Recipient Oyster Research Articles

Lysine acetylation plays a role in the allograft-induced stress response of the pearl oyster (Pinctada fucata martensii)

Implanting a spherical nucleus into a recipient oyster is a critical step in artificial pearl production using the pearl oyster Pinctada fucata martensii. However, little is known about the role of post-translational modifications (PTMs) in the response of the pearl oyster to this operation. Lysine acetylation, a highly conserved PTM, may be an essential adaptive strategy to manage multiple biotic or abiotic stresses. We conducted the first lysine acetylome analysis of the P. f. martensii gill 12 h after nucleus implantation, using tandem mass tags (TMT) labeling and Kac affinity enrichment. We identified 2443 acetylated sites in 1301 proteins, and 1511 sites on 895 proteins were quantitatively informative. We found 25 conserved motifs from all of the identified lysine sites, particularly motifs Kac H, Kac S, and Kac Y were strikingly conserved, of which Kac Y, Kac H, Y Kac, Kac K, Kac *K, Kac R, and Kac F which have been observed in other species and are therefore highly conserved. We identified 58 sites that were significantly differently acetylated in P. f. martensii in response to allograft (|fold change|>1.2, P-value ≤ 0.05); 38 newly acetylated and 20 deacetylated. According to GO functional analysis, subcellar location, and KOG classIfication, these proteins were divided into four categories: cytoskeleton, response to stimulus, metabolism, and other. The differentially acetylated proteins (DAPs) enriched pathways include aminoacyl-tRNA biosynthesis, salmonella infection, and longevity regulating pathway-worm-Caenorhabditis elegans (nematode). Parallel reaction-monitoring (PRM) validation of the differential acetylation of 10 randomly selected differentially acetylated sites from the acetylome analysis. These results indicated that our acetylome analysis results were sufficiently reliable and reproducible. These results provide an essential resource for in-depth exploration of the stress responses and adaptation mechanisms associated with lysine acetylation in marine invertebrates and P. f. martensii.

Comparative proteomics and transcriptomics illustrate the allograft-induced stress response in the pearl oyster (Pinctada fucata martensii)

Implantation of a spherical nucleus into a recipient oyster is a critical step in artificial pearl production. However, the molecular mechanisms underlying the response of the pearl oyster to this operation are poorly understood. In this research, we used transcriptomic and proteomic analyses to examine allograft-induced changes in gene/protein expression patterns in Pinctada fucata martensii 12 h after nucleus implantation. Transcriptome analysis identified 688 differential expression genes (DEGs) (FDR<0.01 and |fold change) ＞ 2). Using a 1.2-fold increase or decrease in protein expression as a benchmark for differentially expressed proteins (DEPs), 108 DEPs were reliably quantified, including 71 up-regulated proteins (DUPs) and 37 down-regulated proteins (DDPs). Further analysis revealed that the GO terms, including "cellular process", "biological regulation" and "metabolic process" were considerably enriched. In addition, the transcriptomics analysis showed that “Neuroactive ligand-receptor interaction”, “NF-kappa B signaling pathway”, “MAPK signaling pathway”, “PI3K-Akt signaling pathway’, “Toll-like receptor signaling pathway”, and “Notch signaling pathway” were significantly enriched in DEGs. The proteomics analysis showed that “ECM-receptor interaction”, “Human papillomavirus infection”, and “PI3K-Akt signaling pathway” were significantly enriched in DEPs. The results indicate that these functions could play an important role in response to pear oyster stress at nucleus implantation. To assess the potential relevance of quantitative information between mRNA and proteins, using Ward's hierarchical clustering analysis clustered the protein/gene expression patterns across the experimental and control samples into six groups. To investigate the biological processes associated with the protein in each cluster, we identified the significantly enriched GO terms and KEGG pathways in the proteins in each cluster. Gene set enrichment analysis (GSEA) was used to reveal the potential protein or transcription pathways associated with the response to nuclear implantation. Thus, the study of P. f. martensii is essential to enhance our understanding of the molecular mechanisms involved in pearl biosynthesis and the biology of bivalve molluscs.

