In this study, we performed an integrative analysis of transcriptomic and metabolomic data to systematically investigate the influence of two key phenotypic factors-sex and body size-on the resistance of Eriocheir sinensis (Chinese mitten crab) to Vibrio alginolyticus infection. Our findings revealed that sex exerts a substantially greater impact on bacterial resistance than body size. Among the four phenotypic groups, large-sized females exhibited the strongest resistance, followed by small-sized females, large-sized males, and small-sized males, indicating a clear trend of "females > males" and "large size > small size" in resistance capacity. Multi-omics analyses further uncovered the molecular basis underlying these phenotypic differences. At the transcriptomic level, female crabs exhibited significantly higher activation of immune-related pathways, including endocytosis, immune signaling, cytoskeletal regulation, and apoptosis, suggesting a stronger innate immune response compared to males. At the metabolomic level, large-sized individuals showed enhanced energy metabolic pathways, potentially providing the energetic support required to sustain robust immune responses.Moreover, this study is the first to comprehensively characterize the endocytic pathway utilized by E. sinensis in response to Vibrio challenge, including pathogen recognition and endocytic initiation, endosomal transport and maturation, and lysosomal degradation. Notably, the regulation of endocytic immune responses was also found to be more strongly influenced by sex than by body size. In summary, this study provides a systems-level understanding of how sex and body size collaboratively modulate the immune resistance of E. sinensis to Vibrio infection, offering valuable insights and molecular markers for disease-resistant breeding and precision aquaculture management in crustaceans.

Photobacterium damselae is a widespread pathogenic bacterium in marine ecosystems, primarily associated with fish skin ulcers. However, infections caused by this bacterium in shrimp are rarely reported. The dominant strain MRY0520, isolated from the hepatopancreas of diseased Litopenaeus vannamei , was identified as Photobacterium damselae subsp. damselae (Phdd) and exhibited high pathogenicity. Elucidating the genetic architecture and infection mechanisms of P. damselae is crucial for the effective prevention and control of related diseases. To characterize its genomic features, whole-genome sequencing and comprehensive functional annotation were performed on the virulent Phdd strain MRY0520, followed by comparative genomic analysis to investigate its evolutionary traits. The genomic sequences have been deposited in GenBank under accession numbers CP113238–CP113240. The genome of strain MRY0520 spans 4,451,849 bp and contains 3,663 coding genes with a combined length of 3,691,779 bp, accounting for 83% of the total genome. It encodes 208 tRNA genes, 62 rRNA genes (including 22 5S rRNA, 20 16S rRNA, and 20 23S rRNA), two CRISPR arrays, and two genomic islands. Comparative analysis revealed that strain MRY0520 exhibits the highest genomic homology with Phdd strain Wu-1. These findings provide a solid theoretical basis for understanding the genomic composition and evolutionary characteristics of MRY0520.

With intensifying global climate change and human activities, and with regional topography interactions, soil and water salinization has intensified, posing major ecological and environmental challenges worldwide. Here, we integrated histology, transmission electron microscopy, RNA sequencing (RNA-seq) and data-independent acquisition (DIA)-based proteomics to profile hepatopancreas responses of Exopalaemon carinicauda during acute sulfate stress (≤48 h). Sulfate exposure disrupted tubular architecture and organelle integrity, consistent with early cellular injury. Multi-omics analyses revealed metabolic reprogramming marked by suppressed glycolysis (e.g., HK2, ENO) and enhanced oxidative phosphorylation (e.g., ATP5F1B), together with activation of calcium signaling (e.g., SLC8A1, ADCY9) and reinforcement of antioxidant/one-carbon and glucose-branch pathways (e.g., SHMT2, PGAM2). These coordinated transcript-protein changes indicate a shift from rapid cytosolic ATP supply to mitochondrial ATP production while buffering Ca2+ overload and reactive oxygen species. Collectively, our results delineate the physiological and molecular adjustments that enable E. carinicauda to cope with sulfate conditions and provide mechanistic targets for selective breeding and water-quality management in saline-alkaline aquaculture.

