Identification of TonB-dependent siderophore receptor inhibitors against Flavobacterium columnare using a structure-based high-throughput virtual screening method.

TonB-dependent siderophore receptors play a critical transport role for Flavobacterium columnare virulence formation and growth, and have become valuable targets for the development of novel antimicrobial agents. Traditional Chinese medicine has demonstrated notable efficacy in the treatment of fish diseases and includes potential antibacterial agents. Herein, we performed molecular docking-based virtual screening to discover novel TonB-dependent siderophore receptor inhibitors from traditional Chinese medicine and provide information for developing novel antibacterial agents. Firstly, we efficiently obtained 11 potential inhibitors with desirable drug-like characteristics from thousands of compounds in the TCM library based on virtual screening and property prediction. The antibacterial activity of Enoxolone, along with its interaction characteristics, were determined via an MIC assay and molecular dynamic simulation. Transcriptional profiling, along with validation experiments, subsequently revealed that an insufficient uptake of iron ions by bacteria upon binding to the TonB-dependent siderophore receptors is the antibacterial mechanism of Enoxolone. Finally, Enoxolone's acceptable toxicity was illustrated through immersion experiments. In summary, we have used virtual screening techniques for the first time in the development of antimicrobial agents in aquaculture. Through this process, we have identified Enoxolone as a promising compound targeting the TonB-dependent siderophore receptor of F. columnare. In addition, our findings will provide new ideas for the advancement of innovative antimicrobial medications in aquaculture.

Exploitation of multiple host-derived nutrients by the yellow catfish epidermal environment facilitates Vibrio mimicus to sustain infection potency and susceptibility.

Infection with Vibrio mimicus in the Siluriformes has demonstrated a rapid and high infectivity and mortality rate, distinct from other hosts. Our earlier investigations identified necrosis, an inflammatory storm, and tissue remodeling as crucial pathological responses in yellow catfish (Pelteobagrus fulvidraco) infected with V. mimicus. The objective of this study was to further elucidate the impact linking these pathological responses within the host during V. mimicus infection. Employing metabolomics and transcriptomics, we uncovered infection-induced dense vacuolization of perimysium; Several genes related to nucleosidase and peptidase activities were significantly upregulated in the skin and muscles of infected fish. Concurrently, the translation processes of host cells were impaired. Further investigation revealed that V. mimicus completes its infection process by enhancing its metabolism, including the utilization of oligopeptides and nucleotides. The high susceptibility of yellow catfish to V. mimicus infection was associated with the composition of its body surface, which provided a microenvironment rich in various nucleotides such as dIMP, dAMP, deoxyguanosine, and ADP, in addition to several amino acids and peptides. Some of these metabolites significantly boost V. mimicus growth and motility, thus influencing its biological functions. Furthermore, we uncovered an elevated expression of gangliosides on the surface of yellow catfish, aiding V. mimicus adhesion and increasing its infection risk. Notably, we observed that the skin and muscles of yellow catfish were deficient in over 25 polyunsaturated fatty acids, such as Eicosapentaenoic acid, 12-oxo-ETE, and 13-Oxo-ODE. These substances play a role in anti-inflammatory mechanisms, possibly contributing to the immune dysregulation observed in yellow catfish. In summary, our study reveals a host immune deviation phenomenon that promotes bacterial colonization by increasing nutrient supply. It underscores the crucial factors rendering yellow catfish highly susceptible to V. mimicus, indicating that host nutritional sources not only enable the establishment and maintenance of infection within the host but also aid bacterial survival under immune pressure, ultimately completing its lifecycle.

Preparation and evaluation of microencapsulated delivery system of recombinant interferon alpha protein from rainbow trout.

