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To determine the prevalence of Helicobacter pylori antigen carriage in stool in the Penan ethnic minority in Malaysian Borneo, we studied 295 Penans 0.6–89.0 years of age from 1) the remote Limbang Division, 2) Mulu regional center, and 3) Belaga village. Overall, 37.7% of the subjects tested positive. Peak prevalence was reached by 10 years of age. There were no differences in age, sex, body mass index, and socioeconomic/domestic variables between antigen-positive and antigen-negative subjects. In a logistic regression analysis, subjects from Limbang were least likely to be antigen-positive (odds ratio [OR] = 0.23, 95% confidence interval [CI] = 0.12–0.44 versus other sites, P < 0.001). Availability of a flushing toilet was protective against H. pylori carriage (OR = 0.51, 95% CI = 0.27–0.95, P = 0.031). Infection with H. pylori among the Penan was less than reported in other low socioeconomic groups. The lowest prevalence in the most remote setting suggests that the infection has been a recent arrival in previously isolated communities.