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

Abstract

BackgroundIn 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.ResultsThere 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-1β (il1β, 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 (il1β, cox2 and igm). However, major histological changes in gut were not detected.ConclusionsUsing 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 β, 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.Electronic supplementary materialThe online version of this article (10.1186/s12917-018-1626-6) contains supplementary material, which is available to authorized users.

Intestinal Explant Cultures from Gilthead Seabream (Sparus Aurata, L.) Allowed Determining the Mucosal Sensitivity to Bacterial Pathogens and the Impact of a Plant Protein Diet

“…IL-1β is a known canonical master regulator of pro-in ammatory processes, it is secreted in response to gram negative bacteria and its release is followed by the production of IL-6 and other cytokines in the pro-in ammatory cascade [46,53]. Modi cation in diet composition changed the expression of IL-1β, in agreement with previous studies, thus indicating that after plant protein diet rendered an already sensitized intestine to in ammatory/stress stimuli [32,[53][54][55].…”

“…In phase II (up to 250 g), there were no signi cant differences between experimental groups (FM and PP), thus supporting that older sh are more tolerant to diet plant proteins than sh in earlier growth stages [16]. It is noteworthy the lack of in ammatory response in PP* group (short PP exposure), supporting that PP long term feeding has deeper alterations in sh gut and contributes to changes in the microbiota and an increase of sh mortality [32], in agreement with previous studies [33,67,68],…”

“…Previous studies in seabream demonstrated that the use of plant proteins induced signi cant alterations of the gut microbiota, gut gene expression and gut proteomic pro le [32,42,43]. Nonetheless, these alterations can be affected to different extent by the feeding period or sh stage.…”

“…Fish, rearing system conditions, diets and feeding conditions A total number of 240 juveniles of gilthead seabream (averagare weigth 7,5 g and 60 days) were obtained from the sh farm BERSOLAZ (Bersolaz Spain, S.L.U, Culmarex Group) located in Port de Sagunt (Valencia, Spain) and transported to the facilities at the Universitat Politècnica de València, where the growth trial was conducted after 15 days of adaptation to experimental conditions. Features of the system and water parameters set were described in previous growth trials carried out in these facilities [32,33]. Lighting conditions were determined by the natural photoperiod.…”

“…Based on the gene expression analysis used in previous studies in this species (Sparus aurata) to evaluate the intestinal in ammatory and immune status [32] tRNA was extracted from intestinal tissue samples using the phenol/chloroform method with Trizol Reagent (Invitrogen, Spain) and treated with DNAse I (Roche) to remove DNases. Total RNA concentration, quality and integrity were assessed using a NanoDrop 2000C Spectrophotometer (Fisher Scienti c SL, Spain).…”