An indigo-producing strain was isolated from activated sludge and identified as Comamonas sp. based on 16S rRNA analysis. It produced indigo at 26.5 mg/l with a conversion of indole to indigo of 47%. Indole at 50 mg/l plus 200 mg naphthalene/l gave 32.2 mg indigo/l with a 58% conversion. A pathway for indigo formation is proposed. This is the first study of indigo biosynthesis by Comamonas sp.