Identification of Legionella pneumophila rcp, a pagP-like gene that confers resistance to cationic antimicrobial peptides and promotes intracellular infection

In the course of characterizing a locus involved in heme utilization, we id entified a Legionella pneumophila gene predicted to encode a protein with h omology to the product of the Salmonella enterica serovar Typhimurium pagP gene. In Salmonella, pagP increases resistance to the bactericidal effects of cationic antimicrobial peptides (CAMPs), Mutants with insertions in the L. pneumophila pagP-like gene were generated and showed decreased resistanc e to different structural classes of CAMPs compared to the wild type; hence , this gene was designated rep for resistance to cationic antimicrobial pep tides, Furthermore, Legionella CAMP resistance was induced by growth in Low -magnesium medium. To determine whether rep had any role in intracellular s urvival, mutants were tested in the two most relevant host cells for Legion naires' disease, i.e., amoebae and macrophages, These mutants exhibited a 1 ,000-fold-decreased recovery during a Hartmannella vermiformis coculture, C omplementation of the infectivity defect could be achieved by introduction of a plasmid containing the intact rep gene. Mutations in rep consistently reduced both the numbers of bacteria recovered during intracellular infecti on and their cytopathic capacity for U937 macrophages. The rep mutant was a lso more defective for lung colonization of A/J mice. Growth of rep mutants in buffered yeast extract broth was identical to that of the wild type, in dicating that the observed differences in numbers of bacteria recovered fro m host cells were not due to a generalized growth defect, However, in low-M g2+ medium, the rep mutant was impaired in stationary-phase survival. This is the first demonstration of a pagP-like gene, involved in resistance to C AMPs, being required for intracellular infection and virulence.