Manila Clam Ruditapes Philippinarum Research Articles

Investigating the role of mitochondrial membrane potential in paternal inheritance of mitochondria

Abstract The process of oxidative phosphorylation (OXPHOS) in mitochondria depends on an electrochemical gradient known as the mitochondrial membrane potential (Δψm). Reflecting high functionality, elevated Δψm usually depicts healthy mitochondria and contributes to organelle selection. This study investigates whether mitochondrial properties linked with bioenergetics, such as Δψm, play a role in paternal inheritance of mitochondria. More specifically, the study looks at how sperm Δψm responds to egg chemoattractants in bivalves characterized by distinct mitochondrial inheritance patterns: strict maternal inheritance (SMI) and doubly uniparental inheritance (DUI), the latter displaying sex-specific transmission of paternal mitochondrial DNA. Sperm Δψm was examined in four bivalve species: the blue mussel (Mytilus edulis) and the Manila clam (Ruditapes philippinarum) (DUI), plus the hard clam (Mercenaria mercenaria) and the soft-shell clam (Mya arenaria) (SMI). In the absence of egg chemoattractants, sperm Δψm did not vary between the two groups. However, there was a trend of increase in Δψm following egg detection only in sperm bearing paternally derived mitochondria (DUI). This suggests, along with bioenergetic changes, that Δψm modulation might be a specific property of at least some DUI species, possibly implicated in their unique ability to transmit their mitochondria in a sex-specific fashion.

Chronic Effects of Waterborne Silver on the Physiology and Reproduction of the Manila Clam Ruditapes philippinarum

Chronic Effects of Waterborne Silver on the Physiology and Reproduction of the Manila Clam Ruditapes philippinarum

Physiological and transcriptomic analysis provides new insights into osmoregulation mechanism of Ruditapes philippinarum under low and high salinity stress

The Manila clam (Ruditapes philippinarum) is a commercially important marine bivalve, which inhabits the estuarine and mudflat areas. The osmoregulation is of great significance for molluscs adaptation to salinity fluctuations. In this study, we investigated the effects of low salinity (10 psu) and high salinity (40 psu) stress on survival and osmoregulation of the R. philippinarum. The results of physiological parameters showed that the ion (Na+, K+, Cl−) concentrations and Na+/K+-ATPase (NKA) activity of R. philippinarum decreased significantly under low salinity stress, but increased significantly under high salinity stress, indicating that there are differences in physiological adaptation of osmoregulation of R. philippinarum. In addition, we conducted the transcriptome analysis in the gills of R. philippinarum exposed to low (10 psu) and high (40 psu) salinity challenge for 48 h using RNA-seq technology. A total of 153 and 640 differentially expressed genes (DEGs) were identified in the low salinity (LS) group and high salinity (HS) group, respectively. The immune (IAP, TLR6, C1QL4, Ank3), ion transport (Slc34a2, SLC39A14), energy metabolism (PCK1, LDLRA, ACOX1) and DNA damage repair-related genes (Gadd45g, HSP70B2, GATA4) as well as FoxO, protein processing in endoplasmic reticulum and endocytosis pathways were involved in osmoregulation under low salinity stress of R. philippinarum. Conversely, the ion transport (SLC6A7, SLC6A9, SLC6A14, TRPM2), amino acid metabolism (GS, TauD, ABAT, ALDH4A1) and immune-related genes (MAP2K6, BIRC7A, CTSK, GVIN1), and amino acid metabolism pathways (beta-Alanine, Alanine, aspartate and glutamate, Glutathione) were involved in the process of osmoregulation under high salinity stress. The results obtained here revealed the difference of osmoregulation mechanism of R. philippinarum under low and high salinity stress through physiological and molecular levels. This study contributes to the assessment of salinity adaptation of bivalves in the context of climate change and provides useful information for marine resource conservation and aquaculture.