Diseases caused by viruses pose a significant risk to the health of aquatic animals, for which there are presently no efficacious remedies. Interferon (IFN) serving as an antiviral agent, is frequently employed in clinical settings. Due to the unique living conditions of aquatic animals, traditional injection of interferon is cumbersome, time-consuming and labor-intensive. This study aimed to prepare IFN microcapsules through emulsion technique by using resistant starch (RS) and carboxymethyl chitosan (CMCS). Optimization was achieved using the Box-Behnken design (BBD) response surface technique, followed by the creation of microcapsules through emulsification. With RS at a concentration of 1.27 %, a water­oxygen ratio of 3.3:7.4, CaCl2 at 13.67 %, CMCS at 1.04 %, the rate of encapsulation can escalate to 80.92 %. Rainbow trout infected with Infectious hematopoietic necrosis virus (IHNV) and common carp infected with Spring vireemia (SVCV) exhibited a relative survival rate (RPS) of 65 % and 60 % after treated with IFN microcapsules, respectively. Moreover, the microcapsules effectively reduced the serum AST levels and enhanced the expression of IFNα, IRF3, ISG15, MX1, PKR and Viperin in IHNV-infected rainbow trout and SVCV-infected carp. In conclusion, this integrated IFN microcapsule showed potential as an antiviral agent for treatment of viral diseases in aquaculture.

Histopathology and transcriptome profiling reveal features of immune responses in gills and intestine induced by Spring viremia of carp virus.

The immune system of bony fish closely resembles that of mammals, comprising both specific (adaptive) and non-specific (innate) components. Notably, the mucosa-associated lymphoid tissue (MALT) serves as the first line of defense within the non-specific immune system, playing a critical role in protecting these aquatic organisms against invading pathogens. MALT encompasses a network of immune cells strategically distributed throughout the gills and intestines, forming an integral part of the mucosal barrier that interfaces directly with the surrounding aquatic environment. Spring Viremia of Carp Virus（SVCV）, a highly pathogenic agent causing substantial harm to common carp populations, has been designated as a Class 2 animal disease by the Ministry of Agriculture and Rural Affairs of China. Utilizing a comprehensive array of research techniques, including Hematoxylin and Eosin (HE)、Alcian Blue Periodic Acid-Schiff (AB-PAS)、transcriptome analysis for global gene expression profiling and Reverse Transcription-Polymerase Chain Reaction (RT-qPCR), this study uncovered several key findings: SVCV is capable of compromising the mucosal architecture in the gill and intestinal tissues of carp, and stimulate the proliferation of mucous cells both in gill and intestinal tissues. Critically, the study revealed that SVCV's invasion elicits a robust response from the carp's mucosal immune system, demonstrating the organism's capacity to resist SVCV invasion despite the challenges posed by the pathogen.

Long-term dietary exposure to 2,2',4,4'-tetrabromodiphenyl ether (BDE-47) reduced feeding in common carp (Cyprinus carpio): Via the JAK-STAT signaling pathway.

Polybrominated diphenyl ethers (PBDEs) are widely present in water ecosystems where they pose a significant threat to aquatic life, but our knowledge about how PBDEs affect feeding is limited. Therefore, this study explored the effects of continuous dietary exposure to 2,2',4,4'-tetrabromodiphenyl ether (BDE-47) (40 and 4000 ng/g) on the feeding in common carp (Cyprinus carpio) and the underlying mechanism. BDE-47 significantly decreased the food intake of carp. Transcriptome analysis of brain tissue showed that BDE-47 mainly affected the nervous, immune, and endocrine systems. Further examination of the expression levels of appetite factors in the brain revealed that BDE-47 caused dysregulation of appetite factors expressions such as agrp, pomc, cart, etc. In addition, the JAK-STAT signaling pathway was activated under BDE-47 exposure. It can be concluded from these findings that BDE-47 activated the JAK-STAT signaling pathway, causing imbalanced expression of appetite factors, leading to disordered feeding behavior and decreased food intake in carp. These results provide an important reference for a more comprehensive understanding of the hazards posed by BDE-47 on animal feeding and the associated mechanisms.

