Curing of common plasmids in gram-negative bacteria using a Cas9-based conjugative vector

Kelly K Yen; Austin J Terlecky; Mingju Hao; Vanessa Cienfuegos; Albert Rojtman; Liang Chen; Barry N Kreiswirth

doi:10.1016/j.mimet.2024.107047

Curing of common plasmids in gram-negative bacteria using a Cas9-based conjugative vector

J Microbiol Methods. 2024 Nov:226:107047. doi: 10.1016/j.mimet.2024.107047. Epub 2024 Sep 18.

Authors

Kelly K Yen¹, Austin J Terlecky², Mingju Hao³, Vanessa Cienfuegos⁴, Albert Rojtman⁵, Liang Chen⁶, Barry N Kreiswirth²

Affiliations

¹ Center for Discovery and Innovation, Hackensack Meridian Health, Nutley, NJ 07110, United States of America. Electronic address: [email protected].
² Center for Discovery and Innovation, Hackensack Meridian Health, Nutley, NJ 07110, United States of America.
³ The First Affiliated Hospital of Shandong First Medical University, Jinan, China.
⁴ School of Microbiology, University of Antioquia, Medellín, Colombia.
⁵ Jersey Shore University Medical Center, Neptune, NJ 07753, United States of America.
⁶ School of Pharmacy and Pharmaceutical Sciences, University at Buffalo, Buffalo, NY 14214, United States of America.

PMID: 39303991
DOI: 10.1016/j.mimet.2024.107047

Abstract

We report the creation of 17 Escherichia coli strains harboring the conjugative plasmid pLCasCureT with a CRISPR-Cas9 system to surgically "cure" the most common plasmids among Enterobacterales species. This approach can create isogenic pairs of strains to study host-plasmid interactions, correlate plasmid genotype and phenotype, and create plasmid-free cloning strains.

Keywords: Antibiotic resistance; CRISPR-Cas9; Gram-negatives; Plasmid curing.

MeSH terms

CRISPR-Cas Systems*
Conjugation, Genetic*
Escherichia coli* / genetics
Genetic Vectors / genetics
Plasmids* / genetics