The Caspase Homologues in Scallop Chlamys farreri and Their Expression Responses to Toxic Dinoflagellates Exposure.

The cysteine aspartic acid-specific protease (caspase) family is distributed across vertebrates and invertebrates, and its members are involved in apoptosis and response to cellular stress. The Zhikong scallop (Chlamys farreri) is a bivalve mollusc that is well adapted to complex marine environments, yet the diversity of caspase homologues and their expression patterns in the Zhikong scallop remain largely unknown. Here, we identified 30 caspase homologues in the genome of the Zhikong scallop and analysed their expression dynamics during all developmental stages and following exposure to paralytic shellfish toxins (PSTs). The 30 caspase homologues were classified as initiators (caspases-2/9 and caspases-8/10) or executioners (caspases-3/6/7 and caspases-3/6/7-like) and displayed increased copy numbers compared to those in vertebrates. Almost all of the caspase-2/9 genes were highly expressed throughout all developmental stages from zygote to juvenile, and their expression in the digestive gland and kidney was slightly influenced by PSTs. The caspase-8/10 genes were highly expressed in the digestive gland and kidney, while PSTs inhibited their expression in these two organs. After exposure to different Alexandrium PST-producing algae (AM-1 and ACDH), the number of significantly up-regulated caspase homologues in the digestive gland increased with the toxicity level of PST derivatives, which might be due to the higher toxicity of GTXs produced by AM-1 compared to the N-sulphocarbamoyl analogues produced by ACDH. However, the effect of these two PST-producing algae strains on caspase expression in the kidney seemed to be stronger, possibly because the PST derivatives were transformed into highly toxic compounds in scallop kidney, and suggested an organ-dependent response to PSTs. These results indicate the dedicated control of caspase gene expression and highlight their contribution to PSTs in C. farreri. This work provides a further understanding of the role of caspase homologues in the Zhikong scallop and can guide future studies focussing on the role of caspases and their interactions with PSTs.

Identification, characterization and expression analyses of cholinesterases genes in Yesso scallop (Patinopecten yessoensis) reveal molecular function allocation in responses to ocean acidification.

Cholinesterases are key enzymes in central and peripheral cholinergic nerve system functioning on nerve impulse transmission in animals. Though cholinesterases have been identified in most vertebrates, the knowledge about the variable numbers and multiple functions of the genes is still quite meagre in invertebrates, especially in scallops. In this study, the complete cholinesterase (ChE) family members have been systematically characterized in Yesso scallop (Patinopecten yessoensis) via whole-genome scanning through in silico analysis. Ten ChE family members in the genome of Yesso scallop (designated PyChEs) were identified and potentially acted to be the largest number of ChE in the reported species to date. Phylogenetic and protein structural analyses were performed to determine the identities and evolutionary relationships of these genes. The expression profiles of PyChEs were determined in all developmental stages, in healthy adult tissues, and in mantles under low pH stress (pH 6.5 and 7.5). Spatiotemporal expression suggested the ubiquitous functional roles of PyChEs in all stages of development, as well as general and tissue-specific functions in scallop tissues. Regulation expressions revealed diverse up- and down-regulated expression patterns at most time points, suggesting different functional specialization of gene superfamily members in response to ocean acidification (OA). Evidences in gene number, phylogenetic relationships and expression patterns of PyChEs revealed that functional innovations and differentiations after gene duplication may result in altered functional constraints among PyChEs gene clusters. Collectively, our results provide the potential clues that the selection pressures coming from the environment were the potential inducement leading to function allocation of ChE family members in scallop.

Identification and characterization of cellouronate (ß-1,4-linked polyglucuronic acid) lyase from the scallop Mizuhopecten yessoensis.

