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The swimming crab, Portunus trituberculatus is one of crucial aquaculture crabs with significant differences in growth and economic performance between male and female swimming crabs. Consequently, the culture of female populations presents higher economic value. The doublesex and mab-3 related transcription factor (Dmrt) gene family are known to play crucial role in gonad differentiation and development. However, there is limited information about this gene family in Portunus trituberculatus. In this study, we identified seven members of the Dmrt gene family in P. trituberculatus based on the published transcriptome and genome data and designated as Ptdmrt-1, Ptdoublesex (Ptdsx), Ptidmrt-1, Ptdmrt-11E, Ptidmrt-2, Ptdmrt-99B, and Ptdmrt-3 based on the homology analysis results, respectively. These Ptdmrt genes distributed across 6 chromosomes and were predicted to encode 283 aa, 288 aa, 529 aa, 436 aa, 523 aa, 224 aa, and 435 aa protein precursors, respectively. The expression patterns of these dmrt genes were characterized by qRT-PCR and gonad transcriptome data. The results showed that five members (Ptdmrt-99B, Ptdsx, Ptdmrt-1, Ptdmrt-3, and Ptdmrt-11E) were differentially expressed between the testis and ovary. In addition, their expression patterns from zoea 2 to juvenile 1 were characterized by published transcriptome data and the results showed that they were lowly expressed and did not exhibit notable difference except for Ptdsx during early development. Overall, majority of Ptdmrt genes were involved in gonad differentiation in the swimming crab. Current findings provide a solid foundation for further exploration of the roles of these genes in gonad development and differentiation in P. trituberculatus., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier B.V.)

Background: Members belonging to the tick genus Hyalomma function as a multi-host reservoir for several pathogens and important parasites infesting large animals, such as camels, goats, cattle and sheep. In Egypt, there is a high risk of pathogen transmission as camels and cattle are imported from Sudan and Ethiopia and shipped to slaughterhouses and animal markets located in populated areas. Hyalomma dromedarii ticks are semi-desert vectors and, similar to other members of the genus Hyalomma, characterized by long-term feeding. During this process, different physiological, biochemical and immunological interactions occur within both the feeding ticks and their hosts. These biological changes affect the different tick developmental phases. The aim of this study was to explore the transcriptome of mixed messenger RNAs (mRNAs) collected from H. dromedarii eggs, larvae, nymphs and fed and unfed adults, using the Gateway cDNA library prepared in pCMV sport6.1 vector METHODS: The clones were sequenced and searched for potential secreted, membrane-associated or transmembrane (SMaT) sequences. The identified SMaT sequences were compared to the National Center for Biotechnology Information (NCBI) non-redundant protein sequence database using Blastx. Annotation and functional classification were achieved by comparison to sequences in the UniProtKB/Swiss-Prot and VectorBase databases and to the publicly available annotated proteomes of six hard tick species (H. asiaticum, Rhipicephalus sanguineus sensu lato, Dermacentor silvarum, Rhipicephalus microplus, Ixodes scapularis and Haemaphysalis longicornis) in addition to the published H. dromedarii sialotranscriptome. For the common sequences, we predicted the physicochemical properties, secondary structures and antigenicity of the fragments similar to matched sequences in the UniProtKB/Swiss-Prot database using three different methods., Results: The quality-trimmed sequences from the cDNA library revealed 319 SMaT transcripts among 1248 sequenced clones. Annotation of the SMaT sequences using the UniProtKB/Swiss-Prot database revealed only 232 non-redundant sequences with at least one match. According to the UniProtKB/Swiss-Prot and Vectorbase databases, the SMaT sequences were either secreted (extracellular) (29 sequences) or cellular (transmembrane and membrane-associated) (203 sequences). These were classified into 10 functional classes: biogenesis (49 sequences), defense (9 sequences), development (36 sequences), signal transduction (28 sequences), transport (15 sequences), protein modification (33 sequences), homeostasis (6 sequences), metabolism (45 sequences) and miscellaneous/uncharacterized (11 sequences). A total of 60 sequences were shared between H. dromedarii SMaT, the sialotransciptome and six other hard tick species. The peptide fragments of these sequences that aligned to proteins from the UniProtKB/Swiss-Prot database were predicted to be promising epitopes and mapped to 10 functional classes at different ratios., Conclusions: Our immuno-informatics analysis identified 60 sequences common among hard tick species and encoded by H. dromedarii salivary glands. These annotated SMaT sequences of H. dromedarii will pave the way for the identification and discovery of novel potential protective antigens that are either secreted, membrane-associated or transmembrane., Competing Interests: Declarations Ethics approval and consent to participate All steps were approved by the Ethical Review Board of the National Research Centre, Egypt (NRC#11127082023), in accordance with the Helsinki Declaration (1975). Consent for publication Not applicable. Competing interests The authors declare no competing interests., (© 2024. The Author(s).)

The salmon louse (Lepeophtheirus salmonis) is an ectoparasite feeding on mucus, skin, and blood of salmonids. On parasitised fish erosions and, at later lice stages, ulcerations appear at the louse feeding site. In susceptible species like Atlantic salmon (Salmo salar) with a limited rejection of lice, only a mild inflammatory response with minor influx of immune cells is seen at these lesions, as the salmon louse secrete proteins that can dampen immune responses. In a previous study, Lepeophtheirus salmonis labial gland protein 3 (LsLGP3) was suggested to dampen cellular responses, and the present study aimed at increasing our understanding of its mode of action. LsLGP3 was found to be secreted on to the host skin, and both in vivo and in vitro experiments were performed to elucidate its function. Histological analysis of the louse attachment site revealed an epidermal and dermal influx of mainly macrophages and granulocytes after 5 days post infestation. The immune cell influx was deeper in the dermis throughout the louse infestation, and LsLGP3 may be involved in dampening this response. Enriched populations of Atlantic salmon B-cells, T-cells, granulocytes, and monocytes were exposed to recombinant LsLGP3 (recLGP3) in vitro, resulting in a significant decrease in cell viability compared to non-exposed controls. An apoptotic cell morphology with "beads-on-a-string" like protrusions was seen in all leukocyte cell fractions after recLGP3 exposure, but not in erythrocytes or keratocytes. A decreased viability was also detected in pink salmon leucocytes, which was not in leucocytes from non-salmonid species. These functional insights suggest that LsLGP3 specifically induces apoptosis of salmonid leukocytes and is likely a key protein secreted by the lice that disables the Atlantic salmon ability to mount an adequate immune response towards the salmon louse. In vivo LsLGP3 knock down studies indicated that the effect is localised primarily at the lice feeding site, without affecting immune cells that are not situated adjacent to the lice-inflicted lesion. The findings from this study could significantly aid in the development of new immune based anti-salmon louse prophylactic measures and treatments., (Crown Copyright © 2024. Published by Elsevier Ltd. All rights reserved.)

Vibrio parahaemolyticus is the main pathogen causing acute hepatopancreatic necrotic disease in crustaceans. To elucidate the epigenetic regulatory mechanism of crustacean resistance to V. parahaemolyticus infection, we conducted artificial infection studies on Portunus trituberculatus. The results showed that the mortality rate reached the highest at 12 h of artificial infection, which was 23.69 %. At 72 h after V parahaemolyticus infection, the expression level of DNA demethylase (ten-eleven-translocation protein) Tet was significantly decreased, the expression of DNA methyltransferase Dnmt3B fluctuated significantly. Based on the differential expression levels of Tet and Dnmt3B. We depict for DNA methylation profiles of the whole genome of P. trituberculatus at single-base resolution by using whole-genome bisulfite sequencing (WGBS) on hemolymph tissues. The overall DNA methylation level was low at 2.16 % in P. trituberculatus hemolymph. A total of 2590 differentially methylated regions (DMRs) were identified, of which 1329 were hypermethylated and 1261 were hypomethylated, and 1389 genes were annotated in these DMRs. Differently methylated genes (DMGs) were significantly enriched in ribosomes (KO03010), protein kinases (KO01001), cell cycle (HSA04110), endocrine resistance (HSA01522) and FoxO signaling pathway (KO04068). Finally, we selected six differentially methylated genes for quantitative analysis. The results showed that DNA methylation not only has a negative regulatory effect on gene expression, but also has a positive regulatory effect. These results indicated that DNA methylation in the regulation of genes involved in immune responses contributes to the resistance of P. trituberculatus to V. parahaemolyticus, which is valuable for understanding how crustaceans regulate the innate immune system to defend against bacterial infections., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

