Rainbow Trout Head Kidney Research Articles

Tenacibaculum dicentrarchi produce outer membrane vesicles (OMV) that are associated with the cytotoxic effect in rainbow trout head kidney macrophages.

Tenacibaculum dicentrarchi is the second most important pathogen in Chilean salmon farming. This microorganism causes severe skin lesions on the body surface of farmed fish. The bacterium can also adhere to surfaces and form biofilm, survive in fish skin mucus, and possess different systems for iron acquisition. However, the virulence mechanisms are still not fully elucidated. Outer membrane vesicles (OMV) are nanostructures released by pathogenic Gram-negative bacteria during growth, but none has been described yet for T. dicentrarchi. In this study, we provide the first reported evidence of the fish pathogen T. dicentrarchi producing and releasing OMV from 24 h after incubation, increasing thereafter until 120 h. Analyses were conducted with T. dicentrarchi TdCh05, QCR29, and the type strain CECT 7612T . The OMV sizes, determined via scanning electron microscopy, ranged from 82.25 nm to 396.88 nm as per the strain and incubation time point (i.e., 24 to 120 h). SDS-PAGE revealed that the number of protein bands evidenced a drastically downward trend among the T. dicentrarchi strains. In turn, the OMV shared five proteins (i.e., 22.2, 31.9, 47.7, 56.3, and 107.1 kDa), but no protein pattern was identical. A heterogeneous amount of protein, RNA, and DNA were obtained, depending on the time at which OMV were extracted. Purified OMV were biologically active and induced a cytotoxic effect in macrophage-enriched cell cultures from rainbow trout (Oncorhynchus mykiss) head kidneys. This is the first step towards understanding the role that OMV could play in the pathogenesis of T. dicentrarchi.

Integrative mRNA-miRNA interaction analysis associated with the immune response in the head kidney of rainbow trout (Oncorhynchus mykiss) after infectious hematopoietic necrosis virus infection

Rainbow trout (Oncorhynchus mykiss) is an important cold-water fish widely cultivated in China. The frequent occurrence of viral diseases caused by infectious hematopoietic necrosis virus (IHNV) seriously restricted the healthy development of the rainbow trout farming industry. However, the immune defense mechanism induced by IHNV in rainbow trout has not been fully elucidated. In the present study, we detected mRNA and miRNA expression profiles in rainbow trout head kidney after IHNV infection using RNA-seq and identified key immune-related genes and miRNAs. The results showed that a total of 7486 genes and 277 miRNAs were differentially expressed, and numerous differentially expressed genes (DEGs) enriched in the immune-related pathways such as Toll-like receptor signaling pathway, RIG-I-like receptor signaling pathway, NOD-like receptor signaling pathway, cytokine-cytokine receptor interaction, and JAK-STAT signaling pathway were significantly up-regulated, including LGP2, MDA5, TRIM25, IRF3, IRF7, TLR3, TLR7, TLR8, MYD88, and IFN1. Integration analysis identified six miRNAs (miR-141-y, miR-200-y, miR-144-y, miR-2188-y, miR-725-y, and miR-203-y) that target at least six key immune-related genes (TRIM25, LGP2, TLR3, TLR7, IRF3, and IRF7). Further, we verified selected immune-related mRNAs and miRNAs through qRT-PCR and confirmed the reliability of the RNA-seq results. These findings improve our understanding of the immune mechanism of rainbow trout infected with IHNV and provide basic data for future breeding for disease resistance in rainbow trout.

Chitosan Microparticles Enhance the Intestinal Release and Immune Response of an Immune Stimulant Peptide in Oncorhynchus mykiss.

