Antimicrobial peptides (AMP) play an essential role in the innate immune system, modulating the defense response. In a previous study, we demonstrated the antimicrobial activity of synthetic hepcidin (hep20) from rainbow trout (Oncorhynchus mykiss), and its protective effect in European sea bass (Dicentrarchus labrax) challenged with Vibrio anguillarum. Additionally, we described the uptake and distribution of hep20 in different tissues and leukocyte cells. Interestingly, various AMPs characterized in high vertebrates, called host defense peptides (HDPs), also possess immunomodulation activity. For that reason, the present study explores the immunomodulatory abilities of hep20 through in vitro and in vivo studies. First, a monocyte/macrophage RTS-11 cell line from rainbow trout was used to evaluate hep20 effects on pro- and anti-inflammatory cytokines in fish leukocyte cells. Next, the European sea bass juveniles were used to determine if hep20 can regulate the expression of cytokines in fish immune tissues. The results show that hep20 was uptake inner to RTS-11 cells and was able to induce the expression of IL-10, IL-1β, and TNFα at transcriptional and protein levels. Then, the European sea bass juveniles were given intraperitoneal injections of the peptide. At 1, 3, 7, 14, and 21 days post-injection (dpi), IL-10, IL -1β, and TNFα mRNA were quantified in the anterior gut, spleen, and head kidney. The hep20 was able to up-regulate cytokine gene expression in these tissues, mainly in the head kidney. Furthermore, the evaluated cytokines showed a cyclical tendency of higher to lesser expression. Finally, a bioinformatics analysis showed that the structure adopted by hep20 is similar to the γ-core domain described for cysteine-stabilized AMP, defined as immunomodulatory and antimicrobial, which could explain the ability of hep20 to regulate the cytokine expression. This study provides new insights into immunomodulatory function complementary to the previously established antimicrobial activity of hep20, suggesting a role as an HDP in teleost fish. These facts are likely to be associated with molecular functions underpinning the protective effect of fish hepcidin against pathogens.

In higher vertebrates, Tripartite Motif (TRIM) proteins modulate the immune response by coordinating processes related to inflammation such as antiviral restriction, autophagy and inflammasome activation. In fish, TRIM proteins have been reported mainly in cyprinids (e.g., carp—Cyprinus carpio and zebrafish—Danio rerio) and salmonids (i.e., rainbow trout—Oncorhynchus mykiss). However, their molecular mechanisms and functions are still being described in aquatic animals. Thus, our study focused on characterizing novel TRIM proteins involved in the innate immunity of gill cells from rainbow trout (RTgill-W1 and primary cultures) stimulated with lipopolysaccharide (LPS) or polyinosinic–polycytidylic acid (poly I:C). Furthermore, an in vivo experiment with rainbow trout was performed to detect TRIM proteins after the challenge with Flavobacterium psychrophilum (a major bacterial pathogen affecting Chilean salmonid industry). In vitro results showed that OmTRIM25 triggered an LPS-induced expression of pro-inflammatory cytokines such as TNF-α2 and IL-1β. Moreover, in the fish experiment, OmTRIM25 and finTRIM2 were up-regulated in the gills two days post-infection (dpi), whereas IL-1β and TNF-α2 had a higher gene expression at four and six dpi, respectively. To investigate the immunological role of OmTRIM25, a gene silencing strategy using RNA interference (RNAi) was used, confirming the immunomodulatory function of OmTRIM25.

Background: Accelerating climate change has intensified hypoxic events in aquatic ecosystems. In aquaculture, high stocking densities make farmed fish particularly vulnerable to these episodes, leading to negative economic repercussions. This has driven interest in selective breeding for hypoxia tolerant fish as a potential mitigation strategy. In this context, the present systematic review synthesizes and critically evaluates current knowledge on genetic variation associated with hypoxia tolerance in farmed fish species. A literature search was conducted in Scopus and Web of Science following the Preferred Reporting Items for Systematic reviews and Meta-Analyses 2020 guidelines. Results: In total, 964 articles were identified, of which 41 met the inclusion criteria, encompassing 26 species and three hybrid lines. Among the farmed fish, the blunt snout bream, rainbow trout, common carp, and Nile tilapia were the most extensively studied. The most commonly used metrics to assess hypoxia tolerance included: (1) time or oxygen level at which loss of equilibrium occurs, (2) survival time or status (alive or dead), and (3) critical oxygen partial tension measured via respirometry. Substantial phenotypic variability in hypoxia tolerance across families, strains, gynogenetic lines, growth transgenic lines, hybrid lines, and species was reported in most studies. Although single nucleotide polymorphism associated with hypoxia tolerance were identified in several studies, heritability estimates were reported in only three of these, ranging from 0.28 to 0.65, underscoring the need for further research to strengthen the basis for selective breeding applications. Furthermore, candidate genes identified across studies were involved in a wide range of biological processes underlying hypoxia responses, including hypoxia signaling and its regulation (HIF-related genes and their inhibitors, such as HIF1αn ), angiogenesis, energy metabolism, oxidative stress defense, erythropoiesis, ion regulation, DNA repair, and immune and apoptotic processes. Conclusions: As aquatic environments become more hypoxic, the findings of this review underscore the potential of the inherent genetic diversity for hypoxia tolerance present in farmed fish populations. In this context, genomic selection and gene editing emerge as promising tools for developing hypoxia tolerant fish lines. Further research under production conditions is essential before implementation of these approaches in practical in breeding programs, particularly to evaluate the most appropriate trait(s) for measuring hypoxia tolerance, potential correlated responses with other economically important traits, and the overall feasibility at an industry scale.

Recent studies have demonstrated that immune-related recombinant proteins can enhance immune function, increasing host survival against infectious diseases in salmonids. This research evaluated inclusion bodies (IBs) of antimicrobial peptides (CAMP^IB and HAMP^IB) and a cytokine (IL1β^IB and TNFα^IB) as potential immunostimulants in farmed salmonids. For this purpose, we produced five IBs (including iRFP^IB as a control), and we evaluated their ability to modulate immune marker gene expression of three IBs in the RTS11 cell line by RT–qPCR. Additionally, we characterized the scale-up of IBs production by comparing two different scale systems. The results showed that CAMP^IB can increase the upregulation of tnfα, il1β, il8, and il10, HAMP^IB significantly increases the upregulation of tnfα, inos, and il10, and IL1β^IB significantly upregulated the expression of tnfα, il1β, and cox2. A comparison of IL1β^IB production showed that the yield was greater in shake flasks than in bioreactors (39 ± 1.15 mg/L and 14.5 ± 4.08 mg/L), and larger nanoparticles were produced in shake flasks (540 ± 129 nm and 427 ± 134 nm, p < 0.0001, respectively). However, compared with its shake flask counterpart, the IL1β^IB produced in a bioreactor has an increased immunomodulatory ability. Further studies are needed to understand the immune response pathways activated by IBs and the optimal production conditions in bioreactors, such as a defined medium, fed-batch production, and mechanical bacterial lysis, to increase yield.