We evaluated the prevalence of, and factors associated with, Helicobacter pylori infection in 222 subjects from 3 distinct communities of native populations (Uru-Eu-Wau-Wau Indians and 2 riverine communities living on the banks of the Machado river and in Portuchuelo) living in isolation in the rainforest of Brazilian Western Amazon. The overall prevalence was 78.8% (95% CI 72.7-83.9). The prevalence was higher in the Machado river community compared with Portuchuelo (chi2 = 3.84, P = 0.05), but no significant difference was observed between the Machado river community and the Uru-Eu-Wau-Wau Indians. Logistic regression showed that residential crowding and age were factors associated with the presence of H. pylori infection. Acquisition of the bacterium started early in life and by the age of 2 years 50% of children were infected. The prevalence increased with age, reaching near universal levels during adulthood (97.9%). Residential crowding was high with a global index of 3.3 persons/room (SD = 1.8), varying significantly between the 3 communities (P = 0.001). These data provide further evidence supporting direct person-to-person spread of the bacterium.