Abstract
Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) are diverse immune systems found in many prokaryotic genomes that target invading foreign DNA such as bacteriophages and plasmids. There are multiple types of CRISPR with arguably the most enigmatic being Type IV. During an investigation of CRISPR carriage in clinical, multi-drug resistant, Klebsiella pneumoniae, a Type IV-A3 CRISPR-Cas system was detected on plasmids from two K. pneumoniae isolates from Egypt (isolated in 2002–2003) and a single K. pneumoniae isolate from the United Kingdom (isolated in 2017). Sequence analysis of all other genomes available in GenBank revealed that this CRISPR-Cas system was present on 28 other plasmids from various Enterobacteriaceae hosts and was never found on a bacterial chromosome. This system is exclusively located on IncHI1B/IncFIB plasmids and is associated with multiple putative transposable elements. Expression of the cas loci was confirmed in the available clinical isolates by RT-PCR. In all cases, the CRISPR-Cas system has a single CRISPR array (CRISPR1) upstream of the cas loci which has several, conserved, spacers which, amongst things, match regions within conjugal transfer genes of IncFIIK/IncFIB(K) plasmids. Our results reveal a Type IV-A3 CRISPR-Cas system exclusively located on IncHI1B/IncFIB plasmids in Enterobacteriaceae that is likely to be able to target IncFIIK/IncFIB(K) plasmids presumably facilitating intracellular, inter-plasmid competition.
Highlights
Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR-Cas) are widespread, adaptive, RNA-mediated, immune systems found in the genomes of prokaryotic organisms that target invading foreign DNA such as bacteriophages and conjugative plasmids (Barrangou et al, 2007; Marraffini and Sontheimer, 2010a)
Type IV-A-3 CRISPR-Cas, based on the gene composition and genetic architecture of the IV-A variants detected in K. pneumoniae as described in Pinilla-Redondo et al (2019), was detected on a total of thirty-one IncHI1B/IncFIB(Mar) plasmid sequences within Enterobacteriaceae (Figure 1 and Supplementary Table S1)
The IncH1B/IncFIB plasmids are large, low copy number, conjugative plasmids with narrow-hostranges, which are found in multiple genera of Enterobacteriaceae (Zhong et al, 2005; Suzuki et al, 2010; Faure et al, 2019a)
Summary
Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR-Cas) are widespread, adaptive, RNA-mediated, immune systems found in the genomes of prokaryotic organisms (bacteria and archaea) that target invading foreign DNA such as bacteriophages and conjugative plasmids (Barrangou et al, 2007; Marraffini and Sontheimer, 2010a). CRISPRCas systems are confirmed, or expected, to provide immunity against viruses and other mobile genetic elements (MGEs), except for transposon-encoded CRISPR-Cas systems that lack the interference module and are predicted to perform functions distinct from adaptive immunity (Makarova et al, 2020). Type IV was previously called the Unknown Type (Type U), due to its rare occurrence and lack of the adaptation module, until an updated classification in 2015 (Makarova et al, 2013; Zhang and Ye, 2017). It was named Type IV (putative) after its identification in Acidithiobacillus ferrooxidans presenting a different genetic arrangement of Type U cas genes (Makarova et al, 2015). It has been hypothesized that Type IV is similar to an ancestral innate immune system that gained adaptive ability by associating with a transposon-like element containing cas and cas (Rath et al, 2015)
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