The semisynthetic polysaccharide cellouronate is a ß-1,4-linked polyglucuronic acid prepared from regenerated cellulose by chemical oxidation. Here, we isolated a novel enzyme, MyAly, as a cellouronate lyase from a scallop Mizuhopecten yessoensis. Its optimum temperature, pH, and NaCl concentration for cellouronate degradation were determined to be 30 °C, 6.9, and 200-500 mM, respectively. MyAly endolytically degraded cellouronate into unsaturated di-, tri-, and tetrasaccharides with kcat of 31.1 s-1. MyAly also showed an alginate-degradation activity with a kcat value of 0.58 s-1. However, there was no significant difference in Km values between cellouronate and alginate. MyAly consisted of 280 amino acids and shared 36.5-44.1 % identity with known marine gastropod alginate lyases belonging to the polysaccharide lyase family 14. This is the first study to identify and characterize a cellouronate-degrading lyase from a marine organism, providing a better understanding of the biodegradability of the industrially important polysaccharide, cellouronate, in marine environments.

Biochemical Characterization of a Novel Endo-1,3-ß-Glucanase from the Scallop Chlamys farreri.

Endo-1,3-ß-glucanases derived from marine mollusks have attracted much attention in recent years because of their unique transglycosylation activity. In this study, a novel endo-1,3-ß-glucanase from the scallop Chlamys farreri, named Lcf, was biochemically characterized. Unlike in earlier studies on marine mollusk endo-1,3-ß-glucanases, Lcf was expressed in vitro first. Enzymatic analysis demonstrated that Lcf preferred to hydrolyze laminarihexaose than to hydrolyze laminarin. Furthermore, Lcf was capable of catalyzing transglycosylation reactions with different kinds of glycosyl acceptors. More interestingly, the transglycosylation specificity of Lcf was different from that of other marine mollusk endo-1,3-ß-glucanases, although they share a high sequence identity. This study enhanced our understanding of the diverse enzymatic specificities of marine mollusk endo-1,3-ß-glucanases, which facilitated development of a unique endo-1,3-ß-glucanase tool in the synthesis of novel glycosides.

Differential responses of a pi-class glutathione S-transferase (CnGSTp) expression and antioxidant status between golden and brown noble scallops under pathogenic stress.

Glutathione S-transferases (GSTs) play important roles in immunity by protecting organisms against the damage of reactive oxygen species (ROS). In this study, a pi-class GST cDNA sequence was first cloned from noble scallop Chlamys nobilis (named CnGSTp). The full length cDNA of CnGSTp was 922 bp, encoding a cytosolic protein of 202 amino acids residues, with predicted molecular masses of 23.1 kDa. Then an acute Vibrio Parahaemolyticus challenge experiment was conducted by using the Golden and Brown noble scallops with different total carotenoids content (TCC), and CnGSTp expression level, TCC and ROS level was separately determined. The results showed that ROS and CnGSTp expression levels were significantly up-regulate under Vibrio Parahaemolyticus challenge than the control group (P < 0.05). The Golden scallops showed significantly higher CnGSTp expression level and lower ROS level in hemocytes than the Brown ones (P < 0.05). Moreover, there is a significantly positive correlation between TCC and ROS in the Golden scallops. The present results revealed that CnGSTp plays important roles in immune response and carotenoids play assistant roles in antioxidant defense system under pathogenic stress in the noble scallop.

The mechanism of Mitogen-Activated Protein Kinases to mediate apoptosis and immunotoxicity induced by Benzo[a]pyrene on hemocytes of scallop Chlamys farreri in vitro.