Exopalaemon carinicauda is a commercially significant aquaculture species in China. However, outbreaks of “zombie disease” caused by Metschnikowia bicuspidata infection have led to substantial economic losses in its farming industry. Despite its growing impact, the molecular basis of E. carinicauda’s response to M. bicuspidata infection remains unexplored. To elucidate the molecular mechanisms involved, this study conducted a transcriptomic analysis of the E. carinicauda hepatopancreas under M. bicuspidata infection and non-infection conditions. Following transcriptome assembly, 67,811 unigenes were generated, exhibiting high N50 value of 1977 base pairs and high complete BUSCO of 94.68%. Among these, 22,561 unigenes were successfully annotated. Comparative gene expression analysis of M. bicuspidata-infected and uninfected samples at 60 h post-infection revealed 1991 DEGs, comprising 1224 upregulated and 767 downregulated transcripts. Functional enrichment analysis revealed that numerous DEGs participate in immune-associated pathways, particularly those related to pattern recognition, lysosomal function, cellular stress responses, and programmed cell death. In addition, a significant proportion of DEGs were linked to metabolic processes such as glycerophospholipid metabolism and protein digestion and absorption. To verify the reliability of the transcriptomic results, eight DEGs were randomly chosen for qRT-PCR validation, and their expression profiles showed strong agreement with the RNA-Seq data. Overall, this study provides a transcriptomic overview of the hepatopancreatic response of E. carinicauda to M. bicuspidata infection, offering insights into the underlying molecular mechanisms and a theoretical foundation for the prevention and management of “Zombie disease.”

C-type lectins (CTLs) are an important class of pattern recognition receptors (PRRs) that exhibit structural and functional diversity in invertebrates. Repetitive DNA sequences are ubiquitous in eukaryotic genomes, representing distinct modes of genome evolution and promoting new gene generation. Our study revealed a new CTL that is composed of two long tandem repeats, abundant threonine, and one carbohydrate recognition domain (CRD) in Exopalaemon carinicauda and has been designated EcTR-CTL. The full-length cDNA of EcTR-CTL was 1242 bp long and had an open reading frame (ORF) of 999 bp that encoded a protein of 332 amino acids. The genome structure of EcTR-CTL contains 4 exons and 3 introns. The length of each repeat unit in EcTR-CTL was 198 bp, which is different from the short tandem repeats reported previously in prawns and crayfish. EcTR-CTL was abundantly expressed in the intestine and hemocytes. After Vibrio parahaemolyticus and white spot syndrome virus (WSSV) challenge, the expression level of EcTR-CTL in the intestine was upregulated. Knockdown of EcTRCTL downregulated the expression of anti-lipopolysaccharide factor, crustin, and lysozyme during Vibrio infection. The recombinant CRD of EcTR-CTL (rCRD) could bind to bacteria, lipopolysaccharides, and peptidoglycans. Additionally, rCRD can directly bind to WSSV. These findings indicate that 1) CTLs with tandem repeats may be ubiquitous in crustaceans, 2) EcTR-CTL may act as a PRR to participate in the innate immune defense against bacteria via nonself-recognition and antimicrobial peptide regulation, and 3) EcTR-CTL may play a positive or negative role in the process of WSSV infection by capturing virions.

Temperature is a limiting factor in the growth of aquatic organisms and can directly affect many chemical and biological processes, including metabolic enzyme activity, aerobic respiration, and signal transduction. In this study, physiological, transcriptomic, and metabolomic analyses were performed to characterize the response of Litopenaeus vannamei to cold stress. We subjected L. vannamei to gradually decreasing temperatures (24 °C, 20 °C, 18 °C, 14 °C, and 12 °C) and studied the changes in the hepatopancreas. The results showed that extreme cold stress (12 °C) caused structural damage to the hepatopancreas of L. vannamei. However, shrimp exhibited response mechanisms to enhance cold tolerance, through regulating changes in key genes and metabolites in amino acid, lipid metabolism, and carbohydrate metabolism, including (a) increased level of methylation in cells to enhance cold tolerance; (b) increased content of critical amino acids, such as proline, alanine, glutamic acid and taurine, to ameliorate energy metabolism, protect cells from cold-induced osmotic imbalance, and promote ion transport and DNA repair; (c) accumulation of unsaturated fatty acids to improve cell membrane fluidity; and (d) regulation of the metabolic pattern shift to rely on anaerobic metabolism with a gradual decrease in aerobic metabolism and enhance glycolysis to produce enough ATP to maintain energy metabolic balance. When the temperature dropped further, cold stress impaired antioxidant and immune defense responses in shrimp. This study provides an integrated analysis of the physiology, transcriptome, and metabolome of L. vannamei in response to cold stress.

