Dissertations / Theses on the topic 'Pearl oysters'

In October 2001, a new intracellular parasite was detected in Western Australian pearl oysters, Pinctada maxima. This ciliated parasite had not previously been seen in the area despite surveillance over preceding years. A study was performed to identify its characteristics and distribution. The information provided by this study could thereby assist the Pearl Producers Association and Government bodies in determining the industry’s response to infections. The morphology of the intracellular ciliate of pearl oysters was investigated using light microscopy, revealing organisms that were approximately 5 x 11 μm in size, with 9 rows of cilia and a lobulated nucleus. The ciliates were located within the apical portion of the cytoplasm of digestive epithelial cells of the digestive gland. Investigations into the ultrastructure of the ciliate using transmission electron microscopy (TEM) supported the findings made using histology. On initial morphological examination, a similarity between the pearl oyster parasite and a ciliate found in Canadian mussels was identified. Samples of Canadian mussels infected with their intracellular ciliate were analysed using light microscopy and TEM. This identified almost identical morphological characteristics to the Western Australian ciliate of pearl oysters, with a slight difference in size that could be attributed to fixation and processing. The successful culture of adductor muscle, mantle tissue and digestive gland derived cells from pearl oysters was developed and maintained in vitro for up to 75 days, using a number of varying cell culture media. This was attempted in order to provide an ability to study the live intracellular organisms in the laboratory. Contamination and slow growth were the main issues affecting the viability of the technique. An investigation into the pathology associated with the presence of the ciliate in the pearl oysters was performed. A positive correlation between the presence of the ciliate and an inflammatory response of the digestive gland was displayed using statistical techniques. Pathological changes to the infected cells were demonstrated using TEM, indicating disrupted cytoplasm, vacuolation and nuclear degeneration. Field trials placing naïve spat in pearl farm leases that had prior ciliate infections were performed. No intracellular ciliates were detected in P. maxima. Other bivalve species from the same regions were examined histologically for health and parasites. With a total of 345 bivalves surveyed during 2005 to 2007, from 8 geographical locations and over 11 species of bivalve included, one oyster contained the intracellular parasite. This oyster was a 20 mm male bastard shell (Pinctada sp.) from Gales Bay, sampled in October 2006. One protozoal parasite was identified in a novel host bivalve species, and a previously undescribed single celled organism was also discovered. An experimental design for a cross infection trial, should the ciliate become available, was completed. The sensitivity of histology to detect ciliates was also examined, revealing its sensitivity as a diagnostic tool in at risk populations was low (38%) to moderate (50%). The historical distribution of the ciliate was examined using archival records, indicating that the highest prevalences occurred in the warmer months of October to February from 2001 to 2006. This also corresponded to the months of reduced rainfall. An attempt at designing a PCR test to determine the molecular characteristics of the ciliate was performed. Formalin fixed, paraffin embedded oyster DNA was successfully extracted, amplified and sequenced, however isolation of ciliate DNA remained elusive, and may have been out-competed with host DNA. In situ hybridisation displayed positive staining with a probe designed from the Ciliophora 16s ssu gene.

Biomineralization is a process encompassing all mineral containing tissues produced within an organism. The most dynamic example of this process is the formation of the mollusk shell, comprising a variety of crystal phases and microstructures. The organic component incorporated within the shell is said to dictate this remarkable architecture. Subsequently, for the past decade considerable research have been undertaken to identify and characterize the protein components involved in biomineralization. Despite these efforts the general understanding of the process remains ambiguous. This study employs a novel molecular approach to further the elucidation of the shell biomineralization. A microarray platform has been custom generated (PmaxArray 1.0) from the pearl oyster Pinctada maxima. PmaxArray 1.0 consists of 4992 expressed sequence tags (ESTs) originating from the mantle, an organ involved in shell formation. This microarray has been used as the primary tool for three separate investigations in an effort to associate transcriptional gene expression from P. maxima to the process of shell biomineralization. The first investigation analyzes the spatial expression of ESTs throughout the mantle organ. The mantle was dissected into five discrete regions and each analyzed for gene expression with PmaxArray 1.0. Over 2000 ESTs were differentially expressed among the tissue sections, identifying five major expression regions. Three of these regions have been proposed to have shell formation functions belonging to nacre, prismatic calcite and periostracum. The spatial gene expression map was confirmed by in situ hybridization, localizing a subset of ESTs from each expression region to the same mantle area. Comparative sequence analysis of ESTs expressed in the proposed shell formation regions with the BLAST tool, revealed a number of the transcripts were novel while others showed significant sequence similarities to previously characterized shell formation genes. The second investigation correlates temporal EST expression during P. maxima larval ontogeny with transitions in shell mineralization during the same period. A timeline documenting the morphologicat microstructural and mineralogical shell characteristics of P. maxima throughout larval ontogeny has been established. Three different shell types were noted based on the physical characters and termed, prodissoconch I, prodissoconch 11 and dissoconch. PmaxArray 1.0 analyzed ESTs expression of animals throughout the larval development of P. maxima, noting up-regulation of 359 ESTs in association with the shell transitions from prodissoconch 1 to prodissoconch 11 to dissoconch. Comparative sequence analysis of these ESTs indicates a number of the transcripts are novel as well as showing significant sequence similarities between ESTs and known shell matrix associated genes and proteins. These ESTs are discussed in relation to the shell characters associated with their temporal expression. The third investigation uses PmaxArray 1.0 to analyze gene expression in the mantle tissue of P. maxima specimens exposed to sub-lethal concentrations of a shell-deforming toxin, tributyltin (TBT). The shell specific effects of TBT are used in this investigation to interpret differential expression of ESTs with respect to shell formation functions. A lethal and sublethal TBT concentration range was established for P. maxima, noting a concentration of 50 ng L- 1 TBT as sub-lethal over a 21 day period. Mantle tissue from P. maxima animals treated with 50 ng L- 1 TBT was assessed for differential EST expression with untreated control animals. A total of 102 ESTs were identified as differentially expressed in association with TBT exposure, comparative sequence identities included an up-regulation of immunity and detoxification related genes and down-regulation of several shell matrix genes. A number of transcripts encoding novel peptides were additionally identified. The potential actions of these genes are discussed with reference to TBT toxicity and shell biomineralization. This thesis has used a microarray platform to analyze gene expression in spatial, temporal and toxicity investigations, revealing the involvement of numerous gene transcripts in specific shell formation functions. Investigation of thousands of transcripts simultaneously has provided a holistic interpretation of the organic components regulating shell biomineralization.

Biomineralization is a process encompassing all mineral containing tissues produced within an organism. The most dynamic example of this process is the formation of the mollusk shell, comprising a variety of crystal phases and microstructures. The organic component incorporated within the shell is said to dictate this remarkable architecture. Subsequently, for the past decade considerable research have been undertaken to identify and characterize the protein components involved in biomineralization. Despite these efforts the general understanding of the process remains ambiguous. This study employs a novel molecular approach to further the elucidation of the shell biomineralization. A microarray platform has been custom generated (PmaxArray 1.0) from the pearl oyster Pinctada maxima. PmaxArray 1.0 consists of 4992 expressed sequence tags (ESTs) originating from the mantle, an organ involved in shell formation. This microarray has been used as the primary tool for three separate investigations in an effort to associate transcriptional gene expression from P. maxima to the process of shell biomineralization. The first investigation analyzes the spatial expression of ESTs throughout the mantle organ. The mantle was dissected into five discrete regions and each analyzed for gene expression with PmaxArray 1.0. Over 2000 ESTs were differentially expressed among the tissue sections, identifying five major expression regions. Three of these regions have been proposed to have shell formation functions belonging to nacre, prismatic calcite and periostracum. The spatial gene expression map was confirmed by in situ hybridization, localizing a subset of ESTs from each expression region to the same mantle area. Comparative sequence analysis of ESTs expressed in the proposed shell formation regions with the BLAST tool, revealed a number of the transcripts were novel while others showed significant sequence similarities to previously characterized shell formation genes. The second investigation correlates temporal EST expression during P. maxima larval ontogeny with transitions in shell mineralization during the same period. A timeline documenting the morphologicat microstructural and mineralogical shell characteristics of P. maxima throughout larval ontogeny has been established. Three different shell types were noted based on the physical characters and termed, prodissoconch I, prodissoconch 11 and dissoconch. PmaxArray 1.0 analyzed ESTs expression of animals throughout the larval development of P. maxima, noting up-regulation of 359 ESTs in association with the shell transitions from prodissoconch 1 to prodissoconch 11 to dissoconch. Comparative sequence analysis of these ESTs indicates a number of the transcripts are novel as well as showing significant sequence similarities between ESTs and known shell matrix associated genes and proteins. These ESTs are discussed in relation to the shell characters associated with their temporal expression. The third investigation uses PmaxArray 1.0 to analyze gene expression in the mantle tissue of P. maxima specimens exposed to sub-lethal concentrations of a shell-deforming toxin, tributyltin (TBT). The shell specific effects of TBT are used in this investigation to interpret differential expression of ESTs with respect to shell formation functions. A lethal and sublethal TBT concentration range was established for P. maxima, noting a concentration of 50 ng L- 1 TBT as sub-lethal over a 21 day period. Mantle tissue from P. maxima animals treated with 50 ng L- 1 TBT was assessed for differential EST expression with untreated control animals. A total of 102 ESTs were identified as differentially expressed in association with TBT exposure, comparative sequence identities included an up-regulation of immunity and detoxification related genes and down-regulation of several shell matrix genes. A number of transcripts encoding novel peptides were additionally identified. The potential actions of these genes are discussed with reference to TBT toxicity and shell biomineralization. This thesis has used a microarray platform to analyze gene expression in spatial, temporal and toxicity investigations, revealing the involvement of numerous gene transcripts in specific shell formation functions. Investigation of thousands of transcripts simultaneously has provided a holistic interpretation of the organic components regulating shell biomineralization.

The study assessed natural levels and patterns of genetic variation in Arabian Gulf populations of a native pearl oyster to define wild population structure considering potential intrinsic and extrinsic factors that could influence any wild structure detected. The study was also the first attempt to develop microsatellite markers and to generate a genome survey sequence (GSS) dataset for the target species using next generation sequencing technology. The partial genome dataset generated has potential biotechnological applications and for pearl oyster farming in the future.

This work places itself in the field of knitted textile design and the context of body and interior. The primary motive is to investigate the tactile and visual properties of oysters and pearls, inspired by Botticelli’s painting Venus. The aim is to explore free-flowing and texture through knitted three-dimensional textile surfaces. Material and colour choices have been made based on the source of inspiration, the oyster, and investigated on industrial circle knit and flat knit machines. The circle knit’s expression has been explored from a hand knitting perspective, using the manual elements to push the machine’s technique to design new expressions. The result of the project is a collection that has four suggestions for a knitted, three-dimensional surface, each inspired and developed from one specific part of the oyster; the shell, the nacre, the flesh, and the pearl. This work investigates the potential of using circle knit machines, commonly used in fast fashion for bulk production, as a tool for handicraft and higher art forms. The final collection pushes the conversation regarding the future uses of the knitting machines and investigates how rigid objects can be expressed through the flexible structure.

The pearl oyster Pinctada martensii or Pinctada fucata is the oyster for produce the South China Sea Pearl, and the production of pearl oyster Pinctada martensii plays a key role for the economic and social welfare of the coastal areas. To guarantee both rich and sustainability of providing pearl oyster productions, addressing the suitable areas for aquaculture is a very important consideration in any aquaculture activities. Relatively rarely, in the case of site selection research, the researchers use GIS analysis to identify suitable sites in fishery industry in China. Therefore, I decided to help the local government to search suitable sites form the view of GIS context. This study was conducted to find the optimal sites for suspended culture of pearl oyster Pinctada martensii using GIS-based multi-criteria analysis. The original idea came from the research of Radiarta and his colleagues in 2008 in Japan. Most of the parameters in the GIS model were extracted from remote sensing data (Moderate Resolution Imaging Spectroradiometer and Landsat 7). Eleven thematic layers were arranged into three sub-models, namely: biophysical model, social-economic model and constraint model. The biophysical model includes sea surface temperature, chlorophyll-α concentration, suspended sediment concentration and bathymetry. The criteria in the social-economic model are distance to cities and towns and distance to piers. The constraint model was used to exclude the places from the research area where the natural conditions cannot be fulfilled for the development of pearl oyster aquaculture; it contains river mouth, tourism area, harbor, salt fields / shrimp ponds, and non-related water area. Finally those GIS sub-models were used to address the optimal sites for pearl oyster Pinctada martensii culture by using weighted linear combination evaluation. In the final result, suitability levels were arranged from 1 (least suitable) to 8 (most suitable), and about 2.4% of the total potential area had the higher levels (level 6 and 7). These areas were considered to be the places that have the most suitable conditions for pearl oyster Pinctada martensii for costal water of Yingpan.

