Eel Liver Research Articles

Expression pattern of the fused in sarcoma gene and its contextual influence on the density-specific response of the growth hormone/insulin-like growth factor 1 axis in zig-zag eels (Mastacembelus armatus)

The fused in sarcoma (FUS) protein is a DNA/RNA binding protein from the ten-eleven translocation protein family that is associated with neurodegeneration, and it has been shown to promote cell proliferation through the growth hormone/insulin-like growth factor 1 (Gh/Igf-1) signaling pathway. The zig-zag eel (Mastacembelus armatus) is a newly discovered species exhibiting sexual dimorphism in growth, and the potential role of fus in the growth and development of this species remains largely unknown. Herein, we analyzed the homology, conserved domains, evolutionary characteristics, and conserved syntenies of fus in several teleost species. The expression of fus was predominant in the brain and exhibited sexual dimorphism in the brain, muscle, and liver of zig-zag eels. We found that microRNA (miR)-146-5p, miR-489-3p, and 24 other miRNAs were targeted to the fus 3′ untranslated region, which might affect muscle and bone development in adults. The igf1, insulin-like growth factor 1 receptor a (igf1ra), insulin-like growth factor 2 receptor (igf2r), growth hormone-releasing hormone-like receptor (ghrhrl), growth hormone secretagogue receptor type 1 (ghsr), and glucocorticoid receptor (gr) genes contained a higher abundance of GU-rich fus motifs compared to the other four genes analyzed in zig-zag eels. We also measured the expression of fus mRNA during fish culture at various stocking densities to further elucidate the relationship between fus expression and the Gh/Igf-1 axis. After 100 days of fish cultivation, the expression of fus and ghrhrl decreased and the expression of ghrh and gr increased as the culture density increased (p < 0.05). The expression of fus exhibited a remarkable positive correlation with a specific growth rate. These results indicate that fus mediates growth differences by regulating the expression of several growth-related genes including Gh/Igf-1 axis genes in zig-zag eels.

Effects of dietary soybean lecithin on growth, tissue fatty acid composition, and gonadal development in the hermaphroditic rice field eel (Monopterus albus)

This study investigates the effects of soybean lecithin (SL) on the growth, fatty acid composition, lipid metabolism, and gonadal development of rice field eels. Fourteen-month-old eels were fed either fish oil diets or varying levels of dietary soybean lecithin (0 %, 1.4 %, and 2.8 %) for 10 weeks. Dietary SL had no significant impact on growth, whereas 1.4 % SL led to a significant decrease in moisture content and an increase in crude protein content in eels. The content of both saturated and polyunsaturated fatty acids increased in muscles fed the higher SL diet, with a concurrent decline in saturated fatty acids and an increase in polyunsaturated fatty acids detected in the liver. The expression levels of genes related to lipid synthesis, including accα and PPARγ, and genes related to polyunsaturated fatty acid synthesis, such as pla2g4a, fad2 and scd, were significantly increased in the liver of eels fed with 1.4 % SL diet. SL supplementation enhances lipid transport and antioxidant capacity in the liver, reduces fat deposition, and protects the liver from damage. Hepatosomatic index of fish in the SL1.4 % and SL2.8 % groups decreased. Supplementation of the diet with 1.4 % SL led to a significant decrease in the content of saturated fatty acids and an increase in the content of polyunsaturated fatty acids in the gonads, with a significant increase in arachidonic acid levels. The gonadosomatic index also increased with increasing dietary SL content. Rice field eels fed diets containing SL exhibited elevated serum levels of E2, delayed ovary-to-testis transition, and a preference for retaining the female state. In conclusion, dietary supplementation with SL increased the proportion of highly unsaturated fatty acids in the tissues, enhanced lipid metabolism in the liver, and improved antioxidant capacity. Furthermore, SL supplementation has been demonstrated to promote ovarian development and delay sex reversal in rice field eels.