The white spot syndrome virus (WSSV) causes white spot disease (WSD), a severe condition in crustacean aquaculture, leading to significant economic losses. Our previous study demonstrated that C7 is an effective therapeutic agent against WSSV infection in aquaculture. It specifically blocked viral horizontal transmission and reduced shrimp mortality in a dose- and time-dependent manner. Here, we report the potential antiviral mechanism of C7 in shrimp. C7 regulated abnormal glycerophospholipid metabolism caused by WSSV and inhibited phosphatidylcholine (PC) synthesis by more than twofold, potentially enhancing shrimp resistance to viral infection. As the primary phospholipid in the cell membrane, PC is one of the main reactants in lipid peroxidation. Our results indicated that C7 significantly reduced the levels of lipid peroxidation products 4-hydroxynonenal (4-HNE) and malondialdehyde (MDA) induced by WSSV, whereas PC had the opposite effect. Accumulation of lipid peroxidation products inhibits stimulator of interferon genes (STING) signaling. Further evidence showed that C7 promoted STING transport from the endoplasmic reticulum to the Golgi apparatus, significantly activating the expression of the shrimp interferon analogue Vago4 gene. In contrast, PC suppressed Vago4 expression. Our results demonstrated that C7 inhibited PC synthesis, reduced the degree of lipid peroxidation, promoted STING translocation, activated Vago4 expression, and ultimately exerted antiviral effects. Therefore, C7 exhibits immunoregulatory activity as a preventative agent for enhancing the innate immunity of shrimp, making it potentially useful for future immunomodulatory approaches., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Chitin-binding proteins (CBPs) play pivotal roles in numerous biological processes in arthropods, including growth, molting, reproduction, and immune defense. However, their function in the antibacterial immune defense of crustaceans remains relatively underexplored. In this study, twenty CBPs were identified and characterized in Penaeus vannamei. Expression profiling highlighted that the majority of CBPs were highly expressed in the intestine and hepatopancreas and responded to challenge by Vibrio parahaemolyticus. To explore the role of these CBPs in innate immunity, six CBPs (PvPrg4, PvKrtap16, PvPT-1a, PvPT-1b, PvExtensin and PvCP-AM1159) were selected for RNAi experiments. Silencing of only PvPrg4 and PvKrtap16 significantly decreased the cumulative mortality of V. parahaemolyticus-infected shrimp. Further studies demonstrated that inhibition of PvPrg4 and PvKrtap16 resulted in a marked upregulation of genes associated with the NF-κB and JAK-STAT signaling pathways, as well as antimicrobial peptides (AMPs), in both the intestine and hepatopancreas. These results collectively suggested that PvPrg4 and PvKrtap16 potentially promote V. parahaemolyticus invasion by negatively regulating the JAK-STAT and NF-κB pathways, thereby inhibiting the expression of AMPs. In addition, SNP analysis identified three SNPs in the exons of PvPrg4 that were significantly associated with tolerance to V. parahaemolyticus. Taken together, these findings are expected to assist in the molecular marker-assisted breeding of P. vannamei associated with anti-V. parahaemolyticus traits, as well as expand our understanding of CBP functions within the immune regulatory system of crustaceans., Competing Interests: Declaration of competing interest The authors declare no financial and commercial conflicts of interest., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

The occurrence of black gill syndrome (BGS) is a serious threat to the healthy culture of Eriocheir sinensis. Studying the innate immune ability of E. sinensis with BGS can help develop new strategies for disease prevention and treatment. Antimicrobial peptides (AMPs) have crucial roles in crustacean humoral immunity. In this study, we found that the expression levels of two antilipopolysaccharide factor (EsALF7 and EsALF-L), one Toll receptor 3 (EsToll3), and one Pelle kinase (EsPelle) were upregulated in E. sinensis with BGS. Moreover, ALFs expressions in E. sinensis with BGS were positively regulated by EsToll3 and EsPelle. The content of hydrogen sulfide (H 2 S) in the gills of E. sinensis with BGS was increased. Further studies showed that the expressions of cystathionine β-synthase (EsCBS) and cystathionine γ-lyase (EsCSE) in the gills of E. sinensis with BGS were upregulated, which positively regulate the production of H 2 S. Whether there was a correlation between the upregulation of ALFs expression and changes in H 2 S content? Further studies showed that 1) the expressions of EsToll3, EsPelle, EsALF7, and EsALF-L in the gills of E. sinensis were upregulated under H 2 S exposure and 2) the knockdown of EsCBS and EsCSE in E. sinensis reduced the transcriptions of EsToll3, EsPelle, EsALF7, and EsALF-L. To sum up, these findings suggest that upregulation of H 2 S content induced by CBS/CSE promotes the expression of ALFs through the Toll pathway in E. sinensis suffering from BGS., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Hypoxia poses a significant challenge to aquatic organisms, especially Litopenaeus vannamei (L. vannamei), which play a vital role in the global aquaculture industry. Hypoxia-inducible factor 1α (HIF-1α) is a pivotal regulator of the organism's adaptation to hypoxic conditions. To understand of how HIF-1α affects the immunity of L. vannamei under hypoxic conditions, we conducted a thorough study involving various approaches. These included observing tissue morphology, analyzing the expression of immune-related genes, assessing the activities of immune-related enzymes, and exploring immune-related pathways. Our study revealed that RNA interference (RNAi)-mediated knockdown of HIF-1α markedly reduced HIF-1α expression in the gill (75-95 %), whereas the reduction ranged from 2 to 43 % in the hepatopancreas. Knockdown of HIF-1α resulted in increased damage to both gill and hepatopancreatic tissues in hypoxic conditions. Additionally, immune-related genes, including Astakine (AST), Hemocyanin (HC), and Ferritin (FT), as well as immune-related enzymes such as Acid Phosphatase (ACP), Alkaline Phosphatase (AKP), and Phenoloxidase (PO), exhibited intricate regulatory patterns in response to hypoxia stress following the knockdown of HIF-1α. Transcriptome analysis revealed that HIF-1α knockdown significantly impacts multiple signaling pathways, including the JAK-STAT signaling pathway, Th17 cell differentiation pathways, PI3K-Akt signaling pathway, ErbB signaling pathway, MAPK signaling pathway, chemokine signaling pathway, ribosomal pathways, apoptosis, lysosomes and arachidonic acid metabolism. These alterations disrupt the organism's immune balance and interfere with normal metabolic processes, potentially leading to various immune-related diseases. We speculate that the weakened immune response resulting from HIF-1 inhibition is due to the reduced metabolic capacity, and the existence of a direct regulatory relationship between them requires further exploration. This study greatly advances our understanding of the vital role that HIF-1α plays in regulating immune responses in shrimp under hypoxic conditions, thereby deepening our comprehension of this critical biological mechanism., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Eriocheir sinensis (Chinese mitten crab) is one of the main economic species in China, which has evolved an extremely sophisticated innate immune system to fend off disease invasions. However, bacterial and viral infections have caused significant financial losses for the E. sinensis aquaculture in recent years. Making well-informed judgments for the control microbial infections would require a thorough understanding and clarification of the intricate innate immune system of E. sinensis. Innate immunity is essential for the host's defense against invasive pathogens. Pattern recognition receptors (PRRs) initially recognize pathogen-associated molecular patterns (PAMPs) and trigger an innate immune response, causing the generation of inflammatory cytokine and promoting the clearance and control of pathogens. In E. sinensis, Toll/Toll-like receptors, lipopolysaccharide and β-1,3-glucan binding proteins, C-type lectins, galactoside-binding lectins, L-type lectins, scavenger receptors, and down syndrome cell adhesion molecules have been identified to be PRRs that are involved in the recognition of bacteria, fungi, and viruses. In this review, we give a comprehensive overview of the literature regarding PRRs' roles in the immunological defenses of E. sinensis, with the aim of providing clues to the mechanisms of innate immunity., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Hypoxic stress, triggered by a multitude of factors, has inflicted significant economic repercussions on the aquaculture of Eriocheir sinensis. In this research, we sequenced a collective of 60 samples from both hypoxia-sensitive and hypoxia-resistant groups utilizing streamlined genome sequencing techniques. Subsequently, we delved into population evolution, scrutinized the selective sweep within these populations, and performed a genome-wide association study (GWAS) focused on the hypoxia tolerance traits within the population, all through the lens of SNPs molecular markers. This comprehensive analysis aimed to uncover the SNPs and pinpoint the pertinent candidate genes that influence the hypoxia tolerance capabilities of E. sinensis. The selective sweep analysis revealed that genes harboring potential genetic variations within the two populations were predominantly enriched in areas such as signaling molecules and interactions, energy metabolism, glycolipid metabolism, and immune response. In the genome-wide association study focusing on hypoxia tolerance traits, we identified four SNPs significantly associated with hypoxia resistance. Furthermore, one potential candidate gene, Dscam2, which is believed to influence hypoxia tolerance, was discovered within a 50 kb vicinity of these SNPs. These identified SNPs can serve as molecular markers for screening hypoxia tolerance, offering valuable insights for the genetic improvement of E. sinensis., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

The microsporidian Enterocytozoon hepatopenaei (EHP) is an emerging pathogen that causes high economic losses in shrimp industry. The knowledge on shrimp's immune response to EHP infection to properly handle this outbreak is poorly understood. The prophenoloxidase (proPO)-activating system is an important invertebrate innate immune systems that produces melanin and toxic reactive intermediates against invading pathogens. In this study, we investigated the role of the proPO-activating system during EHP infection in the Pacific white shrimp Litopenaeus vannamei. The expression of proPO-activating system-related genes was highly responded to the EHP infection and the hemolymph PO activity was significantly increased and tightly regulated during the infection. The melanization products, generated by the proPO activation, exhibit in vitro cytotoxicity effect on the EHP spores and inhibit spore germination. Suppression of the proPO-activating system by RNA interference significantly decreased hemolymph PO activity and resulted in increased EHP copy number and reduced expression of several genes in the JAK/STAT and Toll signaling pathways as well as antimicrobial peptides. Furthermore, suppression of the proPO system also reduced hemocyte adhesion and encapsulation of the EHP spores. These results demonstrated that the proPO system plays a vital role in reducing EHP infectivity and cross-talks with other humoral and cellular responses to coordinately defend the EHP infection., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Forkhead box O (FOXO) proteins are a subgroup of the forkhead family of transcription factors that play important roles in the immune response. In this study, we cloned and identified a FOXO gene named MnFOXO from Macrobrachium nipponense. The full-length cDNA of MnFOXO is 2086 bp and contains a 1302 bp open reading frame, which encodes 433 amino acids. MnFOXO consists of five low-complexity regions and a conserved DNA-binding domain (forkhead domain). Evolutionary analyses indicate that MnFOXO proteins cluster with FOXO proteins from crustaceans. Tissue distribution analysis showed that MnFOXO was expressed in all detected tissues, with relatively higher expression levels in the intestine, eyestalks, stomach, and hemocytes than in the hepatopancreas, gills, and heart. The expression levels of MnFOXO in the hepatopancreas and intestine were significantly up-regulated in M. nipponense infected with white spot syndrome virus (WSSV) at 24 and 48 h. Furthermore, knockdown of MnFOXO increased the expression of WSSV envelope protein VP28 during WSSV infection. Further studies showed that knockdown of the MnFOXO gene in M. nipponense inhibited the synthesis of Dicers (MnDicer1, MnDicer2) and Argonautes (MnArgo1, MnArgo2) during WSSV invasion. These findings suggest that MnFOXO positively regulates the expression of Dicers and Argos, and inhibits the expression of VP28. This study provides new evidence for understanding the role of FOXO in antiviral innate immunity in crustaceans., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

