HIF-1-mediated regulation of LDH gene unravels key insights into MCDV-1 pathogenesis in mud crabs Scylla paramamosain.

Hypoxia-inducible factors -1 (HIF-1) is a crucial transcription factor that regulates the expression of glycolytic genes. Our previous study proved that the Mud crab dicistrovirus-1 (MCDV-1) can induce aerobic glycolysis that favors viral replication in mud crab Scylla paramamosain. However, the role of HIF-1 on key glycolytic genes during the MCDV-1 infection has not been examined. In this study, the intricate interplay between HIF-1 and the key glycolysis enzyme, lactate dehydrogenase (LDH), was investigated after MCDV-1 infection. The expression of LDH was significant increased after MCDV-1 infection. Additionally, the expression of HIF-1α was upregulated following MCDV-1 infection, potentially attributed to the downregulation of prolyl hydroxylase domains 2 expression. Subsequent examination of the SpLDH promoter identified the presence of hypoxia response elements (HREs), serving as binding sites for HIF-1α. Intriguingly, experimental evidence demonstrated that SpHIF-1α actively promotes SpLDH transcription through these HREs. To further elucidate the functional significance of SpHIF-1α, targeted silencing was employed, resulting in a substantial reduction in SpLDH expression, activity, and lactate concentrations in MCDV-1-infected mud crabs. Notably, SpHIF-1α-silenced mud crabs exhibited higher survival rates and lower viral loads in hepatopancreas tissues following MCDV-1 infection. These results highlight the critical role of SpHIF-1α in MCDV-1 pathogenesis by regulating LDH gene dynamics, providing valuable insights into the molecular mechanisms underlying the virus-host interaction.

Identification and functional characterization of activating transcription factor 6 (ATF6) from the mud crab (Scylla paramamosain) in response to hydrogen peroxide and bacterial challenge.

Activating transcription factor 6 (ATF6) is critical for regulation of unfolded protein response (UPR), which is involved in the endoplasmic reticulum (ER) proteostasis maintenance and cellular redox regulation. In the present study, a ATF6 gene from the mud crab (designated as Sp-ATF6) has been cloned and identified. The open reading frame of Sp-ATF6 was 1917 bp, encoding a protein of 638 amino acids. The deduced amino acid sequences of Sp-ATF6 contained a typical basic leucine zipper (BZIP domain). Sp-ATF6 was widely expressed in all tested tissues, with the highest expression levels in the hemocytes and the lowest in the muscle. Subcellular localization showed that Sp-ATF6 was expressed in both nucleus and cytoplasm of S2 cells. The expression level of Sp-ATF6 was induced by hydrogen peroxide and V. parahaemolyticus challenge, indicating that the ATF6 pathway was activated in response to ER stress. In order to know more about the regulation mechanism of the Sp-ATF6, RNA interference experiment was investigated. Knocking down Sp-ATF6 in vivo can decrease the expression of antioxidant-related genes (CAT and SOD) and heat shock proteins (HSP90 and HSP70) after V. parahaemolyticus infection. All these results suggested that Sp-ATF6 played a crucial role in the defense against environmental stress and pathogen infection in crustaceans.

Essential role of the HSC70 in the mud crab Scylla paramamosain in response to Vibrio parahaemolyticus infection.

Heat shock proteins play an important role in host defense, and modulate immune responses against pathogen infection. In this study, a novel HSC70 from the mud crab (designated as SpHSC70) was cloned and characterized. The full length of SpHSC70 contained a 58 bp 5'untranslated region (UTR), an open reading frame (ORF) of 2,046 bp and a 3'UTR of 341 bp. The SpHSC70 protein included the conserved DnaK motif. The mRNA of SpHSC70 was highly expressed in the hemocytes, heart and hepatopancreas, and lowly expressed in the intestine. The subcellular localization results indicated that SpHSC70 was localized in both the cytoplasm and the nucleus. Moreover, SpHSC70 was significantly responsive to bacterial challenge. RNA interference experiment was designed to investigate the roles of SpHSC70 in response to bacterial challenge. V. parahaemolyticus infection induced the expression levels of SpPO, SpHSP70, SpSOD and SpCAT. Knocking down SpHSC70 in vivo can decrease the expression of these genes after V. parahaemolyticus infection. These results suggested that SpHSC70 could play a vital role in defense against V. parahaemolyticus infection via activating the immune response and antioxidant defense signaling pathways in the mud crab.

