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The peritoneum is mainly composed of mesothelial cells, fiber cells, and fibers, with many free cells that come from tissue capillaries. Siniperca chuatsi is a valuable fish commodity and important freshwater aquaculture species in China. With the increase in the scale of its breeding, bacterial and viral disease of the S. chuatsi has become increasingly rampant in recent years. This study analyzed the peritoneal gene expression profile using transcriptome technology for the first time. The results showed that 95,711 unigenes were expressed in the peritoneum with an average length of 920.72 bp. In addition to the housekeeping genes, the genes of extracellular matrix protein 1a (ecm1), Angiopoietin-related protein 4-like (angptl4) and cysteine-rich protein 1 (crip1) were highly expressed genes in the peritoneum, and they are key factors in peritoneal development and the maintenance of morphology. Meanwhile, many immune genes were also expressed in the peritoneum, including beta-2-microglobulin (b2m), MHC II invariant chain (cd74), prostate stem cell antigen-like protein (psca), claudin (cldn), heat shock protein 90 (hsp90), complement component 4 (c4), radical S-adenosyl methionine domain-containing protein 2 (rsad2), and cathepsin S (ctss). The RT-PCR results showed that different genes exhibited different expression profiles. The transcriptome data indicated that b2m, cldn, hsp90, c4, and rsad2 were highly expressed in the peritoneum. b2m, cd74, psca, c4, and ctss were highly expressed after embryonic hatching, whereas cldn, hsp90, and rsad2 were expressed at early embryonic stages. Quantitative PCR demonstrated that b2m, cd74, cldn, hsp90, c4, and ctss genes were highly expressed after immunostimulation. This study provides significant data for studying the morphological and functional diversity of fish peritoneum and lays a foundation for research on mandarin fish immunity. [ABSTRACT FROM AUTHOR]

This review summarizes the current knowledge on immune defence activities of the European sea bass Dicentrarchus labrax by reporting the consistent amount of work done on this economically-important species. A draft genome sequence is available for this species, together with whole transcriptomes from lymphoid and nonlymphoid tissues. Available full-length coding sequences of many immunoregulatory and immune-related genes allow for targeted quantitative PCR analysis, nowadays needed for -omics data verification, ex vivo and in vitro. The first anti-T cells monoclonal antibody teleost-wise was obtained in sea bass, followed by several monoclonal and polyclonal markers of lymphocyte populations, namely T cells (pan-T, CD3e, TcR?, CD45), and B cells (IgM, IgT, IgD). The combined use of molecular and biochemical tools enabled investigations on innate and acquired immune responses of sea bass in unstimulated/stimulated fish, along the development and under variable environmental conditions and food regimes. An overview of sea bass viral and bacterial pathogens and available vaccines against these microorganisms is also provided. The knowledge accumulated in the past 25 years validates the European sea bass as a reference marine model in the field of fish immunology. [ABSTRACT FROM AUTHOR]

The CD28-B7 interaction is required to deliver a second signal necessary for T-cell activation. Additional membrane receptors of the CD28 and B7 families are also involved in immune checkpoints that positively or negatively regulate leukocyte activation, in particular T lymphocytes. BTLA is an inhibitory receptor that belongs to a third receptor family. Fish orthologs exist only for some of these genes, and the potential interactions between the corresponding ligands remain mostly unclear. In this work, we focused on the channel catfish (Ictalurus punctatus), a long-standing model for fish immunology, to analyze these co-stimulatory and co-inhibitory receptors. We identified one copy of cd28, ctla4, cd80/86, b7h1/dc, b7h3, b7h4, b7h5, two btla, and four b7h7 genes. Catfish CD28 contains the highly conserved mammalian cytoplasmic motif for PI3K and GRB2 recruitment, however this motif is absent in cyprinids. Fish CTLA4 share a C-terminal putative GRB2-binding site but lacks the mammalian PI3K/GRB2-binding motif. While critical V-domain residues for human CD80 or CD86 binding to CD28/CTLA4 show low conservation in fish CD80/86, C-domain residues are highly conserved, underscoring their significance. Catfish B7H1/DC had a long intracytoplasmic domain with a P-loop-NTPase domain that is absent in mammalian sequences, while the lack of NLS motif in fish B7H4 suggests this protein may not regulate cell growth when expressed intracellularly. Finally, there is a notable expansion of fish B7H7s, which likely play diverse roles in leukocyte regulation. Overall, our work contributes to a better understanding of fish leukocyte co-stimulatory and co-inhibitory receptors. [ABSTRACT FROM AUTHOR]

Simple Summary: The study of the innate immune system's (IIS) properties is one of the key tasks of modern immunology, since it could help enhance the quality of therapy and drug discovery. This system remains understudied for nonmammalians, and teleost fish are promising model organisms for fundamental studies of IIS organization. The recent single-cell transcriptomics (scRNAseq) experiments carried out on zebrafish and other teleosts brought fundamental knowledge about their innate immunity organization. The aim of this review is to summarize information about the available IIS-related scRNAseq experiments for teleosts and outline further perspectives for studies in this field. We found 89 scRNAseq datasets for zebrafish in different stages and organs suitable for the further meta-analysis of IIS properties and six experiments for other teleosts. Therefore, the obtained data for zebrafish could be further generalized and compared with that of mammals, while for other teleosts, we still have a large gap in knowledge. We believe that in the coming years, new scRNAseq data obtained for nonmodel teleosts will be of particular value and interest in the context of animal immunology. The innate immune system is the first line of defense in multicellular organisms. Danio rerio is widely considered a promising model for IIS-related research, with the most amount of scRNAseq data available among Teleostei. We summarized the scRNAseq and spatial transcriptomics experiments related to the IIS for zebrafish and other Teleostei from the GEO NCBI and the Single-Cell Expression Atlas. We found a considerable number of scRNAseq experiments at different stages of zebrafish development in organs such as the kidney, liver, stomach, heart, and brain. These datasets could be further used to conduct large-scale meta-analyses and to compare the IIS of zebrafish with the mammalian one. However, only a small number of scRNAseq datasets are available for other fish (turbot, salmon, cavefish, and dark sleeper). Since fish biology is very diverse, it would be a major mistake to use zebrafish alone in fish immunology studies. In particular, there is a special need for new scRNAseq experiments involving nonmodel Teleostei, e.g., long-lived species, cancer-resistant fish, and various fish ecotypes. [ABSTRACT FROM AUTHOR]

Simple Summary: Protecting the welfare of animals and the 3Rs rule are very relevance, and for this reason cell culture is an important tool. In fish it has importance in several fields such as virology, toxicology, pathology and immunology. The objective was to carry out a primary culture of the head kidneys of Atlantic salmon (Salmo salar), Pacific salmon (Oncorhynchus kisutch) and rainbow trout (Oncorhynchus mykiss), and to characterize how they respond to bacterial and viral stimuli by analyzing molecules that participate in the innate and adaptive immune response. The primary cell cultures of the head kidney (HK) from the three salmonids studied were cultured and exposed to two substances that mimic molecular patterns of different pathogens, i.e., Lipopolysaccharide (LPS) and Polyinosinic: polycytidylic acid (POLY I:C). The HK primary cell cultures from the three species grown in vitro responded differently to POLY I:C and LPS. This is the first study to demonstrate and characterize the expression of immune genes in head kidney primary cell cultures isolated from three salmonid species. It also indicates their potential role in developing immune responses as defense response agents and targets of immunoregulatory factors. Fish cell culture is a common in vitro tool for studies in different fields such as virology, toxicology, pathology and immunology of fish. Fish cell cultures are a promising help to study how to diagnose and control relevant viral and intracellular bacterial infections in aquaculture. They can also be used for developing vaccines and immunostimulants, especially with the ethical demand aiming to reduce and replace the number of fish used in research. This study aimed to isolate head kidney primary cell cultures from three Chilean salmonids: Salmo salar, Oncorhynchus kisutch, and Oncorhynchus mykiss, and characterize the response to bacterial and viral stimuli by evaluating various markers of the innate and adaptive immune response. Specifically, the primary cell cultures of the head kidney from the three salmonids studied were cultured and exposed to two substances that mimic molecular patterns of different pathogens, i.e., Lipopolysaccharide (LPS) (bacterial) and Polyinosinic: polycytidylic acid (POLY I:C). Subsequently, we determined the mRNA expression profiles of the TLR-1, TLR-8, IgM, TLR-5, and MHC II genes. Head kidney primary cell cultures from the three species grown in vitro responded differently to POLY I:C and LPS. This is the first study to demonstrate and characterize the expression of immune genes in head kidney primary cell culture isolated from three salmonid species. It also indicates their potential role in developing immune responses as defense response agents and targets of immunoregulatory factors. [ABSTRACT FROM AUTHOR]

Further studies should focus not only on the fish immune response but also on the pathogen strain, going deeper into the pathogenesis and depicting the pathogen or fish determinants for susceptibility/resistance. In addition, lamprey BCAP protein levels, and its tyrosine kinase activity, were increased in peripheral blood lymphocytes, gills, and supraneural myeloid bodies upon bacterial infection as well as via lipopolysaccharide (LPS) stimulation, but not phytohemagglutinin (PHA), thus pointing to a role in the B-cell/LPS-mediated immune response [[4]]. Fish is the first group of vertebrates to appear during animal evolution. [Extracted from the article]

