Interplay between OXA-10 ?-Lactamase Production and Low Outer-Membrane Permeability in Carbapenem Resistance in Enterobacterales

Abstract

The OXA-10 class D ?-lactamase has been reported to contribute to carbapenem resistance in non-fermenting Gram-negative bacilli; however, its contribution to carbapenem resistance in Enterobacterales is unknown. In this work, minimum inhibitory concentrations (MICs), whole genome sequencing (WGS), cloning experiments, kinetic assays, molecular modelling studies, and biochemical assays for carbapenemase detection were performed to determine the impact of OXA-10 production on carbapenem resistance in two XDR clinical isolates of Escherichia coli with the carbapenem resistance phenotype (ertapenem resistance). WGS identified the two clinical isolates as belonging to ST57 in close genomic proximity to each other. Additionally, the presence of the blaOXA-10 gene was identified in both isolates, as well as relevant mutations in the genes coding for the OmpC and OmpF porins. Cloning of blaOXA-10 in an E. coli HB4 (OmpC and OmpF-deficient) demonstrated the important contribution of OXA-10 to increased carbapenem MICs when associated with porin deficiency. Kinetic analysis showed that OXA-10 has low carbapenem-hydrolysing activity, but molecular models revealed interactions of this ?-lactamase with the carbapenems. OXA-10 was not detected with biochemical tests used in clinical laboratories. In conclusion, the ?-lactamase OXA-10 limits the activity of carbapenems in Enterobacterales when combined with low permeability and should be monitored in the future.