Shigella is a major cause of morbidity, mortality, and growth retardation for children in developing countries. Emergence of antibiotic resistance among Shigellae demands the development of effective medicines. Previous studies found that the endogenous antimicrobial peptide LL-37 is down-regulated in the rectal epithelium of patients during shigellosis and that butyrate up-regulates the expression of LL-37 in colonic epithelial cells in vitro and decreases severity of inflammation in experimental shigellosis. In this study, Shigella-infected dysenteric rabbits were treated with butyrate (0.14 mmol/kg of body weight) twice daily for 3 days, and the expression levels of the rabbit homologue to LL-37, CAP-18, were monitored in the colon. Butyrate treatment resulted in (i) reduced clinical illness, severity of inflammation in the colon, and bacterial load in the stool, (ii) significant up-regulation of CAP-18 in the surface epithelium, and (iii) disappearance of CAP-18-positive cells in lamina propria. The active CAP-18 peptide was released in stool from its proform by butyrate treatment. In healthy controls, CAP-18 expression was localized predominantly to the epithelial surface of the colon. In infected rabbits, CAP-18 expression was localized to immune and inflammatory cells in the colon, whereas the ulcerated epithelium was devoid of CAP-18 expression. The combination of CAP-18 and butyrate was more efficient in killing Shigella in vitro than CAP-18 alone. Our findings indicate that oral butyrate treatment in shigellosis may be of clinical value because of induction of the endogenous cathelicidin CAP-18 in the colonic epithelium, stimulation of the release of the active peptide CAP-18, and promoting elimination of Shigella.