Helicobacter pylori infection status following experimental inoculation of mice presently requires euthanasia. The purpose of this study was to develop a method for following the time course of H. pylori infection in live experimental animals. Twenty-six C57BL/6, Helicobacter-free female mice were inoculated with H. pylori Sydney strain 1, and 16 mice were sham inoculated. The mice were repeatedly tested during a period of about 1 year with an H. pylori species-specific primer-based PCR analysis of DNA extracted from fecal pellets of mice. The mice were euthanized at 6 months (n = 15) and 10 months (n = 15) to determine their infection status by histology, culture, and PCR of gastric specimens. H. pylori-inoculated mice were tested via the PCR method at 6 and 10 months prior to necropsy. Nine of 13 (69%) and 10 of 13 (77%) mice tested at 6 and 10 months, respectively, were positive. All sham-inoculated mice were negative. These two PCR results suggested a specificity of 100% with a sensitivity range between 69 and 77%. In contrast, sensitivity and specificity rose to 90 and 100% if groups of mice were tested once daily for 4 days. Seventy-seven to 85% of the experimental mice were also positive for H. pylori by culture. The histopathology demonstrated mild to severe gastritis. These findings demonstrate that the persistence or transience of H. pylori infection in live mice can be repeatedly evaluated over time. This method could allow the determination of the time course of infection and the efficacy of medications and/or vaccine without necropsy.