BACKGROUND: Next generation sequencing (NGS) technology allows laboratories to investigate virome composition in clinical and environmental samples in a culture-independent way. There is a need for bioinformatic tools capable of parallel processing of virome sequencing data by exactly identical methods: this is especially important in studies of multifactorial diseases, or in parallel comparison of laboratory protocols. RESULTS: We have developed a web-based application allowing direct upload of sequences from multiple virome samples using custom parameters. The samples are then processed in parallel using an identical protocol, and can be easily reanalyzed. The pipeline performs de-novo assembly, taxonomic classification of viruses as well as sample analyses based on user-defined grouping categories. Tables of virus abundance are produced from cross-validation by remapping the sequencing reads to a union of all observed reference viruses. In addition, read sets and reports are created after processing unmapped reads against known human and bacterial ribosome references. Secured interactive results are dynamically plotted with population and diversity charts, clustered heatmaps and a sortable and searchable abundance table. CONCLUSIONS: The Vipie web application is a unique tool for multi-sample metagenomic analysis of viral data, producing searchable hits tables, interactive population maps, alpha diversity measures and clustered heatmaps that are grouped in applicable custom sample categories. Known references such as human genome and bacterial ribosomal genes are optionally removed from unmapped ('dark matter') reads. Secured results are accessible and shareable on modern browsers. Vipie is a freely available web-based tool whose code is open source.

BioMediTech and Faculty of Medicine and Life Sciences University of Tampere PB 100 FI 33014 Tampere Finland

Department of Pediatrics 2nd Faculty of Medicine Charles University and University Hospital Motol 5 Úvalu 84 150 06 Praha 5 Czech Republic

Fimlab Laboratories Pirkanmaa Hospital District Tampere Finland

School of Social Sciences University of Tampere Kalevantie 4 33100 Tampere Finland

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The Human Microbiome Consortium Structure, function and diversity of the healthy human microbiome. Nature. 2012;486(7402):207–14. doi: 10.1038/nature11234. PubMed DOI PMC

Turnbaugh PJ, Ley RE, Hamady M, Fraser-Liggett CM, Knight R, Gordon JI. The human microbiome project. Nature. 2007;449(7164):804–10. doi: 10.1038/nature06244. PubMed DOI PMC

Houldcroft CJ, Beale MA, Breuer J. Clinical and biological insights from viral genome sequencing. Nat Rev Microbiol. 2017;15(3):183–192. doi: 10.1038/nrmicro.2016.182. PubMed DOI PMC

Tringe SG, Rubin EM. Metagenomics: DNA sequencing of environmental samples. Nat Rev Genet. 2005;6:805–814. doi: 10.1038/nrg1709. PubMed DOI

Shapton TJ. An introduction to the analysis of shotgun metagenomic data. Front Plant Sci. 2014;5:209. PubMed PMC

Flygare S, Simon K, et al. Taxonomer: an interactive metagenomics analysis portal for universal pathogen detection and host mRNA expression profiling. Genome Biol. 2016;201617:111. doi: 10.1186/s13059-016-0969-1. PubMed DOI PMC

Yamashita A, et al. VirusTAP: viral genome-targeted assembly pipeline. Front Microbiol. 2016;7:32. PubMed PMC

Wommack KE, Bhavsar J, et al. VIROME: a standard operating procedure for analysis of viral metagenome sequences. Stand Genomic Sci. 2012;6(3):427–39. doi: 10.4056/sigs.2945050. PubMed DOI PMC

Roux S, Faubladier M, et al. Metavir: a web server dedicated to virome analysis. Bioinformatics. 2011;27(21):3074–5. doi: 10.1093/bioinformatics/btr519. PubMed DOI

Roux S, et al. Metavir 2: new tools for viral metagenome comparison and assembled virome analysis. BMC Bioinf. 2014;15:76. doi: 10.1186/1471-2105-15-76. PubMed DOI PMC

Rampelli S, Soverini M, et al. ViromeScan: a new tool for metagenomic viral community profiling. BMC Genomics. 2016;17:165. doi: 10.1186/s12864-016-2446-3. PubMed DOI PMC

Fosso B. et al. MetaShot: an accurate workflow for taxon classification of host-associated microbiome from shotgun metagenomic data. Bioinform. 2017. doi: 10.1093/bioinformatics/btx036. PubMed PMC

Afgan E, Taylor J, Anton Nekrutenko A, Goecks J, et al. The galaxy platform for accessible, reproducible and collaborative biomedical analyses: 2016 update. Nucleic Acids Res. 2016;44(W1):W3–W10. doi: 10.1093/nar/gkw343. PubMed DOI PMC

Blankenberg D, the Galaxy Team. Taylor J, Nekrutenko A, et al. Dissemination of scientific software with galaxy ToolShed. Genome Biol. 2014;15:403. doi: 10.1186/gb4161. PubMed DOI PMC

Zerbina DR, Birney E. Velvet: algorithms for de novo short read assembly using de Bruijn graphs. Genome Res. 2008;18:821–829. doi: 10.1101/gr.074492.107. PubMed DOI PMC

Namiki T, Hachiya T, Tanaka H, Sakakibara Y. MetaVelvet : An extension of Velvet assembler to de novo metagenome assembly from short sequence reads. Nucleic Acids Res. 2012;40(20):e155. doi: 10.1093/nar/gks678. PubMed DOI PMC

Peng Y, et al. IDBA-UD: a de novo assembler for single-cell and metagenomic sequencing data with highly uneven depth. Bioinformatics. 2013;28:1420–1. doi: 10.1093/bioinformatics/bts174. PubMed DOI

