High-density transposon libraries utilising outward-oriented promoters identify mechanisms of action and resistance to antimicrobials. Issue 22 (13th November 2020)

• Record Type: Journal Article
• Authors: Coward, Chris; Dharmalingham, Gopujara; Abdulle, Omar; Avis, Tim; Beisken, Stephan; Breidenstein, Elena; Carli, Natasha; Figueiredo, Luis; Jones, David; Khan, Nawaz; Malara, Sara; Martins, Joana; Nagalingam, Nabeetha; Turner, Keith; Wain, John; Williams, David; Powell, David; Mason, Clive
• Is Part Of: FEMS microbiology letters. Volume 367:Issue 22(2020)
• Publication Date: 2020-11-13
• Subjects: bacteria -- antibiotic -- development -- resistance -- mechanism -- mutagenesis; Microbiology -- Periodicals; 579
• DOI: 10.1093/femsle/fnaa185
• Languages: English
• ISSNs: 0378-1097

Abstract

ABSTRACT: The use of bacterial transposon mutant libraries in phenotypic screens is a well-established technique for determining which genes are essential or advantageous for growth in conditions of interest. Standard, inactivating, transposon libraries cannot give direct information about genes whose over-expression gives a selective advantage. We report the development of a system wherein outward-oriented promoters are included in mini-transposons, generation of transposon mutant libraries in Escherichia coli and Pseudomonas aeruginosa and their use to probe genes important for growth under selection with the antimicrobial fosfomycin, and a recently-developed leucyl-tRNA synthase inhibitor. In addition to the identification of known mechanisms of action and resistance, we identify the carbon–phosphorous lyase complex as a potential resistance liability for fosfomycin in E. coli and P. aeruginosa . The use of this technology can facilitate the development of novel mechanism-of-action antimicrobials that are urgently required to combat the increasing threat worldwide from antimicrobial-resistant pathogenic bacteria. Abstract : The authors develop a mutagenesis procedure for use in the identification of antimicrobial mechanisms of action and resistance and identify a potential resistance mechanism for fosfomycin.