Pacific Oyster Crassostrea Gigas Research Articles

Genetic Parameters of Growth and Survival in the Tetraploid Larvae of Pacific Oyster Crassostrea gigas

Genetic Parameters of Growth and Survival in the Tetraploid Larvae of Pacific Oyster Crassostrea gigas

DOPA Decarboxylase (DDC) in Pacific Oysters: Characterization and Role in Tyrosine Metabolism and Melanogenesis.

DOPA decarboxylase (DDC) plays a crucial role in the physiological functions of animals by participating in the dopaminergic system. However, the functions of DDC in shellfish remain poorly understood. The Pacific oyster (Crassostrea gigas) is an extensively cultivated shellfish. In this study, we characterized a DDC gene, designated CgDDC, from C. gigas. The CgDDC gene encodes a protein that contains a Pyridoxal_deC domain, which features specific binding sites for pyridoxal-5'-phosphate (PLP) and L-DOPA. CgDDC exhibits a significantly higher expression level in the black shell oyster strain than the white strain. In vitro enzymatic reaction assays demonstrated that CgDDC catalyzes the conversion of L-DOPA to dopamine. In vivo experiments revealed that inhibiting CgDDC activity reduced the expression of genes associated with tyrosine metabolism. Furthermore, the knockdown of CgDDC caused a decline in cAMP level and reduced transcription of genes involved in the cAMP-mediated melanogenesis. Additionally, treatment with L-α-DOPA inhibited CgDDC enzyme activity and cAMP-mediated melanogenesis; however, dopamine supplementation countered this inhibition, maintaining gene expression and melanin content at baseline levels. Collectively, our findings suggest that CgDDC is intricately involved in regulating tyrosine metabolism and melanogenesis in C. gigas.

Comprehensive identification and characterization of ribosomal protein genes in the Pacific oyster (Crassostrea gigas): Potential roles in reproduction and development.

Comprehensive identification and characterization of ribosomal protein genes in the Pacific oyster (Crassostrea gigas): Potential roles in reproduction and development.

Duplicate CgCREBL2β involved in the response of oyster upon high-temperature stress through the induction of glycolysis and haemocyte apoptosis.

Duplicate CgCREBL2β involved in the response of oyster upon high-temperature stress through the induction of glycolysis and haemocyte apoptosis.

Comprehensive analysis of sox genes in the Pacific oyster (Crassostrea gigas): Implications for sex differentiation and development

Comprehensive analysis of sox genes in the Pacific oyster (Crassostrea gigas): Implications for sex differentiation and development

Molecular phylogeny, morphology, and ultrastructure of a Mesomycetozoea member, Sphaeroforma nootkatensis isolated from Pacific oyster, Crassostrea gigas, on the Southern coast of Korea.

Molecular phylogeny, morphology, and ultrastructure of a Mesomycetozoea member, Sphaeroforma nootkatensis isolated from Pacific oyster, Crassostrea gigas, on the Southern coast of Korea.

Optimizing Genome Editing in Mollusks (Crassostrea gigas) in Vitro Validation of sgRNA and Identifying Key Factors Influencing Efficiency.

CRISPR-Cas9 genome editing holds tremendous potential for accelerating genetic improvements in aquaculture. The success of the CRISPR-Cas9 system relies on the specificity and efficiency of engineered single-guide RNAs (sgRNAs). In this study, we optimized an in vitro validation protocol for sgRNAs to streamline the gene editing process, capitalizing on the limited breeding season of the Pacific oyster (Crassostrea gigas). We evaluated the efficiency of 11 sgRNAs targeting four genes both in vitro and invivo in C. gigas. In addition, we found that Cas9 protein differs from Cas9 mRNA in gene editing efficiency at various stages of early development. Cas9 protein proved particular efficacy in achieving early and efficient gene knockout, functioning effectively during the first cell division and facilitating biallelic gene knockouts. Statistical analysis showed that in the protein group, the biallelic editing frequency ranged from 12.5% to 57.8%, and the overall editing frequency reached as high as 75-90.6%. The mRNA group exhibited a biallelic editing frequency of 3.1-14.0% and the overall editing frequency spanning 65.6-78.1%. Contrary to expectations, low-temperature incubation (20°C) of oyster embryos prolonged the time window for the first cell division but did not improve gene editing efficiency, likely due to the high temperature sensitivity of Cas9 enzyme activity. Together, this study provides a comprehensive analysis of factors affecting the efficiency of CRISPR-Cas9 gene editing in C. gigas, providing a robust framework for future gene editing endeavors in mollusks and other marine invertebrates.

