Blood Clam Tegillarca Granosa Research Articles

TAL1-mediated regulation of hemocyte proliferation influences red blood phenotype in the blood clam Tegillarca granosa

Red blood phenotype, indicating the degree of red color in blood or hemolymph, is a significant economic trait in Tegillarca granosa, closely associated with its health condition and nutritive value. However, the molecular regulatory processes that account for the variation in the blood clams' red blood phenotype remain largely unexplored. In our previous studies, weighted gene co-expression network analysis (WGCNA) showed that TAL1 exhibits high intramodular connectivity in red blood-related module. Furthermore, TAL1 serves as a critical transcription factor in the survival and quiescence of hematopoietic stem cells, along with being essential in the terminal maturation processes in vertebrates. Consequently, we investigated the function of the TAL1 in blood clams by conducting a comparative analysis between high total hemocyte count (HTHC: THC ≥ 14 × 104 cells/μL) and low total hemocyte count (LTHC: THC ≤ 2.5 × 104 cells/μL) groups, as well as between negative control and Tg-TAL1 RNA interference groups of blood clams. The results revealed that, compared to LTHC group, there was a significant upregulation of TAL1 mRNA and protein expression levels in the HTHC group, which may be a crucial factor contributing to the alterations observed in THC. RNA interference of the TAL1 in blood clams resulted in a significant decline in THC, causing the blood color to become lighter. EdU staining experiments further confirmed a diminished cell proliferative vitality within T. granosa gills after TAL1 knockdown. Moreover, immunohistochemical analysis and immunofluorescence assays revealed a gradual escalation in TAL1 protein expression and cytoplasmic/nuclear ratio from gill filament chamber endothelial cells to gill filament chamber cells in gills, potentially representing different cell types in the maturation process of erythrocytes in blood clams. It suggesting that the gills might act as one of the hematopoietic tissues in blood clams, with hemocytes potentially originating from the gill filament chamber endothelium of the gills through division and differentiation. This research improves comprehension of the molecular mechanisms behind the variation in T. granosa's red blood phenotype, potentially assisting in the selective breeding of high THC T. granosa populations.

Effect of Extracellular Marennine produced by Haslea Ostrearia on the Blood Clams Tegillarca Granosa

Marennine is a blue-green pigment produced by diatoms of the Haslea genus, one of which is Haslea ostrearia. This marennine pigment is water-soluble and confirmed to contain polyphenols and glycosides. There are two forms of marennine pigment: the intracellular form of marennine (IMn) and the extracellular form of marennine (EMn). Marennine pigments exhibit various biological activities such as antibacterial, antiviral, antioxidant, allelopathic, and inhibiting the growth of other diatoms. With this biological activity, marennine can be used in various fields, one of which is aquaculture.This research aims to determine the level of toxicity and analyze the effect of exposure to the extracellular marennine produced by H. ostrearia on the juvenile blood clam Tegillarca granosa. By using the toxicity test method, Blue water which is supernatant of H. ostrearia (BW) was tested on juvenile of commercially important bivalve species, the blood clams Tegillarca granosa, with three treatments, namely control (0 mg/L), treatment A (0.25 mg/L) and treatment B (0.5 mg/L of BW). The observation parameters of this study were LC50-72 h with one-way ANOVA analysis. The results showed that the BW produced by H. ostrearia increased the survival of juvenile blood clams T. granosa up to 27.7% in treatment B (0.5 mg/L) compared to control within 72 h. The one-way ANOVA analysis revealed that the control and treatment B (0.5 mg/L) were significantly different. This study shows that there is potential to develop the use of marennine in shellfish aquaculture. 

