Reproductive patterns of the Pacific oyster Crassostrea gigas in France

Summary

In France, national management programs focus research on understanding reproductive factors in Crassostrea gigas to confront problems of the oyster industry. However, little information has been documented in which reproductive patterns include sexual changes. The reproductive cycle of oysters at three sites of the Atlantic coast of France was examined from 1996 to 1998, and the seasonal variations in oocyte size-frequencies, and sex ratio were described. The results showed a synchronism within the population concerning reproductive behavior. Young oocytes are generated after spawning and show no apparent changes during winter. Growth of oocytes begins in spring and cells reach maturity in April-May and are ready for a single spawning season in June-July. Oocytes that were not released during spawning are reabsorbed within the gonad. The significant difference between sites is that spawning occurred 1 month later in the southern area. A modal analysis showed that oocyte populations in the sample individuals are primordially bimodal, but with polymodal occurrences in June-July, in some cases. Irregular alternative sexuality was detected at all sites, and hermaphrodites appear to be a transition phase that allows changes from male to female during early spring. Previous observations, together with the study of the development of oocyte cohorts over time, permit a hypothetical model concerning the kinetics of gametogenesis in C. gigas. The model suggests that primary oocytes are generated from energy supplied from degenerating, as well as young oocytes that do not reach the mature stage within the gonad during autumn-winter. It seems that, during vitellogenesis, there is disintegration of smaller cells coupled with transfer of energy to the larger oocytes, which continue to grow and mature.     

Reproduction of the introduced oyster Crassostrea gigas (Bivalvia: Ostreidae) cultured on rafts in Spain

The oyster Crassostrea gigas was introduced in Spain for aquaculture purposes; however, until now, it is not known whether populations are established in the wild, being necessary to define whether this species is spawning and which environmental variables trigger this process. The influence of environmental parameters on the reproduction of C. gigas was evaluated from January 2008 to October 2009 with oysters grown on a raft in the Ría de Arousa (Galicia, NW Spain). Temperature and chlorophyll a are directly correlated to sexual maturation. Oysters can mature at temperatures below 14°C. The temperature necessary for spawning differs between the sexes, requiring a temperature above 15°C for males and 18°C for females. Females had a single massive spawn between June and September, while males had partial spawning from May to December with two peaks, one in May–September and another in October–December, with the second peak more pronounced. The first spawning peak is related to high temperatures and concentrations of chlorophyll a, and the second spawning peak is mainly related to the food availability in the water. The spawning asynchrony may be impeding establishment of wild C. gigas populations in Spain.     

Predation by the endemic whelk Tenguella marginalba (Blainville, 1832) on the invasive Pacific oyster Crassostrea gigas (Thunberg, 1793)

The endemic mulberry whelk (Tenguella marginalba) is a common predator on Australian intertidal rocky shores. The introduced Pacific oyster (Crassostrea gigas), found within the natural range of T. marginalba, is potential prey for the whelk. In experiments designed to increase our understanding of predatory behaviour by the whelk on oysters, we found that adult T. marginalba detected C. gigas and increased movement in the presence of oyster prey. Tenguella marginalba showed a preference for smaller C. gigas, but consumed oysters up to 60?mm in shell height. To access oyster flesh, whelks used their radula to drill holes in the oyster’s shell. These holes were on average 0.68?±?0.09?mm in diameter, most frequently located central to the pericardial cavity on the right (upper) valve. Predation was greatest when predator and prey were both submerged, but was unaffected by a diurnal light cycle. When offered a choice among the native Sydney rock oysters (Saccostrea glomerata), mussels (Trichomya hirsuta) or the invasive C. gigas, whelks displayed no preference among prey. We conclude that the invasive oyster C. gigas represents a viable food source for T. marginalba, which may help to slow the spread of this invasive oyster throughout eastern Australia.     

Arsenic and Arsenic Species in Cultured Oyster (Crassostrea gigas and C. corteziensis) from Coastal Lagoons of the SE Gulf of California, Mexico

The aim of this study was to evaluate the bioavailability of arsenic (As) through cultured oyster Crassostrea gigas and Crassostrea corteziensis from four coastal lagoons (SE Gulf of California). Organisms were collected in two seasons (rainy and dry season), and they were analyzed for total arsenic and chemical speciation of this element. The concentrations of As in oyster soft tissue fluctuated between 5.44 and 9.56 μg/g for rainy season and 6.46 and 8.33 μg/g for dry season (dry weight) in C. gigas. In C. corteziensis, the As concentrations were <5 μg/g for both seasons (dry weight). Arsenic speciation indicated arsenobetaine as the major arseno-compound accounting for 43.2–76.3 % of total content of As. Lower contributions were obtained for non-extractable As (11.3–17.5 %) and other molecules such as arsenocholine and methyl-arsonate (<5 %). Inorganic arsenic was detectable in only two samples, at concentrations lower than <0.1 μg/g. These As data are the first generated for these mollusks in NW Mexico and indicate that C. gigas and C. corteziensis farmed in this area are safe for human consumption in terms of arseno-compounds.     