Benzo [a]pyrene (B [a]P) has received widespread attention for serious pollution in the sea, which may reduce immunity and lead to the outbreak of disease in bivalves. However, the mechanism of immunotoxicity induced by B [a]P in bivalves was still unclear. Previous studies have found that Mitogen-Activated Protein Kinases (MAPKs) including three classic pathways (ERK, p38 and JNK) play an important role in mediating this process. Thus, in order to explore the mechanism of immunotoxicity induced by B [a]P in scallop Chlamys farreri, hemocytes were treated with PD98059 (ERK inhibitor), SB203580 (p38 inhibitor) and SP600125 (JNK inhibitor) for 1 h and then incubation with B [a]P for 24 h at 1 µg/mL. Indexes including oxidative damage, apoptotic rate, and immune indicators were detected in the present study. The results showed that the increase of Reactive Oxygen Species (ROS) and DNA damage induced by B [a]P was inhibited with PD98059 and SB203580. Besides, lysosomal membrane stability (LMS) damage was promoted by PD98059, while it was opposite when treated with SB203580. Moreover, the ascended apoptosis rate induced by B [a]P was increased significantly after treatment with PD98059, but it was remarkably attenuated by SB203580 and SP600125. However, the opposite pattern was showed in phagocytosis compared with apoptosis rate in all of three inhibitors. In addition, antibacterial activity and bacteriolytic activity were enhanced by SB203580 while inhibited by PD98059. Therefore, these results showed that MAPKs directly or indirectly mediate the decrease of oxidative damage, apoptosis and immune defense ability of C. farreri hemocytes, which suggesting ERK/p38/JNK pathways have different functions in the apoptosis and immunity of C. farreri hemocytes after B [a]P exposure. In conclusion, this study intended to enrich the theoretical basis for immunotoxicology of bivalves exposed to pollutants.

Structure and functional analysis reveal an important regulated role of arginine kinase in Patinopecten yessoensis under low pH stress.

Arginine kinase (AK), an important member of the phosphokinase family, is involved in temporal and spatial adenosine triphosphate (ATP) buffering systems. AK plays an important role in physiological function and metabolic regulations, in particular tissues with high and fluctuating energy demands. In present study, four AK genes were firstly identified from Yesso scallop (Patinopecten yessoensis) genome, respectively named PyAK1-4. PyAKs have highly conserved structures with a six-exon/five-exon structure, except for PyAK3. PyAK3 contains an unusual two-domain structure and a "bridge intron" between the two domains, which may originate from gene duplication and subsequent fusion. Phylogenetic analysis showed that all PyAKs belonged to an AK supercluster together with other AK proteins from Mollusca, Platyhelminthes, Arthropoda, and Nematode. A transcriptome database demonstrated that PyAK3 and PyAK4 were the main functional executors with high expression level during larval development and in adult tissues, while PyAK1 and PyAK2 were expressed at a low level. Furthermore, both PyAK2 and PyAK3 showed notably high expression in the male gonad, and PyAK4 was broadly expressed in almost all tissues with the highest level in striated muscle, indicating a tissue-specific expression pattern of PyAKs. In addition, quantitative real-time PCR results demonstrated that the expression of PyAK2, PyAK3 and PyAK4 were significantly upregulated in response to pH stress, especially in an extremely acidifying condition (pH 6.5), revealing the possible involvement of PyAKs in energetic homeostasis during environmental changes. Collectively, a comprehensive analysis of PyAKs was conducted in P. yessoensis. The diversity of PyAKs and their specific expression patterns promote a better understanding of energy metabolism in the growth, development and environmental response of P. yessoensis.

Metabolic responses of adult lion's paw scallops Nodipecten subnodosus exposed to acute hyperthermia in relation to seasonal reproductive effort.

