Pacific Oyster Crassostrea Gigas Research Articles

Environmental Influence on Feeding and Biodeposition Rates of Pacific Oysters (Crassostrea gigas) Throughout Its Culture Cycle in a Coastal Lagoon with Upwelling Influence

Environmental Influence on Feeding and Biodeposition Rates of Pacific Oysters (Crassostrea gigas) Throughout Its Culture Cycle in a Coastal Lagoon with Upwelling Influence

Investigations of the involvement of Vibrio species with Ostreid herpesvirus-1 in mass mortality events in the Pacific oyster Crassostrea gigas

Pacific oyster mortality syndrome (POMS) in Pacific oysters Crassostrea gigas is defined by mass mortality with Ostreid herpesvirus I (OsHV-1). In this study the association of Vibrio species with oysters before, during and after POMS events was investigated at population level to clarify their involvement with OsHV-1 in mass mortality events in the Georges and Hawkesbury River estuaries, Australia. In the POMS affected Georges River estuary, three patterns were observed concurrently at different sites: i) Vibrio counts and OsHV-1 DNA concentrations increased in adults and spat as mortalities started then decreased as mortalities stopped ii) bacterial counts increased in spat but not in adults associated with mortalities due to OsHV-1 iii) bacterial counts increased in the absence of mass mortality or high concentrations of OsHV-1 DNA. Although almost half of the 120 bacterial isolates were identified as an unknown Vibrio species belonging to the Splendidus clade with two predominant biotypes (A and H), there was no association between biotype and mortality. Concurrently in the Hawkesbury River estuary, neither mortality nor OsHV-1 were found, Vibrio counts fluctuated significantly and of 84 isolates identified, V. alginolyticus, V. parahaemolyticus and unidentified Vibrio sp. dominated; only five Splendidus clade isolates were identified. However, when POMS emerged in the Hawkesbury River estuary, the Splendidus-clade isolate became the dominant bacterial species. Mortality also occurred there without OsHV-1 and the dominant bacteria then were V. alginolyticus and V. harveyi, but the Splendidus-clade isolate was not detected. None of the bacterial species were specifically associated with POMS, although the Splendidus clade isolate increased in prevalence after POMS emerged in the Hawkesbury River estuary. V. aestuarianus was not detected in this study. While the findings suggest that bacterial proliferation follows infection with OsHV-1 in POMS, this was not a consistent feature. The number and diversity of bacteria within oyster tissues varied over time, between estuaries, between sites within an estuary and were associated with flooding, growing height, the age of oysters, mortality and OsHV-1.

Assessing selection potential for Pacific oyster (Crassostrea gigas) to a North American OsHV-1 μvar: Comparing two experimental assay methods

Ostreid Herpesvirus - 1 is a global threat to Pacific oyster aquaculture as catastrophic farm losses have occurred where OsHV-1 microvariants (μvars) become endemic. Genetic selection has proven effective for increasing survival to OsHV-1 exposure in multiple Pacific oyster breeding programs, but a novel μvar detected in San Diego Bay California has the potential to severely impact commercial aquaculture along the North American Pacific coast as no OsHV-1 μvar selected population is available commercially. Using a Molluscan Broodstock Program population descending from imports originating in the Miyagi prefecture (Japan), we evaluated the potential for genetic selection to improve survival against the San Diego Bay OsHV-1 μvar using a laboratory trial. Two experimental methods were evaluated, a common garden method where all 15 full-sibling families were pooled, and a petri dish method, where each family was confounded within replicate. No striking differences were found between experimental methods, and both experiments displayed high heritability (0.69 and 0.64) of the tested trait, a broad range of estimated breeding values, and a moderate prediction accuracy for the survival trait. A significant genotype-by-environment interaction occurred, highlighting variability in laboratory-based disease challenges, but the two experiments yielded similar results with minor to moderate re-ranking of mean family survival. Overall, the Miyagi population of Pacific oysters curated by the Molluscan Broodstock Program is a promising population to use for artificial selection to increase survival against San Diego Bay OsHV-1 μvar in attempts to obviate farm losses if the virus expands its currently isolated range.