The aquaculture industry is constantly increasing its fish production to provide enough products to maintain fish consumption worldwide. However, the increased production generates susceptibility to infectious diseases that cause losses of millions of dollars to the industry. Conventional treatments are based on antibiotics and antivirals to reduce the incidence of pathogens, but they have disadvantages, such as antibiotic resistance generation, antibiotic residues in fish, and environmental damage. Instead, functional foods with active compounds, especially antimicrobial peptides that allow the generation of prophylaxis against infections, provide an interesting alternative, but protection against gastric degradation is challenging. In this study, we evaluated a new immunomodulatory recombinant peptide, CATH-FLA, which is encapsulated in chitosan microparticles to avoid gastric degradation. The microparticles were prepared using a spray drying method. The peptide release from the microparticles was evaluated at gastric and intestinal pH, both in vitro and in vivo. Finally, the biological activity of the formulation was evaluated by measuring the expression of il-1β, il-8, ifn-γ, Ifn-α, and mx1 in the head kidney and intestinal tissues of rainbow trout (Oncorhynchus mykiss). The results showed that the chitosan microparticles protect the CATH-FLA recombinant peptide from gastric degradation, allowing its release in the intestinal portion of rainbow trout. The microparticle-protected CATH-FLA recombinant peptide increased the expression of il-1β, il-8, ifn-γ, ifn-α, and mx1 in the head kidney and intestine and improved the antiprotease activity in rainbow trout. These results suggest that the chitosan microparticle/CATH-FLA recombinant peptide could be a potential prophylactic alternative to conventional antibiotics for the treatment of infectious diseases in aquaculture.

Lost and Found: The Family of NF-κB Inhibitors Is Larger than Assumed in Salmonid Fish.

NF-κB signalling is largely controlled by the family of 'inhibitors of NF-κB' (IκB). The relevant databases indicate that the genome of rainbow trout contains multiple gene copies coding for iκbα (nfkbia), iκbε (nfkbie), iκbδ (nkfbid), iκbζ (nfkbiz), and bcl3, but it lacks iκbβ (nfkbib) and iκbη (ankrd42). Strikingly, three nfkbia paralogs are apparently present in salmonid fish, two of which share a high sequence identity, while the third putative nfkbia gene is significantly less like its two paralogs. This particular nfkbia gene product, iκbα, clusters with the human IκBβ in a phylogenetic analysis, while the other two iκbα proteins from trout associate with their human IκBα counterpart. The transcript concentrations were significantly higher for the structurally more closely related nfkbia paralogs than for the structurally less similar paralog, suggesting that iκbβ probably has not been lost from the salmonid genomes but has been incorrectly designated as iκbα. In the present study, two gene variants coding for iκbα (nfkbia) and iκbε (nfkbie) were prominently expressed in the immune tissues and, particularly, in a cell fraction enriched with granulocytes, monocytes/macrophages, and dendritic cells from the head kidney of rainbow trout. Stimulation of salmonid CHSE-214 cells with zymosan significantly upregulated the iκbα-encoding gene while elevating the copy numbers of the inflammatory markers interleukin-1-beta and interleukin-8. Overexpression of iκbα and iκbε in CHSE-214 cells dose-dependently quenched both the basal and stimulated activity of an NF-κB promoter suggesting their involvement in immune-regulatory processes. This study provides the first functional data on iκbε-versus the well-researched iκbα factor-in a non-mammalian model species.

Conserved and divergent arms of the antiviral response in the duplicated genomes of salmonid fishes

Antiviral innate immunity is orchestrated by the interferon system, which appeared in ancestors of jawed vertebrates. Interferon upregulation induces hundreds of interferon-stimulated-genes (ISGs) with effector or regulatory functions. Here we investigated the evolutionary diversification of ISG responses through comparison of two salmonid fishes, accounting for the impact of sequential whole genome duplications ancestral to teleosts and salmonids. We analysed the transcriptomic response of the IFN pathway in the head kidney of rainbow trout and Atlantic salmon, which separated 25–30 Mya. We identified a large set of ISGs conserved in both species and cross-referenced them with zebrafish and human ISGs. In contrast, around one-third of salmonid ISG lacked orthologs in human, mouse, chicken or frog, and often between rainbow trout and Atlantic salmon, revealing a fast-evolving, lineage-specific arm of the antiviral response. This study also provides a key resource for in-depth functional analysis of ISGs in salmonids of commercial significance.