Transcriptome analysis of host anti-Aeromonas hydrophila infection revealed the pathogenicity of A. hydrophila to American eels (Anguilla rostrata)

Aeromonas hydrophila is a commonly pathogenic bacterium in cultivated eels, but its pathogenicity to American eel (Anguilla rostrata) and the molecular mechanism of host anti-A. hydrophila infection remains uncertain. In this study, LD50 of A. hydrophila to American eels was determined and bacterial load in the liver and kidney of eels was assessed post 2.56 doses of LD50 of A. hydrophila infection. The results showed that the LD50 of A. hydrophila to American eels was determined to be 3.9 × 105 cfu/g body weight (7.8 × 106 cfu/fish), and the bacterial load peaked at 36 h post the infection (hpi) in the liver. Then, the histopathology was highlighted by congestion in splenic blood vessels, atrophied glomeruli, and necrotic hepatocytes. Additionally, the results of qRT-PCR revealed that 18 host immune-related genes showed significantly up or downregulated post-infection compare to that of pre-infection. Finally, results of the RNA-seq revealed 10 hub DEGs and 7 encoded proteins play essential role to the anti-A. hydrophila infection in American eels. Pathogenicity of A. hydrophila to American eels and RNA-seq of host anti-A. hydrophila infection were firstly reported in this study, shedding new light on our understanding of the A. hydrophila pathogenesis and the host immune response to the A. hydrophila infection strategies in gene transcript.

The regulation of AMPK pathway in liver abnormal lipid deposition caused by high carbohydrate diet in rice field eel

AMP-activated protein kinase-alpha 1 (AMPKα1) plays a vital role in glucose and lipid metabolism. Five isonitrogenous and isolipid diets with starch levels (15%, 20%, 25%, 30% and 35%) were formulated to study the modulation of AMPKα1 in hepatic lipid deposition induced by high gelatinized starch diet in rice field eel (Monopterus albus) (initial body weight, 30.00 ± 0.10 g) and the growth trial lasted for 8 weeks. The results showed that dietary gelatinized starch did not affect the growth performance of rice filed eel compared with S15 group. High gelatinized starch could induce AMPKα1 expression and glucose catabolism, and inhibit hepatic glycogen synthesis without effecting glucose content. Dietary ≥ 30% gelatinized starch began to dramatically suppress gluconeogenesis and promote the lipid synthesis, followed by a higher triglyceride (TG) and hepatic crude lipid. Dietary 20–30% gelatinized starch induced phosphorylation of AMPKα compared to 15% gelatinized starch diet, whereas 35% starch dramatically suppressed AMPKα phosphorylation together with unnormal increase in phosphorylation of forkhead box O1 A (FOXO1A) and acetyl-CoA carboxylase (ACC), TG content compared to 30% starch. In short, high gelatinized starch could induce AMPKα1 expression and promoted systemic metabolism, whereas dietary ≥ 25% gelatinized starch initially suppressed AMPKα1 activation and caused abnormal lipid deposition in liver of rice field eel.

Effects of compound plant extracts on growth performance, antioxidant capacity, and histomorphology of liver and intestine of rice field eel (Monopterus albus)

AbstractCompound plant extracts (CPE) are beneficial for aquatic animals on growth performance and antioxidant capacity. A 56‐day experiment was conducted to investigate its positive effect on rice field eel. The fish were fed a commercial diet and supplementing CPE (mainly containing eucommia polysaccharides, reducing sugar) at 0, 0.8, 1.6, 2.4, and 3.2 g/kg. Results showed that dietary CPE significantly increased weight gain and specific growth rate (p < 0.05). Liver lipid content in 1.6 and 3.2 g/kg groups was significantly lower than in 0.8 and 2.4 g/kg groups (p < 0.05). Muscle lipid content in 0.8, 2.4, and 3.2 g/kg CPE groups was significantly lower than in control (p < 0.05). Meanwhile, intestinal digestive enzymatic activities in the 3.2 g/kg CPE group were the highest in comparison to all other groups (p < 0.05). Dietary CPE enhanced the antioxidant activities of serum and intestine (p < 0.05). Compared with the control, intestinal fold thickness in CPE groups was significantly increased (p < 0.05). Furthermore, dietary CPE reduced the degree of liver steatosis and the number of lipid droplet vacuoles. This study indicated that dietary CPE was beneficial to growth performance, antioxidant capacity, liver, and intestinal histology. Supplementation with 1.6–3.2 g/kg CPE is optimal for this eel species.

CRFB5a, a Subtype of Japanese Eel (Anguilla japonica) Type I IFN Receptor, Regulates Host Antiviral and Antimicrobial Functions through Activation of IRF3/IRF7 and LEAP2.

