Synergy of histone-derived peptides of coho salmon with lysozyme and flounder pleurocidin

Recent research has identified endogenous cationic antimicrobial peptides a s important factors in the innate immunity of many organisms, including fis h. It is known that antimicrobial activity, as well as lysozyme activity, c an be induced in coho salmon (Oncorhynchus kisutch) mucus after exposure of the fish to infectious agents. Since lysozyme alone does not have antimicr obial activity against Vibrio anguillarum and Aeromonas salmonicida, a four -step protein purification protocol was used to isolate and identify antiba cterial fractions from bacterially challenged coho salmon mucus and blood. The purification consisted of extraction with hot acetic acid, extraction a nd concentration on a C,, cartridge, gel filtration, and reverse-phase chro matography on a C,, column. N-terminal amino acid sequence analyses reveale d that both the blood and the mucus antimicrobial fractions demonstrated id entity with the N terminus of trout HI histone, Mass spectroscopic analysis indicated the presence of the entire histone, as well as fragments thereof , including a 26-amino-acid N-terminal segment, These fractions inhibited t he growth of antibiotic-supersuseptible Salmonella enterica serovar Typhimu rium, as well as A. salmonicida and V. anguillanrm, Synthetic peptides iden tical to the N-terminally acetylated or C-terminally amidated 26-amino-acid fragment were inactive in antimicrobial assays, but they potentiated the a ntimicrobial activities of the flounder peptide pleurocidin, lysozyme, and crude lysozyme-containing extracts from coho salmon. The peptides bound spe cifically to anionic lipid monolayers, However, synergy with pleurocidin di d not appear to occur at the cell membrane level. The synergistic activitie s of inducible histone peptides indicate that they play an important role i n the first line of salmon defenses against infectious pathogens and that w hile some histone fragments may have direct antimicrobial effects, others i mprove existing defenses.