The influence of donor and recipient oyster red and blue shell nacre interference color on Pinctada fucata martensii pearl quality

The influence of shell body color on pearls has been studied extensively, but the effect of shell interference color selection on pearls has not been fully understood yet. Because interference color is an important factor determining pearl quality, it is important to study whether the shell interference color of the donor and recipient oysters has any influence on the resulting pearls' interference color to cope with the market demands. We report that selective breeding of red- and blue-line Pinctada fucata martensii pearl oysters produce high-quality pearls based on shape, flaw, color, size, and luster assessment categories. Saibo from red- and blue-line donors was implanted with a rounded nucleus into the red- and blue-line recipient oysters from the same batch in four treatments which we refer to as DRMR (red-donor, red-recipient), DRMB (red-donor, blue-recipient), DBMR (blue-donor, red-recipient), and DBMB (blue-donor, blue-recipient) oysters. Pearl shape and flaws, body yellowness index (YI) and interference value color categories, luster value, total nacre and tablet thickness, and surface microstructure for these treatments are reported. More DRMR pearls are high grade (H) than DBMB, DBMR, and DRMB pearls. Using electronic pearl quality measuring devices, DRMR pearls have the lowest YI and highest interference color values, rendering them superior in color category. Whole nacre thickness of pearls estimated by the SEM on the pearl section reveal DRMR pearls have significantly (p < .001) thicker nacre than DBMB, DBMR, and DRMB pearls, rendering them generally larger in size. DRMR pearls also have significantly (p < .001) thicker nacre tablets in basal, middle, and surface regions of the nacre layer. Whereas luster does not differ significantly among treatments (p > .05), DRMR pearls are superior to those of other treatments in their shape, reduced number of flaws, color, and size. We consider that P. fucata martensii oysters with red shell nacre interference color improve pearl quality, and for the purposes of pearl culture should be selected for. These results are of commercial importance, especially where P. fucata is cultured for pearl production.

Competition for food reduces disease susceptibility in a marine invertebrate

AbstractCompetition between organisms interfere in host and pathogen dynamics in ways that are difficult to predict. By one side, competitors can reduce the food supply and cause nutritional stress. Such stress could further modulate the susceptibility to infection by altering immune response or metabolic rate of the host. Alternatively, competitors may trap pathogens before they reach the focal host, and therefore reduce, enhance, or have no effect on infection according to the competitor's susceptibility to the infection. To better understand how competition influences host and pathogen interactions, we experimentally assessed the relative importance of competition for pathogens and resources on the severity of a viral disease infecting the Pacific oyster Crassostrea gigas. We designed an open‐flow system where food enriched seawater flowed to filter‐feeding competitors (or empty controls) before being delivered to recipient oysters. We tested a range of competing species that exhibit both low (ascidians, European oysters, mussels) and high (Pacific oysters) susceptibility to the virus. We assessed the physiological condition of the recipient oysters during acclimation, we added virus‐contaminated seawater upstream of the distribution system, and we monitored host and pathogen dynamics. We found that the presence of competitors, regardless of susceptibility to the virus, indirectly reduced the infection rate of hosts by decreasing their food ingestion and growth rates. Although competitors can reduce viral particles from the seawater, this had no effect on the host population. Our data suggest that the effect of competition for food overwhelmed that of competition for pathogens, thus emphasizing the importance of considering resource availability in host and pathogen dynamics. More particularly, resource availability can have positive effects at the individual level, fostering physiological condition and growth, but negative effects at the population level, increasing magnitude of epidemics.