Pearl culture is the second largest aquaculture industry in terms of value in Australia. It is currently worth an estimated Aus$300 million annually and it is anticipated that the industry will be worth Aus$500 million by 2010. The Australian pearl industry is currently based on South Sea pearl production from the silver-lip pearl oyster, Pinctada maxima, for which it is world renowned. However, there has been recent interest in pearl production from two other major marine pearl oyster species, the blacklip pearl oyster, P. margaritifera, and the Akoya pearl oyster, P. fucata, which are both abundant in Australian waters. Production of Akoya pearls, until recently was dominated by the Japanese. However, recent down-scaling of the Japanese pearl oyster industry due to factors that resulted in the death of millions of oysters, has presented an opportunity for other countries to enter the Akoya pearl market. Australia is one such country which has received a lot of interest in Akoya pearl production over the last 5-10 years because of: (1) its reputation as a quality pearl producing nation; (2) the clean non-polluted waters around Australia; and (3) the wide distribution of Akoya oysters along the Australian coastline. Consequently, there was a need for biological information on which the feasibility of Akoya pearl oyster culture in Australia could be assessed. The major objective of the current project was to develop techniques to determine whether Akoya pearl oyster culture is feasible in tropical north Queensland. The results of this study will compliment the results of research with similar goals conducted in temperate Australia (New South Wales). The focus of this study was to produce Akoya pearl oysters in tropical Australia for the first time, before optimizing protocols for hatchery and nursery culture. This information was then utilized to suggest possible sites within Queensland which would be suitable for Akoya pearl oyster production, based on ‘biological performance’. The first successful culture of Akoya pearl oysters in Australia under tropical conditions produced 213 000 larvae which were transferred to settlement tanks. A total of 58 000 spat were subsequently transferred from settlement tank and resulted in 48 000 spat ranging in size from 2-30 mm at 3.5 months of age. These spat were produced using established protocols for other pearl oyster species. After 12 months, Akoya pearl oysters had a mean dorso-ventral shell height (DVH) of 56.2 ± 0.2 mm and showed superior growth rate to those reported for this species in more traditional culture regions (i.e. SE Asia). This project investigated aspects of hatchery production including embryonic and larval development to identify optimal protocols for hatchery culture of Akoya pearl oysters (Chapters 4 and 5). Full orthogonal designs were established to investigate; (1) the effects of water temperature and salinity; and (2) the effects of density and addition of antibiotics on the development of P. fucata embryos into D-stage veligers. Maximum development of P. fucata embryos into D-stage veligers occurs within a water temperature range of 26-28°C and a salinity range of 28-32‰. Further results suggested that antibiotics are not required during embryonic development of P. fucata as development of larvae was not improved in the presence antibiotics. Results have also shown that maximum development of embryos into D-stage veliger occurred when larval stocking densities were low. Suggesting an ideal stocking density is strongly dependant on the individual hatchery and the production goals. These results have obvious implications for the selection of sites for an Akoya oyster hatchery in Queensland. Ideally, a site should be selected in which water parameters are within the above-mentioned ranges. A number of pearl oyster culture techniques were investigated during this project to optimise nursery culture of Akoya pearl oysters under Queensland (tropical) conditions. These included the effects of depth, stocking density, culture apparatus and fouling on the growth and survival of pearl oysters. P. fucata spat were transferred from the hatchery to the long-line and placed in plastic mesh trays at three different depths, 2 m, 4 m, and 6 m. After 8 weeks on the long-line, spat cultured at 2 m were significantly (p<0.05) larger in DVH than spat at either 4 m or 6 m, which were not significantly different from each other. Additionally, greater numbers of ‘large’ spat were recorded when spat cultured at 2 m compared to spat cultured at either 4 m or 6 m. Hatchery-produced P. fucata spat at 3.5 months of age were graded into three size classes, ‘small’, ‘medium’ and ‘large’, which for the purpose of the study were treated as ‘slow’, ‘normal’ and ‘fast’ growers, respectively. This study aimed to determine whether growth rates differed between oysters from the above-mentioned size classes. Results from this study suggest that when oysters are first graded at 3.5 months of age (8 weeks after transfer to the ocean) slow growing oysters should not be discarded (common practice by some pearl farmers within the industry). This is because slow growers, when compared to ‘normal’ growers, only require an additional 2-4 months before reaching pearl production size. The implications of retaining slow growers is discussed. Hatchery-produced spat were cultured at different stocking densities to determine optimal growth and survival of P. fucata. Stocking densities were determined on the basis of percentage of total available net area. In Experiment 1 during early nursery culture, spat were stocked at either 25%, 50% or 75% of total available net area. Maximum growth was recorded for spat cultured at the lowest stocking density (25% of total available net area), which were significantly larger that spat cultured at either 50% or 75% of total available net area. Furthermore, spat cultured at 25% of total available net area had significantly greater numbers of spat in the medium and large size classes than spat cultured at 50% or 75% total available net area. In Experiment 2 during late nursery culture, and based on the results from Experiment 1, spat were cultured at four stocking densities (20, 25, 30 and 40% of total available net area). Similar trends to those in Experiment 1 were recorded in Experiment 2 where spat cultured at the lower stocking densities were significantly larger than spat cultured at the other stocking densities. However, the overall growth performance (Ф’) was greatest in spat cultured at the highest stocking density (40% of total available net area). Survival was not significantly different between treatments. Two experiments were conducted with hatchery-produced P. fucata spat using four different culture units to determine which culture unit supported maximum growth and survival. In Experiment 1, the four treatments used were ‘box’, ‘tray’, ‘pearl net’ and ‘pearl net with noodles’. While maximum growth, was recorded by oysters cultured in pearl nets, there was no significant difference in growth rate to oysters cultured in pearl nets with noodles; however, oysters cultured in the box treatment were significantly smaller than oysters in all other treatments. Survival of oysters in the box treatment was 47%, whereas, survival of spat cultured in the other three treatments was greater than 90%. In Experiment 2, the four treatments included ‘pearl net with small mesh’, ‘pearl net with large mesh’, ‘panel net with small mesh’, and ‘panel net with large mesh’. Maximum growth in terms of DVH was recorded for oysters cultured in panel nets with large mesh, followed by pearl nets with large mesh, pearl nets with small mesh and panel nets with small mesh. Survival was not significantly different between treatments, and all treatments recorded 85% or greater survival. Site did not affect growth and survival in the present study when P. fucata were cultured at Orpheus Island and Magnetic Island for 12 months. Although slight variations in water temperature, salinity and chlorophyll ‘a’ were recorded between the two sites, no significant differences were recorded in overall oyster growth performance (Ф’) of 3.81 and 3.82 for Orpheus Island and Magnetic Island, respectively. Site selection for pearl oyster culture is important if growth and survival are to be maximised during nursery culture. Akoya pearl oysters showed positive growth at all water temperatures experienced throughout this study; however, the range at which optimal growth occurred was between 25.1-28.1°C. Meanwhile, maximum growth occurred within salinity and chlorophyll ‘a’ ranges of 29-33‰ and 3.5-5.3 μg L-1, respectively. This project has produced biological information, which will provide a basis for the development of an Akoya pearl oyster industry in Queensland. Establishment of such an industry would compliment the current valuable pearl industry in Australia. While information generated during this study has answered a number of questions in terms of ‘biological performance’ there is, however, a requirement for further research to appraise pearl production from Akoya oysters in Queensland and factors influencing pearl quality.

Pearl culture is the second largest aquaculture industry in terms of value in Australia. It is currently worth an estimated Aus$300 million annually and it is anticipated that the industry will be worth Aus$500 million by 2010. The Australian pearl industry is currently based on South Sea pearl production from the silver-lip pearl oyster, Pinctada maxima, for which it is world renowned. However, there has been recent interest in pearl production from two other major marine pearl oyster species, the blacklip pearl oyster, P. margaritifera, and the Akoya pearl oyster, P. fucata, which are both abundant in Australian waters. Production of Akoya pearls, until recently was dominated by the Japanese. However, recent down-scaling of the Japanese pearl oyster industry due to factors that resulted in the death of millions of oysters, has presented an opportunity for other countries to enter the Akoya pearl market. Australia is one such country which has received a lot of interest in Akoya pearl production over the last 5-10 years because of: (1) its reputation as a quality pearl producing nation; (2) the clean non-polluted waters around Australia; and (3) the wide distribution of Akoya oysters along the Australian coastline. Consequently, there was a need for biological information on which the feasibility of Akoya pearl oyster culture in Australia could be assessed. The major objective of the current project was to develop techniques to determine whether Akoya pearl oyster culture is feasible in tropical north Queensland. The results of this study will compliment the results of research with similar goals conducted in temperate Australia (New South Wales). The focus of this study was to produce Akoya pearl oysters in tropical Australia for the first time, before optimizing protocols for hatchery and nursery culture. This information was then utilized to suggest possible sites within Queensland which would be suitable for Akoya pearl oyster production, based on ‘biological performance’. The first successful culture of Akoya pearl oysters in Australia under tropical conditions produced 213 000 larvae which were transferred to settlement tanks. A total of 58 000 spat were subsequently transferred from settlement tank and resulted in 48 000 spat ranging in size from 2-30 mm at 3.5 months of age. These spat were produced using established protocols for other pearl oyster species. After 12 months, Akoya pearl oysters had a mean dorso-ventral shell height (DVH) of 56.2 ± 0.2 mm and showed superior growth rate to those reported for this species in more traditional culture regions (i.e. SE Asia). This project investigated aspects of hatchery production including embryonic and larval development to identify optimal protocols for hatchery culture of Akoya pearl oysters (Chapters 4 and 5). Full orthogonal designs were established to investigate; (1) the effects of water temperature and salinity; and (2) the effects of density and addition of antibiotics on the development of P. fucata embryos into D-stage veligers. Maximum development of P. fucata embryos into D-stage veligers occurs within a water temperature range of 26-28°C and a salinity range of 28-32‰. Further results suggested that antibiotics are not required during embryonic development of P. fucata as development of larvae was not improved in the presence antibiotics. Results have also shown that maximum development of embryos into D-stage veliger occurred when larval stocking densities were low. Suggesting an ideal stocking density is strongly dependant on the individual hatchery and the production goals. These results have obvious implications for the selection of sites for an Akoya oyster hatchery in Queensland. Ideally, a site should be selected in which water parameters are within the above-mentioned ranges. A number of pearl oyster culture techniques were investigated during this project to optimise nursery culture of Akoya pearl oysters under Queensland (tropical) conditions. These included the effects of depth, stocking density, culture apparatus and fouling on the growth and survival of pearl oysters. P. fucata spat were transferred from the hatchery to the long-line and placed in plastic mesh trays at three different depths, 2 m, 4 m, and 6 m. After 8 weeks on the long-line, spat cultured at 2 m were significantly (p<0.05) larger in DVH than spat at either 4 m or 6 m, which were not significantly different from each other. Additionally, greater numbers of ‘large’ spat were recorded when spat cultured at 2 m compared to spat cultured at either 4 m or 6 m. Hatchery-produced P. fucata spat at 3.5 months of age were graded into three size classes, ‘small’, ‘medium’ and ‘large’, which for the purpose of the study were treated as ‘slow’, ‘normal’ and ‘fast’ growers, respectively. This study aimed to determine whether growth rates differed between oysters from the above-mentioned size classes. Results from this study suggest that when oysters are first graded at 3.5 months of age (8 weeks after transfer to the ocean) slow growing oysters should not be discarded (common practice by some pearl farmers within the industry). This is because slow growers, when compared to ‘normal’ growers, only require an additional 2-4 months before reaching pearl production size. The implications of retaining slow growers is discussed. Hatchery-produced spat were cultured at different stocking densities to determine optimal growth and survival of P. fucata. Stocking densities were determined on the basis of percentage of total available net area. In Experiment 1 during early nursery culture, spat were stocked at either 25%, 50% or 75% of total available net area. Maximum growth was recorded for spat cultured at the lowest stocking density (25% of total available net area), which were significantly larger that spat cultured at either 50% or 75% of total available net area. Furthermore, spat cultured at 25% of total available net area had significantly greater numbers of spat in the medium and large size classes than spat cultured at 50% or 75% total available net area. In Experiment 2 during late nursery culture, and based on the results from Experiment 1, spat were cultured at four stocking densities (20, 25, 30 and 40% of total available net area). Similar trends to those in Experiment 1 were recorded in Experiment 2 where spat cultured at the lower stocking densities were significantly larger than spat cultured at the other stocking densities. However, the overall growth performance (Ф’) was greatest in spat cultured at the highest stocking density (40% of total available net area). Survival was not significantly different between treatments. Two experiments were conducted with hatchery-produced P. fucata spat using four different culture units to determine which culture unit supported maximum growth and survival. In Experiment 1, the four treatments used were ‘box’, ‘tray’, ‘pearl net’ and ‘pearl net with noodles’. While maximum growth, was recorded by oysters cultured in pearl nets, there was no significant difference in growth rate to oysters cultured in pearl nets with noodles; however, oysters cultured in the box treatment were significantly smaller than oysters in all other treatments. Survival of oysters in the box treatment was 47%, whereas, survival of spat cultured in the other three treatments was greater than 90%. In Experiment 2, the four treatments included ‘pearl net with small mesh’, ‘pearl net with large mesh’, ‘panel net with small mesh’, and ‘panel net with large mesh’. Maximum growth in terms of DVH was recorded for oysters cultured in panel nets with large mesh, followed by pearl nets with large mesh, pearl nets with small mesh and panel nets with small mesh. Survival was not significantly different between treatments, and all treatments recorded 85% or greater survival. Site did not affect growth and survival in the present study when P. fucata were cultured at Orpheus Island and Magnetic Island for 12 months. Although slight variations in water temperature, salinity and chlorophyll ‘a’ were recorded between the two sites, no significant differences were recorded in overall oyster growth performance (Ф’) of 3.81 and 3.82 for Orpheus Island and Magnetic Island, respectively. Site selection for pearl oyster culture is important if growth and survival are to be maximised during nursery culture. Akoya pearl oysters showed positive growth at all water temperatures experienced throughout this study; however, the range at which optimal growth occurred was between 25.1-28.1°C. Meanwhile, maximum growth occurred within salinity and chlorophyll ‘a’ ranges of 29-33‰ and 3.5-5.3 μg L-1, respectively. This project has produced biological information, which will provide a basis for the development of an Akoya pearl oyster industry in Queensland. Establishment of such an industry would compliment the current valuable pearl industry in Australia. While information generated during this study has answered a number of questions in terms of ‘biological performance’ there is, however, a requirement for further research to appraise pearl production from Akoya oysters in Queensland and factors influencing pearl quality.

