Omiganan is an indolicidin analogue with antimicrobial properties that could be beneficial for patients with atopic dermatitis. This randomized, double-blind, placebo-controlled phase II trial explored efficacy, pharmacodynamics and safety of topical omiganan once daily in 36 patients with mild to moderate AD. Patients were randomized to apply topical omiganan 1%, omiganan 2.5% or vehicle gel to one target lesion once daily for 28 consecutive days. Small but significant improvements of the local oSCORAD index and morning itch were observed in the omiganan 2.5% group compared to the vehicle gel group (-18.5%; 95%CI=-32.9,-1.0; p=0.04 and -8.2; 95%CI=-16.3,-0.2; p=0.05 respectively). A shift from lesional to non-lesional skin microbiota was observed in both omiganan treatment groups, in contrast to the vehicle group. In conclusion, treatment with topical omiganan improved dysbiosis in patients with mild to moderate atopic dermatitis and small but statistically significant improvement of clinical scores were detected. Our findings warrant further exploration in future clinical trials.