Phenotypic indicators for cultured pearl size improvement in the black‐lipped pearl oyster (Pinctada margaritifera): towards selection for the recipient growth performance

The black‐lipped pearl oyster, Pinctada margaritifera, is the most important farmed species in French Polynesia and the basis of the most valuable export industry. Mass production of black pearls relies on a surgical operation requiring tissue from a donor pearl oyster to be grafted, together with a nucleus made of shell, into the gonad of a recipient oyster. Improving pearl size through family selection remains one of the main challenges for future aquaculture development. This study analyses the relative contribution of donor and recipient oysters to pearl size. To this end, hatchery‐produced donor oysters of two batches, large and small (based on shell height), were used to supply grafts for recipients, which were then monitored individually for their growth performance by recording shell height, width, and thickness, and total live weight (flesh + shells) every 6 months (four biometric measurement times) over 20 months of culture. Pearls issued from the two batches of donors showed no significant differences in nacre weight or thickness. In contrast, recipient oyster shell height and total weight were increasingly positively correlated with these pearl size parameters over the culture period, becoming significant at 8 months post‐grafting. Potential therefore exists to use shell height and oyster weight as phenotypic indicators for selective breeding of recipient oysters with high growth performance to increase pearl size in P. margaritifera.     

Donor effect on cultured pearl nacre development and shell matrix gene expression in Pinctada margaritifera reared in different field sites

Understanding the respective roles played by donor and recipient pearl oysters in pearl quality determination in relation to the environment is a challenge for the pearl industry. In most Pinctada species, pearl size is mainly related to recipient oyster growth performance but also relies to some extent on the biomineralisation activity of the pearl sac, a tissue that originates from the donor oyster mantle. We examined donor effect on pearl size in response to culture in the lagoons on Arutua and Apataki atolls. Overall, nacre weight and thickness were greater in Arutua than in Apataki, but sensitivity to the environment differed between donors. Some donors were associated with significantly heavier and thicker nacre in Arutua (I group), while others had similar results at the two sites (NI group). On average, up to 20% of the pearl size could be attributed to the donor but, in group I, donor effect was responsible for up to 36% of nacre weight determination. Additionally, a real‐time PCR expression study of eight matrix protein genes related to biomineralisation in the pearl sac showed that MSI60, pearlin and pif177 were significantly and positively correlated with nacre weight and thickness, with the latter two genes explaining the larger pearl size observed in Arutua. Donor oysters in P. margaritifera therefore play a key role in pearl size improvement, related to the role of the shell matrix protein genes. Understanding such contributions could help in the design of genetic selection plans for specific and adapted donor oyster lines.     

Is pearl colour produced from Pinctada margaritifera predictable through shell phenotypes and rearing environments selections?

The black‐lipped pearl oyster, Pinctada margaritifera, is the most important farmed mollusc species in French Polynesia. Donor oyster selection among wild P. margaritifera individuals, chosen according to their inner shell colour, makes it possible to obtain the broadest range of cultured pearl colours of any species. This study demonstrates the relative influence of using black [B] or red [R] outer shell phenotypes, combined with green [G] or yellow [Y] inner shell phenotypes, on pearl darkness level, colour categories and lustre. A large scale grafting experiment was designed and carried out over five grow‐out locations, covering three archipelagos: Tuamotus, Society and Gambier. Results revealed that the [B + G] phenotypes may be used as donors to produce dark green pearl, which suit the demands of the Asian market; whereas, phenotypes incorporating [R] and/or [Y] phenotypes may be used to obtain multicolour pearls of medium/light darkness, which suit the demands of the European market. From an environmental point of view, the 1) [B] phenotype showed no significant variation for light and other pearl colour production, and 2) [Y] phenotype produced both the same rate of pearl darkness level and green colour pearls whatever the grow‐out location. A classification tree model was built to predict, according to shell phenotype and culture location, the colour and darkness level of harvested pearls. Lustre was shown to be more influenced by the environment than by phenotype. These results should be taken into account in pearl farm production management and in selective breeding programmes.     