Real-world evidence: Risdiplam in a patient with spinal muscular atrophy type I with a novel splicing mutation and one SMN2 copy.

Spinal muscular atrophy (SMA), which results from the deletion or/and mutation in the SMN1 gene, is an autosomal recessive neuromuscular disorder that leads to weakness and muscle atrophy. SMN2 is a paralogous gene of SMN1. SMN2 copy number affects the severity of SMA, but its role in patients treated with disease modifying therapies is unclear. The most appropriate individualized treatment for SMA has not yet been determined. Here, we reported a case of SMA type I with normal breathing and swallowing function. We genetically confirmed that this patient had a compound heterozygous variant: one deleted SMN1 allele and a novel splice mutation c.628-3T>G in the retained allele, with one SMN2 copy. Patient-derived sequencing of 4 SMN1 cDNA clones showed that this intronic single transversion mutation results in an alternative exon (e)5 3' splice site, which leads to an additional 2 nucleotides (AG) at the 5' end of e5, thereby explaining why the patient with only one copy of SMN2 had a mild clinical phenotype. Additionally, a minigene assay of wild type and mutant SMN1 in HEK293T cells also demonstrated that this transversion mutation induced e5 skipping. Considering treatment cost and goals of avoiding pain caused by injections and starting treatment as early as possible, risdiplam was prescribed for this patient. However, the patient showed remarkable clinical improvements after treatment with risdiplam for 7 months despite carrying only one copy of SMN2. This study is the first report on the treatment of risdiplam in a patient with one SMN2 copy in a real-world setting. These findings expand the mutation spectrum of SMA and provide accurate genetic counseling information, as well as clarify the molecular mechanism of careful genotype-phenotype correlation of the patient.

Corrigendum: Transcriptome reveals the role of the htpG gene in mediating antibiotic resistance through cell envelope modulation in Vibrio mimicus SCCF01.

[This corrects the article DOI: 10.3389/fmicb.2023.1295065.].

Dietary exposure to 2,2',4,4'-tetrabromodiphenyl ether (BDE-47) induces oxidative damage promoting cell apoptosis primarily via mitochondrial pathway in the hepatopancreas of carp, Cyprinus carpio.

To investigate the mechanisms of BDE-47 on hepatotoxicity in fish, this study examined the effects of dietary exposure to BDE-47 (40 and 4000 ng/g) on carp for 42 days. The results showed that BDE-47 significantly increased carp's condition factor and hepatosomatic index. Pathological results revealed unclear hepatic cord structure, hepatocytes swelling, cellular vacuolization, and inflammatory cell infiltration in the hepatopancreas of carp. Further investigation showed that ROS levels significantly increased on days 7, 14, and 42. Moreover, the activities of antioxidant enzymes SOD, GSH, CAT, and GST increased significantly from 1 to 7 days, and the transcription levels of antioxidant enzymes CAT, Cu-Zn SOD, Mn-SOD, GST, and GPX, and antioxidant pathway genes Keap1, Nrf2, and HO-1 changed significantly at multiple time-points during the 42 days. The results of apoptosis pathway genes showed that the mitochondrial pathway genes Bax, Casp3, and Casp9 were significantly upregulated and Bcl2 was significantly downregulated, while the transcription levels of FADD and PERK were significantly enhanced. These results indicate that BDE-47 induced oxidative damage in hepatopancreas, then it promoted cell apoptosis mainly through the mitochondrial pathway. This study provides a foundation for analyzing the mechanism of hepatotoxicity induced by BDE-47 on fish.

Neglected function of gastrin to reduce feeding in Siberian sturgeon (Acipenser baerii) via cholecystokinin receptor B.