Potential combinations of mabé, keshi and cultured pearl production from colourful hatchery-produced Pinctada margaritifera

Aquaculture of nacreous gems, such as cultured pearls, mabé or keshi, is done mostly using different mollusc species grown in countries of the Indo-Pacific region. To date, no single species has been exploited for the simultaneous generation of more than one of these bioproducts, but all require animals with colourful shells. Historically, Pinctada species have mainly been used for nucleated pearl production, selecting the rarer colourful individuals to be used as graft donors. By contrast, colourful Pteria species have mostly been used for mabé production, as the grafting operation for pearl production is associated with low yield. In this study, we report the potential for cumulating cultured pearl and mabé (MP), or keshi and mabé (MK) production, using a colourful hatchery-produced G2 family of P. margaritifera. For these trials, MP and MK combinations were compared with the operations to produce pearls (P), mabé (M) or keshi (K) alone in an experimental design using groups of small and large recipients from the G2 family. Results showed no significant impact of combining operation types on subsequent pearl weight, keshi weight, or mabé thickness within recipient oyster size group. By contrast, significant differences were observed between the large and small recipients. The small group produced the thickest mabé, while the large group produced the heaviest pearls and keshi. These contrasting results revealed: 1) the relative independence between the two tissues capable of biomineralisation activities, the mantle (shell and mabé growth) and the pearl sac (pearl or keshi growth); 2) the potential compensatory growth of the small recipient oyster group, which had the highest shell growth performance; and 3) the regulation capacity of the larger oyster group of pearl sac activity. With the same growing area and number of cultured oysters, it would be possible for the P. margaritifera pearl industry to benefit from hatchery propagation of selected colourful shell and produce valuable keshi and mabé together with cultured pearls.

Towards a better understanding of allograft-induced stress response in the pearl oyster Pinctada fucata martensii: Insights from iTRAQ-based comparative proteomic analysis

Implantation of a spherical nucleus into a recipient oyster is a critical step in artificial pearl production. The implanted nucleus is known to trigger cellular stress responses at several levels, yet the molecular mechanism underpinning physiological adaptation of the pearl oysters to nucleus implantation is still poorly understood. In this study, we took advantage of the iTRAQ-based proteomics and LC-MS/MS approach to look into allograft induced gene regulation at the protein expression level in the pearl oyster Pinctada fucata martensii, across a period of 30 days following nucleus implantation. A wide variety of proteins, including a group of immune-related proteins such as E3 ubiquitin-ligase and heat shock proteins, exhibited differential expression in response to the surgical operation. Further comparisons between different sampling points revealed that GO terms including “translation” and “oxidation-reduction process” and KEGG pathways including “glycolysis/gluconeogenesis” and “pyruvate metabolism” were significantly enriched at several time points, indicating the important roles of these molecular events in the stress response of pearl oysters to nucleus implantation. In addition, considerable discrepancy between protein expression level and gene transcript abundancy was identified, as only a few genes showed at least 2-fold expression changes at both proteomic and transcriptomic levels. The result implies that post-transcriptional gene regulation for the key proteins may represent an important aspect of allograft-induced stress response in the pearl oysters. Taken together, the data obtained would contribute to a deeper understanding of the molecular mechanisms enabling stress adaptation of the pearl oysters in response to nucleus implantation.

Redescription of Larval Development in Cultured Pearl Oyster Pinctada maxima

Larval development plays a major role in efficient pearl culture. The cultured larvae will be later used for recipient and donor oysters in cultured pearl production. Larval developmental stage of silver lip pearl oyster Pinctada maxima has been reported by several studies. Those studies used female and male oyster parents directly taken from natural habitat.This study aimed to redescribe larval development of P. maxima from commercial pearl oyster culture farm in Indonesia. Larval development of this species whose parents are originated from selected groups in the pearl culture farm has not been reported yet, thus it is necessary to be described. This species undergoes specific larval developmental stage. The larvae were observed under microscope, and then the average shell length (SL) and shell height (SH) were measured. D-shaped veliger larva (77.4±0.3 µm SL; 65.4±1.1 µm SH) appeared 20 h after fertilization. Tenday-old larva (156.2±2.8 µm SL; 149.5±5.6 µm SH) had developed umbo region so it was called umbonal larva. Umbonal larva then developed further into plantigrade larva (411.3±9.8 µm SL; 380.5±6.9 µm SH) in 25 days after fertilization. Developmental stage and larval sizein P. maxima is similar with those observed in P. fucata and P. margaritifera. Keywords: growth, larva, plantigrade, shell, umbo, veliger

Comparison of Two Pearl Sacs Formed in the Same Recipient Oyster with Different Genetic Background Involved in Yellow Pigmentation in Pinctada fucata.

