Effects of dietary protein levels on the disease resistance, immune function and physical barrier function in the gill of grass carp (Ctenopharyngodon idella) after challenged with Flavobacterium columnare

• Optimal level of dietary protein increased fish gill immune function.
• Optimal level of dietary protein prevented oxidative damage in the gills of fish.
• Optimal level of dietary protein inhibited apoptosis in the gills of fish.
• Optimal level of dietary protein strengthened tight junction in the gills of fish.
• Dietary proteins regulated NF-κB, TOR, Nrf2, JNK, MLCK and p38 MAPK signalling.

The effects of dietary protein levels on the disease resistance, gill immune function and physical barrier function of grass carp (Ctenopharyngodon idella) were investigated in this study. A total of 540 grass carp (264.11 ± 0.76 g) were fed six diets containing graded levels of protein (143.1, 176.7, 217.2, 257.5, 292.2 and 322.8 g digestible protein kg−1 diet) for 8 weeks. After the growth trial, fish were challenged with Flavobacterium columnare for 3 days. The results indicated that optimal levels of dietary protein had the following effects: (1) the production of antibacterial components increased, and anti-inflammatory cytokines, inhibitor of κBα, target of rapamycin and ribosomal protein S6 kinases 1 mRNA levels were up-regulated, whereas mRNA levels of pro-inflammatory cytokines, nuclear factor kappa B (NF-κB) P65, NF-κB P52, IκB kinase (IKK) α, IKKβ, IKKγ, eIF4E-binding proteins (4E-BP) 1 and 4E-BP2 were down-regulated in the gills of grass carp (P < 0.05), indicating that fish gill immune function was enhanced at an optimal level of dietary protein; (2) the activities and mRNA levels of antioxidant enzymes and glutathione content increased, the contents of reactive oxygen species, malondialdehyde and protein carbonyl (PC) decreased, and NF-E2-related factor 2, B-cell lymphoma protein-2, inhibitor of apoptosis proteins, myeloid cell leukemia-1 and tight junction complexes mRNA levels were up-regulated, whereas Kelch-like-ECH-associated protein (Keap) 1a, Keap1b, cysteinyl aspartic acid-protease 3, 8, 9, fatty acid synthetase ligand, apoptotic protease activating factor-1, Bcl-2 associated X protein, c-Jun N-terminal protein kinase, myosin light chain kinase and p38 mitogen-activated protein kinase mRNA levels were down-regulated in the gills of grass carp (P < 0.05), indicating that the fish gill physical barrier function improved at an optimal level of dietary protein. Finally, based on the gill rot morbidity, ACP activity and PC content, the optimal levels of dietary protein for grass carp were estimated to be 286.65 g kg−1 diet (253.73 g digestible protein kg−1 diet), 290.46 g kg−1 diet (257.76 g digestible protein kg−1 diet) and 296.25 g kg−1 diet (260.69 g digestible protein kg−1 diet), respectively.