Innate immune response of rainbow trout erythrocytes to spinycterins expressing a downsized viral fragment of viral haemorrhagic septicaemia virus

Recent studies have reported on the importance of RBCs in fish responses to viral infections and DNA vaccines. Surface-displaying recombinant bacterins (spinycterins) are a safe and adaptable prototype for viral vaccination of fish and represent an alternative method of aquaculture prophylaxis, since have been reported to enhance fish immune response. We evaluated the innate immune response of rainbow trout (Oncorhynchus mykiss) red blood cells (RBCs), head kidney, and spleen to spinycterins expressing a fragment of the glycoprotein G of viral haemorrhagic septicemia virus (VHSV), one of the most devastating world-wide diseases in farmed salmonids. We first selected an immunorelevant downsized viral fragment of VHSV glycoprotein G (frg16252–450). Then, spinycterins expressing frg16252–450 fused to Nmistic anchor-motif (Nmistic+frg16252–450) were compared to spinycterins expressing frg16252–450 internally without the anchor motif. Nmistic+frg16252–450 spinycterins showed increased attachment to RBCs in vitro and modulated the expression of interferon- and antigen presentation–related genes in RBCs in vitro and in vivo, after intravenous injection. In contrast, the head kidney and spleen of fish injected with frg16252–450, but not Nmistic+frg16252–450, spinycterins demonstrated upregulation of interferon and antigen-presenting genes. Intravenous injection of Nmistic+frg16252–450 spinycterins resulted in a higher innate immune response in RBCs while frg16252–450 spinycterins increased the immune response in head kidney and spleen. Although more studies are required to evaluate the practicality of using spinycterins as fish viral vaccines, these results highlight the important contribution of RBCs to the fish innate immune response to antiviral prophylactics.

Pathogenic Mechanism of a Highly Virulent Infectious Hematopoietic Necrosis Virus in Head Kidney of Rainbow Trout (Oncorhynchusmykiss) Analyzed by RNA-Seq Transcriptome Profiling.

Infectious hematopoietic necrosis virus (IHNV) is a pathogen that causes high rates of mortality in salmonid fishes. Therefore, an RNA-seq-based transcriptome analysis was performed in the head kidney of rainbow trout infected with a highly virulent IHNV strain to understand the pathogenesis of and defense strategies for IHNV infection in rainbow trout. The results showed that the numbers of DEGs were 618, 2626, and 774 (control vs. IHNV) on days 1, 3, and 5, respectively. Furthermore, the enrichment analysis of gene ontology (GO) annotations to classify DEGs showed that GO terms considerably associated with DEGs were gluconeogenesis, inflammatory response, and cell adhesion in the Biological Process (BP) category, apical plasma membrane, extracellular matrix (ECM) in the Cellular Component category, and transporter activity, integrin binding, and protein homodimerization activity in the Molecular Function category, on days 1, 3, and 5, respectively. Notably, GO terms in the BP category, including the negative regulation of type I interferon production and positive regulation of interleukin-1β secretion, were commonly identified at all time points. In the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis, complement and coagulation cascades were commonly identified at all time points. Importantly, the widely recognized GO terms and KEGG pathways extensively linked to DEGs were related to energy metabolism on day 1, the immune response on day 3, and cell proliferation on day 5. Furthermore, protein–protein interaction networks and centrality analysis showed that the metabolism and signaling transduction pathways were majorly upregulated. Conclusively, the virulent IHNV infection drives pathogenesis by activating the metabolic energy pathway for energy use for viral replication, facilitating necrosis through autophagy, and causing a shutoff response of the host immune system through the downregulation of type I IFN at the initial stage of infection.

RNA-Seq Analysis of the Key Long Noncoding RNAs and mRNAs Related to the Regulation of Acute Heat Stress in Rainbow Trout.