Pearl Oysters of twenty species belonging to four genera of the superfamily Pteriacea, and thirty eight other species of bivalves of thirty-one genera belonging to sixteen superfamilies have been examined to investigate the relationship between particular shell structures and the tissues which secrete the precursors of their organic matrices. A strictly descriptive nomenclature to adequately describe the parts of the mantles, mantle margins, and shells of pearl oysters is suggested based on morphology and histology. Inadequacy of taxonomy of the Australian pearl oysters has been revealed by this study. Shell ultrastructure and histology of the mantle and mantle margins are used to increase the number of species of the genus Pinctada. A large number of the tissues and glands thought to be involved in the secretion of the precursors of the organic matrices of the different shell layers in the animals studied are described, and where known, the roles of their secretions discussed. In particular a detailed light and electron microcopic study of some pearl oysters from Australian waters is presented and forming organic matrices of their nacreous and prismatic layers are described and illustrated. The possible roles played by the pallial blood sinus and the rami of the circum-pallial nerve in pearl oyster shell production are discussed.

Pearl oysters of the genus Pinctada include some economically important species. The taxonomy of some of the species is problematic. Phylogenetic relationship of the species in the genus is also poorly studied. In the present study, phylogenetic relationships of P. chemnitzi, P. fucata, P. margaritifera, P. maxima, P. nigra, P. radiata (from China), P. fucata martensii (from Japan), P. albina and P. imbricata (from Australia) were studied with Pteria penguin as an outgroup, and genetic variation of Chinese P. fucata, Japanese P. fucata martensii and Australian P. imbricata populations were investigated (1) to address the taxonomic confusion and phylogeny of pearl oysters, (2) to understand the genetic connections between the Chinese P. fucata, Japanese P. fucata martensii and Australian P. imbricata in west Pacific and (3) to provide information for the genetic improvement program initiated in China.
Since P. fucata, P. fucata martensii and P. imbricata are synonymous, to study the genetic differentiation and genetic variation of such widely distributed populations is helpful in understanding their genetic connections. For this purpose, five populations, three from China (Daya Bay, Sanya Bay and Beibu Bay), one from Japan (Mie Prefecture) and one from Australia (Port Stephens) were studied using AFLP technique. Three primer pairs generated 184 loci among which 91.8-97.3% is polymorphic. An overall genetic among populations and an average of 0.37 within populations (ranging from 0.35 in Japanese population to 0.39 in Beibu Bay population) were observed. Genetic differentiation among the five populations is low but significant as indicated by pairwise GST (0.0079-0.0404). AMOVA further shows that differentiation is significant among the five populations but is not significant at a broader geographical scale, among the three groups of Chinese. Japanese and Australian populations or among the two groups of Australian and north Pacific populations. The low level of genetic differentiation indicated that P. fucata populations in the west Pacific are genetically linked. Among the five populations, the Australian one is more differentiated from the others, based on both pairwise AMOVA and GST analyses, and is genetically isolated by distance as indicated by Mantel test. However, genetic differences among the three Chinese populations are not correlated with the geographic distances, suggesting that Hainan Island and Leizhou Peninsula may act as barriers blocking gene flow.
The above three wild Chinese populations in southern China were compared with the three adjacent cultured populations using AFLP markers. Three pairs of primers generated 184 loci among 179 individuals in populations from Beibu Bay, Daya Bay and Sanya Bay. A high level of genetic diversity, ranging from 0.363 in a wild population in Sanya Bay to 0.388 in a wild population in Beibu Bay, was observed within both wild and cultured populations, indicating an absence of strong bottleneck effects in the history of cultured P. fucata populations. Yet cultured populations in Sanya Bay and Beibu Bay had more fixed loci than the corresponding wild populations. Genetic differentiation in most pairwise comparisons of populations was significant. AMOVA indicated that genetic variation among populations were very low (1.77%) though significant, while more than 98% variation resided among individuals within population. These findings provide no evidence to show that hatchery practice of pearl oyster in China to date has significantly affected the genetic diversity of the cultured populations, and suggest that all populations are competent for selection. Yet the significant genetic differentiation among populations implies that any translocation of individuals for genetic improvement program should be managed with caution for the preservation of genetic diversity in natural populations.
The internal transcribed spacers (ITS1 and ITS2) of nuclear ribosomal DNA were compared among the above nine taxa, based on sequences determined by the present study and those available from Genl3ank. The phylogenetic analysis indicates that the pearl oysters studied constitute three clades: clade I with the small oysters P. fucata, P. fucata martensii and P. imbricata, clade II with P. albina, P. nigra, P. chemnitzi and P. radiata, and clade III and clade III with the big pearl oysters P. margaritifera and P. maxima forming the basal clade. Clade II is made up two subclades: clade IIA consisting of P. albina and P. nigra and clade IIB consisting of P. chemnitzi and P. radiata. The topology of the phylogenetic tree and substitution pattern of ITS sequences suggest that P. margaritifera and P. maxima are primitive species and P. chemnitzi is a recent species. The genetic divergences between clades ranged from 28% to 76.5%, and between subclades, 8.7-10.2%. In clade I, the interspecific genetic divergences ranged from 0.6% to 1.4%, and overlapped with interspecific divergences (0.6-1.1%), indicating that P. fucata, P. fucata martensii and P. imbricata may be conspecific. Based on amplified fragment length polymorphism (AFLP) markers and ITS sequences from more individuals, analyses of the populations of these three taxa also support the conclusion that Chinese P. fucata, Japanese P. fucata martensii and Australian P. imbricata are the same species, with P. fucata being the correct name. The genetic divergence between P. albina and P. nigra was also very low (1.2%), suggesting that they may represent two subspecies that can only be distinguished by shell color. The genetic divergences between P. maxima and P. margaritifera, and between clade IIA and clade IIB ranged from 8.3% to 10.2%, suggesting that they are closely related, respectively. The ITS1 sequence of P. radiata from GenBank is almost identical to that of P. chemnitzi determined in the present study, suggesting that the specimen used for the P. radiata sequence was possibly misidentified.
Yu Dahui.
"August 2005."
Adviser: Ka Hou Chu.
Source: Dissertation Abstracts International, Volume: 67-11, Section: B, page: 6125.
Thesis (Ph.D.)--Chinese University of Hong Kong, 2005.
Includes bibliographical references (p. 100-124).
Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web.
Electronic reproduction. [Ann Arbor, MI] : ProQuest Information and Learning, [200-] System requirements: Adobe Acrobat Reader. Available via World Wide Web.
Abstracts in English and Chinese.
School code: 1307.

Marine pearl culture is one of the most valuable aquaculture industries in the world with a total estimated value of about US$500 million. The major pearl producing nations are Australia and French Polynesia and, although reliable methods for hatchery culture of pearl oysters were developed in the 1980’s and 1990’s, pearl production in these countries still relies primarily on oysters collected from the wild. Generally, the cultured pearl industry, particularly the ‘black’ pearl industry, has been slow to adopt important advances in aquaculture and other relevant disciplines and is still based on ‘traditional’ methods. Nevertheless, over recent years there has been increasing interest in research relating to general culture methods for pearl oysters and pearl oyster genetics (particularly relating to pearl quality), and there has been increasing reliance on hatchery production. This study addressed important issues relating to broodstock selection, breeding cycle and egg quality in pearl oysters. Pinctada margaritifera and P. fucata were exposed to propylene phenoxetol at a concentration of 2.5 mL L-1 and benzocaine at concentrations of 250, 500 and 1200 mg L-1. Once relaxed, oysters were observed every 5 minutes to evaluate the condition of the mantle and gills. Oysters were classified as either ‘suitable saibo’ or ‘nonsuitable saibo’ depending on their suitability for use as saibo donors for pearl production. Survival of oysters in all treatments was 100%. With the exception of oysters relaxed with 250 mg L-1 of benzocaine, where no relaxation was recorded, oysters exposed to all other treatments became relaxed and showed good condition and acceptable characteristics to be used as saibo donors. To determine whether mantle tissue could be removed from oysters without mortality, P. margaritifera and P. fucata were anaesthetised with 500 mg L-1 of benzocaine and had the ventral margin of either their left, right or both left and right mantle lobes removed. Survival after 4 weeks was 100% for all treatments and oysters showed regeneration of excised mantle tissue. Mantle border grew back to almost its original extent within 60 days after excision. Muscular fibres within the new tissue were not seen until 30 days after mantle excision. Functional (secretory) abilities were presumably recovered before day 15 when conchiolin secretions and secretory cells were seen in the newly regenerated epithelia. Mantle regeneration in P. margaritifera up to 90 days after mantle excision was similar to that for P. fucata. Anaesthetised oysters can provide mantle tissue for pearl seeding and be kept alive for future uses which may include receptors for pearl production (following seeding season) as broodstock (only those providing mantle that produced high quality pearls) and possibly as multiple saibo donors. A biopsy technique to obtain gonad tissue was assessed in P. margaritifera. Prior to biopsy, oysters were anaesthetised with 2 mL L-1 of propylene phenoxetol. Three different 9 cm long biopsy needles (16, 18 and 20 gauge) with a 10 mm sample notch, were compared as a means of obtaining gonad tissue from 20 oysters. Samples were removed from each oyster using each of the 3 biopsy needles. Following the biopsy procedure, each oyster was killed and the gonad sectioned for standard histological preparation. Samples were observed microscopically to assess gonad condition and to compare samples taken using biopsy with those taken using destructive sampling. Oysters showed 100% recovery from the anaesthetic and biopsy procedure after 2 weeks. Non-destructive biopsy sampling was an accurate means of assessing gonad condition in male pearl oysters. However, the use of thicker biopsy needles (e.g. 14 or 12 gauge) may allow better interpretation of gonad stage, particularly in female oysters for which results showed that 16 gauge biopsy needles (the thickest used in this study) were unsatisfactory. Collection of oysters from culture stock held at Magnetic Island and Orpheus Island was conducted from August 2003 to February 2004 to describe biochemical and histological changes associated to gamete development. Six oysters from each site were collected every month for seven months. Samples of mantle, adductor muscle, gonad and digestive gland were obtained for biochemical analyses. Two spawning peaks (winter and summer) were confirmed for P. margaritifera in north Queensland. The adductor muscle played an important role in storage of protein and carbohydrate during gonad development. To analyse changes in micronutrient composition during embryological development of P. margaritifera, samples of eggs, embryos and larvae were taken for GHWHUPLQDWLRQRIFDURWHQRLGDVFRUELFDFLGYLWDPLQ& -tocopherol (vitamin E) and fatty acid content 0, 4, 8, 12, 16, 24 and 46 h after fertilisation. Carotenoids were found only in trace amounts in P. margaritifera eggs and probably do not have an important role in embryo development. The vitamin C and vitamin E contents of P. margaritifera embryos increased with time and may not be limiting nutrients for embryological development. The fatty acids 14:0, 16:0 and most unsaturated C18s were highly utilised during embryological development of P. margaritifera as demonstrated by their decline during development. These fatty acids may be good indicators of egg quality. Much of the research in this study was conducted for the first time with pearl oysters. This study describes new and novel information relating to the breeding cycle, broodstock selection and egg quality of pearl oysters. The results of this study provide a basis for more efficient culture methods and may facilitate significant changes to traditional aspects of pearl oyster culture and pearl production.

