Aminoglycoside resistance in Pseudomonas aeruginosa: the contribution of the MexXY-OprM efflux pump varies between isolates

Introduction. Aminoglycoside antibiotics are widely used to treat infections of Pseudomonas aeruginosa. The MexXY-OprM efflux pump is an important contributor to aminoglycoside tolerance in P aeruginosa reference strains and expression of the mexXY genes is repressed by the MexZ repressor protein. Direct investigation of the role of this efflux pump in clinical isolates is relatively limited. Hypothesis. The contribution of MexXY-OprM to P. aeruginosa aminoglycoside resistance is isolate-specific. Aim. To quantify the role of MexXY-OprM and its repressor, MexZ, in clinical isolates of P. aeruginosa. Methodology. The mexXY genes were deleted from ten clinical isolates of P. aeruginosa. and the mexZ gene from nine isolates. Antimicrobial susceptibility testing was carried out for commonly used antipseudomonal drugs on the engineered mutants and the isogenic wild-type isolates. RT-qPCR was used to measure expression of the mexX gene. Results. All but one of the mexXY mutants were more susceptible to the clinically used aminoglycosides tobramycin, gentamicin and amikacin but the degree to which susceptibility increased varied greatly between isolates. The mexXY mutants were also more susceptible to a fluoroquinolone, ciprofloxacin. In three isolates with functional MexZ, deletion of mexZ increased expression of mexXY and aminoglycoside tolerance. Conversely, deleting mexZ from six clinical isolates with mexZ sequence variants had little or no effect on expression of mexXY or on aminoglycoside susceptibility, consistent with the variants abolishing MexZ function. Genome analysis showed that over 50% of 619 clinical isolates had sequence variants predicted to reduce the affinity of MexZ for DNA, likely increasing mexXY expression and hence efflux of aminoglycosides. Conclusion. Our findings show that the interplay between MexXY, MexZ and the level of mexXY expression plays an important role in aminoglycoside resistance in clinical isolates of P. aeruginosa but the magnitude of the contribution of this efflux pump to resistance is isolate-specific.

Automated summary

The MexXY-OprM efflux pump plays an isolate-specific role in aminoglycoside resistance in clinical isolates of P. aeruginosa. The interplay between MexXY, MexZ, and the level of mexXY expression is important in determining the resistance level. The affinity of MexZ for DNA is a key factor in regulating mexXY expression and aminoglycoside efflux.