Characterization of a novel prolyl hydroxylase 2 gene from mud crab Scylla paramamosain: Insights into its role in the regulation of hypoxia-inducible factor-1α.

Prolyl hydroxylase 2 (PHD2) is the key oxygen sensor that regulates the stability of the hypoxia-inducible factor -1α (HIF-1α). In this study, a novel PHD2 gene from the mud crab Scylla paramamosain, named SpPHD2, was cloned and identified. The full-length transcript of SpPHD2 was found to be 1926 bp, consisting of a 333 bp 5' untranslated region, a 1239 bp open reading frame, and a 354 bp 3' untranslated region. The putative SpPHD2 protein contained a Prolyl 4-hydroxylase alpha subunit homologues (P4Hc) domain in the C-terminal and a Myeloid translocation protein 8, Nervy, and DEAF-1 (MYND)-type zinc finger (zf-MYND) domain in the N-terminal. The mRNA expression of SpPHD2 was found to be widely distributed across all examined tissues. Additionally, the subcellular localization results indicated that the SpPHD2 protein was mainly localized in the cytoplasm. The in vivo silencing of SpPHD2 resulted in the upregulation of SpHIF-1α and a series of downstream genes involved in the HIF-1 pathway, while SpPHD2 overexpression in vitro dose-dependently reduced SpHIF-1α transcriptional activity, indicating that SpPHD2 plays a crucial role in SpHIF-1α regulation. Interestingly, the expression of SpPHD2 increased under hypoxic conditions, which was further inhibited by SpHIF-1α interference. Moreover, four hypoxia response elements were identified in the SpPHD2 promoter, suggesting that a feedback loop exists between SpPHD2 and SpHIF-1α under hypoxia. Taken together, these results provided new insights into the regulation of SpPHD2 in response to hypoxia in S. paramamosain.

Glutaredoxin 2 in the mud crab Scylla paramamosain: Identification and functional characterization under hypoxia and pathogen challenge.

Glutaredoxin (Grx) is a glutathione-dependent oxidoreductase that plays a key role in antioxidant defense. In this study, a novel Grx2 gene (SpGrx2) was identified from the mud crab Scylla paramamosain, which consists of a 196 bp 5' untranslated region, a 357 bp open reading frame, and a 964 bp 3' untranslated region. The putative SpGrx2 protein has a typical single Grx domain with the active center sequence C-P-Y-C. The expression analysis revealed that the SpGrx2 mRNA was most abundant in the gill, followed by the stomach and hemocytes. Both mud crab dicistrovirus-1 and Vibrioparahaemolyticus infection as well as hypoxia could differentially induce the expression of SpGrx2. Furthermore, silencing SpGrx2 in vivo affected the expression of a series of antioxidant-related genes after hypoxia treatment. Additionally, SpGrx2 overexpression significantly increased the total antioxidant capacity of Drosophila Schneider 2 cells after hypoxia, resulting in a reduction of reactive oxygen species and malondialdehyde content. The subcellular localization results indicated that SpGrx2 was localized in both the cytoplasm and the nucleus of Drosophila Schneider 2 cells. These results indicate that SpGrx2 plays a crucial role as an antioxidant enzyme in the defense system of mud crabs against hypoxia and pathogen challenge.

Toxic effects of cadmium exposure on intestinal histology, oxidative stress, microbial community, and transcriptome change in the mud crab (Scylla paramamosain).

Cadmium is one of hazardous pollutants that has a great threat to aquatic organisms and ecosystems. The intestine plays important roles in barrier function and immunity to defend against environmental stress. However, whether cadmium exposure caused the intestine injury is not well studied. Thus, the aim of this study was to explore the potential mechanisms of cadmium toxicity in the intestine of mud crab (Scylla paramamosain) via physiological, histological, microbial community, and transcriptional analyses. Mud crabs were exposed to 0, 0.01, and 0.125 mg/L cadmium. After a 21-day of cadmium exposure, 0.125 mg/L cadmium caused intestine damaged by decreasing superoxide dismutase and catalase activities, and increasing hydrogen peroxide and malondialdehyde levels. Integrated biological index analysis confirmed that the toxicity of cadmium exhibited a concentration-dependent manner. Comparative transcriptional analyses showed that the up-regulations of several genes associated with heat shock proteins, detoxification and anti-oxidant defense, and two key signaling pathways (PI3k-Akt and apoptosis) revealed an adaptive response mechanism against cadmium exposure. Transcriptomic analysis also suggested that cadmium exposure disturbed the expression of ion transport and immune-related genes, indicating that it has negative effects on the immune functions of the mud crab. Furthermore, the intestinal microbial diversity and composition were significantly influenced by cadmium exposure. The abundance of the dominant phyla Fusobacteria and Bacteroidetes significantly changed after cadmium exposure. KEGG pathway analysis demonstrated that cadmium exposure could change energy metabolism and environmental information processing. Overall, we concluded that excessive cadmium exposure could be potentially exerted adverse effects to the mud crab health by inducing oxidative damage, decreasing immune system, disrupting metabolic function, and altering intestinal microbial composition. These results provided a novel insight into the mechanism of cadmium toxicity on crustaceans.