LRSAM1 mediated the degradation of intracellular Vibrio through the ubiquitination-autophagy-lysosome pathway in oyster

The leucine rich repeat and sterile alpha motif containing 1 (LRSAM1) as E3 ligase recognizes bacteria and generates a ubiquitin signal to initiate the autophagy process. In the present study, LRSAM1 was identified from the Pacific oyster Crassostrea gigas (designed as CgLRSAM1), which was able to recognize various pathogen-associated molecular patterns and bacteria and directly ubiquitinate Vibrio splendidus. V. splendidus was co-localized with CgLRSAM1 and ubiquitin after invading haemocytes, and the ubiquitinated V. splendidus was then internalized into haemocyte lysosomes by p62-LC3-mediated autophagy. In haemocytes of CgLRSAM1-RNAi oysters, the activation of CgLC3 was enhanced after V. splendidus stimulation. While the co-localization values of V. splendidus with ubiquitin, CgLC3 and lysosomes all decreased significantly after V. splendidus stimulation. These results indicated that CgLRSAM1 functioned as E3 ligase responsible for anti-Vibrio-associated ubiquitination and regulated the degradation of bacteria through the ubiquitination-autophagy-lysosome pathway.

Evaluating Toxic Interactions of Polystyrene Microplastics with Hazardous and Noxious Substances Using the Early Life Stages of the Marine Bivalve Crassostrea gigas.

Plastics pose a significant threat to marine ecosystems, owing to their slow biodegradability. Microplastics (MPs), in particular, affect marine life and maricultural organisms and can enter the food chain via ingestion by marine organisms, leading to bioaccumulation in predators, including humans. This study assessed the toxic interactions between polystyrene microplastic particles (PSMPs) and cadmium (Cd) and phenanthrene (Phe) using marine bivalves. While PSMPs were non-toxic to Pacific oysters (Crassostrea gigas), the toxicity of Cd and Phe was concentration-dependent. In most conditions, PSMPs reduced the toxicity of Cd and Phe, but in simultaneous exposure, they acted as Cd messengers, altering the toxicity during the adult stage. This study confirms that PSMPs can interact with coastal environmental pollutants, thereby accelerating biotoxicity and posing a significant threat to marine wildlife, mariculture, and human health. It also highlights the need to assess MP toxicity in coastal environments and their interactions with pollutants.

Optimization of Microsatellite Multiplex PCRs for Triploidy Verification and Genetic Diversity Assessment in the Pacific Oyster, Crassostrea gigas.

