Generation of a CRISPR database for Yersinia pseudotuberculosis complex and role of CRISPR‐based immunity in conjugation

Abstract

The clustered regularly interspaced short palindromic repeat - CRISPR-associated genes (CRISPR-Cas) system is used by bacteria and archaea against invading conjugative plasmids or bacteriophages. Central to this immunity system are genomic CRISPR loci that contain fragments of invading DNA. These are maintained as spacers in the CRISPR loci between direct repeats and the spacer composition in any bacterium reflects its evolutionary history. We analysed the CRISPR locus sequences of 335 Yersinia pseudotuberculosis complex strains. Altogether 1902 different spacer sequences were identified and these were used to generate a database for the spacer sequences. Only ∼10% of the spacer sequences found matching sequences. In addition, surprisingly few spacers were shared by Yersinia pestis and Y. pseudotuberculosis strains. Interestingly, 32 different protospacers were present in the conjugative plasmid pYptb32953. The corresponding spacers were identified from 35 different Y. pseudotuberculosis strains indicating that these strains had encountered pYptb32953 earlier. In conjugation experiments, pYptb32953-specific spacers generally prevented conjugation with spacer-positive and spacer-free strains. However, some strains with one to four spacers were invaded by pYptb32953 and some spacer-free strains were fully resistant. Also some spacer-positive strains were intermediate resistant to conjugation. This suggests that one or more other defence systems are determining conjugation efficiency independent of the CRISPR-Cas system.

Comparative analysis of CRISPR‐Cas systems in Klebsiella genomes

“…CRISPR‐Cas system in several pathogens of the family of Enterobacteriaceae, including Yersinia , Escherichia , and Salmonella , has been well investigated . Both the Type I‐E and I‐F CRISPR‐Cas systems have been identified.…”

Dual nuclease activity of a Cas2 protein in CRISPR–Cas subtype I‐B of Leptospira interrogans

“…With the recent advancement of bioinformatics and genetic studies, CRISPR–Cas system have been estimated in virtually all archaea and approximately 40% of the known bacteria , including many pathogens such as Mycobacterium tuberculosis , Streptococcus , Yersinia, Neisseria , Campylobacter jejuni, Clostridium botulinum , Escherichia coli, Listeria monocytogenes , and Corynebacterium . Using similar approach, we propose the existence of a CRISPR–Cas system in bacterium Leptospira interrogans .…”

Phylogeographic separation and formation of sexually discrete lineages in a global population ofYersinia pseudotuberculosis

“…Serotype O:1 strains formed a distinct clade of strains encompassing a large number of sequence type complexes, suggesting a highly diverse population of bacteria within the serotype O:1 group of Y. pseudotuberculosis. In addition to MLST genotyping, recent work has also analysed clustered regularly interspaced short palindromic repeat (CRISPR) loci of 335 Y. pseudotuberculosis isolates 11 . The CRISPR-Cas system is an RNA-based immune system that regulates invasion of plasmids and viruses in bacteria and archaea 12 pseudotuberculosis genomes means that our understanding of the population structure and evolutionary events occurring within this species is poorly informed.…”

“…pseudotuberculosis -specific CRISPR direct repeat sequence (5'tttctaagctgcctgtgcggcagtgaac-3'), its complementary sequence, the 5'-and 3'-flanking sequences of the YP1, YP2 and YP3 loci and their complementary sequences 11 . Identified sequences were submitted to the CRISPRFinder tool at CRISPRs Web Server (http://crispr.u-psud.fr/) together with the spacer dictionary compiled earlier 11 . This analysis increased the number of identified spacers in the Y. pseudotuberculosis spacer dictionary from 1902 to 2969 (Table S2).…”