Cuttlefish are sensitive to the breeding environment, and the low-salinity environment significantly impacts their growth and immunity. So far, it is difficult to breed this species artificially. This study was conducted in Sepia lycidas. And the aim was to investigate the differences in protein phosphorylation in the intestine of S. lycidas under different salinity conditions. Firstly, 999 phosphoproteins (specific peptide = 1), 1928 phosphopeptides, and 2727 phosphorylation sites were identified. Among them were 284 down-regulated expression phosphorylation sites (corresponding to 115 phosphoproteins) and 674 up-regulated expression phosphorylation sites (corresponding to 408 phosphoproteins) in the intestine under a low salinity environment compared with that under a natural salinity environment. Next, GO analysis found that more phosphoproteins corresponding to differentially expressed phosphorylation sites were related to anatomical structure development, multicellular organism development, regulation of the cellular process, etc. The molecular functions of these proteins mainly contain protein binding, transferase activity, catalytic activity, and heterocyclic compound binding. And they are mainly involved in the cellular components of intracellular anatomical structure, organelle, and cytoplasm. KEGG enrichment analysis of the differential phosphoproteins suggested that many significantly enriched pathways were related to the phosphatidylinositol signaling system, cell junction (adherens junction and tight junction), and inositol phosphate metabolism. Finally, changes in environmental salinity can affect the intestinal structure, metabolism, and immune homeostasis of S. lycidas. [ABSTRACT FROM AUTHOR]

Complement (C) is a serum protein with immunity-related functions and enzymatic activities. It can induce inflammatory responses and antibody formation following activation by substances such as pathogens. Moreover, it is a crucial molecule in the congenital immune defense system. Complement manifests earlier than immunoglobulin in the evolutionary process of fish's immune system. Therefore, the complement system of fish is vital for congenital and acquired immunities. A complement system is a polymolecule system comprising more than 30 soluble proteins, membrane-bound proteins, and complement receptors. It can be activated through three pathways for immunoregulation, the clearance of the immune complex, etc. The liver primarily produces the fish complement C3, which is a significant component of the complement system of fishes. C3 is the hub of three activation pathways. Research works concerning the structure, generation process, expression, and functions of C3 in fishes are reviewed in this study. [ABSTRACT FROM AUTHOR]

In recent years, studies on circadian control in immunity have been actively conducted in mammals, but little is known about circadian rhythms in the field of fish immunology. In this study, we aimed to analyse the regulation of the diel oscillation of inflammatory cytokine interleukin-1β (il1b) gene expression by core components of the circadian clock in Japanese medaka (Oryzias latipes). The expression of il1b and clock genes (bmal1 and clock1) in medaka acclimated to a 12:12 light (L): dark (D) cycle showed diel rhythm. Additionally, higher expression of il1b was detected in medaka embryo cells (OLHdrR-e3) overexpressing bmal1 and clock1. A significant decrease in il1b expression was observed in OLHdrR-e3 cells after bmal1 knockdown using morpholino oligos. These changes may be mediated by transcriptional regulation via clock proteins, which target the E-box sequence in the cis -element of il1b as identified using luciferase reporter assays. Moreover, LPS stimulation and pathogenic bacterial infection at different zeitgeber time (ZT) under LD12:12 conditions affected the degree of il1b expression, which showed high and low responsiveness to both immuno-stimulations at ZT2 and ZT14, respectively. These results suggested that fish IL-1β exhibited diel oscillation regulated by clock proteins, and its responsiveness to immune-stimulation depends on the time of day. • Medaka il1b is expressed in a circadian rhythm. • Medaka clock genes bmal1 and clock1 regulate il1b expression. • Immuno-stimulation at different time point impacts il1b expression magnitude. [ABSTRACT FROM AUTHOR]

Pompano fishes have been widely farmed worldwide. As a representative commercial marine species of the Carangidae family, the golden pompano (Trachinotus blochii) has gained significant popularity in China and worldwide. However, because of rapid growth and high-density aquaculture, the golden pompano has become seriously threatened by various diseases. Cell lines are the most cost-effective resource for in vitro studies and are widely used for physiological and pathological research owing to their accessibility and convenience. In this study, we established a novel immortal cell line, GPF (Golden pompano fin cells). GPF has been passaged over 69 generations for 10 months. The morphology, adhesion and extension processes of GPF were evaluated using light and electron microscopy. GPF cells were passaged every 3 days with L-15 containing 20 % fetal bovine serum (FBS) at 1:3. The optimum conditions for GPF growth were 28 °C and a 20 % FBS concentration. DNA sequencing of 18S rRNA and mitochondrial 16S rRNA confirmed that GPF was derived from the golden pompano. Chromosomal analysis revealed that the number pattern of GPF was 48 chromosomes. Transfection experiments demonstrated that GPF could be utilized to express foreign genes. Furthermore, heavy metals (Cd, Cu, and Fe) exhibited dose-dependent cytotoxicity against GPF. After polyinosinic-polycytidylic acid (poly I:C) treatment, transcription of the retinoic acid-inducible gene I-like receptor (RLR) pathway genes, including mda5 , mita , tbk1 , irf3 , and irf7 increased, inducing the expression of interferon (IFN) and anti-viral proteins in GPF cells. In addition, lipopolysaccharide (LPS) stimulation up-regulated the expression of inflammation-related factors, including myd88 , irak1 , nfκb , il1β , il6 , and cxcl10 expression. To the best of our knowledge, this is the first study on the immune response signaling pathways of the golden pompano using an established fin cell line. In this study, we describe a preliminary investigation of the GPF cell line immune response to poly I:C and LPS, and provide a more rapid and efficient experimental material for research on marine fish immunology. • GPF was established and became a stable fin cell line of Trachinotus blochii. • GPF can be used to assess the cytotoxicity of heavy metals. • GPF can be utilized to express foreign genes with 32 % transfection efficiency. • GPF's immune response to LPS and poly I: C challenge proved it can be used for in vitro immunology study. [ABSTRACT FROM AUTHOR]

Antimicrobial peptides (AMPs) are an essential part of the vertebrate innate immune system. Piscidins are a family of AMPs specific in fish. In our previous investigation, we identified four paralogous genes of piscidins in the orange-spotted grouper (Epinephelus coicodes), which exhibited distinct activities against bacteria, fungi, and parasitic ciliated protozoa. Piscidins demonstrated their capability to modulate the expression of diverse immune-related genes; however, their precise immunoregulatory functions remain largely unexplored. In this study, we examined the immunomodulatory properties of putative mature peptides derived from four E. coicodes piscidins (ecPis1S, ecPis2S, ecPis3S, and ecPis4S) in head kidney leukocytes (HKLs) or monocytes/macrophages (MO/MΦ)-like cells isolated from E. coicodes. Our data demonstrate that E. coicodes piscidins exhibit immunomodulatory activities supported by multiple lines of evidence. Firstly, all four piscidins displayed chemotactic activities towards HKLs, with the most potent chemotactic activity observed in ecPis2S. Secondly, stimulation with E. coicodes piscidins enhanced respiratory burst and phagocytic activity in MO/MФ-like cells, with ecPis3S showing the highest efficacy in increasing phagocytosis of MO/MΦ-like cells. Thirdly, mRNA expression levels of chemokine receptors, Toll-like receptors, T cell receptors, and proinflammatory cytokines were modulated to varying extents by the four piscidins in E. coicodes HKLs. Overall, our findings indicate that the immunological activities of these four paralogous piscidins from E. coicodes are exhibited in a paralog-specific and concentration-dependent manner, highlighting their distinct and versatile immunomodulatory properties. This study makes a significant contribution to the field of fish AMPs immunology by elucidating the novel mechanisms through which members of the piscidin family exert their immunomodulatory effects. Moreover, it provides valuable insights for further exploration of fish immunomodulating agents. • E. coicodes paralogous piscidins show the versatile immunomodulatory activities. • E. coicodes piscidin 2 exhibits the strongest chemotactic activity for leucocytes. • E. coicodes piscidin 3 shows the strongest activity to enhance the phagocytosis of MO/MΦ. • Piscidins regulate expression of immune-related genes widely and in paralog-specific manner. [ABSTRACT FROM AUTHOR]

A permanent epithelial‐like cell line, named GrB, was established from brain tissues of the rare minnow (Gobiocypris rarus). GrB cells grow well at temperatures between 22℃ and 27℃ in L‐15 medium supplemented with 10% FBS. Chromosome analysis revealed that GrB cells possessed a diploid chromosome number of 2n = 50 at passage 50, consistent with the number of chromosomes reported for G. rarus. Partial amplification and sequence analysis of the cytochrome oxidase subunit I (COI) confirmed that GrB cells are originated from G. rarus. The cell lines also successfully expressed green fluorescence protein from plasmid pEGFP‐N1 and which makes them suitable for transfection studies. A cytolytic cytopathic effect was observed in the cells after infection with tilapia lake virus 2017A (TiLV‐2017A). A large number of virus particles were detected by electron microscopy. Immunofluorescence and virus titration assays indicated that TiLV‐2017A titres reached 105.67 TCID50/ml at 7 days post‐infection (dpi). Healthy adult rare minnows were challenged with virus obtained from these cells showed that they can be infected by TiLV‐2017A, which causes low mortality but clinical signs. The GrB cell line may represent a valuable model as basics for fish virology and fish immunology. [ABSTRACT FROM AUTHOR]

Granulins are conserved in nearly all metazoans, with an intriguing loss in insects. These pleiotropic peptides are involved in numerous physiological and pathological processes yet have been overwhelmingly examined in mammalian systems. While work in other animal models has been informative, a richer understanding of the proteins should be obtained by integrating knowledge from all available contexts. The main bodies of work described here include 1) the structure-function relationships of progranulin and its cleavage products, 2) the role of expanded granulin gene families and different isoforms in fish immunology, 3) the release of granulin peptides to promote host angiogenesis by parasitic worms, 4) a diversity of molluscan uses for granulins, including immune activation in intermediate hosts to trematodes, 5) knowledge gained on lysosomal functions from C. elegans and the stress-related activities of granulins. We provide an overview of functional reports across the Metazoa to inform much-needed future research. • Granulins are conserved and pleiotropic, yet the evolution of functions is unclear. • Functional patterns and novelties across animal granulins are detailed. • The loss of granulins in some insects is addressed. • Granulin can be released from parasites and hosts simultaneously. • Future directions for each type of organism are suggested throughout. [ABSTRACT FROM AUTHOR]