Genome-wide association study and genomic prediction of resistance to summer mortality in Pacific oyster (Crassostrea gigas) using whole genome resequencing

The Pacific oyster Crassostrea gigas is one of the most important farmed marine shellfish worldwide. However, massive mortality during the summer months caused significant economic losses to the C. gigas farming industry. Understanding the genetic architecture of resistance traits has been an ongoing research issue, and the incorporation of genomic information into breeding programs is expected to accelerate the process of genetic improvement. Genomic selection (GS), as a new approach of genetic improvement, has great potential for breeding new lines with adaptive advantages. In this study, we conducted a genome-wide association study (GWAS) for summer mortality resistance and estimated the accuracy of genomic predictions using different methods. The heritability of the C. gigas summer mortality resistance was low, with heritability of 0.108 ± 0.036, 0.161 ± 0.102 and 0.134 ± 0.043 for PBLUP, GBLUP and ssGBLUP, respectively. In addition, we detected 9,783,674 SNPs and used a mixed linear model to identify 18 significant SNPs associated with summer mortality resistance. The phenotypic variance explained (PVE) for these SNPs ranged from 8.25% to 10.14%. Based on these significantly related SNPs, nine significant candidate genes were identified (TLR4, HEX, SLC22A8, PDE2A, HUWE1, SLMAP, RAD52, TBK1 and RAPH1). The prediction accuracy using PBLUP, GBLUP, ssGBLUP, and weight ssGBLUP (WssGBLUP) was 0.549 ± 0.026, 0.381 ± 0.074, 0.544 ± 0.026, and 0.869 ± 0.018, respectively. Therefore, WssGBLUP models are more suitable for genomic prediction of summer mortality resistance in C. gigas. Our results suggest a polygenic genetic architecture that provide new perspectives for studying candidate genes for resistance to summer mortality, which may facilitate the genetic improvement for resistance lines in C. gigas.

First report of Perkinsus marinus occurrence associated with wild Pacific oysters Crassostrea gigas from the west coast of Korea

This study reports the occurrence of Perkinsus marinus associated with wild Pacific oyster (Crassostrea gigas) specimens collected along the west coast of Korea. Confirmation of P. marinus presence was achieved by conventional PCR using World Organization of Animal Health (WOAH)-recommended primers that specifically targeted regions of the rDNA locus (ITS1, 5.8S, and ITS2). Sequencing of 10 samples revealed two distinct sequences differing by a single base pair, indicating potential haplotype variability. One sequence closely resembled the P. marinus strain found in Maryland, USA, whereas the other exhibited divergence, indicative of species diversity in the Korean strain, as was evident from the haplotype network analysis. Further validation involved the Ray’s Fluid Thioglycollate Medium (RFTM) assay, which initially yielded inconclusive results, possibly due to low infection intensity. Subsequently, RFTM and 2 M NaOH assays conducted on the isolates in the present study, cultured P. marinus cells in standard DMEM/F12 medium, and a positive P. marinus strain (ATCC 50509), revealed characteristic hypnospores of P. marinus upon Lugol’s iodine staining. These comprehensive investigations underscore the conclusive confirmation of P. marinus in Korean waters and mark a significant milestone in our understanding of the distribution and characteristics of this parasite in previously unreported regions.

Antiviral protection in the Pacific oyster Crassostrea (Magallana) gigas against OsHV-1 infection using UV-inactivated virus

The increase of the frequency and severity of marine diseases affecting farmed marine mollusks are currently threatening the sustainability of this aquaculture sector, with few available prophylactic or therapeutic solutions. Recent advances have shown that the innate immune system of invertebrates can develop memory mechanisms allowing for efficient protection against pathogens. These properties have been called innate immune memory, immune priming or trained immunity. Previous results demonstrated the possibility to elicit antiviral immune priming to protect Pacific oysters against the ostreid herpes virus 1 (OsHV-1), currently plaguing M. gigas production worldwide. Here, we demonstrate that UV-inactivated OsHV-1 is also a potent elicitor of immune priming. Previous exposure to the inactivated virus was able to efficiently protect oysters against OsHV-1, significantly increasing oyster survival. We demonstrate that this exposure blocked viral replication and was able to induce antiviral gene expression potentially involved in controlling the infection. Finally, we show that this phenomenon can persist for at least 3 months, suggesting the induction of innate immune memory mechanisms. This study unravels new ways to train the Pacific oyster immune system that could represent an opportunity to develop new prophylactic strategies to improve health and to sustain the development of marine mollusk aquaculture.

