Summary Background The rise in atopic diseases has been linked to disturbances in the intestinal microbiota composition. Objective The purpose of this study was to investigate the intestinal microbiota composition in infants in whom atopic (IgE-associated) eczema was or was not developing, using a molecular fingerprinting technique. Methods Within a prospective birth cohort study, fecal samples have been collected at the infant's age of 1 month. Within the context of this cohort, we conducted a nested case-control study comparing fecal samples of 26 infants who became sensitized and developed eczema within the first year of life with 52 non-sensitized non-eczematous infants. The composition of the fecal samples was examined using PCR combined with denaturing gradient gel electrophoresis. Using real-time PCR, total bacterial counts and bifidobacterial counts were enumerated. Results Neither total bacterial profiles nor the type and proportion of bifidobacteria in the feces were associated with the development of atopic eczema. The similarity of bacterial profiles was low; mean similarity was approximately 33% in both infants with or without atopic eczema. The prevalence of one specific band in total bacterial profiles was significantly higher in infants with atopic eczema compared with controls (96% vs. 71%, P=0.01). Identification of this band revealed that it represented Escherichia coli. Conclusion Although no association was found between the development of IgE-associated eczema and the dominant gut microbiota as a whole or with the bifdobacterial microbiota, the association with E. coli indicates that differences in gut microbiota do precede the development of atopy.