Factors influencing the quality of half‐pearls (mabé) produced by the winged pearl oyster,Pteria penguin (Röding, 1758)

The winged pearl oyster, Pteria penguin, is cultured primarily to produce half‐pearls (mabé). The mabé quality is influenced by culture techniques, but there is limited information in this field. P. penguin with mean (±SE) dorso‐ventral height of 250 ± 6.5 mm were used to investigate the influence of culture period and nucleus position on mabé quality. Oysters were relaxed using 1‐propylene phenoxetol, and five nuclei were glued at different positions to the inner surfaces of the oyster shells; three on the more concave left valve and two on the right valve. Nucleated oysters were then cultured for 10 months under commercial pearl farming conditions at Savusavu in Fiji. Nacre deposited at the base and top of the nuclei was measured monthly, from the 6th to the 10th months of culture and the different qualities of mabé produced at different positions were scrutinized. Nacre thicknesses at the base and top of the resulting mabé were significantly different at different months (P < 0.05) and the rate of nacre deposition was highest during the warmer months. The different positions of nuclei on the valve greatly affected the quality of mabé formed. After a 10‐month culture period, around 1 mm of nacre covered the nuclei although the best quality mabé were obtained after 9 months.     

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

Age‐dependence of cultured pearl grade and colour in the black‐lipped pearl oyster Pinctada 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, six pearl quality traits were recorded and compared: surface defects, lustre, grade, darkness level and visual colour. Analysing 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 lustre, grade (A‐B‐C), darkness level, and green colour – 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. The results showed the same tendencies to a lesser extent, with (i) an improved pearl grade, predominantly through a most important rate of 0 surface defect category, and (ii) a green/grey ratio in favour of the younger donor. A comparison between the graft‐surgreffe and the duplicated graft experiments also highlighted: (i) the indirect role played by the younger recipient oysters, which must be optimized for optimal pearl quality realization, and (ii) the complex interplay between the donor and recipient oysters.     

Impact of season and grafter skill on nucleus retention and pearl oyster mortality rate in Pinctada margaritifera aquaculture

The mollusc Pinctada margaritifera is the top economic aquaculture species in French Polynesia (export value) and forms the basis of the black pearl industry. Mass production of this unique gem, produced by a living organism, relies on a surgical operation requiring tissue from a donor pearl oyster to be grafted, together with a nucleus made of shell, into the gonad of a receiving pearl oyster. This technique is performed by expert grafters, whose work constitutes the first step influencing pearl farm production yield. This study makes the first report of effects mediated by individual grafters and by the season of grafting on three rates scored at 45 days post operation: nucleus retention, nucleus rejection and pearl oyster mortality. These results were obtained in a very large scale grafting experiment, designed and conducted in a single culture site in the atoll of Arutua (Tuamotu Archipelago) and involving a total of 52,910 grafts performed by ten professional grafters, during two contrasting seasons: autumn (four grafting experiments) and spring (three grafting experiments). Statistical analysis using linear mixed-effect model for both univariate and multivariate analysis was performed. The results show a very highly significant season effect (p < 0.001), with a higher rate of nucleus retention in autumn than spring: 91.1 versus 79.4 %, respectively. A very highly significant season effect was also recorded for nucleus rejection rate (p < 0.001), which was greater in spring than in autumn: 12.6 versus 7.7 %. Oyster mortality rate was up to six times higher in spring than in autumn (p < 0.001). Within the two seasons, there was no significant difference between the grafting experiments for any of the three variables, adding robustness to the inter-season results. Taking into account this seasonal variation effect, grafter skill had a significant impact on the three variables (p < 0.001), making it possible to rank grafters from most to least efficient. Aside from skill differences between grafters, grafting in autumn may significantly increase graft success. Water temperature and its indirect consequences could be one of the most important environmental factors implicated in overall efficiency. These findings may help the potential development of an annual grafting schedule according to season/ lagoon temperature as a way to maximize production yield for the black-lipped pearl oyster industry in French Polynesia.     

Macro‐geographical differences influenced by family‐based expression on cultured pearl grade,shape and colour in the black‐lip ‘pearl oyster’ Pinctada margaritifera: a preliminary bi‐local case study in French Polynesia

In French Polynesia, the aquaculture of Pinctada margaritifera is carried out in numerous grow‐out sites, located over three archipelagos (Gambier, Society and Tuamotu). To evaluate the impact of macro‐geographical effects of these growing sites on pearl quality traits, five hatcheries produced families were used as homogeneous donor oysters in an experimental graft. The molluscs were then reared in two commercial locations: Tahaa Island (Society) and Rangiroa atoll (Tuamotu). At harvest, eight pearl quality traits were recorded and compared: surface defects, lustre, grade, circles, shape categories, darkness level, body and secondary colour and visual colour categories. Overall inter‐site comparison revealed that: (1) all traits were affected by grow‐out location except for lustre and round shape, and (2) a higher mean rate of valuable pearls was produced in Rangiroa. Indeed, for pearl grade, Rangiroa showed twice as many A‐B and less reject samples than Tahaa. This was related to the number of surface defects (grade component): in Rangiroa, twice as many pearls had no defects and less pearls had up to 10 defects. Concerning pearl shape, more circled and baroque pearls were found in Tahaa (+10%). For colour variation, 10% more pearls have an attractive green overtone in Rangiroa than in Tahaa, where more grey body colour were harvested. Lustre does not seem to be affected by these two culture site (except at a family scale). This is the first time P. margaritifera donor family have been shown to vary in the quality of pearls they produce depending on their grow‐out location.     

