The study of the gut microbiome has received great attention in the aquaculture sector as an indicator of productivity and fish health, although many interactions among gut microbiome and fish host are still unknown. The promotion of a healthy gut microbiome by dietary intervention is currently of relevant interest, especially in the context of circular ingredients such as single-cell protein, insect meal and marine by-products, which are candidates to replace fishmeal and vegetable protein. Indeed, besides their optimal nutritional values, these novel ingredients may promote gut health by favouring beneficial gut microbiota taxa. The aim of this study is to explore the gut microbiome modulation induced by circular aquafeed ingredients in gilthead sea bream and European sea bass by analysing several dose-response feeding trials. Implications on gut microbiome functions and correlation with fish welfare parameters are discussed.
Materials and methods
Three feeding trials in gilthead sea bream and two in European sea bass were conducted in a closed recirculation aquaculture system (RAS). Gilthead sea bream (initial weight 75, 25 and 98g for trial 1 to 3, respectively were fed: 1) dietary increasing level of bacterial single-cell protein (SCP) from Corynebacterium glutamicum (0%, SCP, 10% SCP, SPC10; 15% SCP, SCP15; 20% SCP, SCP20) to replace vegetal ingredients; 2) dietary SCP from Candida utilis to replace fish meal, FM (0% CTRL, 5% SCP5, 7.5% SCP7.5, and 10% SCP10) and 3) dietary Hermetia illucens (HI) larva meal (0% CTRL, 5% HI5, 10% HI10, and 15% HI15) in partial substitution of FM. European sea bass (initial weight, 75g) were fed dietary increasing levels of fishery and aquaculture by-products and microalgae to totally replace wild-caught FM and soy protein. At the end of each trial, distal intestine content was collected for 16S rRNA gut microbiota analysis on 15 individual fish per treatment. Sequencing was performed on the Illumina MiSeq platform. Growth, feed efficiency parameters and welfare parameters by plasma biochemistry were also assessed. Microbiota biostatistical analysis was produced using R software (https://www.r-project.org/)
Results and Discussion
At the end of the trials, gilthead sea bream fed SCP from Corynebacterium glutamicum and HI larva meal showed a significant (p<0.05) variation compared to their respective control group, in terms of overall gut microbiome composition (Fig.1A and B). In the first case, SCP, even at the lowest content, led to an increase in Bacillus and Oceanobacillus relative abundance, which were the most relevant taxa responsible for a significant separation in the PCoA analyses. Similarly, HI induced a general gut microbiome reconfiguration with significant enrichment of Bacillus, Oceanobacillus and Paenibacillus. As also observed for animals fed with Corynebacterium glutamicum, no dose-response was detected for the ability of both raw materials to shift the gut microbiome structure, which was evident from the lowest dietary inclusion level. In both trials (especially with HI) a general reduction (p<0.05) of alpha diversity indices (i.e., PD_whole_tree, number of ASV and Shannon index) was also detected. In gilthead sea bream fed SCP from Candida utilis and in European sea bass fed with by-products from fisheries and aquaculture (Fig.1C and D), no significant overall impact of diet on the gut microbiome was observed. A common feature of these latest growth trials was a slightly decreasing growth performance (in terms of FCR and SGR) observed at the highest dietary inclusion level of the circular ingredients tested.
The inclusion of novel circular ingredients, such as bacterial single-cell protein and insect meal, clearly modulated the gut microbiome of gilthead sea bream promoting bacteria taxa belonging to Bacillaceae, which may exert beneficial host functions. Specific compounds characterizing these ingredients such as nucleotide/nucleic acids in SCP and chitins in insect meal might induced this modulation. This data match literature on other fish species and confirm the potential role of these ingredients to exert functional properties on gut health besides being a valid alternative for FM and vegetal ingredients at nutritional level.
This research was undertaken under the NextGenProteins (Next Generation Proteins for food and feed) project, grant agreement No 862704, and the NewTechAqua (New Technologies Aquaculture) project, grant agreement No 862658, both funded by European Union’s Horizon 2020, research and innovation programme.