NO production of granulocytes associated with antibacterial immune response in Tegillarca granosa

Nitric oxide (NO) is a signaling molecule and immune effector produced by the nitric oxide synthases (NOS), which involved to various physiological processes of animals. In marine bivalves, hemocytes play important roles in antimicrobial innate immune response. Although hemocyte-derived NO has been detected in several bivalves, the immune function of hemocyte-derived NO is not well understood. Here, we investigated the antibacterial response of hemocyte-derived NO in the blood clam Tegillarca granosa. Two types of hemocytes including erythrocytes and granulocytes were isolated by Percoll density gradient centrifugation, their NO production and TgNOS expression level were analyzed. The results showed that NO was mainly produced in granulocytes and almost no detected in erythrocytes. The granulocytes showed significantly higher NO level and TgNOS expression level than the erythrocytes. And the TgNOS expression level was significantly increased in granulocytes after Vibro parahemolyticus challenge. In addition, the NO donor sodium nitroprusside (SNP) significantly increased the NO production of hemocytes to kill pathogenic bacteria. In summary, the results revealed that granulocytes-derived NO play vital roles in the antimicrobial immune response of the blood clam.

Isolation and functional identification of immune cells in hemolymph of blood clams Tegillarca granosa

Blood clam Tegillarca granosa is a type of economically cultivated bivalve mollusk with red blood, and it primarily relies on hemocytes in its hemolymph for immune defense. However, there are currently no reports on the isolation and identification of immune cells in T. granosa, which hinders our understanding of their immune defense. In this study, we employed single-cell transcriptome sequencing (scRNA-seq) to visualize the molecular profile of hemocytes in T. granosa. Based on differential expression of immune genes and hemoglobin genes, hemocytes can be molecularly classified into immune cells and erythrocytes. In addition, we separated immune cells using density gradient centrifugation and demonstrated their stronger phagocytic capacity compared to erythrocytes, as well as higher levels of ROS and NO. In summary, our experiments involved the isolation and functional identification of immune cells in hemolymph of T. granosa. This study will provide valuable insights into the innate immune system of red-blood mollusks and further deepen the immunological research of mollusks.

CBFA2T3 affects red blood phenotype in the blood clam Tegillarca granosa by regulating hemocyte proliferation

The red blood phenotype of blood clam Tegillarca granosa, is tightly linked to its health conditions and nutritional benefits, making it an economically important trait. Despite its significance, the molecular regulatory mechanisms responsible for red blood phenotype variation in blood clams still poorly understood. Our previous study shows that hemocyte proliferation may affect hemoglobin concentration (HGC) and cause the red blood phenotype variation in T. granosa. Thus, we defined total hemocyte counts (THC) and HGC as red blood-related traits. In this research, we discovered that blood clams with higher THC exhibited elevated HGC and a darker blood color. To gain insights into the genes associated with THC in T. granosa, we conducted RNA-Seq analysis. Our transcriptome analysis identified 682 genes exclusively highly expressed in high THC clams (HTHC:THC ≥ 15.5 × 104 cells/mL), and 709 genes highly expressed in low THC clams (LTHC:THC ≤ 6.5 × 104 cells/mL). Furthermore, the weighted gene co-expression network analysis (WGCNA) revealed that the grey60 module exhibited significant positive correlations with THC and HGC, implying an association of this module with the red-blood phenotype. Subsequently, the red blood-related module exhibited a significant positive correlation with gill tissue, providing an alternative perspective to support the hypothesis that gills serve as the primary hematopoietic organ in T. granosa. By employing differential expression analysis and WGCNA, CBFA2T3, TAL1, and FLI1 are potentially key genes involved in the regulation of hemocyte proliferation and differentiation. RNA interference of CBFA2T3 gene led to a remarkable decrease in HGC and THC in T. granosa, resulting in a lighter blood color. EdU staining further confirmed the decrease of cell proliferation activity in the gills of T. granosa after knockdown of CBFA2T3. This study enhances our apprehending of the molecular mechanisms in red blood phenotype variation in T. granosa, and potential contributes to the selection breeding of high THC blood clams.