Contribution of the TGFβ signaling pathway to growth of Pacific oyster (Crassostrea gigas)

Growth mediated by the transforming growth factor β (TGFβ) signaling pathway is a key trait in understanding environmental adaptability and species stability. The aim of the present study was to identify the TGFβ signaling pathway members and their expression patterns in different sized Pacific oysters (Crassostrea gigas). Nine oysters were selected for each of two groups: group L (shell height 77.57 ± 7.55 mm, shell length 49.96 ± 4.79 mm) and group B (shell height 43.58 ± 2.74 mm, shell length 33.25 ± 2.78 mm). Using a bioinformatics approach, 37 protein sequences of TGFβ signaling pathway members were classified in C. gigas, including 19 species for this species, nine ligands, five receptors, and five Smads. The mRNA expression levels of TGFβ pathway members in oysters of different sizes showed that both the TGFβ/activin/Nodal and BMP/GDF/MIS subfamilies are involved in the growth regulation of C. gigas. In the first subfamily, there was one possible signaling pathway: INHBA - TGFBR1/BMPR1B/ACVR1 - BMPR2 - Smad3/Smad1, 5, 8. In the second subfamily, there were three possible signaling pathways: BMP2 - BMPR1B - BMPR2 - Smad1, 5, 8; BMP4 - BMPR1B - BMPR2 - Smad1, 5, 8; and BMP7 - ACVR1/BMPR1B - BMPR2 - Smad1, 5, 8. Principle components analysis identified Bmp3, I-Smad, Bmp4, INHBA, and Smad3 as biomarkers of growth in the TGFβ signaling pathway of C. gigas, implicating the TGFβ pathway in the growth of C. gigas.

Effect of Neutral Protease on Freshness Quality of Shucked Pacific Oysters at Different Storage Conditions.

The aim of this study was to investigate the effect of neutral protease treatment on the biochemical properties of various parts of Pacific oysters (Crassostrea gigas) under different storage conditions. The mechanism of quality degradation in the mantle, adductor muscle, gill, and trunk of treated oysters stored at -1.5 °C (superchilling) or 4 °C (refrigeration) for several days using different storage methods was studied. The results showed that the oyster treated with the enzyme exhibited higher glycogen content, flavor nucleotide content, and sensory scores compared to the control group. Superchilling at -1.5 °C was observed to slow the increase in total volatile basic nitrogen (TVB-N), total viable count (TVC), and pH, while maintaining sensory scores better than refrigeration at 4 °C. Both wet superchilling (WS) and dry exposed superchilling (DeS) methods effectively preserved freshness and quality at -1.5 °C. The freshness of the oysters' body trunk changed most significantly. K value, K' value, and AEC value, as the evaluation indexes of oyster freshness, were affected by the storage medium. Therefore, neutral protease enhances the flavor of oysters in a short time, and oysters stored in wet superchilling or dry exposed superchilling conditions have an extended shelf life.

Polymorphisms in the cysteine dioxygenase gene and their association with taurine content in the Pacific oyster Crassostrea gigas

The Pacific oyster Crassostrea gigas is rich in taurine, which is crucial for its adaptation to the fluctuating intertidal environment and presents significant potential in improving taurine nutrition and boosting immunity in humans. Cysteine dioxygenase (CDO) is a key enzyme involved in the initial step of taurine biosynthesis and plays a crucial role in regulating taurine content in the body. In the present study, polymorphisms of CDO gene in C. gigas (CgCDO) and their association with taurine content were evaluated in 198 individuals. A total of 24 single nucleotide polymorphism (SNP) loci were identified in the exonic region of CgCDO gene by direct sequencing. Among these SNPs, c.279G>A and c.287C>A were found to be significantly associated with taurine content, with the GG and AA genotype at the two loci exhibiting enhanced taurine accumulation (p < 0.05). Haplotype analysis revealed that the 279GG/287AA haplotype had the highest taurine content of 29.24 mg/g, while the 279AA/287CC haplotype showed the lowest taurine content of 21.19 mg/g. These results indicated that the SNPs of CgCDO gene could influence the taurine content in C. gigas and have potential applications in the selective breeding of high-taurine varieties.

