Aquaculture is steadily increasing in the food industry, but a major challenge in aquaculture is to make the production process more environmentally sustainable
. This means that we need to find ways to protect nature while farming fish and other aquatic life. In recent years, the use of FM and FO in feed has decreased due to the high environmental impact and increased market prices. Therefore , alternative feed ingredients are being sought to meet nutritional needs while minimizing environmental impact and costs. One of the promising alternative nutrient sources for fish feed is insect meal , with Hermetia illucens meal (HIM ) being particularly valuable. It contains biologically active compounds such as chitin, antimicrobial peptides, and short-to-medium chain fatty acids (FAs). These compounds enhance the innate immune response of fish and alter the composition of the gut microbiome, in which has a positive effect on gut health
. Although many studies have explored the use of insects as sustainable protein sources in fish feed, most have been conducted on a small-scale and in controlled environments. There are only a f ew large-scale studies . Therefore, the current study investigated the effect of replacing FM with HIM in the diet of gilthead seabream (Sparus aurata ). Growth performance, gut health and gut microbiota profiles were assessed to evaluate the health of the fish and their response to the diet.
Materials and methods
Approximately 60,000 gilthead seabream with an average initial weight of 131±1.4 g were randomly allocated to four coastal cages (two cages per diet). They were fed either a control diet (FM) or an experimental diet (HIM), with 11% defatted HIM replacing the FM by 35%. Feeding occurred twice daily, six days per week, at rates between 0.6 and 1.3% of biomass. A total of 100 fish per cage were collectively weighed to monitor biomass. After 25 weeks, 88 fish per diet were sacrificed to measure growth parameters . Liver samples from 24 specimens (12 fish per diet, six per cage) were weighed to calculate the hepatosomatic index (HSI). In a ddition, liver and intestinal samples (proximal and distal parts) from ten fish were used for histologic analysis. For the analysis of gut microbiota, ten fish per diet were sacrificed and their intestines (excluding pyloric ceca) were collected. The digesta and mucosal microbiota were collected and mixed. High-throughput sequencing of 16S rRNA gene amplicons (MiSeq platform, Illumina) was used to characterize the gut microbial community profile, as described in detail in
. Fecal samples were collected from nine fish per tank for volatile fatty acid analysis. Statistical tests, including Student’s t-test or Mann–Whitney U-test for data distribution, were used to compare growth performance, alpha diversity indices, and bacterial abundance between groups. PERMANOVA was used to assess variations in beta diversity. Statistical analyses were performed using Past4 v. 4.02 software, with significance set at p < 0.05.
Results and Discussion
The 25-week feeding trial showed that growth parameters, including final body weight, specific growth rate, feed conversion rate, and HSI , were not affected by the diet. In addition , gross examination of the gastrointestinal tract and liver showed no signs of inflammation, indicating that the overall health of the gilthead seabream was not affected by the experimental diets.
Analysis of the gut microbial community profile by high-throughput sequencing showed consistent dominance of phyla Proteobacteria, Fusobacteria, and Firmicutes in the gilthead seabream gut microbiota, regardless of the diet administered. Despite the diet variations, the assessment of alpha diversity showed no significant differences between the two feeding groups. Analysis of the Venn diagram revealed a common core microbiota comprising only eight species, indicating a stable microbial community structure in all groups (Figure 1). In particular , there was a significant decrease in the abundance of Cetobacterium and an increase in the relative abundance of genera Oceanobacillus and Paenibacillus , indicating a modulation of gut microbial populations by the diet.
Further analysis using multivariate permutational analysis revealed statistically significant differences in gut bacterial communities between the two feeding groups, highlighting the effects of dietary interventions on microbial composition. Estimation of the metabolic activities of the microbial community using the PICRUSt software revealed that fish fed the insect diet had an increased number of genes associated with bacterial chemotaxis, motility, and the two-component signal transduction system . This suggests a possible role of insect meal in modulating microbial metabolic pathways associated with host-microbe interactions.
Moreover, insect consumption in the HIM group resulted in a significant increase in short-chain fatty acids (SCFAs), particularly acetic and propionic acids, which play a critical role in host energy metabolism and gut health, compared to the FM diet. In contrast, no significant differences were found in the levels of butyric acid and isobutyric acid in the fecal samples between the two experimental groups, indicating an effect of dietary measures on SCFA production. Overall, the results suggest that the consumption of HIM positively influences the composition of the gut microbiota and metabolic activities in gilthead seabream, possibly contributing to better nutrient utilization and gut health.
Conclusions
Our results show that the inclusion of HIM as an alternative animal protein source has a positive effect on the gut microbiota of sea bream. This includes an increase in the abundance of probiotic and chitinolytic bacterial genera, leading to improved nutrient utilization and production of short-chain fatty acids (SCFAs) that are absorbed by the host. These changes contributed to the energy metabolism of Sparus aurata in coastal farms.
Acknowledgements
This work was funded by the project “BIO = C = O,” PON-MISE I&C 2014–2020 FESR, ASSE I -Innovazione e Azione, Azione 1.1.3. D.M. 05/03/2018, project codex N. F/200078/01–03/X45, CUP B41B20000280005.
References