The ploidy detection is crucial for the oyster industry. The objective of this study was to develop a method that verifies ploidy of the triploid Pacific oyster Crassostrea gigas by analyzing the diversity of triploid through microsatellite multiplex PCRs using fluorescent universal primers. We developed four information-rich multiplex PCR panels, comprising a total of 12 genomic microsatellites located in the genome of the C. gigas, distributed across seven chromosomes with an average of 14 alleles per locus. Each panel used M13(-21) primers labeled with specific fluorochrome dyes, and the forward primers for each locus were appended with M13(-21) sequences. We validated the approach to infer ploidy using flow cytometry as a reference, finding > 95% agreement between these methods, and demonstrated its potential utility to infer aneuploidy. Genotyping of 496 triploid samples from eight populations yielded 10 or more alleles per locus in 99.63% of samples in a single capillary electrophoresis. The correct assignment of triploidy depends on the number of markers with three unique allele fragments (MNM). Using semi-strict criteria of three unique alleles at one or more loci, the detection accuracy rate was 95.26% for triploids. Using the strict criteria of three unique alleles at two or more loci, the detection accuracy rate was 98.34%. Populations with reduced genetic diversity due to selective breeding were better suited for the semistrict criterion, maximizing triploid detection. And cultured populations were more suitable for evaluation using the strict criteria, which effectively reduced false-positive diploid assignment and increased triploid detection accuracy. The markers developed in this study were highly polymorphic and effective for assessing genetic diversity and distinguishing populations, providing a reliable tool for triploid detection and analysis in oyster breeding.

The Regulation of γ-Aminobutyric Acid on Antioxidative Defense Response of Pacific Oyster upon High-Temperature Stress.

Recent studies have found that high temperatures cause oxidative stress and even mass mortality in Pacific oysters (Crassostrea gigas). The role of γ-aminobutyric acid (GABA) in improving antioxidative defense in aquatic animals is increasingly of interest. In the present study, the oxidative stress of Pacific oysters to high-temperature stress was examined, and the regulation of GABA on the antioxidative defense was further investigated. Following 6 h of exposure to 28 °C seawater, a significant increase in the mRNA expression levels of nuclear factor-E2-related factor 2 (Nrf2), superoxide dismutase (SOD), and catalase (CAT), as well as the activities of SOD and CAT, was observed in the gill, compared to those at 0 h. An increase of glutamate decarboxylase (GAD), GABA receptor (GABAAR-α and GABABR-B) mRNA levels, and GABA contents were also detected after 28 °C exposure compared to those at 0 h. Furthermore, the activities and mRNA expression levels of SOD and CAT were significantly upregulated after GABA treatment, while decreased after either GAD inhibitor or GABA receptor inhibitor treatment under high-temperature stress. Meanwhile, the enhanced effects of GABA on antioxidant enzyme activities were reduced when Nrf2 was inhibited by ML385, accompanied by an increase in MDA content. After high-temperature stress, compared with the GABA treatment group, the activities and mRNA expression levels of SOD and CAT were significantly upregulated by GSK-3β inhibitor treatment. Meanwhile, the elevation of antioxidant enzyme activities by GABA was attenuated by the AKT inhibitor treatment. Collectively, GABA first activated GABA receptors under high-temperature stress and then increased the activities of SOD and CAT and reduced MDA content by AKT/GSK-3β and Nrf2 pathways to protect the oysters against oxidative damage upon stress. The present results offer new insights for understanding the regulation mechanisms of antioxidative defense by the neuroendocrine system in molluscs.

CgPPOX and CgFECH Mediate Protoporphyrin IX Accumulation Predominantly in the Outer Mantle Fold of Pacific Oyster Crassostrea gigas.

Mollusk shell is secreted through mantle folds and exhibits color polymorphism predominantly caused by pigments. Mantle tissue consists of three folds, with the outer fold playing crucial roles in shell formation. Among various pigments influencing shell color, porphyrins-especially protoporphyrin IX (PPIX)-significantly contribute to orange-shell phenotype. However, the distinct functions of mantle folds in shell coloration and pigmentation remain poorly understood. In this study, the frozen section of fresh mantles from orange and gold shell-color Pacific oyster Crassostrea gigas was observed to have distinct porphyrin distributions. Significant differences in porphyrin spectrum and precise PPIX concentrations were detected among mantle folds of shell-color strains C. gigas. In addition, key genes from porphyrin pathway: protoporphyrinogen oxidase (PPOX) and ferrochelatase (FECH) were characterized to exhibit distinct expression patterns among mantle folds of different shell-colored oysters. Dual-label fluorescence in situ hybridization analysis of CgPPOX and CgFECH with the PPIX translocator protein (CgTspO) verified the predominant functional location of these genes is the oyster outer mantle fold and periostracal groove. Furthermore, in vivo knockdown of CgPPOX and CgFECH verified their roles in PPIX metabolism, resulting in significant changes in porphyrin metabolic-related gene expression and altered PPIX concentrations in both the outer mantle fold and the newly deposited shell of C. gigas. This study identified the differential mantle fold porphyrin distribution and demonstrated essential roles of CgPPOX and CgFECH in PPIX metabolism, correlating with variations in PPIX content and shell color of C. gigas. The results provide new insights into molecular mechanisms underlying shell-color polymorphism in mollusks.

