Cutaneous leishmaniasis features are dependent on the patient immunological stage, the parasite biology and to some extent are determined by the geographic regions in which it occurs. New World cutaneous infections known as American Cutaneous Leishmaniasis (ACL) are associated with metastatic sequela and treatment failure. In the present study we found that in patients with ACL infected in the Colombian Amazon and Orinoco regions and not responding to Glucantime® treatment, the predominant species was L. braziliensis. In addition 35% of these lesion-isolated parasites show some level of drug resistance. We also found that these resistant parasites display reduced competence when infecting the human monocytic cell line U-937 in ¿in vitro¿ tests and are associated with lesions of reduced size. According to our findings the indication for a second round of treatment with pentavalent antimonials in patients with ACL that did not respond to the first round of treatment is arguable.