Marine pearl culture is one of the most valuable aquaculture industries in the world with a total estimated value of about US$500 million. The major pearl producing nations are Australia and French Polynesia and, although reliable methods for hatchery culture of pearl oysters were developed in the 1980’s and 1990’s, pearl production in these countries still relies primarily on oysters collected from the wild. Generally, the cultured pearl industry, particularly the ‘black’ pearl industry, has been slow to adopt important advances in aquaculture and other relevant disciplines and is still based on ‘traditional’ methods. Nevertheless, over recent years there has been increasing interest in research relating to general culture methods for pearl oysters and pearl oyster genetics (particularly relating to pearl quality), and there has been increasing reliance on hatchery production. This study addressed important issues relating to broodstock selection, breeding cycle and egg quality in pearl oysters. Pinctada margaritifera and P. fucata were exposed to propylene phenoxetol at a concentration of 2.5 mL L-1 and benzocaine at concentrations of 250, 500 and 1200 mg L-1. Once relaxed, oysters were observed every 5 minutes to evaluate the condition of the mantle and gills. Oysters were classified as either ‘suitable saibo’ or ‘nonsuitable saibo’ depending on their suitability for use as saibo donors for pearl production. Survival of oysters in all treatments was 100%. With the exception of oysters relaxed with 250 mg L-1 of benzocaine, where no relaxation was recorded, oysters exposed to all other treatments became relaxed and showed good condition and acceptable characteristics to be used as saibo donors. To determine whether mantle tissue could be removed from oysters without mortality, P. margaritifera and P. fucata were anaesthetised with 500 mg L-1 of benzocaine and had the ventral margin of either their left, right or both left and right mantle lobes removed. Survival after 4 weeks was 100% for all treatments and oysters showed regeneration of excised mantle tissue. Mantle border grew back to almost its original extent within 60 days after excision. Muscular fibres within the new tissue were not seen until 30 days after mantle excision. Functional (secretory) abilities were presumably recovered before day 15 when conchiolin secretions and secretory cells were seen in the newly regenerated epithelia. Mantle regeneration in P. margaritifera up to 90 days after mantle excision was similar to that for P. fucata. Anaesthetised oysters can provide mantle tissue for pearl seeding and be kept alive for future uses which may include receptors for pearl production (following seeding season) as broodstock (only those providing mantle that produced high quality pearls) and possibly as multiple saibo donors. A biopsy technique to obtain gonad tissue was assessed in P. margaritifera. Prior to biopsy, oysters were anaesthetised with 2 mL L-1 of propylene phenoxetol. Three different 9 cm long biopsy needles (16, 18 and 20 gauge) with a 10 mm sample notch, were compared as a means of obtaining gonad tissue from 20 oysters. Samples were removed from each oyster using each of the 3 biopsy needles. Following the biopsy procedure, each oyster was killed and the gonad sectioned for standard histological preparation. Samples were observed microscopically to assess gonad condition and to compare samples taken using biopsy with those taken using destructive sampling. Oysters showed 100% recovery from the anaesthetic and biopsy procedure after 2 weeks. Non-destructive biopsy sampling was an accurate means of assessing gonad condition in male pearl oysters. However, the use of thicker biopsy needles (e.g. 14 or 12 gauge) may allow better interpretation of gonad stage, particularly in female oysters for which results showed that 16 gauge biopsy needles (the thickest used in this study) were unsatisfactory. Collection of oysters from culture stock held at Magnetic Island and Orpheus Island was conducted from August 2003 to February 2004 to describe biochemical and histological changes associated to gamete development. Six oysters from each site were collected every month for seven months. Samples of mantle, adductor muscle, gonad and digestive gland were obtained for biochemical analyses. Two spawning peaks (winter and summer) were confirmed for P. margaritifera in north Queensland. The adductor muscle played an important role in storage of protein and carbohydrate during gonad development. To analyse changes in micronutrient composition during embryological development of P. margaritifera, samples of eggs, embryos and larvae were taken for GHWHUPLQDWLRQRIFDURWHQRLGDVFRUELFDFLGYLWDPLQ& -tocopherol (vitamin E) and fatty acid content 0, 4, 8, 12, 16, 24 and 46 h after fertilisation. Carotenoids were found only in trace amounts in P. margaritifera eggs and probably do not have an important role in embryo development. The vitamin C and vitamin E contents of P. margaritifera embryos increased with time and may not be limiting nutrients for embryological development. The fatty acids 14:0, 16:0 and most unsaturated C18s were highly utilised during embryological development of P. margaritifera as demonstrated by their decline during development. These fatty acids may be good indicators of egg quality. Much of the research in this study was conducted for the first time with pearl oysters. This study describes new and novel information relating to the breeding cycle, broodstock selection and egg quality of pearl oysters. The results of this study provide a basis for more efficient culture methods and may facilitate significant changes to traditional aspects of pearl oyster culture and pearl production.

French Polynesia relies solely on the collection of wild Pinctada margaritifera spat for pearl oyster culture. This was developed to help protect the wild populations from overexploitation, but it is feared that massive spat collection could lead to erosion of genetic diversity both in farmed and wild stocks. Wild and farmed collections of P. margaritifera from four atolls in French Polynesia were genotyped at eight microsatellite loci to determine whether there was a loss of genetic diversity from the wild to adjacent farmed aggregations. The average allelic richness for wild samples was not significantly different from that seen for farmed samples, but there was a significant effect of atoll and locus. Pair-wise genetic differentiation (FST) was not significant between adjacent wild and farmed collections or across atolls. Overall there was no evidence for a loss of genetic variability in farmed oysters. Both farmed and wild individuals analyzed here were adults and could have originated from multiple spawning events in time and space. This could have masked genetic processes linked to recruitment happening at a finer scale. P. margaritifera demonstrates high recruitment variability, but the number of parents contributing to a successful cohort of juveniles recruited on collectors is unknown. Low effective number of breeders and variable recruitment are assumed to be responsible for the genetic patchiness that has been observed at a small spatial scale for this species and this could lead to a loss of genetic diversity in both the farmed and wild stocks. The genetic diversity and family make-up of three groups of 1.5 year old oysters were assessed using 13 microsatellite markers. These individuals were harvested on collectors in three closely located sites of the Takapoto atoll. Higher recruitment density and higher allelic richness was observed in one zone compared to the other two. Significant genetic differentiation was also observed at a small spatial scale. Pair-wise FST estimates between collectors within zone were not significant, but were generally significant across zones. Estimates of effective population size and number of families present for these individuals were larger than expected and suggested that the numbers of parents contributing to the recruits on these collector lines were not limited. Similar results were obtained when assessing monthly cohorts of recruits collected in Takapoto over 5 months with 11 microsatellites. Levels of allelic richness were not significantly different among monthly cohorts, and were comparable to the levels observed in the adult samples above. Small but significant temporal genetic differentiation was observed between the monthly cohorts. Again, there was no evidence for low effective population size or for significant family structuring and it did not appear that a limited number of parents produced these temporal cohorts. Patchy genetic structure was observed, but recruitment on collectors does not seem to be driven by a limited number of successful parents. It does not appear that the current pearl culture practices are negatively impacting the local farmed and wild stocks of P. margaritifera in French Polynesia by reducing their levels of genetic diversity.

Pearl culture has traditionally relied on collecting juvenile pearl oysters (family: Pteriidae) from the wild and growing them to an appropriate size to be used in pearl production; a method that has become increasingly less viable due to a corresponding depletion in wild populations. Hatchery propagation of juvenile pearl oysters is now a necessity in regions where collection from the wild can no longer sustain commercial pearl production. The cultured pearl industry in the Indo-Pacific includes production of half-pearls or 'mabè' from Pteria penguin (Röding 1798). Pearl production from P. penguin has become progressively more reliant on hatchery culture of oysters; however, efficient production is constrained by a lack of knowledge regarding optimal culture techniques. This study aimed to develop hatchery culture techniques that could be implemented by industry to improve the prospects for pearl production from P. penguin and decrease the fishing pressure currently placed on wild populations. The natural spawning season for P. penguin typically spans only a few months of each year, providing a short window for hatchery production of juveniles. A lack of knowledge regarding the diet and conditions required to optimise energy uptake by adult P. penguin has impeded the development of brood-stock conditioning programs that could be used to encourage gametogenesis outside of the natural spawning season. This study examined the pattern of suspension feeding by P. penguin in response to variations in microalgae diet, food concentration and water temperature. Brood-stock were placed in temperature controlled flow-though chambers that supplied individual oysters with a constant concentration of suspended microalgae. Feeding behaviour was quantified as the rate at which water was cleared of algae cells (clearance rate; CR) and the fraction of organic carbon absorbed during digestion (absorption efficiency; AE). The results showed that CR was greater when feeding on the two flagellate species Isochrysis sp. Tahitian (T-Iso) and Pavlova sp. (mean = 32 L h⁻¹ oyster⁻¹) when compared to the diatom Chaetoceros muelleri (27 L h⁻¹ oyster⁻¹). At temperatures of 24-28°CP. penguin maintained a stable CR with increasing food availability up to the maximum concentration tested (50 x 10³ cells mL⁻¹). Mean AE was highest for T-Iso (61%) and not influenced by food concentration. Decreased CR and AE in response to a rapid reduction in water temperature during summer, reflected the 35% lower CR and 47% lower AE observed during the colder austral winter, suggesting temperature contributes to differences in suspension feeding between seasons. The high feeding capacity of P. penguin raised the issue that in order to undertake brood-stock conditioning, hatcheries would require the facilities to culture large volumes of live microalgae. Pearl farms that use P. penguin are typically located in regional areas where the technical capacity for mass algae culture is not available. An experiment was conducted to assess the viability of using commercially available concentrated microalgae and a unique flow-though aquarium system to condition brood-stock prior to the natural spawning season. Fifteen P. penguin of a similar size were distributed between 5 identical 30 L 'flow-though aquaria'. A mixed diet of concentrated microalgae from the Instant Algae® range was supplied to brood-stock for a period of 40 days with periodic increases (10 day intervals) in food concentration and water temperature up to a maximum of 40 x 10 cells mL⁻¹ and 28°C. Histological examination of gonad tissue was conducted at the conclusion of the study so that the reproductive condition of each oyster could be categorized using the five stages for pearl oyster gonad development, ranging from inactive to ripe. The same process was conducted for 15 similar sized P. penguin that were held in ocean culture for the same time period. The reproductive state of conditioned animals suggested that male P. penguin produced spermatozoa at a rate exceeding that observed in a wild environment over the same time period. The production of mature oocytes in experimental females was less reliable, attributable to the period for conditioning being too short for the production of energetically expensive ooyctes. In order to develop techniques for hatchery culture of juvenile P. penguin, it was first necessary to understand the processes of embryogenesis and larval development for this species. Following standard methods used in the hatchery culture of other pearl oyster species, P. penguin eggs were spawned, fertilized and incubated until they hatched into shelled veliger larvae. Larvae were then fed a diet of live microalgae until developing a foot and being deemed competetent to settle. Embryos and larvae were sampled periodically during hatchery culture to be examined under scanning electron microscope (SEM). The resulting high resolution images were then used to map the approximate timing of developmental stages. These stages included the first cleavage (1 h post-fertilisation; hpf), morula (2.5 hpf), blastula (4.5 hpf), gastrula (5.5> hpf), trochophore (7> hpf), D-stage (20 - 22 hpf), prodissoconch II (3 - 6 days post-hatching into D-stage; dph), umbone (10 - 12 dph) and pediveliger (22 dph). Comparison with patterns of embryogenesis and larval development in other oviparous oyster species revealed a similar sequence of key events, with differences occurring in the timing of developmental stages, shell structure and shell shape. Embryo incubation is a period of pearl oyster culture that is typically characterised by excessive mortality. This study addressed the issue of embryo mortality by examining the effects of egg stocking density and the application of antibiotics during incubation. A factorial experimental design combined three egg densities (10, 50 and 100 mL⁻¹) and three antibiotic treatments (Control - no antibiotic; 5 mg mL⁻¹ streptomycin-sulfate; 5 mg mL⁻¹ tetracycline:erythromycin 2.5:2.5 mg mL⁻¹). Antibiotics were added to the culture medium as a single dose and fertilised eggs were incubated for a period of 24 h. Tetracycline:erythromycin (1:1) improved mean survival (23%), but yielded an average of only 9% more veliger larvae than control aquaria due to interference with development. The antibiotic streptomycin-sulfate improved mean survival by 16% when compared to control aquaria, without significantly compromising development. A high egg density of 100 mL⁻¹ did not significantly reduce survival, but resulted in a 5% reduction in normal development to D-stage. It is recommended that eggs be stocked at a density ≤50 mL⁻¹ and mortality be minimised by treating the culture medium with the antibiotic streptomycin-sulfate. Hatchery culture in regional areas is often impossible because farms cannot afford the facilities required to produce the live microalgae used as a food source for larvae. Concentrated algal paste supplied by Instant Algae® has been successfully trialled as an alternative food source during hatchery culture of P. penguin, but the feeding regime that promotes optimal larval growth and development is yet to be determined. Experiment 1 assessed the combined effects of stocking density and feed ration on the survival and growth of P. penguin larvae during D-stage (1-8 days post-fertilisation). Experiment 2 examined the effects of the same treatments on the survival and growth of larvae during umbo-stage (8 - 17 days post-fertilisation). Both experiments used a factorial design combining 3 egg stocking densities (Experiment 1: 2, 6 and 10 larvae mL⁻¹; Experiment 2: 1, 3 and 5 larvae mL⁻¹) and 3 levels of feed ration (Experiment 1: 5, 10 and 15 x 10³ cells mL⁻¹; Experiment 2: 10, 15 and 20 x 10³ cells mL⁻¹). Survival during D-stage was significantly enhanced (by 105%) in aquaria stocked at <10 larvae mL⁻¹, whereby a density of 6 mL⁻¹ maximised larval production per volume of culture medium. An intermediate feed ration of 10 x 10³ cells mL⁻¹ maximised both survival and growth during D-stage. Increasing the initial stocking density of umbostage larvae from 1 to 3 mL⁻¹ resulted in a significant reduction of both survival (360%) and growth (16%). Growth of umbo-stage larvae stocked at 1 mL⁻¹ increased significantly (7%) when feed ration remained below 20 x 10³ cells mL⁻¹. Optimising the rate of larval settlement during pearl oyster hatchery cultivation is critical to maximising the production of juvenile spat for commercial use and is reliant on providing suitable stimuli. This study used two experiments to investigate the effects of (1) treating the culture medium with alternate concentrations of three chemical compounds (Serotonin; GABA; KCl) both in the presence/absence of a bio-film and (2) exposure to five substrate types (red nylon mesh with 5 mm and <1 mm pore sizes; black fibreglass mesh with 3mm and 1mm pore sizes; transparent smooth plastic) both in the presence/absence of a chemical cue (KCl), on recruitment of P. penguin pediveliger larvae. After 48 h, settlement was 65% greater in aquaria containing a substrate covered by a naturally formed bio-film than in control aquaria. After 72 h, settlement of larvae in aquaria treated with serotonin (10⁻³M) or KCl (20 mM) was significantly greater than in control aquaria by 75% and 84%, respectively, while exposure to GABA had no effect. Settlement in response to 20 mM KCl was enhanced by the presence of a red nylon mesh substrate with 5 mm pore size. The findings of this PhD project provide practical knowledge regarding techniques for efficient hatchery culture of P. penguin. The specific aims of this study place emphasis on facilitating hatchery propagation in regional communities within the Indo-Pacific. This research will aid in increasing pearl production from hatchery bred P. penguin and therefore alleviate much of the pressure currently being placed on overexploited wild populations.