Shell shape polymorphism analysis of the Manila clam Ruditapes philippinarum across different geographical populations in China

Morphology of bivalves, a kind of quantitative character, is closely connected with the ecological and physiological behaviors of bivalves, including growth, survival, metabolism and reproduction. In this study, a multivariate statistical method was used to analyze seven indices for shell shape evaluation of Ruditaps philippinarum (R. philippinarum). Polymorphism of R. philippinarum shell shape across seven geographical populations from the southern and northern regions of China was revealed, and decisive effect of shell shape in determining the trait of weight was discussed. In total, three types of R. philippinarum shell shape were observed: globular, flat, and lateral-view round. The Laizhou (LZ) population from the north of China displayed an approximately globular shell shape, with a shell-width-to-shell-length ratio of 0.5496 and an ellipse index ventral view (EIV) value of 0.9543. The Lianjiang (LJ) population from the south of China displayed a flat shell shape, with a shell-width-to-shell-length ratio of 0.4044, an ellipse index lateral view (EIL) value of 1.2518, and an EIV value of 0.7063. The Zhangzidao (ZZD) population from the north of China displayed a lateral view round shell shape, with a shell-width-to-shell-length ratio of 0.7476 and a circle index lateral view (CIL) value of 0.7195. According to the results of principal component analysis (PCA), discriminant analysis and cluster analysis on shell shape parameters, LJ population and LZ population could be grouped separately. However, despite the long geographical distance, Rongcheng (RC) population was highly similar with ZZD population in terms of shell shape traits, implying that these two populations might be more closely related. Lowest degree of divergence in shell shape was observed in Beihai (BH) population and Yingkou (YK) population, both of which belonged to Zuidong (ZD) population. Weight of globular Manila clams was significantly higher than that of flat clams (P < 0.05). This study not only lays a foundation for conservation of R. philippinarum germplasm resources, but also highlights the importance of considering shell morphology in the breeding and culture of R. philippinarum.

The lgi-miR-2d is Potentially Involved in Shell Melanin Synthesis by Targeting mitf in Manila Clam Ruditapes philippinarum.

Shell color as an important economic trait is also the crucial target trait for breeding and production. MicroRNA (miRNA) is an endogenous small non-coding RNA that can post-transcriptionally regulate the expression of target genes, it plays important roles in many life activities and physiological processes, such as shell color, stress response, and disease traits. In this study, we investigated the function of lgi-miR-2d in shell melanin formation and the expression patterns of lgi-miR-2d and target gene Rpmitf in Manila clam Ruditapes philippinarum. We further explored and verified the relationship between Rpmitf and lgi-miR-2d and identified the expression level of shell color-related gene changes by RNAi and injecting the antagomir of lgi-miR-2d, respectively. Our results indicated that lgi-miR-2d antagomir affected the expression of its target gene Rpmitf. In addition, the dual-luciferase reporter assay was conducted to confirm the direct interaction between lgi-miR-2d and Rpmitf. The results showed that the expression levels of melanin-related genes such as Rpmitf and tyr were significantly decreased in the positive treatment group compared with the blank control group after the Rpmitf dsRNA injection, indicating Rpmitf plays a crucial role in the melanin synthesis pathway. Taken together, we speculated that lgi-miR-2d might be negatively modulating Rpmitf, which might regulate other shell color-related genes, thereby affecting melanin synthesis in R. philippinarum.

Unlocking the Therapeutic Potential of a Manila Clam-Derived Antioxidant Peptide: Insights into Mechanisms of Action and Cytoprotective Effects against Oxidative Stress.

Reactive oxygen species (ROS) are implicated in various pathological conditions due to their ability to induce oxidative damage to cellular components. In this study, we investigated the antioxidant properties of a peptide isolated from the hydrolysate of Manila clam (Ruditapes philippinarum) muscle. Purification steps yielded RPTE2-2-4, exhibiting potent scavenging activities against DPPH•, HO•, and O2•-, akin to Vitamin C. Structural analysis showed that the isolated peptide, LFKKNLLTL, exhibited characteristics associated with antioxidant activity, including a short peptide length and the presence of aromatic and hydrophobic amino acid residues. Moreover, our study demonstrated the cytoprotective effects of the peptide against H2O2-induced oxidative stress in HepG2 cells. Pretreatment with the peptide resulted in a dose-dependent reduction in intracellular ROS levels and elevation of glutathione (GSH) levels, indicating its ability to modulate cellular defense mechanisms against oxidative damage. Furthermore, the peptide stimulated the expression of the cytoprotective enzyme heme oxygenase-1 (HO-1), further reinforcing its antioxidant properties. Overall, our findings highlight the potential of the Manila clam-derived peptide as a natural antioxidant agent with therapeutic implications for oxidative stress-related diseases. Further investigation into its mechanisms of action and in vivo efficacy is warranted to validate its therapeutic potential.