Miodrag (Mike) Belosevic and collaborators profoundly influenced the development of primary kidney macrophage culturing system (PKM) to study fish immunology in various aspects of comparative immunology. Their application of using PKM model, opened a new path for studying the development of macrophages, regulation of hematopoiesis, and cell specific response against various pathogens. By measuring histopathological and immunological outcomes, the biological implications of a variety of cytokines and signal transduction molecules could be elucidated with the established PKM system. A variety of growth factors mediating hematopoiesis and cytokines regulating the immune responses were functionally characterized, which served as a fundamental basis for making goldfish an excellent model to study fish immunology. Specifically, using in vivo and PKM based in vitro assays, the Belosevic lab advanced the goldfish- M. marinum model to study the anti-mycobacteria responses in teleosts, thus paving a way for the development of novel therapeutic approaches which could be applied in aquaculture settings or utilized as a model for human disease. In this review, we will look at the contribution of Dr. Mike Belosevic to teleost macrophage development, multiple cytokine functional characterization, and host-pathogen interactions. • Distinct macrophage sub-populations originating from goldfish kidney leukocyte cultures were generated and characterized. • The PKM system of goldfish can serve as a model to investigate teleost hematopoiesis and host-pathogen interactions. • Goldfish was used as a model organism to study the anti-mycobacterial responses in teleosts. [ABSTRACT FROM AUTHOR]

This study evaluated the dietary supplementation of ginger Zingiber of ficinale essential oil for the Neotropical catfish Pseudoplatystoma reticulatum, its effects on fish hematology, immunology, the response of these variables to induction of stress (exposition to air for 3 min), and a bacterial challenge with Aeromonas hydrophila, and the effects on liver histomorphometry. Three levels of the ginger essential oil were evaluated (0.5, 1.0 and 1.5%) in addition to a control group (0%) in a feeding trial for 65 days in triplicate. Fish hematology and immunology were assessed at five different moments: after the feeding trial (0), after the stress induction (stress), and three points after the bacterial challenge (3, 6 and 24 h post-challenge). Fish fed the ginger essential oil supplemented diets did not exhibit significant differences between the hematocrit at the 0-sample point and the 24 h post-challenge point (P = 0.05), while fish fed the control diet presented significantly lower values for the hematocrit when comparing those same two moments (P < 0.05). The thrombocytes count for fish fed the 0.5% essential oil supplemented diet was significantly higher than all other groups after the feeding trial (P < 0.05), and their total leukocytes count after the stress induction was also significantly higher than the other treatments (P < 0.05). Although the results mentioned above indicate that the ginger essential oil had positive health effects on the fish, no significant differences in survival after the challenge with A. hydrophila were observed (P = 0.05). Lastly, histomorphometry suggests that the essential oil did not negatively impact fish hepatocytes. [ABSTRACT FROM AUTHOR]

In this study, we investigated a new application of bubble-eye goldfish (commercially available strain with large bubble-shaped eye sacs) for immunological studies in fishes utilizing the technical advantage of examining immune cells in the eye sac fluid ex vivo without sacrificing animals. As known in many aquatic species, the common goldfish strain showed an increased infection sensitivity at elevated temperature, which we demonstrate may be due to an immune impairment using the bubble-eye goldfish model. Injection of heat-killed bacterial cells into the eye sac resulted in an inflammatory symptom (surface reddening) and increased gene expression of pro-inflammatory cytokines observed in vivo, and elevated rearing temperature suppressed the induction of pro-inflammatory gene expressions. We further conducted ex vivo experiments using the immune cells harvested from the eye sac and found that the induced expression of pro-inflammatory cytokines was suppressed when we increased the temperature of ex vivo culture, suggesting that the temperature response of the eye-sac immune cells is a cell autonomous function. These results indicate that the bubble-eye goldfish is a suitable model for ex vivo investigation of fish immune cells and that the temperature-induced infection susceptibility in the goldfish may be due to functional impairments of immune cells. [ABSTRACT FROM AUTHOR]

Nile tilapia (Oreochromis niloticus) is a freshwater fish, which is extensively cultivated worldwide and constitutes one of the model species for the study of fish immunology. Monoclonal antibodies are very advantageous molecular tools for studying teleost immune system. Specifically, monoclonal antibodies that react with immunoglobulins are used successfully in the study of the humoral immune response of several fish species. In the present study, we produced and characterized a monoclonal antibody against tilapia IgM heavy chain using a peptide-based strategy. The peptide sequence was selected from the surface-exposed region between CH3–CH4 domains. The specificity of the polyclonal serum and the hybridoma culture supernatant obtained by immunization with the peptide conjugated to keyhole limpet hemocyanin were evaluated by western blotting, both showing reactivity against tilapia serum IgM. The purified mAb was able to recognize secreted IgM by western blotting and ELISA and membrane IgM by flow cytometry. We also demonstrated that the antibody doesn't cross-react with a recombinant IgT fragment. This tool allowed us to study for the first time the stimulation of mucosal immunity after Pituitary Adenylate Cyclase Activating Polypeptide administration. Overall, the results demonstrated the utility of this mAb to characterize humoral immune response in O. niloticus. • MAb against Nile tilapia IgM using a peptide-based strategy was produced and characterized. • Nile tilapia IgM purified from serum was successfully identified by mass spectrometry analysis. • The produced mAb was specific for IgM and doesn't cross-react with a recombinant IgT fragment. • The prepared mAb is applicable for IgM detection and quantification by western blotting, ELISA and flow cytometry. • IgM and IgT concentrations increased in tilapia skin mucus after injection of synthetic C. gariepinus PACAP-38. [ABSTRACT FROM AUTHOR]

The bulbus arteriosus tissue of teleosts, which is located at the forefront of the heart, is used to reduce the pulse pressure. In this study, we constructed a permanent cell line (LmAB) for the first time using bulbus arteriosus tissue from spotted sea bass (Lateolabrax maculatus). This cell line has been passaged more than 80 times. Currently, it can be subcultured in L-15 medium with 8 % fetal bovine serum added. The optimal fetal bovine serum concentration and culture temperature for LmAB cells at 62 passages are 20 % and 28 °C, respectively. This cell line consists predominantly of epithelial-like cells. We used 18S rRNA gene sequencing to confirm that LmAB cells originated from spotted sea bass. Karyotype analysis revealed that 43 % of LmAB cells in passage 63 had 48 chromosomes. Exogenous plasmid transfection revealed that LmAB cells can express the green fluorescent protein gene with a transfection efficiency of up to 40 %, indicating that these cells can be used for in vitro genetic research. LmAB cells showed susceptibility to nervous necrosis virus, largemouth bass ulcer syndrome virus, and infectious spleen and kidney necrosis virus, which results in severe cytopathic effects. PCR analysis verified that these viruses can replicate in LmAB cells, and analysis of cytoskeletal F-actin patterns verified that infected cells exhibit serious changes in their actin cytoskeleton. LmAB cells infected with these three viruses showed increased expressions of interferon signaling pathway genes (IFNd , IFNγ-rel , and ISG15), indicating that the host interferon signaling pathway participates in the antiviral immune response. These findings indicate that our newly developed LmAB cell line is a valuable resource for future research in genetics, virology, and immunology. • A novel cell line (LmAB) from bulbus arteriosus tissue in L. maculatus have been established. • LmAB cells are susceptible to LBUSV, NNV and ISKNV virus. • LmAB cells infected viruses exhibit serious changes in their actin cytoskeleton. • Viral infection upregulates the expression of interferon related genes in LmAB cells. [ABSTRACT FROM AUTHOR]

Edwardsiella tarda (E. tarda), an intracellular pathogen, has caused severe economic losses in aquaculture. Effective vaccine development for E. tarda prevention is urgently needed. A previous study indicates that cell-mediated immunity (CMI) might play an important role in E. tarda infection. We believe that the involvement of allograft rejection and CMI has now been well documented in mammals and some fishes. However, there is still little research on the application of blood allograft rejection in vaccine development. In the current study, we investigate the immune response and vaccine effect in fish vaccinated with allogeneic blood + formalin-killed cells vaccine (FKC), allogeneic blood + phosphate-buffered saline (PBS), PBS + FKC and PBS + PBS. In the challenge test, the relative percentage survival (RPS) of the allogeneic + FKC, the allogeneic blood + PBS and the PBS + FKC group was 61.46, 35.41, and 30.63 % respectively. The up-regulated expression of Th1-related genes IFN- γ 1, IFN- γ 1rel2, IL-12p35 and T-bet suggests the protection is via CMI induction. Only in the allogeneic + FKC group, gene expression of IFN- γ 1, IL-12p35 and T-bet is significantly higher, indicating synergy between the two substances. Furthermore, among the fish injected with the allogeneic blood cells, syngeneic blood cells and PBS group, only in the fish of the allogenic blood cells injection group, did expression of IFN- γ 1, IFN- γ 2 and IFN- γ rel2 gene expression significantly increased. The results indicate that the rejection was induced by allogeneic components. Thus, our findings might provide essential information and insights into vaccine development in aquaculture. • Allogeneic adjuvants with FKC showed high survival rates after challenge with Edwardsiella tarda. • Allogeneic adjuvants are potential for CMI induction. • Allogeneic blood cells could elevate survival rate after challenge with E. tarda. [ABSTRACT FROM AUTHOR]