Color is one of the most important factors determining the commercial value of pearls. Pinctada fucata is a well-known pearl oyster producing high-quality Akoya pearls. Phenotypic variation in amount of yellow pigmentation produces white and yellowish pearls. It has been reported that polymorphism of yellow pigmentation of Akoya pearls is genetically regulated, but the responsible gene(s) has remained unknown. Here, we prepared pearl sac pairs formed in the same recipient oyster but coming from donor oysters that differ in their color. These two pearl sacs produced pearls with different yellowness even in the same recipient oyster. Yellow tone of produced pearls was consistent with shell nacre color of donor oysters from which mantle grafts were prepared, indicating that donor oysters strongly contribute to the yellow coloration of Akoya pearls. We also conducted comparative RNA-seq analysis and retrieved several candidate genes involved in the pearl coloration. Whole gene expression patterns of pair sacs were not grouped by pearl color they produced, but grouped by recipient oysters in which they were grown, suggesting that the number of genes involved in the yellow coloration is quite small, and that recipient oyster affects gene expression of the majority of genes in the pearl sac.

Crossing Phenotype Heritability and Candidate Gene Expression in Grafted Black-Lipped Pearl Oyster Pinctada margaritifera, an Animal Chimera.

Grafting mantle tissue of a donor pearl oyster into the gonad of a recipient oyster results in the formation of a chimera, the pearl sac. The phenotypic variations of this chimera are hypothesized to be the result of interactions between the donor and recipient genomes. In this study, the heritability of phenotypic variation and its association with gene expression were investigated for the first time during Pinctada margaritifera pearl production. Genetic variance was evaluated at different levels, 1) before the graft operation (expression in graft tissue), 2) after grafting (pearl sac tissue expression in chimera), and 3) on the product of the graft (pearl phenotype traits) based on controlled biparental crosses and the F1 generation. Donor-related genetic parameter estimates clearly demonstrate heritability for nacre weight and thickness, darkness and color, and surface defects and grade, which signifies a genetic basis in the donor oyster. In graft relative gene expression, the value of heritability was superior to 0.20 in for almost all genes; whereas in pearl sac, heritability estimates were low (h2 < 0.10; except for CALC1 and Aspein). Pearl sac expression seems to be more influenced by residual variance than the graft, which can be explained by environmental effects that influence pearls sac gene expression and act as a recipient additive genetic component. The interactions between donor and recipient are very complex, and further research is required to understand the role of the recipient oysters on pearl phenotypic and gene expression variances.

Cultured Pearl Surface Quality Profiling by the Shell Matrix Protein Gene Expression in the Biomineralised Pearl Sac Tissue of Pinctada margaritifera.

Nucleated pearls are produced by molluscs of the Pinctada genus through the biomineralisation activity of the pearl sac tissue within the recipient oyster. The pearl sac originates from graft tissue taken from the donor oyster mantle and its functioning is crucial in determining key factors that impact pearl quality surface characteristics. The specific role of related gene regulation during gem biogenesis was unknown, so we analysed the expression profiles of eight genes encoding nacreous (PIF, MSI60, PERL1) or prismatic (SHEM5, PRISM, ASP, SHEM9) shell matrix proteins or both (CALC1) in the pearl sac (N = 211) of Pinctada margaritifera during pearl biogenesis. The pearls and pearl sacs analysed were from a uniform experimental graft with sequential harvests at 3, 6 and 9months post-grafting. Quality traits of the corresponding pearls were recorded: surface defects, surface deposits and overall quality grade. Results showed that (1) the first 3months of culture seem crucial for pearl quality surface determination and (2) all the genes (SHEM5, PRISM, ASP, SHEM9) encoding proteins related to calcite layer formation were over-expressed in the pearl sacs that produced low pearl surface quality. Multivariate regression tree building clearly identified three genes implicated in pearl surface quality, SHEM9, ASP and PIF. SHEM9 and ASP were clearly implicated in low pearl quality, whereas PIF was implicated in high quality. Results could be used as biomarkers for genetic improvement of P. margaritifera pearl quality and constitute a novel perspective to understanding the molecular mechanism of pearl formation.