In marine ectotherms, reproduction is an energetically expensive process that affects their thermal window tolerance. For most species, the impacts of hyperthermia during gametogenesis have still not been addressed. Our aim was to assess the metabolic response of adult Nodipecten subnodosus scallops to thermal challenges at early development (spring) and advanced gonad maturation (summer). Scallops collected in both seasons were exposed to acute hyperthermia (26 and 30 °C, 24 h), maintaining a group of scallops at acclimation temperature (22 °C) as a control condition. During the summer, relatively low activity of hexokinase (HK), as well as low levels of ATP and GTP were found in the adductor muscle, suggesting a shift in energy investment for reproduction, although arginine phosphate (ArgP) levels were higher in summer scallops. Hyperthermia (30 °C) induced an increased energy expenditure reflected by a transitory enhanced oxygen consumption (VO2) and relatively high activities of HK and arginine kinase (AK). Moreover, a slight decrease in adenylic energy charge (AEC) was partially compensated by a decrease in ArgP. An increase in nucleotide by-products inosine monophosphate (IMP) and hypoxanthine (HX) indicated a thermal stress at 30 °C. Some of the responses to acute hyperthermia were more pronounced at advanced maturation stages (summer scallops), indicating a possible lack of energy balance, with possible implications in animals challenged to global warming scenario.

Enzymes and non-enzymatic antioxidants responses to sequential cold stress in polymorphic noble scallop Chlamys nobilis with different total carotenoids content.

Noble scallop, an economically important edible marine bivalve displays polymorphism in shells (golden and brown) and flesh colors (orange and white). Mass mortality of noble scallops usually occurs during the winter months. Interestingly, carotenoid-rich golden scallops demonstrated much higher survival rates than brown scallops in winter. In order to understand the response of polymorphic noble scallops to sequential cold stress, the present study aimed to investigate the enzyme and non-enzymatic antioxidant responses of golden and brown scallops under sequential cold stress. Parameters evaluated included total carotenoid content (TCC), fatty acid composition, total antioxidant capacity (TAC), methylenedioxyamphetamine (MDA) content, catalase (CAT) activity, and superoxide dismutase (SOD) enzyme activity. The results of the present study revealed that golden scallops have higher cold tolerance than brown scallops. Golden and brown scallops are well adapted to low water temperature of above 12 °C, but in areas where winter water temperatures are below 12 °C, golden scallops are more suitable for aquaculture than brown scallops. The findings of this study are crucial to understanding the physiological responses of polymorphic scallops to cold stress and identify suitable candidates for winter aquaculture.

A carotenoid oxygenase is responsible for muscle coloration in scallop.

As lipid microconstituents mainly of plant origin, carotenoids are essential nutrients for humans and animals, and carotenoid coloration represents an important meat quality parameter for many farmed animals. Currently, the mechanism of carotenoid bioavailability in animals is largely unknown mainly due to the limited approaches applied, the shortage of suitable model systems and the restricted taxonomic focus. The mollusk Yesso scallop (Patinopecten yessoensis) possessing orange adductor muscle with carotenoid deposition, provides a unique opportunity to research the mechanism underlying carotenoid utilization in animals. Herein, through family construction and analysis, we found that carotenoid coloration in scallop muscle is inherited as a recessive Mendelian trait. Using a combination of genomic approaches, we mapped this trait onto chromosome 8, where PyBCO-like 1 encoding carotenoid oxygenase was the only differentially expressed gene between the white and orange muscles (FDR = 2.75E-21), with 11.28-fold downregulation in the orange muscle. Further functional assays showed that PyBCO-like 1 is capable of degrading ß-carotene, and inhibiting PyBCO-like 1 expression in the white muscle resulted in muscle coloration and carotenoid deposition. In the hepatopancreas, which is the organ for digestion and absorption, neither the scallop carotenoid concentration nor PyBCO-like 1 expression were significantly different between the two scallops. These results indicate that carotenoids could be taken up in both white- and orange-muscle scallops and then degraded by PyBCO-like 1 in the white muscle. Our data suggest that PyBCO-like 1 is the essential gene for carotenoid metabolism in scallop muscle, and its downregulation leads to carotenoid deposition and muscle coloration.

SOX2 participates in spermatogenesis of Zhikong scallop Chlamys farreri.