The Penaeus vannamei aquaculture industry is facing a significant challenge in the form of hepatopancreatic microsporidiosis (HPM) caused by Enterocytozoon hepatopenaei (EHP), resulting in substantial economic losses. However, the extent of knowledge regarding the mechanisms by which shrimp resist EHP is limited. We screened resistant and susceptible shrimp and found that resistant shrimp had lower EHP load and less tissue damage. To gain insight into the molecular mechanisms underlying the EHP resistance of shrimp, a comparison was conducted at the transcriptional level between the resistant and susceptible families. Transcriptomic analysis of shrimp hepatopancreas revealed significant differences between the resistant and susceptible families. Compared to the susceptible family, the immune system of the resistant family was activated. The resistant family showed up-regulation in the expression of cathepsin L, C-type lectin, penaeidin, chitinase genes, and metabolism of xenobiotics by cytochrome P450-related genes. Additionally, the resistant shrimp exhibited a higher capacity for amino acid uptake. The observed differences in the resistant and susceptible family transcriptome may contribute to the shrimp's resistance to EHP., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Myeloid differentiation protein 2 (MD2), generally functions as a coreceptor of Toll-like receptor 4 (TLR4), facilitating the activation of TLR4 and the recognition of lipopolysaccharides (LPS) in host organisms. While the role of MD2 in immune activation is well-documented across various species, the specific role of the MD2 homolog in Scylla paramamosain (SpMD2) remains unidentified. In this study, we applied RNA interference to reduce SpMD2 expression, aiming to elucidate its role in immune system of mud crabs. Notably, SpMD2 interference leded to decrease in the hemocyte counts and phagocytic activity, along with increase in apoptosis rates and level of reactive oxygen species (ROS). Furthermore, the activities of key enzymes related to immune, such as superoxide dismutase (SOD), catalase (CAT), phenoloxidase (PO), peroxidase (POD), lysozyme (LZM), and acid phosphatase (ACP), were reduced by SpMD2 knockdown. Following infection with Vibrio alginolyticus, increase of SpMD2 expression level was observed. This was accompanied by alterations in the expression levels of genes related to immune in mud crabs. Challenge experiment with Vibrio alginolyticus showed a higher mortality rate after SpMD2 interference. Our study underscore the critical role of SpMD2 in enhancing the innate immunity and disease resistant in S. paramamosain, advancing our understanding of the innate immune regulatory mechanisms in crustaceans., Competing Interests: Declaration of competing interest The authors declared that they have no conflicts of interest to this work. We declare that we do not have any commercial or associative interest that represents a conflict of interest in connection with the work submitted., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Spiroplasma eriocheiris is a kind of intracellular pathogen without cell wall and the causative agent of Chinese mitten crab Eriocheir sinensis "tremor disease", which causes significant economic losses in the crustacean aquaculture. However, little is known about the intracellular transport of this pathogen and host innate immune response to this pathogen. Rab GTPases are key regulators for endocytosis and intracellular pathogen trafficking. In this study, we showed that S. eriocheiris infection upregulated the transcription of Rab7 through the downregulation of miR-131-3p. Subsequently, both hemocytes transfected with miR-131-3p mimics and hemocytes derived from Rab7 knockdown crabs exhibited reduced phagocytic activities and increased susceptibility to S. eriocheiris infection. Additionally, Rab7 could interact with the cell shape-determining protein MreB3 of S. eriocheiris, and its overexpression promoted S. eriocheiris internalization and fusion with lysosomes, thereby limiting S. eriocheiris replication in Drosophila S2 cells. Overall, these results demonstrated that Rab7 facilitated host cell phagocytosis and interacted with MreB3 of S. eriocheiris to prevent S. eriocheiris infection. Moreover, miR-131-3p was identified as a negative regulator of this process through its targeting of Rab7. Therefore, targeting miR-131-3p might be an effective strategy for controlling S. eriocheiris in crab aquaculture., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper. The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Qingguo Meng reports financial support was provided by Jiangsu Provincial Department of Agriculture and Rural Affairs. If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Scylla paramamosain, an economically significant crab, is widely cultivated worldwide. In recent years, S. paramamosain has faced a serious threat from viral diseases due to the expansion of culture scale and increased culture density. Among these, mud crab dicistrovirus-1 (MCDV-1) stands out as highly pathogenic, presenting substantial challenges to the healthy development of mud crab aquaculture. Therefore, a comprehensive understanding of the mud crab immune response to MCDV-1 infection is imperative for devising effective disease prevention strategies. In this study, transcriptomic analyses were conducted on the hepatopancreas of mud crabs infected with MCDV-1. The findings revealed a total of 5139 differentially expressed genes (DEGs) between healthy and MCDV-1 infected mud crabs, including 3327 upregulated and 1812 downregulated DEGs. Further analysis showed that mud crabs resist MCDV-1 infection by activating humoral immune-related pathways, including the MAPK signaling pathway, MAPK signaling pathway-fly, and Toll and Imd signaling pathway. In contrast, MCDV-1 infection triggers host metabolic disorders. Several immune-related vitamin metabolism pathways (ascorbate and aldarate metabolism, retinol metabolism, and nicotinate and nicotinamide metabolism) were significantly inhibited, which may create favorable conditions for the virus's self-replication. Notably, endocytosis emerged as significantly upregulated both in GO terms and KEGG pathways, with several viral endocytosis-related pathways showing significant activation. PPI network analysis identified 9 hub genes associated with viral endocytosis within the endocytosis. Subsequent GeneMANIA analysis confirmed the association of these hub genes with viral endocytosis. Both transcriptome data and qPCR analysis revealed a significant upregulation of these hub genes post MCDV-1 infection, suggesting MCDV-1 may use viral endocytosis to enter cells and facilitate replication. This study represents the first comprehensive report on the transcriptomic profile of mud crab hepatopancreas response to MCDV-1 infection. Future investigations should focus on elucidating the mechanisms through which MCDV-1 enters cells via endocytosis, as this may holds critical implications for the development of vaccine targets., Competing Interests: Declaration of competing interest The authors declare that they have no competing interests., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Ecdysis-triggering hormone (ETH) is a neuropeptide hormone characterized by a conserved KxxKxxPRx amide structure widely identified in arthropods. While its involvement in the regulation of molting and reproduction in insects is well-established, its role in crustaceans has been overlooked. This study aimed to de-orphanise a receptor for ETH in the mud crab Scylla paramamosain and explore its potential impact on ovarian development. A 513-amino-acid G protein-coupled receptor for ETH (SpETHR) was identified in S. paramamosain, exhibiting a dose-dependent activation by SpETH with an EC 50 value of 75.18 nM. Tissue distribution analysis revealed SpETH was in the cerebral ganglion and thoracic ganglion, while SpETHR was specifically expressed in the ovary, hepatopancreas, and Y-organ of female crabs. In vitro experiments demonstrated that synthetic SpETH (at a concentration of 10 -8  M) significantly increased the expression of SpVgR in the ovary and induced ecdysone biosynthesis in the Y-organ. In vivo experiments showed a significant upregulation of SpEcR in the ovary and Disembodied and Shadow in the Y-organ after 12 h of SpETH injection. Furthermore, a 16-day administration of SpETH significantly increased 20E titers in hemolymph, gonadosomatic index (GSI) and oocyte size of S. paramamosain. In conclusion, our findings suggest that SpETH may play stimulatory roles in ovarian development and ecdysone biosynthesis by the Y-organ., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier Inc.)