Genome-wide identification and characteristic analysis of ETS gene family in blood clam Tegillarca granosa

BackgroundETS transcription factors, known as the E26 transformation-specific factors, assume a critical role in the regulation of various vital biological processes in animals, including cell differentiation, the cell cycle, and cell apoptosis. However, their characterization in mollusks is currently lacking.ResultsThe current study focused on a comprehensive analysis of the ETS genes in blood clam Tegillarca granosa and other mollusk genomes. Our phylogenetic analysis revealed the absence of the SPI and ETV subfamilies in mollusks compared to humans. Additionally, several ETS genes in mollusks were found to lack the PNT domain, potentially resulting in a diminished ability of ETS proteins to bind target genes. Interestingly, the bivalve ETS1 genes exhibited significantly high expression levels during the multicellular proliferation stage and in gill tissues. Furthermore, qRT-PCR results showed that Tg-ETS-14 (ETS1) is upregulated in the high total hemocyte counts (THC) population of T. granosa, suggesting it plays a significant role in stimulating hemocyte proliferation.ConclusionOur study significantly contributes to the comprehension of the evolutionary aspects concerning the ETS gene family, while also providing valuable insights into its role in fostering hemocyte proliferation across mollusks.

A genome-wide association study to identify the genes associated with growth-related traits in Tegillarca granosa

The blood clam Tegillarca granosa is a commercially important bivalve species that is extensively cultivated worldwide. However, understanding of the molecular mechanisms underlying the growth of this species remains limited. In this study, we conducted genome-wide association studies on the growth traits of T. granosa, including shell and body weight characteristics, using a population of 300 T. granosa individuals. Correlation analysis revealed strong positive associations among the growth traits, with the exception of dressing percentage, suggesting that selective breeding for any of these traits can contribute to overall growth enhancement in T. granosa. A total of 39 single nucleotide polymorphism (SNP) markers associated with the seven growth traits were identified. Further investigation led to the identification of 27 candidate genes located within a 100 kb region upstream or downstream of these SNPs, which are potentially involved in T. granosa growth. Enrichment analysis of the candidate genes highlighted the biological process of transmembrane transport and the pathways related to immune system function, protein family metabolism, and JAK-STAT signaling as potential contributors to T. granosa growth. Furthermore, we discovered a haploblock associated with shell width, and validated the differential expression of four genes (Vha26, Ptprt, Stat5b, and Kmt2c) between fast and slow growing groups, which may play important roles in the growth of T. granosa. Overall, our study presents valuable genetic resources and serves as a reference for future marker-assisted selection strategies for enhancing the growth performance of T. granosa.

ATG13 is involved in immune response of pathogen invasion in blood clam Tegillarca granosa.

Mammalian autophagy-related gene 13 (ATG13) is a vital component of the ATG1 autophagy initiation complex which plays an essential role in autophagy. However, the molecular function of ATG13 in pathogen defense in invertebrates is still poorly understood. In this study, the full-length cDNA sequence of blood clam Tegillarca granosa ATG13 (TgATG13) was obtained, which was 1,918 bp in length, including 283 bp 5' UTR, 252 bp 3' UTR and 1,383 bp open reading frame (ORF) encoding 460 amino acids. Phylogenetic analysis revealed that TgATG13 had the closest relationship with that of Crassostrea Virginica. Quantitative real-time PCR results showed that the transcript of TgATG13 was universally expressed in various tissues of blood clam, with the highest expression level in hemocytes. The expression level of TgATG13 was robustly increased after exposure of both Vibrio alginolyticus and LPS. Fluorescence confocal microscopy further showed that TgATG13 promoted the production of autophagosome. In summary, our study demonstrated that TgATG13 was involved in the immune regulation of blood clam during pathogen invasion, deepening our understanding of the innate immune mechanism of blood clam.

