Antigen Presentation and Autophagy in Teleost Adaptive Immunity

Infectious diseases are a burden for aquaculture. Antigen processing and presentation (APP) to the immune effector cells that fight pathogens is key in the adaptive immune response. At the core of the adaptive immunity that appeared in lower vertebrates during evolution are the variable genes encoding the major histocompatibility complex (MHC). MHC class I molecules mainly present peptides processed in the cytosol by the proteasome and transported to the cell surface of all cells through secretory compartments. Professional antigen-presenting cells (pAPC) also express MHC class II molecules, which normally present peptides processed from exogenous antigens through lysosomal pathways. Autophagy is an intracellular self-degradation process that is conserved in all eukaryotes and is induced by starvation to contribute to cellular homeostasis. Self-digestion during autophagy mainly occurs by the fusion of autophagosomes, which engulf portions of cytosol and fuse with lysosomes (macroautophagy) or assisted by chaperones (chaperone-mediated autophagy, CMA) that deliver proteins to lysosomes. Thus, during self-degradation, antigens can be processed to be presented by the MHC to immune effector cells, thus, linking autophagy to APP. This review is focused on the essential components of the APP that are conserved in teleost fish and the increasing evidence related to the modulation of APP and autophagy during pathogen infection.Versión del editor2,46

Sex steroids modulate fish imnune response

Versión del edito

Shi drum (Umbrina cirrosa) juveniles are susceptible to all Nodavirus genotypes

Nervous necrosis virus (NNV; family Nodaviridae, genus Betanodavirus) is the causative agent of viral retinopathy and encephalopathy (VER) disease, which mainly affects the larval and juvenile stages of fish. The virus has an acute lethal effect in larval stages and juveniles. According to the RNA2 sequence, NNV are mainly divided into four genotypes: RGNNV, SJNNV, BFNNV and TPNNV. The shi drum (Umbrina cirrosa) is a serious candidate for the diversification of Mediterranean aquaculture. Although aspects related to nutrition or reproduction have been elucidated others related to pathology or immunity have been poorly studied. In this regard, the shi drum is a susceptible species for betanodavirus (RGNNV) infection, as several natural outbreaks have been reported. In order to expand the actual knowledge and understand the shi drum-NNV interactions we evaluated whether this fish species is susceptible to all the NNV genotypes. Our data demonstrate that the laboratory infections with all the NNV genotypes produced clinical signs of the VER disease and mortalities in shi drum juveniles. Interestingly, clinical signs and histopathological lesions in the brain and retina were different depending on the genotype used. Finally, viral capsid protein was immunodetected in the brain and retina from all infected fish whilst infective particles were only recovered from RGNNV-, BFNNV- and TPNNV-infected specimens. In conclusion, this work demonstrates that shi drum juveniles are susceptible to all four genotypes of NNV and represent the first step in studying host–NNV interactions and immune responses in this species, which should be further characterized

Identification and Regulation of Interleukin-17 (IL-17) Family Ligands in the Teleost Fish European Sea Bass

Interleukin-17 (IL-17) cytokine comprises a family of six ligands in mammals with proinflammatory functions, having an important role in autoimmune disorders and against bacterial, viral, and fungal pathogens. While IL-17A and IL-17F ligands are mainly produced by Th cells (Th17 cells), the rest of the ligands are expressed by other immune and non-immune cells and have different functions. The identification of IL-17 ligands in fish has revealed the presence of six members, counterparts to mammalian ones, and a teleost-specific form, the fish IL-17N. However, tissue distribution, the regulation of gene expression, and scarce bioactivity assays point to similar functions compared to mammalian ones, though this yet to be investigated and confirmed. Thus, we have identified seven IL-17 ligands in the teleost European sea bass (Dicentrarchus labrax), for the first time, corresponding to IL-17A/F1, IL-17A/F2, IL-17A/F3, IL-17C1, IL-17C2, IL-17D, and IL-17N, according to the predicted protein sequences and phylogenetic analysis. They are constitutively and widely transcribed in sea bass tissues, with some of them being mainly expressed in the thymus, brain or intestine. Upon in vitro stimulation of head-kidney leucocytes, the mRNA levels of all sea bass IL-17 ligands were up-regulated by phytohemagglutinin treatment, a well-known T cell mitogen, suggesting a major expression in T lymphocytes. By contrast, the infection of sea bass juveniles with nodavirus (NNV), a very pathogenic virus for this fish species, resulted in the up-regulation of the transcription of IL-17C1 in the head-kidney and of IL-17C1 and IL-17D in the brain, the target tissue for NNV replication. By contrast, NNV infection led to a down-regulated transcription of IL-17A/F1, IL-17A/F2, IL-17C1, IL-17C2, and IL-17D in the head-kidney and of IL-17A/F1 and IL-17A/F3 in the brain. The data are discussed accordingly with the IL-17 ligand expression and the immune response under the different situations testedVersión del editor2,46