Gastrin is an important intragastrointestinal hormone, but reports on its regulation of feeding behavior in fish are still scarce. This study aimed to determine the feeding regulatory function of gastrin in sturgeon. In this study, a gastrin/cholecystokinin-like peptide was identified in the genomes of sturgeon and proved to be gastrin by evolutionary tree analysis. Tissue distribution of gastrin and its receptor, cholecystokinin receptor B (CCKRB), showed that both had high mRNA abundance in the hypothalamus and gastrointestinal tract. In the duodenum, gastrin and CCKRB mRNAs were reduced at 1 h of fasting, and both were also observed in the stomach and hypothalamus in response to changes in feeding status. Sulfated gastrin 17 is the major form of gastrin in vivo. Therefore, we investigated the effect of sulfated gastrin 17 on feeding by intraperitoneal injection into Siberian sturgeon using sulfated gastrin 17. The results showed that gastrin 17 significantly reduced the cumulative feeding of Siberian sturgeon in the short term (1, 3 and 6 h) and long term (1, 2, 3, 4, 5 and 7 days). Finally, we explored the potential mechanism of feeding inhibition after intraperitoneal injection of gastrin 17 for 7 consecutive days. The results showed that gastrin 17 treatment significantly increased the mRNA levels of anorexigenic peptides (cart, cck and pyy), while it had no significant effect on the mRNA abundance of orexigenic peptides (npy and agrp). In addition, gastrin 17 treatment significantly affected the expression of appetite signaling pathways in the hypothalamus, such that the mRNA expression of ampkα1 was significantly reduced, whereas the mRNA abundance of stat3, mtor and s6k was significantly increased. In conclusion, the present study confirmed the anorectic effect of gastrin on Siberian sturgeon.

The cAMP Receptor Protein (CRP) of Vibrio mimicus Regulates Its Bacterial Growth, Type II Secretion System, Flagellum Formation, Adhesion Genes, and Virulence.

Vibrio mimicus is a serious pathogen in aquatic animals, resulting in significant economic losses. The cAMP receptor protein (CRP) often acts as a central regulator in highly pathogenic pathogens. V. mimicus SCCF01 is a highly pathogenic strain isolated from yellow catfish; the crp gene deletion strain (Δcrp) was constructed by natural transformation to determine whether this deletion affects the virulence phenotypes. Their potential molecular connections were revealed by qRT-PCR analysis. Our results showed that the absence of the crp gene resulted in bacterial and colony morphological changes alongside decreases in bacterial growth, hemolytic activity, biofilm formation, enzymatic activity, motility, and cell adhesion. A cell cytotoxicity assay and animal experiments confirmed that crp contributes to V. mimicus pathogenicity, as the LD50 of the Δcrp strain was 73.1-fold lower compared to the WT strain. Moreover, qRT-PCR analysis revealed the inhibition of type II secretion system genes, flagellum genes, adhesion genes, and metalloproteinase genes in the deletion strain. This resulted in the virulence phenotype differences described above. Together, these data demonstrate that the crp gene plays a core regulatory role in V. mimicus virulence and pathogenicity.

Ano1 regulates embryo transport by modulating intracellular calcium levels in oviduct smooth muscle.

Oviductal smooth muscle exhibits spontaneous rhythmic contraction (SRC) and controls the passage of the ova at the exact time, but its mechanistic regulation remains to be determined. In this study, female mice with Ano1SMKO (smooth muscle-specific deletion of Ano1) had reduced fertility. Deficiency of Ano1 in mice resulted in impaired oviductal SRC function and reduced calcium signaling in individual smooth muscle cells in the oviduct. The Ano1 antagonist T16Ainh-A01 dose-dependently inhibited SRCs and [Ca2+]i in the oviducts of humans and mice. A similar inhibitory effect of SRCs and [Ca2+]i was observed after treatment with nifedipine. In our study, ANO1 acted primarily as an activator or amplifier in [Ca2+]i and contraction of tubal smooth muscle cells. We found that tubal SRC was markedly attenuated in patients with ectopic pregnancy. Then, our study was designed to determine whether chloride channel Ano1-mediated smooth muscle motility is associated with tubal SRC. Our findings reveal a new mechanism for the regulation of tubal motility that may be associated with abnormal pregnancies such as ectopic pregnancies.