The immune system of ticks, along with that of other invertebrates, is comparatively simpler than that of vertebrates, relying solely on innate immune responses. Direct antimicrobial defence is provided by the synthesis of antimicrobial peptides (AMPs), including defensins. The aim of this study was to investigate the differences in defensin genes expression between questing and engorged Ixodes ricinus (def1 and def2) and Dermacentor reticulatus (defDr) ticks, in the presence of selected pathogens: Borrelia spp., Rickettsia spp., Babesia spp., Anaplasma phagocytophilum, and Neoehrlichia mikurensis in the natural environment. After pathogen screening by PCR/qPCR, the expression of defensin genes in pathogen positive ticks and ticks without any of the tested pathogens, was analysed by reverse transcription qPCR. The results showed an increased expression of defensin genes in I. ricinus ticks after blood feeding and I. ricinus and D. reticulatus ticks during in cases of co-infection. In particular, the expression of defensins genes was higher in questing D. reticulatus than in questing and engorged I. ricinus ticks, when borreliae were detected. This study contributes to uncovering the expression patterns of defensin genes in the presence of several known tick pathogens, the occurrence of these pathogens and possible regulatory mechanisms of defensins in tick vector competence., Competing Interests: Declaration of competing interest The authors declare no competing interests., (Copyright © 2024 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Objective: Der f 2, a major allergen derived from Dermatophagoides farinae , is a leading cause of allergic asthma. IL-6 and GM-CSF play essential roles in the exacerbation of asthma. However, the mechanical act by which Der f 2 mediates the expression of IL-6, IL-8, and GM-CSF in airway epithelial cells remains incompletely elucidated. Herein, we aimed to explore the effect of Der f 2 on IL-6 and GM-CSF expression in the human airway epithelial cell BEAS-2B and A549., Methods: Recombinant Der f 2 (rDf2) was acquired using Pichia pastoris . BEAS-2B and A549 cells were used as cell model. The expression of genes and proteins and the involvement of the signaling cascade were assessed using RT-PCR, quantitative real-time PCR (qPCR), Western blotting, and ELISA, respectively., Results: Our findings showed that rDf2 significantly induced mRNA expression and protein production of IL-6 and GM-CSF in BEAS-2B and A549 cells. In contrast, rDf2 did not influence IL-8 expression or production in both cells. Mechanistic studies revealed that rDf2 triggered activation of the p38 MAPK and JNK. Inhibition of p38, but not JNK, significantly attenuated rDf2-induced IL-6 and GM-CSF expression and production., Conclusion: This study demonstrates that Der f 2 promotes the expression and production of the pro-inflammatory cytokines IL-6 and GM-CSF in airway epithelial cells via activation of the p38 signaling pathway. These findings provide insights into the molecular mechanisms that Der f 2 may exacerbate airway inflammation.

Trehalase plays an important role in insect metabolism and development by hydrolyzing blood sugar trehalose, but it seems to perform primarily an immunomodulatory function in crustaceans whose blood sugar is glucose. Metal ions as pollutants seriously affecting crustacean health, but studies on trehalase in metal immunity are still limited. In this study, a soluble trehalase (NdTre1) that could bind to multiple metals was identified from Neocaridina denticulata sinensis for investigating metal resistance. Expression profiling revealed that NdTre1 was mainly expressed in the gill and was significantly decreased following stimulation with copper (Cu²⁺) and cadmium (Cd²⁺). Transcriptomic analysis of gills revealed an increase in ecdysone synthesis after interference with NdTre1. Increased ecdysone activated the endogenous mitochondrial pathway and the mitogen activated protein kinase (MAPK) pathway to further induced apoptosis. In vitro, Escherichia coli overexpressing recombinant NdTre1 had higher survival and faster growth rates to better adapted the metal-containing medium. Overall, NdTre1 exercises an important immune function in shrimp resistance to metal stimulation by regulating apoptosis and molting. Further investigation can further explore specific response mechanisms of NdTre1 to multiple metals., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

The insulin receptor (IR) plays a crucial role in the growth and metabolism of animals. However, there are still many questions regarding the IR in crustaceans, particularly their role in shrimp growth and glucose metabolism. In this study, we identified a novel insulin-like receptor gene in Litopenaeus vannamei and cloned its full length of 6439 bp. This gene exhibited a highly conserved sequence and structural characteristics. Phylogenetic analysis confirmed it as an unreported RTK2-type IR, namely, LvRTK2 . Expression pattern analysis showed that LvRTK2 is primarily expressed in female reproductive and digestive organs. Through a series of in vivo and in vitro experiments, including glucose treatment, exogenous insulin treatment, and starvation treatment, LvRTK2 was confirmed to be involved in the endogenous glucose metabolic pathway of shrimp under different glucose variations. Moreover, long-term and short-term interference experiments with LvRTK2 revealed that the interference significantly reduced the shrimp growth rate and serum glucose clearance rate. Further studies indicated that LvRTK2 may regulate shrimp growth by modulating the downstream PI3K/AKT signaling pathway and a series of glucose metabolism events, such as glycolysis, gluconeogenesis, glycogen synthesis, and glycogenolysis. This report on the characteristics and functions of LvRTK2 confirms the important role of RTK2-type IRs in regulating shrimp growth and glucose metabolism.

Horseshoe crabs are living fossils. In recent decades, the population of horseshoe crabs, especially the tri-spine horseshoe crab Tachypleus tridentatus, has decreased significantly and was listed as an 'endangered species' under the IUCN Red List in 2019. In order to improve the reproduction of T. tridentatus to facilitate stock enhancement, it is important to understand their ovarian development. In this study, a novel TtVtg2-like gene from T. tridentatus was cloned and functionally characterized. The total legth of TtVtg2-like was 5469 bp, encoding a protein consisting of 1822 amino acid with a pI value of 6.51 and a molecular weight of 208.68 KDa. The TtVtg2-like was highly expressed in the ovary and yellow connective tissues, mainly localized in cytoplasm and endoplasmic reticulum vesicles of oocytes and yellow connective tissues, respectively. RNA interference of TtVtg2-like caused the accumulation of ROS, DNA damage, and apoptosis of ovarian primary cells. The results of this study provide useful baseline information for future studies on ovarian development in horseshoe crabs., Competing Interests: Declaration of competing interest The authors declared that there is no conflict of interest in this study. All authors have seen the manuscript and approve it to submit., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Cuticle proteins (CPs) are the vital components of the cuticle and chitin lining covering the digestive tract of crustaceans. In this study, four new CP genes (designated as EsCP3, EsCP4, EsCP5, and EsCP8) were initially cloned and identified from the Chinese mitten crab Eriocheir sinensis. EsCP3/4/5/8 included 375, 411, 381, and 570 bp open reading frame encoding 124, 136, 126, and 189 amino acid proteins, respectively. Except for EsCP8, EsCP3/4/5 all contained a Chitin_bind_4 domain. EsCP3/4/5/8 were clustered into different groups in the phylogenetic tree. Quantitative real-time PCR results indicated that four EsCP genes have different patterns of tissue distribution. Changes in the expression levels of these four EsCP genes were observed in the intestine of crabs under Vibrio parahaemolyticus challenge. RNA interference assay showed that the knockdown of EsCPs in the intestine could inhibit the expression of antimicrobial peptides (AMPs), including crustins and anti-lipopolysaccharide factors. In addition, the knockdown of EsRelish in the intestine decreased the expression levels of these four EsCP genes. These results indicated that EsCPs were involved in regulating the expression of AMPs, and EsCPs were regulated by EsRelish., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Nitrite is a common environmental pollutant in intensive aquaculture systems. In this study, physiological and transcriptomic analyses were performed to investigate nitrite stress responses in the swimming crab Portunus trituberculatus, an important aquaculture species in China. The results revealed that nitrite can affect neurotransmitter signaling via the expression of neurotransmitter receptors such as octopamine receptor (OAR) and 5-hydroxytryptamine receptor (5-HTR), and depress ecdysteroid signaling by downregulating ecdysteroid receptor (EcR) as well as its downstream transcription factors in hepatopancreas. In addition, nitrite suppressed the expression of hemocyanins, the oxygen-transporting protein, which at least partly contributed to tissue hypoxia, resulting in a switchover of energy metabolism from aerobic to anaerobic pathway. To meet the energy demand, glycogens and lipids were mobilized and transported to the hemolymph, and the catabolism of amino acids and fatty acids was enhanced to provide energy for hepatopancreas. β-oxidation of fatty acids, the major process by which fatty acids are oxidized to generate energy, seems to occur mainly not in mitochondria but in peroxisomes. Although the cellular protective mechanisms, including antioxidant defense, heat shock response (HSR), unfolded protein response (UPR), and autophagy, were activated, nitrite-induced cellular stress overwhelmed the repairing capacity and caused significant increase in the levels of apoptosis. These results indicated that nitrite stress influences neurotransmitter and endocrine signaling, disturbs energy metabolism, damages cellular components, and induces apoptosis in P. trituberculatus. The findings of this study provide new insights into nitrite stress response in the swimming crab and provide valuable information for aquaculture management of this species., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Lectins are versatile proteins that specifically recognize and interact with sugar moieties expressed on the cell surface. The potential of lectin in drug targeting and delivery has instigated interest to identify natural lectins. Crabs have been identified as a rich source of lectin because the innate immune system is activated on encounter of pathogens and helps in the production of lectin. Although the presence of lectins in crab's hemolymph is well documented, little information about lectin in hepatopancreas, a vital organ for immunity and digestion in crustaceans, is currently available. A calcium dependent lectin (75 kDa) was purified from the hepatopancreas of the freshwater crab Oziotelphusa naga by bioadsorption and fetuin linked Sepharose 4B affinity chromatography technique. The isolated hepatopancreas lectin is calcium dependent and maximum agglutination was observed with rabbit erythrocytes. The hemagglutinating activity of the hepatopancreas lectin was effectively inhibited by sugars, such as α-lactose, GlcNAc, trehalose and NeuAc. Compared to sialylated N-glycosylated proteins including transferrin and apo transferrin, sialylated O-glycosylated proteins like fetuin exhibited stronger inhibitory effect. The ability of erythrocytes to bind hepatopancreas lectin has been diminished by desialylation of the potent inhibitor, indicating the significance of sialic acid in lectin-ligand interactions. The purified hepatopancreas lectin showed a broad spectrum of antimicrobial activity against bacteria Staphylococcus aureus, Klebsiella pneumoniae, Proteus mirabilis, Pseudomonas aeruginosa, E. coli and fungi Candida albicans and Aspergillus niger. The findings of this study demonstrate the significance of hepatopancreas lectin as a multifunctional defense protein that inhibits the growth of bacteria and fungi., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Background: Dermanyssus gallinae, the poultry red mite (PRM), is a worldwide ectoparasite posing significant economic challenges in poultry farming. The extensive use of pyrethroids for PRM control has led to the emergence of pyrethroid resistance. The objective of this study is to detect the pyrethroid resistance and explore its associated point mutations in the voltage-gated sodium channel (VGSC) gene among PRM populations in China., Results: Several populations of D. gallinae, namely CJF-1, CJP-2, CJP-3, CSD-4 and CLD-5, displayed varying degrees of resistance to beta-cypermethrin compared to a susceptible field population (CBP-5). Mutations of VGSC gene in populations of PRMs associated with pyrethroid resistance were identified through sequencing its fragments IIS4-IIS5 and IIIS6. The mutations I917V, M918T/L, A924G and L925V were present in multiple populations, while no mutations were found at positions T929, I936, F1534 and F1538., Conclusion: The present study confirmed the presence of extremely high levels of pyrethroid resistance in PRM populations in China, and for the first time detected four pyrethroid resistance mutations in the VGSC gene. Identifying pyrethroid resistance in the field population of PRM in China can be achieved through screening for VGSC gene mutations as an early detection method. Our findings underscore the importance of implementing chemical PRM control strategies based on resistance evidence, while also considering the management of acaricide resistance in the control of PRMs. © 2024 Society of Chemical Industry., (© 2024 Society of Chemical Industry.)

