AmpC induction by imipenem in Pseudomonas aeruginosa occurs in the absence of OprD and impacts imipenem/relebactam susceptibility

Abstract

In the United States, carbapenem resistance in Pseudomonas aeruginosa is linked to the regulation of chromosomal resistance determinants, AmpC and OprD. The β-lactamase AmpC requires overexpression and genetic modifications to be capable of inhibiting imipenem activity. The outer membrane porin OprD can be downregulated or undergo genetic modifications that strongly correlate with imipenem non-susceptibility. Co-administration of imipenem and the β-lactamase inhibitor, relebactam, can lower imipenem minimal inhibitory concentrations and restore susceptibility. However, it is not understood how this occurs in P. aeruginosa isolates that do not overproduce AmpC or produce a functional OprD for imipenem entry. Therefore, we investigated whether imipenem could enter P. aeruginosa in the absence of OprD and whether any of the chromosomal β-lactamases (AmpC, OXA-50, and PIB-1) contributed to imipenem and/or imipenem/relebactam non-susceptibility. This investigation evaluated 17 imipenem non-susceptible clinical isolates and three laboratory strains of PAO1, two of which were porin transposon mutants for either oprD or opdP . Expression of OXA-50 and PIB-1 RNA was similar to PAO1. However, all 20 isolates exhibited bla ampC induction under sublethal exposure to imipenem. This occurred in the absence of detectable OprD protein in 18 isolates. Collectively, our data identify that (i) OprD was not the only channel required for imipenem entry and (ii) imipenem susceptibility can be restored by imipenem/relebactam due to the interaction between relebactam and bla ampC overexpression due to imipenem induction. IMPORTANCE Infections caused by Pseudomonas aeruginosa are associated with high mortality and worsened clinical outcomes. The carbapenem, imipenem, has been combined with the β-lactamase inhibitor relebactam to treat carbapenem-resistant P. aeruginosa . Downregulation of the outer membrane porin, OprD is the major mechanism of imipenem resistance; however, relebactam inhibits the chromosomally encoded AmpC β-lactamase. We investigated how relebactam was able to reduce P. aeruginosa imipenem minimal inhibitory concentrations (MICs) in isolates in which OprD was downregulated. Our data show that imipenem is capable of entering the cell in the absence of OprD and capable of inducing the AmpC β-lactamase. The induction of AmpC provides a substrate for relebactam, impacting the imipenem MIC. The data presented support the use of an alternative porin(s) for entry of imipenem. This study provides the basis for further investigation into modifications of imipenem or similar molecules that would increase the affinity for other porins in isolates resistant to imipenem.