In this study, we investigate the effect of the probiotic E. faecium 669 strain on the gut integrity of broilers and the effect on intestinal colonization with Salmonella Enteritidis. In the in vivo experiment, 120-day-old broilers (Ross 308) were divided into 4 equally sized groups. Group A received the probiotic as a single dose by spray at d 18 of incubation and group B received the probiotic in the drinking water daily throughout the experiment. Group C was untreated control. Group D received the antibiotic Apramycin sulfate in the drinking water. Broilers in all four groups were challenged with S. Enteritidis by oral gavage at d 8 of life. From d 9 to 12, a cloacal swab was collected from all broilers for culturing on Salmonella selective media to determine the shedding. At d 12, birds were euthanized and S. Enteritidis in ceca were enumerated and intestinal samples for histology and host gene expression were collected. The group receiving the probiotic in the drinking water shed significantly less S. Enteritidis compared to the untreated control group at all times. The group receiving a single probiotic application before hatch showed a reduced shedding of Salmonella at d 9 and 10. S. Enteritidis was not detected in the ceca of the antimicrobial treated broilers. Histology of jejuni samples and host gene expression showed that intestinal integrity was enhanced by adding probiotic to the drinking water. Overall, the study shows that pre-hatch and daily application of the probiotic strain E. faecium 669 reduces the colonization of broilers with S. Enteritidis and daily application enhances gut integrity. Application of the probiotic E. faecium strain can be recommended as a method to reduce the colonization of broilers with S. Enteritidis and enhance their gut integrity.