IFNAR1, one of the type I IFN receptors, is crucial to mammalian host defense against viral invasion. However, largely unknown is the immunological role of the fish teleost protein IFNAR1, also known as CRFB5. We have successfully cloned the whole cDNA of the Japanese eel's (Anguilla japonica) CRFB5a homolog, AjCRFB5a. The two fibronectin-3 domains and the transmembrane region (238-260 aa) of AjCRFB5a are normally present, and it shares a three-dimensional structure with zebrafish, Asian arowana, and humans. According to expression analyses, AjCRFB5a is highly expressed in all tissues found, particularly the liver and intestine. In vivo, Aeromonas hydrophila, LPS, and the viral mimic poly I:C all dramatically increased AjCRFB5a expression in the liver. Japanese eel liver cells were reported to express AjCRFB5a more strongly in vitro after being exposed to Aeromonas hydrophila or being stimulated with poly I: C. The membranes of Japanese eel liver cells contained EGFP-AjCRFB5a proteins, some of which were condensed, according to the results of fluorescence microscopy. Luciferase reporter assays showed that AjCRFB5a overexpression strongly increased the expression of immune-related genes in Japanese eel liver cells, such as IFN1, IFN2, IFN3, IFN4, IRF3, IRF5, and IRF7 of the type I IFN signaling pathway, as well as one of the essential antimicrobial peptides LEAP2, in addition to significantly inducing human IFN-promoter activities in HEK293 cells. Additionally, RNA interference (RNAi) data demonstrated that knocking down AjCRFB5a caused all eight of those genes to drastically lower their expression in Japanese eel liver cells, as well as to variable degrees in the kidney, spleen, liver, and intestine. Our findings together showed that AjCRFB5a participates in the host immune response to bacterial infection by inducing antimicrobial peptides mediated by LEAP2 and favorably modulates host antiviral immune responses by activating IRF3 and IRF7-driven type I IFN signaling pathways.

Complete genome sequence of Edwardsiella sp. NBRC12716 isolated in 1962 from the liver of diseased eel.

We report the complete genome sequence of Edwardsiella sp. NBRC12716 isolated from a diseased eel in 1962. The genome consists of a single, circular chromosome 3,771,060 bp in length with 59.74% GC content and encodes 25 rRNA, 96 tRNA, and 3,182 protein-coding genes.

Pathogenicity of Edwardsiella anguillarum to American eels (Anguilla rostrata) and RNA-seq analysis of host immune response to the E. anguillarum infection

Edwardsiella anguillarum is a commonly pathogenic bacterium in cultivated eels, but its pathogenicity to American eel (Anguilla rostrata) and the molecular mechanism of host anti-E. anguillarum infection remains uncertain. In this study, LD50 of E. anguillarum to American eels was determined and bacterial load in the liver and kidney of eels was assessed post the LD50 of E. anguillarum infection. The results showed that LD50 of E. anguillarum to American eels was determined to be 2.5 × 105 cfu/g body weight, and the bacterial load peaked at 36 and 72 h post the infection (hpi) in the kidney and liver, respectively. Then, the histopathology was highlighted by congestion in splenic blood vessels, atrophied glomeruli, and necrotic hepatocytes, as well as ultrastructural pathology in the kidney were charactered by acute nephritis, showing necrosis of the renal tubular epithelial cells, glomerular capillaries dilate, mitochondria swelling and ribosomes separate from the endoplasmic reticulum. Furthermore, the results of qRT-PCR revealed that 12 host immune-related genes showed significantly up or downregulated post-infection compare to that of pre-infection. Finally, results of the RNA-seq revealed 6 hub DEGs play essential role to the anti-E. anguillarum infection in American eels. Pathogenicity of E. anguillarum to American eels and hub genes related host anti- E. anguillarum infection were firstly reported in this study, shedding new light on our understanding of the E. anguillarum pathogenesis and the host immune response to the E. anguillarum infection strategies in gene transcript.

Insights into the mechanism of growth and fat deposition by feeding different levels of lipid provided by transcriptome analysis of swamp eel (Monopterus albus, Zuiew 1793) liver.