Temperature effects on plasmalogen profile and quality characteristics in Pacific oyster (Crassostrea gigas) during depuration

The quality of Pacific oyster (Crassostrea gigas) can be affected by many factors during depuration, in which temperature is the major element. In this study, we aim to determine the quality and plasmalogen changes in C. gigas depurated at different temperatures. The quality was significantly affected by temperature, represented by varying survival rate, glycogen content, total antioxidant capacity, alkaline phosphatase activity between control and stressed groups. Targeted MS analysis demonstrated that plasmalogen profile was significantly changed during depuration with PUFA-containing plasmalogen species being most affected by temperature. Proteomics analysis and gene expression assay further verified that plasmalogen metabolism is regulated by temperature, specifically, the plasmalogen synthesis enzyme EPT1 was significantly downregulated by high temperature and four plasmalogen-related genes (GPDH, PEDS, Pex11, and PLD1) were transcriptionally regulated. The positive correlations between the plasmalogen level and quality characteristics suggested plasmalogen could be regarded as a quality indicator of oysters during depuration.

Sudden peak in tetrodotoxin in French oysters during the summer of 2021: Source investigation using microscopy, metabarcoding and droplet digital PCR

Tetrodotoxin (TTX) is a potent neurotoxin causing human intoxications from contaminated seafood worldwide and is of emerging concern in Europe. Shellfish have been shown to contain varying TTX concentrations globally, with concentrations typically higher in Pacific oysters Crassostrea gigas in Europe. Despite many decades of research, the source of TTX remains unknown, with bacterial or algal origins having been suggested. The aim of this study was to identify potential source organisms causing TTX contamination in Pacific oysters in French coastal waters, using three different techniques. Oysters were deployed in cages from April to September 2021 in an estuary where TTX was previously detected. Microscopic analyses of water samples were used to investigate potential microalgal blooms present prior or during the peak in TTX. Differences in the bacterial communities from oyster digestive glands (DG) and remaining flesh were explored using metabarcoding, and lastly, droplet digital PCR assays were developed to investigate the presence of Cephalothrix sp., one European TTX-bearing species in the DG of toxic C. gigas. Oysters analysed by liquid chromatography-tandem mass spectrometry contained quantifiable levels of TTX over a three-week period (24 June–15 July 2021), with concentrations decreasing in the DG from 424 μg/kg for the first detection to 101 μg/kg (equivalent to 74 to 17 μg/kg of total flesh), and trace levels being detected until August 13, 2021. These concentrations are the first report of the European TTX guidance levels being exceeded in French shellfish. Microscopy revealed that some microalgae bloomed during the TTX peak, (e.g., Chaetoceros spp., reaching 40,000 cells/L). Prokaryotic metabarcoding showed increases in abundance of Rubritaleaceae (genus Persicirhabdus) and Neolyngbya, before and during the TTX peak. Both phyla have previously been described as possible TTX-producers and should be investigated further. Droplet digital PCR analyses were negative for the targeted TTX-bearing genus Cephalothrix.

Molecular confirmation and functional study of signal transducer and activator of transcription genes in the Pacific oyster Crassostrea gigas

The JAK (Janus kinase)-STAT (Signal transducer and activator of transcription) is a well-known functional signaling pathway that plays a key role in several important biological activities such as apoptosis, cell proliferation, differentiation, and immunity. However, limited studies have explored the functions of STAT genes in invertebrates. In the present study, the gene sequences of two STAT genes from the Pacific oyster (Crassostrea gigas), termed CgSTAT-Like-1 (CgSTAT-L1) and CgSTAT-Like-2 (CgSTAT-L2), were obtained using polymerase chain reaction (PCR) amplification and cloning. Multiple sequence comparisons revealed that the sequences of crucial domains of these proteins were conserved, and the similarity with the protein sequence of other molluscan STAT is close to 90 %. The phylogenetic analyses indicated that CgSTAT-L1 and CgSTAT-L2 are novel members of the mollusk STAT family. Quantitative real-time PCR results implied that CgSTAT-L1 and CgSTAT-L2 mRNA expression was found in all tissues, and significantly induced after challenge with lipopolysaccharide (LPS), peptidoglycan (PGN), or poly(I:C). After that, dual-luciferase reporter assays denoted that overexpression of CgSTAT-L1 and CgSTAT-L2 significantly activated the NF-κB signaling, and, interestingly, the overexpressed CgSTAT proteins potentiated LPS-induced NF-κB activation. These results contributed a preliminary analysis of the immune-related function of STAT genes in oysters, laying the foundation for deeper understanding of the function of invertebrate STAT genes.