The silver-lip (or gold-lip) pearl oyster, Pinctada maxima (Jameson) is the most highly prized of all pearl oysters. Through production of "south sea pearls", P. maxima is the basis of Australia's most lucrative aquaculture industry and is of great economic importance throughout south east Asia. In recent years there has been a rapid expansion in the cultivation of P. maxima throughout the region brought about by improvements in hatchery technology. Despite this, there is limited published information concerning rearing of P. maxima. This study aims to address this problem by establishing appropriate culture criteria for P. maxima in the specific areas of collection (settlement) and maintenance of spat in the hatchery, and the early nursery rearing and the grow-out of P. maxima at sea. In the first of a series of settlement experiments, the influence of pediveliger stocking density on settlement and subsequent growth of spat was investigated. Pediveligers were placed in 15 L vessels at densities of 0.5, 1.0, 1.5 or 2.0 larvae per mL. The number of spat per 100 cm² in the 1.0 larva per mL treatment was over double, and significantly greater (P< 0.05) than that in the 0.5 larva per rid, treatment. Increases in the number of spat were not significant (P >0.05) between larval densities of 1.0 and 1.5 larvae per mL and 1.5 and 2.0 larvae per mL. Stocking density did not significantly influence survival (P>0.05). By day 44, spat from the 0.5 larva per mL treatment were significantly larger (P <0.05) than spat from all other treatments and spat in the 1.0 larva per mL treatment were significantly larger (P <0.05) than in the 1.5 or 2.0 larvae per mL treatments. Differences in size were most likely due to competition for space and food. Further settlement experiments assessed artificial substrata for the collection of hatchery-reared P. maxima. Pediveliger larvae were settled onto collectors made from: curved PVC slats; polypropylene rope; a combination of PVC slats and polypropylene rope; and mono-filament nylon. Rope and the combined PVC slat and rope collectors had significantly more spat (P <0.05) than either nylon or PVC slat collectors. In a second experiment, significantly more spat (P <0.001) were counted on horizontally positioned PVC slat collectors than those vertically positioned. The concave surface of PVC slats had significantly more (P <0.001) spat than the convex surface, regardless of orientation. In a third experiment, significantly more (P <0.001) spat attached to PVC slats with an epifloral biofilm than clean PVC slats. An adequate diet for settled spat is essential and seventy-five day-old P. maxima were fed for 21 days on the following monospecific micro-algal diets: Isochrysis aff. galbana, (T-ISO), Pavlova lutheri, Chaetoceros muelleri, C. calcitrans, and Tetraselmis suecica. The largest increase in ash free dry weight (AFDW) was for spat fed C. muelleri, which was significantly greater (P <0.05) than for any other species. The mean AFDW of spat fed T suecica and T-ISO did not differ significantly from each other, but were significantly greater than for spat fed C. calcitrans and P. lutheri (P <0.05). The final AFDW of spat fed P. lutheri was not significantly different from that of unfed spat (P >0.05). P. maxima that were ready for transfer to sea were re-settled onto PVC slats at a mean density of 340 per 100 cm² and either left exposed (control) or covered with a mesh sleeve of varying aperture sizes (0.75 mm, 1.5 mm or 3.0 mm) before being suspended from a raft. Two weeks later, there was no significant difference (P >0.05) among the number of spat retained on the covered slats; however, all covered slats had significantly greater (P <0.001) spat retention than controls. Spat were significantly larger (P <0.05) with each increase in mesh size. There was no advantage in using sleeves with a mesh size small enough to retain dislodged. Spat require detaching from their point of attachment during grading. The following stress factors were tested as potential inducers of detachment in P. maxima: salinities of 45 ‰, 40 ‰, 30 ‰ and 25 ‰; pH of 10 and 4, and air exposure. High mortality (>80%) resulted from the use of pH 10 and it was abandoned after 1 h. Hypersaline sea water (45 ‰) resulted in significantly more spat detaching (92.3 ± 0.6 %, mean ± s.e., P<0.05) than in any other treatment. A pH of 4 resulted in 85.6 ± 2.3 % (mean ± s.e.) detaching after 1 h. Exposure to the treatments beyond 1 h, except in the case of exposure to air, did not yield significant increases (P> 0.05) in numbers of detached spat. Spat that had detached in the treatment baths after the first hour began to re-attach during the second hour. After 24 h exposure to treatments, excluding pH 10 and air exposure, spat had firmly re-attached with 100 % survival and no mortality was recorded 24 h after the spat were returned to normal sea water. Byssus regeneration following detachment was studied in P. maxima of six different age classes. In the first experiment, 75 or 120 day old P. maxima were removed from their points of attachment by severing the byssus and byssal thread production and oyster behaviour were monitored for 120 hours. Younger juveniles re-attached faster than older juveniles, but older juveniles produced significantly more (P <0.001) byssal threads after 12 h and significantly more (P <0.001) byssal threads over the 120 h period. Byssus production for the younger juveniles did not increase significantly (P >0.05) after 48 h whereas byssus production from older animals continued to increase significantly (P <0.001). P. maxima were observed to eject the byssal apparatus, move and reattach within 24 h. Re-attachment following ejection of the byssus was faster than that following mechanical severing. In the second experiment, older P. maxima aged 7, 9, 11 or 13 months were placed in nets in strong (2.5-3.5 knots per h) or mild (<1 knot per h) current. Pearl oysters re-attached faster in the mild current. However, after 5 days, oysters aged 13 and 11 months in strong current had produced significantly more threads (P< 0.05) than oysters in mild current. This trend continued after day 5, but was not significant (P>0.05) for pearl oysters aged 9 and 7 months. By day 11, 9-month-old oysters had produced significantly more byssal threads than any other age class. P. maxima are grown in tropical regions affected by monsoonal rains which can alter salinity for extended periods. Juvenile P. maxima were held over a period of 20 days in the following salinities: 45 ‰, 40 ‰, 34 ‰ (ambient), 30 ‰ and 25 ‰. There was no significant difference (P> 0.05) in survival of spat from the different treatments; however, growth was significantly depressed (P< 0.05) at 45 ‰, 40 ‰ and 25 ‰. The best growth was recorded at 30 ‰, where spat were significantly larger (P< 0.05) than those held at ambient salinity. Several experiments investigated the effects of stocking density directly on growth and survival of juvenile P. maxima. Spat were held in suspended nursery culture for six weeks at four stocking densities: 10 juveniles per slat (133 juveniles per m²); 50 juveniles per slat (670 juveniles per m²); 100 juveniles per slat (1330 juveniles per m²) and 150 juveniles per slat (2,000 juveniles per m²). Best growth and survival was recorded at a stocking density of 10 juveniles per slat (80 ± 4.36%: mean ± s.e.), which was significantly higher than the other densities tested (P < 0.05). Survival did not differ significantly between the other densities tested (P >0.05). The incidence of growth deformities increased with increasing stocking density. In a second experiment, juvenile growth was compared at two stocking densities (28 individuals per net: 66 oysters per m² or 48 individuals per net: 99 oysters per m²) with animals held either in suspended or bottom culture. Mean (± s.e.) survival in 28-pocket nets in suspended culture (99.0 ± 1.6 %) was significantly better than any other treatment (P <0.01). Survival was also high in the 48-pocket nets in suspended culture (94.8 ± 3.6 %). Mean survival in bottom culture was significantly lower (P <0.05), being 15.8 ± 7.8 % and 13.3 ± 3.6 %, respectively, for 28 and 48-pocket nets. Oysters held in suspended culture grew significantly larger (P <0.001) than those in bottom culture. In both suspended and bottom culture, P. maxima in the 28-pocket nets grew significantly larger (P <0.001) than those held in 48-pocket nets. The dry weight of suspended solids, and phytoplankton number and diversity were all greater in surface waters indicating greater food availability. In a third experiment, seven month-old P. maxima were graded into four size classes (G1 to G4: largest to smallest, respectively). Three replicates for each size class were stocked into 28-pocket nets and 8-pocket nets (19 pearl oysters per m2). Dorso-ventral shell height (SH), SL and wet weight (WW) were measured monthly for five months. For G2 to G3 no differences in survival or growth were recorded during the experiment. Survival for GI in 8-pocket panels was 100% and significantly better (P<0.01) than GI in 28-pocket panels. Further, by the end of the second month, GI in 8-pocket panels were significantly larger (P<0.001) than G1 in 28-pocket panels. This size advantage was maintained during the course of the experiment. The highest percentage of `runts' resulted from G1 in 28-pocket panels. Cleaning is the major activity on pearl farms. A comparison was made of the growth of one-year-old P. maxima, cleaned every 2, 4 or 8 weeks or after 16 weeks. The diversity of fouling animals was recorded and their dry weight (DW) estimated. The DW of fouling animals increased steadily over the first 10 weeks of the experiment before declining during weeks 10 to 16. Significant (P< 0.05) differences in the DW of fouling animals between treatments was observed and pearl oyster growth was affected by fouling. SH, SL and WW of pearl oysters cleaned every 2 or 4 weeks was significantly greater (P <0.05) than that of pearl oysters cleaned every 8 weeks or after 16 weeks.

The most famous round pearls of the world come from pearl oysters of the genus Pinctada; 'south-sea pearls' are cultured using the silver- or gold-lip pearl oyster, Pinctada maxima, and 'black pearls' are cultured from the black-lip pearl oyster, P. margaritifera. Marine pearl culture is one of the most valuable aquaculture industries in the world with an estimated value of $US475 millions. P. maxima is the basis of Australian pearl industry while P. margaritifera supports cultured pearl industries in southern Japan and the Pacific island nations of French Polynesia and Cook Islands. In French Polynesia, pearl production has an annual value of US$170 millions and, not surprisingly, other nations across the Pacific have shown interest in developing similar industries. The usual way to obtain oysters for pearl farms in Polynesia is by collection of wild spat (or juveniles), activity that relies entirely on natural recruitment which can be unpredictable and unreliable. Hatchery production of pearl oyster is increasing in significance as a source of oyster stock for pearl production and offers many advantages over the collection of oysters from the wild. Hatchery production of pearl oysters is a relatively new field and, as such, techniques and protocols adopted are generally based on those developed for other bivalve species of commercial importance, usually of temperate origin. Much of the research on hatchery production of pearl oyster has focused on P. margaritifera and has covered aspects such as culture systems, feeding and nutrition of larvae and early nursery culture. Microalgae culture is fundamentally important to commercial hatcheries rearing marine molluscs, since they are currently the only suitable food source. Microalgae provided to larvae during culture can affect their growth and survival. Only recently have significant numbers of tropical microalgae species become available to the aquaculture industry. However, many of these have not been assessed for their nutrient content or their nutritional value for culture animals. Microalgae of tropical origin are likely to be better suited to the culture conditions used for tropical species, such as P. maxima and P. margaritifera. This study assessed the nutritional value of eight tropical microalgae species for P. margaritifera larvae. Each was analysed for carbohydrate, lipid and protein content as well as fatty acid and amino acid composition. Each species of microalgae was fed singly to early (D-stage veliger, Chapter 3 (section 3.3.1)) and later (umbo-stage veliger, (section 3.3.2)) P. margaritifera larvae. The results showed Pavlova sp. and P. salina to be the most nutritious of eight microalgae fed to both D-stage and umbone larvae. Relationships between the levels of various nutritional components of microalgae and resulting larval growth were determined. A significant correlation between the growth of D-stage larvae and total protein, lipid and carbohydrate contents of microalgae was found. However, for umbo-stage larvae only carbohydrate contents in microalgae was positively correlated to larval growth. Significant positive correlation between the saturated fatty acid (SFA) and polyunsaturated fatty acid (PUFA) contents of microalgae and larval growth were seen for both larval stages. Bivalve larvae are generally fed a mixture of microalgae species in order to provide a better nutrient balance. This study assessed the more nutritious species of microalgae from Chapter 3 in binary and ternary combinations. D-stage P. margaritifera larvae were fed Pav. salina, Pavlova sp, TISO and Micromonas pusilla in binary and ternary combinations, Chapter 4 (section 4.3.1). Umbo-stage larvae were fed the best binary algae combinations (flagellates) to which one diatom species (either Chaetoceros muelleri, Chaetoceros sp. or Skeletonema sp.) was added per combination (section 4.3.2). The best two ternary algae combinations from the first experiment in Chapter 4 with Dstage larvae (flagellates only), were also used to feed umbo-stage larvae in the second experiment in Chapter 4; these were Pav. salina/ Pavlova sp/TISO and TISO/M. pusilla/Pavlova sp. Greater growth rate was shown by D-stage P. margaritifera larvae fed the ternary combination of Pavlova sp/Pav.salina/TISO followed by the binary combination of Pavlova sp./M. pusilla; however, larvae fed Pavlova sp. as a monospecific diet performed as well as those fed the combination of Pavlova sp/Pav. salina/M. pusilla. Umbo-stage larvae fed ternary combinations containing a diatom showed noticeably greater growth rates than larvae fed combinations without diatoms. The best binary and ternary microalgae combinations for umbo-stage larvae were Pavlova sp./C. muelleri and Pavlova sp./M. pusilla/Skeletonema sp., respectively. However, the growth rate of larvae fed the binary combinations of Pavlova sp./C. muelleri did not differ significantly from those of larvae fed ternary diet combinations. Experiments in Chapters 3 and 4 identified microalgae supporting the greatest growth rates of D-stage and umbo-stage larvae of P. margaritifera. Biochemical analysis of these microalgae (Chapter 2) allowed identification of nutrients within the microalgae that correlate to good growth rates. Chapter 5 investigated the possibility that manipulation of culture conditions could be used as a means of improving (increasing) the levels of these key nutrients in microalgae. Such a development would have clear benefits in the culture of P. margaritifera larvae. TISO, Pav. salina, Pavlova sp., M. pusilla and C. muelleri were cultured under different light regimes. All microalgae tested were easy to grow. Under the culture conditions used during this experiment, photoperiods of 18 hours light and 6 hours dark (18L:6D) and continuous light resulted in greater productivity of algae cultures. Differences in proximate compositions of microalgae were not significant in terms of growth phase; however, general increases in lipid contents of microalgae between the logarithmic and stationary phase growth phases were observed as reported in similar studies. Harvesting microalgae during the stationary growth phase will provide microalgae with high lipid and protein values. Changes in fatty acid contents were species specific; however contents were not significantly different when algae were culture under each light regime. In summary, this study is the first comprehensive assessment of the nutritional value of tropical microalgae species for pearl oyster larvae and the first to relate larval growth rates and survival to the nutrient composition of microalgae. This study has identified species of microalgae that are highly nutritious for P. margaritifera larvae, and the nutrients that impart high nutritional value to them. Assessment of growth rates and changes in the biochemical compositions of microalgae cultures of different ages and under varying culture conditions will allow tropical microalgae to be cultured according to a regime which not only maximizes their productivity, but optimizes the nutritional composition for P. margaritifera larvae. On this basis, the results of this study provide a basis for development of more effective larval culture techniques for P. margaritifera and larvae of different ages.