Essential role of the Cytochrome P450 2 (CYP2) in the mud crab Scylla paramamosain antioxidant defense and immune responses.

Cytochrome P450 (CYPs) enzymes are one of the critical detoxification enzymes, playing a key role in antioxidant defense. However, the information of CYPs cDNA sequences and their functions are lacked in crustaceans. In this study, a novel full-length of CYP2 from the mud crab (designated as Sp-CYP2) was cloned and characterized. The coding sequence of Sp-CYP2 was 1479 bp in length and encoded a protein containing 492 amino acids. The amino acid sequence of Sp-CYP2 comprised a conserved heme binding site and chemical substrate binding site. Quantitative real-time PCR analysis revealed that Sp-CYP2 was ubiquitously expressed in various tissues, and it was highest in the heart followed by the hepatopancreas. Subcellular localization showed that Sp-CYP2 was prominently located in the cytoplasm and nucleus. The expression of Sp-CYP2 was induced by Vibrio parahaemolyticus infection and ammonia exposure. During ammonia exposure, ammonia exposure can induce oxidative stress and cause severely tissue damage. Knocking down Sp-CYP2 in vivo can increase malondialdehyde content and the mortality of mud crabs after ammonia exposure. All these results suggested that Sp-CYP2 played a crucial role in the defense against environmental stress and pathogen infection in crustaceans.

Identification and functional characterization of glutaredoxin 5 from the mud crab (Scylla paramamosain) in response to cadmium and bacterial challenge.

Glutaredoxin (Grx) is a class molecule oxidoreductase, which plays a key role in maintaining redox homeostasis and regulating cell survival pathways. However, the expression pattern and function of Grx remain unknown in the mud crab (Scylla paramamosain). In the present study, a novel full-length of Grx 5 from the mud crab (designated as Sp-Grx 5) was cloned and characterized. The open reading frame of Sp-Grx 5 was 441 bp, which encoded a putative protein of 146 amino acids. The amino acid sequence of Sp-Grx 5 contained a typical C-G-F-S redox active motif and several GSH binding sites. Sp-Grx 5 widely existed in all tested tissues with a high-level expression in hepatopancreas. Subcellular localization showed that Sp-Grx 5 was located in the cytoplasm and nucleus. The expression of Sp-Grx 5 was significantly up-regulated after Vibrio parahaemolyticus infection and cadmium exposure, suggesting that Sp-Grx 5 was involved in innate immunity and detoxification. Furthermore, overexpression of Sp-Grx 5 could improve cells viability after H2O2 exposure. All these results indicated that Sp-Grx 5 played important roles in the redox homeostasis and innate immune response in crustaceans.

Characterization of phosphofructokinase (PFK) from mud crab Scylla paramamosain and its role in mud crab dicistrovirus-1 proliferation.

Phosphofructokinase (PFK), the key enzyme of glycolysis, can catalyze the irreversible transphosphorylation of fructose-6-phosphate forming fructose-1, 6-biphosphate. In the present study, a PFK gene from the mud crab Scylla paramamosain, named SpPFK, was cloned and characterized. The full length of SpPFK contained a 5'untranslated region (UTR) of 249 bp, an open reading frame of 2,859 bp, and a 3'UTR of 1,248 bp. The mRNA of SpPFK was highly expressed in the gill, followed by the hemocytes and muscle. The expression of SpPFK was significantly up-regulated after mud crab dicistrovirus-1 (MCDV-1) infection. Knocking down SpPFK in vivo by RNA interference significantly reduced the expression of lactate dehydrogenase after MCDV-1 infection. Furthermore, silencing of SpPFK in vivo increased the survival rate of mud crabs and decreased the MCDV-1 copies in the gill and hepatopancreas after MCDV-1 infection. All these results suggested that SpPFK could play an important role in the process of MCDV-1 proliferation in mud crab.

