Miguel Ángel García-Álvarez

Th17 is a lymphocyte T helper (Th) subpopulation relevant in the control and regulation of the immune response characterized by the production of interleukin (IL)-17. This crucial cytokine family acts through their binding to the IL-17 receptors (IL-17R), having up to six members. Although the biology of fish Th17 is well-recognized, the molecular and functional characterization of IL-17 and IL-17R has been limited. Thus, our aim was to identify and characterize the IL-17R repertory and regulation in the two main Mediterranean cultured fish species, the gilthead seabream (Sparus aurata) and the European sea bass (Dicentrarchus labrax). Our in silico results showed the clear identification of six members in each fish species, from IL-17RA to IL-17RE-like, with well-conserved gene structure and protein domains with their human orthologues. All of them showed wide and constitutive transcription in naïve tissues but with highest levels in mucosal tissues, namely skin, gill or intestine. In leucocytes, T mitogens showed the strongest up-regulation in most of the il17 receptors though il17ra resulted in inhibition by most stimulants. Interestingly, in vivo nodavirus infection resulted in alterations on the transcription of il17 receptors. While nodavirus infection led to some increments in the il17ra, il17rb, il17rc and il17rd transcripts in the susceptible European sea bass, many down-regulations were observed in the resistant gilthead seabream. Our data identify the presence and conservation of six coding IL-17R in gilthead seabream and European sea bass as well as their differential regulation in vitro and upon nodavirus infection.

The nervous necrosis virus (NNV) leads to viral encephalopathy and retinopathy (VER) disease in more than 170 fish species, mainly from marine habitats. It replicates in the central nervous tissues, reaching up to 100% mortalities after a few days of infection, mainly in the larvae and juvenile stages. This is continuously spreading and affecting more species, both wild and cultured, posing a risk to the development of the aquaculture industry. In the Mediterranean Sea, it mainly affects European sea bass (Dicentrarchus labrax) and some grouper species (Epinephelus spp.). Interestingly, in the gilthead seabream (Sparus aurata), typically resistant to common NNV strains, great mortalities in hatcheries associated with typical clinical signs of VER have been confirmed to be caused by RGNNV/SJNNV reassortants. Thus, we have evaluated the susceptibility of seabream larvae to either RGNNV/SJNNV or SJNNV/RGNNV reassortants, as well as the larval immunity. Based on our results we can conclude that: (i) gilthead seabream larvae are susceptible to infection with both NNV reassortant genotypes, but mainly to RGNNV/SJNNV; (ii) virus replicated and infective particles were isolated; (iii) larval immunity was correlated with larval survival; and (iv) larval resistance and immunity were correlated with age of the larvae. Further investigations should be carried out to ascertain the risks of these new pathogens to Mediterranean larviculture.

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.