Donor and recipient contribution to phenotypic traits and the expression of biomineralisation genes in the pearl oyster model Pinctada margaritifera

Grafting associates two distinct genotypes, each of which maintains its own genetic identity throughout the life of the grafted organism. Grafting technology is well documented in the plant kingdom, but much less so in animals. The pearl oyster, Pinctada margaritifera, produces valuable pearls as a result of the biomineralisation process of a mantle graft from a donor inserted together with a nucleus into the gonad of a recipient oyster. To explore the respective roles of donor and recipient in pearl formation, a uniform experimental graft was designed using donor and recipient oysters monitored for their growth traits. At the same time, phenotypic parameters corresponding to pearl size and quality traits were recorded. Phenotypic interaction analysis demonstrated: 1) a positive correlation between recipient shell biometric parameters and pearl size, 2) an individual donor effect on cultured pearl quality traits. Furthermore, the expressions of biomineralisation biomarkers encoding proteins in the aragonite or prismatic layer showed: 1) higher gene expression levels of aragonite-related genes in the large donor phenotype in the graft tissue, and 2) correlation of gene expression in the pearl sac tissue with pearl quality traits and recipient biometric parameters. These results emphasize that pearl size is mainly driven by the recipient and that pearl quality traits are mainly driven by the donor.

Influence of preoperative food and temperature conditions on pearl biogenesis in Pinctada margaritifera

Trophic conditions and water temperature strongly influence bivalve physiological processes and metabolism. In black-lip pearl oyster Pinctada margaritifera, these parameters have been shown to affect shell biomineralization. The present study investigated the effect of preoperative food level (i.e., microalgal concentration) and temperature on pearl biomineralization. Donor and recipient oysters were conditioned at different levels of food and temperature during the preoperative phase to evaluate the influence of these factors on 1) pearl retention rate (grafting success), 2) expression of genes involved in biomineralization in the mantle and pearl sac and 3) pearl quality traits. Our study confirmed the influence of both microalgal concentration and temperature on shell growth. Food level of donor oysters was decisive for pearl biomineralization, with donors that had been fed at a high microalgal concentration producing pearl sacs with significantly higher biomineralization capabilities and faster nacre establishment during early stages of pearl formation. However, food level showed no effects on quality traits of the pearls harvested 12months postgrafting, while preoperative temperature only influenced the relative expression of two genes in pearl sacs at 12months postgrafting. No significant effects of the preoperative conditioning of recipient oysters were detected in either experiment considering gene expression measurements and pearl quality traits. However, mortality was significantly lower in grafted recipient oysters fed at an intermediate trophic level. Finally, pearl weight was shown to be positively correlated with recipient oyster growth.

