The antibacterial activity of a novel NK-lysin homolog and its molecular characterization and expression in the striped catfish, Pangasianodon hypophthalmus.

Aeromonas hydrophila was a common bacterial pathogen in aquaculture resulting in considerable losses to the striped catfish aquaculture industry. As an emergent antimicrobial peptide (AMP), NK-lysin (NKL) had activity against various microorganisms. However, the antibacterial activity of NKL from striped catfish (Pangasianodon hypophthalmus) both in vitro and vivo remains unclear. In this study, the cDNA sequence of P. hypophthalmus NK-lysin gene (PhNK-lysin) was cloned and characterized. The amino acid sequence of PhNK-lysin contains a signal peptide sequence of 17 amino acid (aa) residues and a mature peptide composed of 130 aa. The saposin B domain of mature peptide comprised six conserved cysteines forming three putative disulfide bonds. Phylogenetic analysis revealed that the PhNK-lysin was most closely related to that of the channel catfish (Ictalurus punctatus) NK-lysin. The transcriptional levels of the PhNK-lysin were significantly upregulated in response to A. hydrophila infection in various tissues including heart, liver, spleen, head kidney, trunk kidney and gill. The synthetic PhNK-lysin-derived peptide consisting of 38aa showed antibacterial activity against Vibrio harveii, Aeromonas hydrophila and Escherichia coli. The MIC for V. harveii, A. hydrophila and E. coli were 15.625 µM, 250 µM and 31.25 µM respectively. Besides, the synthetic PhNK-lysin decreased the bacterial load of liver and trunk kidney in vivo as well as increased the survival rate of A. hydrophila infected striped catfish. Hence, these data suggest that PhNK-lysin had antimicrobial effect and protects the host from pathogenic infection.

Effects of dietary toxic cyanobacteria and ammonia exposure on immune function of blunt snout bream (Megalabrama amblycephala).

Cyanobacterial blooms caused by water eutrophication have become a worldwide problem. During the degradation of toxic cyanobacterial blooms, elevated ammonia and microcystins concentrations co-occur and exert toxicity on fish. Up to now, the combined effect of microcystins and ammonia on fish immunotoxicity has not been reported. The present study investigated immune responses of blunt snout bream (Megalabrama amblycephala) to dietary toxic cyanobacteria and ammonia exposure. Megalobrama amblycephala were exposed to solutions with different concentrations of NH3-N (0, 0.06, 0.12 mg/L) and fed with diets containing 15% and 30% of toxic cyanobacteria lyophilized powder for 30 d. The microcystins concentration in different organs of Megalobrama amblycephala was in the following sequence: head kidney > liver > intestine > gonad > spleen > gill > trunk kidney > brain > muscle > heart. In both head kidney and spleen, the MC-LR and MC-RR concentration increased significantly with increasing NH3-N concentration. It indicates that NH3-N maybe promote the accumulation of microcystins in immune organs of Megalobrama amblycephala. Meanwhile, broadened peripheral interspace of lymphocytes, nucleus shrivel and edematous mitochondria were observed in head kidney lymphocyte of toxic treatment fish. Moreover, there were significant interactions between dietary toxic cyanobacteria and ammonia exposure on head kidney macrophage phagocytosis activity, respiratory burst activities, total number of white blood cells and the transcriptional levels of sIgM, mIgD and sIgZ genes. Our data clearly demonstrated that dietary toxic cyanobacteria combined with ammonia exposure showed a synergistic effect on Megalobrama amblycephala immunotoxicity.