Lipid is an important source of energy in fish feeds, and the appropriate fat content can improve the efficiency of protein utilization. However, excessive lipid content in the feed can lead to abnormal fat deposition in fish, which has a negative effect on the growth of fish. Therefore, the effects of feed lipid levels on swamp eel were studied. Essential functional genes were screened using transcriptomics. We divided 840 fish into seven groups (four replicates). A mixture of fish and soybean oils (1:4), 0%, 2%, 4%, 6%, 8%, 10%, and 12% was added to the basic feed were named groups one to seven (L1-L7), respectively. Isonitrogenous diets were fed swamp eel for 10 weeks. Growth performance, visceral index, nutritional components, and biochemical indexes were measured and analyzed. Livers of the 0%, 6%, and 12% groups were subjected to transcriptome sequencing analysis. The results of our study showed that: the suitable lipid level for the growth of swamp eel was 7.03%; the crude fat content of whole fish, liver, intestine, muscle, and skin increased with the increase of lipid level, with some significant difference, and excess fat was deposited in skin tissue; triglyceride, total cholesterol, and free fatty acid contents increased with the increase of feed lipid level. High-density lipoprotein levels in the L3 and L4 groups were higher than in the other groups. Blood glucose concentrations in the L5, L6, and L7 groups increased; the liver tissue structure was damaged when the lipid level was too high. two-hundred-and-twenty-eight differentially expressed genes were found. Several critical pathways regulating glucose metabolism and energy balance (e.g., glycerolipid metabolism, glycolysis synthesis, degradation of ketone bodies, and Janus Kinase/Signal Transducer and Activator of Transcription signaling pathway) were enriched in swamp eel compared with the Kyoto Encyclopedia of Genes and Genomes (KEGG) database. Suitable lipid levels (7.03%) can promote the growth of swamp eel, and excessive lipid levels can cause elevated blood lipids and lead to liver cell damage. Regulatory mechanisms may involve multiple metabolic pathways for glucose and lipid metabolism in eels. This study provides new insights to explain the mechanism of fat deposition due to high levels of lipid and provides a basis for the production of efficient and environmentally friendly feed for swamp eel.

Analysis of Alternative Splicing and Long Noncoding RNAs After the Edwardsiella anguillarum Infected the Immunized European Eels (Anguilla anguilla) Revealed the Role of Outer Membrane Protein A in OmpA Subunit Vaccine.

Edwardsiella anguillarum is a bacterium that commonly infects cultivated eels. Outer membrane protein A (OmpA) emulsified with Freund's adjuvant has been shown to be an effective fishery vaccine against this pathogen. However, the specific roles of OmpA in the vaccine have not been fully explored. In this study, we performed RNA-seq in the liver of a European eel (Anguilla anguilla) after challenge with E. anguillarum in eels previously immunized with an OmpA subunit vaccine. Our aim was to elucidate the differentially alternative splicing (DAS) and differentially expressed long noncoding RNAs (DE-lncRNAs) using a genome-wide transcriptome. The results showed after that at 28days post-immunization, eels challenged with E. anguillarum (Con_inf)exhibited severe pathological changes in the liver. In contrast, the OmpA infused eels (OmpA_inf group) showed infiltrated lymphocytes, while Freund's adjuvant-inoculated eels (FCIA_inf group) showed edema of hepatocytes and blood coagulation. The relative percent survival (RPS) was 77.7% and 44.4% for OmpA_inf and FCIA_inf compared to the Con_inf group. We identified 37 DE-lncRNAs and 293 DAS genes between OmpA_inf and FCIA_inf. Interactions between DAS gene-expressed proteins indicated that 66 expressed proteins formed 20 networks. Additionally, 33 DE-lncRNAs interacted with 194 target genes formed 246 and 41 networks in co-expression and co-location. Taken together, our findings demonstrate that the OmpA subunit vaccine elicits a higher RPS and provides novel insights into the role of OmpA through DAS genes and DE-lncRNAs perspective. These results are significant for the development of fishery subunit vaccines.

IKKε positively regulates NF-κB, MAPK, and IRF3-mediated type I IFN signaling pathways in Japanese eel (Anguilla japonica)

IKKε is an IκB kinase participating in the control of NF-κB and type I IFN signal pathways in mammals. However, the function of IKKε in regulating immune response is largely unknown in teleost. Herein, an IKKε homologue named AjIKKε was characterized in Japanese eel (Anguilla japonica). AjIKKε has an N-terminal kinase domain, a ubiquitin-like domain, and a coiled coil-containing domain (CC), which is conserved and similar to its counterpart in mammals. Expression analysis showed that AjIKKε could be up-regulated in kidney, spleen, and particularly in liver under the stimulation of poly I:C, LPS, and Aeromonas hydrophila infection. In vitro, the mRNA levels of AjIKKε were significantly provoked in eel liver cells stimulated by LPS and poly I:C, or the different concentrations of A. hydrophila. The overexpression of AjIKKε could not only induce a significantly higher level of promoter activity of human NF-κB, AP-1, and IFN-β in a dose-dependent manner but also up-regulate the activation of promoters of Japanese eel cRel, AP1, IL6, IFN4, IRF3, and IRF7 in HEK293 cells. RNAi studies showed that after AjIKKε was knocked down, the expression levels of IL1, IL6, TNFα, c-Jun, IFN2, IFN3, MX1, MX2, and IRF3 genes were significantly down-regulated in liver, spleen, and kidney of Japanese eels. In addition, the mutants of AjIKKε-K39A, AjIKKε-S174A, and AjIKKε-ΔCC failed to activate Japanese eel IFN4, IRF3 and human IFN-β promoters in HEK293 cells. Collectively, these results suggest that AjIKKε may function as a positive regulator of NF-κB, MAPK, and IRF3-mediated type I IFN signaling pathways related to immune response evoked by bacterial and viral infection.

