Shannon J. Williamson

From Bacterial to Microbial Ecosystems (Metagenomics)

Metagenomics is revolutionizing the field of microbial ecology through techniques that eliminate the prerequisite of culturing. Metagenomic studies of microbial populations in different environments reveal the incredible diversity and adaptive capabilities of these organisms. With the advent of cheaper, high-throughput sequencing technologies, these studies are also producing vast amounts of sequence data. Here, we discuss the different components of a metagenomic study including sample collection, DNA extraction, sequencing, and informatics. We highlight their issues and challenges, and review the solutions that are currently in use. We conclude with examples of metagenomic studies conducted on environments of varying complexities.

Separation of single-stranded DNA, double-stranded DNA and RNA from an environmental viral community using hydroxyapatite chromatography.

Viruses, particularly bacteriophages (phages), are the most numerous biological entities on Earth. Viruses modulate host cell abundance and diversity, contribute to the cycling of nutrients, alter host cell phenotype, and influence the evolution of both host cell and viral communities through the lateral transfer of genes. Numerous studies have highlighted the staggering genetic diversity of viruses and their functional potential in a variety of natural environments. Metagenomic techniques have been used to study the taxonomic diversity and functional potential of complex viral assemblages whose members contain single-stranded DNA (ssDNA), double-stranded DNA (dsDNA) and RNA genotypes. Current library construction protocols used to study environmental DNA-containing or RNA-containing viruses require an initial nuclease treatment in order to remove nontargeted templates. However, a comprehensive understanding of the collective gene complement of the virus community and virus diversity requires knowledge of all members regardless of genome composition. Fractionation of purified nucleic acid subtypes provides an effective mechanism by which to study viral assemblages without sacrificing a subset of the communitys genetic signature. Hydroxyapatite, a crystalline form of calcium phosphate, has been employed in the separation of nucleic acids, as well as proteins and microbes, since the 1960s. By exploiting the charge interaction between the positively-charged Ca(2+) ions of the hydroxyapatite and the negatively charged phosphate backbone of the nucleic acid subtypes, it is possible to preferentially elute each nucleic acid subtype independent of the others. We recently employed this strategy to independently fractionate the genomes of ssDNA, dsDNA and RNA-containing viruses in preparation of DNA sequencing. Here, we present a method for the fractionation and recovery of ssDNA, dsDNA and RNA viral nucleic acids from mixed viral assemblages using hydroxyapatite chromatography.

EXPLORING THE OCEAN'S MICROBES: SEQUENCING THE SEVEN SEAS

Marvin E. Frazier,Douglas B. Rusch, Aaron L. Halpern, Karla B. Heidelberg, Granger Sutton, Shannon Williamson, Shibu Yooseph, Dongying Wu, Jonathan A. Eisen, Jeff Hoffman, Charles H. Howard, Cyrus Foote, Brooke A. Dill, Karin Remington, Karen Beeson, Bao Tran, Hamilton Smith, Holly Baden-Tillson, Clare Stewart, Joyce Thorpe, Jason Freemen, Cindy Pfannkoch, Joseph E. Venter, John Heidelberg, Terry Utterback, Yu-Hui Rogers, Shaojie Zhang, Vineet Bafna, Luisa Falcon, Valeria Souza,German Bonilla, Luis E. Eguiarte , David M. Karl, Ken Nealson, Shubha Sathyendranath, Trevor Platt, Eldredge Bermingham, Victor Gallardo, Giselle Tamayo, Robert Friedman, Robert Strausberg, J. Craig Venter 1 J. Craig Venter Institute, Rockville, Maryland, United States Of America 2 The Institute For Genomic Research, Rockville, Maryland, United States Of America 3 Department of Computer Science, University of California San Diego 4 Instituto de Ecologia Dept. Ecologia Evolutiva, National Autonomous University of Mexico Mexico City, 04510 Distrito Federal, Mexico 5 University of Hawaii, Honolulu, United States of America 6 Dept. of Earth Sciences, University of Southern California, Los Angeles, California, United States of America 7 Dalhousie University, Halifax, Nova Scotia, Canada 8 Smithsonian Tropical Research Institute, Balboa, Ancon, Republic of Panama 9 University of Concepcion, Concepcion, Chile 10 University of Costa Rica, San Pedro, San Jose, Republic of Costa Rica