Identificaiton and characterization of a novel hemoglobin gene (Tgr-HbIII) from blood clam Tegillarca granosa

Hemoglobin (Hb) is the major protein component of red blood cells (hemocytes) of the blood clam Tegillarca granosa. Three T. granosa hemoglobin genes have been mentioned in the literature, designated Tgr-HbI, Tgr-HbIIA and Tgr-HbIIB. Previously, our group identified another novel gene, Tgr-HbIII, in the Hb cluster of the chromosome-level genome but the issue of whether this Hb gene expresses functional protein remains unclear. In the current study, phylogenetic analysis revealed that Tgr-HbIII resembles an ancient Hb gene. Sequence alignment and three-dimensional structural modeling results showed that Tgr-HbIII does not bind heme due to the completely different structure at amino acid position 96–100 and replacement of the N100 residue in known Tgr-Hbs with Q100, what causes loss of a single hydrogen bond linking heme with the globin fold. Interface prediction data suggest that Tgr-HbIII forms a homodimer (ΔG = −5.6 kcal/mol) with a similar conformation to the Tgr-HbI homodimer (ΔG = −3.5 kcal/mol). In adult T. granosa, mRNA expression of Tgr-HbIII was lower than that of Tgr-HbIIA and Tgr-HbIIB (up to 100 × ), but comparable to that of Tgr-HbI. Notably, protein expression of Tgr-HbIII was extremely low. Single-cell RNA sequencing analysis of Hb expression showed that all adult hemocytes expressed Tgr-HbI, Tgr-HbIIA and Tgr-HbIIB, while only 43 % (3872 of 8978) expressed Tgr-HbIII. Based on the collective data, we speculate that Tgr-HbIII carried oxygen prior to mutation of N100 to Q100 and subsequently evolved into a known functional remnant of Hb with an adequate mRNA/low protein expression profile. The current study provides a foundation for further research on the origin, evolution and function of molluscan Hbs.

Comparative metabarcoding analysis of phytoplankton community composition and diversity in aquaculture water and the stomach contents of Tegillarca granosa during months of growth

Filter-feeder bivalves and phytoplankton are interdependent. Their interaction plays important role in estuarine and coastal ecosystem. The correlation between bivalve feeding and phytoplankton is highly species specificity and environment dependent. In the background of miniature and nondiatom trend of phytoplankton in coastal seawaters, how bivalve respond and how the response play roles in the phytoplankton community are poorly known. In the present study, by applying DNA metabarcoding approach based on plastid 23S rDNA, this question was addressed by comparing the phytoplankton composition in the seston and the stomach content of blood clam Tegillarca granosa sampled during the growth period from March to November 2020 in an experimental farm on tidal flat in Xiangshan Bay, East China Sea. The result showed that, a total of seven phyla, 55 genera and 73 species of phytoplankton were identified for all samples. Chlorophyta, Bacillariophyta, and Cyanobacteria were found to be three dominant phyla both in the stomach contents and seston. High diversity of pico-sized phytoplankton, which was easy overlooked by microscopy, was revealed both in seston and stomach contents. This result indicated that the clam was able to feed on the pico-sized algae. At the genus level, the most abundant genera were the pico-sized green alga Ostreococcus (6.12 %–67.88 %) in seston and Picochlorum (4.07 %–35.33 %) in the stomach contents. In addition, microalgae of high nutritional value showed trend of higher proportion in stomach contents than that in seston, especially in July and September when significant growth of T. granosa was observed during this period (the body size increased 155 %). Biodiversity of phytoplankton in the seston was totally higher than that in stomach content, however, the changes among the months showed respective trend. Especially in July when the biodiversity was the lowest in seston, that in the stomach content showed the highest. The results indicated that blood clam farming might influence the phytoplankton composition, including those of pico-sized level, although the particular species in seston were mainly correlating with the dominant environmental factors such as temperature, salinity, pH respectively. These results extend the understanding of roles that bivalve aquaculture may play in the changing of coastal phytoplankton community.