As an important transcription factor, SOX2 involves in embryogenesis, maintenance of stem cells and proliferation of primordial germ cell (PGC). However, little was known about its function in mature gonads. Herein, we investigated the SOX2 gene profiles in testis of scallop, Chlamys farreri. The level of C. farreri SOX2 (Cf-SOX2) mRNA increased gradually along with gonadal development and reached the peak at mature stage, and was located in all germ cells, including spermatogonia, spermatocytes, spermatids and spermatozoa. Knockdown of Cf-SOX2 using RNAi leaded to a mass of germ cells lost, and only a few spermatogonia retained in the nearly empty testicular acini after 21 days. TUNEL assay showed that apoptosis occurred in spermatocytes. Furthermore, transcriptome profiles of the testes were compared between Cf-SOX2 knockdown and normal scallops, 131,340 unigenes were obtained and 2,067 differential expression genes (DEGs) were identified. GO and KEGG analysis showed that most DEGs were related to cell apoptosis (casp2, casp3, casp8), cell proliferation (samd9, crebzf, iqsec1) and spermatogenesis (htt, tusc3, zmynd10, nipbl, mfge8), and enriched in p53, TNF and apoptosis pathways. Our study revealed Cf-SOX2 is essential in spermatogenesis and testis development of C. farreri and provided important clues for better understanding of SOX2 regulatory mechanisms in bivalve testis.

Molecular identification of steroidogenesis-related genes in scallops and their potential roles in gametogenesis.

Sex steroids are crucial for controlling gametogenesis and germ cell maturation in vertebrates. It has been proposed that Yesso scallop (Mizuhopecten yessoensis) has the same sex steroids as those animals, but the scallop biosynthetic pathway is unclear. In this study, we characterized several steroidogenesis-related genes in M. yessoensis and proposed a putative biosynthetic pathway for sex steroids that is similar to that of vertebrates. Specifically, we identified several steroidogenesis-related gene sequences that encode steroid metabolizing enzymes: StAR-related lipid transfer (START) protein, 17α-hydroxylase, 17,20-lyase (cyp17a), 17ß-hydroxysteroid dehydrogenase (hsd17b), and 3ß-hydroxysteroid dehydrogenase (hsd3b). We sampled adult scallops throughout their reproductive phase to compare their degree of maturation with their intensity of mRNA expression. Semi-quantitative RT-PCR analysis revealed a ubiquitous expression of transcripts for steroid metabolizing enzymes (i.e., star, cyp17a, hsd17b, and hsd3b) in peripheral and gonadal tissues. Real-time PCR analysis revealed a high level of expression of star3 and cyp17a genes in gonadal tissues at the early stage of cell differentiation in scallops. Interestingly, mRNA expression of hsd3b and hsd17b genes showed a synchronous pattern related to degree of gonad maturity. These results indicate that both hsd3b and hsd17b genes are likely involved in steroidogenesis in scallops. We therefore believe that these steroid-metabolizing enzymes allow scallops to endogenously produce sex steroids to regulate reproductive events.

A mitochondrial manganese superoxide dismutase involved in innate immunity is essential for the survival of Chlamys farreri.

Superoxide dismutase (SOD) ubiquitously found in both prokaryotes and eukaryotes functions as the first and essential enzyme in the antioxidant system. In the present study, a manganese SOD (designated as CfmtMnSOD) was cloned from Zhikong scallop Chlamys farreri. The complete cDNA sequence of CfmtMnSOD contained a 681 bp open reading frame (ORF), encoding a peptide of 226 amino acids. A SOD_Fe_N domain and a SOD_Fe_C domain were found in the deduced amino acid sequence of CfmtMnSOD. The mRNA transcripts of CfmtMnSOD were constitutively expressed in all the tested tissues, including gill, gonad, hepatopancreas, hemocytes, mantle and muscle, with the highest expression level in hemocytes. After the stimulation of Vibrio splendidus, Staphylococcus aureus and Yarrowia lipolytica, the mRNA transcripts of CfmtMnSOD in hemocytes all significantly increased. The purified rCfmtMnSOD protein exhibited Mn2+ dependent specific and low stable enzymatic activities. After Vibrio challenge, the cumulative mortality of CfmtMnSOD-suppressed scallops was significantly higher than those of control groups and the semi-lethal time for CfmtMnSOD-suppressed scallops was rather shorter than those of control groups either. Moreover, the final mortality rate of CfmtMnSOD-suppressed group was significant higher than those of control groups, even without Vibrio challenge. All these results indicated that CfmtMnSOD was efficient antioxidant enzyme involved in the innate immunity, and also essential for the survival of C. farreri.