Salvia miltiorrhiza polysaccharide promotes the health of crayfish (Procambarus clarkii) by promoting hemocyte phagocytosis, protecting hepatopancreas and enhancing intestinal barrier function.

Plant polysaccharides as immunomodulators are considered one of the effective measures to reduce antibiotic therapy in aquaculture. The immunomodulatory function of Salvia miltiorrhiza polysaccharides (SMP) has been demonstrated and begun to be applied in vertebrates, but its potential effect on crustaceans is unclear. In this study, crayfish (Procambarus clarkii) was fed with 0 %, 0.3 %, 0.7 %, 1.1 %, and 1.5 % SMP for 4 weeks to investigate the effects of SMP on hemocytes phagocytosis, hepatopancreatic function, and intestinal barrier function. The results revealed that hemocyte phagocytic activity was increased in all SMP groups. During the process of hemocytes phagocytic recognition and formation of phagosomes and phagolysosomes, the mRNA expression levels of mas, hem, rab3, ctsb, and lamp-1 were up-regulated mainly in the 0.3 % SMP group. During the clearance phase of phagocytosis, respiratory burst activity, ROS level, T-SOD, CAT, GST, and LZM activities were mainly increased in the 1.5 % SMP group. Hepatopancreas AKP and GOT activity were no significant change in all SMP groups. ACP activity was significantly enhanced in the 1.1 % SMP group. The GPT activity of 0.3-0.7 % SMP group was significantly decreased. The 0.7 % SMP group had the highest intestinal fold height. The highest index values of OTUs, Ace, Chao, and Shannon were in the 0.3 % SMP group. The dietary addition of 0.3 % SMP led to a tendency of increased relative abundance of Firmicutes and Bacteroidota at the phylum level, while the relative abundance of Proteobacteria at the phylum level decreased. In conclusion, dietary SMP could promote crayfish health by enhancing phagocytosis, protecting hepatopancreas and enhancing intestinal barrier function. This study contributes to the theoretical foundation for exploring the potential application of plant polysaccharides in crustaceans.

Identification, characterization and the inflammatory regulating effect of NOD1/2 in sturgeon.

As an ancient species with both conservation and commercial value, Sturgeon's inflammatory regulation mechanism is a research point. Nucleotide-binding and oligomerization domain-containing proteins 1 and 2 (NOD1/2) are classical intracellular pattern recognition receptors (PRRs) in immunity of anti-bacterial infection. However, the characterization and function of NOD1/2 in Sturgeon are still unclear. In this study, we analyzed the synteny relationship of NOD1/2 genes between Acipenser ruthenus and representative fishes at the genome-level. Results showed that the ArNOD2 collinear genes pair was present in all representative fishes. The duplicated ArNOD1/2 genes were under purifying selection during evolution as indicated by their Ka/Ks values. To explore the function of NOD1/2, we further investigated their expression patterns and the effects of pathogenic infection, PAMPs treatment, and siRNA interference in Acipenser baerii, the sibling species of A. ruthenus. Results showed that both AbNOD1/2 were expressed at early developmental stages and in different tissues. Pathogenic infection in vivo and PAMPs treatment in vitro demonstrated that AbNOD1/2 could respond to pathogen stimulation. siRNA interference with AbNOD1/2 inhibited expression levels of RIPK2 and inflammatory cytokines compared to the control group after iE-DAP or MDP treatment. This study hinted that the AbNOD1/2 could stimulate the inflammatory cytokines response during evolutionary processes.

Microbiome analysis reveals the intestinal microbiota characteristics and potential impact of Procambarus clarkii.