Substituted kynurenic acid derivatives as fluorophore-based probes for D- and L-amino acid oxidase assays and their in vitro application in eels.

High levels of D-amino acid oxidase (DAO) are associated with neurological and psychiatric disorders, while L-amino acid oxidase (LAO) exhibits antimicrobial and antitumor properties. The enzymatic conversion of the non-fluorescent kynurenine (KYN) into the endogenous weak fluorescent kynurenic acid (KYNA) by the action of DAO has previously been reported. However, the fluorescence of KYNA can be improved by changing the substituents on the aromatic rings. In this study, we prepared different 6-phenyl-substituted KYNA derivatives and investigated their fluorescence properties. Among them, 2-MePh-KYNA showed the maximum fluorescence quantum yield of 0.881 at 340 nm excitation and 418 nm emission wavelengths. The effects of solvent properties (dielectric constant, pKa, viscosity, and proticity) on the fluorescence intensity (FLI) of the KYNA derivatives were explored. The FLI of 2-MePh-KYNA was significantly large in protic solvents. Subsequently, 2-MePh-D-KYN and 2-MePh-L-KYN were prepared with high enantiopurity (>99.25%) for the enzymatic conversion. 2-MePh-D-KYN exhibited high sensitivity (∼19 times that of a commercial DAO substrate and ∼60 times that of the previously reported MeS-D-KYN) and high selectivity, as it was not cross-reactive towards LAO, while 2-MePh-L-KYN was also converted into 2-MePh-KYNA by LAO. Furthermore, the 2-MePh-D-KYN probe successfully detected DAO in eel liver, kidney, and heparin-anticoagulated plasma in the in vitro study.

Comprehensive Relationship Analysis of the Long Noncoding RNAs (lncRNAs) and the Target mRNAs in Response to the Infection of Edwardsiella anguillarum in European eel (Anguilla anguilla) Inoculated with Freund's Adjuvant.

Freund's complete adjuvant (FCA) and incomplete adjuvant (FIA), generally applied in subunit fishery vaccine, have not been explored on the molecular mechanism of the non-specific immune enhancement. As long noncoding RNAs (lncRNAs) play vital regulating roles in various biological activities, in this study, we examined the genome-wide expression of transcripts in the liver of European eel (Anguilla anguilla, Aa) inoculated with FCA and FIA (FCIA) to elucidate the regulators of lncRNAs in the process of Edwardsiella anguillarum (Ea) infection and Aa anti-Ea infection using strand-specific RNA-seq. After eels were challenged by Ea at 28days post the first inoculation (dpi), compared to the control uninfected eels (Li group), the control infected eels (Con_Li group) showed severe bleeding, hepatocyte atrophy, and thrombi formed in the hepatic vessels of the liver, although eels inoculated with FCIA (FCIA_Li group) also formed slight thrombi in the hepatic vessels. Compared to the FCIA_Li group, there was about 10 times colony-forming unit (cfu) in the Con_Li group per 100μg liver tissue, and the relative percent survival (RPS) of eels was 50% in FCIA_Li vs Con_Li. Using high-throughput transcriptomics, differential expressed genes (DEGs) and transcripts were identified and the results were verified using fluorescence real-time polymerase chain reaction (qRT-PCR). Interactions between the differential expressed lncRNAs (DE-lncRNAs) and the target DEGs were explored using Cytoscape according to their co-expression and co-location relationship. We found 13,499 lncRNAs (10,176 annotated and 3423 novel lncRNAs) between 3 comparisons of Con_Li vs Li, FCIA_Li vs Li, and FCIA_Li vs Con_Li, of which 111, 110, and 129 DE-lncRNAs were ascertained. Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis of DEGs targeted by DE-lncRNAs revealed these DEGs mainly involved in single-organism cellular process in BP, membrane in CC and binding in MF, and KEGG pathways showed that the target DEGs in co-expression and co-location enriched in cell adhesion molecules. Finally, 118 DE-lncRNAs target 1161 DEGs were involved in an interaction network of 8474 co-expression and 333 co-location-related links, of which 16 DE-lncRNAs play vital roles in anti-Ea infection. Taken together, the interaction networks revealed that DE-lncRNAs underlies the process of Ea infection and Aa anti-Ea infection.

