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Epidemiologic studies were conducted during the period 1986–1988 in a small rural community in Colombia (El Callejon) where visceral leishmaniasis is highly endemic. In this community of 185 people, 14 cases of infantile visceral leishmaniasis were diagnosed in the 9 years 1981–1988. Leishmanin skin testing of a sample of the human residents showed that prevalence of Leishmania chagasi infection increased with age; overall, 51.2% of the subjects had a positive reaction. A canine surveillance program was instituted, using introduced sentinel dogs as well as the indigenous dog population. Eleven of 16 sentinel dogs were infected within 8 months of exposure; mean seroconversion time was 4.4 months. Eleven of 25 seronegative local dogs were also infected during the 26 month period; mean seroconversion time was 8 months. Parasites identified by isozyme electrophoresis as L. chagasi were recovered from 18 of 22 seropositive dogs. Collections of wild animals using baited live traps yielded mainly the neotropical opossum, Didelphis marsupialis. Leishmania chagasi was recovered from 12 of 37 (32.4%) opossums. Six of 681 female Lutzomyia longipalpis collected in the community had flagellates in their guts; cultures from 4 were identified as L. chagasi. These data confirmed that active parasite transmission occurred. The relatively high prevalence of L. chagasi infection found among D. marsupialis captured near human dwellings suggests that these animals may be an important peridomestic reservoir.