Pilon: An Integrated Tool for Comprehensive Microbial Variant Detection and Genome Assembly Improvement

TLDR

Modern sequencing technologies enable rapid analysis of hundreds of bacterial genomes per run, creating a need for fully automated methods to generate high‑quality assemblies and variant calls. The study introduces Pilon, an automated, all‑in‑one tool for correcting draft assemblies and detecting sequence variants of various sizes, including large insertions and deletions. Pilon operates on diverse sequence data, performing best with paired‑end reads from two Illumina libraries of small (≈180 bp) and large (≈3–5 kb) insert sizes. Pilon markedly improves draft assemblies by correcting bases, fixing mis‑assemblies, and filling gaps, producing more contiguous, error‑free genomes and enabling detection of more biologically relevant genes, while accurately identifying small and large variants and is already used to enhance thousands of bacterial genomes and variant catalogs. Pilon is freely available as open‑source software.

Abstract

Advances in modern sequencing technologies allow us to generate sufficient data to analyze hundreds of bacterial genomes from a single machine in a single day. This potential for sequencing massive numbers of genomes calls for fully automated methods to produce high-quality assemblies and variant calls. We introduce Pilon, a fully automated, all-in-one tool for correcting draft assemblies and calling sequence variants of multiple sizes, including very large insertions and deletions. Pilon works with many types of sequence data, but is particularly strong when supplied with paired end data from two Illumina libraries with small e.g., 180 bp and large e.g., 3-5 Kb inserts. Pilon significantly improves draft genome assemblies by correcting bases, fixing mis-assemblies and filling gaps. For both haploid and diploid genomes, Pilon produces more contiguous genomes with fewer errors, enabling identification of more biologically relevant genes. Furthermore, Pilon identifies small variants with high accuracy as compared to state-of-the-art tools and is unique in its ability to accurately identify large sequence variants including duplications and resolve large insertions. Pilon is being used to improve the assemblies of thousands of new genomes and to identify variants from thousands of clinically relevant bacterial strains. Pilon is freely available as open source software.