1887

Abstract

Resistance to the polycationic antibiotic polymyxin B and expression of the outer-membrane protein OprH in the opportunistic pathogen both involve the PhoP-PhoQ two-component regulatory system. The genes for this system form an operon with , , that responds to Mg starvation and PhoP levels. In this study, the Mg-regulated promoter for this operon was mapped upstream of by primer-extension experiments. An ::-Gm mutant H855 was constructed and measurement of the catechol 2,3-dioxygenase activity expressed from this transcriptional fusion provided evidence for a second, weak promoter for . Wild-type PAO1 strain H103 was found to exhibit Mg-regulated resistance to the α-helical antimicrobial cationic peptide CP28 in addition to its previously characterized resistance to polymyxin B. Resistance to this peptide was unchanged in the OprH-null mutant H855 and a PhoP-null mutant H851. In contrast, PhoQ-null mutant H854 demonstrated constitutive CP28 resistance. Northern blot analysis revealed constitutive expression of in this strain, implicating PhoP-PhoQ in the resistance of to cationic peptides. Furthermore, all three null-mutant strains demonstrated increased resistance to the aminoglycoside antibiotics streptomycin, kanamycin and amikacin. Two additional mutant strains, H895 and H896, were constructed that carried unmarked deletions in and were found to exhibit aminoglycoside susceptibility equivalent to that of the wild-type. This result provided definitive evidence that OprH is not involved in aminoglycoside resistance and that the changes in resistance in strain H855 and a previously reported mutant were due to polar effects on rather than loss of OprH expression. A role for PhoP-PhoQ in resistance to aminoglycosides is envisaged that is distinct from that in resistance to cationic peptides and polymyxin B.

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2000-10-01
2019-10-16
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