Role of ornibactin biosynthesis in the virulence of Burkholderia cepacia: Characterization of pvdA, the gene encoding L-ornithine N-5-oxygenase

Citation
Pa. Sokol et al., Role of ornibactin biosynthesis in the virulence of Burkholderia cepacia: Characterization of pvdA, the gene encoding L-ornithine N-5-oxygenase, INFEC IMMUN, 67(9), 1999, pp. 4443-4455
Citations number
68
Language
INGLESE
art.tipo
Article
Categorie Soggetti
Immunology
Journal title
INFECTION AND IMMUNITY
ISSN journal
0019-9567 → ACNP
Volume
67
Issue
9
Year of publication
1999
Pages
4443 - 4455
Database
ISI
SICI code
0019-9567(199909)67:9<4443:ROOBIT>2.0.ZU;2-4
Abstract
Burkholderia cepacia is a frequent cause of respiratory infections in cysti c fibrosis patients. B, cepacia has been shown to produce at least four sid erophores which may play a role in the virulence of this organism. To chara cterize genes involved in the synthesis of siderophores, Tn5-OT182 mutants were isolated in strain K56-2, which produces two siderophores, salicylic a cid (SA) and ornibactins. Two mutants were characterized that did not produ ce zones on Chrome Azurol S agar in a commonly used assay to detect siderop hore activity, These mutants were determined to produce sevenfold more SA t han K56-2 yet did not produce detectable amounts of ornibactins. These muta nts, designated I117 and T10, had a transposon insertion in genes with sign ificant homology to pyoverdine biosynthesis genes of Pseudomonas aeruginosa , I117 contained an insertion in a pvdA homolog, the gene for the enzyme L- ornithine N-5-oxygenase, which catalyzes the hydroxylation of L-ornithine. Ornibactin synthesis in this mutant was partially restored when the precurs or L-N-5-OH-Orn was added to the culture medium. T10 contained an insertion in a pvdD homolog, which is a peptide synthetase involved in pyoverdine sy nthesis. P-Galactosidase activity was iron regulated in both I117 and T10, suggesting that the transposon was inserted downstream of an iron-regulated promoter. Tn5-OT182 contains a lacZ gene that is expressed when inserted d ownstream of an active promoter, Both I117 and T10 were deficient in uptake of iron complexed to either ornibactins or SA, suggesting that transposon insertions in ornibactin biosynthesis genes also affected other components of the iron transport mechanism. The B. cepacia pvdA homolog was approximat ely 47% identical and 59% similar to L-ornithine N-5-oxygenase from P. aeru ginosa, Three clones were identified from a K56-2 cosmid library that parti ally restored ornibactin production, SA production, and SA uptake to parent al levels but did not affect the rate of Fe-59-ornibactin uptake in I117, A chromosomal pvdA deletion mutant was constructed that had a phenotype simi lar to that of I117 except that it did not hyperproduce SA, The pvdA mutant s were less virulent than the parent strain in chronic and acute models of respiratory infection. A functional pvd4 gene appears to be required for ef fective colonization and persistence in B, cepacia lung infections.