To explore the molecular mechanisms of the Litopenaeus vannamei response to infection by Photobacterium damselae, reveal its immune response and energetic metabolic effect, and provide a valuable genetic data source for the scientific prevention and control of Vibrio infection, transcriptomic analysis, RT-qPCR, and physiological and biochemical tests were conducted. The results showed that the expression of key genes involved in lipid and carbohydrate transport, such as apolipoprotein and TPS, was upregulated after pathogenic infection, which brought the accumulation of triacylglycerol and trehalose into the hemolymph. Additionally, the pathogenic infection selectively triggered an immune response in infected L. vannamei, activating certain immune pathways, such as the serpins and MAPK pathways. The pathogenic infection suppressed the activity of phenoloxidase (PO), and the prophenoloxidase (PPO) cascade responses were suppressed by the invasive bacteria. This paper will help us understand the energetic metabolism, immune response, and activation of the immune recognition response after pathogenic infection by P. damselae, and it lays a theoretical foundation for the biological prevention and control of P. damselae infection.

As a typical G protein-coupled receptor, the adipokinetic hormone receptor (AKHR) has seven transmembrane domains (TMDs), and its structure and function are similar to the gonadotropin-releasing hormone receptor (GnRHR) in vertebrates. However, there is a dearth of information on other components of the AKHR signaling pathway and how it functions in the interaction between insect hosts and parasitoids. In this study, we cloned and analyzed the multifunctional Ostrinia furnacalis AKHR (OfAKHR) cDNA (GenBank accession number MF797868). OfAKHR has a 2206 bp full-length cDNA, which includes an open reading frame containing 1194 bp. OfAKHR contains the typical seven TMDs, and a "DRY" motif. OfAKHR has the highest relative expression in the fat body and the fifth instar larvae. The results revealed that ApoLpⅢ, PPO2, GS, TPS, Cecropin, and Moricin decreased the transcription levels from 48 to 72 h after the knockdown of OfAKHR expression by dsOfAKHR injection in the fourth instar O. furnacalis larvae. The parasitization of Macrocentrus cingulum selectively upregulated the expression levels of nutrition metabolism and immune-related genes in parasitized O. furnacalis larvae, stimulated lysozyme activity, and obviously raised the concentrations of triglyceride and trehalose in the hemolymph of O. furnacalis larvae. However, they inhibited the activities of PO and trehalase. This study is conducive to a deeper cognition of the roles of OfAKHR in nutrition and immune homeostasis, coevolution, and coexistence between parasitic wasps and hosts. It also sheds light on the potential as the target of pest control reagents.

The aim of this study was to investigate the dynamic changes in the microbial communities of both the environment and gut of Litopenaeus vannamei, as well as to elucidate the mechanisms underlying microbial community assembly in greenhouse farming. 16S rDNA high-throughput sequencing and bioinformatics methods were used to carry out the research on the community structure of the microorganisms under greenhouse culture conditions in water, sediment, and gut microorganisms; correlations pertaining to environmental factors; the feasibility of using Source Tracker; and the mechanisms of community construction. The results show that the dominant microorganisms in water, sediment, and gut farming in a greenhouse environment varied and were subject to dynamic change. A variety of beneficial microbiota such as Bacillus were found in the gut, whereas a variety of microorganisms such as Marivita and Pseudomonas, which function as nitrogen and phosphorus removers, were present in water. Source Tracker and environmental correlation analyses showed that changes in the gut were associated with eutrophication indicators (total nitrogen, total phosphorus, ammonia nitrogen) and changes in environmental microorganisms (in water and sediment). The results of the community-building mechanism analysis show that stochastic processes determine the community-building directions of environmental and gut microorganisms. These findings will help us to understand the microbiota characteristics of shrimp ponds under greenhouse farming conditions, and the complex interactions between the shrimp gut and the environmental microbiota and environmental variables, as well as revealing the changing rules of the gut microbiota.

