ISMapper: identifying transposase insertion sites in bacterial genomes from short read sequence data

Abstract

BackgroundInsertion sequences (IS) are small transposable elements, commonly found in bacterial genomes. Identifying the location of IS in bacterial genomes can be useful for a variety of purposes including epidemiological tracking and predicting antibiotic resistance. However IS are commonly present in multiple copies in a single genome, which complicates genome assembly and the identification of IS insertion sites. Here we present ISMapper, a mapping-based tool for identification of the site and orientation of IS insertions in bacterial genomes, directly from paired-end short read data.ResultsISMapper was validated using three types of short read data: (i) simulated reads from a variety of species, (ii) Illumina reads from 5 isolates for which finished genome sequences were available for comparison, and (iii) Illumina reads from 7 Acinetobacter baumannii isolates for which predicted IS locations were tested using PCR. A total of 20 genomes, including 13 species and 32 distinct IS, were used for validation. ISMapper correctly identified 97 % of known IS insertions in the analysis of simulated reads, and 98 % in real Illumina reads. Subsampling of real Illumina reads to lower depths indicated ISMapper was able to correctly detect insertions for average genome-wide read depths >20x, although read depths >50x were required to obtain confident calls that were highly-supported by evidence from reads. All ISAba1 insertions identified by ISMapper in the A. baumannii genomes were confirmed by PCR. In each A. baumannii genome, ISMapper successfully identified an IS insertion upstream of the ampC beta-lactamase that could explain phenotypic resistance to third-generation cephalosporins. The utility of ISMapper was further demonstrated by profiling genome-wide IS6110 insertions in 138 publicly available Mycobacterium tuberculosis genomes, revealing lineage-specific insertions and multiple insertion hotspots.ConclusionsISMapper provides a rapid and robust method for identifying IS insertion sites directly from short read data, with a high degree of accuracy demonstrated across a wide range of bacteria.

Impact of insertion sequences on convergent evolution ofShigellaspecies

“…These numbers are lower (median 53-86%) than those identified in completed reference chromosomes (Supplementary Figure 1) 13 . This underestimation is expected because ISMapper detects insertion sites relative to an IS-free reference sequence for each species 28 , and cannot detect insertions of IS within other IS (which does occur in the reference genomes) 13 . IS are also present in the virulence plasmid sequences of finished Shigella reference genomes 13 ( Supplementary Table 2); however as the plasmid is frequently lost during culture 29 and was lacking from many of the short read data sets 23 , IS variation in the virulence plasmid was not further examined in this study.…”

Challenges and opportunities for whole‐genome sequencing–based surveillance of antibiotic resistance

“…Alternatively, transposon insertions can increase the expression of intrinsic resistance genes, as illustrated by transposon‐mediated upregulation of ampC in A. baumannii , causing third‐generation cephalosporin resistance, or increased expression of an efflux system in E. coli , which contributes to fluoroquinolone resistance . In cases where transposon‐mediated resistance has been experimentally verified, these events may be predicted from genome data, using either read‐based tools (ISMapper) or assembly‐based analysis (ISseeker) to identify transposon insertion sites in microbial genomes.…”

Multiple Genetic Mutations Associated with Polymyxin Resistance in Acinetobacter baumannii

“…To identify insertion sequences (IS) that differed between the isolates in each pair, we screened all draft genome assemblies to identify every IS present using the ISFinder database (21), determining the presence or absence of these sequences in each isolate read set using ISMapper (22).…”