Benchmarking informatics approaches for virus discovery: caution is needed when combining <i>in silico</i> identification methods

Abstract

The identification of viruses from environmental metagenomes using informatics tools has offered critical insights in microbial ecology. However, it remains difficult for researchers to know which tools optimize viral recovery for their specific study. In an attempt to recover more viruses, studies are increasingly combining the outputs from multiple tools without validating this approach. After benchmarking combinations of six viral identification tools against mock metagenomes and environmental samples, we found that these tools should only be combined cautiously. Two to four tool combinations maximized viral recovery and minimized non-viral contamination compared with either the single-tool or the five- to six-tool ones. By providing a rigorous overview of the behavior of <i>in silico</i> viral identification strategies and a pipeline to replicate our process, our findings guide the use of existing viral identification tools and offer a blueprint for feature engineering of new tools that will lead to higher-confidence viral discovery in microbiome studies.