CULTURE OF THE FRESHWATER CLADOCERAN, DAPHNIA PULEX F\ UTILIZING SCENEDESMUS OBLIQUUS GROWN IN DAIRY WASTE MEDIUM by NELSON M. CASTILLO B. Sc., University of the Philippines, Quezon City Philippines, 1975 A THESIS SUBMITTED IN PARTIAL FULFILLMENT OF THE REQUIREMENTS FOR THE DEGREE OF MASTER OF SCIENCE in THE FACULTY OF GRADUATE STUDIES (Department of Agricultural Mechanics) We accept this thesis as conforming to the required standard THE UNIVERSITY OF BRITISH COLUMBIA October 1981 ©Nelson M. Castillo, 1981 In presenting this thesis in partial fulfilment of the requirements for an advanced degree at the University of British Columbia, I agree that the Library, shall make it freely available for reference and study. I further agree that permission for extensive copying of this thesis for scholarly purposes may be granted by the head of my department or by his or her representatives. It is understood that copying or publication of this thesis for financial gain shall not be allowed without my written permission. Department of Q^O^!Sn^ QUx&qV\flg*> The University of British Columbia 2075 Wesbrook Place Vancouver, Canada V6T 1W5 Date SAW \&j R?\ J ii ABSTRACT Scenedesmus obliquus was grown in batch cultures at varying concentrations of digested dairy medium ranging from 200 to 2,000 ug-at • N-l1. Higher growth rates were observed at low N- concentrations while higher cell yields were observed at high N- concentrations. Aeration enhanced both algal growth rates and biomass yields. Results show an advantage in adjusting the nitrogen to phosphorus atomic ratio of the medium. More biomass was produced in cultures with higher N:P ratios. The algal biomass produced was used as food for the freshwater cladoceran, Daphnia pulex F. Three feeding levels -1 were used: 50,000, 100,000 and 150,000 cells-ml . However, no significant differences were observed in both Daphnia biomass yields and biomass conversion efficiencies. The tendency of Scenedesmus cells to settle down in the bottom and cling to the sides of the tank presented a major problem in the study.- Intensive feeding did not increase the biomass production of Daphnia, although larger-sized adults with larger brood sizes were produced. Animals in culture reached a density of 1.24 animals-mfAand obtained conversion efficiencies as high as 40- 50%. Dr. J,W. Zahradnik Thesis Supervisor iii TABLE OF CONTENTS Page ABSTRACT ii TABLE OF CONTENTS . iii LIST OF TABLES v LIST OF FIGURES . vi ACKNOWLEDGEMENTS ix INTRODUCTION 1 LITERATURE REVIEW '.. 3 Recycling of Wastes through Aquaculture 3 Algal Production from Waste 5 Daphnia as a Potential Fish Food in Aquaculture 8 Present State-of-the-Art on Daphnia Culture 12 MATERIALS AND METHODS 14 Algae 14 Daphnia pulex 14 Media Preparation 15 Algal Culture Medium 15 Daphnia Culture Medium 15 Description of Culture Units 16 Algal Culture Unit 16 Daphnia Culture Unit 18 Culture Methods 18 Algal-Culture Method 18 Daphnia Culture Method 19 Chemical Analysis 21 Kheldahl Nitrogen 21 Ammonia/Nitrate/Nitrite 21 iv Total Phosphate 22 . Ortho-Phosphate 22 Algal Cell Weight 22 Growth Rate Experiments ( Daphnia ) 23 Length-Weight Relationship ( Daphnia ) 23 RESULTS 25. Dairy Waste Analysis .25 Algal Experiments 26 Daphnia Experiments 37 Algal Cell Weight 68 Growth Rate Experiments 68 Length-Weight Relationship 71 Brood Size in Relation to Daphnia length 71 DISCUSSION 77 Livestock Waste as a Nutrient Source in Algal Production 77 Algal Growth on Manure Medium 78 Daphnia Biomass Production 81 Population Growth and Size Structure 82 Growth Rate Experiments 84 Length-Weight Relationship 85 Nutritional Inadequacy of Certain Algae Daphnia Culture 85 Toxicity in Daphnia Culture 87 Scale-up Considerations 89 SUMMARY AND CONCLUSIONS 91 REFERENCES 94 APPENDICES 103 V LIST OF TABLES Table Title Page 1 Summary of chemical composition of each media and the algal experiment in which they were used 25 2 Summary of the algal experiments, the different- treatments and the results 27 3 Chemical composition of media before and after algal growth .33 4 Amount of algae consumed by Daphnia at 3-day intervals .42 ,5 Fecundity of Daphnia cultures in all five experiments 69 6 Ammonia, pH and DO levels measured daily in Daphnia culture tanks in D^ Expts. #4 #5 70 7 Daphnia brood size in relation to total length 75 vi LIST OF FIGURES Figure Title Page 1 A generalized view of the Algal Culture Unit 17 2 Daily cell density of S.obiiquus grown in varying N-concentrations of dairy waste medium (Algal Expt.