Immunomodulatory effects of a probiotic combination treatment to improve the survival of Pacific oyster (Crassostrea gigas) larvae against infection by Vibrio coralliilyticus.

The culture of Pacific oysters (Crassostrea gigas) is of significant socio-economic importance in the U.S. Pacific Northwest and other temperate regions worldwide, with disease outbreaks acting as significant bottlenecks to the successful production of healthy seed larvae. Therefore, the current study aims to describe the mechanisms of a probiotic combination in improving the survival of C. gigas larvae. Specifically, we investigate changes in C. gigas larval gene expression in response to V. coralliilyticus infection with or without a pre-treatment of a novel probiotic combination. Treatment groups consisted of replicates of Pacific oyster larvae exposed to a) a combination of four probiotic bacteria at a total concentration of 3.0 x 105 CFU/mL at 18 hours post-fertilization (hpf), b) pathogenic V. coralliilyticus RE22 at a concentration of 6.0 x 103 CFU/mL at 48 hpf, and c) the probiotic combination at 18 hpf and V. coralliilyticus RE22 at 48 hpf. RNA was extracted from washed larvae after 72 hpf, and transcriptome sequencing was used to identify significant differentially expressed genes (DEGs) within each treatment. Larvae challenged with V. coralliilyticus showed enhanced expression of genes responsible for inhibiting immune signaling (i.e., TNFAIP3, PSMD10) and inducing apoptosis (i.e., CDIP53). However, when pre-treated with the probiotic combination, these genes were no longer differentially expressed relative to untreated control larvae. Additionally, pre-treatment with the probiotic combination increased expression of immune signaling proteins and immune effectors (i.e., IL-17, MyD88). Apparent immunomodulation in response to probiotic treatment corresponds to an increase in the survival of C. gigas larvae infected with V. coralliilyticus by up to 82%. These results indicate that infection with V. coralliilyticus can suppress the larval immune response while also prompting cell death. Furthermore, the results suggest that the probiotic combination treatment negates the deleterious effects of V. coralliilyticus on larval gene expression while stimulating the expression of genes involved in infection defense mechanisms.

The expression patterns of exosomal miRNAs in the Pacific oyster after high-temperature stress or Vibrio stimulation

The exosomal miRNA plays a crucial role in the intercellular communication response to environmental stress and pathogenic stimulation. In the present study, the expression of exosomal miRNAs in the Pacific oyster Crassostrea gigas after high-temperature stress or Vibrio splendidus stimulation was investigated through high-throughput sequencing. The exosomes were identified to be teardrop-like vesicles with the average size of 81.7 nm by transmission electron microscopy. There were 66 known miRNAs and 33 novel miRNAs identified, of which 10 miRNAs were differentially expressed after both high-temperature stress and Vibrio stimulation compared to the control group. A total of 1868 genes were predicted as the putative targets of miRNAs, of which threonine aspartase 1-like was targeted by the highest number of related miRNAs. The robustness and reliability of miRNA expression from the sRNA sequencing data were verified by employing eight miRNAs for qPCR. GO and KEGG clustering analyses revealed that apoptosis was significantly enriched by the target genes of differentially expressed exosomal miRNAs after high-temperature stress, and autophagy and cytokine activity were significantly enriched after Vibrio stimulation. Energy metabolism was found to be significantly shared in the target gene enrichments after both high-temperature stress and Vibrio stimulation. These findings would improve our understanding of the regulatory mechanisms of exosomal miRNAs in C. gigas after high-temperature stress or Vibrio stimulation.

