Metagenomics

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Metagenomics (also Environmental Genomics, Ecogenomics or Community Genomics) is the study of genomes recovered from environmental samples as opposed to from clonal cultures. This relatively new field of genetic research allows the genomic study of organisms that are not easily cultured in a laboratory. Early molecular work in the field was conducted by Norm Pace and colleagues, who used PCR to explore the diversity of ribosomal RNA sequences from organisms present in uncultured environmental samples. Considerable efforts ensured that these were not PCR false positives and supported the existence of a complex community of unexplored species. Although this methodology was limited to exploring highly conserved, non-protein coding genes, it did support early microbial morphology-based observations that diversity was far more complex than was known by culturing methods.

The insights gained from these breakthrough studies led Pace to propose the idea of cloning DNA directly from environmental samples as early as 1985 (ASM News 51:4). This led to the first report of isolating and cloning bulk DNA from an environmental sample, published by Pace and colleagues in 1991 (J. Bacteriol. 173: 4371) while Pace was in the Department of Biology at Indiana University. Soon after that, Healy reported the metagenomic isolation of functional genes from "zoolibraries" constructed from a complex culture of environmental organisms grown in the laboratory on dried grasses in 1995 (Appl. Microbiol. Biotechnol. 43: 667). After leaving the Pace laboratory, Ed DeLong continued in the field and has published work that has largely laid the groundwork for contemporary metagenomics based on signature 16S sequences, beginning with his group's construction of libraries from marine samples (1996 J. Bacteriol. 178:591) in collaboration with scientists from Diversa (formerly Recombinant BioCatalysis, Inc.). Construction of libraries from soil DNA faced many technical challenges before the first successes were reported from the laboratories of Radomski (USPN 5,849.491 filed in 1995), Handelsman, Pernodet, and Gottschalk.

Metagenomics for isolation of expressed gene and gene pathways products (functional screening of libraries derived directly from environmental samples, without culturing) was first conceived and practiced by Jay M. Short, head of research and a founder of Diversa Corporation, at least as early as 1994. Short published articles, and filed and was issued many U.S. patents on the discovery of genes, gene pathways and their products from gene libraries made from communities of uncultured organisms, using a variety of functional and sequence based methods for screening. Following earlier patent filings, one of the earlier publications related to Short’s inventions appeared in SIM News (46:3-8) in 1996 and was titled “The Discovery of New Biocatalysts from Microbial Diversity”. There are over a dozen publications and patents between 1996 and 2005. This work by Short and coworkers extended substantially the analysis beyond 16S sequences and was a precursor to functional genome-wide analysis such as that appearing in 2005 in Science (308:554-557) entitled “Comparative Metagenomics of Microbial Communities”.

The term "metagenomics" was first used by Jo Handelsman and others in the University of Wisconsin Department of Plant Pathology, and appeared in publication in 1998 (see references below). Kevin Chen and Lior Pachter (researchers at the University of California, Berkeley) defined metagenomics as "the application of modern genomics techniques to the study of communities of microbial organisms directly in their natural environments, bypassing the need for isolation and lab cultivation of individual species."

1 Sequences from environmental samples
2 Community metabolism
3 References
4 External links

[edit] Sequences from environmental samples

Conventional sequencing begins with a culture of identical cells as a source of DNA. However early metagenomic studies revealed that there are probably large groups of microorganisms in many environments that cannot be cultured and thus cannot be sequenced. These early studies focused on 16S ribosomal RNA sequences which are relatively short, often conserved within a species, and generally different between species. Many 16S rRNA sequences have been found which do not belong to any known cultured species, indicating that there are numerous unisolated organisms out there.

Recovery of DNA sequences longer than a few thousand base pairs from environmental samples was very difficult until recent advances in molecular biological techniques, particularly related to constructing libraries in bacterial artificial chromosomes (BACs), provided better vectors for molecular cloning. In addition, advances in bioinformatics, refinements of DNA amplification, and proliferation of computational power have greatly aided the analysis of DNA sequences recovered from environmental samples by metagenomics. In 2002, Mya Breitbart, Forest Rohwer, and collegues used metagenomic to show that 200 liters of seawater contains over 5000 different viruses. Subsequent studies showed that there are >1000 viral species in human stool and possibly a million different viruses per kilogram of marine sediment. Essentially all of the viruses in these studies were new species. A 2004 metagenomic study of the Sargasso Sea found DNA from nearly 2000 different species including 148 types of bacteria never seen before. Another study, also from 2004, revealed the genomes of bacteria and archaea from an acid mine drainage system that had resisted attempts to culture them.

