Background: Acne vulgaris is a chronic inflammatory disease of pilosebaceous unit. The role of Pityrosporum ovale (P.ovale) in the exacerbation of acne is still under debate. Objective: To determine the efficacy of topical 2% ketoconazole cream in comparison to placebo in the treatment of acne vulgaris in patients with positive smear for P.ovale. Patients and Methods: A smear was obtained from lesions of 84 patients with acne vulgaris referred to Razi Hospital, Tehran. After determination of clinical and mycological grading, patients with positive smear for P.ovale were randomly treated with topical 2% ketoconazole cream or placebo. After 4 weeks the patients were evaluated for the clinical and mycological grading and treated with traditional anti-acne drugs. Results: Fifty-six of 84 patients had positive smears, 29 of them were treated with ketoconazole cream and 27 patients received placebo. Clinical grading before treatment was 2.60 and 2.21 in ketoconazole and placebo groups, which was reduced to 2.16 and 2.05, respectively. Mycological grading reduced from 1.4 to 0.44 in ketoconazole group, but did not change in placebo group (1.42). Conclusion: 2% ketoconazole cream was more effective than placebo in reduction of mycological grading but did not have any effect on clinical grading of acne.