Effect of air blast freezing and frozen storage on inactivation of Vibrio vulnificus in Pacific oysters (Crassostrea gigas)

Air-blast freezing followed by frozen storage at different temperatures was assessed for inactivation of Vibrio vulnificus in Pacific oysters (Crassostrea gigas). Pacific oysters were allowed to bioaccumulate a cocktail of six strains of V. vulnificus. They were then air-blast frozen at −55 °C followed by frozen storage at −8 °C, −13 °C, −18 °C, −23 °C and −28 °C. V. vulnificus reductions were enumerated at different sampling times from 0 to 360 days. Air-blast freezing and frozen storage inactivated V. vulnificus in Pacific oysters in a two-phase inactivation pattern, with rapid inactivation during the first 3 days after air-blast freezing (short-term effect) followed by slower log-linear declines during frozen storage (long-term effect). The effects of storage temperature on V. vulnificus inactivation (log10 MPN/day) were assessed using a combined model that includes effects from both short- and long-term storage. The US National Shellfish Sanitation Program requires that postharvest processes to reduce Vibrio concentrations achieve ≥3.52 log reductions. At −18 °C this was achieved after an average of 90 days, but 95% confidence was not achieved until 123 days. Lower frozen storage temperatures caused slower declines of V. vulnificus. As well as artificially bioaccumulated V. vulnificus, we modeled inactivation of naturally occurring V. parahaemolyticus and showed that the results of a previous North American study fitted with our data. Their validated treatment for this organism in half-shell oysters can therefore be applied to New Zealand whole-shell oysters. This study will help guide oyster producers/processors aiming to achieve significant inactivation of Vibrio bacteria in their product, by providing a starting point for developing suitable frozen storage time/temperature conditions for Vibrio inactivation.

Potential Function of 3,5-Dihydroxy-4-Methoxybenzyl Alcohol from Pacific Oyster (Crassostrea gigas) in Brain of Old Mice.

3,5-Dihydroxy-4-methoxybenzyl alcohol (DHMBA) is found in oyster extracts in recent years and is reported to have antioxidant activity. Although it has been reported to be protective in various models of oxidative stress, the therapeutic effect of DHMBA on neurological damage caused by agingremains to be demonstrated. The present study investigates the potential functions of DHMBA in brain of oldC57BL/6J mice and aging cell model. Administration of DHMBA improves working memory, reduces anxiety behavior, decreases the expression levels of cell cycle proteins, cycin-dependent kinase inhibitor 1(P21) and peptidase inhibitor 16(P16) and inhibits neuronal loss in old mice. The data obtained from the aging cell model are consistent with those from the old mice. The interaction between DHMBA and Kelch-like ECH-associated protein 1 (Keap1) is predicted by molecular docking assay, and then it is verifiedby co-immunopricipitation (CoIP) that factor red lineage 2-related factor 2 (Nrf2)-Keap1 protein-protein interactionis inhibited by DHMBA. Protein levels of Nrf2 and its target genes, such as glutathione peroxidase 4(GPX4) and heme oxygenase 1 (HO-1), are detected in old mice and aging cell model. This study provides new evidence that explores the antioxidant mechanism of DHMBA and implies a potential role of DHMBA on antiaging in brain.

CgPHB2 involved in the haemocyte mitophagy in response to Vibrio splendidus stimulation in Pacific oyster Crassostrea gigas

Prohibitin2 (PHB2) is recently identified as a novel inner membrane mitophagy receptor to mediate mitophagy. In the present study, the function of CgPHB2 in mediating mitophagy in response to Vibrio splendidus stimulation was investigated in Crassostrea gigas. CgPHB2 protein was mainly distributed in the cytoplasm of three subpopulations of haemocytes. After V. splendidus stimulation, the expressions of CgPHB2 mRNA in haemocytes were up-regulated significantly at 6, 12 and 24 h, and the abundance of CgPHB2 protein was also enhanced at 12–24 h compared to control group. Furthermore, the green signals of CgPHB2 were colocalized respectively with the red signals of mitochondria and CgLC3 in the haemocytes at 12 h after V. splendidus stimulation, and the co-localization value of CgPHB2 and mtphagy Dye was significantly increased. The direct interaction between CgPHB2 and CgLC3 was simulated by molecular docking. In PHB2-inhibitor Fluorizoline-treated oysters, the mRNA expressions of mitophagy-related genes and the ratio of mitophagy were significantly decreased in haemocytes of oysters after V. splendidus stimulation. All the results collectively suggested that CgPHB2 participated in mediating the haemocyte mitophagy in the antibacterial immune response of oysters.