The Macrocentrus cingulum is a vital endoparasitic wasp that parasites in Ostrinia furnacalis larvae. It has been widely utilized for biological control of corn borer. Although previous studies have explored how the wasps suppress the host's immunity, further research is required to determine the impact of the wasps on the host's lipid metabolism and activated immune response. In this study, high-throughput transcriptome sequencing, quantitative real-time PCR, and physiological and biochemical methods were used to investigate the balance between energy metabolism and immunity in parasitic systems. The results showed that specific genes linked to lipid metabolism and transportation were up-regulated in parasitized O. furnacalis larvae, including AKHR, AKH, apoLpIII, and FAS. Additionally, M. cingulum triggered immune responses in the parasitized larvae, leading to rapid recognition and the production of immunity effectors, such as Lysozyme and antimicrobial peptides, to protect against the invading wasps. The phenoloxidase (PO) activity was inhibited as a result of parasitization. Parasitism selectively promoted lipid metabolism and activated the immune recognition response, significantly up-regulated immune pattern recognition receptors (PRRs) and effector genes, but suppressed the PPO cascade response in O. furnacalis larvae. This study can shed light on lipid metabolism, immune response, and activating the immune recognition response for parasitoid and host co-evolution and provide insight into the biological control of O. furnacalis.

Temperature is an important factor in the physiological processes of aquatic organisms and can seriously affect several chemical and biological processes in their bodies, including respiratory metabolism, antioxidant capacity, immune capacity, and signal transduction. In this study, physiological, transcriptomic, and metabolomic analyses were used to investigate the response of Penaeus vannamei to cold stress. The results indicated that cold stress disrupted nucleotide metabolism and inhibited gluconeogenesis. However, shrimp exhibited response mechanisms in order to enhance cold tolerance, regulating changes in key genes and metabolites in amino acid and lipid metabolism to increase energy supply and protect cell membrane stability. In addition, the response included regulating Ca2+ pumps and ion channels to maintain intracellular ion homeostasis and osmotic balance. When the temperature dropped further, oxidative damage occurred due to overwhelming of the antioxidant defense system, and immune function was inhibited. This research provides some references regarding the molecular mechanisms involved in responding to cold stress and potential strategies to improve cold tolerance in P. vannamei; these are important references for studying the cold stress response of shrimp.

以虾肝肠胞虫(EHP)、致急性肝胰腺坏死病副溶血性弧菌(VpAHPND)、十足目虹彩病毒1(DIV1)和白斑综合征病毒(WSSV)4种常见病原为研究对象,将微流控芯片技术与LAMP技术有机结合,建立了相应的基因芯片联检系统,并开展了优化了基因芯片的反应体系及条件研究.结果表明,该联检系统可在30 min反应时间内同时检测上述4种病原,特异性好.各病原指标检出限分别为 EHP 0.57 copies·μL-1、DIV1 0.43 copies·μL-1、WSSV 2.12 copies·μL-1 和 VpAHPND 3.18 copies·μL-1,已达到甚至超越了 LAMP检测的敏感性,且该系统检测试剂盒具有操作简便,检测灵敏,特异性强,检测时间短,设备要求简单等特点,适合养殖全程中凡纳滨对虾EHP、VpAHPND、DIV1和WSSV的监测与筛查工作.

To evaluate the performance of microfluidic chip method in the joint detection of four common pathogens of Litopenaeus vannamei including Enterocytozoon hepatopenaei（EHP）, acute hepatopancreas necrosis disease caused by Vibrio parahaemolyticus（VpAHPND）, Decapod iridescent virus 1（DIV1） and white spot syndrome virus（WSSV）, primers for the amplification reactions of the above four pathogens were designed, new functional divisions were made for the existing chips, and the reaction system and constant temperature amplification conditions of the microfluidic chip were optimized. The results showed that, the optimized combined detection method could detect the above four pathogens simultaneously with good specificity, and the joint detection results could be obtained within 30 min reaction time. The detection limits of EHP, DIV1, WSSV and VpAHPND were 0.57 copies/μl, 0.43 copies/μl, 2.12 copies/μl and 3.18 copies/μl, respectively. Some of the limits had reached or even exceeded the testing sensitivity of loop-mediated isothermal amplification（LAMP） method. The detection kit used in the joint detection method of microfluidic chip for the four pathogens of L. vannamei in this test is characterized by simple operation, sensitive detection, strong specificity, short detection time and simple equipment requirements, and is suitable for the monitoring and screening of EHP, VpAHPND, DIV1 and WSSV of L. vannamei in the whole breeding process.