GATA3 belongs to the GATA family, and it could interact with the target gene promoter. It has been reported to play a central role in regulating lymphocyte differentiation. In this study, the GATA3 cDNA sequence was identified by a homologous clone and the RACE technology from Japanese flounder (Paralichthys olivaceus). The full-length of the GATA3 cDNA sequence was 2904 bp, including 1332 bp open reading frame (ORF), 265 bp 5 ′-untranslated region (5′ UTR), and 1308 bp 3 ′-UTR, encoding 443 amino acids. GATA3 protein sequence was conserved in vertebrates and invertebrates, including two zinc finger domains. qRT-PCR showed that the expression of GATA3 was high in the gill, kidney, and spleen. Expression of GATA3 slowly increased at the earlier stages and culminated at the late gastrula and somatic stages. Immunohistochemistry (IHC) results showed that the GATA3 protein was expressed in lymphocyte cells, undifferentiated basal and pillar cells of the gills, as well as lymphocyte cells and melanin macrophages of the kidney. The expression of GATA3 was significantly regulated in tissues and different types of lymphocytes after stimulation with Edwardsiella tarda. Dual-luciferase reporter assay indicated that the GATA3 protein could directly interact with promoters of target genes involved in the immune response. These findings suggested that GATA3 plays a major role in regulating the immune response. This study provided a theoretical basis for the immune response mechanism of teleost and a useful reference for later research on fish immunology. • The full-length of GATA3 cDNA was identified from Japanese flounder. • GATA3 was highly expressed in the gill. • A positive signal of GATA3 was detected in lymphocyte-like cells of the gill and kidney. • GATA3 was significantly down-regulated after being challenged with Edwardsiella tarda in tissues and lymphocytes. • GATA3 could interact with target genes involved in immune response as a transcription factor. [ABSTRACT FROM AUTHOR]

Increasing water temperatures due to climate change have resulted in more frequent high mortality events of New Zealand Greenshell™ mussels (Perna canaliculus Gmelin 1791). These events have significant impacts within mussel farms which support a major shellfish industry for New Zealand. The present study investigates metabolic responses of farmed mussels during a summer mortality event in order to identify health impacts and elucidate mechanistic effects of external stressors on mussels. A gas chromatography–mass spectrometry (GC–MS)-based metabolomics approach was used to identify metabolic perturbations and flow cytometry assays were used to assess viability, oxidative stress and apoptosis of haemocytes from healthy and unhealthy mussels during a summer mortality event. The results showed significantly higher mortality and apoptosis of haemocytes in unhealthy mussels compared to healthy mussels. Reactive oxygen species (ROS) production, which is an indicator of oxidative stress was very high in both mussel groups, but no differences were observed between the two mussel groups. Metabolomics revealed alterations of many metabolites in both haemolymph and hepatopancreas (digestive gland) of unhealthy mussels compared to healthy mussels, reflecting perturbations in several molecular pathways, including energy metabolism, amino acid metabolism, protein degradation/tissue damage and oxidative stress. An increased level of itaconic acid which is an antimicrobial metabolite and biomarker of pathogen infection was observed in haemolymph, but not in hepatopancreas samples. This investigation provides the first detailed metabolic characterization of mussel immune responses to a summer mortality event and illustrates the benefits of using an integrated metabolomics and flow cytometry workflow for mussel health assessment and biomarker identification for summer mortality early detection. Image 1 • Unknown summer mortality causes perturbations in mussel metabolism. • Metabolomics allows classification of mussels from different affected levels. • Accumulation of itaconic acid may indicate the involvement of pathogen infections. • Itaconic acid has potential to be a biomarker for pathogen infection. [ABSTRACT FROM AUTHOR]

Like terrestrial vertebrates, bony fishes have a nasopharynx-associated lymphoid tissue (NALT) that protects the host against invading pathogens. Despite nasal immunity being a relatively new field in fish immunology, the investigation of nasal immune systems has already illuminated fundamental aspects of teleost mucosal immune systems as well as neuroimmunology. In this review, we highlight the importance of nasal infections in bony fish and the progress that has been made towards understanding how fish respond locally and systemically to nasal infection or vaccination. We also want to highlight the complex interactions between neurons and immune cells that occur in the olfactory organ during the course of an immune response. We predict that similar neuroimmune interactions govern immune responses at all mucosal tissues in bony fish. Understanding the principles of mucosal immune responses in teleost NALT has therefore revealed important aspects of fish mucosal immunity that are critical for mucosal vaccination in aquaculture. • Teleosts have a nasopharynx-associated lymphoid tissue (NALT). • Rainbow trout NALT is now well-studied in the context of nasal vaccination. • Nasal vaccination has unveiled fundamental neuroimmune interactions in teleosts. • Repertoire analyses of fish NALT reveal local B cell responses to vaccination. [ABSTRACT FROM AUTHOR]

Mucosal surfaces such as fish gills interface between the organism and the external environment and as such are major sites of foreign Ag encounter. In the gills, the balance between inflammatory responses to waterborne pathogens and regulatory responses toward commensal microbes is critical for effective barrier function and overall fish health. In mammals, IL-4 and IL-13 in concert with IL-10 are essential for balancing immune responses to pathogens and suppressing inflammation. Although considerable progress has been made in the field of fish immunology in recent years, whether the fish counterparts of these key mammalian cytokines perform similar roles is still an open question. In this study, we have generated IL-4/13A and IL-4/13B mutant zebrafish (Danio rerio) and, together with an existing IL-10 mutant line, characterized the consequences of loss of function of these cytokines. We demonstrate that IL-4/13A and IL-4/13B are required for the maintenance of a Th2-like phenotype in the gills and the suppression of type 1 immune responses. As in mammals, IL-10 appears to have a more striking anti-inflammatory function than IL-4-like cytokines and is essential for gill homeostasis. Thus, both IL-4/13 and IL-10 paralogs in zebrafish exhibit aspects of conserved function with their mammalian counterparts. [ABSTRACT FROM AUTHOR]

Proliferative kidney disease (PKD), caused by the myxozoan Tetracapsuloides bryosalmonae, is one of the most serious parasitic diseases of salmonids in which outbreaks cause severe economic constraints for the aquaculture industry and declines of wild species throughout Europe and North America. Given that rainbow trout (Oncorhynchus mykiss) is one of the most widely farmed freshwater fish and an important model species for fish immunology, most of the knowledge on how the fish immune response is affected during PKD is from this organism. Once rainbow trout are infected, PKD pathogenesis results in a chronic kidney immunopathology mediated by decreasing myeloid cells and increasing lymphocytes. Transcriptional studies have revealed the regulation of essential genes related to T‐helper (Th)‐like functions and a dysregulated B‐cell antibody type response. Recent reports have discovered unique details of teleost B‐cell differentiation and functionality and characterized the differential immunoglobulin (Ig)‐mediated response. These studies have solidified the rainbow trout T. bryosalmonae system as a sophisticated disease model capable of feeding key advances into mainstream immunology and have contributed essential information to design novel parasite disease prevention strategies. In our following perspective, we summarize these efforts to evaluate the immune mechanisms of rainbow trout during PKD pathogenesis. [ABSTRACT FROM AUTHOR]

In Chile, the salmon and trout farmed fishing industries have rapidly grown during the last years, becoming one of the most important economic sources for the country. However, infectious diseases caused by bacteria, virus, mycoses and parasites, result in losses of up to 700 million dollars per year for the Chilean aquaculture production with the consequent increase of antibiotic and antiparasitic usage. After 30 years of its first appearance, the main salmon health problem is still the salmonid rickettsial septicaemia (SRS), which together with other disease outbreaks, reveal that vaccines do not provide acceptable levels of long-lasting immune protection in the field. On the other hand, due to the large dependence of the industry on salmonids production, the Chilean government promoted the Aquaculture diversification program by 2009, which includes new species such as Merluccius australis , Cilus gilberti and Genypterus chilensis , however, specific research regarding the immune system and vaccine development are issues that still need to be addressed and must be considered as important as the farm production technologies for new fish species. Based on the experience acquired from the salmonid fish farming, should be mandatory an effort to study the immune system of the new species to develop knowledge for vaccination approaches, aiming to protect these aquaculture species before diseases outbreaks may occur. This review focuses on the current status of the Chilean aquaculture industry, the challenges related to emerging and re-emerging microbial pathogens on salmonid fish farming, and the resulting needs in the development of immune protection by rational designed vaccines. We also discussed about what we have learn from 25 years of salmonid researches and what can be applied to the new Chilean farmed species on immunology and vaccinology. • Summarize the epidemiological situation in the Chilean aquaculture. • Review the surveillance of emerging and re-emerging infectious diseases. • Characterize vaccines against piscirickettsiosis, infectious salmon anemia, infectious pancreatic necrosis and caligidosis. • Adaptive immune response in salmonids need a deep understanding of cellular and molecular interactions. • Discuss future trends for fish immunology and vaccinology in Chile. [ABSTRACT FROM AUTHOR]

Polyinosinic:polycytidylic acid [poly (I:C)] was administered in vivo to Chinook salmon (Oncorhynchus tshawytscha) post‐smolts to determine the immune responses on haematological and cellular functional parameters, including spleen (SP), head kidney (HK) and red blood cell (RBC) cytokine expression, as well as serum metabolomics. Poly (I:C) in vivo (24 h exposure) did not affect fish haematological parameters, leucocyte phagocytic activity and phagocytic index, reactive oxygen species and nitric oxide production. Gas chromatography–mass spectrometry–based metabolomics revealed that poly (I:C) significantly altered the serum biochemistry profile of 25 metabolites. Metabolites involved in the branched‐chain amino acid/glutathione and transsulphuration pathways and phospholipid metabolism accumulated in poly (I:C)–treated fish, whereas those involved in the glycolytic and energy metabolism pathways were downregulated. At cytokine transcript level, poly (I:C) induced a significant upregulation of antiviral ifnγ in HK and Mx1 protein in HK, SP and RBCs. This study provides evidence for poly (I:C)–induced, immune‐related biomarkers at metabolic and molecular levels in farmed O. tshawytschain vivo. These findings provide insights into short‐term effects of poly (I:C) at haematological, innate and adaptive immunity and metabolic levels, setting the stage for future studies. [ABSTRACT FROM AUTHOR]