The muscle LIM protein (MLP) is a member of the cysteine and glycine-rich protein (CSRP) family, composed of CSRP1, CSRP2 and CSRP3/MLP. MLP is involved in a multitude of functional roles, including cytoskeletal organization, transcriptional regulation, and signal transduction. However, the molecular mechanisms underlying its involvement in immune and stress responses remain to be elucidated. This study identified an MnMLP in the freshwater crustacean Macrobrachium nipponense. The isothermal titration calorimetry assay demonstrated that recombinant MnMLP was capable of coordinating with Zn 2+ . Upon challenge by Aeromonas veronii or WSSV, and exposure to CdCl 2 , up-regulation was recorded in the muscle and intestinal tissues, suggesting its involvement in immune and anti-stress responses. MnMLP protein was predominantly expressed in the cytoplasm of the transfected HEK-293T cells, but after treatment with LPS, Cd 2+ or H 2 O 2 , the MnMLP was observed to be transferred into the nucleus. The comet assay demonstrated that the overexpression of MnMLP could mitigate the DNA damage induced by H 2 O 2 in HEK-293T cells, suggesting the potential involvement of MnMLP in the DNA repair process. These findings suggest that DNA repair may represent a possible mechanism by which MnMLP may be involved in the host's defense against pathogens and stress., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Antimicrobial peptides (AMPs) are crucial in the humoral immunity aspect of invertebrates' innate immune systems. However, studies on AMP discovery in the Pacific white shrimp (Litopenaeus vannamei) using omics data have been limited. Addressing the growing concern of antibiotic resistance in aquaculture, this study focused on the identification and characterization of AMPs in L. vannamei using advanced genomic and transcriptomic techniques. The genome of L. vannamei was performed to predict and identify a total of 754 AMP-derived genes, distributed across most chromosomes and spanning 24 distinct AMP families, and further identified 236 AMP-derived genes at the mRNA level in hemocytes. A subset of 20 chemically synthesized peptides, derived from these genes, exhibited significant antimicrobial activity, with over 85% showing effectiveness against key bacterial strains such as Staphylococcus aureus and Vibrio parahaemolyticus. The expression patterns of these AMPs were also investigated in different shrimp tissues and at various infection stages, revealing dynamic responses to pathogenic challenges. These findings highlight the significant potential of AMPs in L. vannamei as novel, effective alternatives to traditional antibiotics in aquaculture, offering insights into their diverse structural properties and biological functions. Together, this comprehensive characterization of the AMP repertoire in L. vannamei demonstrates the efficacy of using omics data for AMP discovery and lays the groundwork for their potential applications., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

The mannose receptor (MR) plays a key role in the innate immune system as a pattern recognition receptor. Here, a novel type of mannose receptor, named PvMR2, was identified from Penaeus vannamei (P. vannamei). The PvMR2 coding sequence (CDS) obtained was 988 base pairs in length, encoding a protein consisting of 328 amino acids. This protein includes a signal peptide and two classical C-type lectin domains (CTLD). Quantitative real-time PCR showed that PvMR2 was distributed in all detected tissues, with the highest levels in the intestines and stomach. Following a bacterial challenge with Vibrio anguillarum (V. anguillarum), PvMR2 showed significant up-regulation in both the intestines and stomach of shrimp. To validate the function of PvMR2, recombinant proteins were extracted and purified using a His-tag. The resulting rPvMR2 demonstrated binding capability with lipopolysaccharides (LPS) and peptidoglycan (PGN) in a dose-dependent manner, affirming its binding affinity. The purified rPvMR2 demonstrated calcium-independent binding activity towards both Gram-positive bacteria (V. anguilliarum and Vibrio parahaemolyticus) and Gram-negative bacteria (Escherichia coli and Aeromonas Veronii). Antibacterial assays confirmed that rPvMR2 inhibits bacterial growth. Intestinal adhesion and adhesion inhibition experiments confirmed that the rPvMR2 can be used to reduce the adhesion capacity of harmful bacteria in the gut. Phagocytosis experiments have shown that rPvMR2 promotes phagocytosis in hemocytes and protects the host from external infection. Treatment with recombinant PvMR2 significantly bolstered bacterial clearance within the hemolymph and markedly augmented shrimp survival post-infection with V. anguillarum. These results suggest that PvMR2 has agglutination, growth inhibition, adhesion inhibition, clearance promotion, and phagocytosis effects on harmful bacteria, and plays a crucial role in the antimicrobial immune response of P. vannamei., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Mannose-binding lectin (MBL) is a vital member of the lectin family, crucial for mediating functions within the complement lectin pathway. In this study, following the cloning of the mannose-binding lectin (MBL) gene in the ridgetail white prawn, Exopalaemon carinicauda, we examined its expression patterns across various tissues and its role in combating challenges posed by Vibrio parahaemolyticus. The results revealed that the MBL gene spans 1342 bp, featuring an open reading frame of 972 bp. It encodes a protein comprising 323 amino acids, with a predicted relative molecular weight of 36 kDa and a theoretical isoelectric point of 6.18. The gene exhibited expression across various tissues including the eyestalk, heart, gill, hepatopancreas, stomach, intestine, ventral nerve cord, muscle, and hemolymph, with the highest expression detected in the hepatopancreas. Upon challenge with V. parahaemolyticus, RT-PCR analysis revealed a trend of MBL expression in hepatopancreatic tissues, characterized by an initial increase followed by a subsequent decrease, peaking at 24 h post-infection. Employing RNA interference to disrupt MBL gene expression resulted in a significant increase in mortality rates among individuals challenged with V. parahaemolyticus. Furthermore, we successfully generated the Pet32a-MBL recombinant protein through the construction of a prokaryotic expression vector for conducting in vitro bacterial inhibition assays, which demonstrated the inhibitory effect of the recombinant protein on V. parahaemolyticus, laying a foundation for further exploration into its immune mechanism in response to V. parahaemolyticus challenges., Competing Interests: Declaration of competing interest The authors declare that they have no competing interests., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Background: A GPCR (G protein-coupled receptor) target-based approach was applied to identify antagonists of the arthropod-specific tick kinin receptor. These small molecules were expected to reproduce the detrimental phenotypic effects that had been observed in Rhipicephalus microplus females when the kinin receptor was silenced by RNA interference. Rhipicephalus microplus, the southern cattle tick, cattle fever tick, or Asian blue tick, is the vector of pathogenic microorganisms causing the deadly bovine babesiosis and anaplasmosis. The widespread resistance to acaricides in tick populations worldwide emphasizes that exploring novel targets for effective tick control is imperative., Results: Fifty-three structural analogs of previously identified tick kinin antagonists were screened in a 'dual-addition' calcium fluorescence assay using a CHO-K1 cell line expressing the tick kinin receptor. Seven molecules were validated as non-cytotoxic antagonists, four of which were partial (SACC-0428764, SACC-0428780, SACC-0428800, and SACC-0428803), and three were full antagonists (SACC-0428799, SACC-0428801, and SACC-0428815). Four of these antagonists (SACC-0428764, SACC-0428780, SACC-0428799, and SACC-0428815) also inhibited the tick midgut contractions induced by the myotropic kinin agonist analog 1728, verifying their antagonistic bioactivity. The small molecules were tested on recombinant human neurokinin (NK) receptors, the one most similar to the invertebrate kinin receptors. Most molecules were inhibitors of the NK1 receptor, except SACC-0412066, a previously identified tick kinin receptor antagonist, which inhibited the NK1 receptor only at the highest concentration tested (25 μm). None of the molecules inhibited the NK3 human receptor., Conclusion: Molecules identified through this approach could be useful probes for studying the tick kinin signaling system and midgut physiology. © 2024 The Author(s). Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry., (© 2024 The Author(s). Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.)

