As a foodborne pathogen of global importance, serovar Enteritidis ( Enteritidis) is a threat to public health that is mainly spread by poultry products. Intestinal Enterobacteriaceae can inhibit the colonization of Enteritidis and are regarded as a potential antibiotic substitute. We investigated, in chicks, the anti- Enteritidis effects of () Nissle 1917, the most well-known probiotic member of Enterobacteriaceae. Eighty 1-d-old healthy female AA broilers were randomly divided into 4 groups, with 20 in each group, namely the negative control (group P), the Nissle 1917-treated group (group N), the . Enteritidis-infected group (group S) and the Nissle 1917-treated and . Enteritidis-infected group (group NS). From d 5 to 7, chicks in groups N and NS were orally gavaged once a day with Nissle 1917 and in groups P and S were administered the same volume of sterile PBS. At d 8, the chicks in groups S and NS were orally gavaged with . Enteritidis and in groups P and N were administered the same volume of sterile PBS. Sampling was conducted 24 h after challenge. Results showed that gavage of Nissle 1917 reduced the spleen index, loads, and inflammation ( < 0.05). It improved intestinal morphology and intestinal barrier function ( < 0.05). Enteritidis infection significantly reduced mRNA expression of angiotensin-converting enzyme 2 () and solute carrier family 6-member 19 () in the cecum and the content of Gly, Ser, Gln, and Trp in the serum ( < 0.05). Pretreatment with Nissle 1917 yielded mRNA expression of and in the cecum and levels of Gly, Ser, Gln, and Trp in the serum similar to that of uninfected chicks ( < 0.05). Additionally, Nissle 1917 altered cecum microbiota composition and enriched the abundance of , Lactobacillales, and Lachnospiraceae. These findings reveal that the probiotic Nissle 1917 reduced Enteritidis infection and shows enormous potential as an alternative to antibiotics.