In vitro transposition system for efficient generation of random mutants of Campylobacter jejuni

Campylobacter jejuni is the most common cause of food-borne illnesses in th e United States. Despite the fact that the entire nucleotide sequence of it s genome has recently become available, its mechanisms of pathogenicity are poorly understood. This is in part due to the lack of an efficient mutagen esis system. Here we describe an in vitro transposon mutagenesis system bas ed on the Staphylococcus aureus transposable element Tn552 that allows the efficient generation of insertion mutants of C. jejuni. Insertions occur ra ndomly and throughout the entire bacterial genome. We have tested this syst em in the isolation of nonmotile mutants of C. jejuni. Demonstrating the ut ility of the system, six nonmotile mutants from a total of nine exhibited i nsertions in genes known to be associated with motility. An additional muta nt had an inactivating insertion in sigma 54, implicating this transcriptio n factor in flagellum regulation. The availability of this efficient system will greatly facilitate the study of the mechanisms of pathogenesis of thi s important pathogen.