Mitochondrial and nuclear DNA phylogeography of Crassostrea angulata, the Portuguese oyster endangered in Europe

The respective status of the Portuguese oyster, Crassostrea angulata, and the Pacific oyster, Crassostrea gigas, has long been a matter of controversy. Morphological and physiological similarities, homogeneity of allozyme allelic frequencies between populations of the two taxa and the demonstration of hybridization lead most authors to suggest that they should be regrouped within the same species. The risk of introgression and the present expansion of C. gigas aquaculture in Europe raises the question of the need for preservation of C. angulata in Europe, as only a few populations remain. We studied European and Asian populations of C. gigas and C. angulata using microsatellite and mitochondrial DNA markers to estimate their genetic diversity and differentiation. The analysis of genetic distances and the distribution of allelic and haplotype frequencies revealed significant genetic differences between taxa, showing two clusters: (1) C. gigas French and Japanese populations and (2) C. angulata Portuguese and Taiwanese populations. The Asian origin of the Crassostrea angulata taxa is therefore confirmed. Unlike previous studies based on allozymes, significant nuclear genome differences were noted between C. angulata and C. gigas. Despite the presumed history of the introduction of C. angulata into Southern Europe, these populations did not show any significant reduction of variability compared to Taiwanese populations. Any conservation plans for European C. angulata populations should take its non-native origin into account. They represent a valuable genetic resources for European breeding program.     

<Emphasis Type="Italic">Crassostrea gigas</Emphasis> in natural oyster banks in southern Brazil

We report on the invasion of Brazil by the Pacific oyster Crassostrea gigas, and discuss the likely routes of invasion. Because this phenotypically diverse oyster sometimes resembles the native species C. brasiliana and C. rhizophorae, its invasion went unnoticed until it was detected through the analysis of DNA sequences for ribosomal 16S and the ribosomal second internal transcribed spacer. C. gigas was found amongst the native species in oyster banks up to 100 km south of oyster farms in South Brazil. Under most circumstances, water temperatures in the coastal southerly Brazil current would be too high to allow for the establishment of stable populations of C. gigas, but the production of spat in oyster farm laboratories has probably selected for resistance to warmer temperatures, which would promote invasion by C. gigas.     

The density and spatial arrangement of the invasive oyster Crassostrea gigas determines its impact on settlement of native oyster larvae

Understanding how the density and spatial arrangement of invaders is critical to developing management strategies of pest species. The Pacific oyster, Crassostrea gigas, has been translocated around the world for aquaculture and in many instances has established wild populations. Relative to other species of bivalve, it displays rapid suspension feeding, which may cause mortality of pelagic invertebrate larvae. We compared the effect on settlement of Sydney rock oyster, Saccostrea glomerata, larvae of manipulating the spatial arrangement and density of native S. glomerata, and non‐native C. gigas. We hypothesized that while manipulations of dead oysters would reveal the same positive relationship between attachment surface area and S. glomerata settlement between the two species, manipulations of live oysters would reveal differing density‐dependent effects between the native and non‐native oyster. In the field, whether oysters were live or dead, more larvae settled on C. gigas than S. glomerata when substrate was arranged in monospecific clumps. When, however, the two species were interspersed, there were no differences in larval settlement between them. By contrast, in aquaria simulating a higher effective oyster density, more larvae settled on live S. glomerata than C. gigas. When C. gigas was prevented from suspension feeding, settlement of larvae on C. gigas was enhanced. By contrast, settlement was similar between the two species when dead. While the presently low densities of the invasive oyster C. gigas may enhance S. glomerata larval settlement in east Australian estuaries, future increases in densities could produce negative impacts on native oyster settlement. Synthesis and applications: Our study has shown that both the spatial arrangement and density of invaders can influence their impact. Hence, management strategies aimed at preventing invasive populations reaching damaging sizes should not only consider the threshold density at which impacts exceed some acceptable limit, but also how patch formation modifies this.     