Serum immune response of pearl oyster Pinctada fucata to xenografts and allografts

The mantle piece from the donor pearl oyster would be rejected by the immune system of recipient oyster in pearl culture practice, especially in the case that the donor and receptor are different species. Thus, investigation of the immune response of recipient oyster to grafted mantle pieces, particularly to xenografts, is of importance in creating xenograft transplantation technology for pearl culture industry. The humoral immune responses of P. fucata to allograft (mantle piece of P. fucata) and xenografts (mantle pieces of P. maxima and P. margaritifera, respectively) were studied in this paper. The oysters receiving no transplantations were served as the control group. The serum was collected from recipient P. fucata at 1 d, 2 d, 3 d, 4 d, 5 d, 7 d, 9 d, 11 d, 13 d, and 15 d, respectively after transplantation, and the serum antibacterial activity, lysozyme activity (LZM), alkaline phosphatase (AKP), acid phosphatase (ACP), total antioxidant capacity (TAC), and agglutination to rabbit red blood cells were investigated. The result indicated that serum of both the experimental groups and the control group can agglutinate rabbit red blood cells, with variation between groups and between time points, respectively. The antibacterial activity in the experimental group was significantly higher than that in the control group at 2–4 d, but lower at 5–11 d and returned back to normal at 15 d, with significant differences among experimental groups (P < 0.05). The LZM in the experimental group was significantly higher than that in the control group at 3–7 d, with significant differences in bacteriolytic activity among various groups (P < 0.05). Both the ACP and AKP activity levels in the experimental groups were higher than those in the control group at 2–9 d, with significant differences among various groups at 3–9 d (P < 0.05). The TAC level in the experimental groups was higher than that in the control group at 1–7 d, with significant differences among various groups at 4–7 d (P < 0.05). The above results indicated that all of the humoral immune factors investigated showed immune responses to both allografts and xenografts, with no specific to any of them. Thus, it is necessary to further screen immune rejection factors specific to xenografts, including cellular immune components.

Differentially expressed immune-related genes in hemocytes of the pearl oyster Pinctada fucata against allograft identified by transcriptome analysis

The pearl oyster Pinctada fucata is commonly cultured for marine pearls in China. To culture pearls, a mantle piece from a donor pearl oyster is grafted with a nucleus into a receptor. This transplanted mantle piece may be rejected by the immune system of the recipient oyster, thus reducing the success of transplantation. However, there have been limited studies about the oyster's immune defense against allograft. In this study, hemocyte transcriptome analysis was performed to detect the immune responses to allograft in P. fucata at 0 h and 48 h after a transplant. The sequencing reaction produced 92.5 million reads that were mapped against the reference genome sequences of P. fucata. The Gene Ontology (GO) annotation and the Kyoto Encyclopedia of Genes and Genomes (KEGG) were used to identify all immune-related differentially expressed genes (DEGs). Compared with patterns at 0 h, a total of 798 DEGs were identified, including 410 up-regulated and 388 down-regulated genes at 48 h. The expression levels of interleukin receptor and toll-like receptor in hemocytes were increased significantly 48 h post-transplant, indicating that the oyster immune response was induced. Finally, altered levels of 18 randomly selected immune-related DEGs were confirmed by quantitative real-time PCR (qRT-PCR). Our results provide the basis for further analysis of the immune rejection of allotransplantation.

Shell Growth Performance of Hatchery Produced Pinctada margaritifera: Family Effect and Relation with Cultured Pearl Weight

Size is the most important and valuable quality trait of cultured pearls produced by the black-lipped pearl oyster, Pinctada margaritifera. In French Polynesia, several breeding programmes have been started that aim to improve this size trait, which is highly related to shell growth rate in both recipient and donor oysters. Shell growth rate dictates the time of grafting, size of implanted nuclei and bio-mineralisation potential of the mantle and pearl sac. We assessed shell growth rate through routine digital shell biometric analysis on 22 hatchery families produced between 2005 and 2008. These included full-sib families and half-sib families derived from polyandry (one dam crossed with two or more sires). Results showed that: 1) a significant family effect was recorded for growth performance, analysed according to the Von Bertalanffy model, 2) a significant male effect was observed for some of the half-sib families and 3) a relationship was found between the shell growth performances of five families randomly selected and used as graft donors in a grafting experiment and the final weight of the cultured pearls produced. These results have important implications for the breeding of pearl oysters with high growth capacities: it may be possible to select oyster lines for the potential to produce large pearls using shell equivalent diameter estimated by the digital method as a selection criterion.

