We investigated the seroprevalence of Trypanosoma cruzi infection in an indigenous Kariña population in eastern Venezuela. A total of 175 serum samples were collected in the community of Piñantal during February 2009. Interviews targeting socioeconomic and environmental factors associated with the T. cruzi transmission were also conducted. Samples were evaluated using trypomastigote excreted/secreted antigens (TESAs) in an ELISA format. TESA-ELISA positive samples were confirmed by indirect haemagglutination (HAI) (Wiener). A nonsystematic collection of vectors was also undertaken. T. cruzi seroprevalence was 7.43% according to both assays, and the mean age of infected patients was 48.61±10.40 years (range 34 to 73 years). The vector infection rate was 20.00% (2/10). T. cruzi seropositivity was associated with a history of triatomine bites, the ability to recognize the vector and poor knowledge about Chagas disease, but no associations were found with gender, house type, knowledge of how the disease is transmitted, or the presence of vectors or animals inside dwellings. To our knowledge, this is the first study of the seroprevalence of T. cruzi in an indigenous population in eastern Venezuela. All of the epidemiological variables required for the establishment of active vectorial transmission of T. cruzi were present in this community.