Evaluating studies over the past almost 40 years, this review outlines the current knowledge and research gaps in the use of isolated leucocytes in salmonid immunology understanding. This contribution focuses on the techniques used to isolate salmonid immune cells and popular immunological assays. The paper also analyses the use of leucocytes to demonstrate immunomodulation following dietary manipulation, exposure to physical and chemical stressors, effects of pathogens and parasites, vaccine design and application strategies assessment. We also present findings on development of fish immune cell lines and their potential uses in aquaculture immunology. The review recovered 114 studies, where discontinuous density gradient centrifugation (DDGC) with Percoll density gradient was the most popular leucocyte isolation method. Fish head kidney (HK) and peripheral blood (PB) were the main sources of leucocytes, from rainbow trout (Oncorhynchus mykiss) and Atlantic salmon (Salmo salar). Phagocytosis and respiratory burst were the most popular immunological assays. Studies used isolated leucocytes to demonstrate that dietary manipulations enhance fish immunity, while chemical and physical stressors suppress immunity. In addition, parasites, and microbial pathogens depress fish innate immunity and induce pro-inflammatory cytokine gene transcripts production, while vaccines enhance immunity. This review found 10 developed salmonid cell lines, mainly from S. salar and O. mykiss HK tissue, which require fish euthanisation to isolate. In the face of high costs involved with density gradient reagents, the application of hypotonic lysis in conjunction with mico-volume blood methods can potentially reduce research costs, time, and using nonlethal and ethically flexible approaches. Since the targeted literature review for this study retrieved no metabolomics study of leucocytes, indicates that this approach, together with traditional technics and novel flow cytometry could help open new opportunities for in vitro studies in aquaculture immunology and vaccinology. Image 1 • Leucocytes mostly isolated by discontinuous density gradient centrifugation (DDGC). • Head kidney (HK) and peripheral blood (PB) were the main sources of leucocytes. • Phagocytosis and respiratory burst are the most popular immunological assays. • Isolated leucocytes show that dietary manipulations and vaccines enhance immunity. • Leucocytes reveal that chemical, physical & pathogenic stressors suppress immunity. [ABSTRACT FROM AUTHOR]

We investigated cellular functional and targeted immune cytokine responses of farmed Chinook salmon (Oncorhynchus tshawytscha) peripheral blood mononuclear cells (PBMCs) in vitro to LPS from Escherichia coli (E. coli) serotypes O111: B4 and O55: B5, and a phorbol ester phorbol 12-myristate 13-acetate (PMA). Bacterial LPS and PMA significantly (p < 0.05) induced reactive oxygen species (ROS) production in O. tshawytscha PBMCs, and enhanced by interferon (IFN)-inducible cytokine production. Cellular phagocytosis was significantly enhanced with PMA and E. coli serotype O111: B4 LPS after 1 and 2 h respectively. At the molecular level, LPS and PMA significantly (p < 0.05) upregulated pro-inflammatory cytokine gene transcripts for IFNγ, TNF-α, and anti-inflammatory IL-10, 24 h post-stimulation. This response is postulated to be mediated via the MyD88 and TRIF pathways in TLR4, or synergistic TLR1 and TLR2 receptors. This is the first report of LPS induced immune related in vitro responses in farmed O. tshawytscha PBMCs. • Bacterial LPS and PMA significantly stimulated fish PBMC ROS production. • LPS and PMA significantly enhanced chinook salmon PBMC phagocytosis. • LPS and PMA upregulated PBMC pro-inflammatory IFNγ and TNF-α cytokines. • LPS and PMA stimulated PBMC ant-inflammatory IL-10 cytokine production. [ABSTRACT FROM AUTHOR]

Apolipoprotein A-I (ApoA-I) is a lipoprotein involved in a variety of physiological and pathological processes. However, the immunomodulatory functions of ApoA-I in fish are not well understood. In this study, ApoA-I from Nile tilapia (Oreochromis niloticus) (On-ApoA-I) was identified, and its function in bacterial infection was investigated. The open reading frame of On-ApoA-I is 792 bp, which codes for a protein containing 263 amino acids. On-ApoA-I shared over 60% sequence similarity with other teleost fish and more than 20% with mammalian ApoA-I. On-ApoA-I was found to be highly expressed in the liver and significantly induced during Streptococcus agalactiae infection by qRT‒PCR analysis. Furthermore, in vivo studies revealed that recombinant On-ApoA-I protein could suppress inflammation and apoptosis and improve the likelihood of surviving bacterial infection. Additionally, On-ApoA-I showed in vitro antimicrobial properties against Gram-positive and Gram-negative bacteria. These findings offer a theoretical basis for further investigations into the role of ApoA-I in fish immunology. • Apolipoprotein A-I (ApoA-I) was identified from Nile Tilapia (Oreochromis niloticus). • ApoA-I was induced during bacterial infection. • ApoA-I protects tilapia against S. agalactiae infection in vivo. [ABSTRACT FROM AUTHOR]

The Yellow River carp (Cyprinus carpio haematopterus) is a vital economically farmed fish of the Cyprinidae family. With the development of intensive aquaculture, carp production has increased dramatically, leading to the frequent occurrence of various diseases. Cell lines are considered the most cost-effective resource for in vitro studies and are widely used for physiological and pathological studies because of accessibility and convenience. This research established a novel immortal cell line CCM (Yellow River carp muscle cells) derived from the carp muscle. CCM has been passed over 71 generations for 1 year. The morphology of CCM and the adhesion and extension processes were captured by light and electron microscopy. CCM were passaged every 3 days with 20% FBS DMEM/F12 at 1:3. The optimum conditions for CCM growth were 28 °C and 20% FBS concentration. DNA sequencing of 16S rRNA and COI showed that CCM was derived from carp. CCM positively reacts to anti-PAX7 and anti-MyoD antibodies of carp. Analysis of chromosomes revealed that the chromosomal pattern number of CCM was 100. Transfection experiment demonstrated that CCM might be utilized to express foreign genes. Furthermore, cytotoxicity testing showed that CCM was susceptible to Aeromonas hydrophila , Aeromonas salmonicida , Aeromonas veronii , and Staphylococcus Aureus. The organophosphate pesticides (chlorpyrifos and glyphosate) or heavy metals (Hg, Cd, and Cu) exhibited dose-dependent cytotoxicity against CCM. After LPS treatment, the MyD88-IRAKs-NFκB pathway stimulates inflammatory-related factor il1β , il8 , il10 , and nfκb expression. LPS did not seem to cause oxidative stress in CCM, and the expression of cat and sod was not affected. Poly (I:C) through TLR3-TRIF-MyD88-TRAF6-NFκB and TRIF-TRAF3-TBK1-IRF3 activated the transcription of related factors, increased expression of anti-viral protein, but no changes in apoptosis-related genes. To our knowledge, this is the first muscle cell line in Yellow River carp and the first study on the immune response signal pathways of Yellow River carp based on the muscle cell line. CCM cell line provides a more rapid and efficient experimental material for fish immunology research, and this study preliminarily elucidated its immune response strategy to LPS and poly (I:C). • CCM was established and became the first stable muscle cell line of Yellow River carp. • CCM positively reacts to anti-PAX7 and anti-MyoD antibodies of Yellow River carp. • CCM can be used to assess the cytotoxicity of pesticides and heavy metals and are sensitive to bacterial. • Immune-responsive pathways in response to LPS and poly (I: C) challenge in CCM were evidenced. • CCM cell line can be used as a cell model for virology and immunology. [ABSTRACT FROM AUTHOR]

Since the 2000 s, the pearl oyster Pinctada imbricata (Röding, 1798) has become established along the transitional waterways of the "Capo Peloro Lagoon" natural reserve, where it is now abundant due to its adaptability to different hydrological, climatic, environmental, and pollution conditions. This study aims to evaluate haemocyte immune-mediated responses in vitro to quaternium-15, a common pollutant in aquatic ecosystems. Cell viability and phagocytosis activity decreased when exposed to 0.1 or 1 mg L− 1 of quaternium-15. Moreover, decreasing phagocytosis was confirmed by gene expression modulation of actin, involved in cytoskeleton rearrangement. Effects on oxidative stress-related genes were also assessed (Cat , MnSod , Zn/CuSod , GPx). The qPCR data revealed alterations in antioxidant responses through gene dose- and time-dependent modulation. This study presents insights into the physiological responses and cellular mechanisms of P. imbricata haemocytes to environmental stressors, indicating that this species is useful as a novel bioindicator for future toxicological studies. [Display omitted] • In vitro analysis of P. imbricata haemocytes immunostimulation using Quaternium-15. • Quaternium-15 haemocyte exposure leads to alterations in immune responses. • Proposal of P. imbricata as a new bioindicator in temperate Mediterranean waters. [ABSTRACT FROM AUTHOR]

This article is part of Topical Collection on Fish Immunology Since 2016, I Immunogenetics i has published a yearly special issue with articles by leading experts in a particular field. The subsequent five articles are on the immune system of teleost (modern bony) fishes. Sutoh and Kasahara ([10]) contributed "The immune system of jawless vertebrates: insights into the prototype of the adaptive immune system." 10.1007/s00251-020-01183-5 10 Sutoh Y, Kasahara M. The immune system of jawless vertebrates: insights into the prototype of the adaptive immune system. [Extracted from the article]

