Lactobacillus reuteri ATCC 55730 is a probiotic (health-promoting) bacterium widely used as a dietary supplement. This study was designed to examine local colonization of the human gastrointestinal mucosa after dietary supplementation with L. reuteri ATCC 55730 and to determine subsequent immune responses at the colonized sites. In this open clinical investigation, 10 healthy volunteers and 9 volunteers with ileostomy underwent gastroscopy or ileoscopy and biopsy samples were taken from the stomach, duodenum, or ileum before and after supplementation with 4 ؋ 10 8 CFU of live L. reuteri ATCC 55730 lactobacilli per day for 28 days. Biopsy specimen colonization was analyzed using fluorescence in situ hybridization with a molecular beacon probe, and immune cell populations were determined by immunostaining. Endogenous L. reuteri was detected in the stomach of 1 subject and the duodenum of 3 subjects (out of 10 subjects). After L. reuteri ATCC 55730 supplementation, the stomachs of 8 and the duodenums of all 10 subjects were colonized. Three ileostomy subjects (of six tested) had endogenous L. reuteri at baseline, while all six displayed colonization after L. reuteri supplementation. Gastric mucosal histiocyte numbers were reduced and duodenal B-lymphocyte numbers were increased by L. reuteri ATCC 55730 administration. Furthermore, L. reuteri administration induced a significantly higher amount of CD4-positive T-lymphocytes in the ileal epithelium. Dietary supplementation with the probiotic L. reuteri ATCC 55730 induces significant colonization of the stomach, duodenum, and ileum of healthy humans, and this is associated with significant alterations of the immune response in the gastrointestinal mucosa. These responses may be key components of a mechanism by which L. reuteri ATCC 55730 exerts its well-documented probiotic effects in humans.Lactobacillus reuteri (13) is a heterofermentative bacterium that resides in the gastrointestinal tract of humans and animals (2, 11) and is considered to be one of the few true autochthonous (indigenous) Lactobacillus species in humans (14). L. reuteri ATCC 55730 (also designated SD2112) has been extensively studied and is widely used as a food additive to improve human gastrointestinal health. Oral administration delivers L. reuteri ATCC 55730 to the gastrointestinal tract, leading to shedding of live bacteria in the feces (16,20). Clinical trials have shown that L. reuteri ATCC 55730 administration is safe (20, 21), significantly reduces the incidence and the severity of diarrhea of different origins (16, 17), reduces gastrointestinal illness and infections (G. Asli, A. Alsheikh, and Z. Weizman, Abstr. 36th Annu. Meet. Eur. Soc. Paediatr. Gastroenterol. Hepatol. Nutr., abstr. O041, 2003), and may be useful in the relief of constipation in elderly subjects (12).Fecal colonization does not, however, provide information on the specific sites of colonization of L. reuteri ATCC 55730 along the gastrointestinal tract. Our aim was to document these specific sites and to examine possib...