Molting is a crucial biological process of crustaceans. Crustaceans go through three separate stages throughout their molting process, including pre-molt, post-molt and inter-molt. However, the exact mechanism of immunological modulation during molting remains unclear. Tumor necrosis factor receptor-associated factor 6 (TRAF6) has been extensively documented to participate in immune defense. In the present study, a TRAF6 gene with two TRAF-type zinc finger domains was identified from Eriocheir sinensis (designed as EsTRAF6), and its role in regulating immune response during molting process was explored. The mRNA expression level of EsTRAF6 at pre-molt stage was higher than that at post-molt stage and inter-molt stage. After Aeromonas hydrophila stimulation, the expression levels of EsTRAF6, EsRelish and anti-lipopolysaccharide factors (ALFs) genes exhibited a considerable increase at three molting stages. Subsequently, the expression patterns of EsTRAF6 and EsRelish in response to the treatment with 20-hydroxyecdysone (20E) were examined. The mRNA expression of EsTRAF6 and EsRelish were significantly increased at 12 h after 20E injection. Additionally, the protein expression level of TRAF6 was also up-regulated in 20E group compared to control group. Furthermore, the role of EsTRAF6 in regulating the anti- ALFs expression at pre-molt stage post A. hydrophila stimulation was investigated. Following the inhibition of the EsTRAF6 transcript using RNAi or the injection of inhibitor (TMBPS), there was a notable decrease of the EsALF1, EsALF2 and EsALF3 transcripts. Moreover, a significant reduction in the phosphorylation level of NF-κB at pre-molt stage was observed after A. hydrophila stimulation in TRAF6-inhibited crabs. Collectively, our results suggest that EsTRAF6 could be induced by 20E and promoted the EsALFs expression by activating NF-κB at pre-molt stage, which provides a novel insight into the research of immune regulatory mechanism during the process of molting of crustaceans., Competing Interests: Declaration of competing interest The authors declare no competing interest, (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Research has shown that voraxin α derived from male ticks stimulates blood feeding to engorge in female ticks. Whereas, the oviposition rate, egg weight, and body weight of female ticks were reduced in animals vaccinated with recombinant (r-) voraxin α. These data suggest a potential role of r-voraxin α as a functional anti-tick antigen in Rhipicephalus appendiculatus and Amblyomma hebraeum tick infestation. This study investigated the immunogenicity of r-voraxin α protein from Hyalomma anatolicum (H. anatolicum) tick as an anti-tick vaccine in rabbits. The H. anatolicum voraxin α sequence was optimized according to the codon usage in E. coli before being sub-cloned into pQE30. The gene sequence of the voraxin α was synthesized, verified by DNA sequencing, cloned in a pQE30 vector, and transformed into E. coli. Then, the expression of the r-voraxin α protein was confirmed by SDS-PAGE and Western blot analysis. Subsequently, three rabbits were immunized with the r-voraxin α as the vaccinated group, whereas three rabbits without injection were considered the control group. The result indicated the success of cloning of codon-optimized H. anatolicum voraxin α gene. Moreover, the expression of the r-voraxin α protein (approximately 18 kDa) in the bacterial expression system was confirmed by SDS-PAGE and Western blot analysis. The results of this study showed that the mortality rate in vaccine recipients increased compared to the control group (P < 0.01). Also, the egg weight, oviposition rate, and engorgement weight of female ticks fed from vaccinated animals were significantly reduced compared to the control group (P < 0.01). The results confirmed that the codon-optimized H. anatolicum voraxin α gene expressed in the bacterial expression system could be a suitable anti-tick vaccine against H. anatolicum tick infestation., Competing Interests: Declaration of Competing Interest The authors confirm that there are no known conflicts of interest associated with this publication and there has been no significant financial support for this work that could have influenced its outcome., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Ticks are notable vectors of diseases affecting both humans and animals, with Hyalomma anatolicum being of particular significance due to its wide distribution and capability to transmit a variety of pathogens, including Theileriaannulata and Crimean-Congo haemorrhagic fever virus. This study aimed to investigate the inhibitory effects of H. anatolicum salivary gland extract (HaSGE) and the identification of its key component on the complement system of the host's innate immune defense. We demonstrated that HaSGE exerts a dose-dependent inhibition on the complement activation in a host-specific manner. Mechanistic studies revealed that HaSGE interferes with deposition and cleavage of complement proteins C3 and C5, thus preventing the formation of the membrane attack complex. Further, we identified a serine protease inhibitor, Hyalomma anatolicum serpin-1 (HAMpin-1), from the HaSGE through proteomic analysis and characterized its structure, function, and interaction with complement proteins. HAMpin-1 exhibited potent inhibitory activity against chymotrypsin and cathepsin-G, and notably, it is the first serpin from ticks shown to inhibit the classical and lectin pathways of the complement system. The expression of HAMpin-1 was highest in the salivary glands, suggesting its crucial role in blood feeding and immune evasion. Our findings revealed one of the potential mechanisms used by H. anatolicum to modulate host immune responses at the interface, offering new insights into tick-host interactions., Competing Interests: Conflict of interest The authors declare that they have no conflicts of interest with the contents of this article., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Sinopotamon Henanense expresses two metal‒induced metallothioneins (MTs), Cd‒induced MT and Cu‒induced MT (ShCuMT). The Cd‒induced MT has been characterized as a Cd‒thiolate MT. However, it is unknown whether ShCuMT is a Cu‒thiolate MT. In the present study, ShCuMT was expressed heterologously in Escherichia coli and purified by Ni‒NTA column and superdex‒75 column. And its metal‒binding feature was evaluated by DTNB reaction, circular dichroism spectroscopy (CD), isothermal microtitration (ITC), electrospray flight mass spectrometry (ESI‒TOF‒MS), and matrix‒assisted laser desorption ionization flight mass spectrometry (MALDI‒TOF‒MS). Bioinformatics analysis demonstrated that ShCuMT possessed the cysteine‒triplet motif of a Cu‒specific MT. Expression and purification of ShCuMT illustrated that SUMO tag used as the production system for ShCuMT resulted in a high production yield. The stability order of ShCuMT binding metal ions were Cu (Ⅰ) > Cd (Ⅱ) > Zn (Ⅱ). The CD spectrum indicated that ShCuMT binding with Cu (I) exhibited a compact thiol metal clusters structure. Besides, there emerged no a visible nickel‒thiol absorption after Ni‒NTA column affinity chromatography. The ITC results implied that Cu‒ShCuMT possessed the optimal thermodynamic conformation and the highest stoichiometric number of Cu (Ⅰ). Overall, the results suggested that SUMO fusion system is a robust and inexpensive approach for ShCuMT expression and Ni‒NTA column had no influence on metal binding of ShCuMT and Cu(Ⅰ) was considered its cognate metal ion, and ShCuMT possessed canonical Cu‒thiolate characteristics. The metal binding feature of ShCuMT reported here contributes to elucidating the structure‒function relationship of ShCuMT in S. Henanense., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Lysozymes are hydrolytic enzymes, and they are ubiquitous among all living organisms. They are mostly associated with antibacterial properties through their muramidase activity, while other properties such as iso-peptidase activity are also common. Invertebrate-type (i-type) lysozymes include the enzyme Destabilase, which is present in the salivary secretions of the medicinal leach Hirundo medicinalis. Destabilase has the ability to hydrolyse the ε-(γ-glutamyl)-lysine iso-peptide bonds formed by transglutaminase in fibrin of vertebrate blood, thereby destabilising blood clots. We have identified an i-type lysozyme from the hemocytes of the freshwater crayfish Pacifastacus leniusculus, which was found to be upregulated at the protein level in response to an injection of the β-1,3-glucan laminarin. Based on its sequence we predicted that this lysozyme would lack muramidase activity, and therefore we decided to determine its putative immune function. The P. leniusculus i-type lysozyme (Pl-ilys), is a protein with 159 amino acid residues, including a 29 residue signal peptide, with a predicted molecular weight of 16 kDa and a predicted pI of 5.6. It is expressed primarily in the hemocytes and to a lesser extent in the hematopoietic tissue. A recombinant mature Pl-ilys using an E. coli expression system was produced, and we could ascertain that this enzyme was deficient of muramidase activity. Moreover, no iso-peptidase activity could be detected against the substrate l-γ-glutamine-p-nitroanilide. Analysis of the conserved domains in Pl-ilys showed a putative destabilase domain, and thus we tested the clot dissolving activity of this enzyme. We could show that the purified P. leniusculus clotting protein which had been coagulated and clotted with transglutaminase was dissolved by the addition of Pl-ilys. Taken together our results indicate that Pl-ilys has a clot dissolving or destabilising activity in crustacean blood., (Copyright © 2024 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Background: Tetranychus cinnabarinus is one of the most common polyphagous arthropod herbivores, and is primarily controlled by the application of acaricides. The heavy use of acaricides has led to high levels of resistance to acaricides such as cyflumetofen, which poses a threat to global resistance management programs. Cyflumetofen resistance is caused by an increase in metabolic detoxification; however, the role of uridine diphosphate (UDP)-glycosyltransferase (UGT) genes in cyflumetofen resistance remains to be determined., Results: Synergist 5-nitrouracil (5-Nul) significantly enhanced cyflumetofen toxicity in T. cinnabarinus, which indicated that UGTs are involved in the development of cyflumetofen resistance. Transcriptomic analysis and quantitative (q)PCR assays demonstrated that the UGT genes, especially UGT201H1, were highly expressed in the YN-CyR strain, compared to those of the YN-S strain. The RNA interference (RNAi)-mediated knockdown of UGT201H1 expression diminished the levels of cyflumetofen resistance in YN-CyR mites. The findings additionally revealed that the recombinant UGT201H1 protein plays a role in metabolizing cyflumetofen. Our results also suggested that the aromatic hydrocarbon receptor (AhR) probably regulates the overexpression of the UGT201H1 detoxification gene., Conclusion: UGT201H1 is involved in cyflumetofen resistance, and AhR may regulates the overexpression of UGT201H1. These findings provide deeper insights into the molecular mechanisms underlying UGT-mediated metabolic resistance to chemical insecticides. © 2024 Society of Chemical Industry., (© 2024 Society of Chemical Industry.)

