Dataset Information

A Transposon-Associated CRISPR/Cas9 System Specifically Eliminates both Chromosomal and Plasmid-Borne mcr-1 in Escherichia coli.

ABSTRACT: The global spread of antimicrobial-resistant bacteria has been one of the most severe threats to public health. The emergence of the mcr-1 gene has posed a considerable threat to antimicrobial medication since it deactivates one last-resort antibiotic, colistin. There have been reports regarding the mobilization of the mcr-1 gene facilitated by ISApl1-formed transposon Tn6330 and mediated rapid dispersion among Enterobacteriaceae species. Here, we developed a CRISPR/Cas9 system flanked by ISApl1 in a suicide plasmid capable of exerting sequence-specific curing against the mcr-1-bearing plasmid and killing the strain with chromosome-borne mcr-1. The constructed ISApl1-carried CRISPR/Cas9 system either restored sensitivity to colistin in strains with plasmid-borne mcr-1 or directly eradicated the bacteria harboring chromosome-borne mcr-1 by introducing an exogenous CRISPR/Cas9 targeting the mcr-1 gene. This method is highly efficient in removing the mcr-1 gene from Escherichia coli, thereby resensitizing these strains to colistin. The further results demonstrated that it conferred the recipient bacteria with immunity against the acquisition of the exogenous mcr-1 containing the plasmid. The data from the current study highlighted the potential of the transposon-associated CRISPR/Cas9 system to serve as a therapeutic approach to control the dissemination of mcr-1 resistance among clinical pathogens.

SUBMITTER: He YZ

PROVIDER: S-EPMC8448152 | biostudies-literature | 2021 Sep

REPOSITORIES: biostudies-literature

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Publications

A Transposon-Associated CRISPR/Cas9 System Specifically Eliminates both Chromosomal and Plasmid-Borne mcr-1 in Escherichia coli.

He Yu-Zhang YZ Yan Jin-Ru JR He Bing B Ren Hao H Kuang Xu X Long Teng-Fei TF Chen Cai-Ping CP Liao Xiao-Ping XP Liu Ya-Hong YH Sun Jian J

Antimicrobial agents and chemotherapy 20210802 10

The global spread of antimicrobial-resistant bacteria has been one of the most severe threats to public health. The emergence of the mcr-1 gene has posed a considerable threat to antimicrobial medication since it deactivates one last-resort antibiotic, colistin. There have been reports regarding the mobilization of the mcr-1 gene facilitated by ISApl1-formed transposon Tn6330 and mediated rapid dispersion among Enterobacteriaceae species. Here, we developed a C ...[more]

PMID: 34339270

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Project description:Many Candida species that cause infection have diploid genomes and do not undergo classical meiosis. The application of clustered regularly interspaced short palindromic repeat-Cas9 (CRISPR-Cas9) gene editing systems has therefore greatly facilitated the generation of gene disruptions and the introduction of specific polymorphisms. However, CRISPR methods are not yet available for all Candida species. We describe here an adaption of a previously developed CRISPR system in Candida parapsilosis that uses an autonomously replicating plasmid. Guide RNAs can be introduced in a single cloning step and are released by cleavage between a tRNA and a ribozyme. The plasmid also contains CAS9 and a selectable nourseothricin SAT1 marker. It can be used for markerless editing in C. parapsilosis, C. orthopsilosis, and C. metapsilosis We also show that CRISPR can easily be used to introduce molecular barcodes and to reintroduce wild-type sequences into edited strains. Heterozygous mutations can be generated, either by careful selection of the distance between the polymorphism and the Cas9 cut site or by providing two different repair templates at the same time. In addition, we have constructed a different autonomously replicating plasmid for CRISPR-Cas9 editing in Candida tropicalis We show that editing can easily be carried out in multiple C. tropicalis isolates. Nonhomologous end joining (NHEJ) repair occurs at a high level in C. metapsilosis and C. tropicalis IMPORTANCE Candida species are a major cause of infection worldwide. The species associated with infection vary with geographical location and with patient population. Infection with Candida tropicalis is particularly common in South America and Asia, and Candida parapsilosis infections are more common in the very young. Molecular methods for manipulating the genomes of these species are still lacking. We describe a simple and efficient CRISPR-based gene editing system that can be applied in the C. parapsilosis species group, including the sister species Candida orthopsilosis and Candida metapsilosis We have also constructed a separate system for gene editing in C. tropicalis.

Dataset Information

A Transposon-Associated CRISPR/Cas9 System Specifically Eliminates both Chromosomal and Plasmid-Borne mcr-1 in Escherichia coli.

Publications

A Transposon-Associated CRISPR/Cas9 System Specifically Eliminates both Chromosomal and Plasmid-Borne <i>mcr-1</i> in Escherichia coli.

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