Fecal excretion of hepatitis A virus (HAV) in 18 patients with HAV infection was evaluated by enzyme immunoassay (EIA) to detect viral antigen and by reverse transcription-PCR amplification followed by ethidium bromide staining (PCR-ETBr) or nucleic acid hybridization (PCR-NA) to detect viral genetic material. A gradation of sensitivity was observed in the detection of virus by the three methods. In persons who had detectable virus, serial stool samples were found to be positive by EIA for up to 24 days after the peak elevation of liver enzymes. Viral genetic material could be detected by PCR-ETBr for up to 34 days and by PCR-NA for up to 54 days after the peak elevation of liver enzymes. After intravenous inoculation of tamarins with stool suspensions categorized as highly reactive for HAV (positive by EIA, PCR-ETBr, and PCR-NA), moderately reactive (positive by PCR-ETBr and PCR-NA), or weakly reactive (positive by PCR-NA), only tamarins infected with highly reactive stool suspensions (EIA positive) developed HAV infection. We conclude that positivity of stool specimens for HAV by PCR-ETBr or PCR-NA indicates a lower potential for infectivity, compared to that of EIA-positive stools.