Mycobiota of the giant oyster <Emphasis Type="Italic">Crassostrea gigas</Emphasis> (Thunberg, 1787) (Bivalvia) from the Peter the Great Bay of the Sea of Japan

Mycological investigation of the Pacific (giant) oyster Crassostrea gigas (Thunberg, 1793) (Bivalvia) from the Peter the Great Bay of the Sea of Japan was carried out. The taxonomic composition of filamentous fungi associated with C. gigas was studied. The taxonomic composition of the fungi associated with the giant oyster included 22 species of filamentous fungi of which 17 species were identified. The latter belonged to six genera: Alternaria, Aspergillus, Botrytis, Fusarium, Penicillium, and Trichoderma. The distribution of filamentous fungi in the internal organs of the bivalve mollusk was studied.     

Spatio-temporal variations in biological performances and summer mortality of the Pacific oyster Crassostrea gigas in Normandy (France)

Mortality and biological performances of half-grown Crassostrea gigas were studied from spring 2000 to autumn 2001 at six instrumented stations located in two areas (Gefosse and Grandcamp) of the Bay of Veys (Normandy). Shell and meat growth, condition indexes and a macroscopic maturity index were determined on oysters deployed at the six stations in order to assess spatial variability in the influence of environmental conditions. In addition, histological and biochemical analyses were performed in order to determine the sex and establish the reproductive cycle (at all six sites) and the biochemical composition (at four stations). The data set including monthly mean temperatures and data provided by examination of 2,837 oysters were analysed by Principal Component Analysis and a Hierarchical Ascending Clustering which resulted in the formation of four clusters. The highest station on the shoreline belonged to a cluster characterized notably by low total weight due to a short immersion/feeding period, whereas all other stations belonged to another single cluster. Nevertheless, various biological differences were found between these stations, e.g. the reproductive cycle was generally synchronized throughout the bay but some differences relative to spawning occurrence were observed. In 2000, oyster mortality was higher at Gefosse than at Grandcamp, the latter being a more marine area. In 2001, oyster mortalities were significantly higher and all stations were strongly affected. In the Bay of Veys, oyster biological performances and mortality thus showed spatio-temporal variations which were worthy to be discussed.     

Characterization of 79 microsatellite DNA markers in the Pacific oyster Crassostrea gigas

We characterized 79 microsatellite DNA markers, which were obtained from genomic libraries enriched for CA, GA, ATG and TAGA motif repeats, in the Pacific oyster Crassostrea gigas. For eight F₁ grandparents or great‐grandparents of mapping families, the average heterozygosity, 0.705, and average number of alleles per locus, 5.7, did not vary among motif‐repeat or motif‐complexity categories. Non‐amplifying polymerase chain reaction null alleles, which were confirmed by segregation in the mapping families, were detected at 41 (51.9%) of the 79 loci. Cross‐species amplifications from C. angulata, C. sikamea, C. ariakensis and C. virginica showed a precipitous decline with distance from the focal species C. gigas.     

Enzyme-linked immunosorbent assay (ELISA) used in quantification of reproductive output in the pacific oyster,Crassostrea gigas,in Korea

Polyclonal antibody developed from egg proteins of the Pacific oysters, Crassostrea gigas, was used in quantification of the eggs. Quantity of the egg protein was measured using ELISA and a weight-normalized gonadosomatic index (GSI) was then established. Histology indicated that the oysters collected from Goseong Bay, Korea, in 2000 initiated gametogenesis as early as late February and spawned mostly in late June to early July when water temperature reached 23–25 °C. A second spawning peak was also observed in late August and the spawning activity continued to late October. Oyster egg proteins could be detected by ELISA in all months except March and November; a few oysters collected during December and January contained a measurable amount of eggs. The highest monthly mean GSI, 0.423, was observed in mid-June when the oysters were ready to spawn. The maximum GSI recorded in this study was 0.667. A positive correlation was found between oyster size and fecundity; the number of eggs increased as oyster biomass increased (r²=0.7497). Our data suggest that oysters in Goseong Bay produce 3–196 million eggs during the spawning season. The immunological method developed in this study was sensitive enough to measure a small quantity of eggs and considered to be method of choice for studying bivalve reproduction.     

A cDNA Microarray for Crassostrea virginica and C. gigas

The eastern oyster, Crassostrea virginica, and the Pacific oyster, C. gigas, are species of global economic significance as well as important components of estuarine ecosystems and models for genetic and environmental studies. To enhance the molecular tools available for oyster research, an international group of collaborators has constructed a 27,496-feature cDNA microarray containing 4460 sequences derived from C. virginica, 2320 from C. gigas, and 16 non-oyster DNAs serving as positive and negative controls. The performance of the array was assessed by gene expression profiling using gill and digestive gland RNA derived from both C. gigas and C. virginica, and digestive gland RNA from C. ariakensis. The utility of the microarray for detection of homologous genes by cross-hybridization between species was also assessed and the correlation between hybridization intensity and sequence homology for selected genes determined. The oyster cDNA microarray is publicly available to the research community on a cost-recovery basis.     

