The seroprevalence of Helicobacter pylori among secluded Indian populations of South America was determined to gain insight into the evolutionary history and possible transmission patterns of the organism. Serum samples obtained from 1024 donors in 22 different villages were tested by enzyme-linked immunosorbent assay for immunoglobulin G antibodies, and the results were confirmed by Western blot. The overall seroprevalence was 92%: >80% of children tested positive by 3 years of age, the highest prevalence in populations studied to date. Comparison of H. pylori prevalence with that of herpes simplex virus type 1, which is known to be transmitted orally, demonstrated a linear correlation in their prevalence rates, suggesting that these pathogens share risk factors. However, H. pylori seroprevalence was consistently higher, indicating that additional routes of transmission exist and/or that the organism is more transmissible. Seroprevalence did not correlate with the length of contact with the outside world. These results suggest that H. pylori was indigenous to the South American Indians and was not introduced by contact with outsiders.