Accumulation and transformation of azaspiracids in scallops (Chlamys farreri) and mussels (Mytilus galloprovincialis) fed with Azadinium poporum, and response of antioxidant enzymes.

Azaspiracid (AZA) producing microalgae have been reported internationally and could potentially impact a variety of seafood. Scallops (Chlamys farreri) and mussels (Mytilus galloprovincialis) from China were fed with the AZA2 producer, Azadinium poporum, to study uptake, metabolism and oxidative stress in the shellfish. LC-MS/MS showed significant accumulation and differential metabolism of AZA2 in the scallops and mussels. In mussels AZA2 was metabolized to AZA19, with subsequent decarboxylation to AZA6. In scallops no AZA19 or AZA6 was observed, however, a novel AZA metabolite was formed that is isobaric with AZA19 ([M+H]+, m/z 886), but elutes at a different retention time. In addition it was noted that the scallop metabolite was stable during heating, while AZA19 has been shown to decarboxylate. Concentrations of reactive oxygen species (ROS) and activities of antioxidant enzymes were monitored. ROS levels increased slightly in the meat of scallops and mussels due to starvation in the acclimation and depuration periods, but reduced in the feeding periods with non-toxic Isochrysis galbana or toxic A. poporum. No obvious variations were found in activities for a range of antioxidant enzymes. These results provide new insights on the potential for accumulation and metabolism of AZAs in bivalve species relevant to this area of China, which is of importance considering the recent finding of AZA producing microalgae in the region.

Multi-biomarker approach in the scallop Chlamys farreri to assess PAHs pollution in Qingdao coastal areas of China.

A multi-biomarker approach was conducted in the scallop Chlamys farreri from three sites, denoted here as S1, S2, and S3, in Qingdao coastal areas of China in March, June, September and December 2014 to assess pollution from polycyclic aromatic hydrocarbons (PAHs) and to select appropriate biomarkers. A suite of biological responses of the gills and digestive glands of the scallops was assayed, including: (i) phase I detoxification enzymes of 7-ethoxyresorufin-O-deethylase (EROD), epoxide hydrolase (EH), and dihydrodiol dehydrogenase (DD) and phase II detoxification enzymes of glutathione-S-transferase (GST) and sulfotransferase (SULT); (ii) antioxidant enzymes: catalase (CAT), superoxide dismutase (SOD), and glutathione peroxidase (GPx); (iii) oxidative damage parameters: lipid peroxidation (LPO) expressed by malondialdehyde (MDA) contents, protein carbonylation (PC) and DNA damage (F value); and (iv) the metabolism-related genes of EH, DD, GST, SULT and SOD. Simultaneously, the concentrations of total PAHs along with 16 types of PAHs previously identified by the US Environmental Protection Agency (USEPA) and environmental parameters, including temperature and salinity together with pH, were measured. Using Principle Component Analysis (PCA), it was revealed that S2 was the most PAH-contaminated site, while S1 was identified as the least PAH-polluted site, which was consistent with the results utilizing the Biomarker Response Index (BRI); in other words, the biological health status of S2 was worse than S1 and S3. Moreover, the most suitable biomarkers to assess PAH pollution in Qingdao coastal areas proved to be DD mRNA expression and the F value in both the gills and digestive glands for the total PAHs, DD activity and PC contents or PC and MDA contents in the gills or digestive glands for 5 + 6 rings PAHs and DD mRNA expression in both the gills and digestive glands for 2 + 3 rings and 4 rings PAHs. Moreover, this study highlighted the possible use of the scallop Chlamys farreri for studying contamination due to PAHs and provided valuable information on environmental assessment.

