We describe two cases of duodenal leishmaniasis in patients with human immunodeficiency virus (HIV) infection, diagnosed by light and electron microscopy. The patients presented nonspecific signs and symptoms, blood cultures were sterile, and serological tests for Leishmania spp. were negative. Endoscopy showed normal-appearing mucosa in one patient and possible peptic duodenitis in the other patient. In these patients, the parasite was only detected in a duodenal biopsy specimen. In view of the unusual location of the parasite and the fact that the diagnostic and dissemination of the disease was established by means of conventional biopsy, this is not a routine procedure for the diagnosis of leishmaniasis because the classic procedures require the demonstration of antibodies and visualization in bone marrow, lymph nodes, liver and/or spleen aspirates. We decided to report these two cases to call attention to the possible finding of Leishmania amastigotes in biopsies from intestinal mucosa in HIV infected patients.