SummaryThe nucleotide sequence of the pertussis toxin operon ofBordetella pertussis, Bordetella parapertussisandBordetella bronchiseptica, has shown that the last two species contain many common mutations and are likely to derive from a common ancestor (Arico and Rappuoli, 1987). To elucidate further the evolutionary relationships between theBordetellaspecies, we have cloned and sequenced the promoter region and the gene coding for the S1 subunit of pertussis toxin from additionalB. pertussisstrains, such as the type strain BP 18323 and two recent clinical isolates, namely strain BP 13456 from Sweden and strain BP SA1 from Italy. While the strains BP SA1 and BP 13456 are shown to differ from the publishedB. pertussissequences by only one base pair, the type strain BP 18323 contains a total of 11 base‐pair substitutions. Remarkably, 9 of the 11 substitutions found in BP 18323 are also common toB. parapertussisandB. bronchiseptica, strongly suggesting that this strain derives from the same ancestor asB. parapertussisandB. bronchiseptica.Computer analysis of the sequence data allows the construction of an evolutionary ‘tree’ showing that theB. pertussisstrains are very homogeneous and significantly distant fromB. parapertussisandB. bronchiseptica.Therefore the proposed conversion fromB. parapertussistoB. pertussisappears highly impro