How cultured pearls acquire their colour

Round nucleated pearls are produced through a surgical operation, where a round nucleus and a mantle tissue ‘saibo’ from donor oyster are inserted into the gonad of the host oyster. The epithelial cells in the mantle tissue proliferate around the nucleus, and thus, the pearl sac is formed. Pearl sac secrets nacre and forms a pearl. The quality and economic value of pearls are assessed by pearl features such as colour, brightness, lustre and shape. Among all these features, colour has been reported as an important economic indicator and has been widely studied by researchers. Generally, pearl colour is affected by the donor oyster which is determined genetically and biological pigments (melanin and carotenoid). Organic matrices, metal ions and other factors have also been reported to influence the colour of a cultured pearl. Recently, multi‐omics methods have been used to study the colour formation of pearl, and some key genes and signal pathways related to the colour formation of pearls have been identified. Nevertheless, the specific mechanism of pearl formation needs further research. The review combines both fresh and sea water pearls focusing on Hyriopsis cumingii and pearl oysters to provide a general overview and understanding for pearl colour formation.     

Pearl formation in the Japanese pearl oyster (Pinctada fucata) by CaCO3 polymorphs: Pearl quality‐specific biomineralization processes and their similarity to shell regeneration

Pearl oyster shell consists of two layers: a calcite prismatic layer (outer layer) and an aragonite nacreous layer (inner layer). Calcite and aragonite are CaCO₃ polymorphs, and their formations are controlled by shell‐forming tissue called mantle. Pearl sacs originating in the mantle form cultured pearls. Therefore, it has been widely accepted that pearl and shell are produced by the same process. However, this idea has been called into question by some recent mineralogical studies indicating microstructural and crystal‐polymorphic diversity in pearls. The pearl biomineralization process is still not fully understood in detail. Thus, in this study, we focused on the diversity of CaCO₃ polymorphism of non‐nacreous structures (NNSs) underlying the nacreous layer in pearl and regenerated shell, to reveal the biomineralization process of the Japanese pearl oyster (Pinctada fucata). Using Meigen's stain and scanning electron microscope‐energy dispersive X‐ray (SEM‐EDX), NNSs polymorphs in valuable and valueless pearls, in addition to regenerated shell, were compared. Aragonite was exclusively observed in the NNSs of valuable pearls, whereas calcite was dominant in those of valueless pearls. The same analysis of NNSs of regenerated shells was carried out. As in valueless pearls, almost all regenerated shell NNSs consisted of calcite, but one NNS was composed of aragonite. Accordingly, it seems that pearls are formed by the same biomineralization process as shell regeneration rather than shell formation.     

Cost–benefit analysis of two culture methods that influence pearl production from the black‐lip pearl oyster,Pinctada margaritifera

The black‐lip pearl oyster, Pinctada margaritifera, used for round pearl production in Polynesia, is generally cultured using “ear‐hanging” where they are attached to a rope to form “chaplets.” In other countries, pearl oysters are cultured using panel (pocket) nets that are more expensive than chaplets but afford more protection to cultured oysters. Prior research has shown panel nets produce pearls of higher quality and value, potentially generating higher profits. This study used cost–benefit analysis to compare pearl production using chaplet‐based and panel net‐based culture methods. Whole farm data, including gross revenues and annual production costs, fixed and variable, were analyzed. Average production cost per pearl using panel net‐based culture was USD 22.47 and for chaplet‐based culture was USD 21.55. However, use of panel nets saved around 3,430 hr (USD 6,860) of labor a year, offsetting the greater capital investment. A chaplet‐based pearl farm generated USD 65,738 in annual profits compared to USD 88,774 for a panel net‐based farm. Positive cash flow was achieved 1 year earlier (Year 7) for the panel net‐based farm. This is the first economic analysis of different pearl culture methods for P. margaritifera and evidence of profitability will support further development of the black‐lip pearl industry in the Indo‐Pacific region.     

