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Pathogenic micro-organisms have evolved many strategies to counteract the antimicrobial peptides (AMPs) that they encounter upon entry into host systems. These strategies play vital roles in the virulence of pathogenic micro-organisms. The Salmonella enterica serovar Typhimurium genome has a gene cluster consisting of yejA, yejB, yejE and yejF genes, which encode a putative ATP-binding cassette (ABC) transporter. Our study shows that these genes constitute an operon. We deleted the yejF gene, which encodes the ATPase component of the putative ABC transporter. The [DELTA]yejF strain showed increased sensitivity to protamine, melittin, polymyxin B, human defensin (HBD)-1 and HBD-2, and was compromised in its capacity to proliferate inside activated macrophages and epithelial cells. Inside Intestine 407 cells, Salmonella was found to co-localize with human defensins HD-5 and HBD-1 ; this suggests that the ability to counteract AMPs in the intracellular milieu is important for Salmonella. In a murine typhoid model, the AyejF strain displayed decreased virulence when infected intragastrically. These findings suggest that the putative transporter encoded by the yejABEF operon is involved in counteracting AMPs, and that it contributes to the virulence of Salmonella.