Microplastic and an associated metal contaminant (Palladium) impair the immune response against pathogenic bacteria of the marine bivalve Mytilus galloprovincialis

Microplastics (MPs) are widely distributed in marine environments and have been reported to cause harmful physiological effects in marine bivalves including immune modulation. While Mytilus galloprovincialis is a model species in environmental monitoring studies, little is known regarding the effects of MPs and palladium (Pd), an emerging contaminant, metal on the immune functioning of this species. In this study, gills and gland samples of specimens of M.ytilus galloprovincialis which were exposed to the same particle concentration (2.5 x 10 66 μm3μm /mL) of Control (microalgae), MP (virgin microplastics) and MP-Pd (Pd spiked microplastics) once an hour during 4 hours. Thus, mussels were exposed to 6095 ng of Pd/mussel. Samples analyzed were collected after 4 and 24 hours of exposure and after 24, 48 and 144 hours of depuration. Several innate immune activities (lysozymes, peroxidase, protease, antiprotease, and bactericidal activities) were analyzed. Our data demonstrated that after 24 hours of exposure to MP, a decrease in lysozymes and peroxidase activities occurred, but not in the bactericidal activity which increased. However, when MP-Pd was used, an increase in all these activities was observed compared to MP levels group. These data suggest that when MP are spiked with Pd, the latter compound might induce an inflammatory process that will results in higher levels of most of all the immune activities analyzed. In the gland, however, most of the activity levels were decreased upon MP-Pd treatment compared to MP levels at different time point of exposure depending on the activity. Interestingly, after 24 hours of removing the pollutants from the water, most of the activity levels in both tissues, gills and gland, were recovered to control levels, but not the bactericidal activity. Considering this impairment of the bactericidal activity against possible pathogenic bacteria, a potential threat to mussels population in a polluted scenario is highly plausible

Role of estrogens in fish immunity with special emphasis on GPER1

It is well accepted that estrogens, the primary female sex hormones, play a key role in modulating different aspects of the immune response. Moreover, estrogens have been linked with the sexual dimorphism observed in some immune disorders, such as chronic inflammatory and autoimmune diseases. Nevertheless, their effects are often controversial and depend on several factors, such as the pool of estrogen receptors (ERs) involved in the response. Their classical mode of action is through nuclear ERs, which act as transcription factors, promoting the regulation of target genes. However, it has long been noted that some of the estrogen-mediated effects cannot be explained by these classical receptors, since they are rapid and mediated by non-genomic signaling pathways. Hence, the interest in membrane ERs, especially in G protein-coupled estrogen receptor 1 (GPER1), has grown in recent years. Although the presence of nuclear ERs, and ER signaling, in immune cells in mammals and fish has been well documented, information on membrane ERs is much scarcer. In this context, the present manuscript aims to review our knowledge concerning the effect of estrogens on fish immunity, with special emphasis on GPER1. For example, the numerous tools developed over recent years allowed us to report for the first time that the regulation of fish granulocyte functions by estrogens through GPER1 predates the split of fish and tetrapods more than 450 million years ago, pointing to the relevance of estrogens as modulators of the immune responses, and the pivotal role of GPER1 in immunity.Versión del editor3,26

RGNNV and SJNNV reassortants produce mortality and replicate in gilthead seabream larvae