Enhancing Seed Harvest of Wild Manila Clam (Ruditapes philippinarum) through Fiberglass Mesh Installation in the Tidal Flat of Jugyo, West Coast of Korea

Enhancing Seed Harvest of Wild Manila Clam (Ruditapes philippinarum) through Fiberglass Mesh Installation in the Tidal Flat of Jugyo, West Coast of Korea

New evidence of fraudulent mislabeling and illegal harvesting of Manila clams (Ruditapes philippinarum) through elemental fingerprints of their shells and chemometric analyses

Seafood is traded globally and is increasingly vulnerable to mislabeling and illegal, unregulated, and unreported (IUU) harvesting. These illicit activities prevail within the Manila clams (Ruditapes philippinarum) industry in the Tagus estuary (TE) in Portugal. This scenario prompted the use of elemental fingerprints (EF) of bivalve shells to confirm (or refute) claims on the geographic origin of this pricey seafood. To expose fraudulent practices, we mimicked real case scenarios and assessed the EF of R. philippinarum shells from three Portuguese estuaries: Ria de Aveiro (RAv), TE, and the Sado estuary (SE), as well as from two blind samples (B1 and B2) suspected of having been illegally harvested from TE, but mislabeled as originating from SE. Two reference models were developed using the EF of specimens from RAv, TE, and SE using CAP and Random Forests (RF). Both models confirmed the geographic origin of these clams (100% accuracy for CAP and 97.8% for RF). The same models predominantly allocated specimens from B1 and B2 to TE (93.3% and 100% for CAP; 100% and 93.3% for RF, respectively). Manganese and Phosphorus contributed to the classification of reference models, being present in high levels in the shells of clams from TE, B1 and B2, thereby being key for the allocation of blind samples to TE. This study confirms the potential of using EF of bivalve shells to trace the geographic origin of Manila clams under real-case scenarios, making it possible to strengthen the fighting against the mislabeling of seafood and IUU fishing.

Genome-wide sequencing reveals geographical variations in the thermal adaptation of an aquaculture species with frequent seedling introductions

Climate change and human activity are essential factors affecting marine biodiversity and aquaculture, and understanding the impacts of human activities on the genetic structure to increasing high temperatures is crucial for sustainable aquaculture and marine biodiversity conservation. As a commercially important bivalve, the Manila clam Ruditapes philippinarum is widely distributed along the coast of China, and it has been frequently introduced from Fujian Province, China, to other regions for aquaculture. In this study, we collected four populations of Manila clams from different areas to evaluate their thermal tolerance by measuring cardiac performance and genetic variations using whole-genome resequencing. The upper thermal limits of the clams showed high variations within and among populations. Different populations displayed divergent genetic compositions, and the admixed population was partly derived from the Zhangzhou population in Fujian Province, implying a complex genomic landscape under the influence of local genetic sources and human introductions. Multiple single nucleotide polymorphisms (SNPs) were associated with the cardiac functional traits, and some of these SNPs can affect the codon usage and the structural stability of the resulting protein. This study shed light on the importance of establishing long-term ecological and genetic monitoring programs at the local level to enhance resilience to future climate change.

Exploring the mechanism of nonylphenol-induced ovarian developmental delay of manila clams, Ruditapes philippinarum: Applying RNAi to toxicological analysis

Nonylphenol (NP) contamination in the coastal environment of China poses ecological risks to aquatic organisms. However, the endocrine disruptive impacts of NP on bivalves, particularly on ovarian development, remain poorly understood. In this study, Manila clams Ruditapes philippinarum at the developing stage of gonad were exposed to 1.0 μg/L NP for 21 days. Utilizing RNA interference (RNAi) to suppress ER gene expression, we observed a delay in ovarian development as evidenced by histological observations under both NP and NPRi (NP with ER-RNAi) treatment, with Vtg elevation exclusive to the NP group. Comprehensive analyses encompassing transcriptomics, real-time quantitative PCR, and steroid hormone measurement revealed significant alterations in aldosterone synthesis, estrogen signaling, and thyroid hormone synthesis. These pathways showed similar perturbations in both NP and NPRi groups compared to controls. Notably, the NPRi group exhibited distinct enrichment in PPAR and insulin signaling pathways, may implicating these in ER function suppression. Steroid hormone biosynthesis was notably reduced in both treatments, pointing to a profound impact on hormone synthesis. The contrast between in vivo and in vitro findings suggests that NP's detrimental effects on ovarian development may primarily involve neuroendocrine regulation of steroidogenesis. This investigation highlights the complex dynamics of NP-induced endocrine disruption in bivalves, emphasizing the pivotal role of ER and associated pathways.

