DNase treatment improves viral enrichment in agricultural soil viromes

Abstract

The small genomes of most viruses make it difficult to fully capture viral diversity in metagenomes dominated by DNA from cellular organisms. Viral size-fraction metagenomics (viromics) protocols facilitate enrichment of viral DNA from environmental samples, and these protocols typically include a DNase treatment of the post-0.2 m viromic fraction to remove contaminating free DNA prior to virion lysis. However, DNase may also remove desirable viral genomic DNA (e.g., contained in virions compromised due to frozen storage or laboratory processing), suggesting that DNase-untreated viromes might be useful in some cases. In order to understand how virome preparation with and without DNase treatment influences the resultant data, here we compared 15 soil viromes (7 DNase-treated, 8 untreated) from 8 samples collected from agricultural fields prior to tomato planting. DNase-treated viromes yielded significantly more assembled viral contigs, contained significantly less non-viral microbial DNA, and recovered more viral populations (vOTUs) through read mapping. However, DNase-treated and untreated viromes were statistically indistinguishable, in terms of ecological patterns across viral communities. Although results suggest that DNase treatment is preferable where possible, in comparison to previously reported total metagenomes from the same samples, both DNase-treated and untreated viromes were significantly enriched in viral signatures by all metrics compared, including a ~225 times greater proportion of viral reads in untreated viromes compared to total metagenomes. Thus, even without DNase treatment, viromics was preferable to total metagenomics for capturing viral diversity in these soils, suggesting that preparation of DNase-untreated viromes can be worthwhile when DNase treatment is not possible. ImportanceViromics is becoming an increasingly popular method for characterizing soil viral communities. DNase treatment of the viral size fraction prior to DNA extraction is meant to reduce contaminating free DNA and is a common step within viromics protocols to ensure sequences are of viral origin. However, some samples may not be amendable to DNase treatment due to viral particles being compromised either in storage (i.e. frozen) or during other sample processing. To date, the effect of DNase treatment on the recovery of viruses and downstream ecological interpretations of soil viral communities is not thoroughly understood. This work sheds light on these questions and indicates that while DNase treatment of soil viromes improves recovery of viral populations, this improvement is modest in comparison to the gains made by viromics over total soil metagenomics. Further, DNase treatment may not be necessary to observe the ecological patterns structuring soil viral communities.