Data on the molecular response of the intestine to the food borne pathogen Salmonella are derived from in vitro studies, whereas in vivo data are lacking. We performed an oral S. enteritidis infection study in Wistar rats to obtain insight in the in vivo response in time. Expression profiles of ileal mucosa (IM) and Peyer's patches (PP) were generated using DNA microarrays at days 1, 3 and 6 post infection. An overview of Salmonella regulated processes was obtained and confirmed by quantitative real-time PCR on individual and pooled samples. Salmonella-induced gene expression responses in vivo are fewer and smaller than observed in vitro and the response develops over a longer period of time. Few effects are seen at day 1 and mainly occur in IM, suggesting the mucosa as the primary site of invasion. Later a bigger response is observed, especially in PP. Decreased expression of anti-microbial peptides genes (in IM at day 1) suggests inhibition of this process by Salmonella. Newly identified target processes are carbohydrate transport (increased expression in IM at day 1) and phase I and phase II detoxification (decreased expression at day 3 and 6). Increase of cytokine and chemokine expression occurs at later time points, both in PP and IM. Pancreatitis-associated protein, lipocalin 2 and calprotectin, potential inflammatory marker proteins, showed induced expression from day 3 onwards. We conclude that the in vivo gene expression response of the ileum to Salmonella differs to a large extend from the response seen in vitro.