The urea breath test is sensitive and specific for detection of chronic infection with H. pylori. We sought to determine the sensitivity of the [14C]urea breath test for detection of acute H. pylori infection using experimentally infected rhesus monkeys. Eighteen monkeys were inoculated with H. pylori. Serial [14C]urea breath tests and cultures of gastric biopsies were performed before and up to 10 weeks after inoculation. Cultures from all 18 monkeys were positive for H. pylori at each time point. The sensitivity of the [14C]urea breath test increased systematically from 43% at two weeks after inoculation up to 93% at 10 weeks after inoculation. Quantitative cultures of H. pylori showed a tendency to decline over time following inoculation. We conclude that the [14C]urea breath test is not sensitive for detection of acute H. pylori infection in rhesus monkeys until 10 weeks after inoculation. While this may reflect a gradual increase in bacterial load that was not detected by limited sampling, our data are not consistent with this hypothesis.