Nervous Necrosis Virus (NNV) is one of the most challenging pathogens for aquaculture development nowadays, mainly affecting marine teleost fish of major interest to the aquaculture industry and causing great economic losses. NNV consist in four genotypes, which seem to have a tropism for certain teleost fish species. Among them, gilthead seabream (Sparus aurata) has been considered as a non-susceptible species to the disease produced by traditional NNV genotypes. However, there are some evidences that indicate seabream is able to develop the disease in the presence of certain reassortant strains of NNV, called RGNNV/SJNNV, which possesses the RNA1 segment of the RGNNV genotype and the RNA2 segment of the SJNNV genotype, which may cause a new threat to aquaculture. Therefore, the main objective of this study was to evaluate the susceptibility of gilthead seabream larvae to the reassortant strains RGNNV/SJNNV and SJNNV/RGNNV. For this purpose, larvae were exposed to 104 TCDI50/mL in triplicate tanks with the reassortant strains. Samples of 5 individual larvae were collected at different days post-infection and used for gene expression and infective NNV isolation. Our data show that both reassortants produced mortalities, although the RGNNV/SJNNV was the one which produced the highest mortality and viral gene transcription, which significantly increased from 1 to 7 days post-infection. In conclusion, our study demonstrate that seabream larvae are susceptible to both RGNNV/SJNNV and SJNNV/RGNNV reassortants under laboratory conditions. Further studies should be performed to understand the pathogenicity of the NNV reassortant strains to prevent and control future outbreaks in aquaculture farms

Profile of Innate Immunity in Gilthead Seabream Larvae Reflects Mortality upon Betanodavirus Reassortant Infection and Replication

Historically, gilthead seabream (Sparus aurata) has been considered a fish species resistant to nervous necrosis virus (NNV) disease. Nevertheless, mortality in seabream hatcheries, associated with typical clinical signs of the viral encephalopathy and retinopathy (VER) disease has been confirmed to be caused by RGNNV/SJNNV reassortants. Because of this, seabream larvae at 37 and 86 days post-hatching (dph) were infected by immersion with RGNNV/SJNNV and SJNNV/RGNNV reassortants under laboratory conditions, and mortality, viral replication and immunity were evaluated. Our results show that gilthead seabream larvae, mainly those at 37 dph, are susceptible to infection with both NNV reassortant genotypes, with the highest impact from the RGNNV/SJNNV reassortant. In addition, viral replication occurs at both ages (37 and 86 dph) but the recovery of infective particles was only confirmed in 37 dph larvae,; this value was also highest with the RGNNV/SJNNV reassortant. Larvae immunity, including the expression of antiviral, inflammatory and cell-mediated cytotoxicity genes, was affected by NNV infection. Levels of the natural killer lysin (Nkl) peptide were increased in SJNNV/RGNNV-infected larvae of 37 dph, though hepcidin was not. Our results demonstrate that the seabream larvae are susceptible to both NNV reassortants, though mainly to RGNNV/SJNNV, in an age-dependent manner.Versión del editor2,46

Vacunas DNA frente a nodavirus. Administración oral y transferencia materna para proteger los estadios larvarios más vulnerables de lubina

Nodavirus (NNV) es el agente causal de la retinopatía y encefalopatía viral (VER) que genera elevadas mortalidades en larvas y juveniles de lubina y de otras especies potencialmente importantes para la acuicultura Mediterránea. Por ello, en este trabajo hemos evaluado dos formas distintas de administración de una vacuna DNA: (i) la vacunación oral de una vacuna encapsulada en partículas de quitosano y (ii) la vacunación de hembras para potenciar la transferencia materna de inmunidad a la descendencia. Hemos obteniendo resultados favorables en ambos procedimientos, aunque estudios más exhaustivos serían necesarios para determinar los mecanismos implicados en estas respuestas y poder así mejorar la eficacia de los tratamientos

Molecular identification and characterization of haptoglobin in teleosts revealed an important role on fish viral infections

Haptoglobin (Hp) molecule has been cloned and characterized in two marine teleosts (gilthead seabream and European sea bass), obtaining putative proteins of 319 residues encoded by an ORF of 960 bp in both species. However, the matrix of similarity revealed low identities among bony fish species 78.9% (seabream-sea bass), 43% (seabream/seabass-zebrafish) and lower than 20% with sharks and human. The protein sequences showed a signal peptide from the position 1 to 23, a trypsin domain from 47 to 297, and several predicted disulfide bridges and glycosylation sites. The expression of hp transcript levels during ontogeny showed a progressive increase of expression in seabream whilst remained almost unaltered in sea bass. By tissues, this gene was found constitutively expressed with the highest levels on liver in both species. The main results on hp transcript levels showed the up-regulation in gilthead seabream suffering from naturally occurring lymphocystis disease; and the down-regulation and up-regulation after nodavirus infection in the resistant gilthead seabream and the susceptible European sea bass, respectively. These findings demonstrate for the first time an important role of haptoglobin against viral infections, operating differently in two of the most important marine farmed fish species.Postprint3,26