on Tilapia in Aqua culture Aviv Israel. Jobling, S. 1983. A short review and critique of methodology used in fish nutrition studies. J. Fish Bio. ( 23 ): 685 703.

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107 Khawaja, T., S. H. Khan, N. N. Ansari. 2007 Effect of different levels of blood meal on broiler performanc e duri ng two phases of growth. International Journal of Poultry Science 6 (12) : 860 865. Kleemann, G. K., Barros, M.M., Pezzato, L.E., Sampaio, F.G., Ferrari, J.C., Valle, J.B., Freire, E.S., Zuanon, J.A. 2003 Iron requirements for Nile tilapia Oreochromois nilo ticus World Aquaculture Society, baton Rouge, Louisiana (Vol. vol1, p. 82). B aton Rouge, Louisiana Li, M. H. 2006 Feed formulation and manufacture. Pages 517 545 i n C. E. Lim & C. D. Webster e d itors. Tilapia: Biology, Culture, and Nutrition Food Product s Press. Li, Z., Lei, W., Ye, J. and He, X. 1991. The nutritional value of commercial feed in ingredients for Nile tilapia in China. Pages 101 106 i n S. S. De Silva e d itors Fish nutrition research in Asia M anila, Philippines: Asian Fisher ies Society Spe cial Publication. Lim E. Ch h orn and Carl D. Webster. 2006. Nutrient requirements. Pages 469 501 i n W. C. David, e d itors Tilapia: Biology, Culture, and Nutrition Food Produ ct Press. Lim E. Ch h orn and Klesius P.H. 2001 Influence of dietary levels of f olic acid on growth response and resistance of Nile tilapia Oreochromis niloticus to Streptococcus iniae Pages 150 i n Sixth Asian Fisheries Forum. Book of abstracts Quezon City, Philippines: Asian Fisheries Society. Lovell, R. T. 1988. Nutrition and Feed ing of Fish (p. 224). Kluwer Academic Publishers. MacLennan, D. N. 1995 Technology in capture fis heries. Paper presented at the Government of Japan/FAO International Conference on Sustainable Contribution of Fisheries to Food Security, Kyoto, Japan 4 9 D ecember 1995; and 1997. Review of th e state of World Aquaculture. FAO Fisheries Circular No. 886, Rev. 1. Rome, Italy (December 1995). Madalla, N. 2008. Novel feed ingredients for Nile Tilapia ( O reochromis niloticus L.). Doctoral thesis, Institute of Aqua culture, University of Sterling, Scotland, December 2008. Madu, C. T., Sogbesan, O.A. and Ibiyo, L.M.O. 2003 Some Non conventional f ish feed resources in Nigeria. Pages 73 82 i n A.A. Eyo, editors: p roceeding of the Joint Fisheries Society of Nigeria/Natio nal Institute For Freshwater Fisheries Research/FAO National Special Programme For Food Security National workshop on Fish feed development and Feeding Practices in Aquaculture held New Bussa. Makkar, H.P.S. and Becker K. 1997. Nutrients and antiquality factors in different morphological parts of the Morninga oleifera tree. Journal of Agricultural Science 128(3):311 322.

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113 BIOGRAPHICAL SKETCH Pascale was born in Jrmie, Haiti to Guy and Mervela Fran ois. She is mom to be autiful 3 year old girl named Jeisha Jenny. She earned her Bachelor of Science at the State University of Haiti, where she pursued an a gronomy degree with minor in natural resources and environment c onservation. She presented and defended with honor her u n evices (FAD) at Anse ity of Florida she worked with national program of c atchment po nds ( p rogramme nationale de lacs collinaires PNLC) f ou nded by the Ministry of Natural Resources and Environme nt with a Master of Science in i nterdisciplinary e colo gy with concentration in F isheries and Aquatic Sciences.