Clustered Regularly Interspaced Short Palindromic Repeats (CRISPRs) for the Genotyping of Bacterial Pathogens

Grissa, Ibtissem; Vergnaud, Gilles; Pourcel, Christine

doi:10.1007/978-1-60327-999-4_9

Cited by 25 publications

(14 citation statements)

References 21 publications

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“…The sequences were analysed with the software CRISPRcompar and CRISPRtionary via the CRISPR website http://crispr.u-psud.fr/ [30]–[33]. The previously published CRISPR data from Y. pestis strains was used as reference (“spacers dictionary” Table S2, this report and Table S1 in Cui et al [18]).…”

Section: Methodsmentioning

confidence: 99%

Yersinia pestis Lineages in Mongolia

et al. 2012

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BackgroundWhole genome sequencing allowed the development of a number of high resolution sequence based typing tools for Yersinia (Y.) pestis. The application of these methods on isolates from most known foci worldwide and in particular from China and the Former Soviet Union has dramatically improved our understanding of the population structure of this species. In the current view, Y. pestis including the non or moderate human pathogen Y. pestis subspecies microtus emerged from Yersinia pseudotuberculosis about 2,600 to 28,600 years ago in central Asia. The majority of central Asia natural foci have been investigated. However these investigations included only few strains from Mongolia.Methodology/Principal FindingsClustered Regularly Interspaced Short Prokaryotic Repeats (CRISPR) analysis and Multiple-locus variable number of tandem repeats (VNTR) analysis (MLVA) with 25 loci was performed on 100 Y. pestis strains, isolated from 37 sampling areas in Mongolia. The resulting data were compared with previously published data from more than 500 plague strains, 130 of which had also been previously genotyped by single nucleotide polymorphism (SNP) analysis. The comparison revealed six main clusters including the three microtus biovars Ulegeica, Altaica, and Xilingolensis. The largest cluster comprises 78 isolates, with unique and new genotypes seen so far in Mongolia only. Typing of selected isolates by key SNPs was used to robustly assign the corresponding clusters to previously defined SNP branches.Conclusions/SignificanceWe show that Mongolia hosts the most recent microtus clade (Ulegeica). Interestingly no representatives of the ancestral Y. pestis subspecies pestis nodes previously identified in North-western China were identified in this study. This observation suggests that the subsequent evolution steps within Y. pestis pestis did not occur in Mongolia. Rather, Mongolia was most likely re-colonized by more recent clades coming back from China contemporary of the black death pandemic, or more recently in the past 600 years.

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Section: Methodsmentioning

confidence: 99%

Yersinia pestis Lineages in Mongolia

et al. 2012

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“…Cells that harbor these systems can be immunized against the attack of viruses by the integration of a virus-derived genome fragment into the host genome (1). The genetic memory of previous infections is mediated by CRISPR loci, which consist of a series of short repeat sequences (typically 24–37 bp) that are separated by spacer sequences (2–4). Cas proteins are often encoded in proximity to the CRISPR loci and are key players during all phases of immunization and protection of the cell (5,6).…”

Section: Introductionmentioning

confidence: 99%

In vitro assembly and activity of an archaeal CRISPR-Cas type I-A Cascade interference complex

Plagens

Tripp

Daume

et al. 2014

Nucleic Acids Research

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Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)-CRISPR-associated (Cas) systems of type I use a Cas ribonucleoprotein complex for antiviral defense (Cascade) to mediate the targeting and degradation of foreign DNA. To address molecular features of the archaeal type I-A Cascade interference mechanism, we established the in vitro assembly of the Thermoproteus tenax Cascade from six recombinant Cas proteins, synthetic CRISPR RNAs (crRNAs) and target DNA fragments. RNA-Seq analyses revealed the processing pattern of crRNAs from seven T. tenax CRISPR arrays. Synthetic crRNA transcripts were matured by hammerhead ribozyme cleavage. The assembly of type I-A Cascade indicates that Cas3′ and Cas3′′ are an integral part of the complex, and the interference activity was shown to be dependent on the crRNA and the matching target DNA. The reconstituted Cascade was used to identify sequence motifs that are required for efficient DNA degradation and to investigate the role of the subunits Cas7 and Cas3′′ in the interplay with other Cascade subunits.

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“…They appear to be transferable in an intra and interspecies manner [35]. CRISPR polymorphism has been used to genotype strains and to perform phylogenetic analyses in a number of bacterial species [36]. Two CRISPR-cas systems have been found in the genome of several A. baumannii strains and in the A. bayli ADP1 strain [37].…”

Section: Introductionmentioning

confidence: 99%

Diversity of Acinetobacter baumannii in Four French Military Hospitals, as Assessed by Multiple Locus Variable Number of Tandem Repeats Analysis

et al. 2012

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BackgroundInfections by A. calcoaceticus-A. baumannii (ACB) complex isolates represent a serious threat for wounded and burn patients. Three international multidrug-resistant (MDR) clones (EU clone I-III) are responsible for a large proportion of nosocomial infections with A. baumannii but other emerging strains with high epidemic potential also occur.Methodology/Principal FindingsWe automatized a Multiple locus variable number of tandem repeats (VNTR) analysis (MLVA) protocol and used it to investigate the genetic diversity of 136 ACB isolates from four military hospitals and one childrens hospital. Acinetobacter sp other than baumannii isolates represented 22.6% (31/137) with a majority being A. pittii. The genotyping protocol designed for A.baumannii was also efficient to cluster A. pittii isolates. Fifty-five percent of A. baumannii isolates belonged to the two international clones I and II, and we identified new clones which members were found in the different hospitals. Analysis of two CRISPR-cas systems helped define two clonal complexes and provided phylogenetic information to help trace back their emergence.Conclusions/SignificanceThe increasing occurrence of A. baumannii infections in the hospital calls for measures to rapidly characterize the isolates and identify emerging clones. The automatized MLVA protocol can be the instrument for such surveys. In addition, the investigation of CRISPR/cas systems may give important keys to understand the evolution of some highly successful clonal complexes.

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Clustered Regularly Interspaced Short Palindromic Repeats (CRISPRs) for the Genotyping of Bacterial Pathogens

Cited by 25 publications

References 21 publications

Yersinia pestis Lineages in Mongolia

Yersinia pestis Lineages in Mongolia

In vitro assembly and activity of an archaeal CRISPR-Cas type I-A Cascade interference complex

Diversity of Acinetobacter baumannii in Four French Military Hospitals, as Assessed by Multiple Locus Variable Number of Tandem Repeats Analysis

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