CRISPR Visualizer: rapid identification and visualization of CRISPR loci via an automated high-throughput processing pipeline

Abstract

A CRISPR locus, defined by an array of repeat and spacer elements, constitutes a genetic record of the ceaseless battle between bacteria and viruses, showcasing the genomic integration of spacers acquired from invasive DNA. In particular, iterative spacer acquisitions represent unique evolutionary histories and are often useful for high-resolution bacterial genotyping, including comparative analysis of closely related organisms, clonal lineages, and clinical isolates. Current spacer visualization methods are typically tedious and can require manual data manipulation and curation, including spacer extraction at each CRISPR locus from genomes of interest. Here, we constructed a high-throughput extraction pipeline coupled with a local web-based visualization tool which enables CRISPR spacer and repeat extraction, rapid visualization, graphical comparison, and progressive multiple sequence alignment. We present the bioinformatic pipeline and investigate the loci of reference CRISPR-Cas systems and model organisms in 4 well-characterized subtypes. We illustrate how this analysis uncovers the evolutionary tracks and homology shared between various organisms through visual comparison of CRISPR spacers and repeats, driven through progressive alignments. Due to the ability to process unannotated genome files with minimal preparation and curation, this pipeline can be implemented promptly. Overall, this efficient high-throughput solution supports accelerated analysis of genomic data sets and enables and expedites genotyping efforts based on CRISPR loci.

Comparative genomics of eight Lactobacillus buchneri strains isolated from food spoilage

“…To bolster our understanding of the environmental interaction between these strains and invasive nucleic acids, we analyzed their CRISPR-Cas systems in detail. Location and identification of CRISPR-Cas systems were not hindered by the highly fragmented genome assemblies, and loci were successfully assigned a canonical type and subtype using standard tools and references (37, 38). Across the 10 strains analyzed, we found CRISPR-Cas systems belonging to both II-A and I-E canonical subtypes [37].…”

“…CRISPR-Cas loci were identified, visualized, and aligned using CRISPRviz [38], and type was determined using the canonical definitions defined by Koonin et al [68] after inspecting flanking cas genes and their corresponding annotations. Spacer identity was investigated using BLAST+ [39] against the nt, env_nt, and gss remote databases with the following flags: -task blastn-short -dust no -outfmt 5 -evalue 1e-5.…”

Cronobacter sakazakii, Cronobacter malonaticus, and Cronobacter dublinensis Genotyping Based on CRISPR Locus Diversity

“…Considered the temporal organization of spacers, the sequencing of CRISPR arrays has been a extremely useful tool to genotype bacteria like Yersinia species, E. coli , and Salmonella enterica (Cui et al, 2008; Fricke et al, 2011; Yin et al, 2013; Li et al, 2014; Bugarel et al, 2018), and it has also been used to investigate bacterial diversity based on metagenomic data (Berg Miller et al, 2012; Sun et al, 2016). Recently, some useful tools to extract spacers and visualize the spacer content with color schemes were developed (Biswas et al, 2016; Couvin et al, 2018; Dion et al, 2018; Nethery and Barrangou, 2019).…”

Harnessing the type I CRISPR‐Cas systems for genome editing in prokaryotes

“…The subtype is characterized by the signature subtype gene cas8f (formerly csy1 ) and the unique cas2 ‐ cas3 fusion (Fig. 3) (Richter et al, 2012; Nethery and Barrangou, 2019b). Type I‐F CRISPR‐Cas‐mediated chromosome targeting at dispensable genomic islands has been initially demonstrated to cause rapid and spontaneous genomic alterations including the remodeling or deletion of entire pathogenicity islands in Pectobacterium atrosepticum (Vercoe et al, 2013).…”