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Prokaryotic CRISPR-Cas systems limit the acquisition of genetic elements and provide immunity against invasive bacteriophage. The characteristics of CRISPR-Cas systems in clinical  Klebsiella pneumoniae  isolates are still unknown. Here, 97  K. pneumoniae  genomes retrieved from the Integrated Microbial Genomes & Microbiomes genome database and 176 clinical isolates obtained from patients with bloodstream (BSI,  n  = 87) or urinary tract infections (UTI,  n  = 89) in Taiwan, were used for analysis. Forty out of ninety-seven genomes (41.2%) had CRISPR-Cas systems identified by the combination of CRISPRFinder and  cas1  gene sequence alignment. The phylogenetic trees revealed that CRISPR-Cas systems in  K. pneumoniae  were divided into two types (type I-E, 23; subtype I-E ∗ , 17) based on the sequences of Cas1 and Cas3 proteins and their location in the chromosome. The distribution of type I-E and I-E ∗  CRISPR-Cas systems was associated with the multilocus sequence typing and the pulsed-field gel electrophoresis results. Importantly, no CRISPR-Cas system was identified in published genomes of clonal complex 258 isolates (ST11 and ST258), which comprise the largest multi-drug resistant  K. pneumoniae  clonal group worldwide. PCR with  cas -specific primers showed that 30.7% (54/176) of the clinical isolates had a CRISPR-Cas system. Among clinical isolates, more type I-E CRISPR-Cas systems were found in UTI isolates (BSI, 5.7%; UTI, 11.2%), and subtype I-E ∗  CRISPR-Cas systems were dominant in BSI isolates (BSI, 28.7%; UTI, 15.7%) ( p  = 0.042). Isolates which had subtype I-E ∗  CRISPR-Cas system were more susceptible to ampicillin-sulbactam ( p  = 0.009), cefazolin ( p  = 0.016), cefuroxime ( p  = 0.039), and gentamicin ( p  = 0.012), compared to the CRISPR-negative isolates. The strains containing subtype I-E ∗  CRISPR-Cas systems had decreased numbers of plasmids, prophage regions, and acquired antibiotic resistance genes in their published genomes. Here, we first revealed subtype I-E ∗  CRISPR-Cas system in  K. pneumoniae  potentially interfering with the acquisition of phages and plasmids harboring antibiotic resistance determinants, and thus maintained these isolates susceptible to antibiotics.

Characterization of CRISPR-Cas Systems in Clinical Klebsiella pneumoniae Isolates Uncovers Its Potential Association With Antibiotic Susceptibility