Culture site dependence on pearl size realization in Pinctada margaritifera in relation to recipient oyster growth and mantle graft biomineralization gene expression using the same donor phenotype

Size is the most important and valuable quality of the cultured black-lip pearl, Pinctada margaritifera. As this pearl aquaculture is carried out at numerous grow-out sites, this study analyzes the environmental influence on pearl size parameters (nacre weight and thickness) in relation to the recipient oyster biometric parameters (shell thickness, height, width, and oyster weight) at harvest time. Toward this end, an experimental graft was designed by using a homogeneous donor oyster phenotype. The recipient oysters were randomly and equally transferred and reared in five commercial and contrasting grow-out locations. Overall inter-site comparisons revealed that the cultured pearl size (N = 2168) and the biometric parameters of the recipient oysters were highest for sites with warmer temperatures with low seasonal variation in comparison to the southern latitude sites. These results were supported by positive correlations between pearl nacre thickness and recipient oyster shell thickness, height, and width. In parallel, the biomineralization potential of the mantle graft was screened through four genes encoding aragonite (Pif 177, MSI60) and calcite (shematrin 9, aspein). As the gene expression levels were the same among all the donor oysters, this finding demonstrates that: 1) the pearl sac that originated from the mantle graft was not isolated from environmental variations during the culture period and 2) the phenotypic expressions of the two biomineralizing tissues in the recipient oyster were consistent (shell and pearl). In the near future, this knowledge will be helpful at the production sites of genetically selected donor oyster lines for growth produced in hatchery systems.

Distribution of Ostreid herpesvirus-1 (OsHV-1) microvariant in seawater in a recirculating aquaculture system

Ostreid herpesvirus-1 (OsHV-1) microvariant presents a significant threat to the commercial production of Crassostrea gigas worldwide. Investigating the transmission of the virus in the estuarine environment where oysters are grown has proven to be difficult due to the complexity of the ecosystem. The aims of this trial were to (1) investigate the distribution of OsHV-1 in seawater over time in a recirculation system during an experimental disease outbreak; (2) assess the effects of the biofiltration and ultraviolet light components of the recirculation system on the detection of OsHV-1; and (3) assess the potential for OsHV-1 to associate with particles in the artificial seawater matrix. The source of virus was experimentally infected donor and cohabitating recipient oysters. Both the presence of infected oysters and the biofilter and UV light source significantly affected the detection rate of OsHV-1 in seawater. It was detected more frequently after high dose exposure (n=6 donors per tank) compared with low dose exposure (n=2 donors per tank), and when the biofilter and UV were disconnected from the treatment tanks. The odds of detecting OsHV-1 in the treatment tanks, which were periodically disconnected from the biofilter and UV, were 2.56 that of the system sumps which were always connected to the biofilter and UV. It may not be possible to remove OsHV-1 completely from the seawater of a recirculation system during a disease event using biofiltration and UV irradiation alone. The presence of food did not significantly affect the detection rate of OsHV-1 in seawater. The pellet produced by low speed centrifugation of seawater had ~2 to 5 fold greater odds of being positive for OsHV-1 by qPCR, in comparison to unprocessed seawater and the supernatant fraction. These results are consistent with the hypothesis that OsHV-1 may be attached to particles, which in this model may include algal feed, oyster faeces, fragments of oyster tissues or microorganisms introduced to the system with the oysters, but confirmation of this is required. Statement of relevanceIt is important to understand the nature and persistence of OsHV-1 microvariant within seawater in order to undertake aquaculture in hatchery environments, where water reticulation, recirculation, biofiltration, UV and other water treatments may be used. Understanding the distribution and mode of transmission of OsHV-1 within seawater has proven difficult. Experimental studies in controlled environments are important to furthering this understanding.