The intestinal microbiota interacts with the host and plays an important role in the immune response, digestive physiology, and regulation of body functions. In addition, it is also well documented that the intestinal microbiota of aquatic animals are closely related to their growth rate. However, whether it resulted in different sizes of crayfish in the rice-crayfish coculture model remained vague. Here, we analyzed the intestinal microbiota characteristics of crayfish of three sizes in the same typical rice-crayfish coculture field by high-throughput sequencing technology combined with quantitative real-time polymerase chain reaction (qRT-PCR) and enzyme activity, investigating the relationship between intestinal microbiota in crayfish and water and sediments. The results showed that the dominant intestinal microbiota of crayfish was significantly different between the large size group (BS), normal size group (NS), and small size group (SS), where Bacteroides and Candidatus_Bacilloplasma contributed to the growth of crayfish by facilitating food digestion through cellulolysis, which might be one of the potential factors affecting the difference in sizes. Follow-up experiments confirmed that the activity of lipase (LPS) and protease was higher in BS, and the relative expression of development-related genes, including alpha-amylase (α-AMY), myocyte-specific enhancer factor 2a (MEF2a), glutathione reductase (GR), chitinase (CHI), and ecdysone receptor (EcR), in BS was significantly higher than that in SS. These findings revealed the intestinal microbiota characteristics of crayfish of different sizes and their potential impact on growth, which is valuable for managing and manipulating the intestinal microbiota in crayfish to achieve high productivity in practice. KEY POINTS: • Significant differences in the dominant microflora of BS, NS, and SS in crayfish. • Cellulolysis might be a potential factor affecting different sizes in crayfish. • Adding Bacteroides and Candidatus_Bacilloplasma helped the growth of crayfish.

Identification of C1q/TNF-related protein 4 as a novel appetite-regulating peptide that reduces food intake in Siberian sturgeon (Acipenser baerii).

Emerging findings point to a role for C1q/TNF-related protein 4 (CTRP4) in feeding in mammals. However, it remains unknown whether CTRP4 regulates feeding in fish. This study aimed to determine the feeding regulation function of CTRP4 in Siberian sturgeon (Acipenser baerii). In this study, the Siberian sturgeon ctrp4 (Abctrp4) gene was cloned, and Abctrp4 mRNA was shown to be highly expressed in the hypothalamus. In the hypothalamus, Abctrp4 mRNA decreased during fasting and reversed after refeeding. Subsequently, we obtained the AbCTRP4 recombinant protein by prokaryotic expression and optimized the expression and purification conditions. Siberian sturgeon (81.28 ± 14.75 g) were injected intraperitoneally using 30, 100, and 300 ng/g Body weight (BW) AbCTRP4 to investigate its effect on feeding. The results showed that 30, 100, and 300 ng/g BW of the AbCTRP4 significantly reduced the cumulative food intake of Siberian sturgeon at 1, 3, and 6 h. Finally, to investigate the potential mechanism of CTRP4 feeding inhibition, 300 ng/g BW AbCTRP4 was injected intraperitoneally. The findings demonstrated that AbCTRP4 treatment for 1 h significantly promoted the mRNA levels of anorexigenic peptides (pomc, cart, and leptin) while suppressing the mRNA abundances of orexigenic peptides (npy and agrp).In addition, the jak2/stat3 pathway in the hypothalamus was significantly activated after 1 h of AbCTRP4 treatment. In conclusion., this study confirms the anorexigenic effect of CTRP4 in Siberian sturgeon.

Genotype diversity and antibiotic resistance risk in Aeromonas hydrophila in Sichuan, China.

