The effect of acquired resistance to bile on the antimicrobial sensitivity of two Bifidobacterium strains (Bifidobacterium animalis IPLA4549 and Bifidobacterium longum NIZO B667) was studied. The MICs of 23 different antibiotics belonging to the most clinically important groups were determined by using the Etest method, which comprises nonporous plastic strips calibrated with a predefined gradient of antibiotic concentrations covering 15 twofold dilutions. The strains were sensitive to most antibiotics assayed, although they tolerated relatively high concentrations of gentamicin, kanamycin, streptomycin, polymyxin B, and ciprofloxacin (from 32 to more than 1,024 microg/ml). One of the bile-adapted strains was more strongly resistant to ceftazidime than was its parent bile-sensitive strain, and the other bile-adapted strain had increased resistance to tetracyclines. Therefore, to test the possibility that the acquisition of stable resistance to bile could be associated with a general increase in resistance to some antibiotics, we analyzed the sensitivities of four additional pairs of parent strains and their bile-adapted derivatives to ceftazidime and three tetracyclines (doxycycline, minocycline, and tetracycline). Three of the bile-resistant derivatives had increased resistance to ceftazidime (more than 256-fold) compared with their parents, and two had enhanced resistance to tetracyclines (at least 12-fold). Thus, the acquisition of bile salts resistance in Bifidobacterium induced modifications of the antibiotic resistance patterns. These results suggest that adaptation of probiotics to bile could also change their potential impact on intestinal microbiota, and this possibility deserves further attention.