Promoting effect of Manila clam Ruditapes philippinarum (Adams & Reeve, 1850) on the establishment of eelgrass Zostera marina L. seedlings

It is necessary to restore the declined eelgrass Zostera marina L. beds which have multiple functions. The utilization of seedlings to recover eelgrass beds can maintain the genetic eelgrass diversity; however, the seedling establishment rate that is affected by many habitat factors is very low. Manila clam Ruditapes philippinarum (Adams & Reeve, 1850) transform habitats through several ways. Thus, it is inferred that the clam affects the establishment of eelgrass seedlings in multiple ways, and the seedling establishment rate of eelgrass might not simply increase or decrease under the presence of clams. To verify this hypothesis, a five-month clam-mediated seedling establishment experiment was conducted. The contents of total nitrogen (TN) and exchangeable ammonia NH4-N (Ex-NH) in sediment increased as the clam body size increased, and TN decreased with the increase in clam abundance. There was a positive relationship between the content of organic matter in sediment (OM) and the interaction of body size and abundance of clam. The emergence rate of germinated seeds and the seedling establishment rate increased with increasing clam biomass. They were positively correlated with the interaction of OM and Ex-NH. The seedling establishment rate was negatively correlated with the interaction of water content in sediment (W) and medium diameter of sediment (Ø50), OM and TN. The presence of clams affected the height, chlorophyll b content and superoxide dismutase (SOD) activity of seedlings. The clam body size affected the SOD activity of seedlings by influencing Ex-NH. Thus, the habitat modification of clams with moderate biomass improves the establishment and traits of seedlings in multiple ways. The seedling establishment mediated by Manila clam is useful for eelgrass bed restoration.

Establishment and application of multiplex microsatellite genotyping for parentage assignment in Manila clam Ruditapes philippinarum

The manila clam Ruditapes philippinarum has become an important bivalve species in worldwide aquaculture due to its delicious taste and high commercial value. With the rapid expansion of clam farming, there is an urgent need to obtain genetic improvement by selective breeding, such as family-based selection of improved varieties. In this study, 10 microsatellite loci were selected to establish four multiplex PCR groups using four different fluorescences (FAM, ROX, HEX, TAM). The accuracy of these loci was verified by the 100% success rate of parentage assignment in a full-sib family. Parentage assignment was carried out in the mixed-mating parents and their offspring with unknown pedigree information. The genotyping of ten microsatellite loci generated 90 alleles, having the observed heterozygosity (Ho) ranging from 0.333 to 0.729. The expected heterozygosity (He) varied from 0.358 to 0.794. The PIC ranged from 0.342 to 0.763, with a mean value of 0.620. The theoretical combined parentage exclusion (cEP) power for all ten microsatellites was 97% with known genotypes of one parent (P1) and 99% for given the genotypes of both parents (P2). The actual successful assignment rate reached 98% by using all four sets of multiplex microsatellites, serving as the effective method for parentage assignment in the clams. The development and application of multiplex microsatellite genotyping methods provide a cost-effective method for pedigree reconstruction, population genetics and selective breeding of the commercially important clams.

Transcriptome analysis provides new insights into the immune response of Ruditapes philippinarum infected with Vibrio alginolyticus

Manila clam (Ruditapes philippinarum) is a bivalve species with commercial value, but it is easily infected by pathogenic microorganisms in aquaculture, which restricts the shellfish industry. Notably, the impact of Vibrio alginolyticus on clam culture is obvious. In this study, RNA-seq was performed to analyze clam hepatopancreas tissue in 48 h (challenge group, G48h) and 96 h (challenge group, G96h) after infection with V. alginolyticus and 0 h after injection of PBS (control group, C). The results showed that a total of 1670 differentially expressed genes were detected in the G48h vs C group, and 1427 differentially expressed genes were detected in the G96h vs C group. In addition, KEGG analysis showed that DEGs were significantly enriched in pathways such as Lysosome and Mitophagy. Moreover, 15 immune related DEGs were selected for qRT-PCR analysis to verify the accuracy of RNA-seq, and the results showed that the expression level of DEGs was consistent with that of RNA-seq. Therefore, the results obtained in this study provides a preliminary understanding of the immune defense of R. philippinarum and molecular insights for genetic breeding of V. alginolyticus resistance in Manila clam.