Simple SummaryAt present, climate warming is a very serious environmental problem. A sudden and large increase or decrease in temperature is likely to cause stress response in animals. Rainbow trout is a kind of cultured cold-water fish, which is very sensitive to high temperature. Therefore, it is very vulnerable to heat waves during production. The current study found that the behavior, antioxidant capacity, and natural immune function of rainbow trout under acute heat stress were significantly enhanced in the early stages of stress response, but its anti-stress ability decreased with an increase in stress intensity and duration. Transcriptome sequencing and bioinformatics analysis showed that some non-coding RNAs could competitively bind to target genes, and jointly participate in metabolism, apoptosis, and the immune regulation of rainbow trout under stress environments. In conclusion, our study can lay a theoretical foundation for the breeding of heat-resistant rainbow trout varieties.As the global climate warms, more creatures are threatened by high temperatures, especially cold-water fish such as rainbow trout. Evidence has demonstrated that long noncoding RNAs (lncRNAs) play a pivotal role in regulating heat stress in animals, but we have little understanding of this regulatory mechanism. The present study aimed to identify potential key lncRNAs involved in regulating acute heat stress in rainbow trout. lncRNA and mRNA expression profiles of rainbow trout head kidney were analyzed via high-throughput RNA sequencing, which exhibited that 1256 lncRNAs (802 up-regulation, 454 down-regulation) and 604 mRNAs (353 up-regulation, 251 down-regulation) were differentially expressed. These differentially expressed genes were confirmed to be primarily associated with immune regulation, apoptosis, and metabolic process signaling pathways through Gene Ontology and Kyoto Encyclopedia of Genes and Genomes pathway enrichment analysis and coding-noncoding co-expression network analysis. These results suggested that 18 key lncRNA-mRNA pairs are essential in regulating acute heat stress in rainbow trout. Overall, these analyses showed the effects of heat stress on various physiological functions in rainbow trout at the transcriptome level, providing a theoretical basis for improving the production and breeding of rainbow trout and the selection of new heat-resistant varieties.

Transcriptome Analysis in the Head Kidney of Rainbow Trout (Oncorhynchus mykiss) Immunized with a Combined Vaccine of Formalin-Inactivated Aeromonas salmonicida and Vibrio anguillarum.

This study aimed to identify the molecular mechanisms regulated by a combined vaccine against Aeromonas salmonicida and Vibrio anguillarum (O1 serotype). These bacteria cause furunculosis and vibriosis, respectively, and are associated with a high mortality in rainbow trout in Korea. The vaccine upregulated gene expression of TCRα, T-bet, sIgM, and mIgM, markers of an activated adaptive immune response. On days 1, 3, and 5, transcriptome analysis revealed 862 (430 up- and 432 downregulated), 492 (204 up- and 288 downregulated), and 741 (270 up- and 471 downregulated) differentially expressed genes (DEGs), respectively. Gene ontology (GO) enrichment analysis identified 377 (108 MF, 132 CC, 137 BP), 302 (60 MF, 180 CC, 62 BP), and 314 (115 MF, 129 CC, 70 BP) GOs at days 1, 3, and 5, respectively. Kyoto Encyclopedia of Genetic and Genomic enrichment analysis identified eight immune system-related pathways like cytokine-cytokine receptor interaction, NF-kappaB signaling pathway, TNF signaling pathway, NOD-like receptor signaling pathway, cytosolic DNA sensing pathway, cell adhesion molecule, complement and coagulation cascade, and antigen processing and presentation. In the analysis of the protein–protein interaction of immune-related DEGs, a total of 59, 21, and 21 interactional relationships were identified at days 1, 3, and 5, respectively, with TNF having the highest centrality at all three time points.

Differential MHC-П, IL-8 and IgM genes expression in the liver, head Kidney and gill tissues of juvenile rainbow trout (Oncorhynchus mykiss) immunized by inactive trophonts of Ichthyophthirius multifiliis

Differential MHC-П, IL-8 and IgM genes expression in the liver, head Kidney and gill tissues of juvenile rainbow trout (Oncorhynchus mykiss) immunized by inactive trophonts of Ichthyophthirius multifiliis

Transcriptome profiling in head kidney of rainbow trout (Oncorhynchus mykiss) after infection with the low-virulent Nagano genotype of infectious hematopoietic necrosis virus.

