Sodium salt medium-chain fatty acids and Bacillus-based probiotic strategies to improve growth and intestinal health of gilthead sea bream (Sparus aurata)

Abstract

Background: The increased demand for fish protein has led to the intensification of aquaculture practices which are hampered by nutritional and health factors affecting growth performance. To solve these problems, antibiotics have been used for many years in the prevention, control and treatment against disease as well as growth promoters to improve animal performance. Nowadays, the use of antibiotics in the European Union and other countries has been completely or partially banned as a result of the existence of antibiotic cross-resistance. Therefore, a number of alternatives, including enzymes, prebiotics, probiotics, phytonutrients and organic acids used alone or in combination have been proposed for the improvement of immunological state, growth performance and production in livestock animals. The aim of the present study was to evaluate two commercially available feed additives, one based on medium-chain fatty acids (MCFAs) from coconut oil and another with a Bacillus-based probiotic, in gilthead sea bream (GSB, Sparus aurata), a marine farmed fish of high value in the Mediterranean aquaculture.

Methods: The potential benefits of adding two commercial feed additives on fish growth performance and intestinal health were assessed in a 100-days feeding trial. The experimental diets (D2 and D3) were prepared by supplementing a basal diet (D1) with MCFAs in the form of a sodium salt of coconut fatty acid distillate (DICOSAN^®; Norel, Madrid, Spain), rich on C-12, added at 0.3% (D2) or with the probiotic Bacillus amyloliquefaciens CECT 5940, added at 0.1% (D3). The study integrated data on growth performance, blood biochemistry, histology and intestinal gene expression patterns of selected markers of intestinal function and architecture.

Results: MCFAs in the form of a coconut oil increased feed intake, growth rates and the surface of nutrient absorption, promoting the anabolic action of the somatotropic axis. The probiotic (D3) induced anti-inflammatory and anti-oxidant effects with changes in circulating cortisol, immunoglobulin M, leukocyte respiratory burst, and mucosal expression levels of cytokines, lymphocyte markers and immunoglobulin T.

Discussion: MCFA supplementation showed positive effects on GSB growth and intestinal architecture acting mainly in the anterior intestine, where absorption takes place. The probiotic B. amyloliquefaciens CECT 5940 exhibited key effects in the regulation of the immune status inducing anti-inflammatory and anti-oxidant effects which can be potentially advantageous upon infection or exposure to other stressors. The potential effects of these feed additives in GSB are very promising to improve health and disease resistance in aquaculture.

Keywords: Bacillus amyloliquefaciens; DICOSAN; Intestinal health; Medium-chain fatty acid; Probiotic; Teleost.

Figure 1. Histological effects of experimental diets.

Photomicrographs of liver (A–C), anterior (D–I) and posterior (J–L) intestinal segments of gilthead sea bream fed with D1 (control) (A, D, G, J), D2 (supplemented with DICOSAN) (B, E, H, K) or D3 (supplemented with the probiotic Bacillus amyloliquefaciens) (C, F, I, L). Staining, scale bars: A–F = H&E, 100 µm; G–L = Giemsa, 10 µm. White arrows point to goblet cells. Black arrowheads point to submucosa with abundant eosinophilic granulocytes (pink cells). Inset in J (scale bar = 10 µm) shows the characteristic irregular and granular shaped eosinophilic granulocytes.

Figure 2. Effects of experimental diets on mast cell abundance in the intestinal submucosa.

Representative photomicrographs of anterior (A–C) and posterior (D–F) intestinal segments of gilthead sea bream fed with D1 (control) (A, D), D2 (supplemented with DICOSAN) (B, E) or D3 (supplemented with the probiotic Bacillus amyloliquefaciens) (C, F) and stained with an anti-histamine antibody. Histamine positive cells are located in the submucosa and appear brown. All pictures were taken using the same magnification (40×) and the scale bars are 10 µm. White arrows point to some representative mast cells in the submucosa. Inset in D (scale bar = 10 µm) shows a typical histamine positive cell with irregular shape and cytoplasmic granules.

Figure 3. Effects of experimental diets on genes related to intestinal architecture, absorption and immune response.

Hierarchical heatmap of fold-changes (experimental group (D2, D3) vs. control (D1)) for differentially expressed genes in at least one of the intestinal sections. AI, anterior intestine; PI, posterior intestine.