Liquid Chromatography-Tandem Mass Spectrometry Analysis Demonstrates a Decrease in Porins and Increase in CMY-2 β-Lactamases in Escherichia coli Exposed to Increasing Concentrations of Meropenem

Dimard E. Foudraine*, Camiel N.M. Aarents, Agnes A. Wattel, Ria van Boxtel, Nikolaos Strepis, Marian T. ten Kate, Annelies Verbon, Theo M. Luider, Corné H.W. Klaassen, John Hays, Lennard J.M. Dekker, Jan Tommassen, Wil H.F. Goessens

*Corresponding author for this work

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Abstract

While Extended-Spectrum β-Lactamases (ESBL) and AmpC β-lactamases barely degrade carbapenem antibiotics, they are able to bind carbapenems and prevent them from interacting with penicillin-binding proteins, thereby inhibiting their activity. Further, it has been shown that Enterobacterales can become resistant to carbapenems when high concentrations of ESBL and AmpC β-lactamases are present in the bacterial cell in combination with a decreased influx of antibiotics (due to a decrease in porins and outer-membrane permeability). In this study, a targeted liquid chromatography-tandem mass spectrometry (LC-MS/MS) assay was developed for the detection of the Escherichia coli porins OmpC and OmpF, its chromosomal AmpC β-lactamase, and the plasmid-mediated CMY-2 β-lactamase. BlaCMY–2–like positive E. coli isolates were cultured in the presence of increasing concentrations of meropenem, and resistant mutants were analyzed using the developed LC-MS/MS assay, Western blotting, and whole genome sequencing. In five strains that became meropenem resistant, a decrease in OmpC and/or OmpF (caused by premature stop codons or gene interruptions) was the first event toward meropenem resistance. In four of these strains, an additional increase in MICs was caused by an increase in CMY-2 production, and in one strain this was most likely caused by an increase in CTX-M-15 production. The LC-MS/MS assay developed proved to be suitable for the (semi-)quantitative analysis of CMY-2-like β-lactamases and porins within 4 h. Targeted LC-MS/MS could have additional clinical value in the early detection of non-carbapenemase-producing carbapenem-resistant E. coli.

Original languageEnglish
Article number793738
JournalFrontiers in Microbiology
Volume13
DOIs
Publication statusPublished - 28 Feb 2022

Bibliographical note

Funding Information:
This work was supported by both Netherlands Enterprise Agency and the European Union Through Horizon 2020 by means of a Eurostars grant (project number 18128). In addition, this project was in part supported by a research grant from the Dutch ZonMw organization (project number 205200006).

Publisher Copyright: Copyright © 2022 Foudraine, Aarents, Wattel, van Boxtel, Strepis, ten Kate, Verbon, Luider, Klaassen, Hays, Dekker, Tommassen and Goessens.

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