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Phospholipases D (PLDs) are virtually ubiquitous in eukaryotic organisms; however, they are relatively uncommon in prokaryotes. In this report, we demonstrate that the environmentally acquired, opportunistic pathogen Pseudomonas aeruginosa expresses PLD activity. A gene designated pldA was identified in the genomic database of P. aeruginosa PAO1 encoding a protein with significant homology to eukaryotic PLDs, but not to any prokaryotic PLDs. PldA is most homologous to PLDs from mammals and yeast. The pldA gene was cloned and shown to express an [almost equal to] 116 kDa protein with calcium-regulated PLD activity that is localized to the periplasm. Interestingly, not all strains of P. aeruginosa carry pldA. When present, pldA is always linked to an open reading frame (ORF), ORF4, and a gene (vgr A1 ) encoding a protein homologous to Vgr from Escherichia coli. Vgr proteins contain regularly repeated dipeptide motifs (v aline-g lycine r epeats). In E. coli, genes encoding Vgr are associated with multicopy genetic elements designated Rhs (r earrangement h ot-s pots). P. aeruginosa PAO1 has 10 vgr homologues dispersed throughout its genome, but the copy number of these genetic elements varies considerably in different strains. Neither vgr A1 nor ORF4 is present in strains lacking pldA. Furthermore, sequences flanking vgr A1 , pldA and ORF4 in the P. aeruginosa strains examined are highly conserved, suggesting a specific site of insertion. These and other data suggest that vgr A1 , pldA and ORF4 constitute an [almost equal to] 7 kb mobile genetic element and that pldA was acquired horizontally, perhaps from a eukaryotic organism. Competition studies between a PldA knock-out mutant and the parental wild-type strain indicate that PldA contributes to the ability of P. aeruginosa PAO1 to persist in a chronic pulmonary infection model in rats.

(C) 2001 Blackwell Science Ltd.