CRISPR-Cas3 and type I restriction-modification team up against bla-IncF plasmid transfer in Klebsiella pneumoniae.

Objective: We explored whether the Clustered regularly interspaced short palindromic repeat (CRISPR)-Cas and restriction-modification (R-M) systems are compatible and act together to resist plasmid attacks.

Methods: 932 global whole-genome sequences from GenBank, and 459 K. pneumoniae isolates from six provinces of China, were collected to investigate the co-distribution of CRISPR-Cas, R-M systems, and bla plasmid. Conjugation and transformation assays were applied to explore the anti-plasmid function of CRISPR and R-M systems.

Results: We found a significant inverse correlation between the presence of CRISPR and R-M systems and bla plasmids in K. pneumoniae, especially when both systems cohabited in one host. The multiple matched recognition sequences of both systems in bla-IncF plasmids (97%) revealed that they were good targets for both systems. Furthermore, the results of conjugation assay demonstrated that CRISPR-Cas and R-M systems in K. pneumoniae could effectively hinder bla plasmid invasion. Notably, CRISPR-Cas and R-M worked together to confer a 4-log reduction in the acquisition of bla plasmid in conjugative events, exhibiting robust synergistic anti-plasmid immunity.

Conclusions: Our results indicate the synergistic role of CRISPR and R-M in regulating horizontal gene transfer in K. pneumoniae and rationalize the development of antimicrobial strategies that capitalize on the immunocompromised status of KPC-KP.