Hypoxia Affects the Antioxidant Activity of Glutaredoxin 3 in Scylla paramamosain through Hypoxia Response Elements.

Hypoxia is a major environmental stressor that can damage the oxidation metabolism of crustaceans. Glutaredoxin (Grx) is a key member of the thioredoxin superfamily and plays an important role in the host's defense against oxidative stress. At present, the role of Grx in response to hypoxia in crustaceans remains unclear. In this study, the full-length cDNA of Grx3 (SpGrx3) was obtained from the mud crab Scylla paramamosain, which contains a 129-bp 5' untranslated region, a 981-bp open reading frame, and a 1,183-bp 3' untranslated region. The putative SpGrx3 protein contains an N-terminal thioredoxin domain and two C-terminal Grx domains. SpGrx3 was expressed in all tissues examined, with the highest expression in the anterior gills. After hypoxia, SpGrx3 expression was significantly up-regulated in the anterior gills of mud crabs. The expression of Grx2 and glutathione S-transferases was decreased, while the expression of glutathione peroxidases was increased following hypoxia when SpGrx3 was silenced in vivo. In addition, the total antioxidant capacity of SpGrx3-interfered mud crabs was significantly decreased, and the malondialdehyde content was significantly increased during hypoxia. The subcellular localization data indicated that SpGrx3 was predominantly localized in the nucleus when expressed in Drosophila Schneider 2 (S2) cells. Moreover, overexpression of SpGrx3 reduced the content of reactive oxygen species in S2 cells during hypoxia. To further investigate the transactivation mechanism of SpGrx3 during hypoxia, the promoter region of the SpGrx3 was obtained by Genome Walking and three hypoxia response elements (HREs) were predicted. Dual-luciferase reporter assay results demonstrated that SpGrx3 was likely involved in the hypoxia-inducible factor-1 (HIF-1) pathway during hypoxia, which could be mediated through HREs. The results indicated that SpGrx3 is involved in regulating the antioxidant system of mud crabs and plays a critical role in the response to hypoxia.

The role of caspase 3 in the mud crab (Scylla paramamosain) after Vibrio parahaemolyticus infection.

Apoptosis plays essential roles in the immune defense mechanism against pathogen infection. Caspase 3 is a family of cysteine proteases involved in apoptosis and the immune response. In this study, the full-length of mud crab (Scylla paramamosain) caspase 3 (designated as Sp-caspase 3) was cloned and characterized. The open reading frame of Sp-caspase 3 was comprised a 1035 bp, which encoded a putative protein of 344 amino acids. Sp-caspase 3 was ubiquitously expressed in various tissues with a high-level expression in hemocytes. Cellular localization analysis revealed that Sp-caspase 3 was located in the cytoplasm and nucleus. Over-expression of Sp-caspase 3 could induce cell apoptosis. In addition, V. Parahaemolyticus infection induced the relative expression of caspase-3 mRNA and increased caspase-3 activity. Knocking down Sp-caspase 3 in vivo significantly reduced cell apoptosis and increased mortality of mud crab after V. parahaemolyticus infection. These results indicated that Sp-caspase 3 played important roles in the immune response and apoptosis against bacterial infection.

Oxidative stress, DNA damage, and cellular response in hydrogen peroxide-induced cell injury of mud crab (Scylla paramamosain).

Oxidative stress is considered as the toxicity mechanism of environmental stressors on aquatic organisms. This study aims to explore the effects of oxidative stress on physiological responses, DNA damage and transcriptional profiles of the mud crabs Scylla paramamosain. In the present study, mud crabs were injected with 0.1% and 1% hydrogen peroxide (H2O2) for 72 h. The results showed that superoxide dismutase and catalase activities significantly decreased after H2O2 injection. Malondialdehyde content, H2O2 content, aspartate aminotransferase, alanine aminotransferase and lactate dehydrogenase activity significantly increased after H2O2 injection. Moreover, DNA damage occurred after H2O2 injection. Transcriptome analysis showed that 531 and 372 differentially expressed genes (DEGs) were identified after 0.1% and 1% H2O2 injection, respectively. These DEGs were mainly involved in the oxidative stress response and immune functions. All these results indicated that oxidative stress could impair both antioxidant defense systems and immune systems. Transcriptome analysis provided valuable information on gene functions associated with the response to oxidative stress in the mud crab.

Hypoxia-induced oxidative stress and transcriptome changes in the mud crab (Scylla paramamosain).

