Nervous necrosis virus (NNV) has spread throughout the world, affecting more than 120 freshwater and marine fish species. While vaccination effectively prevents disease outbreaks, the difficulty of producing sufficient viruses using cell lines continues to be a significant disadvantage for producing inactivated vaccines. This study, therefore, explored the application of synthetic peptides as potential vaccine candidates for the prevention of NNV in Asian seabass (Lates calcarifer). Using the epitope prediction tool and molecular docking, three predicted immunogenic B cell epitopes (30-32 aa) derived from NNV coat protein were selected and synthesised, corresponding to amino acid positions 5 to 34 (P1), 133 to 162 (P2) and 181 to 212 (P3). All the predicted peptides interact with Asian sea bass’s MHC class II by docking. The antigenicity of these peptides was determined through ELISA and all peptides were able to react with NNV-specific antibodies. Subsequently, the immunogenicity of these synthetic peptides was investigated by immunisation of Asian seabass with individual peptides (30 μg/fish) and a peptide cocktail (P1+P2+P3, 10 μg each/fish) by intraperitoneal injection, followed by a booster dose at day 28 post-primary immunisation. There was a subset of immunised fish that were able to induce upregulation of CD4 in the head kidney and spleen. Importantly, antibodies derived from fish immunised with synthetic peptides reacted with whole NNV virions. Taken together, these findings indicate that synthetic linear peptides based on predicted B cell epitopes exhibited both antigenic and immunogenic properties, suggesting that they could be potential vaccine candidates for the prevention of NNV in fish.

This study addresses the urgent need for sustainable protein sources in aquaculture due to the depletion of marine resources and rising costs. Animal protein sources, particularly poultry by-product meal (PBM) and insect exuviae meal, are investigated as viable alternatives to fishmeal (FM). Research confirms the successful replacement of FM with a combination of PBM and insect exuviae meal (up to 50%) in the diet of European seabass without compromising growth, feed conversion, gut health and liver fat content. In particular, growth performance was robust with both PBM formulations, with the 25% PBM diet showing better results. Histological examinations showed good gut and liver health, contradicting the concerns of previous studies. The study emphasizes the importance of holistic analyzes that go beyond growth parameters and include histomorphological investigations. The results show that PBM in combination with insect/exuviae meal is well tolerated by seabass, which is consistent with literature reports of mitigating negative effects on gut health. A detailed analysis of the microbiota revealed a decrease in the Firmicutes/Proteobacteria ratio due to an increase in potentially pathogenic bacteria. However, the formulation containing insect exuviae partially counteracted this effect by preserving the beneficial Lactobacillus and promoting the synthesis of short-chain fatty acids (SCFA), particularly butyrate. Chitin-rich components from insect exuviae were associated with improved gut health, which was supported by increased production of SCFA, which are known for their anti-inflammatory properties. The study concludes that a combination of PBM and insect/exuviae meal can replace up to 50% of FM in the diet of seabass, supporting sustainable aquaculture practices. Despite some changes in the microbiota, the negative effects are mitigated by the addition of insect exuviae, highlighting their potential as a prebiotic to increase fish productivity and contribute to a circular economy in aquaculture.

In the present study, bivalent nanovaccines of S. iniae and F. covae were administered by immersion vaccination at 30 and 40 days after hatching (DAH), and the third vaccination was orally administered by feeding at 50 DAH. Five different groups were designed: group 1 was set as the control group, and groups 2-4 were vaccinated using the same protocols. ELISA revealed that the levels of total IgM and specific IgM to S. iniae, and F. covae were significantly elevated in groups 2-4 at 10, 20, and 30 days after vaccination (DAV). A qRT‒PCR analysis of immune-related genes revealed significantly higher IgT expression in the vaccinated groups compared with the control group, as revealed by 44-100-fold changes in the vaccinated groups compared with the control (p < 0.001) at every tested time point after vaccination. All vaccinated groups expressed IgM, MHCIIα and TCRα at significantly higher levels than the control group at 10 and/or 20 DAV (p < 0.05). In the S. iniae challenge tests, the survival of vaccinated groups 2-4 ranged from 62.15 ± 2.11 to 75.70 ± 3.36%, which significantly differed from that of the control group (44.44 ± 1.92%). Similarly, all vaccinated groups showed higher survival rates of 68.89 ± 3.85 to 77.78 ± 5.09% during F. covae challenge than the control groups (50.00 ± 3.33%) (p < 0.05).

The aim of the present study was to evaluate the effects on the presence and distribution of oxyntopeptic cells (OPs) and enteroendocrine cells (EECs) expressing GHR, NPY and SOM in the gastric mucosa of European seabass and seabream of dietary levels of bioactive peptides (BPs) obtained from salmon processing by-products (in silico approach). In this study, 27 seabass and 27 seabreams were divided into three experimental groups: a control group (CTR) fed a control diet and two groups fed different levels of BP to replace fishmeal: 5% BP (BP5%) and 10% BP (BP10%). The stomach of each fish was sampled and processed for immunohistochemistry. Some SOM, NPY and GHR-IR cells exhibited alternating “open type” and “closed type” EECs morphologies. In seabream gastric mucosa, the BP10% group (16.8 ± 7.5) showed a significant change in NPY-IR cell number compared to CTR (CTR 8.5 ± 4.8) and BP5% (BP10% vs. CTR P ≤ 0.01; BP10% vs. BP5% P ≤ 0.05). In seabream, SOM-IR cells in the BP 10% diet (16.8 ± 3.5) differed from those in CTR (12.5 ± 5) (CTR vs. BP 10% P ≤ 0.05) and BP 5% (12.9 ± 2.5) (BP 5% vs. BP 10% P ≤ 0.01). In seabass, EEC SOM-IR cells increased at 10 % BP (5.3 ± 0.7) compared to 5 % BP (4.4 ± 0.8) (5 % BP vs. 10 % BP P ≤ 0.05). The results obtained suggest that salmon BPs have a promising impact as circulating ingredients for seabass and seabream.

Using qRT-PCR analysis, the LcIL-8 expression level indicated broad expression in most detected tissues, with the most predominant expression in whole blood at 6 to 24 h after infection with S. iniae at concentrations of 105 colony-forming unit (CFU)/fish (p < 0.05). After immersion with F. covae, the LcIL-8 transcript was upregulated in the gills, liver and intestine, with the highest expression level found in the gills. However, LcIL-8 was downregulated in all tested tissues at 48 and 96 h after infection with the two pathogenic strains, indicating that Lc-IL8 has a short half-life during the early immune responses to pathogens. MIC analysis of rLcIL-8 protein against S. iniae was 10.42 ± 3.61 µg/mL. Furthermore, the functional analyses clearly demonstrated that 10 and 100 µg of rLcIL-8 protein efficiently enhanced the phagocytic activity of Asian seabass phagocytes in vitro (p < 0.05). Additionally, in vivo injection of S. iniae following the rLcIL-8 protein indicated that 50 and 100 µg of rLc-IL-8 were highly effective in protecting fish from the tested pathogen (p < 0.001). These results indicate that rLcIL-8 plays a crucial role in the defense mechanisms against bacterial infections in the target fish.