Immune responses of hemocytes in the blood clam Tegillarca granosa in response to in vivo Vibrio harveyi infection

Aquaculture of the blood clam Tegillarca granosa accounts for approximately 50% of Arcidae (ark shell) production in China. Vibrio infection severely threatens the sustainability of the clam aquaculture industry. Exposure to Vibrio induces an immune response in blood clams. However, the underlying mechanism remains poorly understood. In this study, immune responses of hemocytes in blood clams were detected after Vibrio infection; the immersion method was used in vivo to mimic the clam's natural infection process. After 24 h of exposure to Vibrio infection, the Vibrio load in hemolymph fluid in both the treatment Ⅰ (25,033.33 ± 19,563.11 CFU/mL) and treatment Ⅱ (122,163.33 ± 194,409.49 CFU/mL) groups were significantly higher, than that in the control group (13.67 ± 37.73 CFU/mL) (P < 0.05). Correspondingly, the production of intracellular reactive oxygen species was approximately 1.40 (treatment Ⅰ) and 2.12 (treatment Ⅱ) fold higher than that in the control group (P < 0.05), and the induced DNA damage showed a similar trend (P < 0.05). Vibrio infection also significantly increased lysozyme content, adenosine triphosphate content, and peroxidase isozyme activity, in both the serum and hemocyte lysates (P < 0.05). The expression of immune-associated genes (ABCA3, c-Myc, Caspase 3, and HSP70) was upregulated under infection conditions. The phagocytic activity was approximately 1.99 (treatment Ⅰ) and 2.57 (treatment Ⅱ) fold that in control clams (P < 0.05). In addition, the total hemocyte count and red granulocyte percentage both significantly decreased by approximately 75–90% after Vibrio infection. These results provided novel insights into the mechanism of hemocyte immunity in T. granosa against Vibrio infection, which may aid in the future prevention and control of Vibrio infection in vivo.

Genome-wide identification and immune response analysis of serine protease inhibitor genes in the blood clam Tegillarca granosa

Serine protease inhibitors (SPIs) are the main regulators of serine protease activities. In this study, we present a genome-wide identification of SPI genes in T. granosa（TgSPI genes）and their expression characteristics in respond to Vibrio stress. A total of 102 TgSPI genes belonging to eight families, including Serpin, TIL (trypsin inhibitor like cysteine rich domain), Kunitz, Kazal, I84, Pacifastin, WAP (whey acidic protein) and A2M (Alpha-2-macroglobulin) were identified, while no genes belonging to Bowman-Birk, amfpi and Antistasin families were identified. The Kazal family has the most TgSPI genes with 38, and 11 TgSPI genes belong to the mollusc-specific I84 family. The TgSPI genes were found to be randomly distributed on 17 chromosomes with 12 tandem duplicate gene pairs. Expression profiles showed that most TgSPI genes were mainly expressed in immune-related tissues such as hepatopancreas, gill and mantle. In the hepatopancreas, most of TgSPI genes were sensitive to Vibrio stress, 28 and 29 TgSPI genes were up-regulated and down-regulated, respectively. Some up-regulated genes with signal peptides, such as the TgSPIs of I84 family, may act as a mechanism to directly prevent Vibrio from invasion. Six Kazal-type TgSPIs (TgSPI29, 45, 49, 50, 51 and 52) were intracellular proteins and their expression was down-regulated in hemocytes after Vibrio stress. This may have boosted protease activity in hemocytes to the point that more hemoglobin derived peptides were produced and secreted into the hemolymph to exert their anti-Vibrio effects. These findings may provide valuable information for further clarifying the roles of SPIs in the immune defense and will benefit future exploration of the immune function of SPIs in molluscs.

Acute sulfide exposure induces hemocyte toxicity and microbiota dysbiosis in blood clam Tegillarca granosa.

