Temperature is an important factor affecting the growth, development and survival of marine organisms. A short episode of high temperature has been proven to be a severe threat to sustainable shellfish culture. Zhikong scallop (Chlamys farreri), a shellfish with broad economic and biological value in North China, has frequently experienced heat stress in summer in recent years. To understand the effects of heat stress on shellfish, the metabolism of C. farreri was analyzed after exposure to 27 °C for either 6 h or 30 d. After 6 h of heat stress exposure, a total of 326 and 264 significantly different metabolites (SDMs) were identified in gill and mantle tissues, respectively. After 30 d of heat stress exposure, a total of 381 and 341 SDMs were found in the gill and mantle tissues, respectively. These SDMs were mainly related to the metabolism of amino acids, carbohydrates, lipids and nucleotides. A decline in pyruvic acid, and an increase in citric acid and fumaric acid in the gills and mantle of C. farreri indicated an alteration in energy metabolism, which may be attributed to increased ATP production in order to overcome the heat stress. Among the SDMs, 33 metabolites, including pyruvic acid, glycine and citric acid, were selected as potential biomarkers for heat stress response in C. farreri. In addition, a decline in glutamine and β-Alanine levels indicated oxidative stress in C. farreri exposed to heat, as well as an increase in the total antioxidant capacity (T-AOC). Our findings suggested C. farreri have the potential to adapt to heat stress by regulating energy metabolism and antioxidant capacity.

Background: Gonadal development is driven by a complex genetic cascade in vertebrates. However, related information remains limited in molluscs owing to the long generation time and the difficulty in maintaining whole life cycle in the lab. The dwarf surfclamMulinia lateralisis considered an ideal bivalve model due to the short generation time and ease to breed in the lab.Results: To gain a comprehensive understanding of gonadal development inM. lateralis, we conducted a combined morphological and molecular analysis on the gonads of 30 to 60 dpf. Morphological analysis showed that gonad formation and sex differentiation occur at 35 and 40–45 dpf, respectively; then the gonads go through gametogenic cycle. Gene co-expression network analysis on 40 transcriptomes of 35–60 dpf gonads identifies seven gonadal development-related modules, including two gonad-forming modules (M6, M7), three sex-specific modules (M14, M12, M11), and two sexually shared modules (M15, M13). The modules participate in different biological processes, such as cell communication, glycan biosynthesis, cell cycle, and ribosome biogenesis. Several hub transcription factors includingSOX2,FOXZ,HSFY, FOXL2andHES1are identified. The expression of top hub genes from sex-specific modules suggests molecular sex differentiation (35 dpf) occurs earlier than morphological sex differentiation (40–45 dpf).Conclusion: This study provides a deep insight into the molecular basis of gonad formation, sex differentiation and gametogenesis inM. lateralis, which will contribute to a comprehensive understanding of the reproductive regulation network in molluscs.

Heat shock proteins 90 (HSP90s) are a class of ubiquitous, highly conserved, and multi-functional molecular chaperones present in all living organisms. They assist protein folding processes to form functional proteins. In the present study, three HSP90 genes, CfHSP90, CfGRP94 and CfTRAP1, were successfully identified in the genome of Chlamys farreri. The length of CfHSP90, CfGRP94 and CfTRAP1 were 7211 bp, 26,457 bp, and 28,699 bp, each containing an open reading frame (ORF) of 2181 bp, 2397 bp, and 2181 bp, and encoding proteins of 726, 798, and 726 amino acids, respectively. A transcriptomic database demonstrated that CfHSP90 and CfGRP94 were the primary functional executors with high expression during larval development and in adult tissues, while CfTRAP1 expression was low. Furthermore, all of the three CfHSP90s showed higher expression in gonads and ganglia as compared with other tissues, which indicated their probable involvement in gametogenesis and nerve signal transmission in C. farreri. In addition, under heat stress, the expressions of CfHSP90 and CfGRP94 were significantly up-regulated in the mantle, gill, and blood, but not in the heart. Nevertheless, the expression of CfTRAP1 did not change significantly in the four tested tissues. Taken together, in coping with heat stress, CfHSP90 and CfGRP94 could help correct protein folding or salvage damaged proteins for cell homeostasis in C. farreri. Collectively, a comprehensive analysis of CfHSP90s in C. farreri was conducted. The study indicates the functional diversity of CfHSP90s in growth, development, and environmental response, and our findings may have implications for the subsequent in-depth exploration of HSP90s in invertebrates.

