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Concerns over the increasing emergence of antibiotic-resistant pathogenic microorganisms due to the overuse of antibiotics and the lack of effective antibiotics for livestock have prompted efforts to develop alternatives to conventional antibiotics. Antimicrobial peptides (AMPs) with a broad-spectrum activity and rapid killing, along with little opportunity for the development of resistance, represent one of the promising novel alternatives. Their high production cost and cytotoxicity, however, limit the use of AMPs as effective antibiotic agents to livestock. To overcome these problems, we developed potent antimicrobial  Escherichia coli  displaying multimeric AMPs on the cell surface so that the AMP multimers can be converted into active AMP monomers by the pepsin in the stomach of livestock. Buf IIIb, a strong AMP without cytotoxicity, was expressed on the surface of  E. coli  as Lpp-OmpA-fused tandem multimers with a pepsin substrate residue, leucine, at the C-terminus of each monomer. The AMP multimers were successfully converted into active AMPs upon pepsin cleavage, and the liberated Buf IIIb-L monomers inhibited the growth of two major oral infectious pathogens of livestock,  Salmonella enteritidis  and  Listeria monocytogenes . Live antimicrobial microorganisms developed in this study may represent the most effective means of providing potent AMPs to livestock, and have a great impact on controlling over pathogenic microorganisms in the livestock production.

Display of Multimeric Antimicrobial Peptides on the Escherichia coli Cell Surface and Its Application as Whole-Cell Antibiotics