Dietary bile acids supplementation mainly regulates the amino acid metabolic pathways without decreasing bile acids levels in the liver of farmed European eel (Anguilla anguilla) juveniles

This study was conducted to investigate the effects of dietary bile acids (BAs) supplementation on the liver metabolism of farmed European eel (Anguilla anguilla) juveniles. Fish with similar body weight (140 g per fish) in six tanks (681.7 ± 22.7 kg fish per tank) were randomly divided into the control group fed a basal diet and the BAs group fed a basal diet supplemented with 500 mg/kg BAs. The experiment lasted for 15 weeks. The untargeted metabolomics method was used to evaluate global metabolic profiles, while the targeted metabolomics method was utilized for BAs profile analysis in the liver. Compared with the control group, there were 33 upregulated metabolites and 15 downregulated metabolites in the BAs group. The differential metabolites belonged to amino acids, carbohydrates, lipids, nucleotides, vitamins, etc. Among the top ten differential metabolic pathways affected by BAs supplementation, the main pathways were relevant to amino acids metabolisms. These included aminoacyl-tRNA biosynthesis, phenylalanine, tyrosine and tryptophan biosynthesis, alanine, aspartate and glutamate metabolism, histidine metabolism, arginine and proline metabolism, beta-alanine metabolism, phenylalanine metabolism, D-glutamine and D-glutamate metabolism, sphingolipid metabolism, and pantothenate and CoA biosynthesis. Only the levels of deoxycholic acid, glycodeoxycholic acid, and glycoursodeoxycholic acid were significantly increased in the liver of the European eel in the BAs group (P < 0.05). These results indicated that exogenous bile acids might mainly regulate the metabolism of amino acids without decreasing the bile acids levels in the liver of farmed European eel juveniles.

Rag1 and Rag2 Gene Expressions Identify Lymphopoietic Tissues in Larvae of Rice-Field Eel (Monopterus albus).

In immature lymphocytes, recombination activating genes 1 and 2 are necessary for antigen receptor V (D) J recombination, representing immature lymphocyte biomarkers. Herein, we cloned and sequenced rice-field eel rag1 and rag2 genes. Their expressions in the thymus, liver, and kidney were significant from 0 days post hatching (dph) to 45 dph, peaking at 45 dph in these three tissues. In situ hybridization detected high rag1 and rag2 expressions in the liver, kidney, and thymus of rice-field eel from 2 to 45 dph, suggesting that multiple tissues of rice-field eel contain lymphocyte lineage cells and undergo lymphopoiesis. Tissue morphology was used to observe lymphopoiesis development in these three tissues. The thymus primordium began to develop at 2 dph, while the kidney and liver have generated. Our findings verified that the thymus is the primary lymphopoietic tissue and suggested that, in rice-field eel, lymphocyte differentiation also occurs in the liver and kidney.

Life-stage dependent histological characterization of liver HIF-1α expression in the Indonesian shortfin eel (Anguilla bicolor bicolor McClelland, 1884)

The Indonesian shortfin eel (Anguilla bicolor bicolor McClelland, 1884) is catadromous fish. The developmental phase of the fish is influenced by the availability and type of feed, and dissolved oxygen levels (DO). Oxygen level fluctuations during migration from sea to fresh waters can cause hypoxia and trigger activation of the Hypoxia-inducible factor 1 alpha gene (HIF-1α). Therefore, undertanding HIF-1α expression is very important in understanding the life cycle and physiology of this organism. This study aimed to perform a histological comparison between the liver structure of eel in the elver eel phase to silver eel and compared the expression of HIF-1α in the liver. The eels were collected from Kulon Progo Regency, Segara Anakan in Cilacap, and Cangkringan in Sleman, respectively. The eel livers were prepared with standard paraffin method. The samples were stained with Hematoxylin-Eosin, Mallory Acid Fuchsin and Periodic Acid-Schiff Alcian Blue for histological observation. The HIF-1α protein expression was detected using immunohistochemistry. The results showed that the structure of liver elver, yellow, and silver eel shared similar tubular hepatocytes. Moreover, there was no significant differences in the parenchymal structure of liver of all stage of eel. Mucin levels were different in each life cycle phase. HIF-1α expression varied by eel phase, with the elver eel showing weak expression, yellow stage showing moderate expression, and silver eels displaying the highest levels of expression. We conclude that the highest expression of HIF-1α was found in the silver eel, which resides in freshwater environments and is found in freshwater while returning to the sea for reproduction.