Expression profiles of different glutathione S-transferase isoforms in scallop Chlamys farreri exposed to benzo[a]pyrene and chrysene in combination and alone.

Aquatic organisms are increasingly exposed to polycyclic aromatic hydrocarbons (PAHs) due to anthropogenic pressure. This study aimed at evaluating the response of Glutathione S-transferases (GSTs) in scallop Chlamys farreri against benzo[a]pyrene (BaP) and chrysene (CHR) exposure under laboratory conditions. Nine published GST genes were classified into six subfamilies and a new member of rho family was identified for the first time. Twelve GSTs (including nine published GST genes and three in transcriptome established by our laboratory) mRNA transcript levels in the gills, digestive glands, adductor muscle, mantle, testis, ovaries, blood cells of scallops were measured by real-time PCR. The results showed that the mRNA transcript levels of twelve GSTs, except GST-zeta, GST-mu and GST-microsomal, were highest in digestive gland. Accordingly, the mRNA expression levels of GSTs were measured in digestive glands of scallops exposed to BaP (0.1µg/L and 1µg/L), CHR (0.1µg/L and 1µg/L) and their mixtures (0.1µg/L BaP +0.1µg/L CHR and 1µg/L BaP +1µg/L CHR). The results indicated that different GST had specific response to different pollution exposure. In BaP exposure experiment, the mRNA expression level of GST-theta was a potential suitable biomarker. GST-sigma-2 and GST-3, which belonged to sigma class, were sensitive to CHR exposure while GST-microsomal was considered a potential ideal bioindicator to joint exposure of BaP and CHR. In summary, this study investigated the classification of GSTs and provided information about the expression profiles of different class GSTs after PAHs exposure.

Effects of TiO2 nanoparticles at predicted environmental relevant concentration on the marine scallop Chlamys farreri: An integrated biomarker approach.

Manufactured nanoparticles (NPs) have caused extensive concern about their toxic effects on the marine environment. However, the chronic toxicity of NPs at predicted environmental relevant concentration on the marine organisms is poorly understood. In this study, we investigated the oxidative stress, neurotoxicity and histopathological effects of TiO2 NPs at predicted environmental relevant concentration (1mg/L) to marine scallop Chlamys farreri. The results showed that TiO2 NPs caused obviously oxidative damage on the scallops as evidenced by the significantly elevated superoxide dismutase (SOD), catalase (CAT) activities and malondialdehyde (MDA) contents. The increased acetylcholine esterase (AChE) activities reflected neurotoxicity of TiO2 NPs. The histopathological analysis revealed alterations in the gill and digestive gland, such as dysplastic and necrosis. Additionally, integrated biomarker response (IBR) values indicated that TiO2 NPs can cause strong toxic effects on the scallop. These results suggested that predicted environmental relevant TiO2 NPs can cause adverse effects on scallops and IBR analysis can be used as an effective approach for risk assessment of NPs on the marine organisms.

Immune response of the bay scallop, Argopecten irradians, after exposure to the algicide palmitoleic acid.

