Escherichia coli F Plasmid Transfers to and Replicates within Legionella pneumophila: An Alternative to Using an RP4-Based System for Gene Delivery

Abstract

Derivatives of the self-transmissible F plasmid of Escherichia coli can be introduced into Legionella pneumophila by conjugation and maintained within only upon selection. In L. pneumophila. F-based replicons seem to exist as extrachromosomal elements since they were readily lost when F-containing L. pneumophila was grown on nonselective medium. The F-based plasmids were not self-transmissible in L. pneumophila. The mating defect may be due to an inability to form the F pilus since F-containing strains of L. pneumophila could neither be infected with the pilus-specific phage M13 nor transduced with fl-packaged ColE1 replicons. Currently, the most commonly used transfer system for introducing genetic information into L. pneumophila employs E. coli donors with a chromosomally integrated copy of RP4::Mu to mobilize plasmids bearing the RK2 origin of transfer ( oriT). Use of this system to deliver Tn phoA for mutagenesis of the L. pneumophila chromosome led to transconjugants that all contained cryptic DNA alterations that involved the plasmid RP4 and phage Mu. No Tn phoA transposition was observed in L. pneumophila. The fact that F-mediated conjugation can be used to efficiently transfer plasmids containing the oriT of F to L. pneumophila provides an important alternative to the RP4-based plasmid transfer system and may avoid DNA anomalies in transconjugants that impede genetic analysis. Furthermore, our results demonstrate the promiscuous nature of the F conjugal transfer and replication systems.