Specific Detection of Pacific Oyster (Crassostrea gigas) Larvae in Plankton Samples Using Nested Polymerase Chain Reaction

Management of sustainable Pacific oyster fisheries would be assisted by an early, rapid, and accurate means of detecting their planktonic larvae. Reported here is an approach, based on polymerase chain reaction (PCR), for the detection of Pacific oyster larvae in plankton samples. Species-specific primers were designed by comparing partial mitochondrial cytochrome oxidase subunit I (COI) sequences from Crassostrea gigas, with other members of the family Ostreidae including those of Crassostrea angulata. Assay specificity was empirically validated through screening DNA samples obtained from several species of oysters. The assay was specific as only C. gigas samples returned PCR-positive results. A nested PCR approach could consistently detect 5 or more D-hinge-stage larvae spiked into a background of about 146 mg of plankton. The assay does not require prior sorting of larvae. We conclude that the assay could be used to screen environmental and ballast water samples, although further specificity testing against local bivalve species is recommended in new locations.     

New Resources for Marine Genomics: Bacterial Artificial Chromosome Libraries for the Eastern and Pacific Oysters (Crassostrea virginica and C. gigas)

Large-insert genomic bacterial artificial chromosome (BAC) libraries of two culturally and economically important oyster species, Crassostrea virginica and C. gigas, have been developed as part of an international effort to develop tools and reagents that will advance our ability to conduct genetic and genomic research. A total of 73,728 C. gigas clones with an average insert size of 152 kb were picked and arrayed representing an 11.8-fold genome coverage. A total of 55,296 clones with an average insert size of 150 kb were picked and arrayed for C. virginica, also representing an 11.8-fold genome coverage. The C. gigas and C. virginica libraries were screened with probes derived from selected oyster genes using high-density BAC colony filter arrays. The probes identified 4 to 25 clones per gene for C. virginica and 5 to 50 clones per gene for C. gigas. We conducted a preliminary analysis of genetic polymorphism represented in the C. gigas library. The results suggest that the degree of divergence among similar sequences is highly variable and concentrated in intronic regions. Evidence supporting allelic polymorphism is reported for two genes and allelic and/or locus specific polymorphism for several others. Classical inheritance studies are needed to confirm the nature of these polymorphisms. The oyster BAC libraries are publicly available to the research community on a cost-recovery basis at     

Microsatellite Analysis of 6-Hour-Old Embryos Reveals No Preferential Intraspecific Fertilization Between Cupped Oysters Crassostrea gigas and Crassostrea angulata

Experimental examination of reproductive isolation is the first step in understanding hybridization processes. Here, we studied preferential fertilization between 2 cupped oyster taxa, Crassostrea angulata and Crassostrea gigas, as a potential prezygotic reproductive isolation. Early examination of sperm competition is now possible by molecular analysis of oyster embryos. This avoids the confounding effect of differential mortality during the larval stage. Six hundred embryos were sampled from 2 crosses. Three microsatellite loci were enough to determine without ambiguity the taxa of contributing sires of embryos. No evidence of preferential fertilization between gametes from the same taxa was shown. A significantly higher contribution of the C. gigas males was revealed with the C. angulata females, but not with the C. gigas females, which might suggest early heterosis or interaction differences between gametes. In the light of these results, natural hybridization between both taxa can be expected in cases of their geographical coexistence, as in the Southern European populations in which both taxa are in contact as a result of aquaculture development. Received May 6, 2000; accepted March 6, 2001.     

Cloning of cDNAs and hybridization analysis of lysozymes from two oyster species, Crassostrea gigas and Ostrea edulis

In bivalve molluscs including oysters, lysozymes play an important role in the host defense mechanisms against invading microbes. However, it remains unclear in which sites/cells the lysozyme genes are expressed and which subsequently produced the enzyme. This study cloned lysozyme cDNAs from the digestive organs of Pacific oyster Crassostrea gigas and European flat oyster Ostrea edulis. Both complete sequences of two oysters' lysozymes were composed of 137 amino acids. Two translated proteins present a high content in cysteine residues. Phylogenetic analyses showed that these oysters' lysozymes clustered with the invertebrate-type lysozymes of other bivalve species. In the Pacific oyster, lysozyme mRNA was expressed in all tissues except for those of the adductor muscle. In situ hybridization analyses revealed that lysozyme mRNA was expressed strongly in basophil cells in the digestive gland tubule of C. gigas, but not in digestive cells. Results indicated that the basophil cells of the oyster digestive gland are the sites of lysozyme synthesis.