Sulfide are widely accumulated in aquatic environments under anaerobic conditions, which cause health problems of aquatic animals, yet their toxic effects to benthic bivalves are not well understood. We investigated the effects of sulfide on innate immunity of the blood clam Tegillarca granosa. Immunity-related indicators and hemolymph microbiota were investigated in the clams exposed to sulfide (via 10, 100 and 1000 μmol/L of Na2S) over a 7-day period. The results showed that cellular immune responses in T. granosa were affected by exposure to high sulfide concentration (1000μmol/L Na2S), as indicated by total counts of hemocytes (THC), cell viability, ROS levels and phagocytic activities, suggesting that sulfide stress induces T. granosa more vulnerable to pathogen challenges. In addition, the Na2S-induced stress also reshaped the hemolymph microbial community structure of T. granosa that some original genera decreased, such as Lactobacillus, Desulfovibrio and Akkermansia; some genera increased, such as Vibrio and Pseudoalteromonas in sulfide stress group. Sulfide exposure promoted the proliferation of opportunistic pathogen and reduced the diversity of microbial community in the hemolymph of T. granosa. In summary, sulfide stress had marked hemocytotoxicity, reduced immune-cell activity and increased bacterial infections in the blood clam.

Molecular cloning, characterization, and tissue distribution of c-Myc from blood clam Tegillarca granosa and its role in cadmium-induced stress response

Cadmium (Cd) pollution threatens the cultivation of the blood clam Tegillarca granosa (T. granosa) in coastal regions of the East China Sea. The molecular mechanisms regulating Cd stress response and detoxification in blood clams are largely unclear. In the present study, the full-length T. granosa c-Myc (Tgc-Myc) cDNA was cloned for the first time. The 3063-bp cDNA consisted of a 129-bp 5′ untranslated region (UTR), a 1746-bp 3′ UTR, and a 1188-bp open reading frame encoding a predicted protein of 395 amino acid residues. The predicted protein had a calculated molecular weight of 44.9 kDa and an estimated isoelectric point of 6.82. The predicted protein contained an N-terminal transactivation domain and a C-terminal basic helix-loop-helix leucine zipper domain, which are conserved functional domains of c-Myc proteins. Tgc-Myc showed broad tissue distribution in blood clams, with the highest expression detected in the gill and hepatopancreas. Exposure to Cd, a major heavy metal pollutant in coastal regions of the East China Sea, induced Tgc-Myc expression in gill tissues. Tgc-Myc knockdown led to reduced expression of a variety of stress response/detoxification genes in blood clams cultivated in Cd-contaminated seawater. Tgc-Myc knockdown also led to decreased expression of IGF1R, a proto-oncogene that promotes cell proliferation. These findings indicated that Tgc-Myc regulates Cd-induced stress response and detoxification in blood clams. The upregulation of Tgc-Myc may serve as an approach to generate strains with an enhanced detoxification response and consequently a low heavy metal buildup.

Bisphenol A and microplastics weaken the antimicrobial ability of blood clams by disrupting humoral immune responses and suppressing hemocyte chemotactic activity

Robust antimicrobial capability is crucial for marine organisms survival in complex ocean environments. Although the detrimental impacts of emergent pollutants on cellular immune response of marine bivalve mollusks were increasingly documented, the effects of bisphenol A (BPA) and microplastics (MPs) on humoral immune response and hemocyte chemotactic activity remain unclear. Therefore, in this study, the toxicities of BPA and MPs, alone or in combination, to the antimicrobial ability, humoral immune response, and hemocyte chemotactic activity were investigated in the blood clam Tegillarca granosa. Our data demonstrated that exposure of blood clams to BPA, MPs, and BPA-MPs for 2 weeks lead to significant reductions in their survival rates upon pathogenic bacterial challenge, indicating evident impairment of antimicrobial ability. Compared to control, the plasma of pollutant-incubated blood clams exhibited significantly less antimicrobial activity against the growth of V. harveyi, suggesting significant reduction in humoral immune effectors including defensin, lysozyme (LZM), and lectin. Moreover, hemocytes migration across the polycarbonate membrane to the serum containing chamber was markedly arrested by 2-week exposure to BPA, MPs, and BPA-MPs, suggesting a hampered chemotactic activity. In addition, the intracellular contents of ROS and protein carbonyl in hemocytes were markedly induced whereas the expression levels of key genes from the MAPK and actin cytoskeleton regulation pathways were significantly suppressed upon exposure. In this study, it was also found that BPA-MP coexposure was significantly more toxic than single exposures. In summary, our findings revealed that exposure to the pollutants tested possibly impair the antimicrobial ability of blood clam through (1) reducing the inhibitory effect of plasma on bacterial growth, the contents of humoral immune effectors, and the chemotactic activity of hemocytes, (2) interrupting IL-17 activation of MAPK signal pathway, (3) inducing intracellular ROS, elevating protein carbonylation levels, and disrupting actin cytoskeleton regulation in hemocytes.

