Polymerase chain reaction (PCR) assays that amplify segments of a repeated gene element and a toxin promoter gene of Bordetella pertussis were compared with a culture established for the diagnosis of pertussis. Of 44 nasopharyngeal (NP) aspirates collected during a pertussis outbreak, repeated gene element PCR showed a positive result in 21 (48%), including all three patients with positive culture results. Results of toxin promoter gene PCR were positive in eight (18%) cases, and the pathogen was not detected in one patient with a positive culture result. A more sensitive nested PCR assay, based on repeated gene element PCR, was then developed. During a second outbreak two different transportation systems were tested in 146 duplicate NP swabs. Transportation of swabs in empty tubes proved to be better than in transport media for PCR. A total of 190 NP specimens from the two outbreaks were tested, and in 56 the results were shown to be positive by PCR, including all 16 cases confirmed as positive by culture. We conclude that the PCR assay is more sensitive than culture in the diagnosis of pertussis; NP swabbing is a simple, practical, and reliable method of collecting clinical specimens for PCR assays and cultures.