碩士
國立臺南藝術大學
造形藝術研究所
107
The study focused on four series of works created by myself between 2015 and 2017. It attempted to discuss the statuses of objects in my works and organized the issue of how “images” and “space” influenced the appearances of my works with “pearl-oyster,” an image that perfectly mixes various contradictory properties. In the process of writing the thesis, I depicted each image of pearl-oyster respectively: pearls, shells and flesh, flesh and shells as if I kneaded and shaped them with the hands of words. Last but not the least, I hoped to construct a complete image of pearl-oyster to enable readers to read the thesis as if they are viewing how each work is created at the same time.

Pearl oysters are not only farmed for their gemstone quality pearls worldwide, but are also becoming important model organisms for investigating genetic mechanisms of biomineralisation and bivalve evolution. However, despite their economic and scientific significance, limited genomic resources are available for this important group of bivalves, hampering investigations aiming to identify genes that regulate important pearl quality traits and unique biological characteristics (i.e. biomineralisation). The silver-lipped pearl oyster, Pinctada maxima, is one species where there is interest in understanding genes that regulate commercially important pearl traits, but presently there is a dearth of genomic information. Recently, the market for the highly valuable South Sea pearl produced by P. maxima has expanded, creating strong interest in stock improvement to increase the production of high quality South Sea pearls. Molecular stock improvement techniques such as marker assisted selection (MAS) have great potential in accelerating selective breeding programs for animal production industries as they bypass limitations that have so far restricted the success of phenotypic breeding programs. Genetic breeding programs not only require accurate phenotypes, but they also need established high density genomic resources including genome-wide molecular markers and dense genomic maps, of which none are available for any pearl oyster species. These resources allow the dissection of complex phenotypic traits of commercial importance and the identification of genetic variation and marker associations that disproportionately influence variation in these traits, both of which are integral before genetic breeding programs can be established. Once marker associations to traits are identified, stock populations can be screened for negative or beneficial genetic variants, which can either be avoided or selected. The overarching objective of this thesis is to utilise recent advances in genome sequencing and genotyping technologies to develop genome-wide genomic resources necessary for incorporating genetic selection breeding programs into the Pinctada maxima pearling industry. Specifically, I describe the large scale sequencing and annotation of the P. maxima transcriptome, the development of thousands of genomewide microsatellite and single nucleotide polymorphism (SNP) loci, the construction of the first moderate-density genetic linkage map for a pearl oyster species, and the identification of preliminary quantitative trait loci (QTL) and marker associations to two commercially important groups of pearl oyster traits; oyster shell growth and pearl quality. Firstly, the large-scale sequencing (~1.3 million sequences) of the P. maxima mantle transcriptome allowed the subsequent development and validation of two large suites of type I genome-wide molecular markers; SNPs, and microsatellites. This thesis describes molecular marker development, along with the assembly (96,794 contigs) and sequence annotation of the mantle transcriptome database. The SNP discovery effort resulted in the de novo identification of 172,625 SNPs, of which 9,108 were identified as high-value (MAF ≥ 0.15, read depth ≥ 8). Validation of 2,782 of these SNPs using Illumina iSelect Infinium genotyping technology returned some of the highest assay conversion (86.6%) and validation (59.9%; mean MAF 0.28) rates observed in aquaculture species to date. In addition, results show that subsets of the SNP loci will have widespread use as parentage and genetic diversity markers across species within Pinctada, which may potentially compliment the use of current suites of microsatellite genetic tools. Likewise, a total of 2,322 unique microsatellite loci were identified throughout the transcriptome database, with 360 shown to be polymorphic in silico using allelic motif variation. The mantle transcriptome database and molecular marker suites presented here form a strong foundation for genome mapping studies and have been pivotal to research investigating the biological mechanisms behind biomineralisation, quantitative trait loci mapping and association analysis as demonstrated within this thesis. Secondly, this thesis describes the construction of a dense genetic linkage map for P. maxima, including investigations into segregation distortions, family-specific heterogeneity and sex-specific recombination rates. The construction of a moderate- to high-density genetic linkage map for P. maxima is not only essential for mapping QTL and unraveling the genomic architecture of complex pearl quality traits, but also provides indispensable information on the genome structure of pearl oysters. A total of 335 oysters from eight full-sib families (six phase known and two phase unknown) were genotyped over 2,782 genome-wide SNPs. Of the 2,782 SNPs, 1,189 were informative and incorporated into linkage map construction. The final linkage map consisted of 887 SNPs in 14 linkage groups and spans a total genetic distance of 831.7 centimorgans (cM). The average marker interval (excluding intervals of 0 cM) was 2 cM and the map covers an estimated 96% of the P. maxima genome. Assessments of sex-specific recombination revealed mild heterochiasmy, but pronounced localised differences between male and female recombination throughout the linkage groups, whereby male recombination was suppressed near the centromeres compared to female recombination, but inflated towards telomeric regions. Numerous candidate genes for nacre biomineralisation were also localised providing some of the first positional information for these genes. Finally, the genetic architecture of complex oyster growth (i.e. shell height, length, width and weight) and pearl quality traits (pearl size, weight, surface complexion and colour) were explored using genome-wide association tests and QTL approaches. This thesis documents, for the first time, QTL and genetic associations to these commercially important pearl oyster growth and pearl quality traits and provides important insights into the genetic control of these traits. A total of 16 QTL and 32 genetic associations were detected for all oyster shell growth and pearl quality traits that explained from 27.4% to 46.1% of the phenotypic variation of a trait per family. The majority of QTL and marker associations were detected for the traits oyster shell width and pearl colour. This study confirms previous quantitative genetic studies providing conclusive evidence that oyster growth and pearl quality traits have a low to moderate additive genetic component and are complex and polygenic in nature. As a body of work, this thesis presents the most comprehensive genomic resources produced for any pearl oyster to date and presents the first quantitative genetic dissection of performance traits within P. maxima. Such work is pivotal to enabling the incorporation of genetic breeding programs within the P. maxima pearling industry and for investigating broader biological questions including the dynamic process of biological-mediated biomineralisation and bivalve evolution.

The thesis addressed various aspects of the reproductive biology and growth of the winged pearl oyster, Pteria penguin, in the Great Barrier Reef, north-eastern Queensland. The study was supported by the Australian Centre for International Agricultural Research (ACIAR), as part of ACIAR Project FIS/2006/172, 'Winged pearl oyster industry development in Tonga', for which James Cook University is the commissioned organization. The overall aim of this project was to provide information to support further development of P. penguin culture for pearl production, particularly as a basis for supporting livelihoods in Tonga and other developing countries. P. penguin were collected at Orpheus Island, north-eastern Queensland to observe sexual development, sex ratio and changes in morphometric relationships during growth. P. penguin were found to be protandrous hermaphrodites, with all sexually mature oysters below 88.8 mm dorso-ventral measurement (DVM) being males. Evidence of male primary gonad development was seen in oysters with a DVM of ≥33.9 mm. Sexual maturity of males was first seen in oysters at ≥56.0 mm DVM, but was common in oysters at ≥70.0 mm DVM. Mean female size was 150.1 ± 3.6 DVM (mm ± 1 SE), and mean male size was 111.3 ± 2.2 mm DVM. In oysters <170 mm DVM, males outnumbered females. The female to male sex ratio in the population tended towards 1:1 with increasing size. Female to male sex ratio of oysters between 170-180 mm DVM was 1:1, and females outnumbered males in oysters >180 mm DVM. Significant linear correlations were found between antero-posterior measurement of the shell (APM) and DVM (r² = 0.97, p <0.001) and between shell thickness (ST) and DVM (r² = 0.95, p <0.001), and a curvilinear relationship was recorded between adductor muscle weight (AW) and DVM (r² = 0.93, p <0.001). These findings have industry applications relating to collection of juvenile stock, spawning induction procedures and determining optimal shell size for nucleus implanting. The spawning cycle of P. penguin, was studied to observe seasonal changes in gonad development, mean oocyte diameter and sex ratio over time. A total of 201 oysters were collected over ten sampling periods, approximately every five to six weeks (mean = 20 oysters/sample). Gonad development stages for both males and females were identified as developing (with small follicles separated by connective tissue), ripe (swollen follicles containing fully developed eggs or sperm), spawning (partially empty follicles with walls partly broken), and spent (almost empty follicles with no signs of gametogenesis present). Oysters during periods of quiescence were described as inactive (with mainly connective tissue present and sex not distinguishable). Histological analysis of gonads showed that spawning activity occurred mainly through the austral mid-spring to late summer (November to March), when mean monthly water temperature ranged between 27.5–29.4 ºC. There was a primary spawning peak at the beginning of the spawning season in December, followed by a secondary peak in March. Mean oocyte diameter (μm) was highest during the warmer months, ranging from 31.2 ± 2.4 in November, to 36.2 ± 3.2 in March, and sex ratio analysis showed the percentage of females in the population also peaked between 30- 50% during this period. Recruitment of P. penguin spat in relation to season, substrate type and depth was investigated over a period of 27 months, from February 2008 to April 2010. Two substrate types (70 % shade cloth and open weave polypropylene mesh bags) were deployed at two depths (4 m and 6 m) and checked every six weeks over three spawning seasons to determine any differences in quantity of spatfall between these factors. No significant difference was found in spat recruitment between substrate types (p = 0.158) or depth (p = 0.349), while there was a significant seasonal effect on spat recruitment (p < 0.001), with a peak in the quantity of spatfall in late summer, from February to March, and no spat collected in the winter to spring (July to October). Maximum settlement of spat was 10.2 per mesh bag collector in February 2008. Recruitment was significantly reduced (p < 0.001) during the 2010 spawning season due to disturbance from severe storms generated by tropical cyclone Olga in late January. Juvenile P. penguin were cultured for six months in three commonly used culture units (panel pocket nets, plastic mesh trays and pyramidal pearl nets) at two dissimilar sites, Pioneer Bay (a coral reef environment in which P. penguin are naturally present) and Cape Ferguson (a coastal semi-estuarine area with high levels of silt deposition), to determine the effects of site and culture method on growth, survival and fouling. Mean growth increases (mm) were recorded for DVM, APM, ST and WW (whole weight, g). At Pioneer Bay, five replicates for each of the three culture unit types were suspended at 3 m and at 6 m, and mean initial DVM was 28.0 ± 0.6 mm (n = 190). At Cape Ferguson, there were five replicates for each culture unit at 3 m only, and mean initial DVM was 28.0 ± 0.6 mm (n = 86). Mean growth increase at the end of the experiment for oysters at Pioneer Bay and Cape Ferguson (respectively) were: 26.6 ± 1.0 and 32.6 ± 2.4 mm DVM, 29.6 ± 1.2 and 34.0 ± 2.9 mm APM, 6.7 ± 0.3 and 9.7 ± 0.5 mm ST, and 16.5 ± 0.7 and 23.0 ± 2.1 g WW. For all growth parameters, mean increase was significantly higher under high turbidity conditions at Cape Ferguson (p < 0.05). Culture unit also affected growth, with oysters held in mesh trays showing significantly higher growth at both sites (p < 0.05). Mean survival of oysters at Cape Ferguson (96.5%) was significantly higher than at Pioneer Bay (79.4%). Depth had no significant effect on growth, survival or fouling. The results indicated that site selection and culture unit are important parameters for optimizing growth and survival during nursery phase culture of P. penguin. Growth of P. penguin was monitored over 20 months, from April 2009 to November 2010, to investigate differences in growth performance at three dissimilar sites; Pioneer Bay, Cape Ferguson and Horseshoe Bay in the Great Barrier Reef lagoon. Growth parameters generated with the von Bertalanffy growth function ranged from K = 0.09 - 0.32 and L∞ = 283.6 - 822.5 (mm DVM). Overall growth performance (Φ') ranged between 4.40 - 4.77 and time to reach commercial size (T₁₀₀) was between 1.38 - 1.54 years, and T₁₂₀ was between 1.74 - 1.92 years. A more accurate estimate of L∞ = 213.4 mm DVM was obtained at Pioneer Bay by using larger data set which incorporated a wider size range of oysters. Overall monthly increase in DVM of oysters held at Horseshoe Bay (5.3 ± 0.2 mm) was significantly higher (p < 0.05) than those at Pioneer Bay (4.8 ± 0.1 mm) and Cape Ferguson (4.9 ± 0.1 mm), which were not significantly different. Monthly DVM increase was fastest (7.2 ± 0.1 mm), in small oysters (50 - 70 mm DVM) in the spring-summer and was lowest (2.2 ± 0.4 mm) in larger oysters (105 - 110 mm DVM) during the spring. Regression analysis showed APM, ST and WW were significantly correlated with DVM for all groups (p < 0.001). In the DVM size class of 100-120 mm, mean WW of oysters at Cape Ferguson was significantly higher (p < 0.01), and APM:DVM ratio was also significantly higher (p < 0.01) for oysters at Cape Ferguson and Horseshoe Bay, while the there were no significant differences between groups in ST:DVM ratio. At all three sites, the highest mortalities (%) were recorded for small oysters (25-50 mm DVM) during the winter period. Suspended inorganic matter (PIM, g) levels were significantly different between sites (p < 0.001). Overall results show that P. penguin are able to tolerate and even thrive under a wide range of turbidity levels. However, site selection must consider the risk of exposure to low salinity and turbulent wave action, which may have negative impacts on growth. Comparison between growth rates obtained in the study demonstrate that there is significant variability in growth between sites in the Great Barrier Reef lagoon.