Decapod iridescent virus 1 (DIV1) stands as a significant pathogen affecting crustaceans, posing a grave threat to the shrimp industries in aquaculture dependent nations. Within the Iridoviridae family, the conserved envelope protein DIV1-168L plays a pivotal role in virion entry. Nonetheless, the host factors that interact with 168L remain unidentified. To address this gap, we established a cDNA library derived from Litopenaeus vannamei gill tissue and conducted yeast two-hybrid screening to identify host factors that interact with 168L. Additionally, we performed co-immunoprecipitation assays to verify the interaction between cuticle protein 8 (CP8) and 168L. Expression pattern analysis revealed the presence of CP8 transcripts in the gill and epidermis. Furthermore, immunohistochemistry results demonstrated the expression of CP8 in gill cells and its localization in the gill filament epithelium. Fluorescence analysis indicated that full-length CP8 colocalized with 168L in the cytoplasm of Sf9 cells. Removal of the signal peptide from the N-terminal of CP8 eliminated its concentration in the cytoplasm. Additionally, CP8 expression was significantly inhibited during DIV1 infection. Therefore, our research contributes to a better understanding of the entry mechanism of iridovirids. The GenBank accession number for the DIV1 sequence is MF197913.1., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Inc. All rights reserved.)

LRR-only protein (LRRop) is an important class of immune molecules that function as pattern recognition receptor in invertebrates, however, the bacterial inhibitory activity of this proteins remain largely unknown. Herein, a novel LRRop was cloned from Eriocheir sinensis and named as EsLRRop2. The EsLRRop2 consists of six LRR motifs and formed a horseshoe shape three-dimension structure. EsLRRop2 was mainly expressed in intestine and hepatopancreas. The transcripts of EsLRRop2 in the intestine and hepatopancreas were induced by Vibrio parahaemolyticus and Staphylococcus aureus, and displayed similar transcriptional profiles. The expression levels of EsLRRop2 responded more rapidly and highly to V. parahaemolyticus than S. aureus in the intestine and hepatopancreas. Although the basal expression level of EsLRRop2 in hemocytes was relatively low, its transcripts in hemocytes were significantly induced by V. parahaemolyticus and S. aureus. The recombinant proteins of EsLRRop2 (rEsLRRop2) displayed a wide range of binding spectrum against vibrios, including V. parahaemolyticus, V. alginolyticus, and V. harveryi. The rEsLRRop2 showed dose- and time-dependent inhibitory activity against V. parahaemolyticus and S. aureus, and it could agglutinate the two bacteria. Furthermore, the inhibitory activities of rEsLRRop2 against V. parahaemolyticus, V. alginolyticus, V. harveryi and S. aureus was slightly affected by pH and salinity, and the rEsLRRop2 displayed the strongest inhibitory activity against all the three vibrios when the salinity was 20 ‰ and pH was 8.0. Collectively, these results elucidate the bacterial binding and inhibitory activities of EsLRRop2, and provide theoretical foundations for the application of rEsLRRop2 in prevention and control of vibrio diseases in aquaculture., Competing Interests: Declaration of competing interest The authors declare that they have no competing interests., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Endocrine-disrupting chemicals (EDCs) are toxic pollutants generated by artificial activities. Moreover, their hormone-like structure induces disturbances, such as mimicking or blocking metabolic activity. Previous studies on EDCs have focused on the adverse effect of the endocrine system in vertebrates, with limited investigations conducted on ion channels in invertebrates. Thus, in this study, we investigated the potential adverse effects of exposure to bisphenol-A (BPA) and di-(2-ethylhexyl) phthalate (DEHP) at the molecular level on the ryanodine receptor (RyR), a calcium ion channel receptor in Macrophthalmus japonicus. In the phylogenetic analysis, the RyR amino acid sequences in M. japonicus clustered with those in the Crustacean and formed separated branches for RyR in insects and mammals. When exposed to 1 μg L -1 BPA, a significant increase in RyR mRNA expression was observed in the gills on day 1, although a similar level to the control group was observed from day 4 to day 7. However, the RyR expression due to DEHP exposure decreased on days 1 and 4, although it increased on day 7 following exposure to 10 μg L -1 . The RyR expression pattern in the hepatopancreas increased for up to 4 days, depending on the BPA concentration. However, there was a tendency for the expression to decrease gradually after the statistical significance increased during the early stage of DEHP exposure (D1). Hence, the transcriptional alterations in the M. japonicus RyR gene observed in the study suggest that exposure toxicities to EDCs, such as BPA and DEHP, have the potential to disrupt calcium ion channel signaling in the gills and hepatopancreas of M. japonicus crabs., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Aquatic environments are subject to ultraviolet B (UVB) radiation incidence, and its effects on organisms are dose-dependent. Besides DNA, mitochondria are an important target of this radiation that causes structural damage and impairs its functional dynamics. Here, we hypothesize that mitophagy acts as an organelle quality control mechanism to mitigate UVB impacts in embryonic cells. Then, freshwater prawn Macrobrachium olfersii embryos was used as a model to investigate the effects of UVB on genes (Tomm20, Opa1, Pink, Prkn, Sqstm1, and Map1lc3) and proteins (TOM20, PINK1, p62 and LC3B) involved in mitophagy modulation. The choice of genes and proteins was based on the identification of mitochondrial membrane (Tomm20, Opa1 and TOM20), mediation of mitophagy (Pink1, Prkn and PINK1), and recognition of mitochondria by the autophagosome membrane (Sqstm1, Map1lc3, p62 and LC3B). First, the phylogeny of all genes presented bootstrap values >80 and conserved domains among crustacean species. Gene expression was inherently modulated during development, with transcripts (Tomm20, Opa1, Pink, Prkn, Sqstm1, and Map1lc3) overexpressed in the initial and final stages of development. Moreover, UVB radiation induced upregulation of Tomm20, Opa1, Pink, Prkn, Sqstm1, and Map1lc3 genes at 6 h after exposure. Interestingly, after 12 h, the protein content of PINK1, p62, and LC3B increased, while TOM20 was not responsive. Despite UVB radiation's harmful effects on embryonic cells, the chronology of gene expression and protein content indicates rapid activation of mitophagy, serving as an organelle quality control mechanism, given the analyzed cells' integrity., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Evelise Maria Nazari reports financial support and equipment, drugs, or supplies were provided by National Council for Scientific and Technological Development. If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023. Published by Elsevier Inc.)

This study investigated the impact of hyperthermal (34 °C) and hypothermal (14 °C) stress on the expression of the octopamine/tyramine receptor (LvOA/TA-R) and immune parameters in Litopenaeus vannamei, which is a species critical to the aquaculture industry. Given the sensitivity of aquatic organisms to climate change, understanding the physiological and immune responses of L. vannamei to temperature variations is essential for developing strategies to mitigate adverse effects. This research focuses on the immune response and expression changes of LvOA/TA-R under acute (0.5, 1, and 2 h) and chronic (24, 72, and 168 h) thermal stress conditions. Our findings reveal that thermal stress induces changes in LvOA/TA-R expression and impacts immune responses. Immune parameters such as total haemocyte count, differential haemocyte count, phenoloxidase activity, respiratory bursts, lysozyme activity, clearance efficiency, and phagocytosis exhibited a general trend of significant decline under the stress conditions. LvOA/TA-R had a higher expression in haemocyte under hyperthermal stress. The study elucidated that thermal stress modifies the expression of the LvOA/TA-R and diminishes immune functionality in L. vannamei, underscoring the potential influence of climate change on industry., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

In this study, we evaluated the role of the gnathosoma (mouthparts) in chemosensing of the most devastating honey bee parasite, Varroa destructor mite. Through transcriptomic analysis, we compared the expression of putative chemosensory genes between the body parts containing the main chemosensory organs (the forelegs), gnathosoma and the rest of the body devoid of these two body parts. Furthermore, we checked the presence of chemosensory-related transcripts in the proteome of the gnathosoma. Our comparative transcriptomic analysis revealed the presence of 83 transcripts with known characteristic conserved domains belonging to eight chemosensory gene families in the three Varroa transcriptomes. Among these transcripts, 11 were significantly upregulated in the mite's forelegs, compared to 8 and 10 in the gnathosoma and body devoid of both organs, respectively. Whilst the gnathosoma and the forelegs share similar expression of some putative lipid carrier proteins, membrane-bound receptors, and associated proteins, they also differ in the expression profiles of some transcripts belonging to these protein families. This suggests two functional chemosensory organs that may differ in their chemosensory function according to specific characteristics of compounds they detect. Moreover, the higher expression of some chemosensory transcripts in the body devoid of forelegs and gnathosoma compared to the gnathosoma alone, may suggest the presence of additional function of these transcripts or alternatively presence of additional external or internal chemosensory organs. Insights into the functional annotation of a highly expressed gustatory receptor present in both organs using RNA interference (RNAi) are also revealed., (© 2024. The Author(s).)

