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Updated: Apr 3, 2026
In vitro Investigation of the MexAB Efflux Pump From Pseudomonas aeruginosa
Published on: February 17, 2014
Polymyxin Susceptibility in Pseudomonas aeruginosa Linked to the MexXY-OprM Multidrug Efflux System
Keith Poole1, Calvin Ho-Fung Lau2, Christie Gilmour2
1Department of Biomedical and Molecular Sciences, Queen's University, Kingston, Ontario, Canada poolek@queensu.ca.
Abstract:
The ribosome-targeting antimicrobial, spectinomycin (SPC), strongly induced the mexXY genes of the MexXY-OprM multidrug efflux system in Pseudomonas aeruginosa and increased susceptibility to the polycationic antimicrobials polymyxin B and polymyxin E, concomitant with a decrease in expression of the polymyxin resistance-promoting lipopolysaccharide (LPS) modification loci, arnBCADTEF and PA4773-74. Consistent with the SPC-promoted reduction in arn and PA4773-74 expression being linked to mexXY, expression of these LPS modification loci was moderated in a mutant constitutively expressing mexXY and enhanced in a mutant lacking the efflux genes. Still, the SPC-mediated increase in polymyxin susceptibility was retained in mutants lacking arnB and/or PA4773-74, an indication that their reduced expression in SPC-treated cells does not explain the enhanced polymyxin susceptibility. That the polymyxin susceptibility of a mutant strain lacking mexXY was unaffected by SPC exposure, however, was an indication that the unknown polymyxin resistance 'mechanism' is also influenced by the MexXY status of the cell. In agreement with SPC and MexXY influencing polymyxin susceptibility as a result of changes in the LPS target of these agents, SPC treatment yielded a decline in common polysaccharide antigen (CPA) synthesis in wild-type P. aeruginosa but not in the ΔmexXY mutant. A mutant lacking CPA still showed the SPC-mediated decline in polymyxin MICs, however, indicating that the loss of CPA did not explain the SPC-mediated MexXY-dependent increase in polymyxin susceptibility. It is possible, therefore, that some additional change in LPS promoted by SPC-induced mexXY expression impacted CPA synthesis or its incorporation into LPS and that this was responsible for the observed changes in polymyxin susceptibility.
Insights
Spectinomycin (SPC) increases polymyxin susceptibility in Pseudomonas aeruginosa by inducing mexXY genes. This effect on polymyxin resistance is MexXY-dependent and linked to changes in lipopolysaccharide (LPS) structure, not just reduced LPS modification.
Area of Science:
- Microbiology
- Molecular Biology
- Drug Resistance
Background:
- Pseudomonas aeruginosa possesses intrinsic resistance to polymyxins, mediated partly by lipopolysaccharide (LPS) modifications.
- Multidrug efflux systems, such as MexXY-OprM, play a role in bacterial defense mechanisms.
- Understanding the interplay between efflux pumps and LPS modification is crucial for combating antibiotic resistance.
Purpose of the Study:
- To investigate the mechanism by which spectinomycin (SPC) enhances polymyxin susceptibility in Pseudomonas aeruginosa.
- To determine the role of the mexXY efflux genes and LPS modification loci (arnBCADTEF, PA4773-74) in SPC-induced polymyxin susceptibility.
- To elucidate the impact of SPC and mexXY on lipopolysaccharide (LPS) structure and its contribution to polymyxin resistance.
Main Methods:
- Treatment of wild-type and mutant Pseudomonas aeruginosa strains with spectinomycin (SPC).
- Gene expression analysis of mexXY, arnBCADTEF, and PA4773-74 loci.
- Assessment of polymyxin B and polymyxin E susceptibility using minimum inhibitory concentration (MIC) assays.
- Analysis of common polysaccharide antigen (CPA) synthesis and its incorporation into LPS.
Main Results:
- Spectinomycin strongly induced mexXY gene expression and increased susceptibility to polymyxins.
- Reduced expression of LPS modification loci (arn and PA4773-74) was observed upon SPC treatment, linked to mexXY.
- Polymyxin susceptibility increase was MexXY-dependent and associated with decreased common polysaccharide antigen (CPA) synthesis, but not solely explained by reduced LPS modification or CPA loss.
Conclusions:
- The MexXY efflux system is a key mediator of spectinomycin-induced polymyxin susceptibility in Pseudomonas aeruginosa.
- While SPC-induced mexXY expression reduces polymyxin resistance-associated LPS modifications and CPA synthesis, these factors alone do not fully explain the observed susceptibility increase.
- An uncharacterized change in LPS structure, influenced by MexXY activity, likely underlies the enhanced polymyxin susceptibility.
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