Li D, et al. MEGAHIT: an ultra-fast single-node solution for large and complex metagenomics assembly via succinct de Bruijn graph. Bioinformatics. 2015;31(10):1674–6. doi: 10.1093/bioinformatics/btv033. PubMed DOI

Simpson K, et al. ABySS: A parallel assembler for short read sequence data. Genome Res. 2009;19(6):1117–1123. doi: 10.1101/gr.089532.108. PubMed DOI PMC

Paszkiewicz K, Studholme DJ. De novo assembly of short sequence reads. Brief Bioinform. 2010;11(5):457–472. doi: 10.1093/bib/bbq020. PubMed DOI

Tritt A, Eisen JA, Facciotti MT, Darling AE. An integrated pipeline for de novo assembly of microbial genomes. PLoS One. 2012;7(9):e42304. doi: 10.1371/journal.pone.0042304. PubMed DOI PMC

Li Y, et al. VIP: an integrated pipeline for metagenomics of virus identification and discovery. Sci Rep. 2016;6:23774. doi: 10.1038/srep23774. PubMed DOI PMC

Altschul SF, et al. Basic local alignment search tool. J Mol Biol. 1990;215:403. doi: 10.1016/S0022-2836(05)80360-2. PubMed DOI

Li H, Durbin R. Fast and accurate short read alignment with Burrows-Wheeler transform. Bioinformatics. 2009;25(14):1754–1760. doi: 10.1093/bioinformatics/btp324. PubMed DOI PMC

Szymanski M, Zielezinski A, et al. 5SRNAdb: an information resource for 5S ribosomal RNAs. Nucleic Acids Res. 2016;44(D1):D180–3. doi: 10.1093/nar/gkv1081. PubMed DOI PMC

Edgar RC. Search and clustering orders of magnitude faster than BLAST. Bioinformatics. 2010;26(19):2460–1. doi: 10.1093/bioinformatics/btq461. PubMed DOI

Cock PA, Antao T, Chang JT, Bradman BA, Cox CJ, Dalke A, Friedberg I, Hamelryck T, Kauff F, Wilczynski B, de Hoon MJL. Biopython: freely available python tools for computational molecular biology and bioinformatics. Bioinformatics. 2009;25:1422–1423. doi: 10.1093/bioinformatics/btp163. PubMed DOI PMC

Wickham H. ggplot2: elegant graphics for data analysis. New York: Springer; 2009.

Kimura H, et al. A food-borne outbreak of gastroenteritis due to genotype G1P[8] rotavirus among adolescents in Japan. Microbiol Immunol. 2014;58(9):536–9. doi: 10.1111/1348-0421.12176. PubMed DOI

DNA Data bank of Japan http://getentry.ddbj.nig.ac.jp/(DRA004165) Accessed 01 Dec 2016.

Rodríguez-Diaz J, et al. Presence of human enteric viruses in the stools of healthy Malawian 6-month-old infants. J Pediatr Gastroenterol Nutr. 2014;58(4):502–4. doi: 10.1097/MPG.0000000000000215. PubMed DOI

Mangani C, et al. Effect of complementary feeding with lipid-based nutrient supplements and corn-soy blend on the incidence of stunting and linear growth among 6- to 18-month-old infants and children in rural Malawi. Matern Child Nutr. 2015;11(Suppl 4):132–43. doi: 10.1111/mcn.12068. PubMed DOI PMC

Vipie project SourceForge https://sourceforge.net/projects/vipie/files/data/Accessed 15 Mar 2017

Shannon CE. A mathematical theory of communication. Bell Syst Tech J. 1948;27:379–423 and 623–656. doi: 10.1002/j.1538-7305.1948.tb01338.x. DOI

Simpson EH. Measurement of diversity. Nature. 1949;163:688. doi: 10.1038/163688a0. DOI

Dutilh BE, Edwards RA, et al. A highly abundant bacteriophage discovered in the unknown sequences of human faecal metagenomes. Nat Commun. 2014;5:4498. doi: 10.1038/ncomms5498. PubMed DOI PMC

NIH Human Microbiome Project website. http://www.hmpdacc.org/HMASM/HMASM-690.csv. Accessed 01 Jan 2017

Wylie KM, Mihindukulasuriya KA, Zhou Y, Sodergren E, Storch GA, Weinstock GM. Metagenomic analysis of double-stranded DNA viruses in healthy adults. BMC Biol. 2014;12:71. doi: 10.1186/s12915-014-0071-7. PubMed DOI PMC

Huang W, et al. ART: a next-generation sequencing read simulator. Bioinformatics. 2012;28:593–594. doi: 10.1093/bioinformatics/btr708. PubMed DOI PMC

McMurdie PJ, Holmes S. Phyloseq: an R package for reproducible interactive analysis and graphics of microbiome census data. PLoS One. 2013;8(4):e61217. doi: 10.1371/journal.pone.0061217. PubMed DOI PMC

Love MI, Huber W, Anders S. Moderated estimation of fold change and dispersion for RNA-seq data with DESeq2. Genome Biol. 2014;15:550. doi: 10.1186/s13059-014-0550-8. PubMed DOI PMC

Audano P, Vannberg F. KAnalyze: a fast versatile pipelined k-mer toolkit. Bioinformatics. 2014;30:2070–2. doi: 10.1093/bioinformatics/btu152. PubMed DOI PMC

Alonso-Alemany D, et al. Further steps in TANGO: improved taxonomic assignment in metagenomics. Bioinformatics. 2014;30(1):17–23. doi: 10.1093/bioinformatics/btt256. PubMed DOI

Sayers EW, et al. Database resources of the national center for biotechnology information. Nucleic Acids Res. 2009;37(Database issue):D5–15. doi: 10.1093/nar/gkn741. PubMed DOI PMC