Infectious hematopoietic necrosis virus (IHNV) causes clinical diseases and mortality in a wide variety of salmonid species. Here, we studied transcriptional responses in rainbow trout infected by the IHNV-Nagano strain isolated in Korea. RNA-seq-based transcriptome analysis of head kidney tissues cataloged differentially expressed genes. Enrichment analysis of gene ontology annotations was performed, and a total of fifteen biological process terms were commonly identified at all time points. In the Kyoto Encyclopedia of Genes and Genomes pathway analysis, pathogen recognition receptor (PRR) signaling pathways such as the retinoic-acid-inducible gene-I-like receptor signaling pathway and the Toll-like receptor signaling pathway were identified at all time points. The nucleotide-binding oligomerization-domain-like receptor signaling pathway and cytosolic DNA-sensing pathway were identified at days 1 and 3. Protein-protein interaction network and centrality analyses revealed that the immune system, signaling molecules, and interaction pathways were upregulated at days 1 and 3, with the highest centrality of tumor necrosis factor. Cancer, cellular community, and endocrine system pathways were downregulated, with the highest centrality of fibronectin 1 at day 5. STAT1 was upregulated from days 1 to 5 with a high centrality. The reproducibility and repeatability of the transcriptome analysis were validated by RT-qPCR. IHNV-Nagano infection dynamically changed the transcriptome profiles in the head kidney of rainbow trout and induced a defense mechanism by regulating the immune and inflammatory pathways through PRR signaling at an early stage. Downregulated pathways involved in extracellular matrix formation and focal adhesion at day 5 indicated the possible failure of wound healing, which is important in the pathogenesis of IHNV infection.

Genome-wide identification of Hsp70/110 genes in rainbow trout and their regulated expression in response to heat stress.

Heat shock proteins (Hsps) play an important role in many biological processes. However, as a typical cold water fish, the systematic identification of Hsp70/110 gene family of rainbow trout (Oncorhynchus mykiss) has not been reported, and the role of Hsp70/110 gene in the evolution of rainbow trout has not been described systematically. In this study, bioinformatics methods were used to analyze the Hsp70/110 gene family of rainbow trout. A total of 16 hsp70/110 genes were identified and classified into ten subgroups. The 16 Hsp70/110 genes were all distributed on chromosomes 2, 4, 8 and 13. The molecular weight is ranged from 78.93 to 91.39 kD. Gene structure and motif composition are relatively conserved in each subgroup. According to RNA-seq analysis of rainbow trout liver and head kidney, a total of four out of 16 genes were significantly upregulated in liver under heat stress, and a total of seven out of 16 genes were significantly upregulated in head kidney. RT-qPCR was carried out on these gene, and the result were consistent with those of RNA-seq. The significantly regulated expressions of Hsp70/110 genes under heat stress indicats that Hsp70/110 genes are involved in heat stress response in rainbow trout. This systematic analysis provided valuable information about the diverse roles of Hsp70/110 in the evolution of teleost, which will contribute to the functional characterization of Hsp70/110 genes in further research.

Possible effect of lesser galangal (Alpinia officinarum) extracts encapsulated into mesoporous silica nanoparticles on the immune status of rainbow trout (Oncorhynchusn)

Using plant extracts as immunostimulants in aquaculture has prove effectiv in resisting infectious disease, in addition to being safe and inexpensive. The present study is concerned about the prospective mode of action of using lesser galangal extract (Alpinia officinarum) encapsulated into amine surface functionalized mesoporous silica nanoparticles (MSN) in order to elevate the immune status of rainbow trout (Oncorhynchus mykiss). Fish were randomly divided into 6 groups and fed for 4 weeks with commercial diets supplemented with 0% (control), 0.5% and 1.5% of lesser galangal extract, and with previous concentrations encapsulated with MSN (0.5% + MSN, 1.5% + MSN) and MSN (without extract). The effects of the feeding trial on the innate humoural immune parameters (total protein, myeloperoxidase content, lysozyme antiproteases and bactericidal activities) in rainbow trout were examined. Also, the influence of dietary supplement on some immune-related genes in rainbow trout head kidney (TNF, IL-8, IL-1b, LYZ2 and CD4) after challenge with Yersinia ruckeri was examined. The results revealed enhancement in immune parameters in all treatment groups compared to the control, especially in the fish group fed with 1.5% + MSN which showed the highest significant difference (p < .05) in total protein, lysozyme and antiproteases activities. Also, feeding lesser galangal extract encapsulated into MSN led to an increase in myeloperoxidase content and bactericidal activity. An improvement in the expression of immune-related genes has been recorded in fish groups fed doses of lesser galangal extract and lesser galangal extract encapsulated into MSN compared to the control or to the group fed with MSN only. Particularly, the group fed with 0.5% + MSN showed a significant up-regulation in most of the immune-related genes. The current investigation supports using lesser galangal extract encapsulated into MSN in fish diets as a supplement to enhance the immune response of rainbow trout and elevate its resistance to infectious diseases.