Transcriptome analysis of Manila clam (Ruditapes philippinarum) under Vibrio parahaemolyticus challenge in a low-salinity environment provides molecular insights into immune response

Bacterial infections caused high mortality of Manila clam (Ruditapes philippinarum) after the sudden drop in salinity of seawater, which caused serious economic losses. Therefore, we explored the molecular mechanism of V. parahaemolyticus-mediated immunity of R. philippinarum under low-salinity condition. In this study, Manila clams treated with low-salinity (CD), V. parahaemolyticus-treated under low-salinity condition (HCD), and a blank group (CC) were sequenced through the Illumina HiSeq platform, and we verified 15 immune-related DEGs (12 up-regulated genes and 3 down-regulated genes) through quantitative real-time PCR (qRT-PCR), the results were consistent with those detected from the transcriptome. In addition, the results of KEGG pathway enrichment analysis showed that the different expression genes (DEGs) were mostly enriched in the lysosome and phagosome pathway exhibited upregulated expression throughout the V. parahaemolyticus infection. Among them, lysosome and phagosome-related genes were induced after R. philippinarum treated with V. parahaemolyticus. Thus, our results proved that the important role of regulatory mechanism of immune homeostasis mediated by the lysosome and phagosome pathway in R. philippinarum against V. parahaemolyticus infection. Altogether, this work provides a new information about the immune regulatory mechanisms of Manila clam response to V. parahaemolyticus challenge in a low-salinity environment.

Characterization of Eighty-Eight Single-Nucleotide Polymorphism Markers in the Manila Clam Ruditapes philippinarum Based on High-Resolution Melting (HRM) Analysis.

Single-nucleotide polymorphisms (SNPs) are the most commonly used DNA markers in population genetic studies. We used the Illumina HiSeq4000 platform to develop single-nucleotide polymorphism (SNP) markers for Manila clam Ruditapes philippinarum using restriction site-associated DNA sequencing (RAD-seq) genotyping. Eighty-eight SNP markers were successfully developed by using high-resolution melting (HRM) analysis, with a success rate of 44%. SNP markers were analyzed for genetic diversity in two clam populations. The observed heterozygosity per locus ranged from 0 to 0.9515, while the expected heterozygosity per locus ranged from 0.0629 to 0.4997. The value of FIS was estimated to be from -0.9643 to 1.0000. The global Fst value was 0.1248 (p < 0.001). After Bonferroni correction, 15 loci deviated significantly from the Hardy-Weinberg equilibrium (p < 0.0006). These SNP markers provide a valuable resource for population and conservation genetics studies in this commercially important species.

Behavioral responses of intertidal clams to compound extreme weather and climate events

Rapidly increasing concentration of carbon dioxide (CO2) in the atmosphere not only results in global warming, but also drives increasing seawater acidification. Infaunal bivalves play critical roles in benthic-pelagic coupling, but little is known about their behavioral responses to compound climate events. Here, we tested how heatwaves and acidification affected the behavior of Manila clams (Ruditapes philippinarum). Under acidified conditions, the clams remained capable of burrowing into sediments. Yet, when heatwaves attacked, significant decreases in burrowing ability occurred. Following two consecutive events of heatwaves, the clams exhibited rapid behavioral acclimation. The present study showed that the behavior of R. philippinarum is more sensitive to heatwaves than acidification. Given that the behavior can act as an early and sensitive indicator of the fitness of intertidal bivalves, whether, and to what extent, behavioral acclimation can persist under scenarios of intensifying heatwaves in the context of ocean acidification deserve further investigations.

Occurrence and seasonal variation of Perkinsus sp. Infection in wild mollusk populations from coastal waters of Qingdao, northern China

Perkinsosis has been recognized as one of the major threats to natural and farmed bivalve populations, many of which are of commercial as well as environmental significance. Three Perkinsus species have been identified in China, and the Manila clam (Ruditapes philippinarum) was the most frequently infected species in northern China. Although the occurrence and seasonal variation of Perkinsus spp. have previously been examined, the pathological characteristics of these infections in wild Manila clams and sympatric species in China have seldom been reported. In the present study, the prevalence and intensity of Perkinsus infection in wild populations of Manila clams and 10 sympatric species from three sites were investigated by Ray’s fluid thioglycolate medium (RFTM) assay seasonally across a single year. Perkinsus infection was only identified in Manila clams, with a high prevalence (274/284 = 96.48 %) and low intensity (89.8 % with a Mackin value ≤ 2, suggesting generally low-intensity infections) throughout the year. Heavily infected clams were mainly identified in Tianheng in January, which displayed no macroscopic signs of disease. An overview of the whole visceral mass section showed that the trophozoites mostly aggregated in gills and connective tissue of the digestive tract, to a lesser extent in the mantle and foot, and even less frequently in adductor muscle and connective tissues of the gonad. PCR and ITS-5.8S rRNA sequencing of 93 representative RFTM-positive samples revealed a 99.69 to 100 % DNA sequence identity to Perkinsus olseni. Unexpectedly, significantly higher infection intensities were usually identified in January and April when the Condition Index (CI) was relatively high. We propose that factors associated with the anthropogenic harvesting pressure and irregular disturbances should be responsible for the uncommon seasonal infection dynamics of perkinsosis observed in the present study.

