Penicillin-binding protein 3 sequence variations reduce susceptibility of Pseudomonas aeruginosa to β-lactams but inhibit cell division

Abstract

Abstract Background β-lactam antibiotics, which inhibit penicillin-binding protein 3 (PBP3) that is required for cell division, play a key role in treating P. aeruginosa infections. Some sequence variations in PBP3 have been associated with β-lactam resistance but the effects of variations on antibiotic susceptibility and on cell division have not been quantified. Antibiotic efflux can also reduce susceptibility. Objectives To quantify the effects of PBP3 variations on β-lactam susceptibility and cell morphology in P. aeruginosa. Methods Nineteen PBP3 variants were expressed from a plasmid in the reference strain P. aeruginosa PAO1 and genome engineering was used to construct five mutants expressing PBP3 variants from the chromosome. The effects of the variations on β-lactam minimum inhibitory concentration (MIC) and cell morphology were measured. Results Some PBP3 variations reduced susceptibility to a variety of β-lactam antibiotics including meropenem, ceftazidime, cefepime and ticarcillin with different variations affecting different antibiotics. None of the tested variations reduced susceptibility to imipenem or piperacillin. Antibiotic susceptibility was further reduced when PBP3 variants were expressed in mutant bacteria overexpressing the MexAB-OprM efflux pump, with some variations conferring clinical levels of resistance. Some PBP3 variations, and sub-MIC levels of β-lactams, reduced bacterial growth rates and inhibited cell division, causing elongated cells. Conclusions PBP3 variations in P. aeruginosa can increase the MIC of multiple β-lactam antibiotics, although not imipenem or piperacillin. PBP3 variations, or the presence of sub-lethal levels of β-lactams, result in elongated cells indicating that variations reduce the activity of PBP3 and may reduce bacterial fitness.