LazypipeX: customizable virome analysis pipeline enabling fast and sensitive virus discovery from NGS data

Abstract Metagenomic next-generation sequencing (mNGS) is pivotal for detecting known and novel viruses, however, its efficacy depends on applied bioinformatics pipelines. We present Lazypipe Extended (LazypipeX), an mNGS pipeline employing modular “annotation strategies” to optimize the balance between speed and sensitivity as required by different applications. Using a synthetic benchmark, we show that a simple two-round strategy designed for speed and low false positive rate reduces execution time to a fraction of traditional BLASTN searches with no loss in accuracy. Additionally, using real data, we show that strategies based on combinations of fast-to-accurate homology searches reduce execution time from 5- to 20-fold compared to the BLASTN/BLASTP baseline. Using one of these “annotation chains”, we characterized multiple novel complete viral genomes that were missed by the Lazypipe v1/v2 and CZ ID mNGS pipelines. Overall, the LazypipeX offers customizable and transparent workflows that can facilitate virus discovery and identification in diverse mNGS applications.