The Roles of Plasticity and Selection in Rapid Phenotypic Changes at the Pacific Oyster Invasion Front in Europe.

Invasive species present significant management challenges worldwide due to their ability to rapidly adapt to novel environments. The Pacific oyster Crassostrea gigas, a globally distributed invasive species, arrived in western Sweden in 2006 but has not yet colonised the low salinity waters of the Baltic Sea, presumably because low salinities act as a barrier to reproduction. We used classic mating designs to investigate fertilisation rates and heritability of embryonal salinity tolerance (in 8‰-33‰) in oysters from three locations with different invasion history and salinity (established, 33‰; past invasion front, 23.5‰; and present invasion front, 16‰). We found that fertilisation rates at lower salinities increased with proximity to the range front, with a pronounced heritable component. We then used whole-genome sequencing of oysters from the present invasion front to identify genomic regions showing stronger deviations from Mendelian inheritance in larval full-sib families reared in low salinity compared to controls. These regions contained coding sequences for Histones and ribosomal DNA, with the paternal genotype explaining a significant proportion of the deviation, suggesting the involvement of sperm in modulation of low-salinity tolerance at fertilisation and early development. Furthermore, we found no evidence of recent bottlenecks along the invasion front. We conclude that the Pacific oyster has developed low-salinity tolerant reproductive phenotypes at the present invasion front through acclimation and natural selection. Given the strong heritability for tolerance to low-salinities at fertilisation, the species likely has the potential to adapt further to low-salinity conditions and may invade the Baltic Sea.

Histone lysine demethylase 5 regulates haemocyte proliferation during immune priming in the oyster Crassostreagigas.

Histone lysine demethylase 5 regulates haemocyte proliferation during immune priming in the oyster Crassostreagigas.

Toxic effects of exposure to Polymethyl methacrylate and polyvinyl chloride microplastics in Pacific oysters (Crassostrea gigas)

Toxic effects of exposure to Polymethyl methacrylate and polyvinyl chloride microplastics in Pacific oysters (Crassostrea gigas)

Temporal and spatial co-occurrence of pacific oyster mortality and increased planktonic Vibrio abundance.

Oyster mortality and human food poisoning events are linked to pathogens from the Vibrio genus. However, the link between these events, planktonic bacterial dynamics and environmental variables has not yet been resolved. In Port Stephens, Australia, we characterized the microbial community and quantified the abundance of total Vibrio, Vibrio harveyi, and Vibrio parahaemolyticus in a (i) 27-month seawater planktonic microbial time-series; (ii) samples of Pacific oysters (Crassostrea gigas) during a mortality event and (iii) seawater samples following the mortality event. Vibrio harveyi and V.parahaemolyticus exhibited seasonal abundance, peaking during the summer months. Total Vibrio and V.harveyi in seawater were significantly greater at sites with high levels of oyster mortality and decreased 5-fold in the weeks following oyster mortality. Our findings provide evidence for the role of Vibrio in oyster mortality events and indicate that ocean warming and elevated phytoplankton may stimulate putative pathogens in the Vibrio genus.

Multi-scale influences on Escherichia coli concentrations in shellfish: From catchment to estuary.

