Intestinal homeostasis of juvenile tiger puffer Takifugu rubripes was sensitive to dietary arachidonic acid in terms of mucosal barrier and microbiota

Abstract

A 74-day feeding trial was conducted to investigate the effects of different levels of dietary arachidonic acid (ARA) on mucosal barrier and microbiota in the intestine of tiger puffer Takifugu rubripes. Three isonitrogenous and isolipidic experimental diets were formulated to contain different ARA levels, i.e., 0.33 (Control), 2.45 (L-ARA), and 23.41 (H-ARA) g kg−1 dry matter. Each diet was fed to triplicate tanks. The results showed that the integrity of intestinal epithelial cells was impaired by increasing ARA levels. The leucocyte infiltration in the lamina propria was also enhanced by dietary ARA supplementation. Compared to the control group, group H-ARA had significantly down-regulated gene expression of intestinal tight junction proteins (claudin-4, claudin-7 and zonula occludens-1), but significantly up-regulated expression of myosin light chain kinase (MLCK). Gene expression of claudin-7 was significantly down-regulated also in group L-ARA. The intestinal lysozyme activity significantly decreased with increasing dietary ARA levels. Compared to the Control, diet H-ARA significantly up-regulated the gene expression of the pro-inflammatory cytokines tumor necrosis factor alpha (TNF-α) and interleukin 8 (IL-8), and down-regulated that of anti-inflammatory cytokines transforming growth factor-β1 (TGF-β1) in the hind intestine. Intestinal gene expression of Toll-like receptor 3 (TLR-3) and nuclear factor kappa B p100 (NF-κB p100) was significantly up-regulated by H-ARA. Sequencing of bacterial 16 s rRNA V4 region showed that Proteobacteria, Tenericutes and Firmicutes were dominant in the intestine of tiger puffer. Fish fed diet H-ARA showed significantly decreased alpha diversity of microbiota and increased relative abundance of Arcobacter, a potential pathogenic genus. These results suggested that dietary ARA tended to induce the dysfunction of intestinal mucosal barrier and modulate the intestinal bacterial community, and MLCK and TLR3/NF-κB signaling were possibly involved in these processes. The ARA content in diets for tiger puffer needs to be carefully managed considering the intestinal sensitivity to dietary ARA of this fish.