Genome-Wide Identification of hsp90 Gene in Rainbow Trout (Oncorhynchus mykiss) and Their Regulated Expression in Response to Heat Stress.

In this study, we analyzed the gene structure, chemical characterizations, chromosome locations, evolutionary relationship, and expression profile of hsp90 genes with online database. In addition, the expression levels of hsp90s were also investigated under heat stress by quantitative real-time (qRT)-PCR. A total of eight hsp90 genes were identified from the rainbow trout genome. They were all distributed on chromosomes 2, 4, 8, and 13. The molecular weight ranged from 78.93 to 91.39 kDa, and the isoelectric point ranged from 4.84 to 4.96. The eight hsp90 genes were clustered into six subfamilies (A, B, C, D, E, and F). Genetic structure and conserved domain analysis revealed that all eight hsp90 genes had only one exon, and motif 1-motif 10 was shared by most genes. According to RNA-seq analysis of rainbow trout liver and head kidney, a total of seven out of eight genes were significantly upregulated under heat stress, and qRT-PCR was carried out on these seven genes; the expression levels of these genes were significantly upregulated under heat stress. The significantly regulated expressions of hsp90 genes under heat stress indicated that hsp90 genes are involved in heat stress response in rainbow trout. This study provides a theoretical basis for further study on the role of hsp90 in the heat stress tolerance of rainbow trout.

Recombinant VP1 and VP2 of infectious pancreatic necrosis virus trigger lymphoid cell changes and induced cytokine transcriptional expression in rainbow trout head kidney

Recombinant VP1 and VP2 of infectious pancreatic necrosis virus trigger lymphoid cell changes and induced cytokine transcriptional expression in rainbow trout head kidney

Physiological response and miRNA-mRNA interaction analysis in the head kidney of rainbow trout exposed to acute heat stress.

The rainbow trout is a cold-water fish cultured in China. Heat stress has a serious impact on the summer survival and the yield of rainbow trout. A better understanding of the regulatory response of rainbow trout to heat stress will help in determining the relationship between heat stress signaling pathways and adaption mechanisms and help contribute to breeding new high-temperature tolerant strains of rainbow trout. In this study, the 48-h median lethal temperature (48h-LT50) of rainbow trout was determined as 22.5°C. We developed control (16°C) and heat-treated (22.5°C) groups and extracted RNA from the head kidney tissues for high-throughput sequencing to study the microRNA (miRNA) expression profiles. Twelve up-regulated and five down-regulated miRNAs were identified between the control and heat-treated groups. A total of 22 target genes were predicted for 6 of the differentially expressed miRNAs, including 31 negative miRNA-mRNA interactions. Important regulatory pathways under heat stress are related to the metabolism and immune responses of the rainbow trout. Our findings provide preliminary data for investigating the high-temperature molecular mechanism of the rainbow trout and can help producers to reduce the economic losses caused by high temperature weather.

Glutamine protects against LPS-induced inflammation via adjusted NODs signaling and enhanced immunoglobulins secretion in rainbow trout leukocytes

To evaluate effects of glutamine (GLN) on fish immune responses, leukocytes were isolated from head kidney of rainbow trout and cultured in GLN-free DMEM media supplemented with different combinations of lipopolysaccharide (LPS) and GLN. LPS significantly increased expression of pro-inflammatory cytokines, while GLN supplementation alleviated LPS-induced inflammation. Leukocytes in +GLN + LPS group showed more active GLN anabolism and catabolism, which signals could be sensed by O-GlcNAcylation, and then affected LPS binding to cell surface (LBP) and adjusted NODs signaling. The mRNA expression of immunoglobulins (Igs) and their receptor (pIgR) was also significantly increased after GLN supplementation. Further analysis showed that GLN increased the percentage of IgM+ B cells and IgT+ B cells, accompanied with the increased IgM and IgT secretion in culture media, which further increased complement C3 expression to perform effector functions. All these results illustrated the regulating mechanism of GLN against LPS-induced inflammation both via adjusted NODs signaling and increased Igs+ B cells to secrete Igs.