Transcriptomic analysis using dual RNA sequencing revealed a Pathogen–Host interaction after Edwardsiella anguillarum infection in European eel (Anguilla anguilla)

Many studies have explored differentially expressed genes (DEGs) between some pathogens and hosts, but no study has focused on the interaction of DEGs between Edwardsiella anguillarum (Ea) and Anguilla anguilla (Aa). In this study, we examined the interactions of DEGs during Ea infection and Aa anti-infection processes by dual RNA sequencing. Total RNA from in vitro and in vivo (Aa liver) Ea culture was extracted. Using high-throughput transcriptomics, significant DEGs that were expressed between Ea cultured in vitro versus in vivo and those in the liver of the infected group versus control group were identified. Protein–protein interactions between the pathogen and host were explored using Cytoscape according to the HPIDB 3.0 interaction transcription database. The results showed that the liver in the infection group presented with severe bleeding and a large number of thrombi in the hepatic vessels. We found 490 upregulated and 398 downregulated DEGs of Ea in vivo versus Ea cultured in vitro, and 2177 upregulated and 970 downregulated genes in the liver of the infected eels. Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis of the pathogen DEGs revealed that the upregulated genes were mainly enriched in migration, colonization, biofilm formation, and significantly enriched in ABC transport and quorum sensing; the downregulated genes were mainly involved in metabolism, information transduction, organelle formation, enzyme catalysis, molecular transport, and binding. GO of the host DEGs showed that metabolic process, catalytic activity, single organism metabolic process, small molecule binding, nucleotide binding, nucleotide phosphate binding, and anion binding were markedly enriched. Finally, we found that 79 Ea and 148 Aa proteins encoded by these DEGs were involved in an interaction network, and some pathogen (DegP, gcvP, infC, carB, rpoC, trpD, sthA, and FhuB) and host proteins (MANBA, STAT1, ETS2, ZEP1, TKT1, NMI and RBPMS) appear to play crucial roles in infection. Thus, determining the interaction networks revealed crucial molecular mechanisms underlying the process of pathogenic infection and host anti-infection.

Histological alterations, oxidative stress, and inflammatory response in the liver of swamp eel (Monopterus albus) acutely exposed to copper.

Copper (Cu) is widely used as an essential trace element in diets as well as a therapeutic chemical. However, excessive Cu has deleterious effects on organisms, including teleosts. Although numerous toxic effects of Cu have been reported, the effects of Cu exposure on the swamp eel (Monopterus albus) as well as the underlying mechanisms have not yet been elucidated. In this study, swamp eels were acutely exposed to 100, 200, and 400μg/L of Cu for 96h to evaluate liver histopathology, oxidative stress, and inflammation. Dissolution of hepatocyte membrane, vacuolar degeneration, and inflammatory cell infiltration were detected in the livers of the Cu-treated swamp eels, especially in the 400μg Cu/L group. Cu-induced hepatic dysfunction was further verified by the elevated activities of glutamate oxaloacetate transaminase (GOT) and glutamate pyruvate transaminase (GPT) and transcript levels of GOT and GPT genes. In addition, Cu exposure decreased the activities of total superoxide dismutase T-SOD and catalase (CAT) and the contents of glutathione (GSH) and total antioxidant capacity (T-AOC) and increased the levels of malondialdehyde (MDA). Cu exposure also significantly decreased the transcript levels of glutathione synthetase (GSS) and increased the transcript levels of SOD1, SOD2, CAT, and heme oxygenase-1 (HO-1) genes. Furthermore, pro-inflammatory genes such as interleukin (IL)-1β, tumor necrosis factor-α (TNF-α), and IL-8 were significantly upregulated. These results indicate that Cu induces oxidative stress and inflammatory response and causes pathological changes in the liver of the swamp eel.