#1) 28 3 Daily cell density of S.obiiquus grown in varying N-concentrations of.dairy waste medium (Algal Expt.#2) 30 4 Daily cell density of S.obiiquus grown in varying N-concentrations of dairy waste medium (Algal Expt.#3) 31 5 Daily cell density of S.obliquus grown in varying . N-concentrations of dairy waste medium (Algal Expt.#4). .32 6 Daily cell density of S.obiiquus grown in varying N-concentrations of dairy waste medium (Algal Expt.#5) • 35 7 Daily cell density of S.obiiquus grown in varying N-concentrations of dairy waste medium (Algal Expt.#6) 36 8 Daily cell density of S.obliquus grown in dairy waste medium with Nitrogen to Phosphorus atomic ratios of 17 and 64 (Algal Expt.#7) 38 9 Daily cell density of S.obliquus grown in dairy waste medium with Nitrogen to Phosphorus atomic ratios of 4, 22 and 69 (Algal Expt.#9) 39 10 Daily cell density of S.obiiquus grown in dairy waste medium with and without aeration (Algal Expt.#8) 40 11 N-consumed vs. Algal yield 41 12 Daphnia biomass at the three feeding levels at 3-day intervals ( D^ Expt.#l) 45' 13 Biomass conversion efficiency of Scenedesmus to Daphnia at the three feeding levels at 3-day intervals ( EL Expt.#l) 45 14 Daphnia biomass at the three feeding levels at 3-day intervals ( D^ Expt.#2) 46 15 Biomass conversion efficiency of Scenedesmus to Daphnia at the three feeding levels at 3-day •VI1 intervals ( IK Expt'.#2) 46 16 Daphnia biomass at the three feeding levels..at 3-day intervals (-'IK Expt. #3)' 47 17 Biomass conversion efficieny of Scenedesmus to Daphnia at the three feeding levels at 3-day . intervals (. IK" Expt. #3) 47 18 Daphnia biomass at 3-day intervals fed with 100,000 Scenedesmus cells ml 2 to 3 times daily (IK. Expt.#4 and #5) ; 48 19 Biomass conversion efficiency of Scenedesmus to Daphnia in EK_ Expts.#4 and #5 at.intervals 48. 20 Proportion of juvenile, young adult and adult Daphnia in terms of number and biomass in cultures at the three feeding levels at 3-day intervals ( EL Expt.#1) 49 21 Proportion of juvenile, young adult and adult Daphnia in terms of number and biomass in cultures at the three feeding levels at 3-day intervals ( IK Expt.=2) 50 22 Proportion of juvenile, young adult and adult Daphnia in terms of number and biomass in cultures at the three feeding levels at 3-day intervals ( IK Expt.#3) 51 23 Proportion of juvenile, young adult, and adult Daphnia in terms of number and biomass at 3-day intervals in EK_ Expt.#4 and #5 52 24 Daphnia size-frequency structure at 50,000 cells ml feeding level at 3-day intervals ( IK Expt.#1) 53 25 Daphnia size-frequency structure at 100,000 cells ml feeding level at 3-day intervals ( IK Expt.#1) 54 26 Daphnia size-frequency structure at 150,000 cells ml feeding level at 3-day intervals ( IK Expt.#1) 55 27 Daphnia size-frequency structure at 50,000 cells ml feeding level at 3-day intervals ( IK Expt.#2) 56 28 Daphnia size-frequency structure at 100,000 cells ml feeding level at 3-day intervals ( IK Expt.#2) 57 29 Daphnia size-frequency structure at 150,000 VI11 cells ml feeding level at 3-day intervals ( EL Expt.#2) 58 30 Daphnia size-frequency structure at 50,000 cells ml feeding level at 3-day intervals ( EL Expt.#3) 59 31 Daphnia size-frequency structure at 100,000 cells ml feeding level at 3-day intervals ( EL_ Expt.#3) 60 32 Daphnia size-frequency structure at 150,000 cells ml feeding level at 3-day intervals ( EL Expt.#3) .61 33 Daphnia size-frequency structure at 3-day intervals fed. with 100,000 cells ml 2 to 3 times daily ( IK Expt.#4- Repl ."l) 62 34 Daphnia size-frequency structure at 3-day intervals fed with 100,000 cells ml 2 to 3 times daily ( IL Expt.#4- Repl.2) 63 35 Daphnia size-frequency structure at 3-day intervals fed with 100,000 cells ml 2 to 3 times daily ( IL Expt.#4- Repl.3) 64 36 Daphnia size-frequency structure at 3-day intervals fed with 100,000 cells ml 2 to 3 times daily ( Expt.#5- Repl.1) 65 37 Daphnia size-frequency structure at 3-day intervals fed with 100,000 cells ml 2 to 3 times daily ( EL Expt.#5- Repl.2) 66 38 Daphnia size-frequency structure at 3-day intervals fed with 100,000 cells ml 2 to 3 times daily ( EL Expt.#5- Repl.3) 67 39 Daily total length of D.pulex at the three food concentrations (Growth Rate Expt.#1) 72 40 Daily total length of D.pulex at the three food concentrations (Growth Rate Expt.#2) 73 41 Length-weight relationship of D.pulex 74 ACKNOWLEDGEMENT My sincerest gratitude to Dr. J.W. Zahradnik, my advisor, Department of Bio-Resource Engineering. I am deeply indebted to all members of my committee: Dr. P.J. Harrison, Department of Oceanography and Dr.