Palmitoleic acid (PA) is an effective algicide against the toxin-producing dinoflagellate Alexandrium tamarense; however, its effects on the immune system of the edible bay scallop Argopecten irradians are unclear. Therefore, we investigated the effects of PA on the immune response in A. irradians by assessing total haemocyte counts (THC), alkaline phosphatase activity (ALP), nitrite oxide (NO), glutathione (GSH), and lactate dehydrogenase (LDH) levels, as well as the expression of immune-related genes (FREP, PGRP, HSP90, MnSOD, and Cu/ZnSOD) at various hours post-exposure (hpe) to the compound. THC decreased in PA-treated groups, whereas ALP increased significantly in all of the PA treatment groups at 3 hpe, after which it significantly decreased. The LDH and NO levels were significantly enhanced in the high and medium concentration group. Notably, the GSH level increased in all PA treatment groups at each time interval. Our study revealed that after treatment with different concentrations of PA, variable effects on the expression of genes involved in the immune system response were observed. The results of our study demonstrate that immersing scallops in PA at effective concentrations could result in differential effects on immune system responses and expression of immune-related genes. Specifically, PA may disrupt the endocrine system or affect signal transduction pathways in the scallops. Therefore, the present study highlights the potential risk of using the PA as an algicide to control algal bloom outbreaks in the marine environment.

Characterization of three mitogen-activated protein kinases (MAPK) genes reveals involvement of ERK and JNK, not p38 in defense against bacterial infection in Yesso scallop Patinopecten yessoensis.

Mitogen-activated protein kinases (MAPKs) are protein Ser/Thr kinases that play a vital role in innate immune responses by converting extracellular stimuli into a wide range of cellular responses. Although MAPKs have been extensively studied in various vertebrates and invertebrates, our current understanding of MAPK signaling cascade in scallop is in its infancy. In this study, three MAPK genes (PyERK, PyJNK, and Pyp38) were identified from Yesso scallop Patinopecten yessoensis. The open reading frame of PyERK, PyJNK, and Pyp38 was 1104, 1227, and 1104 bp, encoding 367, 408, and 367 amino acids, respectively. Conservation in some splicing sites was revealed across the three PyMAPKs, suggesting the common descent of MAPKs genes. The expression profiles of PyMAPKs over the course of ten different developmental stages showed that they had different expression patterns. In adult scallops, PyMAPKs were primarily expressed in muscles, hemocytes, gill, and mantle. To gain insights into their role in innate immunity, we investigated their expression profiles after infection with Gram-positive bacteria (Micrococcus luteus) and Gram-negative bacteria (Vibrio anguillarum). Significant difference in gene expression was only found in PyERK and PyJNK, but not Pyp38, suggesting Pyp38 may not participate in immune response to bacterial infection. Besides, PyERK and PyJNK exhibited more drastic change against the invasion of V. anguillarum than M. luteus, suggesting they could be more sensitive to Gram-negative bacteria than Gram-positive bacteria. This study provides valuable resource for elucidating the role of MAPK signal pathway in bivalve innate immune response.

Biological effects of mechanically and chemically dispersed oil on the Icelandic scallop (Chlamys islandica).

This study aimed to simulate conditions in which dispersant (Dasic NS) might be used to combat an oil spill in coastal sub-Arctic water of limited depth and water exchange in order to produce input data for Net Environmental Benefit Analysis (NEBA) of Arctic and sub-Arctic coastal areas. Concentration dependent differences in acute responses and long-term effects of a 48h acute exposure to dispersed oil, with and without the application of a chemical dispersant, were assessed on the Arctic filter feeding bivalve Chlamys islandica. Icelandic scallops were exposed for 48h to a range of spiked concentrations of mechanically and chemically dispersed oil. Short-term effects were assessed in terms of lysosomal membrane stability, superoxide dismutase, catalase, gluthatione S-transferases, glutathione peroxidases, glutathione reductase, glutathione, total oxyradical scavenging capacity, lipid peroxidation and peroxisomal proliferation. Post-exposure survival, growth and reproductive investment were followed for 2 months to evaluate any long-term consequence. Generally, similar effects were observed in scallops exposed to mechanically and chemically dispersed oil. Limited short-term effects were observed after 48h, suggesting that a different timing would be required for measuring the possible onset of such effects. There was a concentration dependent increase in cumulative post-exposure mortality, but long-term effects on gonadosomatic index, somatic growth/condition factor did not differ among treatments.