Recently, the gut microbiome has been at the focus of nutritional aquaculture research because it has been shown to regulate nutrient uptake
and changes in fish metabolism
. Such functional flexibility of the gut microbiota likely increases the adaptability of fish digestion, and studies of the gut microbiome are becoming a necessity for aquaculture nutrition research. In this study, the effects of experimental diets on the gut microbiome of juvenile and subadult European seabass were investigated after a 120- and 147-day feeding trial, respectively.
Material and methods
In both experiments, diets were formulated to be iso-proteic (45%), iso-lipidic (20%), isoenergetic (20.3 MJ kg-1 ) and to meet the dietary requirements of juvenile and subadult European seabass. We used plant protein-based diet (CV), two plant-based diets in which graded amounts of plant protein mixtures were replaced with partially defatted Hermetia illucens pupae meal alone (VH10) or in combination with poultry byproduct meal (VH10P30), a fish meal diet (CF), and a fish meal diet supplemented with H. illucens (FH10).
Bacterial DNA was extracted from the whole intestine content of 15 juvenile and 20 subadult experimentally fed European seabass (3 juvenile replicates and 4 subadult replicates per treatment) using the Invitrogen PureLink Microbiome DNA Purification K it (Carlsbad, CA, USA) following the manufacturer’s protocol. The commercial services of Microsynth AG (Balgach , Switzerland) were used for library preparation based on Nextera two-step PCR. Variable region V4 of the 16S rRNA gene was successfully amplified from 15 juvenile and 18 subadult DNA extracts using the MiSeq2000 Next Generation system (Illumina, San Diego, CA, USA). The USEARCH algorithm
was applied to generate clusters (OTUs) and taxonomic assignment for the representative sequence of each OTU based on the RDP database
. Downstream analysis was performed using the Phyloseq package
for R (version 4.2.2).
Results and discussion
The results suggest that the gut microbiota of juvenile European seabass is more stable than that of subadult European seabass using the same experimental diet formulations, regardless of the diet tested. Although the majority of phyla detected were the same in juveniles and subadults (Actinobacteria, Bacteroidetes, Cyanobacteria/Chloroplast, Firmicutes, and Proteobacteria), the observed richness was lower in juveniles and there were no statistically significant differences between treatments for alpha and beta diversity. Interestingly, the overall growth performance of juvenile fish was worse on plant-based diets than on fish-based diets, whereas the opposite was observed in subadult fish. There is a possibility that changes in the gut microbiota of subadult fish increased their digestive adaptability and contributed to better performance on alternative plant-based diets containing poultry by-products and/or insect meal, while this effect was absent in juvenile European seabass. These results are important for the development of sustainable aquafeeds and the aquaculture industry.
Financial support for this study has been provided by Interreg AdriAquaNet (Project ID 10045161).
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