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Updated: Aug 12, 2025

Author Spotlight: Advancing Antibiotic Resistance Research Using an Efflux-Deficient Bacterial Strain and a Single-Copy Gene Expression System
05:06

Author Spotlight: Advancing Antibiotic Resistance Research Using an Efflux-Deficient Bacterial Strain and a Single-Copy Gene Expression System

Published on: January 5, 2024

684

Exploiting a conjugative endogenous CRISPR-Cas3 system to tackle multidrug-resistant Klebsiella pneumoniae

Ying Zhou1, Yang Yang2, Xiaobin Li3

  • 1Department of Laboratory Medicine, Huashan Hospital, Shanghai Medical College, Fudan University, Shanghai, China; Department of Clinical Laboratory, Shanghai Pulmonary Hospital, Tongji University School of Medicine, Shanghai, China.

Ebiomedicine
|January 25, 2023
PubMed
Abstract

Insights

We utilized a native CRISPR-Cas3 system to eliminate high-risk plasmids from multidrug-resistant Klebsiella pneumoniae. This approach effectively re-sensitizes bacteria to antibiotics and prevents further plasmid spread.

Area of Science:

  • Microbiology
  • Genetics
  • Molecular Biology

Background:

  • Mobile plasmids drive the global spread of multidrug-resistant (MDR) Klebsiella pneumoniae.
  • Plasmid curing is a potential strategy to combat antimicrobial resistance (AMR).
  • This study focuses on targeting high-risk IncFII plasmids within K. pneumoniae.

Purpose of the Study:

  • To investigate the efficacy of a native CRISPR-Cas3 system for curing IncFII plasmids in K. pneumoniae.
  • To evaluate the potential of CRISPR-mediated plasmid elimination as a therapeutic strategy against MDR bacteria.

Main Methods:

  • Examined protospacers in 725 sequenced IncFII plasmids from K. pneumoniae.
  • Re-engineered a native CRISPR-Cas3 system and delivered it via conjugation.
  • Assessed plasmid curing efficiency in vitro and in vivo using a Galleria mellonella infection model.

Main Results:

  • Identified conserved protospacer regions in IncFII plasmids, targetable by the native CRISPR-Cas3 system.
  • Achieved high plasmid curing efficiency in vitro (8-log decrease) and in vivo (~100% in G. mellonella).
  • Demonstrated that the system provides immunization against IncFII plasmid invasion in susceptible hosts.

Conclusions:

  • Native CRISPR-mediated plasmid curing is applicable for re-sensitizing drug-resistant K. pneumoniae to antibiotics.
  • Utilizing native CRISPR-Cas systems is a promising approach to combat AMR in K. pneumoniae.
  • This strategy offers a novel way to control the spread of antibiotic resistance.
Keywords:
Antimicrobial resistanceCRISPR-Cas systemIncFII plasmidK. pneumoniae

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