Impact of high dietary plant protein with or without marine ingredients in gut mucosa proteome of gilthead seabream (Sparus aurata, L.).

The digestive tract, particularly the intestine, represents one of the main sites of interactions with the environment, playing the gut mucosa a crucial role in the digestion and absorption of nutrients, and in the immune defence. Previous researches have proven that the fishmeal replacement by plant sources could have an impact on the intestinal status at both digestive and immune level, compromising relevant productive parameters, such as feed efficiency, growth or survival. In order to evaluate the long-term impact of total fishmeal replacement on intestinal mucosa, the gut mucosa proteome was analysed in fish fed with a fishmeal-based diet, against plant protein-based diets with or without alternative marine sources inclusion. Total fishmeal replacement without marine ingredients inclusion, reported a negative impact in growth and biometric parameters, further an altered gut mucosa proteome. However, the inclusion of a low percentage of marine ingredients in plant protein-based diets was able to maintain the growth, biometrics parameters and gut mucosa proteome with similar values to FM group. A total fishmeal replacement induced a big set of underrepresented proteins in relation to several biological processes such as intracellular transport, assembly of cellular macrocomplex, protein localization and protein catabolism, as well as several molecular functions, mainly related with binding to different molecules and the maintenance of the cytoskeleton structure. The set of downregulated proteins also included molecules which have a crucial role in the maintenance of the normal function of the enterocytes, and therefore, of the epithelium, including permeability, immune and inflammatory response regulation and nutritional absorption. Possibly, the amino acid imbalance presented in VM diet, in a long-term feeding, may be the main reason of these alterations, which can be prevented by the inclusion of 15% of alternative marine sources. SIGNIFICANCE: Long-term feeding with plant protein based diets may be considered as a stress factor and lead to a negative impact on digestive and immune system mechanisms at the gut, that can become apparent in a reduced fish performance. The need for fishmeal replacement by alternative ingredients such as plant sources to ensure the sustainability of the aquaculture sector has led the research assessing the intestinal status of fish to be of increasing importance. This scientific work provides further knowledge about the proteins and biologic processes altered in the gut in response to plant protein based diets, suggesting the loss of part of gut mucosa functionality. Nevertheless, the inclusion of alternative marine ingredients was able to reverse these negative effects, showing as a feasible option to develop sustainable aquafeeds.

Long-term feeding with high plant protein based diets in gilthead seabream (Sparus aurata, L.) leads to changes in the inflammatory and immune related gene expression at intestinal level.

BACKGROUND: In order to ensure sustainability of aquaculture production of carnivourous fish species such as the gilthead seabream (Sparus aurata, L.), the impact of the inclusion of alternative protein sources to fishmeal, including plants, has been assessed. With the aim of evaluating long-term effects of vegetable diets on growth and intestinal status of the on-growing gilthead seabream (initial weight = 129 g), three experimental diets were tested: a strict plant protein-based diet (VM), a fishmeal based diet (FM) and a plant protein-based diet with 15% of marine ingredients (squid and krill meal) alternative to fishmeal (VM+). Intestines were sampled after 154 days. Besides studying growth parameters and survival, the gene expression related to inflammatory response, immune system, epithelia integrity and digestive process was analysed in the foregut and hindgut sections, as well as different histological parameters in the foregut. RESULTS: There were no differences in growth performance (p = 0.2703) and feed utilization (p = 0.1536), although a greater fish mortality was recorded in the VM group (p = 0.0141). In addition, this group reported a lower expression in genes related to pro-inflammatory response, as Interleukine-1ß (il1ß, p = 0.0415), Interleukine-6 (il6, p = 0.0347) and cyclooxigenase-2 (cox2, p = 0.0014), immune-related genes as immunoglobulin M (igm, p = 0.0002) or bacterial defence genes as alkaline phosphatase (alp, p = 0.0069). In contrast, the VM+ group yielded similar survival rate to FM (p = 0.0141) and the gene expression patterns indicated a greater induction of the inflammatory and immune markers (il1ß, cox2 and igm). However, major histological changes in gut were not detected. CONCLUSIONS: Using plants as the unique source of protein on a long term basis, replacing fishmeal in aqua feeds for gilthead seabream, may have been the reason of a decrease in the level of different pro-inflammatory mediators (il1 ß, il6 and cox2) and immune-related molecules (igm and alp), which reflects a possible lack of local immune response at the intestinal mucosa, explaining the higher mortality observed. Krill and squid meal inclusion in vegetable diets, even at low concentrations, provided an improvement in nutrition and survival parameters compared to strictly plant protein based diets as VM, maybe explained by the maintenance of an effective immune response throughout the assay.

Inclusion of alternative marine by-products in aquafeeds with different levels of plant-based sources for on-growing gilthead sea bream (Sparus aurata, L.): effects on digestibility, amino acid retention, ammonia excretion and enzyme activity.

The search for new sustainable aquafeeds for the species with greater economic importance, such as the gilthead sea bream in Europe, is one of the main challenges in the aquaculture sector. The present work tested fishmeal replacement by a mixture of plant meals at different levels, as well as the use of marine by-products with attractant properties and high-quality protein in high plant protein diets. In order to do that, effects on growth and biometric parameters, digestibility, amino acid retention, excreted ammonia and proteases and amylase activity were assessed, using six different diets: FM100 (100% of protein provided by fishmeal), FM50 (50% of replacement), FM25 (75% of replacement) and FM0 (100% of replacement), but also FM25+ (75% of replacement and 15% of squid and krill meal inclusion), and FM0+ (100% of replacement and 15% of squid and krill meal inclusion). In group FM0, a clear impact of dietary changes was observed on growth, survival and ammonia excretion. Amino acid retention in group FM0+ was also significantly affected, which can be explained by the limited content of certain amino acids in this diet. On the other hand, no significant differences were observed in most biometric parameters or in enzyme activity. In conclusion, complete fishmeal replacement can be achieved by using a mixture of plant-based sources, but supplementation with complementary marine ingredients can prevent detrimental effects on growth, survival, nutritional parameters and protein metabolism.

Potential use of high levels of vegetal proteins in diets for market-sized gilthead sea bream (Sparus aurata).

The effect of partial or total dietary substitution of fishmeal (FM) by vegetal protein sources on growth and feed efficiency was carried out in on-growing gilthead sea bream (mean initial weight 131 g). The Control diet (FM 100) contained FM as the primary protein source, while in Diets FM 25 and FM 0 the FM protein was replaced at 75% and 100%, respectively, by a vegetable protein mixture consisting of wheat gluten, soybean meal, rapeseed meal and crystalline amino acids. Diets FM 25 and FM 0 also contained krill meal at 47 g/kg in order to improve palatability. At the end of the trial (after 158 d), fish survival was above 90%. Final weight and the specific growth rate were statistically lower in fish fed the Control diet (361 g and 0.64%/d), compared with 390-396 g and 0.69-0.70%/d after feeding vegetal diets. No significant differences were found regarding feed intake and feed conversion ratio. The digestibility of protein and amino acids (determined with chromium oxide as indicator) was similar in all diets. The blood parameters were not significantly affected by treatments. The activity of trypsin and pepsin was significantly reduced after feeding Diet FM 0. In the distal intestine, the villi length in fish fed Diet FM 25 was significantly longer and the intestine of the fish fed the FM 100 diet showed a smaller number of goblet cells. In conclusion, a total FM substitution by a vegetal mix supplemented with synthetic amino acids in on-growing sea bream is feasible.