Background: Glucantime is regarded as the first-line treatment of cutaneous leishmaniasis (CL); however, failure to treatment is a problem in many cases. Aim: The aim was to evaluate the therapeutic effect of glucantime in CL complicated with secondary bacterial infection compared to uncomplicated lesions. Methods: This experimental study was performed in Skin Diseases and Leishmaniasis Research Center, Isfahan, Iran. A total of 161 patients enrolled in the study had CL confirmed by positive smear of lesions. All the patients were treated with systemic glucantime for 3 weeks and followed for 2 months. Response to treatment was defined as loss of infiltration, reepithelization, and negative smear. Depending on the results of bacterial cultures, the lesions were divided into two groups and the efficacy of glucantime was compared. Results: A total of 123 patients (76.4%) were negative, and 38 patients (23.6%) were positive for secondary bacterial infection. In groups with negative bacterial culture response to treatment was 65% (80 patients) and in the other positive group, it was 31.6% (12 patients), with a difference (χ2 = 13.77, P < 0.01). Conclusion: Therapeutic effect of glucantime showed a decrease in CL lesions with secondary bacterial infection. Therefore, in the cases of unresponsiveness to treatment, the lesions should be evaluated for bacterial infection, before repeating the treatment.