碩士
東方設計大學
文化創意設計研究所
107
The Executive Yuan has organized a one-of-a-kind and value-added cultural creative industry with such local features as OTOP (one town one product program) in various parts of Taiwan to train local talents and develop industries of local specialties in the hopes of driving business and employment opportunities. For example, Changhua County features small and medium-sized enterprises that play an important role in the developing the regional economy. While the coastal areas of Xianxi primarily features oyster farmers who raise and sell their own products, the cultivation of pearls has gradually declined due to competition of other products from other townships, and the lack of funding and sales platforms have resulted in a non-benign cycle. This study focuses on creative packaging design that integrates the culture and humanity of Xianxi Township in Changhua County, with the theme of the oyster pearl. Then, we adopt the evaluation grid method (EGM) to explore the local style of the theme-based oyster-raising industry in order to analyze and encourage the four major primal attractive factors of packaging design, local specialties, local association, and fresh ingredients, as well as six specific attractive factors and nine abstract attractive factors. In this study, we used figurative graphics to compare old oyster farmers at the production end and fresh oysters at the consumption end as the primary vision, using such abstract graphics as totemized waves and windmill backgrounds as the creative theme. Doing so has rendered a new artistic sentiment from the perspective of this research, helping to demonstrate oyster pearl packaging in another way. Since the scene of thee two pictures, which greatly differ from each other as realism versus totem, we expect to establish a new creative concept. Furthermore, appreciating consumer preferences and satisfaction can help graphic designers integrate such consumer opinions and references in their creations with regard to the styles for local industry.

The silver/gold-lipped pearl oyster, Pinctada maxima (Jameson 1901), is one of the most important pearl producing species throughout Southeast Asia and northern Australia. The commercial production of high value "South Sea" pearls is based on the culture of P. maxima, and stands to benefit substantially from the implementation of long-term selective breeding programs. Industry-wide interest towards genetic improvement of P. maxima is rapidly growing, however, several fundamental issues must be addressed before selection commences. To achieve sustained response to selective breeding, it is essential that adequate genetic diversity is present within the population; yet currently there is little known about how the culture process affects diversity in P. maxima. This thesis addresses key issues concerning the capture and maintenance of genetic diversity in cultured P. maxima populations, as well as identifying and understanding patterns of genetic structure and diversity distribution throughout its natural range. As the source of broodstock to create base populations for selective breeding, it is important to understand the genetic properties of wild P. maxima populations. Analysis of microsatellite and mitochondrial DNA (mtDNA) variation in wild P. maxima populations throughout its natural distribution revealed a gradient in genetic diversity across its range, with decreasing levels of variation seen in peripheral populations when compared to those situated more centrally (i.e. central Indonesia). Significant genetic structuring and differentiation was also observed amongst populations, and is attributed to historic and contemporary biogeographic influences. Comparisons between wild and cultured P. maxima populations indicate a high level of genetic erosion has occurred in hatchery-propagated populations, with effective populations sizes (Nₑ) as low as 3.5 and reductions in microsatellite variation as high as 44% occurring as early as two generations beyond wild progenitors. The practice of mass spawning was identified as a major factor in the reduction of diversity, although diversity was not necessarily maintained when a more controlled spawning approach was utilised. Subsequent investigation using DNA parentage analyses revealed highly variable broodstock contributions have played a significant role towards an increase in genetic relatedness and low Nₑ in cultured P. maxima and is likely to be exaggerated by variable survival rates amongst different pearl oyster families. Upon further investigation and experimentation, it was determined that highly variable family survival will affect Nₑ in communally reared P. maxima and the practice of equalising family sizes in order to reduce family size variance (and maximise Nₑ) may only become consistently beneficial once further progress is made towards understanding and then reducing variation in family survival rates. Culture practices related to variation in growth, such as size grading, culling and mass-selection were assessed for the ability to contribute to diversity losses. It was cautioned that broodstock selection for subsequent generations was potentially far more influential on diversity maintenance than culling or grading. Outcomes of this thesis have provided a substantial advancement in the understanding of factors influencing genetic diversity in wild and cultured P. maxima populations. Population structuring and differentiation found in wild P. maxima provides grounds for further investigation into possible hybrid vigour or outbreeding depression when crossbreeding different stocks, and whether population genetic differences translate into phenotypic variation in commercially significant traits that could be exploited by selective breeding. This thesis also highlights important culture practices that must be improved (or avoided) in order to capture and retain genetic diversity and reduce inbreeding within closed populations, which will increase the likelihood of sustained response to selection programs. It is recommended that to ensure the maintenance of genetic variation and long-term sustainability of future P. maxima selection programs, culturists should employ the use of molecular tools for parentage assignment of candidate broodstock to avoid mating related individuals, or implement structured breeding designs intended to conserve genetic variability whilst maximising genetic response to selection.

This thesis addressed various growth aspects of cultivating P. maxima in a commercial farm in Indonesia with special emphasis on the influence of the environment. P. maxima of three age-classes were grown at three sites (Ganan, Manselo and Batu Terio) and two depths over a period of 18 months and various aspects of somatic growth (linear and weight measurements), gonad growth (visual and histological observations) and factors which influence growth (environmental and biological) were monitored. Environmental monitoring of the three sites and two depths showed that seawater parameters varied spatially between sites and depths as well as temporally throughout the sampling period. Some of the parameters measured were physico-chemical properties (water temperature, salinity, pH) and particulate matter (suspended particulate matter, particulate organic matter and chlorophyll a, b and c). These environmental descriptors provide the basis for comparison of growth rates of P. maxima held at the three sites in subsequent chapters. Total growth (GT) and monthly instantaneous growth (G30) of both length and weight were computed as part of the study on growth of P. maxima. G30 is a better indicator of growth as it is a standardised measure and permits comparison to be made between various age-classes of oysters with different shell sizes as well as at a specific point in time. Growth studies showed age has an inverse relationship on P. maxima growth, with both GT and G30 decreasing with increasing age. Multivariate testing showed that growth of all age classes was affected by culture depth, but not culture site. Growth rate at a depth of 5 m was higher than at 15 m. When oysters were partitioned by size before analysis, there was a site effect on growth rate for medium and small oysters. Spatial differences in P. maxima growth was shown to be linked to the local culture environment, where variation in somatic grow was influenced by varying pH, salinity and pH levels, and gonad growth was influenced by water temperature, pH, SPM, POM. Growth of P. maxima was fitted into five mathematical models i.e Special Von Bertalanffy Growth Function (VBGF), General VBGF, Gompertz, Richards and Logistic and various growth parameters computed. The criteria used for best fit was low mean residual sum of squares (MRSS), high coefficient of determination (r2) and low deviation of the asymptotic length (L!) from the maximum length (Lmax). Based on these criteria, the Special VBGF and the General VBGF equally provided the best fit to length-at-age data for all the pearl oysters grown at the area. However, when data was plotted, General VBGF tended to underestimate L!. The Special VBGF best described the growth of P. maxima cultured at the farm, with growth parameters estimates of L! = 168.38 mm, K = 0.930 y-1, t0 = 0.126. Biofouling studies showed that six classes of macro-fouling which settled on the shells of P. maxima were invertebrates from the classes Maxillopoda, Polychaeta, Bivalvia, Demospongiae, Foraminifera and Ascidicea. The quantity and diversity of biofouling was found to affect growth in medium and small oysters. The spatial and temporal variation in quantity and diversity of the six classes of biofouling was in turn affected by various environmental parameters. Regression analysis provided information on environmental parameters acting in concert to affect biofouling while principal component analysis showed the interaction between different biofouling taxa and environmental parameter. Together, they allowed examination of the interaction between various parameters, apportionment of environmental factors towards taxa of fouling and the degree a particular environmental variable affects fouling. Chlorophyll levels, pH and salinity were found to have a greater affect on biofouling settlement than SPM, POM and seawater temperature. Macroscopic investigation of gonads and comparison to histological data in this study support previous reports that gonad colour and appearance may be used to determine sex and stage of development in P. maxima. A fundamental difference in the colour and the area occupied by the developing gametes made it possible to distinguish between the gender and various stages of development of P. maxima oysters with relative ease. While most of the oysters observed appeared to be of indeterminate sex, enough male and female oysters were observed to show that gametogenesis in cultured P. maxima occurred between August to February, with spawning occurring twice during that period; once in October/November and again in February. Sex ratio in cultured P. maxima was overwhelmingly biased towards maleness, with no spatial difference in sex ratio between oysters cultured at various sites and depths. The expression of maleness was weakly correlated to water temperature, pH and rainfall, while there was no correlation between femaleness and environmental descriptors. Size, and not age, was more important in determining the sex of P. maxima. In summary, this research presented new data on growth of different age classes of P. maxima cultured in a farm situation in Indonesia. It has added to our knowledge the importance of various environmental factors and biofouling on somatic and gonadal growth of P. maxima. This information can be utilised to improve farming management practices through judicious selection of future culture sites. It is hoped that this will form a basis for further study into grow-out of P. maxima in the pearling industry in Indonesia and South-East Asia and lead to further improvement and expansion in the industry for the future.

