Cultivation of the New Zealand Geoduck Clam
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CULTIVATION OF THE NEW ZEALAND GEODUCK CLAM, Panopea zelandica Dung Viet Le A dissertation submitted to Auckland University of Technology in fulfilment of the requirements for the degree of Doctor of Philosophy (PhD) Faculty of Health and Environmental Sciences School of Applied Sciences 2016 Abstract The geoduck Panopea zelandica has been signalled as a new emerging species for aquaculture in New Zealand. To pave the way for the establishment of a geoduck aquaculture industry, information on how to grow this species over its life cycle needs to be determined. The aim of this study was to identify conditions that optimize P. zelandica broodstock conditioning, fertilization, larval growth and metamorphosis, and juvenile and young adult growth. P. zelandica broodstock were conditioned within a combination of three water temperatures (7–8, 11–12, and 16–17°C) and three feeding rations (10,000, 50,000, and 100,000 cells mL-1 of Chaetoceros muelleri and Tisochrysis lutea, 1:1 cell counts) for 73 days. After conditioning, similar percent matured and dry condition index values were observed on geoducks among temperatures. However, significantly higher dry gonadosomatic indices (GSIdw) were recorded at 8 and 12°C. Although no difference was detected in the percentage of spawned individuals and connective tissue occupation indices, a higher percent of matured individuals was recorded when fed 10,000 and 50,000 cells mL-1. Glycogen, protein, and lipid analyses indicated that geoducks within all treatments achieved a positive energy balance, except for those in the treatments combining the highest temperature and lowest feeding ration. Comparisons of fatty acid profiles of animals among treatments and with the reference group (pond water conditioned) revealed that eicosapentaenoic (EPA, C20:5n-3), docosahexaenoic (DHA, C22:6n-3) and arachidonic (ARA, C20:4n- 6) fatty acids were important contributors to gametogenic development for geoduck conditioning. i The development of P. zelandica embryos at 15°C and 35 ppt and the optimal sperm:egg ratios for fertilization under hatchery conditions were investigated. Fertilization was conducted at sperm:egg ratios of: 50:1, 100:1, 500:1, 1000:1, and 10,000:1 with a sperm-egg contact time of 40 min. The optimal sperm:egg ratio was determined to be < 500:1 and the normal embryo yield at 3 and 18 h post-fertilization ranged from 83-96%. P. zelandica eggs (~ 80 µm diameter) developed the first and second polar bodies within 15 - 20 and 50 - 55 min post- fertilization, respectively. The blastula appeared at ~ 8 hpf, including the XR and XL cells and the presumptive shell field depression. Gastrulation occurred at 12 - 18 hpf with organic material shell apparent at the shell field depression. The mid-stage trochophore, which appeared at around 35 hpf had an apical plate with an apical tuft. The shell field spread to form the periostracum, which expanded and folded into right and left segments covering the late trochophore. The early D-stage veliger appeared at 45 hpf with the soft body being enclosed by two valves and the appearance of the velum. The physiological, morphological, and behavioral characteristics throughout the larval developmental process were determined for P. zelandica larvae, which were reared in a flow-through system at 17°C and 35 ppt. The initial veliger stocking densities ranged from 50 - 200 larvae mL-1 and geoduck larvae were fed continuously with Tisochrysis lutea and Chaetoceros calcitrans at residual algal levels of 20,000 to 80,000 cells mL-1 in three rearing batches. The larval development took 16 - 19 days from first D-veliger and metamorphosis occurred across a wide size range (300 - 375 µm shell length). The increase in shell length was linear over time and correlated with the deposition of striae in the prodissoconch II. The ingestion rate followed a power function with time and was closely correlated with the development of the alimentary system. Rearing ii with an initial stocking density of 100 larvae mL-1 and residual algal background of 20,000 cells mL-1 resulted in about 76% survival and 15 µm day-1 growth rate. The metamorphic induction of larval P. zelandica was tested with different neuroactive compounds. Two batches of competent hatchery-reared larvae were exposed to acetylcholine chloride, epinephrine hydrochloride, and excess potassium ions in the form of KCl and K2SO4 for 3 and 24 h. None of the tested chemicals increased the proportion of metamorphosed geoducks, and in some cases the chemical inhibited metamorphosis and caused significant mortality, despite having been used extensively with other species, such as mussels and oysters. The allometric coefficients (β) of respiration rate RR and clearance rate CR in P. zelandica were 0.73±0.03 and 0.62±0.07, respectively. P. zelandica juvenile and young adults were acutely thermally challanged at five different temperatures representative of potential farming conditions (8, 11, 15, 19, and 23°C). Their aerobic scope for activity and clearance rates were determined at all temperatures. Comparisons of aerobic scope for activity and clearance rates between size classes revealed that juvenile geoducks had a narrower thermal optimum than young adults (15 – 19°C versus 11 – 19°C, respectively). Temperatures higher than 19°C resulted in a reduction of aerobic scope for activity and clearance rate for both juvenile and young adults, which may lead to reduced performance and elevated mortality. P. zelandica juvenile and young adults were exposed to normoxia, mild hypoxia, and severe hypoxia. The respiration, aerobic scope, critical oxygen partial pressure (PcO2), and oxygen regulation capacity in two size classes of fed and starved animals were determined. The PcO2 was determined to be ~4 kPa for iii all geoduck groups. The respiration rates of fed small geoducks decreased significantly from normoxia (16.7 - ~21 kPa) to mild hypoxia (PcO2 - 16.7 kPa). Conversely, the respiration rates of starved geoducks from both size classes, and large fed geoducks were maintained at a constant level when exposed to the same change in oxygen concentration. However, all geoducks experienced decreased respiration rates during severe hypoxia (0 kPa - PcO2). In addition, overall oxyregulatory capacity, assessed using a regulation index, was affected by size rather than by nutritional stress. Large geoducks maintain oxygen consumption across an oxygen gradient more effectively than small geoducks. Also, the aerobic scope of small geoducks decreased significantly with declining PO2, while large geoducks maintained their aerobic scope under hypoxia. iv Table of Contents Abstract .............................................................................................................. i Table of Contents .............................................................................................. v List of Tables .................................................................................................. xiii List of Figures ................................................................................................. xv Attestation of Authorship .............................................................................. xxi Authorship Agreement Form ........................................................................xxii Acknowledgements ......................................................................................xxiv CHAPTER 1 - Introduction................................................................................ 1 1.1 General Introduction ............................................................................... 1 1.2 Literature review ...................................................................................... 5 1.2.1 Current status of geoduck hatchery knowledge and techniques . 5 1.2.2 Current status of geoduck field grow-out ..................................... 13 1.3 Present Study ........................................................................................ 15 1.4 Thesis Structure .................................................................................... 18 CHAPTER 2 - Broodstock conditioning of New Zealand geoduck (Panopea zelandica) within different temperature and feeding ration regimes .......... 28 Abstract ........................................................................................................ 29 2.1 Introduction ........................................................................................... 30 2.2 Methods and Materials .......................................................................... 33 2.2.1 Source of broodstock ..................................................................... 33 v 2.2.2 Production of microalgae ............................................................... 34 2.2.3 Conditioning experiment ................................................................ 34 2.2.4 Sampling .......................................................................................... 35 2.2.5 Histological analysis ...................................................................... 35 2.2.6 Connective tissue occupation index ............................................. 36 2.2.7 Condition index ............................................................................... 36 2.2.8 Gonadosomatic index .................................................................... 37 2.2.9 Statistical analyses ......................................................................... 37 2.3 Results ..................................................................................................