title: Host Subtraction, Filtering and Assembly Validations for Novel Viral Discovery Using Next Generation Sequencing Data.
abstract: The use of next generation sequencing (NGS) to identify novel viral sequences from eukaryotic tissue samples is challenging. Issues can include the low proportion and copy number of viral reads and the high number of contigs (post-assembly), making subsequent viral analysis difficult. Comparison of assembly algorithms with pre-assembly host-mapping subtraction using a short-read mapping tool, a k-mer frequency based filter and a low complexity filter, has been validated for viral discovery with Illumina data derived from naturally infected liver tissue and simulated data. Assembled contig numbers were significantly reduced (up to 99.97%) by the application of these pre-assembly filtering methods. This approach provides a validated method for maximizing viral contig size as well as reducing the total number of assembled contigs that require down-stream analysis as putative viral nucleic acids.
authors.affiliation: Lab of Viral Zoonotics, Department of Veterinary Medicine, University of Cambridge, Madingley Road, Cambridge, CB30ES, United Kingdom.
authors.forename: Gordon M
authors.initials: GM
authors.lastname: Daly
authors.affiliation: The Genome Analysis Centre (TGAC), Norwich Research Park, Norwich, NR47UH, United Kingdom.
authors.forename: Richard M
authors.initials: RM
authors.lastname: Leggett
authors.affiliation: Lab of Viral Zoonotics, Department of Veterinary Medicine, University of Cambridge, Madingley Road, Cambridge, CB30ES, United Kingdom.
authors.forename: William
authors.initials: W
authors.lastname: Rowe
authors.affiliation: Lab of Viral Zoonotics, Department of Veterinary Medicine, University of Cambridge, Madingley Road, Cambridge, CB30ES, United Kingdom.
authors.forename: Samuel
authors.initials: S
authors.lastname: Stubbs
authors.affiliation: Lab of Viral Zoonotics, Department of Veterinary Medicine, University of Cambridge, Madingley Road, Cambridge, CB30ES, United Kingdom.
authors.forename: Maxim
authors.initials: M
authors.lastname: Wilkinson
authors.affiliation: The Genome Analysis Centre (TGAC), Norwich Research Park, Norwich, NR47UH, United Kingdom.
authors.forename: Ricardo H
authors.initials: RH
authors.lastname: Ramirez-Gonzalez
authors.affiliation: The Genome Analysis Centre (TGAC), Norwich Research Park, Norwich, NR47UH, United Kingdom.
authors.forename: Mario
authors.initials: M
authors.lastname: Caccamo
authors.affiliation: Institute of Liver Studies, King's College Hospital, Denmark Hill, London, SE59RS, United Kingdom.
authors.forename: William
authors.initials: W
authors.lastname: Bernal
authors.affiliation: Lab of Viral Zoonotics, Department of Veterinary Medicine, University of Cambridge, Madingley Road, Cambridge, CB30ES, United Kingdom.
authors.forename: Jonathan L
authors.initials: JL
authors.lastname: Heeney
mesh_terms: D000465:Algorithms; D020451:Contig Mapping / Q000379:methods*; D059018:DNA Contamination; D004279:DNA, Viral / Q000737:chemistry*; D059014:High-Throughput Nucleotide Sequencing / Q000379:methods*; D006801:Humans; D008099:Liver / Q000821:virology; D017422:Sequence Analysis, DNA / Q000379:methods
chemical_list: D004279:DNA, Viral