We describe a novel insertion sequence (IS) element, IS1002, which was found to be closely related to IS481, which is found only in Bordetella pertussis; we found that these two IS elements have a level of sequence identity of 61.5% and also have almost identical terminal inverted repeats. IS1002 was present in both B. pertussis and Bordetella parapertussis strains isolated from humans. In contrast, IS1002 was absent from B. parapertussis strains isolated from sheep. A DNA fingerprint analysis performed with another IS element, IS1001, which is present in B. parapertussis and Bordetella bronchiseptica, revealed that B. parapertussis isolates obtained from sheep are distinct from human isolates. Thus, human and ovine B. parapertussis strains comprise two distinct populations, indicating that little or no transmission occurs between sheep and humans. An IS-associated restriction fragment length polymorphism analysis revealed by B. parapertussis strains isolated from sheep are genetically more polymorphic than the human B. parapertussis population, which is genetically very homogeneous. This suggests that human B. parapertussis strains diverged from a single clone only recently. IS1001 is present in a subset of B. bronchiseptica strains that were derived mainly from pigs and rabbits, suggesting that these strains had a common ancestry. On the basis of the results of a comparison of IS1001 band patterns and IS1001 sequences, ovine and human B. parapertussis strains appear to have evolved independently from B. bronchiseptica strains and to have adapted to different hosts (sheep and humans). Once in the human host, B. Parapertussis probably acquired IS1002 from B. pertussis. In contrast to human B. parapertussis isolates, B. pertussis strains produced polymorphic IS1002-related DNA fingerprint patterns.