Mud crab (Scylla paramamosain) is an economically important cultured species in China. Hypoxia is a major environmental stressor during mud crab culture. In the present study, we investigated the oxidative stress and transcriptome changes in the gills of mud crab after intermediate hypoxia stress with dissolved oxygen (DO) 3.0 ± 0.2 mg/L (named as "DO3") and acute hypoxia stress with DO 1.0 ± 0.2 mg/L (named as "DO1") for 0, 3, 6, 12 and 24 h. The superoxide dismutase (SOD) activity of DO1 increased significantly at 3, 6 and 24 h after hypoxia stress, while SOD activity of DO3 increased significantly at 6 and 24 h. The total antioxidant capacity (T-AOC) increased significantly at 6, 12 and 24 h after hypoxia stress. The malondialdehyde (MDA) concentration of DO1 increased significantly at 6, 12 and 24 h after hypoxia stress, while MDA concentration of DO3 only increased significantly at 6 h. The lactate dehydrogenase (LDH) activity of DO1 increased significantly at 3, 6, 12 and 24 h after hypoxia stress, while LDH activity of DO3 increased significantly at 12 and 24 h. Transcriptomic analysis was conducted at 24 h of gill tissues after hypoxia stress. A total of 1052 differentially expressed genes (DEGs) were obtained, including 394 DEGs between DO1 and DO3, 481 DEGs between DO1 and control group, 177 DEGs between DO3 and control group. DEGs were enriched in the pathways related to metabolism, immune functions, ion transport, and signal transduction. Transcriptional analysis showed that glycolysis and tricarboxylic acid cycle genes were the key factors in regulating the adaptation of mud crab to hypoxia stress.

The role of tumor suppressor protein p53 in the mud crab (Scylla paramamosain) after Vibrio parahaemolyticus infection.

The tumor suppressor protein p53 plays important roles in DNA repair, cell cycle and genetic stability. In the present study, a p53 gene in the mud crab (Scylla paramamosain) (designated as Sp-53) was identified and characterized. The open reading frame of Sp-53 was comprised a 1383 bp, which encoded a putative protein of 460 amino acids. Sp-53 is expressed in all examined tissues, with the highest expression in hepatopancreas and hemocytes. Vibrio parahaemolyticus infection induced oxidative stress, and led to DNA damage. The Sp-53 transcriptions in hepatopancreas were significantly up-regulated after V. parahaemolyticus infection. RNA interference (RNAi) experiment was used to understand the roles of Sp-53 in response to V. parahaemolyticus infection. Knocking down Sp-53 in vivo significantly reduced the expression of the Mn-SOD, Gpx3 and caspase 3 after V. parahaemolyticus infection. Moreover, the mortality of mud crabs and DNA damage in Sp-53-silenced mud crab challenged with V. parahaemolyticus were significantly higher than those in the control group. All these results suggested that Sp-53 played an important role in responses to V. parahaemolyticus infection through its participation in regulation of antioxidant defense, DNA repair and apoptosis.

Oxidative stress, cell cycle arrest, DNA damage and apoptosis in the mud crab (Scylla paramamosain) induced by cadmium exposure.

Cadmium is one of the most common heavy metal pollutants in the aquatic environment. Mud crab (Scylla paramamosain) is considered a model organism to monitor the impact of heavy metals. However, knowledge about toxicological mechanism of cadmium in crustaceans still remains limited. In this study, mud crabs were exposed to different concentrations of cadmium (0, 1.25, 2.5, 5 and 10 mg/L) for 72 h. Cadmium exposure significantly decreased superoxide dismutase (SOD) activity, catalase (CAT) activity and total antioxidative capacity (T-AOC), and significantly increased malondialdehyde (MDA) and H2O2 levels. Aspartate aminotransferase (AST), alanine aminotransferase (ALT) and lactate dehydrogenase (LDH) activity significantly increased after cadmium exposure. Moreover, integrated biological responses version 2 (IBRv2) analysis suggested that cadmium exposure exerted stronger toxicity on mud crab. Furthermore, oxidative stress induced by cadmium exposure could decrease total hemocyte count (THC), interrupt Ca2+ homeostasis, and lead to cytological damage. Cadmium exposure induced DNA damage, which activated DNA damage response signaling ATR-CHK1-p53 pathway. Our results also showed that cadmium exposure significantly increased the apoptosis and caspase-3 mRNA levels, which implied that cadmium induced apoptosis through a caspase-3 pathway.

Effects of Vibrio parahaemolyticus infection on physiological response, histopathology and transcriptome changes in the mud crab (Scylla paramamosain).