Sichuan is a significant aquaculture province in China, with a total aquaculture output of 1.72 × 106 tons in 2022. One of the most significant microorganisms hurting the Sichuan aquaculture is Aeromonas hydrophila, whose genotype and antibiotic resistance are yet unknown. This study isolated a total of 64 strains of A. hydrophila from various regions during September 2019 to June 2021 within Sichuan province, China. The technique of Multi-Locus Sequence Typing (MLST) was used for the purpose of molecular typing. Meanwhile, identification of antibiotic resistance phenotype and antibiotic resistance gene was performed. The findings of the study revealed that 64 isolates exhibited 29 sequence types (ST) throughout different regions in Sichuan, with 25 of these ST types being newly identified. Notably, the ST251 emerged as the predominant sequence type responsible for the pandemic. The resistance rate of isolated strains to roxithromycin was as high as 98.3%, followed by co-trimoxazole (87.5%), sulfafurazole (87.5%), imipenem (80%), amoxicillin (60%), and clindamycin (57.8%). Fifteen strains of A. hydrophila exhibited resistance to medicines across a minimum of three categories, suggesting the development of multidrug resistance in these isolates. A total of 63 ARGs were detected from the isolates, which mediated a range of antibiotic resistance mechanisms, with deactivation and efflux potentially serving as the primary mechanisms of antibiotic resistance. This study revealed the diversity of A. hydrophila genotypes and the risk of antibiotic resistance in Sichuan, providing reference for scientific and effective control of A. hydrophila infection.

The potential mechanism of concentrated mannan-oligosaccharide promoting goldfish's (Carassius auratus Linnaeus) resistance to Ichthyophthirius multifiliis invasion.

Because of the low host specificity, Ichthyophthirius multifiliis (Ich) can widely cause white spot disease in aquatic animals, which is extremely difficult to treat. Prior research has demonstrated a considerable impact of concentrated mannan-oligosaccharide (cMOS) on the prevention of white spot disease in goldfish, but the specific mechanism is still unknown. In this study, transcriptome sequencing, histological analysis, immunofluorescence analysis, phagocytosis activity assay and qRT-PCR assay were used to systematically reveal the potential mechanism of cMOS in supporting the resistance of goldfish (Carrasius auratus) to Ich invasion. According to the transcriptome analysis, the gill tissue of goldfish receiving the cMOS diet showed greater expression of mannose-receptor (MRC) related genes, higher phagocytosis activity, up-regulated expression of phagocytosis-related genes and inflammatory-related genes compared with the control, indicating that cMOS can have an effect on phagocytosis and non-specific immunity of goldfish. After the Ich challenge, transcriptome analysis revealed that cMOS fed goldfish displayed a higher level of phagocytic response, whereas non-cMOS fed goldfish displayed a greater inflammatory reaction. Besides, after Ich infection, cMOS-fed goldfish displayed greater phagocytosis activity, a stronger MRC positive signal, higher expression of genes associated with phagocytosis (ABCB2, C3, MRC), and lower expression of genes associated with inflammation (IL-1ß, IL-17, IL-8, TNF-α, NFKB). In conclusion, our experimental results suggest that cMOS may support phagocytosis by binding to MRC on the macrophage cell membrane and change the non-specific immunity of goldfish by stimulating cytokine expression. The results of this study provide new insights for the mechanism of cMOS on parasitic infection, and also suggest phagocytosis-related pathways may be potential targets for prevention of Ich infection.

Spring Viremia of Carp Virus Infection Induces Carp IL-10 Expression, Both In Vitro and In Vivo.