Recently, elevated seawater temperatures have resulted numerous adverse effects, including significant mortality among bivalves. The dwarf surf clam, Mulinia lateralis, is considered a valuable model species for bivalve research due to its rapid growth and short generation time. The successful cultivation in laboratory setting throughout its entire life cycle makes it an ideal candidate for exploring the potential mechanisms underlying bivalve responses to thermal stress. In this study, a total of 600 clams were subjected to a 17-day thermal stress experiment at a temperature of 30 °C which is the semi-lethal temperature for this species. Ninety individuals who perished initially were classified as heat-sensitive populations (HSP), while 89 individuals who survived the experiment were classified as heat-tolerant populations (HTP). Subsequently, 179 individuals were then sequenced, and 21,292 single nucleotide polymorphisms (SNPs) were genotyped for downstream analysis. The heritability estimate for survival status was found to be 0.375 ± 0.127 suggesting a genetic basis for thermal tolerance trait. Furthermore, a genome-wide association study (GWAS) identified three SNPs and 10 candidate genes associated with thermal tolerance trait in M. lateralis. These candidate genes were involved in the ETHR/EHF signaling pathway and played pivotal role in signal sensory, cell adhesion, oxidative stress, DNA damage repair, etc. Additionally, qPCR results indicated that, excluding MGAT4A, ZAN, and RFC1 genes, all others exhibited significantly higher expression in the HTP (p < 0.05), underscoring the critical involvement of the ETHR/EHF signaling pathway in M. lateralis' thermal tolerance. These results unveil the presence of standing genetic variations associated with thermal tolerance in M. lateralis, highlighting the regulatory role of the ETHR/EHF signaling pathway in the bivalve's response to thermal stress, which contribute to comprehension of the genetic basis of thermal tolerance in bivalves.

Bivalves hold an important role in marine aquaculture and the identification of growth-related genes in bivalves could contribute to a better understanding of the mechanism governing their growth, which may benefit high-yielding bivalve breeding. Somatostatin receptor (SSTR) is a conserved negative regulator of growth in vertebrates. Although SSTR genes have been identified in invertebrates, their involvement in growth regulation remains unclear. Here, we identified seven SSTRs (PySSTRs) in the Yesso scallop, Patinopecten yessoensis, which is an economically important bivalve cultured in East Asia. Among the three PySSTRs (PySSTR-1, -2, and -3) expressed in adult tissues, PySSTR-1 showed significantly lower expression in fast-growing scallops than in slow-growing scallops. Then, the function of this gene in growth regulation was evaluated in dwarf surf clams (Mulinia lateralis), a potential model bivalve cultured in the lab, via RNA interference (RNAi) through feeding the clams Escherichia coli containing plasmids expressing double-stranded RNAs (dsRNAs) targeting MlSSTR-1. Suppressing the expression of MlSSTR-1, the homolog of PySSTR-1 in M. lateralis, resulted in a significant increase in shell length, shell width, shell height, soft tissue weight, and muscle weight by 20%, 22%, 20%, 79%, and 92%, respectively. A transcriptome analysis indicated that the up-regulated genes after MlSSTR-1 expression inhibition were significantly enriched in the fat digestion and absorption pathway and the insulin pathway. In summary, we systemically identified the SSTR genes in P. yessoensis and revealed the growth-inhibitory role of SSTR-1 in bivalves. This study indicates the conserved function of somatostatin signaling in growth regulation, and ingesting dsRNA-expressing bacteria is a useful way to verify gene function in bivalves. SSTR-1 is a candidate target for gene editing in bivalves to promote growth and could be used in the breeding of fast-growing bivalves.

Previous studies have shown that post-thaw sperm performance is affected by multiple stressors during cryopreservation, such as those induced by physical, chemical, mechanical and physiological changes. One of these is the balance disturbance between the antioxidant defense system and reactive oxygen species (ROS) production. This study investigated whether this disturbance could be alleviated by the addition of different antioxidants to cryoprotective solution [8% dimethyl sulfoxide (DMSO) in 1 µm filtered seawater] optimized for the sperm in dwarf surf clam Mulinia lateralis, the model bivalve species used in many different types of studies. Results showed that the addition of 20 μM coenzyme Q10 (Q10) to 8% DMSO achieved a D-stage larval rate similar to that of the fresh control at a sperm-to-egg ratio at least 50% less than the 8% DMSO treatment alone. The addition of other antioxidants (glycine, melatonin and polyvinylpyrrolidone) did not have any positive effects. The improvement in post-thaw sperm quality by Q10 could be due to its ability to significantly decrease ROS production and lipid peroxidation and significantly increase the motility, plasma membrane integrity, mitochondrial membrane potential, acrosome integrity, DNA integrity and activities of catalase and glutatione. In this study, 37 fatty acids (FAs) were quantified in dwarf surf clam sperm, with 21 FAs being significantly impacted by the cryopreservation with 8% DMSO. Thirteen of these 21 FAs were changed due to the addition of 20 μM Q10 to 8% DMSO, with approximately half of them being improved significantly toward the levels of fresh control, while the remaining half extended further from the trends shown with 8% DMSO treatment. However, no significant difference was found in the percentage of each FA category sum and the ratio of unsaturated/saturated FAs between the two treated groups. In conclusion, the antioxidant Q10 has shown the potential to further improve the sperm cryopreservation technique in bivalves.