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International journal of molecular sciences
Jun 26, 2024;25 (13):

Screening of the PA14NR Transposon Mutant Library Identifies Genes Involved in Resistance to Bacteriophage Infection in .

Peiying Ho, Linh Chi Dam, Wei Ren Ryanna Koh, Rui Si Nai, Qian Hui Nah, Faeqa Binte Muhammad Rajaie Fizla, Chia Ching Chan, Thet Tun Aung, Shin Giek Goh, You Fang, Zhining Lim, Ming Guang Koh, Michael Demott, Yann Felix Boucher, Benoit Malleret, Karina Yew-Hoong Gin, Peter Dedon, Wilfried Moreira
Author Information
  1. Peiying Ho: Antimicrobial Resistance Interdisciplinary Research Group (AMR IRG), Singapore-MIT Alliance for Research and Technology (SMART) Centre, Singapore 117576, Singapore. ORCID
  2. Linh Chi Dam: Antimicrobial Resistance Interdisciplinary Research Group (AMR IRG), Singapore-MIT Alliance for Research and Technology (SMART) Centre, Singapore 117576, Singapore.
  3. Wei Ren Ryanna Koh: Antimicrobial Resistance Interdisciplinary Research Group (AMR IRG), Singapore-MIT Alliance for Research and Technology (SMART) Centre, Singapore 117576, Singapore.
  4. Rui Si Nai: Antimicrobial Resistance Interdisciplinary Research Group (AMR IRG), Singapore-MIT Alliance for Research and Technology (SMART) Centre, Singapore 117576, Singapore.
  5. Qian Hui Nah: Antimicrobial Resistance Interdisciplinary Research Group (AMR IRG), Singapore-MIT Alliance for Research and Technology (SMART) Centre, Singapore 117576, Singapore.
  6. Faeqa Binte Muhammad Rajaie Fizla: Antimicrobial Resistance Interdisciplinary Research Group (AMR IRG), Singapore-MIT Alliance for Research and Technology (SMART) Centre, Singapore 117576, Singapore.
  7. Chia Ching Chan: Antimicrobial Resistance Interdisciplinary Research Group (AMR IRG), Singapore-MIT Alliance for Research and Technology (SMART) Centre, Singapore 117576, Singapore.
  8. Thet Tun Aung: Immunology Translational Research Programme, Department of Microbiology and Immunology, Yong Loo Lin School of Medicine, National University of Singapore, Singapore 117597, Singapore. ORCID
  9. Shin Giek Goh: Energy & Environmental Sustainability Solutions for Megacities (E2S2) Program, Campus for Research Excellence and Technological Enterprise (CREATE), Singapore 117576, Singapore.
  10. You Fang: Energy & Environmental Sustainability Solutions for Megacities (E2S2) Program, Campus for Research Excellence and Technological Enterprise (CREATE), Singapore 117576, Singapore. ORCID
  11. Zhining Lim: Immunology Translational Research Programme, Department of Microbiology and Immunology, Yong Loo Lin School of Medicine, National University of Singapore, Singapore 117597, Singapore.
  12. Ming Guang Koh: Immunology Translational Research Programme, Department of Microbiology and Immunology, Yong Loo Lin School of Medicine, National University of Singapore, Singapore 117597, Singapore.
  13. Michael Demott: Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA 02139, USA.
  14. Yann Felix Boucher: Singapore Centre for Environmental Life Science Engineering (SCELSE), Singapore 637551, Singapore. ORCID
  15. Benoit Malleret: Immunology Translational Research Programme, Department of Microbiology and Immunology, Yong Loo Lin School of Medicine, National University of Singapore, Singapore 117597, Singapore. ORCID
  16. Karina Yew-Hoong Gin: Energy & Environmental Sustainability Solutions for Megacities (E2S2) Program, Campus for Research Excellence and Technological Enterprise (CREATE), Singapore 117576, Singapore. ORCID
  17. Peter Dedon: Antimicrobial Resistance Interdisciplinary Research Group (AMR IRG), Singapore-MIT Alliance for Research and Technology (SMART) Centre, Singapore 117576, Singapore. ORCID
  18. Wilfried Moreira: Antimicrobial Resistance Interdisciplinary Research Group (AMR IRG), Singapore-MIT Alliance for Research and Technology (SMART) Centre, Singapore 117576, Singapore. ORCID
PMID: 39000118 DOI: 10.3390/ijms25137009

Abstract

Multidrug-resistant infections pose a serious public health threat due to the rise in antimicrobial resistance. Phage therapy has emerged as a promising alternative. However, has evolved various mechanisms to thwart phage attacks, making it crucial to decipher these resistance mechanisms to develop effective therapeutic strategies. In this study, we conducted a forward-genetic screen of the PA14 non-redundant transposon library (PA14NR) to identify dominant-negative mutants displaying phage-resistant phenotypes. Our screening process revealed 78 mutants capable of thriving in the presence of phages, with 23 of them carrying insertions in genes associated with membrane composition. Six mutants exhibited total resistance to phage infection. Transposon insertions were found in genes known to be linked to phage-resistance such as and a glycosyl transferase gene, as well as novel genes such as , , and two hypothetical proteins. Functional experiments demonstrated that these genes played pivotal roles in phage adsorption and biofilm formation, indicating that altering the bacterial membrane composition commonly leads to phage resistance in . Importantly, these mutants displayed phenotypic trade-offs, as their resistance to phages inversely affected antibiotic resistance and hindered biofilm formation, shedding light on the complex interplay between phage susceptibility and bacterial fitness. This study highlights the potential of transposon mutant libraries and forward-genetic screens in identifying key genes involved in phage-host interactions and resistance mechanisms. These findings support the development of innovative strategies for combating antibiotic-resistant pathogens.

Keywords

Pseudomonas aeruginosa bacteriophage bacteriophage resistance mechanisms transposon mutagenesis

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Grants

  1. NRF2018-ITS003-021/National Research Foundation
  2. NUHSRO/2018/006/SU/01/National University Health System
  3. NUHSRO/2019/046/PDF/19/National University Health System
  4. OFIRG21jun-0038/National Medical Research Council

MeSH Term

Pseudomonas aeruginosa
DNA Transposable Elements
Mutation
Gene Library
Biofilms
Bacteriophages

Chemicals

DNA Transposable Elements
Journal Article

Word Cloud

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