Spatio-Temporal Variation in Perkinsus olseni Infection Intensity in Manila Clam Ruditapes philippinarum in Anmyeondo and Cheonsu Bay Tidal Flats on the West Coast of Korea

Spatio-Temporal Variation in Perkinsus olseni Infection Intensity in Manila Clam Ruditapes philippinarum in Anmyeondo and Cheonsu Bay Tidal Flats on the West Coast of Korea

Bioconcentration, biotransformation, and transcriptomic impact of the UV-filter 4-MBC in the manila clam Ruditapes philippinarum

Ultraviolet filters (UV-filters) are compounds extensively used in personal care products. These compounds are produced at increasing rates and discharged into marine ecosystems in unknown quantities and with no regulation, making them emerging contaminants. Among those, the UV-filter 4-Methylbenzylidene camphor (4-MBC) is used in a variety of personal care products such as sunscreens, soaps, or lipsticks. This high consumption has resulted in its presence in various environmental matrices at in concentrations ranging from ng to μg L−1. Very little is known, however, about the possible adverse effects in exposed non-target organisms. Our study presents novel data on the bioconcentration, toxicokinetics, and molecular effects of 4-MBC in a marine bivalve species of commercial interest, Ruditapes philippinarum (Manila clam). Organisms were exposed at two different concentrations (1.34 and 10.79 μg L−1) of 4-MBC for 7 days, followed by a 3-day depuration period (clean sea waters). Bioconcentration factors (BCF) were 3562 and 2229 L kg−1 for the low and high exposure concentrations, respectively, making this pollutant bioaccumulative according to REACH criteria. Up to six 4-MBC biotransformation products (BTPs)were identified, 2 of them for the first time. Transcriptomic analysis revealed between 658 and 1310 differently expressed genes (DEGs) after 4-MBC exposure. Functional and enrichment analysis of the DEGs showed the activation of the detoxification pathway to metabolize and excrete the bioconcentrated 4-MBC, which also involved energy depletion and caused an impact on the metabolism of carbohydrates and lipids and in the oxidative phosphorylation pathways. Oxidative stress and immune response were also evidenced through the activation of cathepsins and the complement system. Such elucidation of the mode of action of a ubiquitous pollutant such as 4-MBC at the molecular level is valuable both from an environmental point of view and for the sustainable production of Manila clam, one of the most cultivated mollusk species worldwide.

Metabolic adaptation of the clam Ruditapes philippinarum during air exposure and the positive effects of sodium nitroprusside pretreatment.

The Manila clam (Ruditapes philippinarum), as one of the shellfish living in the intertidal zone, is known for its strong ability to withstand air exposure. Sodium nitroprusside (SNP), a donor of nitric oxide (NO), has been shown to be useful for antioxidant and immune regulation in aquatic animals. In this study, an untargeted metabolomics (LC-MS/MS) technique was employed for the first time in Manila clam to analyze the metabolic and histological impacts after air exposure and the positive effects of SNP pretreatment. During air exposure, a significant increase in taurine, L-glutamate, and several polyunsaturated fatty acids in clams was detected, which indicates that clams may experience inflammatory reactions, oxidative stress, and an increase in blood ammonia content. When clams were exposed to SNP for 6h, arginine, spermine, L-glutamic acid, and glutathione content were all upregulated, indicating that the SNP exposure induced NO production and improved antioxidant capacity in clams. When the clams were exposed to air after SNP pretreatment, there were no significant differences in the levels of taurine, L-glutamate, or aliphatic acids between the experimental and control groups. Gill tissue was more severely damaged in clams directly exposed to air than in those that experienced air exposure after SNP pretreatment, especially in clams exposed to air for a long time (72h). Both metabolomics and tissue section structure indicated that SNP pretreatment decreased the stress responses caused by air exposure in R. philippinarum. These findings provided fresh insights and a theoretical foundation for understanding the tolerance to air exposure and physiological functions of SNP (or NO) in R. philippinarum.