We have shown recently that dietary calcium phosphate (CaPi) has a trophic effect on the intestinal microflora and strongly protects against salmonella infection. It was speculated that precipitation by CaPi of intestinal surfactants, such as bile acids and fatty acids, reduced the cytotoxicity of intestinal contents and favored growth of the microflora. Because lactobacilli may have antagonistic activity against pathogens, the main purpose of the present study was to examine whether this CaPi-induced protection coincides with a reinforcement of the endogenous lactobacilli. In vitro, Salmonella enteritidis appeared to be insensitive to bile acids and fatty acids, whereas Lactobacillus acidophilus was killed by physiologically relevant concentrations of these surfactants. Additionally, after adaptation to a purified diet differing only in CaPi concentration (20 and 180 mmol CaHPO4. 2H2O/kg), rats (n = 8) were orally infected with S. enteritidis. Besides reducing the cytotoxicity and the concentration of bile acids and fatty acids of ileal contents and fecal water, CaPi notably changed the composition of ileal bile acids in a less cell-damaging direction. Significantly greater numbers of ileal and fecal lactobacilli were detected in noninfected, CaPi-supplemented rats. As judged by the lower urinary NOx excretion, which is a biomarker of intestinal bacterial translocation, dietary CaPi reduced the invasion of salmonella. Additionally, the colonization resistance was improved considering the reduction of excreted fecal salmonella. In accordance, fewer viable salmonella were detected in ileal contents and on the ileal mucosa in the CaPi group. In conclusion, reducing the intestinal surfactant concentration by dietary CaPi strengthens the endogenous lactobacilli and increases the resistance to salmonella.