Characterization of a novel activating protein-1 (AP-1) gene and the association of its single nucleotide polymorphisms with vibrio resistance in Tegillarca granosa

The blood clam Tegillarca granosa is a commercial marine bivalve of economic value, accounting for approximately 50% of clam production in China. In recent years, the yield of blood clams has been threatened by bacterial infections caused by marine Vibrio species that thrive under a rising sea temperature. The transcription factor activating protein-1 (AP-1) is emerging as an important player in the innate immunity of marine bivalves against viral or bacterial infections. In this study, the full-length cDNA of a novel T. granosa AP-1 (TgAP-1) was cloned for the first time. The 1591-bp cDNA encoded a protein of 292 amino acid residues with a calculated molecular weight of 32.8 kDa. The TgAP-1 protein contained an N-terminal Jun domain and a C-terminal basic region leucine zipper domain typically found in Jun proteins (a subfamily of AP-1 proteins). TgAP-1 was ubiquitously expressed in T. granosa, with the highest expression detected in the gill and foot, followed by the mantle, hemolymph, and hepatopancreas. Exposure to Vibrio harveyi induced TgAP-1 expression in gill tissues and the expression levels of TgAP-1 of resistant blood clams were always lower than that of control population whether Vibro infection or not. A total of 18 single nucleotide polymorphisms (SNPs) of TgAP-1 were detected in T. granosa. SNP-typing and haplotyping of resistant and susceptible populations revealed that six SNPs (AG type of TgSNP-1, GA type of TgSNP-2, TG type of TgSNP-4, CT type of TgSNP-7, AG type of TgSNP-11, and GA type of TgSNP-12) and four haplotypes (fHap2, fHap3, fHap6, and fHap7) were significantly associated with V. harveyi resistance. Risk assessment showed that fHap2 (CG) and fHap7 (GA) were associated with an increased resistance, while fHap3 (CT) and fHap6 (AG) were associated with an increased susceptibility. The results from this study supported a potential role of TgAp-1 in the anti-Vibro immunity of T. granosa. The discovery of the genetic molecular markers and haplotypes related to Vibrio resistance can provide guidance for selective breeding of T. granosa in the future.

Hypoxia-mediated immunotoxicity in the blood clam Tegillarca granosa

In marine ecosystems, dissolved oxygen (DO) is essential for maintaining intracellular energy balance during aerobic metabolism. Bivalve mollusks are frequently exposed to hypoxia environments due to tides, temperature changes, and anthropogenic activities. The blood clam, Tegillarca granosa, mainly inhabits intertidal mudflats and is more susceptible to low oxygen events. In this study, we investigated the effect of hypoxia on immune responses in clams, and showed that hypoxia exposure reduced total hemocyte counts (THC), hemoglobin concentrations, and intracellular reactive oxygen species (ROS) levels. Also, phagocytic and cell activities of hemocyte were significantly inhibited. Furthermore, immune-related gene expression was also down-regulated. In conclusion, hypoxia greatly affected immune functions in blood clams, and our research provided the foundation for further mechanistic studies on hypoxia tolerance in clams.

