Identification of Bordetella pertussis in nasopharyngeal swabs by PCR amplification of a region of the adenylate cyclase gene

Abstract

The polymerase chain reaction (PCR) was used to amplify a 522-bp region of the adenylate cyclase toxin (cyaA) gene of Bordetella pertussis. As few as 100 cfu from a suspension of B. pertussis could be detected by this procedure when the amplified PCR product was detected by ethidium bromide staining of agarose gels. However, simulated clinical specimens, prepared from swabs impregnated with known numbers of B. pertussis cells, only yielded a positive reaction with > or = 10(4) cfu. Hybridisation of a Southern blot of the PCR products from the swab samples with a cya-specific probe gave a positive reaction with as few as 8 cfu, but the hybridisation signal was uniformly weak with fewer than 10(4) cfu. Nevertheless, three of 13 nasopharyngeal swabs, taken from suspected clinically defined cases of whooping cough and stored frozen for up to 18 months, gave a positive PCR reaction.