In order to generate Salmonella enterica serovar Enteritidis fimbriae antigens (rSEF21), the intact region encoding SEF21 was amplified from Salmonella Enteritidis by PCR and subcloned into a prokaryotic expression vector pET-28a(+) to yield pET-28a(+)-SEF21. The rSEF21 protein was highly expressed and purified by nickel affinity chromatography. Liposomeassociated rSEF21 was prepared for oral immunization to seek protective efficacy for intestinal infection with Salmonella Enteritidis. Evidence of IgA and IgG responses were found in the intestinal tracts and in the sera of a group of chickens immunized. Two weeks after the booster immunization, the chickens were challenged orally with 2 x 10(6) colony-forming units of live Salmonella Enteritidis, and fecal samples were examined for bacterial excretion from the intestinal tract. Significantly less fecal excretion of bacteria was observed in immunized chickens for 4 wk after challenge. The numbers of bacteria in the intestinal contents (cecum and rectum) were also significantly lower in immunized chickens than in unimmunized controls. Therefore, oral immunization with liposome-associated rSEF21 elicits both systemic and mucosal antibody responses, leading to a reduction in bacterial colonization in the intestinal tract and excretion of Salmonella Enteritidis in the feces.