Interleukin-10 (IL-10) is a pleiotropic cytokine with both immune enhancement and immunosuppression activities, but the main role is immunosuppression and anti-inflammatory ability. In order to use the immunosuppressive function of IL-10, many viruses, such as SARS-CoV-2, hepatitis B virus and EB virus, can evade the host's immune surveillance and clearance by increasing the expression of host IL-10. However, it has not been reported whether the aquatic animal infection virus can upregulate the expression of host IL-10 and the mechanisms are still unknown. Spring viremia of carp (SVC) is a fatal viral disease for many fish species and is caused by spring viremia of carp virus (SVCV). This disease has caused significant economic losses in the aquaculture industry worldwide. In this study, the expression of carp IL-10 with or without infection of SVCV in epithelioma papulosum cyprinid (EPC) cells, carp head kidney (cHK) primary cells and common carp tissues were analyzed using RT-PCR and ELISA. The results show that SVCV infection induced carp IL-10 mRNA and protein expression, both in vitro and in vivo. However, the upregulation of carp IL-10 by SVCV was hindered by specific inhibitors of the JAK inhibitor (CP-690550), STAT3 inhibitor (STA-21), NF-κB inhibitor (BAY11-7082) and p38 MAPK (mitogen-activated protein kinase) inhibitor (SB202190), but not JNK inhibitor (SP600125). Furthermore, the results demonstrated that JAK1, JAK2, JAK3, TYK2 and STAT5 played important roles in carp IL-10 production induced by SVCV infection. Taken together, SVCV infection significantly induced carp IL-10 expression and the upregulation trigged in JAK-STAT, NF-κB and p38MAPK pathways. To our knowledge, this is the first time that a fish infection virus upregulated the host IL-10 expression through the JAK-STAT, NF-κB and p38MAPK pathways. Altogether, fish viruses may have a similar mechanism as human or other mammalian viruses to escape host immune surveillance and clearance.

Ammonia nitrogen stress damages the intestinal mucosal barrier of yellow catfish (Pelteobagrus fulvidraco) and induces intestinal inflammation.

Nitrogen from ammonia is one of the most common pollutants toxics to aquatic species in aquatic environment. The intestinal mucosa is one of the key mucosal defenses of aquatic species, and the accumulation of ammonia nitrogen in water environment will cause irreversible damage to intestinal function. In this study, histology, immunohistochemistry, ultrastructural pathology, enzyme activity analysis and qRT-PCR were performed to reveal the toxic effect of ammonia nitrogen stress on the intestine of Pelteobagrus fulvidraco. According to histological findings, ammonia nitrogen stress caused structural damage to the intestine and reduced the number of mucous cells. Enzyme activity analysis revealed that the activity of bactericidal substances (Lysozyme, alkaline phosphatase, and ACP) had decreased. The ultrastructure revealed sparse and shortened microvilli as well as badly degraded tight junctions. Immunohistochemistry for ZO-1 demonstrated an impaired intestinal mucosal barrier. Furthermore, qRT-PCR revealed that tight junction related genes (ZO-1, Occludin, Claudin-1) were downregulated, while the pore-forming protein Claudin-2 was upregulated. Furthermore, as ammonia nitrogen concentration grew, so did the positive signal of Zap-70 (T/NK cell) and the expression of inflammation-related genes (TNF, IL-1ß, IL-8, IL-10). In light of the above findings, we conclude that ammonia nitrogen stress damages intestinal mucosal barrier of Pelteobagrus fulvidraco and induces intestinal inflammation.

Starvation induces hepatopancreas atrophy in Chinese mitten crab (Eriocheir sinensis) by inhibiting angiogenesis.

BACKGROUND: The hepatopancreas of crustaceans serves as a significant organ for both the synthesis and secretion of digestive enzymes, as well as energy storage. In the event of food shortage, the hepatopancreas can provide energy for survival. To investigate the potential regulatory mechanisms of the hepatopancreas in response to starvation in Eriocheir Sinensis, transcriptome analysis, histological study and qRT-PCR were performed. RESULTS: The results showed that starvation caused a decrease in the hepatopancreas index of E. sinensis, which had certain effects on the tissue structure, metabolism and angiogenesis in the hepatopancreas. In addition, WGCNA and linear regression analysis showed that the genes significantly related to the hepatopancreas index were mainly enriched in the angiogenesis pathway, in which AKT signaling played an important role. Starvation may inhibit AKT signaling pathway by reducing the expression of TGFBI, HSP27, HHEX, and EsPVF1, thereby hindering angiogenesis, promoting apoptosis, and leading to hepatopancreas atrophy. CONCLUSION: These results indicate that AKT plays an important role in the angiogenesis pathway and apoptosis of the starvation induced hepatopancreas index reduction, which is beneficial to further understand the effect of starvation stress on hepatopancreas of Chinese mitten crab.