小型温棚养殖模式(下称小棚模式)是近年来凡纳滨对虾(Litopenaeus vannamei)养殖的热点模式.为阐明小棚模式养殖水体微生物群落的特点,探究该模式高产、高效的原因,基于16SrRNA基因的测序结果,对第2、第44、第69和第96天小棚模式养殖水体的微生物群落组成和功能进行了分析.结果表明:凡纳滨对虾小棚养殖后期,水体微生物群落的丰富度和多样性显著高于前期;在整个养殖过程中,门水平主要优势菌为变形菌门、拟杆菌门和放线菌门,其中变形菌门丰度在第69天显著增加,放线菌门丰度前期较高,后期下降;属水平优势菌属中Candidatus_Aquiluna的丰度在第2天最高(28.7％);海命菌属(Marivita)在中期富集,在第69天丰度最高(9.94％);黄杆菌属(Flavobacterium)的丰度随着养殖时间逐渐增加,在第96天达到最高(11.63％).通过PICRUSt2预测微生物群落的功能,丰度前20的代谢功能项在第69和第96天的丰度显著高于第2天,尤其萜类和聚酮类代谢、脂类代谢、异种生物降解和代谢等功能均高度富集,通过FAPROTAX鉴定得出,化能异养类菌的丰度在养殖后期显著增加.环境因子关联分析发现,总氮(TN)和化学需氧量(COD)对小棚模式水体微生物群落结构的影响最大,海命菌属、Candidatus_Aquiluna、红小梨形菌属(Rhodo-pirellula)等微生物发挥了固碳、降氮、降磷的作用.综上所述,在高密度、高氮磷的环境条件下,小棚模式的水体微生物在保持水环境稳定和增强对虾免疫与抗病能力方面发挥了重要作用.

Salinity is an important factor in the aquatic environment, and its fluctuations always result in osmotic stress, which affects the survival, distribution, and physiological activities of crustaceans. Crustaceans counter them through osmoregulation, which consists of many mechanisms. Palaemon gravieri is an important economic species in Palaemonidae, widely distributed in the southern East China Sea and the China Yellow Sea, and has a good adaptability to salinity stress. Currently, there are only a few studies on the effects of salinity on P. graviera. Therefore, it is particularly important to study the molecular responses of P. gravieri to salinity fluctuations. In this study, P. gravieri was treated with salinities of 10, 25, and 40, and the hepatopancreas and gills of shrimp in the different salinity groups were sampled after 24 h. The samples were used for RNA extraction and transcriptome analysis. In total, 80,994 unigenes were obtained, of which 19,114 were annotated. The differences in gene expression between different tissues at the same salinity were more significant. Many metabolism-related genes were downregulated in the gills, such as beta-hexosaminidase subunit alpha (HEXA), 10-formyltetrahydrofolate dehydrogenase (ALDH1L1), and Alcohol dehydrogenase class-3 (ADH5). Scanning transmission electron microscope analysis showed that the expression levels of some stress-(but not salinity stress) related genes changed after stress (mostly upregulated), suggesting the existence of secondary stress. Gene set enrichment analysis (GSEA) focused on the expression of transporters in osmoregulation, and the results showed that they mainly played a role in the gills, but ATP-binding cassette (ABC) transporters were more active in the hepatopancreas. This study showed that the response of P. gravieri to salinity change was different not only between the hepatopancreas and gills, but also between low salinity and higher salinity, and the ion transport-related genes were mainly expressed in the gills. Overall, these results improve our understanding of salt tolerance mechanism in P. gravieri.