Expression profiles of nine biomineralization genes and their relationship with pearl nacre thickness in the pearl oyster,Pinctada fucata martensii Dunker

Pinctada fucata martensii is an ideal animal for study of biomineralization. Although dozens of genes have been identified, the molecular mechanism of biomineralization remains still unclear. The purpose here was to discover the expression profiles of nine biomineralization genes in related tissues: mantle edge (ME), mantle centre (MC) and pearl sac (PS), and to explore the relationships between expression level and nacre thickness. The expression levels of seven genes (ACCBP, aspein, CaM, EFCBP, KRMP, nacrein and prismalin‐14) showed no significant difference between ME and MC, but were significantly higher than those in PS. The expression level of N19 was the same in ME and PS, significantly higher than that in MC. However, msi60 in ME, MC and PS showed no significant differences in expression level. In addition, the correlations of expression levels between ME, MC and PS were highly significant, indicating that there are some similar function and mechanism among the three tissues. Also the nine genes had significant correlation of expression levels between one another. Furthermore, no other genes from oysters which produced pearls of different grades showed significant difference in the expression level, except CaM in MC, which had significantly higher expression level in the medium pearl grade than in the plain nucleus grade. Moreover, the expression level of msi60 in MC of host oyster was significantly correlated with produced pearl layer thickness, suggesting that msi60 might play an important role in pearl formation and host oyster may also participate in the development of pearl.     

Growth of wild pearl oysters <Emphasis Type="Italic">Pinctada fucata,Pinctada margaritifera</Emphasis> and <Emphasis Type="Italic">Pinctada sugillata</Emphasis> (Bivalvia: Pteriidae) in Taiwan

In order to understand growth features of pearl oysters in the genus Pinctada, i.e. Pinctada fucata, Pinctada margaritifera, and Pinctada sugillata in Taiwan, a total of 3062 wild individuals of these species from juvenile to adult were collected monthyly from March 2001 to April 2002 in Jukeng, Pingtung County, south-west Taiwan. Quantitative measurements of live oysters were conducted for shell height (SH), shell length (SL), shell width (SW), hinge length (HL), and wet weight (WW). Different cohorts were identified through multiple length frequency analysis on SH of P. fucata and P. margaritifera, and growth curves with seasonal variation were estimated for these species. Pinctada fucata in Taiwan had a different seasonal growth pattern from the Japanese population, but had similar growth rates during the high growth period. The growth rate of P. margaritifera in Taiwan was slower than in French Polynesia, the Solomon Islands, and the Red Sea. Comparisons of morphological growth features among the three species show large differences in the SW-related features. Pinctada fucata in Taiwan had larger SW than in Japan and Korea. The differences in growth rates and morphological features suggested that the wild Taiwanese oysters may retain genetically pristine characteristics, thus genetic conservation might be urgently needed.     

Two epizootic Perkinsus spp. events in commercial oyster farms at Santa Catarina,Brazil

Perkinsus spp. have been detected in various bivalve species from north‐east Brazil. Santa Catarina is a South Brasil state with the highest national oyster production. Considering the pathogenicity of some Perkinsus spp., a study was carried out to survey perkinsosis in two oyster species cultured in this State, the mangrove oyster Crassostrea gasar and the Pacific oyster Crassostrea gigas. Sampling involved eight sites along the state coast, and oyster sampling was collected during the period between January 2013 and December 2014. For the detection of Perkinsus, Ray's fluid thioglycollate medium (RFTM) and histology were used, and for the identification of the species, PCR and DNA sequencing were used. Perkinsus spp. was found by RFTM in C. gigas and C. gasar from São Francisco do Sul. This pathology was also detected in C. gasar from Balneário Barra do Sul both, by RFTM and histology. Perkinsus marinus was identified in C. gigas and C. gasar from São Francisco do Sul and Perkinsus beihaiensis in C. gasar from Balneário Barra do Sul. This is the first report of P. marinus in C. gigas from South America. Results of this preliminary study suggest that both oyster species tolerate the species of Perkinsus identified, without suffering heavy lesions.     

Growth performance and biochemical composition of the oysters Crassostrea sikamea,Crassostrea angulata and their hybrids in southern China