Alkaline phosphatase (AP) is a major, recently recognized component of innate immunity. The intestinal AP (IAP) isoform plays a pivotal role in controlling gastrointestinal and systemic inflammation in terrestrial mammals. This is so essentially through detoxification (by dephosphorylation) of proinflammatory microbial components that can no longer be recognized by so-called toll-like receptors, thus preventing cellular inflammatory cascade activation. A unique feature of fish is the presence of AP in skin and epidermal mucus (skin mucus AP) but its actual functions and underlying mechanisms of action are presently unknown. Here, we gather and analyse knowledge available on skin mucus AP in order to provide a holistic view of this important protective enzyme. Our main conclusions are that skin mucus AP is responsive to biotic and abiotic factors, including nutrients and bioactive feed components, prebiotics and probiotics. Importantly, both skin mucus AP and IAP appear to correlate, thus raising the interesting possibility that skin mucus AP be used as a proxy for IAP in future nutritional studies. Blood serum AP also seems to correlate with skin mucus AP, though biological interpretation for such relationship is presently unknown. Finally, the precise isoform/s of AP present in skin should be identified and underlying molecular mechanisms of skin mucus AP actions deciphered. Image 1 • LALLES JP – Review in Fish and Shellfish Immunology – MS FSIM-D-19-00026. • Fish produce skin mucus which contains alkaline phosphatase (AP) activity. • The actual functions of skin mucus AP are presently unknown. • Skin mucus AP could be anti-inflammatory as intestinal AP is in mammals and fish. • Skin mucus AP activity can be increased by various dietary compounds. [ABSTRACT FROM AUTHOR]

Abstract Simultaneous and sequential infections often occur in wild and farming environments. Despite growing awareness, co-infection studies are still very limited, mainly to a few well-established human models. European salmonids are susceptible to both Proliferative Kidney Disease (PKD), an endemic emergent disease caused by the myxozoan parasite Tetracapsuloides bryosalmonae , and Viral Haemorrhagic Septicaemia (VHS), an OIE notifiable listed disease caused by the Piscine Novirhabdovirus. No information is available as to how their immune system reacts when interacting with heterogeneous infections. A chronic (PKD) + acute (VHS) sequential co-infection model was established to assess if the responses elicited in co-infected fish are modulated, when compared to fish with single infections. Macro- and microscopic lesions were assessed after the challenge, and infection status confirmed by RT-qPCR analysis, enabling the identification of singly-infected and co-infected fish. A typical histophlogosis associated with histozoic extrasporogonic T. bryosalmonae was detected together with acute inflammation, haemorrhaging and necrosis due to the viral infection. The host immune response was measured in terms of key marker genes expression in kidney tissues. During T. bryosalmonae /VHSV-Ia co-infection, modulation of pro-inflammatory and antimicrobial peptide genes was strongly influenced by the viral infection, with a protracted inflammatory status, perhaps representing a negative side effect in these fish. Earlier activation of the cellular and humoral responses was detected in co-infected fish, with a more pronounced upregulation of Th1 and antiviral marker genes. These results reveal that some brown trout immune responses are enhanced or prolonged during PKD/VHS co-infection, relative to single infection. Highlights • First comprehensive transcriptional study on a sequential heterogeneous co-infection in fish. • Large primer set optimised for immune gene expression screenings in brown trout. • Significant correlations between pathogenesis and innate and adaptive immune genes expression. • Favourable modulations of T helper subsets activity seen in PKD/VHS co-infected fish. • Enhanced antiviral response seen in PKD/VHS co-infected fish. [ABSTRACT FROM AUTHOR]

Graphical abstract Highlights • 4-nonylphenol induced cytopathological changes in MMCs of Clarias gariepinus. • 4-NP has immunological effects on the hepatic MMC of C. gariepinus. • SOD2 totally increased in 4-NP treated catfish to overcome 4-NP induced mitochondrial oxidative stress. Abstract Melanomacrophage centres (MMCs) play a key role in the immune response in fish. They are considered sensitive bio-monitoring structures with roles in the assessment of toxicant impacts. The aim of this study was to examine the potential histopathological effect of 4-nonylphenol (4-NP) on hepatic MMCs in Clarias gariepinus. To achieve this objective, adult male fish were divided randomly and equally into two groups: a control group and a group that was exposed to 4-NP (dissolved in water at a dose of 0.1 mg/L) for 21 days. The 4-NP-intoxicated hepatic MMCs contained numerous necrotic macrophages. Superoxide dismutase 2 was immuno-expressed in the hepatic MMCs in both groups, with no significant difference. Histomorphometric examination revealed that the sizes and numbers of MMCs were dramatically higher in the livers of 4-NP-exposed C. gariepinus than in control fish. Following 4-NP challenge, in the liver, the abundance of lipofuscin and haemosiderin pigments increased, and single-pigmented macrophages, aggregated groups of deformed red blood cells (RBCs) and macrophages were present near blood vessels and hepatic sinusoids. These results reveal that 4-NP exerts immunological effects on hepatic MMCs in C. gariepinus and support the utility of MMCs as a cytological biomarker for aquatic exposure to 4-NP. [ABSTRACT FROM AUTHOR]

Abstract Polymeric immunoglobulin receptor (pIgR) plays important roles in transport of polymeric immunoglobulins across mucosal epithelial cells and prevention of access of pathogens into the organisms. In this study, a pIgR ortholog (OnpIgR) was cloned and identified from Nile tilapia (Oreochromis niloticus). The open reading frame (ORF) of OnpIgR is 1023 bp, encoding a predicted protein of 341 amino acids. The deduced amino acid sequence of OnpIgR is highly homologous to other teleost's pIgRs, containing two Ig-like domains (ILDs), a transmembrane region and an intracellular region. Expression analysis revealed that OnpIgR was exhibited in all tested tissues including liver, spleen, skin and intestines, and highly expressed in the liver. Immunofluorescence detection indicated that the natural pIgR expressed on the membrane of the epithelial cells. After challenge of bacterial pathogen (Streptococcus agalactiae), OnpIgR expression was significantly up-regulated in intestines and skin, as well as intestinal epithelial cells. In addition, recombinant OnIFN-γ induced the expression of pIgR in the intestinal epithelial cells. Moreover, recombinant OnpIgR protein was able to bind both S. agalactiae and A. hydrophila in vitro , and interact with (r)OnIgM and (r)OnIgT. Taken together, the results indicated that OnpIgR, involving in the polymeric immunoglobulin transport, might play an important role in mucosal immunity against bacterial infection. Highlights • The pIgR gene was identified in Nile tilapia (OnpIgR). • OnpIgR expression was significantly up-regulated in intestines and skin, and intestinal epithelial cells following bacterial challenges. • Recombinant OnIFN-γ could significantly enhance the pIgR expression in epithelial cells. in vitro. • Recombinant OnpIgR was able to bind S. agalactiae and A. hydrophila , and interact with (r)OnIgM and (r)OnIgT. [ABSTRACT FROM AUTHOR]

Abstract Grobiotic®-A (GA) is a unique prebiotic that contains a combination of partially autolyzed brewer's yeast, dairy components, and dried fermentation products. This study was conducted to dynamically investigate the effects of dietary GA (1%, G1) on growth, lipid metabolism, inflammatory responses in liver and visceral adipose tissues (VAT), and gut microbiota in largemouth bass (Micropterus salmoides). The results demonstrated that GA inclusion significantly improved growth during the 8w feeding, which could be related to higher proportion of Firmicutes and Bacillus in the gut. Dual responses of lipid metabolism in liver and VAT were observed. The different pattern of lipid metabolism induced lipid homeostasis in liver but high fat accumulation in VAT when fed G1 diet. GA inclusion induced the short-term (2w) inflammatory responses but returned to same or lower levels in a longer term (8w). GA improved liver health by decreasing hepatic glycogen and ameliorating lipid metabolism genes expression, further inhibiting the inflammatory and apoptosis responses. Highlights • Prebiotic GA inclusion improved growth of largemouth bass by altering gut microbiota. • Different patterns of lipid metabolism were observed in liver and VAT. • GA inclusion induced inflammation at 2w, and returned to same or lower levels at 4, 8w. • GA inclusion protected liver health by reducing glycogen accumulation. • Long-term GA inclusion positively regulated lipid metabolism and alleviated inflammation and apoptosis. [ABSTRACT FROM AUTHOR]

Abstract For many years, feed-borne mycotoxin contamination has been a serious and unavoidable problem in the aquaculture industry. This problem is extensively aggravated due to the increasing replacement of fish meal by plant protein. Aflatoxin B1 (AFB1) is a mycotoxin that has received a great deal of attention, owing to its prevalence in plant feedstuffs and detrimental effects on animals. The objective of this study was to investigate the effect of AFB1 on growth and health. The growth performance and structural integrity of immune organs were evaluated after feeding grass carp diets with 0, 29, 59, 86, 110 and 147 μg AFB1/kg for 60 days. The results indicated that dietary AFB1 caused poor growth performance and deformities in juvenile grass carp. In addition, we are the first to observe that AFB1 was detectable in the head kidney and spleen of juvenile grass carp and damages the structural integrity of those organs. And we, for the first time, explore potential mechanisms of this destruction effect to immune organs of fish, which might be partly involved in (1) attenuating antioxidant ability through up-regulation of Keap1a (not Keap1b) to suppress Nrf2 signalling leading to decrease of mRNA levels and activities of antioxidant enzymes (except GSTP1 mRNA level); (2) aggravating apoptosis partly by activating p38 MAPK signalling to activate mitochondria pathway and death receptor pathway; and (3) weakening TJs structure by promoting MLCK signalling to down-regulate the mRNA of TJs protein (claudin-12 was up-regulated) in the head kidney and spleen. Finally, based on the oxidative injury-related indexes (contents of MDA in the head kidney and PC in the spleen), the safe upper doses of AFB1 for grass carp were first estimated to be 30 and 29 μg/kg diet, respectively. Highlights • Dietary aflatoxin B1 caused poor growth performance and deformities of fish. • Dietary aflatoxin B1 could residue in the head kidney and spleen of juvenile grass carp. • Dietary aflatoxin B1 attenuated antioxidant ability, and thus impaired structural integrity of fish immune organs. • Dietary aflatoxin B1 aggravated apoptosis leading to the damage of structural integrity of fish immune organs. • Dietary aflatoxin B1 weakened intercellular integrity of immune organs resulting in destruction the tight junction in fish. [ABSTRACT FROM AUTHOR]

