CRESSENT: a Bioinformatic Toolkit to Explore and Improve ssDNA Virus Annotation

Single-stranded DNA (ssDNA) viruses are important components of diverse ecosystems, however, it remains challenging to systematically identify and classify them. This is in part due to their broad host range and resulting genomic diversity, structure, and rapid evolution rates. In addition, distinguishing genuine ssDNA genomes from contaminating sequences in metagenomic datasets (e.g., from commercial kits) has been an unresolved issue for years. Here, we present CRESSENT ( CRESS -DNA E xtended a N notation T oolkit), a comprehensive and modular bioinformatic pipeline focused on ssDNA virus genome-to-analysis and annotation. The pipeline integrates multiple functionalities organized into six modules: sequence dereplication, decontamination, phylogenetic analysis, motif discovery, stem-loop structure prediction, and recombination detection. Each module can be used independently or in combination with others, allowing researchers to customize their analysis workflow. With this tool, researchers can comprehensively and systematically include ssDNA viruses in their viromics workflows and facilitate comparative genomic studies, which are often limited to dsDNA viruses, therefore leaving behind a crucial component of the microbiome community under study.