The forkhead box transcription factor O family protein (FOXO) acts as a transcription factor that regulates biological processes regarding DNA repair, immunity, cell cycle regulation, and other biological processes. In this study, EcFOXO was identified from the ridgetail white prawn, Exopalaemon carinicauda. EcFOXO protein contains multiple low-complexity regions and a forkhead (FH) domain. Phylogenetic tree showed that EcFOXO is clustered with crustacean FOXOs. The amino acid sequences of its FH domain are highly similar to the FH domain of FOXOs from other crustaceans. The expression of EcFOXO is altered after white spot syndrome virus (WSSV) stimulation in hepatopancreas and gills. The relationship between EcFOXO and EcRelish was explored by RNA interference (RNAi). Results showed that EcFOXO and EcRelish could positively regulate each other’s expression. The expression levels of various antimicrobial peptides (AMPs) significantly reduced after interfering with EcFOXO or EcRelish. These results suggest a positive regulatory loop between EcFOXO and EcRelish, which participates in the innate immunity of ridgetail white prawn by regulating the expression of AMPs during WSSV infection. This study enriches the knowledge about the regulatory mechanism of FOXO in the innate immunity of crustaceans.

为确定 2018 年冬季以来江苏省沿海地区养殖脊尾白虾患"僵尸病"的病原及流行病学特点,实验采用LB培养基和PDA培养基从病虾血淋巴中分离得到直径为 1～3 mm、边缘整齐、米黄色隆起菌落;人工回感实验结果显示,回感后的脊尾白虾表现出与自然患病脊尾白虾相同的症状,并在回感脊尾白虾体内也分离出了相同的菌株,符合科赫氏法则.对该菌株进行形态观察结合 18S rRNA序列对比及系统发育分析,发现分离菌株MQ2101具有酵母的典型形态,且与二尖梅奇酵母相似度达 99.82%,结果表明菌株MQ2101 为二尖梅奇酵母.致病性结果初步分析显示,MQ2101 对脊尾白虾的半致死浓度(LD50)为1.39×107 CFU/尾.病理学观察发现,患病脊尾白虾鳃、肌肉和肝胰腺均发生不同程度的病变,其中肝胰腺病变最为严重,肝小管呈现空泡化,管腔体积变大;在鳃和肝胰腺组织中均存在大量定殖的菌体.流行病学调查结果显示,每年 2-5 月发病迅速,发病率为5%～30%,死亡率为 3%～10%.本研究确定了二尖梅奇酵母为江苏沿海地区脊尾白虾"僵尸病"的病原,其对脊尾白虾具有较强致病性,主要侵染组织为肝胰腺和鳃.以上研究结果为脊尾白虾"僵尸病"的防控提供了相关科学依据.

近年来,急性肝胰腺坏死病(Acute hepatopancreatic necrosis disease,AHPND)造成了凡纳滨对虾(Litopenaeus vannamei)养殖业巨大的经济损失,其病原为一类含有致病基因PirAB的弧菌(Vibrio),主要为副溶血弧菌(Vibrio parahaemolyticus,VPAHPND).本研究以VPAHPND为受试菌株,从江苏海域捕获的黄姑鱼(Nibea albiflora)体内分离筛选对VPAHPND具有抑制作用的拮抗菌株.通过形态学、生理生化及分子生物学方法进行鉴定,研究了其拮抗效果、拮抗活性物质,并对生物安全性和被动保护效果进行评估.结果表明,分离筛选出1株具有良好拮抗效果的菌株JSHY97,经鉴定该菌株为铜绿假单胞菌(Pseudomonas aeruginosa).该菌株对3株具有不同分型的VPAHPND均具有良好的拮抗效果,其抑菌圈直径达19.25 mm～21.80 mm,初步判断其活性物质来源于胞外产物.口服JSHY97的凡纳滨对虾的7 d保护率也达到80％,拮抗组的凡纳滨对虾体内的毒力基因PirA拷贝数显著低于对照组(P<0.05).安全性实验表明JSHY97对斑马鱼(Brachydanio reriovar)和凡纳滨对虾无毒力.由此表明,本研究得到的铜绿假单胞菌菌株JSHY97对VPAHPND具有较好的抑制作用,为AHPND的防治提供了新的菌株资源.

细菌耐药性是全球关注的热点,但水产动物致病菌耐药性的产生和对养殖、人类及环境的影响具有滞后性,容易被忽略.本研究从水产动物耐药性致病菌的含义、污染现状、产生原因、危害、存在问题、预防和削减方法等角度进行剖析,以期引起业界重视.