Immunoglobulins are globulins with antibody activity or antibody-like structures, and fish were the first vertebrates to develop immunoglobulins. There are three major types of immunoglobulins (IgM, IgD and IgZ) in teleost fish, which play an important role in resisting pathogen infection. For fish, the intestinal tract is not only the place for nutrient digestion but also the largest endocrine organ. The intestinal mucosal barrier system can block pathogenic microorganisms and toxins and is important for immunologic defense. Therefore, the study of the molecular mechanism of fish immunoglobulin in intestinal mucosa is of great significance to the selection of immune prevention strategies for fish diseases. At present, information on the immune system of intestinal mucosa in fish is scarce, and there are still many problems in the function of newly identified antibodies, which need further research. In this study, we have summarized current knowledge about the types, gene structure, biological function, and immune response of immunoglobulins in teleost fish, and intestinal mucosal immune tissue of fish. The scientific questions in this field were also presented according to the progress from the latest research in fish immunology. The aim of present study is to provide a theoretical basis for the study of fish immunology. [ABSTRACT FROM AUTHOR]

Abstract This study examined the effects of replacing fish meal (FM) with three different types of soybean meal (SM) including untreated SM, Bacillus pumillus SE5 (BP) fermented SM (BPFSM) and Pseudozyma aphidis ZR1 (PA) fermented SM (PAFSM) in diets for Japanese seabass (Lateolabrax japonicus). A basal diet was formulated using FM (FM diet), and six other diets were produced by substituting 40 or 80% of FM with SM, BPFSM or PAFSM (SM40, SM80, BPFSM40, BPFSM80, PAFSM40 and PAFSM80 diets). Each diet was fed to triplicate groups of fish (7.14 ± 0.05 g) twice daily for eight weeks. Replacing 40% of FM with SM sources did not significantly influence growth (P > 0.05), while increasing the substitution level to 80% led to reduced growth rates (P < 0.05). The groups received SM80 and PAFSM80 diets showed significantly higher feed conversion ratio and lower protein digestibility than FM group. Furthermore, notably lower dry matter digestibility was detected in SM80 group. Remarkably lower serum total antioxidant capacity was found in the SM80 group, and catalase activity did not significantly differ between FM and BPFSM40 groups. Serum malondialdehyde concentration was enhanced by increasing FM replacement level and the highest value was observed in the SM80 fed fish. FM and PAFSM40 groups showed significantly higher lysozyme activity than the SM80 group. Fish fed the BPFSM40 diet exhibited the highest complement C3 activity and the lowest value was observed in the SM80 group. Expression of lysozyme gene in spleen was down-regulated in the SM80 group, and no significant difference in expression of C3 gene was found among FM, BPFSM40 and PAFSM40 groups. Digestive enzymes activity and gut morphology were significantly influenced by FM replacement. Expression of HSP70 and pro-inflammatory genes including TNF-α and IL-1β were up-regulated by FM replacement and relatively lower expression levels were found by using fermented SM. An opposite trend was observed for the anti-inflammatory TGF-β gene expression. Serum d -lactate concentration was significantly increased by replacing 80% of FM with any of the SM sources. These findings indicated that using fermented SM, particularly BPFSM, beneficially influences feed utilization, antioxidant capacity, innate immunity and gut health in juvenile Japanese seabass. Highlights • Yeast and bacterial fermented soybean meal were tested in Japanese seabass feed. • Fermentation with Bacillus pumillus SE5 improved digestibility of soybean meal. • Fermented soybean meal enhanced antioxidant capacity and innate immune response. • Fish meal replacement with soybean meal suppressed digestive enzymes activity. • Fermentation of soybean meal ameliorated the damaging effects on gut structure. [ABSTRACT FROM AUTHOR]

Abstract The present study investigated the effects of dietary blackberry syrup on growth performance, haematological, non-specific immune and spleen gene expression responses of Nile tilapia, Oreochromis niloticus. Five experimental groups of fish with mean weights of 26.75 ± 2.67 g were used in the study; three of them were fed with blackberry syrup incorporated diets (7.5 g kg−1- BBRY7.5, 15 g kg−1- BBRY15, 30 g kg−1- BBRY30), whereas an additive free basal diet served as the control. Additionally, the fifth group was an antibiotic medicated diet (0.02 g kg−1- ABTC), prepared with the florfenicol. Dietary blackberry syrup especially at 15 g kg−1 significantly increased growth performance, respiratory burst activity, potential killing activity, phagocytic activity, phagocytic index, lysozyme activity, myeloperoxidase activity, total immunoglobulin levels, serum SOD activity and serum CAT activity (p < 0.05). Furthermore, dietary blackberry syrup increased the expression levels of immune [heat shock protein 70 (HSP70), interleukin 1, beta (IL-1β), tumor necrosis factor (TNF-α), interferon gamma (IFN-γ), immunoglobulin M (IgM)] and antioxidant [glutathione peroxidase (GPx)] related genes in the spleen of fish fed with especially 15 g kg−1 blackberry syrup (p < 0.05). At the end of the 20-day challenge period the survival rates were significantly higher in the BBRY15 and ABTC groups compared to all other treatment groups (p < 0.05). As a result, feeding Nile tilapia with a diet containing 15 g kg−1 blackberry syrup over a period of 90 days might be adequate to improve growth performance, fish immune parameters, antioxidant status, as well as survival rate against P. shigelloides , similar to antibiotic treatment. Hence, blackberry syrup can be used as an antibiotics replacer for controlling P. shigelloides in tilapia feed. Highlights • Blackberry syrup increased the innate immune responses of Nile tilapia. • Dietary supplementation of 15 g kg−1 blackberry syrup significantly improved growth performance. • Dietary supplementation of 15 g kg−1 blackberry syrup enhanced the protection of tilapia against P. shigelloides. • Dietary supplementation of 15 g kg−1 blackberry syrup enhanced the protection of Nile tilapia against Plesiomonas shigelloides. [ABSTRACT FROM AUTHOR]

Abstract In order to elucidate the influence of T lymphocytes subpopulations on B lymphocytes immune response, in this paper, CD4-1+, CD4-2+, CD8+ T lymphocytes and B lymphocytes responses to thymus-independent (TI) or thymus-dependent (TD) antigen plus immunosuppressant were investigated in flounder (Paralichthys olivaceus). The results showed that in LPS-immunized group, the percentages of CD4-1+, CD4-2+, CD8β+ T (PCD4-1+ T, PCD4-2+ T and PCD8β+ T) lymphocytes in peripheral blood leucocytes (PBLs) had no significant variations, the percentages of IgM+ B (PIgM+ B) lymphocytes and LPS-specific antibodies (LA) significantly increased and peaked at 3rd or 4th week post-injection; CsA had no inhibition on both T/B lymphocytes and LA; RaPa only suppressed the PIgM+ B lymphocytes and LA, and the inhibition maximum (Imax) were about 35% and 20%, respectively. In KLH-immunized group, the PCD4-1+, PCD4-2+ and PCD8β+ T lymphocytes significantly increased and peaked at 3rd or 5th day, successively the PIgM+ B lymphocytes and KLH-specific antibodies (KA) significantly increased to the peak at 5th week; the PCD4-1+, PCD4-2+ T and PIgM+ B lymphocytes and LA were inhibited significantly by both CsA and RaPa, and the Imax on them were 13%–33%, 11%–25%, 19%–34%, 22%–26%, respectively, while the PCD8β+ T lymphocytes showed no significant suppression. The results indicated that the suppression of PIgM+ B lymphocytes in KLH + CsA group was not directly derived from CsA, but due to the suppression of T lymphocytes, especially CD4+ T lymphocytes subpopulations. The results showed for the first time that, similar to higher vertebrates, T lymphocytes didn't respond to TI antigen, moreover, T lymphocyte subpopulations had a regulation on the immune response of B lymphocyte for TD antigen in flounder. Highlights • KLH activated CD4-1+, CD4-2+, CD8+ T and B lymphocytes responses. • LPS only induced B lymphocytes response and antibodies production. • Cyclosporine A (CsA) specifically inhibited T lymphocytes. • CD4-1+, CD4-2+ T lymphocytes influenced the response of B lymphocytes for TD antigen. [ABSTRACT FROM AUTHOR]

Abstract Toll-like receptors (TLRs) are well-known pattern recognition receptors that play key immunological roles in a diverse range of organisms. In this study, two novel invertebrate TLRs from disk abalone (designated as AbTLR-A and AbTLR-B) were identified and functionally characterized for the first time. AbTLR-A and AbTLR-B comprised the typical TLR domain architecture containing an extracellular leucine-rich repeat domain, transmembrane domain, and Toll/interleukin-1 receptor domain. Expressional analysis revealed that both TLRs were constitutively expressed at all the early embryonic stages of disk abalone analyzed, with the highest level of AbTLR-A found at the 16-cell stage and AbTLR-B at the trochophore stage. According to tissue distribution analysis, prominent mRNA expression of AbTLR-A and AbTLR-B was detected in the hemocytes and gills, respectively. AbTLR-A and AbTLR-B mRNAs were significantly up-regulated in response to Gram-negative Vibrio parahemolyticus , Gram-positive Listeria monocytogenes , and viral hemorrhagic septicemia virus injections in abalone hemocytes and gills. Overexpression of AbTLR-A and AbTLR-B in HEK293T cells directly activated nuclear factor-κB (NF-κB) and activator protein-1 (AP-1) responsive reporters. Neither TLRs showed a high response to pathogen-associated molecular patterns in vitro. Co-expression of AbTLR-A and AbTLR-B with AbMyD88-2 and AbMyD88-X activated NF-κB-responsive reporters in a synergetic manner. These findings demonstrate the involvement of AbTLR-A and AbTLR-B in abalone innate immunity. Highlights • Two novel Toll Like Receptors (TLRs) were identified from Disk abalone (AbTLR-A and AbTLR-B). • Both abalone TLRs comprised with typical TLR structural features. • Constitutive expressed mRNA of AbTLR-A and AbTLR-B was detected at early embryonic developmental stages of disk abalone. • AbTLR-A and AbTLR-B expression was modulated upon immune challenge. • Overexpression of both TLRs in HEK293T cells directly activated NF-κB and AP-1. [ABSTRACT FROM AUTHOR]