Hyalomma anatolicum is a tick of significant one-health importance due to its role as a vector for various pathogens affecting humans, animals and the environment, such as Theileria annulata, which causes tropical theileriosis in cattle, leading to severe economic losses. When infected with pathogens like T. annulata, the salivary glands of H. anatolicum undergo gene expression changes, secrete modified proteins and activate immune responses, all of which facilitate pathogen survival and transmission by modulating the host immune response and optimizing conditions for pathogen development. Understanding these responses is crucial for developing control strategies for tick-borne diseases. To understand the interaction between H. anatolicum and T. annulata, we performed a differential gene expression analysis of H. anatolicum salivary glands. An average of approximately 25 million raw sequencing reads were generated in each replicate using Illumina Sequencing. The sequenced reads were de novo assembled and the assembled transcriptome yielded approximately 50,231 non-redundant transcripts after clustering with CD-HIT using a sequence identity of 95% and alignment coverage of 90%. The assembly quality was evaluated with BUSCO analysis and found to be 86% complete using the Arachnida dataset and then blasted against non-redundant protein sequence database from NCBI followed by counting of reads and differential expression analysis. Overall, around 2400 and 400 genes were found differentially expressed with logFC > 1 and logFC > 2 respectively at FDR < 0.05. Top up-regulated genes included Calpain, Papilin, Neprilysin, and Ankyrin repeat-containing protein. Top down-regulated genes included Scoloptoxin, and Selenoprotein S and other uncharacterized proteins. Many other up-regulated proteins with high significance were uncharacterized suggesting room for further H. anatolicum functional and structural characterization studies. To our best knowledge, this is the first study of H. anatolicum sialotranscriptome which greatly contributes to sialotranscriptome information not only as sequence database but also indicates the potential targets for development of vaccine against ticks and transmission-blocking vaccines against T. annulata., (© 2024. The Author(s), under exclusive licence to Springer Nature Switzerland AG.)

Litopenaeus vannamei is a widely distributed euryhaline aquatic animal, affected by low salinity, which can impact its disease resistance and immunity. However, there is a limited understanding of the adaptation mechanisms of L. vannamei with different genetic backgrounds to low salinity. Therefore, the present study aimed to compare the immunity characteristics and transcriptomics of L. vannamei low salt-tolerant (FG I/J) and low salt-sensitive (control) families. Also, the disease resistance and immune parameters (including [THC], hemolymph cell viability, lysozyme activity [LZM], phenoloxidase content [PO], interleukin-6 [IL-6], and tumor necrosis factor-alpha [TNF-α]) of the FG I/J and control families of L. vannamei under low salinity (5‰) and ambient salinity (24‰) were examined. Additionally, hepatopancreas transcriptomics of the FG I/J and control families were analyzed at a salinity of 5‰. The results showed that the FG I/J family had higher disease resistance to Vibrio parahaemolyticus and stronger immunological capacity than the control family. Transcriptomic analysis showed significantly enriched energy metabolism and immune regulation pathways. Therefore, we speculated that energy metabolism provides sufficient energy for immunological modulation in the FG I/J family to deal with long-term low-salt stress and achieve high growth and survival rates., Competing Interests: Declaration of competing interest The authors declare that they have no conflict of interest., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

This study investigates the prolonged effect of immune disease resistance in Litopenaeus vannamei through the administration of tyramine (TA) formulated with polyethylene glycol (PEG). Facing the challenges of intensive farming, environmental stress, and global climate changes, innovative approaches to improve shrimp health are essential. The research focuses on the role of biogenic amines in stress response and immune regulation, demonstrating that TA, especially when combined with PEG, significantly prolongs immunity and resistance against Vibrio alginolyticus. The experimental design included administering TA, PEG, and TA-PEG, followed by evaluations of immunity, lactate and glucose levels, and immune-related gene expressions. Results showed notable prolonged effects in total hemocyte count, phenoloxidase activity, and phagocytic activity in the TA-PEG group, indicating enhanced immune activation period. Additionally, the expression of prophenoloxidase system-related genes was significantly upregulated in the TA-PEG group. Furthermore, the TA-PEG group exhibited a significantly higher survival rate in a susceptibility test against V. alginolyticus. The results of this study confirm that the combined use of PEG can effectively extend the immunostimulatory duration of TA., (Copyright © 2024. Published by Elsevier Ltd.)

Polycyclic aromatic hydrocarbons (PAHs), such as phenanthrene (PHE), are common pollutants found in coastal areas where shrimp farming is developed. Even though PAHs can have adverse effects on physiology, shrimp can detoxify and metabolize toxic compounds and neutralize the reactive oxygen species (ROS) produced during this process. This requires the activation of multiple antioxidant enzymes, including peroxiredoxin 6 (Prx6). Prx6 uses glutathione (GSH) to reduce phospholipid hydroperoxides, a function shared with GSH peroxidase 4 (GPx4). Prx6 has been scarcely studied in crustaceans exposed to pollutants. Herein, we report a novel Prx6 from the shrimp Penaeus vannamei that is abundantly expressed in gills and hepatopancreas. To elucidate the involvement of Prx6 in response to PAHs, we analyzed its expression in the hepatopancreas of shrimp sub-lethally exposed to PHE (3.3 μg/L) and acetone (control) for 24, 48, 72, and 96 h, along with GPx4 expression, GSH-dependent peroxidase activity, and lipid peroxidation (indicated by TBARS). We found that GPx4 expression is not affected by PHE, but Prx6 expression and peroxidase activity decreased during the trial. This might contribute to the rise of TBARS found at 48 h of exposure. However, maintaining GPx4 expression could aid to minimize lipid damage during longer periods of exposure to PHE., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Tropomyosin (TM) is the main allergen of Macrobrachium nipponense. Recombinant allergens have great prospects in the detection, diagnosis, and treatment of food allergens. The purpose of this study was to compare the differences in structure and allergenicity between natural TM and recombinant TM. Recombinant TM of M. nipponense with a molecular weight of 38 kDa was successfully expressed in the Escherichia coli system. The amino acid sequence as well as secondary structure between natural and recombinant TM were similar, which were verified by mass and CD spectrometry, respectively. Studies showed that both natural TM and recombinant TM had strong allergenicity, and recombinant TM was more allergenic, which could be used as a substitute for natural TM in the diagnosis and treatment of shrimp allergy. This study provided stable and reliable allergen components for the detection of crustacean allergens and the diagnosis and treatment of food allergies caused by crustacean allergens., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

The two-spotted spider mite (Tetranychus urticae Koch, TSSM) is recognized as one of the most problematic spider mite pests. However, the precise gene expression patterns across its key developmental stages remain elusive. Here, we performed a comprehensive transcriptome analysis of TSSM eggs, nymphs and adult females using publicly available RNA sequencing (RNA-seq) data to elucidate the overarching transcriptomic differences between these developmental stages. Principal component analysis and hierarchical clustering analysis unveiled distinct separations among samples across different developmental stages, regardless of their Wolbachia infection status. Differential expression analysis revealed 4,089,2,762, and 1,282 core genes specifically enriched in eggs, nymphs, and adults, respectively. KEGG and GO enrichment analyses showed upregulation of genes in eggs are associated with proteolysis, Wnt signaling pathway, DNA transcription, RNA biosynthetic and metabolic processes, as well as protein folding, sorting, and degradation pathways. Meanwhile, nymphs exhibited increased abundance of genes related to chitin/amino sugar metabolic processes, G protein-coupled receptor signaling pathways, monoatomic ion transport, and neurotransmitter transport pathways. Pathways involving sphingolipid and carbohydrate metabolic processes, proteolysis, lipid transport, and localization were particularly enriched in older females. Altogether, our findings suggest that the egg stage exhibits higher activity in cell differentiation processes, the nymph stage is more involved in chitin development, and the adult stage shows increased metabolic and reproductive activity. This study enhances our understanding of the molecular mechanisms underlying TSSM development and paves the way for further research into the intricate physiological processes of TSSM., (© 2024. The Author(s), under exclusive licence to Springer Nature Switzerland AG.)

Dragon fruit oligosaccharide (DFO) is an indigestible prebiotic that enhances the growth and reproduction of Daphnia magna, increases the expression of genes involved in immunity, and reduces oxidative stress. This study investigated the effects of DFO on the expression of innate immunity- (Toll, Pelle, proPO, A2M, and CTL), oxidative stress- (Mn-SOD), and nitric oxide (NO) synthesis-related genes (NOS1, NOS2, and arginase) as well as NO localization and number of hemocytes in D. magna. For this ten-day-old D. magna were treated with 0 or 9 mg l -1 of DFO for 24 and 85 h. Gene expression levels, NO intensity and localization, and total hemocytes were evaluated. After 24 h, the expression of Toll and proPO increased significantly (p < 0.05), while that of C-type lectins (CTL) was reduced (p < 0.05). At 85 h, Mn-SOD and CTL expressions were markedly suppressed (p < 0.05). NO was mostly localized in the foregut, midgut, hindgut, and carapace. The expression of NOS1 was reduced after 24 h (p < 0.05). In addition, NO intensity at 24 h was insignificantly lower than the control (p > 0.05). At 85 h, the expression of NOS1, NOS2, and arginase was higher than control, but NO intensity did not differ significantly (p > 0.05). Furthermore, the total hemocyte count elevated remarkably at 85 h (p < 0.05). Our study suggested that 9 mg l -1 of DFO could alter the expression of the genes related to innate immunity, oxidative stress, and NO synthesis in D. magna and significantly stimulate hemocyte production., Competing Interests: Declare of competing interest none., (Copyright © 2024 Elsevier Ltd. All rights reserved.)