Cutting performance of bionic cutting tools based on surface microstructures of blood clam Tegillarca granosa in dry cutting of CFRP

Cutting performance of bionic cutting tools based on surface microstructures of blood clam Tegillarca granosa in dry cutting of CFRP

Molecular Characterization and Expression of the LAP3 Gene and Its Association with Growth Traits in the Blood Clam Tegillarca granosa

Leucine aminopeptidase 3 (LAP3) is a metallopeptidase that cleaves N-terminal residues and is involved in protein maturation and degradation. In this study, we characterized the leucine aminopeptidase 3 (LAP3) gene from Tegillarca granosa (Tg-LAP3 for short), which appeared to consist of 15,731 nucleotides encoding 530 amino acids. We identified 12 introns and 13 exons in the Tg-LAP3 gene, suggesting a highly conserved genomic structure. The proximal promoter sequence consists of 1922 bps with a typical TATA box structure, which is the general structural characteristic of core promoters in eukaryotes. We found two functional domains in the Tg-LAP3 protein, including an N-terminal domain (41–174aa) and a peptidase_M17 catalytic domain (209–522aa). Multiple alignment showed that Tg-LAP3 shares 73.4% identity with LAP3 of Mizuhopecten yessoensis and 55.2–70.7% identity with LAP3 of other species. Quantitative analysis of Tg-LAP3 in embryos/larvae and adult tissues indicated that the highest expression occurred in eyebot larva, with limited expression in other stages; among tissues, the highest expression was found in the liver (p &lt; 0.05). Association analysis found that three single-nucleotide polymorphisms (SNPs) (g.-488A &gt; G, g.-1123C &gt; T, and g.-1304C &gt; A) in the proximal promoter were successfully typed, but there was no significant difference in growth traits (body weight, shell length, shell width, and shell height) among these genotypes. The results of our study demonstrate the functional roles of the Tg-LAP3 gene and provide valuable information for molecular marker-assisted selection (MAS) of the blood clam.

Acute benzo[a]pyrene exposure induced oxidative stress, neurotoxicity and epigenetic change in blood clam Tegillarca granosa

The blood clam (Tegillarca granosa) is being developed into a model bivalve mollusc for assessing and monitoring marine pollution on the offshore seabed. However, the information on the response of blood clam to PAHs, an organic pollutant usually deposited in submarine sediment, remains limited. Herein, we employed multiple biomarkers, including histological changes, oxidative stress, neurotoxicity and global DNA methylation, to investigate the effects of 10 and 100 μg/L Bap exposure on the blood clams under laboratory conditions, as well as the potential mechanisms. Acute Bap exposure can induce significant morphological abnormalities in gills as shown through hematoxylin–eosin (H.E) staining, providing an intuitive understanding on the effects of Bap on the structural organization of the blood clams. Meanwhile, the oxidative stress was significantly elevated as manifested by the increase of antioxidants activities of superoxide dismutase (SOD), catalase (CAT), peroxidase (POD) and glutathione-s-transferase (GST), lipid peroxidation (LPO) level and 8-hydroxy-2′-deoxyguanosine (8-OHdG) content. The neurotoxicity was also strengthened by Bap toxicity manifested as inhibited acetylcholinesterase (AChE) and choline acetyltransferase (ChAT) activities. In addition, the global DNA methylation level was investigated, and a significant DNA hypomethylation was observed in Bap exposed the blood clam. The correlation analysis showed that the global DNA methylation was negatively correlated with antioxidants (SOD, CAT and POD) activities, but positively correlated choline enzymes (AChE and ChAT) activities. These results collectively suggested that acute Bap exposure can cause damage in gills structures in the blood clam possibly by generating oxidative stress and neurotoxicity, and the global DNA methylation was inhibited to increase the transcriptional expression level of antioxidants genes and consequently elevate antioxidants activities against Bap toxicity. These results are hoped to shed some new light on the study of ecotoxicology effect of PAHs on marine bivalves.