Molecular characterization, expression patterns, and subcellular localization of a classical and a novel nonclassical MHC class I α molecules from Japanese eel Anguilla japonica

Major histocompatibility complex (MHC) molecules play a critical role in the immune response of vertebrate animals by presenting foreign antigens to T lymphocytes. In this study, we first cloned and identified a classical and a novel nonclassical MHC I α molecules from Japanese eel (Anguilla japonica), named as AjMHC I-UBA and AjMHC I-L, respectively. The full-length cDNA of AjMHC I-UBA contains an open reading frame (ORF) of 1047 bp encoding a predicted protein of 348 amino acids, while AjMHC I-L 1089 bp encodes 362 amino acids. The multiple alignment of the amino acid sequence showed that AjMHC I-UBA and AjMHC I-L consist of an N-terminal MHC I superfamily domain within α1 and α2 helices, an IgC-MHC I α3 domain, and a transmembrane region close to the C-terminal, which are similar to other fish and mammal species. Molecular polymorphism analysis showed that eight different major alleles of AjMHC I-UBA, named as AjMHC I-UBA*0101～1001, were identified from six Japanese eel individuals. Furthermore, a distinguishing signature of the nonclassical L-lineage genes specific motif “HINMTL”, including an N-glycosylation site (NXS/T), was present in the α3 domain of AjMHC I-L sequences. Although the predicted three-dimensional structures of AjMHC I-UBA and AjMHC I-L are similar to that of human MHC I α, phylogenetic analysis showed that these two protein molecules belong to classical MHC I UBA gene of U-lineage and nonclassical MHC I gene of L-lineage, respectively. Quantitative real-time polymerase chain reaction (qRT-PCR) analysis revealed that the highest expression of AjMHC I-UBA and AjMHC I-L was found in the intestine and the expression level of AjMHC I-UBA in all of the examined tissues was significantly higher than that of AjMHC I-L. The expressions of AjMHC I-UBA and AjMHC I-L in liver, kidney and spleen were significantly induced following injection with the viral mimic poly I:C, LPS, and Aeromonas hydrophila infection. In vitro, the AjMHC I-UBA and AjMHC I-L transcripts of Japanese eel liver cells were significantly enhanced by the treatment of poly I:C or the stimulation of the high concentration of A. hydrophila. Subcellular localization showed that under natural conditions, AjMHC I-UBA and AjMHC I-L were uniformly distributed in the cytoplasm, and aggregated into spots or flakes in the cytoplasm after the stimulation of poly I:C or LPS. These results collectively suggested that AjMHC I-UBA and AjMHC I-L are important components possibly involved in Japanese eel defense against viral and bacterial infection. Taken together, these findings provide valuable insight into the immune function of MHC class I in the immune system of teleost.

Mercury accumulation in muscle and liver tissue and human health risk assessment of two resident freshwater fish species in Flanders (Belgium): a multilocation approach.

Detrimental effects of chemical pollution-primarily caused by human activities-on aquatic ecosystems have increasingly gained attention. Because of its hydrophobic qualities, mercury is prone to easily bioaccumulate and biomagnify through the food chain, decreasing biodiversity and eventually also affecting humans. In the present study, accumulated mercury concentrations were measured in muscle and liver tissue of perch (Perca fluviatilis) and European eel (Anguilla anguilla) collected at 26 sampling locations in Flemish (Belgian) waterbodies, allowing a comparison of these species within a variety of environmental situations. Furthermore, effects of size and weight have been assessed, expected to influence accumulation and storage of pollutants. Mercury concentrations in perch ranged up to 1.7 μg g-1 dw (median: 0.29 μg g-1 dw) in muscle and from 0.02 to 0.77 μg g-1 dw (median: 0.11 μg g-1 dw) in liver tissue. For eel, these concentrations were between 0.07 and 1.3 μg g-1 dw (median: 0.39 μg g-1 dw) and between 0.08 and 1.4 μg g-1 dw (median: 0.55 μg g-1 dw) respectively. We found a correlation of accumulated mercury with length in perch, independent of location. Furthermore, a significant difference in accumulated mercury concentrations between the targeted species was measured, with the highest mean concentrations per dry weight in eel liver and muscle tissue. In perch, higher concentrations were found in muscle compared to liver tissue, while in eel, liver tissue showed the highest concentrations. These findings were further considered with concentrations corrected for lipid content, excluding the fat compartment, which is known to a hold negligible portion of the total and methyl mercury concentrations. This confirmed our previous conclusions, except for mercury concentrations in eel. Here there was no longer a significant difference between muscle and liver concentrations. Finally, health risk analyses revealed that only frequent consumption of local eel (> 71 g day-1) could pose risks to humans.