Next-generation sequencing and metagenomic analysis: a universal diagnostic tool in plant virology.

A novel, unbiased approach to plant viral disease diagnosis has been developed which requires no a priori knowledge of the host or pathogen. Next-generation sequencing coupled with metagenomic analysis was used to produce large quantities of cDNA sequence in a model system of tomato infected with Pepino mosaic virus. The method was then applied to a sample of Gomphrena globosa infected with an unknown pathogen originally isolated from the flowering plant Liatris spicata. This plant was found to contain a new cucumovirus, for which we suggest the name 'Gayfeather mild mottle virus'. In both cases, the full viral genome was sequenced. This method expedites the entire process of novel virus discovery, identification, viral genome sequencing and, subsequently, the development of more routine assays for new viral pathogens.

[1]  M. Breitbart,et al.  Using pyrosequencing to shed light on deep mine microbial ecology , 2006, BMC Genomics.

[2]  M. Nei,et al.  MEGA4: Molecular Evolutionary Genetics Analysis (MEGA) software version 4.0. , 2007, Molecular biology and evolution.

[3]  James R. Knight,et al.  Genome sequencing in microfabricated high-density picolitre reactors , 2005, Nature.

[4]  Reinhard Simon,et al.  Complete viral genome sequence and discovery of novel viruses by deep sequencing of small RNAs: a generic method for diagnosis, discovery and sequencing of viruses. , 2009, Virology.

[5]  H. Pospieszny,et al.  Complete genomic RNA sequence of the Polish Pepino mosaic virus isolate belonging to the US2 strain , 2008, Virus Genes.

[6]  J. Cairney,et al.  A simple and efficient method for isolating RNA from pine trees , 1993, Plant Molecular Biology Reporter.

[7]  P. Salamon,et al.  Metagenomic Analyses of an Uncultured Viral Community from Human Feces , 2003, Journal of bacteriology.

[8]  Peter Salamon,et al.  Metagenomic and Small-Subunit rRNA Analyses Reveal the Genetic Diversity of Bacteria, Archaea, Fungi, and Viruses in Soil , 2007, Applied and Environmental Microbiology.

[9]  Claire Cowie,et al.  Phylogenetic structure of Xanthomonas determined by comparison of gyrB sequences. , 2009, International journal of systematic and evolutionary microbiology.

[10]  Jay D. Evans,et al.  A Metagenomic Survey of Microbes in Honey Bee Colony Collapse Disorder , 2007, Science.

[11]  J. Simons,et al.  A new arenavirus in a cluster of fatal transplant-associated diseases. , 2008, The New England journal of medicine.

[12]  M. Egholm,et al.  Novel Borna Virus in Psittacine Birds with Proventricular Dilatation Disease , 2008, Emerging infectious diseases.

[13]  K. Dupuis,et al.  Rapid Identification of Known and New RNA Viruses from Animal Tissues , 2008, PLoS pathogens.

[14]  Alexander F. Auch,et al.  MEGAN analysis of metagenomic data. , 2007, Genome research.

[15]  W. Ian Lipkin,et al.  Genetic Detection and Characterization of Lujo Virus, a New Hemorrhagic Fever–Associated Arenavirus from Southern Africa , 2009, PLoS pathogens.

[16]  A. Halpern,et al.  The Sorcerer II Global Ocean Sampling Expedition: Metagenomic Characterization of Viruses within Aquatic Microbial Samples , 2008, PloS one.

[17]  J. B. Jones,et al.  Heterogeneity of Xanthomonas campestris pv. hederae Strains from Araliaceous Hosts. , 1999, Phytopathology.

[18]  S. Weller,et al.  Classification and identification of plant-associated bacteria by fatty acid profiling* , 2000 .

[19]  A. Gibbs,et al.  A primer pair for amplifying part of the genome of all potyvirids by RT-PCR. , 1997, Journal of virological methods.

[20]  Tao Zhang,et al.  RNA Viral Community in Human Feces: Prevalence of Plant Pathogenic Viruses , 2005, PLoS biology.

[21]  Jun Kawai,et al.  Metagenomic Diagnosis of Bacterial Infections , 2008, Emerging infectious diseases.

[22]  N. Boonham,et al.  First report of Broad bean wilt virus 2 in the UK: findings in foxglove and salvia , 2006 .

[23]  N. Pradel,et al.  Genomic Subtraction To Identify and Characterize Sequences of Shiga Toxin-Producing Escherichia coli O91:H21 , 2002, Applied and Environmental Microbiology.

[24]  Forest Rohwer,et al.  Metagenomic and stable isotopic analyses of modern freshwater microbialites in Cuatro Ciénegas, Mexico. , 2009, Environmental microbiology.

[25]  D. Toney,et al.  Identification of a Novel Astrovirus (Astrovirus VA1) Associated with an Outbreak of Acute Gastroenteritis , 2009, Journal of Virology.

[26]  Florent E. Angly,et al.  The Marine Viromes of Four Oceanic Regions , 2006, PLoS biology.

[27]  D. Golino,et al.  Deep sequencing analysis of RNAs from a grapevine showing Syrah decline symptoms reveals a multiple virus infection that includes a novel virus. , 2009, Virology.

[28]  J. Kawai,et al.  Direct Metagenomic Detection of Viral Pathogens in Nasal and Fecal Specimens Using an Unbiased High-Throughput Sequencing Approach , 2009, PloS one.

[29]  J. Maniloff,et al.  Virus taxonomy : eighth report of the International Committee on Taxonomy of Viruses , 2005 .

[30]  T. Candresse,et al.  trategies for simultaneous detection of multiple plant viruses , 2006 .

[31]  Benjamin J. Raphael,et al.  The Sorcerer II Global Ocean Sampling Expedition: Expanding the Universe of Protein Families , 2007, PLoS biology.

[32]  M. Breitbart,et al.  Discovery of a Novel Single-Stranded DNA Virus from a Sea Turtle Fibropapilloma by Using Viral Metagenomics , 2008, Journal of Virology.

[33]  W. Ian Lipkin,et al.  Newly Discovered Ebola Virus Associated with Hemorrhagic Fever Outbreak in Uganda , 2008, PLoS pathogens.

[34]  P. Susi.  Black currant reversion virus, a mite-transmitted nepovirus. , 2004, Molecular plant pathology.

[35]  David L. Wheeler,et al.  GenBank , 2015, Nucleic Acids Res..

[36]  O. White,et al.  Environmental Genome Shotgun Sequencing of the Sargasso Sea , 2004, Science.

[37]  Joel Dudley,et al.  MEGA: A biologist-centric software for evolutionary analysis of DNA and protein sequences , 2008, Briefings Bioinform..

[38]  N. Boonham,et al.  Microarrays for rapid identification of plant viruses. , 2007, Annual review of phytopathology.

[39]  E. Delwart,et al.  Metagenomic Analyses of Viruses in Stool Samples from Children with Acute Flaccid Paralysis , 2009, Journal of Virology.

[40]  R. Shafer,et al.  A highly prevalent and genetically diversified Picornaviridae genus in South Asian children , 2008, Proceedings of the National Academy of Sciences.

[41]  M. Crowell,et al.  Human gut microbiota in obesity and after gastric bypass , 2009, Proceedings of the National Academy of Sciences.

[42]  Susan M. Huse,et al.  Microbial diversity in the deep sea and the underexplored “rare biosphere” , 2006, Proceedings of the National Academy of Sciences.