The black-lip pearl oyster Pinctada margaritifera (L.) is a bivalve mollusc highly valued for cultured pearl and pearlshell production throughout its extensive Indo-Pacific natural distribution, where it makes substantial contributions to local economies and supports coastal community livelihoods. Despite its commercial importance, substantial knowledge gaps exist for this species, particularly regarding genetic structure and population connectivity at both local and regional scales, as well as its taxonomic identity. This information is required for the development of sustainable fishery management strategies, as well as responsible aquaculture practices, to ensure the persistence of healthy wild populations, and continued commercial production. The overarching goal of the research undertaken for this thesis was to investigate the stock structure, connectivity and taxonomy of P. margaritifera, to inform fishery management and aquaculture practices across the extent of its Indo-Pacific distribution, with a particular focus on the Fiji Islands. Specifically, over four separate investigations, I develop novel genomewide single nucleotide polymorphism (SNP) markers for this species, and use them to investigate population genetic structure, diversity, connectivity and local adaptation of Fijian oysters, as well as for populations sampled from across the broader ~18,000 km species distribution. I also compare estimates of population connectivity derived from genomic analyses with an independent hydrodynamic particle dispersal model, to corroborate patterns of larval transport between study sites. Finally, I utilise phylogenomic analyses to assess the evolutionary relationships of the black-lip pearl oyster across its natural distribution, and also to ascertain its taxonomic identity among other members of the family Pteriidae. The first investigation developed 5,243 novel genome-wide SNP markers for P. margaritifera, and tested their utility by assessing population structure, genetic diversity, as well as detecting regions of the genome underlying functional differences among populations. It involved 156 Fijian oysters sampled from three wild, and one hatchery produced population. Shallow but significant genetic structure was revealed among all wild populations (average pairwise Fₛₜ = 0.046), with clear evidence of a genetic bottleneck in the hatchery population (Nₑ(LD) = 6.1), compared to wild populations (Nₑ(LD) >192.5). Fₛₜ outlier detection to differentiate individuals between the orange and black tissue colour morphotypes characteristic of this species, revealed 42-62 highly differentiated SNPs (p<0.02), while casecontrol association discovered up to 152 SNPs (p<0.001). Database searches revealed that five of these SNPs were associated with a melanin biosynthesis pathway, demonstrating their biological relevance. This investigation demonstrated the utility of genome-wide SNP data for assessment of genetic structure and diversity in P. margaritifera, with transferability to other highly-dispersive marine taxa for their conservation and management. The second investigation utilised 4,123 genome-wide SNPs, together with an independent hydrodynamic particle dispersal model to assess genetic structure, diversity, local adaptation and population connectivity at 6 farm and 5 wild Fijian sites. Weak fine-scale patterns of population structure indicative of broad-scale admixture were observed among wild oysters, while a hatchery-sourced farmed population exhibited a higher degree of genetic divergence (hatchery oysters cf. all other populations Fₛₜ=0.085–0.102). This hatchery-produced population had also experienced a bottleneck (Nₑ(LD)=5.1; 95% C.I.=[5.1-5.3]); compared to infinite Nₑ(LD) estimates for all wild oysters. Simulation of larval transport pathways confirmed the existence of broad-scale admixture by surface ocean currents, correlating well with finescale patterns of population structure discovered. Fₛₜ outlier tests failed to detect genetic signatures supportive of selection, with only 2-5 directional outlier SNPs identified (average Fₛₜ=0.116). The lack of biologically significant population genetic structure, absence of evidence for local adaptation and larval dispersal simulation, all indicated the existence of a single genetic stock of P. margaritifera in the Fiji Islands for management purposes. The combined use of independent high resolution genomic and oceanographic data as demonstrated here is a novel approach that can be applied to other broadcast spawning taxa. The third investigation examined the microevolutionary forces influencing genetic structure, connectivity and adaptive variation across the ~18,000 km Indo-Pacific distribution of P. margaritifera. Concordance with a theoretical population model known as the Core- Periphery Hypothesis (CPH), was used as a framework for this assessment. The CPH predicts that genetic diversity is expected to be highest at the centre of a species' distribution, progressively decreasing with increased differentiation towards outer range limits, as populations become increasingly isolated, fragmented and locally adapted. Analyses utilising 9,624 genome-wide SNPs and 580 oysters sampled from 14 sites, discovered differing patterns of significant and substantial broad-scale genetic structure between the Indian and Pacific Ocean basins. Indian Ocean populations were markedly divergent (Fₛₜ=0.253-0.418, p<0.001), compared to Pacific Ocean oysters, where basin-wide gene flow was much higher (Fₛₜ=0.001-0.109, p<0.001). Visualisation of population structure at selectively neutral loci resolved three and five discrete genetic clusters for the Indian and Pacific Oceans respectively, while evaluation of genetic structure at adaptive loci for Pacific populations (89 SNPs under directional selection; Fₛₜ=0.101-0.437, FDR=0.05), revealed five clusters identical to those detected at neutral SNPs, suggesting environmental heterogeneity within the Pacific. Patterns of structure and connectivity were supported by Mantel tests of isolation by distance (IBD) and independent hydrodynamic particle dispersal simulations. These findings have revealed that population organisation in this species is highly complex and far more elaborate than generalised CPH predictions, with structuring being produced by the interaction of ocean currents, IBD and seascape features at a broad scale, together with habitat geomorphology and local adaptation at regional levels. The fourth and final investigation examined evolutionary relationships and the taxonomic identity of P. margaritifera. This study was required as the current species classification is not supported by molecular data, and includes a total of six subspecies that are described exclusively using morphological characters. Here, 69 oysters were sampled from 14 populations in both the Indian and Pacific Oceans. Samples were also collected from the congeneric taxa P. maxima and P. mazatlanica (n=29 and n=10, respectively), and phylogenetic reconstruction carried out using both 8,308 genome-wide SNPs and 10,000 dominant loci. Reconstructions using neighbour-joining (Nei's 1972 unbiased distances), maximum likelihood and Bayesian approaches all indicate that the taxonomy of P. margaritifera is more complex than previously indicated, with distinct evolutionary significant units (ESUs) identified within Tanzanian and Iranian populations, correlating with type localities for two Indian Ocean morphological subspecies descriptions. Contrastingly, phylogenies generated for Pacific Ocean P. margaritifera resolved a large monophyletic clade, suggesting little support for two of three morphological subspecies classifications reported from this ocean basin. Furthermore, P. mazatlanica specimens all formed a basal clade closest to French Polynesian P. margaritifera, suggesting it may not constitute a separate species. Collectively, these findings provide evidence to support a suggestion by previous studies that P. margaritifera comprises a species complex; however, further investigation involving finer-scale sampling with higher sample densities is required to resolve regional ESU boundaries. Collectively, this thesis presents the most comprehensive evaluation of genetic structure, population connectivity and evolutionary relationships for P. margaritifera to date. The data generated have permitted fundamental insights into the stock and taxonomic structure of this species, which are invaluable for its sustainable fishery management and aquaculture, with extension to other taxa possessing similar biological attributes.

碩士
國立澎湖科技大學
水產資源與養殖研究所
101
For a long time, the collection of black pearl oyster (pinctada margaritifera) as a food source in Penghu islands, has led to the depletion of the species. There were insufficient studies on black pearl oyster stock and species conservation in Taiwan. This study aim to assess black pearl oyster stock of 9 survey sites along the coastline of southern Penghu Island. The transect line survey method was conducted to investigate the population density of P. margaritifera, substrate preferences and the composition of epibiotic community on the shells. A total of 70 black pearl oysters were collected from 4860 m2 survey area. The depth of distribution ranged from 3 to 9 m under sea water level. Mean population density was 1.42 ± 0.36 individual•100 m-2. The highest population density was 8.7 ± 0.9 individual•100 m-2 in Shan Shuei 1 survey site, while the lowest population density was 0.19 ± 0.1 individual•100 m-2 in Shan Shuei port survey site. Mean shell height of P. margaritifera among sampling sites ranged from 31 to 132 mm, the overall mean was 81 mm. For substrate preferences, P. margaritifera was more likely to settle on the hard rock substratum (90%) than coral reef (10%). There were 30 taxa of the composition of epibiotic community on the shells, including Chlorophyta, Phodophyta, Cyanobactera, Protozoa, Porifera, Coelenterata, Platyhelminthes, Mollusca, Echiurioidea, Annelida, Arthopoda, Echinodermata and Chrodata. MDS ordination analysis revealed no difference on the composition of epibiotic community among sampling sites. In general, in comparesion with other studies, this survey suggested the population density of P. margaritifera is relative low in Southern Penghu island. There was also no enough evidence demonstrating that the epibiotic community had negative effect on P. margaritifera. Therefore, over fishing activity might be the major reason to interpret low population density of P. margaritifera and Penghu county government need to protect and conserve P. margaritifera resource in the future.

The silver-lipped pearl oyster, Pinctada maxima, is cultivated in Australia and throughout South East Asia for the production of "South Sea pearls". Pearls from P. maxima are the largest and most valuable pearl produced out of all pearl producing oyster species, with the industry being valued at ~US$ 412 million farm-gate globally. The commercial production of pearls from this species is well established and hatchery techniques have been developed to the stage where the life cycle is considered closed. A standard commercial pearl production cycle takes 4 years from the time oysters are spawned in the hatchery to the harvesting of pearls. The first two years is a "prepearl" grow-out phase for the oyster that will become the host oyster, and then a further two years in a "pearl growth" phase whereby the oyster is "seeded" and the pearl nacre is laid down. After this 4 year cycle only ~20% of pearls harvested fall within the category of being "gem quality", where the combination of the commercially important traits lustre, size, shape, weight, colour and complexion are all within the accepted standards. It is this long phase of production and the low proportion of gem quality pearls harvested that makes it very attractive for pearling companies to improve the consistency of production through the use of genetic breeding approaches. A current impediment to adoption of genetic approaches, however, is a lack of knowledge on the role additive genetic factors play in the realization of a pearl quality trait. Another consideration for future breeding programs is that pearl companies generally have several farm sites situated in geographically distinct locations, with one hatchery supplying all the seedstock to these locations. As such, as well as understanding the genetic basis underpinning pearl traits, it is important for companies to determine the effect disparate environmental influences due to site location may have on both oyster growth and pearl quality, and to establish whether the realization of genetic potential in improved oysters will be affected by genotype by environment (G x E) interactions. Like that for genetic parameters governing pearl quality and growth traits, however, data on the potential impact G x E may have on selected oysters reared under different environmental conditions is lacking. In response to this critical missing information this thesis aimed to establish genetic parameters and estimate genotype by environment interactions for both growth traits and pearl quality in the silver-lipped pearl oyster. Investigations commenced in this thesis by considering the effect long-line location (and perceived differences in micro-environment) at four sites within a pearl farm have on the realization of growth and pearl quality. Variance of these traits due to genetic differences as a consequence of using different cohorts of broodstock was also considered. In this experiment long-line site was shown to have a significant impact on the overall growth rate of oysters, with oysters reared on long-lines at the Sasanaflapo site growing significantly faster than those at the other three sites examined. These growth differences were also shown to manifest regardless of the age or genetic composition of the cohort evaluated. This shows that oyster growth is a trait that may be influenced by environmental parameters within farm locations, and that long-line location may override individual genetic effects on growth. The influence site has within a farm on pearl quality was, however, less conclusive. Only differences in pearl colour overtones and lustre appeared to be modified by site effects within a farm. For example, pearl colour and lustre could be both partitioned using classification tree analyses by site, with the Duyef and Wulu sites producing on average more white pearls with pink overtones, while Maratlap and Sasanaflapo produced higher numbers of silver pearls with pink overtones. Likewise for lustre, splits in classification trees were related to the fact that the Wulu site produced more pearls exhibiting higher lustre grades. Despite these differences though no disparities were found in the economic value of pearls harvested from the various sites within the farm evaluated. With the suggestion that local-scale environmental effects may modify oyster growth traits the thesis then goes on to test if genetic differences represented by oysters from different families could be easily modified through the manipulation of defined environmental parameters. Here the relative performance in shell growth of spat from five full-sib families when spat were communally reared at different salinities (29, 34 and 40 ppt), food availability (high, medium and low), food quality (high, medium and low), and in a hatchery vs. ocean environment for 43 days, were compared. In support of the first experiment, rearing environment was again found to significantly influence growth expression, with significant differences evident when spat were reared at different salinities, in the ocean instead of hatchery, or when fed algae of differing nutritional quality. Additionally, comparative family growth was also altered when the environment changed, with significant environment by family interactions (G x E) apparent under food quality, food availability and hatchery vs. ocean rearing conditions. These results indicate, that at least during early oyster growth phases, that growth and relative family performance in P. maxima may change dependent on local environmental conditions. To further examine the effect environment and genetics has on oyster growth, spat were produced for a large commercial scale trial using broodstock originating from three distinct Indonesian populations (Bali, West Papua, Aru Islands). These spat from different genetic backgrounds were communally on-grown to adult sizes for two years at each of two Indonesian commercial farming locations (Bali and Lombok). Microsatellite based parentage determination analyses were used to retrospectively sort out oysters to both their family and population of origin. Significant size differences were observed in all shell growth traits measured (dorsal-ventral measurement DVM, anterior-posterior measurement APM, shell width SW and wet weight WW), with oysters originating from Bali and West Papua (DVM (mm) = 103.7±0.9 and 101.0±0.6 respectively) growing faster than those from the Aru population (93.5±0.5) at both grow-out locations. Family level differences within these populations were also present for shell traits, indicating a large amount of genetic variability present for potential breeding programs. However, although there were significant familial size differences for shell traits, unlike the earlier spat growth experiment, genetic correlation analyses showed little evidence for re-ranking of family performance among the two culture sites (r(g) = 0.89–0.99). This implies that under the commercial conditions oysters were evaluated that insignificant genotype by environment deviations among sites were evident. Heritability analyses based on these families were also conducted for shell traits, with DVM and APM found to be moderately heritable (0.15 ±0.00 (DVM), 0.23 ±0.03 (APM)). Thus selection for faster growing host oysters should be possible which would advantage the industry by decreasing the amount of time it takes to grow oysters to a size suitable for nucleus implantation. The final experiment undertaken in this thesis estimated for the first time genetic parameters and G x E for pearl quality traits when multiple families were again reared at Bali and Lombok. Here significant differences in the size and value of pearls produced at the two locations were observed, with pearls produced at Lombok generally bigger and more valuable than their Bali counterparts. Comparisons of pearls produced by the various families jointly reared at these two sites also indicated adverse genetic correlations for size (r(g) = -0.22), colour (r(g) = 0.28) and weight (r(g) = 0.38), and less so shape (r(g) = 0.56) and lustre (r(g) = 0.59); thus the occurrence of genotype by environment modifications for these pearl quality traits needs to be factored into improvement programs. Heritability analyses based on the donor-oyster additive genetic contribution showed that all pearl traits except that of shape exhibited low to moderate heritabilities (size h² = 0.13, lustre h² = 0.14, weight h² = 0.15, colour h² = 0.15 and complexion h² = 0.25). As a consequence these traits could be improved through the practice of selection. The findings of this thesis have substantially advanced our knowledge of the respective role genetics and the environment play in the realization of commercially important traits in the pearl oyster P. maxima. Pearl quality and oyster growth traits have been shown to have a heritable basis, thereby making them amiable to improvement through selection approaches. Results also have shown that when designing future breeding programs considerations of large-scale site induced environmental effects and associated genotype by environment modifications will need to be factored. Through implementation of the information gathered in this thesis the P. maxima pearling industry now has a sound basis for the future design of efficient selection programs aimed at improving the productivity and profitability of their industry.