Over-Expression of Hypochlorite Inducible Major Facilitator Superfamily (MFS) Pumps Reduces Antimicrobial Drug Susceptibility by Increasing the Production of MexXY Mediated by ArmZ in Pseudomonas aeruginosa

Punyawee Dulyayangkul, Aim Satapoomin, Matthew B Avison*, Nisanart Charoenlap, Paiboon Vattanaviboon, Skorn Mongkolsuk*

*Corresponding author for this work

Research output: Contribution to journalArticle (Academic Journal)peer-review

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Abstract

Pseudomonas aeruginosa, a well-known cause of nosocomial infection, is frequently antibiotic resistant and this complicates treatment. Links between oxidative stress responses inducing antibiotic resistance through over-production of RND-type efflux pumps have been reported in P. aeruginosa, but this has not previously been associated with MFS-type efflux pumps. Two MFS efflux pumps encoded by mfs1 and mfs2 were selected for study because they were found to be sodium hypochlorite (NaOCl) inducible. Antibiotic susceptibility testing was used to define the importance of these MFS pumps in antibiotic resistance and proteomics was used to characterize the resistance mechanisms involved. The results revealed that mfs1 is NaOCl inducible whereas mfs2 is NaOCl, N-Ethylmaleimide and t-butyl hydroperoxide inducible. Deletion of mfs1 or mfs2 did not affect antibiotic or paraquat susceptibility. However, over-production of Mfs1 and Mfs2 reduced susceptibility to aminoglycosides, quinolones, and paraquat. Proteomics, gene expression analysis and targeted mutagenesis showed that over-production of the MexXY RND-type efflux pump in a manner dependent upon armZ, but not amgRS, is the cause of reduced antibiotic susceptibility upon over-production of Mfs1 and Mfs2. mexXY operon expression analysis in strains carrying various lengths of mfs1 and mfs2 revealed that at least three transmembrane domains are necessary for mexXY over-expression and decreased antibiotic susceptibility. Over-expression of the MFS-type efflux pump gene tetA(C) did not give the same effect. Changes in paraquat susceptibility were independent of mexXY and armZ suggesting that it is a substrate of Mfs1 and Mfs2. Altogether, this is the first evidence of cascade effects where the over-production of an MFS pump causes over-production of an RND pump, in this case MexXY via increased armZ expression.

Original languageEnglish
Article number592153
Number of pages12
JournalFrontiers in Microbiology
Volume11
DOIs
Publication statusPublished - 12 Jan 2021

Bibliographical note

Funding Information:
We thank Wirongrong Whangsuk from the Laboratory of Biotechnology, Chulabhorn Research Institute for technical assistance and Dr. Kate Heesom from the Proteomics Facility, University of Bristol for performing the mass spectrometry analysis. Funding. PD was supported by the Royal Golden Jubilee (RGJ) Ph.D. Program (grant numbers PHD/0036/2558) and Chulabhorn Graduate Scholarship (CGS) Commemorating the 84th Birthday Anniversary of His Majesty King Bhumibol Adulyadej the Great. This work was funded by grant MR/S004769/1 from the Antimicrobial Resistance Cross Council Initiative supported by the seven United Kingdom research councils and the National Institute for Health Research to SM and MA, and grants from Chulabhorn Research Institute to SM.

Funding Information:
PD was supported by the Royal Golden Jubilee (RGJ) Ph.D. Program (grant numbers PHD/0036/2558) and Chulabhorn Graduate Scholarship (CGS) Commemorating the 84th Birthday Anniversary of His Majesty King Bhumibol Adulyadej the Great. This work was funded by grant MR/S004769/1 from the Antimicrobial Resistance Cross Council Initiative supported by the seven United Kingdom research councils and the National Institute for Health Research to SM and MA, and grants from Chulabhorn Research Institute to SM.

Publisher Copyright:
© Copyright © 2021 Dulyayangkul, Satapoomin, Avison, Charoenlap, Vattanaviboon and Mongkolsuk.

Keywords

  • Pseudomonas aeruginosa
  • MFS
  • efflux pump
  • paraquat
  • ethidium bromide
  • MexXY
  • ArmZ

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