High-throughput sequencing reveals microRNAs in response to heat stress in the head kidney of rainbow trout (Oncorhynchus mykiss).

Recently, the research of animal microRNAs (miRNAs) has attracted wide attention for its regulatory effect in the development process and the response to abiotic stresses. Rainbow trout is a commercially and cold water fish species, and usually encounters heat stress, which affects its growth and leads to a huge economic loss. But there were few investigations about the roles of miRNAs in heat stress in rainbow trout. In this study, miRNAs of rainbow trout which were involved in heat stress were identified by high-throughput sequencing of six small RNA libraries from head kidney tissues under control (18°C) and heat-treated (24°C) conditions. A total of 392 conserved miRNAs and 989 novel miRNAs were identified, of which 78 miRNAs were expressed in different response to heat stress. Ten of these miRNAs were further validated by quantitative real-time PCR. In addition to, including 393 negative correlation miRNA-target gene pairs, several important regulatory pathways were involved in heat stress of the potential target genes, including protein processing in endoplasmic reticulum, NOD-like receptor signaling pathway, and phagosome. Our data significantly advance understanding of heat stress regulatory mechanism of miRNA in the head kidney of rainbow trout, which provide a useful resource for the cultivation of rainbow trout.

Proteome Profiles of Head Kidney and Spleen of Rainbow Trout (Oncorhynchus Mykiss).

The head kidney and spleen are major lymphoid organs of the teleost fish. The authors identify proteome profiles of head kidney and spleen of rainbow trout (Oncorhynchus mykiss) using a shotgun proteomic approach. Gene ontology annotation of proteins is predicted using bioinformatic tools. This study represents detailed proteome profiles of head kidney and spleen of rainbow trout, with a total of 3241 and 2542 proteins identified, respectively. It is found that lymphoid organs are equipped with a variety of functional proteins related to defense, receptor, signal transduction, antioxidant, cytoskeleton, transport, binding, and metabolic processes. The identified proteome profiles will serve as a template for understanding lymphoid organ functions in salmonids and will increase the amount of spectra information of rainbow trout proteins in the public data repository PRIDE. This data can be accessed via ProteomeXchange with identifiers PXD008473 and PXD008478.

Protein Nanoparticles Made of Recombinant Viral Antigens: A Promising Biomaterial for Oral Delivery of Fish Prophylactics.

In the search for an eminently practical strategy to develop immunostimulants and vaccines for farmed fish, we have devised recombinant viral antigens presented as “nanopellets” (NPs). These are inclusion bodies of fish viral antigenic proteins produced in Escherichia coli. Soluble recombinant proteins are too labile to endure the in vivo environment and maintain full functionality, and therefore require encapsulation strategies. Yet when they are produced as nanostructures, they can withstand the wide range of gastrointestinal pH found in fish, high temperatures, and lyophilization. Moreover, these nanomaterials are biologically active, non-toxic to fish, cost-effective regarding production and suitable for oral administration. Here, we present three versions of NPs formed by antigenic proteins from relevant viruses affecting farmed fish: the viral nervous necrosis virus coat protein, infectious pancreatic necrosis virus viral protein 2, and a viral haemorrhagic septicemia virus G glycoprotein fragment. We demonstrate that the nanoparticles are taken up in vitro by zebrafish ZFL cells and in vivo by intubating zebrafish as a proof of concept for oral delivery. Encouragingly, analysis of gene expression suggests these NPs evoke an antiviral innate immune response in ZFL cells and in rainbow trout head kidney macrophages. They are therefore a promising platform for immunostimulants and may be candidates for vaccines should protection be demonstrated.