A truncated peptide Sp-NPFin from the neuropeptide FII SpNPFII of Scylla paramamosain exhibiting potent antimicrobial activity

Abstract

Neuropeptide (NP) F is a homolog of vertebrate neuropeptide Y, and mounting evidence suggests that it may play a role in innate immunity in addition to its function in invertebrate neurobiology. However, so far, there have been few reports on the immunological properties of crustacean NPs. Here, we cloned a neuropeptide FII gene, naming it SpNPFII, from the mud crab Scylla paramamosain. The full-length cDNA sequence of SpNPFII was 551 bp, encoding 124 amino acids. Structural prediction suggested that the SpNPFII mature peptide mainly formed an α-helix. SpNPFII was predominately expressed in male crab gonadal tissues, with the highest expression in the anterior vas deferens and seminal vesicle. The expression level of SpNPFII changed significantly under challenge by lipopolysaccharide (LPS) or Vibrio alginolyticus. After bioinformatic analysis, a truncated 45-amino acid peptide derived from SpNPFII and having potent antimicrobial activity was identified and named Sp-NPFin. Sp-NPFin exerted strong, broad-spectrum antibacterial (minimum inhibitory concentration [MIC] ranged from 1.5 to 12 μM) and antimildew activity (MIC ranged from 1.5 to 48 μM), a low fungicidal concentration (e.g., minimum bactericidal concentration [MBC] for Fusarium oxysporum was 1.5 to 3 μM) and rapid bactericidal kinetic (killed all bacteria within 1 to 6 h). Sp-NPFin could bind to several microbial surface components (e.g. LPS, lipoteichoic acid, peptidoglycan and glucan), and induce significant damage to microbial membranes and cause leakage of cell contents in a concentration-dependent manner. Confocal microscopy indicated that Sp-NPFin was localized at microbial surfaces. In addition, Sp-NPFin exhibited no cytotoxicity to primary cultured crab hemocytes and mammalian cells, and in vivo study demonstrated that it could significantly improve the survival rate of S. paramamosain under the challenge of V. alginolyticus. Taken together, SpNPFII might play a role in the immune system of S. paramamosain, and its truncated peptide, Sp-NPFin, would be a promising antibacterial and antimildew agent with broad application prospects in the fields of aquaculture, veterinary practices and medicine.