The Kumamoto oyster (Crassostrea sikamea) and the Portuguese oyster (C. angulata) are important aquaculture species which naturally coexist along the southern coast of China. To understand the potential feasibility of hybridization between the two species, we conducted two‐by‐two factorial cross‐experiments in Beihai (Guangxi province), and also compared the survival and growth of the hybrids to that of the two parental progenies during the grow‐out period from July 2014 to July 2015. Genetic analysis confirmed that the hybrid spats were true hybrids. Additionally, the biochemical composition of the 1‐year‐old oyster progenies was determined. In July 2015, the mean shell height of the hybrids was 42.98 ± 6.29 mm, which was higher than that of the Kumamoto oyster progeny. The cumulative survival rate of the hybrids was 26.37 ± 1.32%, which was higher than that of the progeny of the Portuguese oyster. Mean lipid content of the hybrids was 13.65 ± 1.63% of dry weight, which showed obvious heterosis compared to those of the two parental progenies. Observation of gonads revealed that all hybrids were completely fertile. Furthermore, relative expression of the lipid homeostasis genes, SREBP (sterol regulatory element‐binding proteins), PPARα (peroxisome proliferator‐activated receptor) and INSIG (insulin‐induced gene) were found to vary between parental progenies and the hybrids, thus providing a possible reason for difference in the lipid contents of these experimental groups. Overall, the hybrids were viable, rich in lipid and completely fertile and thus could serve as a promising aquaculture stock for oyster breeding in southern China.     

Contribution of donor and host oysters to the cultured pearl colour in Pinctada martensii

Cultured free round pearls are produced by implanting a spherical nucleus with a small piece of nacre‐secreting mantle graft from a donor oyster into the gonad of a recipient oyster (host). To examine the possible contribution of host and donor oysters to the colouration of the harvested pearls, the CSE‐1 Imaging and Color‐Measuring System were used to quantitatively measure the L*a*b* values of donor and host shells and the produced pearls in Pinctada martensii. Results showed that the colour of pearls had significant positive correlation (r = 0.1–0.22, P = 0.00) with that of donors, but had no correlation with that of host oysters, thus convincingly confirmed the contribution of nacre colour of donor to the realization of pearl quality of colour. To further clarify the relationship between the donor and the pearl colour, the donors from pearls of good and poor colour quality were further compared and the results demonstrated the significant difference in L* values (P < 0.05) and insignificant difference in a* and b* values, suggesting the necessity of selecting donors with bright and lustrous nacre in pearl production.     

Effects of replacing microalgae with an artificial diet on pearl production traits and mineralization‐related gene expression in pearl oyster Pinctada fucata martensii

The effects of replacing dietary microalgae with artificial diet on pearl production traits and biomineralization‐related gene expression were investigated in pearl oyster (Pinctada fucata martensii). Three experiment groups (EG1, EG2 and EG3) were set. EG1 and EG3 were separately fed Platymonas subcordiformis and artificial diet (D4), respectively, and EG2 was fed with mixed P. subcordiformis and D4. A control group (CG) was cultured in natural sea. All groups were continuously fed for 150 days. The results showed that the survival rates of EG1, EG2 and EG3 were significantly higher than that of CG. No significant differences in retention rate and pearl thickness were observed among the four groups, but CG had the lowest values of both parameters. The relative expression levels of EGFR and FGF18 mRNA did not significantly differ among EG1, EG2 and EG3. Moreover, the relative expression levels of GHITM and TβR I mRNA significantly varied among the groups and EG3, and EG1 had the highest and lowest relative expression levels of GHITM and TβR I mRNA respectively. The relative expression levels of nacrein and chitin‐binding mRNA significantly differed among the groups, and EG1 had the lowest expression levels of nacrein and chitin‐binding mRNA. No significant differences in the relative expression levels of MSI60 and TIMP mRNA were observed among the groups. The results suggested that D4 can replace part of microalgae, which is helpful for further studies on developing artificial diet of pearl oyster.     

Extreme ocean acidification reduces the susceptibility of eastern oyster shells to a polydorid parasite

Ocean acidification poses a threat to marine organisms. While the physiological and behavioural effects of ocean acidification have received much attention, the effects of acidification on the susceptibility of farmed shellfish to parasitic infections are poorly understood. Here we describe the effects of moderate (pH 7.5) and extreme (pH 7.0) ocean acidification on the susceptibility of Crassostrea virginica shells to infection by a parasitic polydorid, Polydora websteri. Under laboratory conditions, shells were exposed to three pH treatments (7.0, 7.5 and 8.0) for 3‐ and 5‐week periods. Treated shells were subsequently transferred to an oyster aquaculture site (which had recently reported an outbreak of P. websteri) for 50 days to test for effects of pH and exposure time on P. websteri recruitment to oyster shells. Results indicated that pH and exposure time did not affect the length, width or weight of the shells. Interestingly, P. websteri counts were significantly lower under extreme (pH 7.0; ~50% reduction), but not moderate (pH 7.5; ~20% reduction) acidification levels; exposure time had no effect. This study suggests that extreme levels – but not current and projected near‐future levels – of acidification (?pH ~1 unit) can reduce the susceptibility of eastern oyster shells to P. websteri infections.