Background: Egg white must provide nutrients and protection to the developing avian embryo. One way in which this is achieved is an arsenal of antimicrobial proteins and peptides which are essentially extensions of the innate immune system. Gallin is a recently identified member of a family of peptides that are found in egg white. The function of this peptide family has not been identified and they are potentially antimicrobial. Results: We have confirmed that there are at least 3 forms of the gallin gene in the chicken genome in 3 separate lines of chicken, all the forms are expressed in the tubular cells of the magnum region of the oviduct, consistent with its presence in egg white. mRNA expression levels are in the order 10,000 times greater in the magnum than the shell gland. The conservation between the multiple forms of gallin in the chicken genome compared with the conservation between gallin and other avian gallin like peptides, suggests that the gene duplication has occurred relatively recently in the chicken lineage. The gallin peptide family contains a six cysteine motif (C-X(5)-C-X(3)-C-X(11)-CX(3)-C-C) found in all defensins, and is most closely related to avian beta-defensins, although the cysteine spacing differs. Further support for the classification comes from the presence of a glycine at position 10 in the 41 amino acid peptide. Recombinant gallin inhibited the growth of Escherischia coli (E. coli) at a concentration of 0.25 mu M confirming it as part of the antimicrobial innate immune system in avian species. Conclusions: The relatively recent evolution of multiple forms of a member of a new defensin related group of peptides that we have termed ovodefensins, may be an adaptation to increase expression or the first steps in divergent evolution of the gene in chickens. The potent antimicrobial activity of the peptide against E. coli increases our understanding of the antimicrobial strategies of the avian innate immune system particularly those of the egg white and the evolution of the defensin family. The potential of this peptide and others in the family can now be investigated in a number of novel antimicrobial roles.