1. Probiotics are beneficial microbes that are currently considered as alternatives to antimicrobial growth promoters (AGP) in animal nutrition. In addition, they are purported to suppress pathogens in the gut via the concept of competitive exclusion (CE). The aim of this work was to evaluate the efficacy of a multistrain probiotic against Salmonella enteritidis (SE) in broilers. 2. Three hundred and four, one-day-old, male Cobb broilers were fed a corn-soybean basal diet and were allocated to four experimental treatments for 6 weeks. Three of the four treatments were challenged with SE. Treatments were: C- (Negative Control, no SE challenge), C+ (Positive control, SE challenge, no other additives), AB (SE challenge + Avilamycin used as AGP) and PFW (SE challenge + probiotic in feed and water). Morbidity, mortality, incidence of Salmonella positive broilers, cecal microflora composition and antibody response (IgA and IgG) in the blood and intestine were determined as biomarkers of probiotic CE efficacy against SE. 3. The three SE challenged treatments had a significantly higher morbidity compared to C- during the first 2 weeks of age, while there were no differences between treatments regarding mortality. Salmonella shedding was evidenced in 75 and 50% of the birds at 5 and 21 d post-challenge, respectively. Microbiological analysis of cecal digesta in 42-d-old broilers revealed that while broilers in treatment C+ were 100% Salmonella positive, the broilers in treatments PFW and AB were 50% positive and in addition they had lower Salmonella levels (CFU/g digesta) by 27 logs compared to C+. At the age of 42 d significantly higher IgA and IgG specific immune responses against SE were detected at systemic and at intestinal level only for the positive control treatment (C+). 4. In conclusion, the lack of significant levels of specific IgA and IgG against SE at systemic and intestinal level combined with the lower prevalence of SE positive broilers and the lower cecal SE levels in treatments AB and PFW compared to C+, suggest that treatments PFW and AB were efficacious at reducing SE.