Age-dependence of cultured pearl grade and colour in the black-lipped pearl oysterPinctada margaritifera

Pinctada margaritifera is an economically important marine bivalve species for cultured pearl production in French Polynesian aquaculture. In order to evaluate the influence of donor oyster age on pearl quality traits, experiments were conducted over 6 years using both grafts and surgreffe operations. At harvest, 6 pearl quality traits were recorded and compared: surface defects, luster, grade, darkness level, and visual color. Analyzing the quality traits of pearls harvested in the initial graft process and those of pearls obtained from surgreffe experiments allowed a comparison of the influence of pearl sac cells originating from the initial mantle graft, which aged together with their recipient oysters. The results demonstrated a significant decrease between these successive grafts in luster, grade (A-B-C,) darkness level, and green color – traits that are of major importance in the pearl market. The duplicated graft experiment allowed the comparison of donor oyster families at 2 and 5 years old, where a mantle graft was inserted into recipient oysters aged 2.5 years old. The results showed the same tendencies to a lesser extent, with 1) an improved pearl grade, predominantly through a most important rate of 0 surface defect category, and 2) a green / grey ratio in favor of the younger donor. A comparison between the graft-surgreffe and the duplicated graft experiments also highlighted: 1) the indirect role played by the younger recipient oysters, which must be optimized for optimal pearl quality realization, and 2) the complex interplay between donor and recipient oysters.

Half-sib family effects on cultured pearl quality traits in the black-lipped pearl oyster Pinctada margaritifera: testing for indirect benefits of polyandry and polygyny

Breeding strategies and parental effects play a major role in genetic selection programmes for quality trait improvement. The black-lip pearl oyster, Pinctada margaritifera, is a protandrous hermaphrodite species in which the number of mature females is low, and these only appear after 4 years of culture. This difference underlines the importance of studying the contributions of dams and sires in the production and selection of donor pearl oyster families to choose the most appropriate breeding strategies for cultured pearl quality trait improvement. Laboratory crosses were made using wild dams and sires to produce half-sib families from polyandry and polygyny mating designs. The resulting half-sib families were used as donor oysters in a large grafting experiment of 900 grafts with random recipient oysters, using nuclei of the same size and brand, the same operating technician and the same grow-out site. After 18 months of culture, 497 cultured pearls were harvested. Comparison between half-sib pairs revealed that for nacre thickness, weight and shape, a polygyny effect exists, whereas no significant effect was observed for the polyandry mating. By contrast, no evidence of dam- or sire-related benefits was found for the presence/absence of circles, darkness level and colour categories or grade and its components: lustre and surface defects. These results should have important implications for the selective breeding of donor pearl oysters for hatchery management.

Comparison of harvested rate and nacre deposition parameters between cultured pearls issued from initial graft and second nucleus insertion inP. margaritifera

Cultured pearls produced with Pinctada margaritifera, using the surgreffe method (implantation of a second nucleus following pearl harvest) were studied for the first time to: (1) examine family effect on nacre thickness, nacre weight and nacre deposition speed and (2) compare variation in these three traits with that obtained from the cultured pearls previously harvested after the corresponding initial grafts. A surgreffe experiment using 783 recipient oysters was realized in Rangiroa atoll (French Polynesia). After 24 months of culture, 389 cultured pearls were harvested. Significant donor family effect was found for the harvested pearl rate from surgreffe (P = 0.046). Highly significant donor family effect was recorded for nacre thickness (P = 0.004). Very highly significant donor family effects were recorded for nacre weight and nacre deposition speed (P < 0.0001). Comparison between surgreffe and initial graft showed: (1) no significant effect for the average cultured pearl rate harvested (P = 0.052) and average cultured pearl nacre deposition speed (P = 0.622) and (2) very highly significant differences (P < 0.0001) for the average cultured pearl nacre thickness and nacre weight. This study highlighted three major implications for pearl industry management: (1) donor family effect was maintained from initial graft to surgreffe, for nacre thickness, weight and deposition speed, (2) the persistence of the pearl sac metabolic activity over three years of culture and (3) the relation between harvested pearl rate and the size of the nucleus inserted in the pearl sac.