Transcriptome analysis reveals polyploidy-related differential gene expression among diploid, triploid, and tetraploid Pacific oysters (Crassostrea gigas) based on growth-related phenotypes

Tetraploid oysters hold a significant position in the global oyster industry, particularly as core germplasm resources for the production of triploids. Triploids exhibit desirable characteristics such as reduced gonadal development, larger adult size, and superior flesh quality compared with diploids. However, tetraploids often face challenges of slow growth and chromosomal instability during the breeding process, and the mechanisms underlying the formation of these polyploid characteristics remain unclear. This study aimed to elucidate these mechanisms by constructing a polyploid oyster system and comparing the differences in growth-related phenotypes and gene expression among diploids, triploids, and tetraploids. We identified 1404 differentially expressed genes (DEGs) between diploids and triploids, 1814 DEGs between triploids and tetraploids, and 933 DEGs between diploids and tetraploids. Furthermore, we observed distinct gene expression patterns related to innate immunity response and biomineralization between different ploidy levels. Through trend analysis, we clustered genes with similar expression level changes across different ploidy levels and conducted gene ontology (GO) enrichment analysis on genes with different expression profiles. The results indicated that genes associated with the innate immunity response exhibited significantly upregulated expression with increasing ploidy, while genes related to biomineralization showed higher or lower expression levels in triploid oysters. As biomineralization is closely linked to oyster shell growth, this highlights the crucial role of genes associated with biomineralization in regulating the rapid shell growth observed in triploid oysters. Confronted with environmental challenges, tetraploid oysters may display a stronger innate immune response than diploids and triploids. Our study provides valuable resources for investigating the functional aspects of genes related to polyploid phenotype differences and offers new insights into the molecular mechanisms of existing issues in tetraploid oyster breeding.

An LPS-induced TNF-α factor involved in immune response of oyster Crassostrea gigas by regulating haemocytes apoptosis

LPS induced TNF-α Factor (LITAF) is a transcription factor widely involving in activation of Tumor Necrosis Factor (TNF) and other cytokines in the inflammatory response. In the present study, a homologue of LITAF with a conserved LITAF domain was identified from the Pacific oyster Crassostrea gigas. The transcripts of CgLITAF were detected in all examined tissues with highest expression in hepatopancrease. The immunofluorescence assay and Western blot showed that LPS stimulation induced an obvious nucleus translocation of CgLITAF protein in haemocytes. While the mRNA level of CgLITAF changed slightly after LPS stimulation. When the siRNA of CgLITAF was injected to inhibit its expression, the apoptotic level of haemocytes decreased observably after LPS stimulation. Consistently, the transcripts of CgTNF3 and CgTNF4 (LOC105343080, LOC105341146), the apoptotic-related molecules including CgBax, CgCytochrome c, CgCaspase9 and CgCaspase3, were significantly suppressed in the CgLITAF-RNAi oysters. While the mRNA expression level of CgBcl was enhanced significantly in the CgLITAF-RNAi oysters. These results indicated that CgLITAF promoted haemocyte apoptosis by regulating the expression of apoptotic-related factors, suggesting its important role in the immune response of oysters.

Cross-talk and mutual shaping between the immune system and the microbiota during an oyster's life.

The Pacific oyster Crassostrea gigas lives in microbe-rich marine coastal systems subjected to rapid environmental changes. It harbours a diversified and fluctuating microbiota that cohabits with immune cells expressing a diversified immune gene repertoire. In the early stages of oyster development, just after fertilization, the microbiota plays a key role in educating the immune system. Exposure to a rich microbial environment at the larval stage leads to an increase in immune competence throughout the life of the oyster, conferring a better protection against pathogenic infections at later juvenile/adult stages. This beneficial effect, which is intergenerational, is associated with epigenetic remodelling. At juvenile stages, the educated immune system participates in the control of the homeostasis. In particular, the microbiota is fine-tuned by oyster antimicrobial peptides acting through specific and synergistic effects. However, this balance is fragile, as illustrated by the Pacific Oyster Mortality Syndrome, a disease causing mass mortalities in oysters worldwide. In this disease, the weakening of oyster immune defences by OsHV-1 µVar virus induces a dysbiosis leading to fatal sepsis. This review illustrates the continuous interaction between the highly diversified oyster immune system and its dynamic microbiota throughout its life, and the importance of this cross-talk for oyster health. This article is part of the theme issue 'Sculpting the microbiome: how host factors determine and respond to microbial colonization'.