We evaluated the diagnostic performance of a genomic DNA amplification method for Bordetella pertussis and Bordetella parapertussis compared with culture isolation. Aliquots from B. pertussis and B. parapertussis cultures were added to sterile physiological saline or sterile distilled water to give bacterial suspensions of 108 cells/ml and serial dilutions were prepared. Suspensions in physiological saline were cultured on charcoal agar medium; bacterial growth was observed up to dilutions of 10–7. Suspensions in distilled water were subjected to DNA extraction and nested polymerase chain reaction (PCR) was performed on the extracts; the PCR was positive up to dilutions of 10–8 for B. pertussis and 10–9 for B. parapertussis. Since the efficacy of culture isolation, regarded as the standard for the detection of B. pertussis and B. parapertussis, declines after the first stage of pertussis or with prior vaccination or antibiotic therapy, PCR, although not yet standardized, may provide an alternative diagnostic tool.