Mud crab (Scylla paramamosain) is an important economic species in China. Vibrio parahaemolyticus infection have caused a great economic loss in mud crab farming. The mechanism involved in the immune responses of mud crab to V. parahaemolyticus is unclear. In this study, the physiological and immune response to V. parahaemolyticus infection were investigated in S. paramamosain. The results showed that V. parahaemolyticus infection decreased total hemocyte counts, led to cytological damage, and caused high mortality. Transcriptome analysis showed that 1327 differentially expressed genes (DEGs), including 809 up-regulated and 518 down-regulated ones, were obtained after V. parahaemolyticus challenge. These DEGs were mainly involved in the immune response and infectious disease. Additionally, transcriptome analysis revealed that Toll, immune deficiency (IMD), and prophenoloxidase signalling pathways played essential roles in antibacterial immunity against V. parahaemolyticus infection in mud crab.

Immune and physiological responses of mud crab (Scylla paramamosain) under air exposure.

The immune and physiological responses of mud crab (Scylla paramamosain) under air exposure were studied. The results showed that air exposure increased plasma activities of AST, ALT, ALP. There was a significant increase in glucose (GLU) and malondialdehyde (MDA) levels after air exposure. The transcript levels of SOD, CAT, HSP90, HSP70, p53, and hypoxia-inducible factor-1 (HIF-1) were induced by air exposure. Furthermore, caspase-3 transcript significantly increased at 48 and 72 h, while it significantly decreased at 96 h and 120 h under air exposure. These results suggested that oxidative stress occurred in the prolonged period of air exposure. HIF-1 and p53 signaling pathways played an important role under air exposure.

The role of Mu-type glutathione S-transferase in the mud crab (Scylla paramamosain) during ammonia stress.

Glutathione S-transferase (GST) plays important roles in cellular detoxification and antioxidant defense. A Mu-type glutathione S-transferase (designated as SpMu-GST) was obtained from the mud crab Scylla paramamosain. The open reading frame of SpMu-GST was comprised a 690 bp, which encoded a putative protein of 229 amino acids. Quantitative real-time PCR (qRT-PCR) revealed that the SpMu-GST mRNA was expressed in all examined tissues, with highest expression in hepatopancreas. During ammonia exposure, the SpMu-GST transcriptions in hepatopancreas and gill were significantly up-regulated at early exposure time. Moreover, RNA interference (RNAi) experiment was designed to understand the roles of SpMu-GST under ammonia exposure. Ammonia exposure reduced the levels of glutathione S-transferase (GST), superoxide dismutase (SOD), catalase (CAT) and total antioxidative capacity (T-AOC), and increased the formation of malondialdehyde (MDA). After knockdown of the SpMu-GST level, GST activity and T-AOC were significantly decreased at some exposure time after ammonia exposure. However, the mortality of mud crabs and malondialdehyde (MDA) contents significantly increased under ammonia exposure. These results further suggested that SpMu-GST played a critical role in mud crab antioxidant defenses in response to environmental stress.

Effect of nitrite exposure on oxidative stress, DNA damage and apoptosis in mud crab (Scylla paramamosain).

Nitrite is one of major environmental pollutants that can impact immunological parameters in aquatic organisms. In the present study, we investigated the effects of nitrite exposure on oxidative stress, DNA damage and apoptosis in mud crab (Scylla paramamosain). Mud crab were exposed to 0, 5, 10 and 15 mg L-1 nitrite for 72 h. These data showed that acid phosphatase (ACP) and alkaline phosphatase (ALP) activity significantly decreased in treatments with various concentrations of nitrite (5, 10 and 15 mg L-1) after 24 and 48 h, while the levels of nitric oxide (NO) significantly increased in these treatments. Nitrite exposure could suppress superoxide dismutase (SOD) and catalase (CAT) activity, and increase the formation of malondialdehyde (MDA) after 48 and 72 h of exposure. In addition, nitrite exposure decreased total haemocyte counts after 48 and 72 h of exposure. Cytological damage, DNA damage and apoptosis was observed obviously at 72 h after nitrite exposure. Moreover, nitrite exposure significantly induced the mRNA levels of phosphorylated Jun N-terminal kinases (JNK), and eventually activated p53 signaling and caspase-3. These results indicated that nitrite exposure could induce oxidative stress, which further caused DNA damage and apoptosis in mud crab. Our results will be helpful to understand the mechanism of nitrite toxicity on crustaceans.