Fluctuations in copy number of the class A carbapenemase gene blaFRI appear to confer high-level carbapenem resistance

OBJECTIVES: French imipenemase (FRI) is a carbapenemase that confer a wide range of MICs of carbapenems in blaFRI-harbouring isolates. In some isolates, duplication of the blaFRI-containing region is reported with high carbapenem MICs. A clinical Enterobacter asburiae isolate harbouring blaFRI-9 on plasmid showed high-level carbapenem resistance. This study aimed to prove that this high-level carbapenem resistance is attributed to antimicrobial-induced duplication of blaFRI-9. METHODS: Carbapenem-resistant and carbapenem-susceptible revertants derived from the clinical isolate were selected by culturing with and without antimicrobials, respectively. WGS was performed to identify the genetic background of those strains. The number of repeats of the 35 422-bp region surrounding blaFRI-9, designated as the FRI-9-long repeat region (FRI9-LR), was estimated by the ratio of the average sequence depth of FRI9-LR to that of other regions excluding FRI9-LR. RESULTS: Revertants obtained by culturing with and without antimicrobials, resulting in strains with varying carbapenem MICs. WGS revealed correlation between the number of repeats of FRI9-LR and the carbapenem MICs. Exposing the carbapenem-susceptible revertant to ampicillin did not increase the number of FRI9-LR repeats and only minimally increased the carbapenem MICs, possibly due to overexpression of AmpC β-lactamase. However, exposure to meropenem after ampicillin significantly increased both the number of FRI9-LR repeats and the carbapenem MIC, resulting in multidrug resistance. CONCLUSIONS: The high-level carbapenem resistance in this clinical isolate was attributed to multiple duplications of FRI9-LR. Furthermore, the flexibility in the duplication of this region resulted in changes in antimicrobial susceptibility, without loss or acquisition of resistance genes.