ExplorePipolin: reconstruction and annotation of piPolB-encoding bacterial mobile elements from draft genomes

Abstract

Motivation Detailed and accurate analysis of mobile genetic elements (MGEs) in bacteria is essential to deal with the current threat of multiresistant microbes. The overwhelming use of draft, contig-based genomes hinder the delineation of the genetic structure of these plastic and variable genomic stretches, as in the case of pipolins, a superfamily of MGEs that spans diverse integrative and plasmidic elements, characterized by the presence of a primer-independent DNA polymerase. Results ExplorePipolin is a Python-based pipeline that screens for the presence of the element and performs its reconstruction and annotation. The pipeline can be used on virtually any genome from diverse organisms and of diverse quality, obtaining the highest-scored possible structure, and reconstructed out of different contigs if necessary. Then, predicted pipolin boundaries and pipolin encoded genes are subsequently annotated using a custom database, returning the standard file formats suitable for comparative genomics of this mobile element. Availability All code is available and can be accessed here: github.com/pipolinlab/ExplorePipolin Contact modesto.redrejo@uam.es Supplementary information Supplementary data are available at Bioinformatics Advances online.

Pipolins are bimodular platforms that maintain a reservoir of defense systems exchangeable with various bacterial genetic mobile elements

“…In order to understand the diversity and evolution of pipolins, we performed a complete screening of the bacteria assemblies in GenBank Datasets, using the tool ExplorePipolin, based in profile searches against a piPolB curated HMM profile 26 . We detected putative pipolins in 11,431 of >1.3M genome assemblies, comprising a prevalence of nearly 0.9% of bacterial genomes ( Figure 1 ).…”

“…Using ExplorePipolin 26 we were able to extract 8,646 bona fide complete integrated pipolins in bacteria assemblies, where the known direct repeats (DR) and adjacent tRNA integration site were successfully predicted, being around 6,000 the most-likely reconstruction from different contigs ( Figure 2B-C ). Additionally, it annotated 2,896 varied incomplete pipolins or piPolB-containing fragments where known flanking DRs and/or integration sites could not be identified due to deletions near the ends of the element, the presence of alternative structures (e.g., circular plasmids), or integration into a site other than a tRNA, as discussed below.…”

“…All bacterial genome assemblies from the GenBank database (1,310,495 genomes, accessed on November 11, 2022) were downloaded using the NCBI Datasets command line tool (version 13.43.2, https://www.ncbi.nlm.nih.gov/datasets) with the “--exclude-atypical” option and analyzed for the presence of pipolins using ExplorePipolin 26 . Briefly, ExplorePipolin is a pipeline that analyzes the presence of pipolins in multi-fasta nucleotide sequence files and returns several files describing pipolin genome coordinates, flanking direct repeats, genes encoded in the element, and other useful information.…”

“…Then pipolin protein function was inferred both at individual level, using EggNOG-mapper v2 35 for a general annotation, and at cluster level by taking representatives with CD-HIT (-c 0.9), aligning them with MAFFT and running hhblits 3.3.0 36 against the Pfam35 37 database (-n 5 iterations, top 3 hits with E-value < 0.001 kept). Additionally, we used specific profiles to annotate relaxases (through MOBScan 38 ), integrases 39,40 , and piPolBs 26 , applying the methods and thresholds used in the reference articles.…”