Insect meal has shown promise as a potentially sustainable source of nutrients for aquafeeds, offering an alternative to expensive and ecologically undesirable ingredients, in the context of population explosion and climate change. Despite this promising outlook, its effects on fish growth and immune responses remain to be thoroughly investigated. Our scientific goal was to experimentally test responses to replacements of the fish meal with a protein source derived from black soldier fly larvae meal (BSFLM). Possible impacts on growth, immunological response, and the expression of selected immune-system related genes were evaluated in Koi carp (Cyprinus carpio var. koi) using a biofloc culture system. Three hundred fish (20.0 ± 0.2 g) were allocated into five groups: a control group receiving a basal diet containing 0 g kg⁻¹ BSFLM and four experimental groups in which fish meal was replaced with 50, 100, 150, and 200 g kg⁻¹ BSFLM for eight weeks. After 4 weeks of feeding, there were no statistically significant differences in specific growth rate (SGR), feed conversion ratio (FCR), and survival rate between fish fed BSFLM-enriched diets at 50, 100, 150 g kg⁻¹ and a control (0 g kg⁻¹ BSFLM) diet. However, fish fed 200 g kg⁻¹ BSFLM showed significantly improved weight gain (WG) and SGR compared to the control after 4 weeks; this difference persisted through 8 weeks (p < 0.05). After eight weeks, there was a moderate to weak negative linear regression shown in FCR (r = 0.470) and SR (r = 0.384), respectively, with the BSFLM levels, but significant and highly correlated linear relationships were observed in WG (r = 0.917) and SGR (r = 0.912). Immunological response analysis showed slight changes in lysozyme and peroxidase levels by replacing fish meal with BSFLM, but these apparent differences were not significantly related to experimental diets. Interestingly, mRNA transcripts of immune-related genes (TNF-α, TGF-β, IL1, IL10, and hsp70) were upregulated in the groups receiving higher amounts of BSFLM, with statistically significant differences observed in certain comparisons. Our findings reveal that fish meal can be effectively replaced by BSFLM, and that this not only has a positive effect on immune-related gene expression in Koi carp, but also on growth rate, pointing to the future potential role of BSFLM as an alternative fish meal protein in aquafeed formulation.

Chitosan (CH) shows great potential as an immunostimulatory feed additive in aquaculture. This study evaluates the effects of varying dietary CH levels on the growth, immunity, intestinal morphology, and antioxidant status of Nile tilapia (Oreochromis niloticus) reared in a biofloc system. Tilapia fingerlings (mean weight 13.54 ± 0.05 g) were fed diets supplemented with 0 (CH0), 5 (CH5), 10 (CH10), 20 (CH20), and 40 (CH40) mL·kg⁻¹ of CH for 8 weeks. Parameters were assessed after 4 and 8 weeks. Their final weight was not affected by CH supplementation, but CH at 10 mL·kg⁻¹ significantly improved weight gain (WG) and specific growth rate (SGR) compared to the control (p < 0.05) at 8 weeks. Skin mucus lysozyme and peroxidase activities were lower in the chitosan-treated groups at weeks 4 and 8. Intestinal villi length and width were enhanced by 10 and 20 mL·kg⁻¹ CH compared to the control. However, 40 mL·kg⁻¹ CH caused detrimental impacts on the villi and muscular layer. CH supplementation, especially 5–10 mL·kg⁻¹, increased liver and intestinal expressions of interleukin 1 (IL-1), interleukin 8 (IL-8), LPS-binding protein (LBP), glutathione reductase (GSR), glutathione peroxidase (GPX), and glutathione S-transferase (GST-α) compared to the control group. Overall, dietary CH at 10 mL·kg⁻¹ can effectively promote growth, intestinal morphology, innate immunity, and antioxidant capacity in Nile tilapia fingerlings reared in biofloc systems.

Nanobubble technology has shown appealing technical benefits and potential applications in aquaculture. We recently found that treatment with ozone nanobubbles (NB-O₃) activated expression of several immune-related genes leading to effective response to subsequent exposure to fish pathogens. In this study, we investigated whether pre-treatment of Nile tilapia (Oreochromis niloticus) with NB-O₃ can enhance specific immune responses and improve efficacy of immersion vaccination against Streptococcus agalactiae. Spleen and head kidney of fish in the vaccinated groups showed a substantial upregulation in expression levels of pro-inflammatory cytokine genes (IL-1β, TNF-α, IL-6) and immunoglobulin classes (IgM, IgD, IgT) compared with the unvaccinated control groups. The mRNA transcript of pro-inflammatory cytokine genes was greatest (approx. 2.8-3.3 folds) on day 7 post-vaccination, whereas the relative expression of immunoglobulin genes was greatest (approx. 3.2-4.1 folds) on day 21 post-immunization. Both systemic and mucosal IgM antibodies were elicited in vaccinated groups. As the result, the cumulative survival rate of the vaccinated groups was found to be higher than that of the unvaccinated groups, with a relative percent survival (RPS) ranging from 52.9 to 70.5%. However, fish in the vaccinated groups that received pre-treatment with NB-O₃, bacterial antigen uptakes, expression levels of IL-1β, TNF-α, IL-6,IgM, IgD, and IgT, as well as the specific-IgM antibody levels and percent survival, were all slightly or significantly higher than that of the vaccinated group without pre-treatment with NB-O₃. Taken together, our findings suggest that utilizing pre-treatment with NB-O₃ may improve the immune response and efficacy of immersion vaccination in Nile tilapia.

Vaccines prepared from formalin-killed Streptococcus agalactiae were administered to Nile tilapia (Oreochromis niloticus) via three different routes: immersion in a water-based vaccine, injection with an oil-based vaccine, and as a water-based oral vaccine. All vaccination treatments increased lysozyme and peroxidase activity in skin mucus of Nile tilapia by 1.2- to 1.5-fold compared to their activities in unvaccinated control fish. Likewise, alternative complement, phagocytosis, and respiratory burst activities in the blood serum of the vaccinated fish were 1.2- to 1.5-times higher than in the unvaccinated fish. In addition, the expression transcripts of interleukin-1 (IL-1), interleukin-8 (IL-8), and lipopolysaccharide-binding protein (LBP) were 2.3- to 2.9-fold higher in the vaccinated fish compared to those in the unvaccinated control. The unvaccinated fish challenged with Streptococcus agalactiae had a survival rate of 25% compared to a survival rate of 78–85% for the vaccinated fish. The differences between the unvaccinated and vaccinated fish were all statistically significant, but there was no significant difference in any of the indicators of immunity between the three vaccinated groups. Collectively, these results confirm that vaccination with formalin-killed Streptococcus agalactiae significantly improved the resistance of Nile tilapia to infection by the pathogen. Overall, the efficacy of oral administration of the vaccine was comparable to that of vaccine administered via injection, indicating that oral vaccination is a viable cost-effective alternative to administering vaccines by injection.