A cross section of a human population (501 individuals) selected at random, and living in a Bolivian community, highly endemic for Chagas disease, was investigated combining together clinical, parasitological and molecular approaches. Conventional serology and polymerase chain reaction (PCR) indicated an active transmission of the infection, a high seroprevalence (43.3%) ranging from around 12% in < 5 years to 94.7% in > 45 years, and a high sensitivity (83.8%) and specificity of PCR. Abnormal ECG tracing was predominant in chagasic patients and was already present among individuals younger than 13 years. SAPA (shed acute phase antigen) recombinant protein and the synthetic peptide R-13 were used as antigens in ELISA tests. The reactivity of SAPA was strongly associated to Trypanosoma cruzi infection and independent of the age of the patients but was not suitable neither for universal serodiagnosis nor for discrimination of specific phases of Chagas infection. Anti-R-13 response was observed in 27.5% only in chagasic patients. Moreover, anti-R13 reactivity was associated with early infection and not to cardiac pathology. This result questioned previous studies, which considered the anti-R-13 response as a marker of chronic Chagas heart disease. The major clonets 20 and 39 (belonging to Trypanosoma cruzi I and T. cruzi II respectively) which circulate in equal proportions in vectors of the studied area, were identified in patients’ blood by PCR. Clonet 39 was selected over clonet 20 in the circulation whatever the age of the patient. The only factor related to strain detected in patients’ blood, was the anti-R-13 reactivity: 37% of the patients infected by clonet 39 (94 cases) had anti-R13 antibodies contrasting with only 6% of the patients without clonet 39 (16 cases).