Bacterial resistance to CRISPR-Cas antimicrobials

Ruben V. Uribe, Christin Rathmer, Leonie Johanna Jahn, Mostafa Mostafa Hashim Ellabaan, Simone S. Li, Morten Otto Alexander Sommer*

*Corresponding author for this work

Research output: Contribution to journalJournal articleResearchpeer-review

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Abstract

In the age of antibiotic resistance and precise microbiome engineering, CRISPR-Cas antimicrobials promise to have a substantial impact on the way we treat diseases in the future. However, the efficacy of these antimicrobials and their mechanisms of resistance remain to be elucidated. We systematically investigated how a target E. coli strain can escape killing by episomally-encoded CRISPR-Cas9 antimicrobials. Using Cas9 from Streptococcus pyogenes (SpCas9) we studied the killing efficiency and resistance mutation rate towards CRISPR-Cas9 antimicrobials and elucidated the underlying genetic alterations. We find that killing efficiency is not correlated with the number of cutting sites or the type of target. While the number of targets did not significantly affect efficiency of killing, it did reduce the emergence of chromosomal mutations conferring resistance. The most frequent target of resistance mutations was the plasmid-encoded SpCas9 that was inactivated by bacterial genome rearrangements involving translocation of mobile genetic elements such as insertion elements. This resistance mechanism can be overcome by re-introduction of an intact copy of SpCas9. The work presented here provides a guide to design strategies that reduce resistance and improve the activity of CRISPR-Cas antimicrobials.
Original languageEnglish
Article number17267
JournalScientific Reports
Volume11
Issue number1
Number of pages9
ISSN2045-2322
DOIs
Publication statusPublished - 2021

Keywords

  • Anti-Infective Agents
  • CRISPR-Cas Systems
  • Drug Resistance, Bacterial
  • Escherichia coli
  • Gene Editing
  • Genome, Bacterial
  • Microbial Viability
  • Mutation
  • Plasmids
  • Streptococcus pyogenes
  • Whole Genome Sequencing

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