Schematic of bioinformatics pipeline used for processing of NGS libraries.

<p>High quality reads, excluding ribosomal and mitochondrial sequences, were aligned against the taxonomy databases of NCBI using BLASTn (taxonomic classification). Unclassified or ambiguously classified reads, together with virus, phage, and HERV sequences were assembled into scaffolds. Scaffolds were used to query the non-redundant protein database of NCBI using BLASTx to identify viral proteins with similarity to predicted polypeptides in our scaffolds (finding novel viruses). Given the large genomes of NCLDVs, hits to this class of viruses were reanalyzed with the profile hidden Markov model-based algorithm HHblits. PCR and Sanger sequencing were used to confirm the presence of novel viral-like sequences in our samples.