Envelope proteome changes driven by RamA overproduction in Klebsiella pneumoniae that enhance acquired β-lactam resistance

Juan-Carlos Jiménez-Castellanos, Wan Nur Ismah Wan Ahmad Kamil, Yuiko Takebayashi, Jacqueline Findlay, Thamarai Schneiders, Kate J Heesom, Matthew B Avison*

*Corresponding author for this work

Research output: Contribution to journalArticle (Academic Journal)

13 Citations (Scopus)
260 Downloads (Pure)

Abstract

Objectives: In Klebsiella pneumoniae, overproduction of RamA results in reduced envelope permeability and reduced antimicrobial susceptibility but clinically relevant resistance is rarely observed. Here we have tested whether RamA overproduction can enhance acquired β-lactam resistance mechanisms in K. pneumoniae and have defined the envelope protein abundance changes upon RamA overproduction during growth in low and high osmolarity media. Methods: Envelope permeability was estimated using a fluorescent dye accumulation assay. β-Lactam susceptibility was measured using disc testing. Total envelope protein production was quantified using LC-MS/MS proteomics and transcript levels were quantified using real-time RT-PCR. Results: RamA overproduction enhanced β-lactamase-mediated β-lactam resistance, in some cases dramatically, without altering β-lactamase production. It increased production of efflux pumps and decreased OmpK35 porin production, though micF overexpression showed that OmpK35 reduction has little impact on envelope permeability. A survey of K. pneumoniae bloodstream isolates revealed ramA hyperexpression in 3 of 4 carbapenemase producers, 1 of 21 CTX-M producers and 2 of 19 strains not carrying CTX-M or carbapenemases. Conclusions: Whilst RamA is not a key mediator of antibiotic resistance in K. pneumoniae on its own, it is potentially important for enhancing the spectrum of acquired β-lactamase-mediated β-lactam resistance. LC-MS/MS proteomics analysis has revealed that this enhancement is achieved predominantly through activation of efflux pump production.

Original languageEnglish
Article numberdkx345
Pages (from-to)88-94
Number of pages7
JournalJournal of Antimicrobial Chemotherapy
Volume73
Issue number1
Early online date6 Oct 2017
DOIs
Publication statusPublished - 1 Jan 2018

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