Multi-scale influences on Escherichia coli concentrations in shellfish: From catchment to estuary.

The Potential Mechanism of Cuproptosis in Hemocytes of the Pacific Oyster Crassostrea gigas upon Elesclomol Treatment.

Cuproptosis is a novel cell death dependent on mitochondrial respiration and regulated by copper. While the study of it is mainly focused on tumor therapy, in the present study, two key cuproptosis-related genes, ferredoxin (FDX1) and dihydrolipoamide S-acetyltransferase (DLAT) homologs (designated as CgFDX1 and CgDLAT), were identified from Crassostrea gigas. CgFDX1 has a Fer2 domain with a 2Fe-2S cluster forming a unique ferredoxin. CgDLAT is composed of a biotin_lipoyl domain, an E3-binding domain, and a 2-oxoacid_dh domain. CgFDX1 and CgDLAT mRNA were expressed in all the examined tissues. After elesclomol treatment, both mRNA and protein expressions of them were reduced in the hemocytes. The mortality rate of the hemocytes increased significantly, and the hemocytes were accompanied with noticeable adhesive abnormalities and heightened secretion after elesclomol treatment. Additionally, the accumulation or depletion of actin was observed in the hemocytes. The integrity of the double membrane structure of the mitochondria was compromised, and the organization of mitochondrial cristae was disrupted. The contents of copper, malondialdehyde (MDA), pyruvic acid and mitoSOX as well as the ratio of cells with low mitochondrial potential increased significantly in the hemocytes upon elesclomol treatment and the content of citric acid decreased significantly. These findings suggest the potential presence of cuproptosis in oysters and its activation mechanism is relatively conserved in evolution.

GABA mitigates mitochondrial apoptosis induced by high temperature stress in the Pacific oyster (Crassostrea gigas).

High temperature is a critical environmental factor leading to mass mortality in oyster aquaculture in China. Recent advancements highlight the physiological regulation function of γ-aminobutyric acid (GABA) in the adaptation of environmental stress. This study examined the physiological responses of the Pacific oyster (Crassostrea gigas) upon high temperature exposure, focusing on the histopathological changes in gill, the GABA concentration, the mRNA expression and activities of apoptosis-related genes. Following 24h of exposure to seawater at 28°C, notable histopathological changes, including cellular swelling and vacuolization, along with an increase in TUNEL-positive cells were observed in the oyster gill, compared to the control group maintained at 18°C. Moreover, there was a significant increase in CgCaspase-3 transcripts, Caspase-3 and Caspase-9 activities in the gills, glutamate decarboxylase CgGAD transcripts in the haemocytes, and GABA concentrations in the haemolymph supernatant. Intervention with GABA markedly ameliorated these responses, including reducing the mRNA expression levels of CgBax, CgBak, CgCaspase-3, and CgCaspase-9, as well as the activities of Caspase-3/9. Furthermore, after the treatment with GABAA and GABAB receptor antagonists, the activities and expression levels of Caspase-3 and Caspase-9 significantly up-regulated under hightemperature stress. GABA treatment also significantly diminished the increased Caspase-3 activity by mitochondrial pathway apoptosis inducers. High temperature induced mitochondrial pathway apoptosis via increased caspase activities. The transcripts of CgGAD in haemocytes and GABA concentration in hemolymph supernatant also increased after high-temperature stress. GABA countered these effects through the activation of GABAA and GABAB receptors, reducing both caspase activity and expression of apoptosis-related genes.

General and Specific Combining Abilities of Pacific Oyster (Crassostrea gigas) and Fujian Oyster (C. angulata) for Growth, Survival and Thermal Tolerance of Their Reciprocal Hybrids

General and Specific Combining Abilities of Pacific Oyster (Crassostrea gigas) and Fujian Oyster (C. angulata) for Growth, Survival and Thermal Tolerance of Their Reciprocal Hybrids