Antimicrobial activity was detected in the ovary and oviduct tissues of healthy mature White Legborn hens, Gallas gallas. Two anti-microbial proteins were purified to homogeneity using acid extraction followed by multiple steps of chromatography and the pure proteins were further characterized biochemically. Peptide mixtures obtained after enzymatic digestion of the chicken antimicrobial proteins were analyzed using peptide mass fingerprinting and partial sequencing by tandem nanoelectrospray mass spectrometry and the proteins were identified as histones H1 and H2B. Chicken historic antimicrobial proteins were active against, both Gram-positive and Gram-negative bacteria. The abundance of these proteins in the reproductive tissues and their broad-spectrum antimicrobial nature may indicate their defensive role against pathogens during the follicle development in the ovary and egg formation in the oviduct. The discovery of antimicrobial histones in chicken reproductive system provides further evidence that histones may play a role in innate immunity against microorganisms in a wide range of animal species. (c) 2005 Elsevier Inc. All rights reserved.