Abstract The tripartite motif (TRIM) family involves many cellular processes, including fundamental functions in antiviral immunity. Antiviral activities of TRIMs are reported in a variety of patterns, and one of the most significant channels is related to the activation of the type-I interferon (IFN) pathway. In this study, we described a fintrim (ftr) gene named ftr36 , which is mainly expressed in the gills, skin, and intestines. This study shows that ftr36 encodes a protein affording a potent antiviral effect. In vitro , overexpression of FTR36 mediated an upregulated pattern of recognition receptor retinoic acid–inducible gene I (RIG-I), interferon regulatory factor 3/7(IRF3/7), IFN, and IFN-stimulated genes (ISGs) expression. Thereby, FTR36 expression could afford host defense against the spring viremia of carp virus (SVCV) and the giant salamander iridovirus (GSIV). With the deletion of the RING domain or B30.2 domain separately, the antiviral ability of FTR36 was abolished partially and almost lost its ability to activate the IFN-pathway. These findings indicate that both RING and B30.2 domains are indispensable for the antiviral activity of FTR36. Altogether, this study described a finTRIM FTR36, which can activate IFN-pathways and stimulate ISGs to provide host defense against viral infections. Highlights • The mRNA expression of ftr36 was induced due to SVCV infection in vivo. • Overexpression of FTR36 significant inhibits SVCV replication in EPC and FHM cells. • FTR36 positively regulated the key genes expression of the interferon pathway. • RING and B30.2 domains were required for antiviral effects of FTR36. [ABSTRACT FROM AUTHOR]

Abstract Japanese eel (Anguilla japonica) has become a commercially important fish species all over the world. High-density aquaculture has led to congestion and contributed to bacterial infection outbreaks that have caused high mortality. Therefore a 56-days feeding trial was conducted to determine the effects of dietary Bacillus amyloliquefaciens (GB-9) and Yarrowia lipolytica lipase2 (YLL2) on growth performance, digestive enzymes activity, innate immunity and resistance to pathogens of A. japonica. Fish growth performance was significantly affected by dietary YLL2 supplementation but not by GB-9. Fish fed diets with YLL2 at 2.0 g/kg diet in combination of high and low levels of GB-9 (5.0 g/kg and 2.0 g/kg) produced the highest growth. For digestive enzyme, lipase and trypsin activities was promoted by dietary containing YLL2, while amylase activities was increased by dietary containing YLL2, GB-9 single or combination. For innate immunity, the mucus lysozyme activity, leukocytes phagocytosis activity and reactive oxygen species level of skin, peroxidase and lysozyme activity of serum were enhanced in fish fed with GB-9 compared to those in control group (p < 0.05). The highest resistance to Vibrio anguillarum and Aeromonas hydrophila was determined in fish fed with 5.0 g kg−1 GB-9 + 2.0 g/kg YLL2. This study demonstrated that GB-9 and YLL2 enhanced non-specific immune defense system of A. japonica , providing them with higher resistance to pathogens. The present results suggested that the combination of these supplements could be considered as potential biological additives for aquaculture farmed fish. Highlights • The supplementations of YLL2 resulted in a significant increase in growth performance of Japanese eel. • The lysozyme and peroxidase activity, phagocytosis rates, ROS level were significantly higher in supplemented group. • The digestive enzymes activity were significantly higher in GB-9 supplemented group. • The survival rate of Japanese eel challenged with V. anguillarum and A. hydrophila was significantly improved. [ABSTRACT FROM AUTHOR]

Abstract The global aquaculture has shown an impressive growth in the last decades contributing with a major part of total food fish supply. However, it also helps in the spread of diseases that in turn, causes great economic losses. The White Spot Syndrome Virus (WSSV) is one of the major viral pathogen for the shrimp aquaculture industry. Several attempts to eliminate the virus in the shrimp have been addressed without achieving a long-term effectiveness. In this work, we determine the capacity of the commercial non-toxic PVP-coated silver nanoparticles to promote the response of the immune system of WSSV-infected shrimps with or without an excess of iron ions. Our results showed that a single dose of metallic silver in the nanomolar range (111 nmol/shrimp), which is equivalent to 12 ng/mL of silver nanoparticles, produces 20% survival of treated infected shrimps. The same concentration administered in healthy shrimps do not show histological evidence of damage. The observed survival rate could be associated with the increase of almost 2-fold of LGBP expression levels compared with non-treated infected shrimps. LGBP is a key gene of shrimp immunological response and its up-regulation is most probably induced by the recognition of silver nanoparticles coating by specific pathogen-associated molecular pattern recognition proteins (PAMPs) of shrimp. Increased LGBP expression levels was observed even with a 10-fold lower dose of silver nanoparticles (1.2 ng/shrimp, 0.011 nmol of metallic silver/shrimp). The increase in LGBP expression levels was also observed even in the presence of iron ion excess, a condition that favors virus proliferation. Those results showed that a single dose of a slight amount of silver nanoparticles were capable to enhance the response of shrimp immune system without toxic effects in healthy shrimps. This response could be enhanced by administration of other doses and might represent an important alternative for the treatment of a disease that has still no cure, white spot syndrome virus. [ABSTRACT FROM AUTHOR]

Abstract Thymosins β are actin-binding proteins that play a variety of different functions in inflammatory responses, wound healing, cell migration, angiogenesis, and stem cell recruitment and differentiation. In crayfish, thymosins participate in antiviral immunology. However, the roles of thymosin during bacterial infection in shrimp remain unclear. In the present study, four thymosins were identified from kuruma shrimp, Marsupenaeus japonicus , and named as Mj thymosin2, Mj thymosin3, Mj thymosin4, and Mj thymosin5 according the number of their thymosin beta actin-binding motifs. Mj thymosin3 was selected for further study because its expression level was the highest in hemocytes. Expression analysis showed that Mjthymosin3 was upregulated in hemocytes after challenged by Vibrio anguillarum or Staphylococcus aureus. The recombinant Mj thymosin3 protein could inhibit the growth of certain bacteria in an in vitro antibacterial test. Mj thymosins could facilitate external bacterial clearance in shrimp, and were beneficial to shrimp survival post V. anguillarum or S. aureus infection. The results suggested that Mj thymosins played important roles in the antibacterial immune response of kuruma shrimp. Highlights • Four thymosins were identified from kuruma shrimp, Marsupenaeus japonicus. • Mjthymosin3 was upregulated in hemocytes after challenge by V. anguillarum or S. aureus. • R Mj thymosin3 could inhibit the growth of certain bacteria. • Mj thymosins could facilitate external bacterial clearance in shrimp. • Mj thymosins were beneficial to shrimp survival post V. anguillarum or S. aureus infection. [ABSTRACT FROM AUTHOR]

In recent years, investigations of microbial flora associated with fish gut have deepened our knowledge of the complex interactions occurring between microbes and host fish. The gut microbiome not only reinforces the digestive and immune systems in fish but is itself shaped by several host-associated factors. Unfortunately, in the past, majority of studies have focused upon the structure of fish gut microbiome providing little knowledge of effects of these factors distinctively and the immense functional potential of the gut microbiome. In this review, we have highlighted the recently gained insights into the diversity and functions of the fish gut microbiome. We have also delved on the current approaches that are being employed to study the fish gut microbiome with an aim to collate all the knowledge gained and make accurate conclusions for their application based perspectives. The literature reviewed indicated that the future research should shift towards functional microbiomics to improve the maximum sustainable yield in aquaculture. [ABSTRACT FROM AUTHOR]

An 8‐week feeding trial was conducted to assess the suitability of fenugreek seed meal (FKSM; Trigonella foenum‐graecum) to replace fishmeal (FM) in diets of African catfish Clarias gariepinus. Five isonitrogenous and iso‐energetic diets were formulated with increasing amounts of FKSM, at the expense of FM, at 0 (Control), 100 (FKSM100), 180 (FKSM180), 260 (FKSM260) or 340 g/kg (FKSM340). Each treatment was triplicated, with 30 fish in each replicate (initial mean ± SD of 9.3 ± 0.5 g). Growth performance, haematology, serum biochemistry, carcass and fillet proximate composition, some innate immunological parameters and hepatic insulin‐like growth factor (IGF) I and II were measured. Growth, feeding efficiencies and hepatic IGF‐I and IGF‐II gene expression were unchanged in the dietary FKSM100 or FKSM180 treatments, compared to the control. In the FKSM180 treatment, blood haemoglobin, haematocrit, red blood cell counts, respiratory burst activity and lysozyme activity of catfish significantly increased. Carcass crude protein, ash and hepatic IGF‐I gene expression were significantly lower in the FKSM260 and FKSM340 treatments. Results show that up to 180 g/kg FKSM can replace FM at 350 g/kg, on a total protein basis in the diets of C. gariepinus. [ABSTRACT FROM AUTHOR]