Partial purification of antibacterial proteinaceous factors from erythrocytes of Oncorhynchus mykiss

Antimicrobial peptides are natural antibiotics known to be present in both myeloid cells and epithelial surfaces of vertebrates. Nevertheless, the reports of antimicrobial peptides isolated from blood cells of teleosts are scarce. In this paper we show that acid-soluble erythrocyte extracts from rainbow trout, Oncorhynchus mykiss, display antibacterial activity against Planococcus citreus on a radial diffusion assay. Following tC18 solid phase extraction, cationic exchange chromatography and C18 reversed phase HPLC, two groups of fractions with antibacterial properties were obtained. This antibacterial activity is thermostable and susceptible to digestion by proteinase K, thus showing that the antibacterial agents have a proteinaceous nature. The factors eluted from a C18 column with circa 33% acetonitrile are active against P. citreus and Escherichia coli, with minimal inhibitory concentrations in the range 7-14 microg ml(-1) and 14-28 microg ml(-1), respectively; the ones eluted with approximately 44% acetonitrile on the same column only displayed activity against P. citreus, with a minimal inhibitory concentration of 1-2 microg ml(-1). These results raise the possibility that trout erythrocytes may contain antimicrobial factors not previously considered to be part of the innate immune system.