It is generally held that with rare exception Leishmania braziliensis braziliensis is the parasite responsible for the metastatic development of mucocutaneous leishmaniasis in the New World. Yet the infrequency of mucocutaneous disease compared with cutaneous manifestations, and the difficulty of isolating parasites from mucocutaneous lesions have restricted the study of the organisms involved. We here report the biologic, isoenzymatic, and monoclonal antibody specificity characteristic of eight Leishmania isolates obtained from the mucosal lesions of the same number of patients. Individually and collectively, the identifying criteria implicate at least two L. braziliensis subspecies as etiologic agents of mucocutaneous leishmaniasis in Colombia and suggest that a spectrum of intrinsically distinguishable organisms may be involved in this disease form.