The microbiome may influence disease severity in atopic dermatitis. The skin of atopic dermatitis patients and healthy individuals was sampled in a standardized manner and the microbial composition analysed using next-generation sequencing. Optical density measurements were used to investigate bacterial growth under defined conditions in vitro. Lesional skin from patients with atopic dermatitis had a higher abundance of Staphylococcus aureus and reduced quantities of Propionibacterium acnes and Lawsonella clevelandensis compared with non-lesional skin. The abundance of P. acnes correlated negatively with that of S. aureus (ρ= -0.6501, p < 0.0001). Fermentation products of P. acnes inhibited the growth of S. aureus and S. epidermidis. Serum from patients with atopic dermatitis inhibited the growth of S. aureus to a greater extent than did serum from healthy individuals. These results suggest that selective modification of the skin microbiome could potentially be used as a therapeutic strategy in atopic dermatitis.