Because the collection of DNA from an environment is largely uncontrolled, large samples, often sometimes prohibitively so, are needed to fully resolve the genomes of underrepresented members of a microbial community. On the other hand, many such underrepresented organisms might never be noticed without metagenomic analysis if they are difficult to isolate using traditional culturing techniques.

[edit] Community metabolism

Many bacterial communities show significant division of labor in metabolism. Waste products of some organisms are metabolites for others. Working together they turn raw resources into fully metabolised waste. Using comparative gene studies and expression experiments with microarrays or proteomics researchers can piece together a metabolic network that goes beyond species boundaries. Such studies require detailed knowledge about which versions of which proteins are coded by which species and even by which strains of which species. So, community genomic information is fundamental to the study of how metabolites move through a community to be processed.

[edit] References

Chen K, Pachter L (2005) Bioinformatics for whole-genome shotgun sequencing of microbial communities. PLoS Comp Biol 1(2): e24.
Breitbart et al. (2004). Diversity and population structure of a nearshore marine sediment viral community. Proceedings of the Royal Society B 271: 565-574.
Handelsman J. (2004). Metagenomics: application of genomics to uncultured microorganisms. Microbiology and Molecular Biology Reviews 68:669-685.
Tyson et al. (2004). Insights into community structure and metabolism by reconstruction of microbial genomes from the environment. Nature 428:37-43.
Venter et al. (2004). Environmental Genome Shotgun Sequencing of the Sargasso Sea. Science 304:66-74.
Rodriguez-Valera. (2004). Environmental genomics, the big picture?. FEMS Microbiology Letters 231:153-158.
Diversa Corporation (2004). Diversa Scientists First to Link Protein Function to Genetic Sequence Information Derived from Complex Microbial Communities. April 21st Press Release.
Breitbart et al. (2003). Metagenomic analyses of an uncultured viral community from human feces. Journal of Bacteriology 85:6220-6223.
Diversa Corporation (2003). Diversa and Department of Energy’s Joint Genome Institute Announce Large-scale Microbial Sequencing Collaboration. May 5th Press Release.
Breitbart et al. (2002). Genomic analysis of uncultured marine viral communities. Proceedings of the National Academy USA 99:14250-14255.
Handelsman et al. (1998). Molecular biological access to the chemistry of unknown soil microbes: a new frontier for natural products. Chemistry Biology 5:R245-R249.
Short, JM (2001) Sequence Based Screening. U.S. Patent Number 6,455,245. (Diversa Corporation)
Short, JM (1997) Screening Methods for Enzymes and Enzyme Kits. U.S. Patent Number 6,168,919. (Diversa Corporation)
Short, JM (1997) Protein Activity Screening of clones having DNA from Uncultured Microorganisms. U.S. Patent Number 5,958,672. (Diversa Corporation)
Short, JM (1997) Production of Enzymes Having Desired Activities by Mutagenesis. U.S. Patent Number 5,939,250. (Diversa Corporation)
Short, JM (1997) Recombinant Approaches for Accessing Biodiversity. Nature Biotechnology 15:1322-1323
Short, JM (2001) Gene Expression Library Produced from DNA from Uncultivated Microorganisms and Methods for Making the Same. U.S. Patent Number 6,280,926 (Diversa Corporation)
Short, JM et al. (1999) Production and use of normalized DNA libraries. U.S. Patent Number 6,001,574 (Diversa Corporation)
Short, JM (2003) Protein activity screening of clones having DNA from uncultivated microorganisms. U.S. Patent Number 6,528,249 (Diversa Corporation)
Short, JM (2004) Protein activity screening of clones having DNA from uncultivated microorganisms. U.S. Patent Number 6,677,115 (Diversa Corporation)
Abulencia, C. B., Wyborski, D. L., Garcia, J. A., Podar, M., Chen, W., Chang, S. H. et al. (2006). Environmental whole-genome amplification to access microbial populations in contaminated sediments. Applied and Environmental Microbiology 72[5], 3291-3301. (Diversa Corporation)
Green, B. D. & Keller, M. (2006). Capturing the uncultivated majority. Current Opinion in Biotechnology 17[3], 236-240. (Diversa Corporation)
Keller, M. & Sengler, K. (2004). Tapping into microbial diversity. Nature Reviews Microbiology 2[2], 141-150. (Diversa Corporation)
Mathur, E., Toledo, G., Green, B. D., Podar, M., Richardson, T. H., Kulwiec (2005). A biodiversity-based approach to development of performance enzymes: Applied metagenomics and directed evolution. Industrial Biotechnology, 1, 283-287. (Diversa Corporation)
Zengler, K., Toledo, G., Rappe, M., Elkins, J., Mathur, E. J., Short, J. M. (2002). Cultivating the uncultured. Proceedings of the National Academy of Sciences of the United States of America 99[24], 15681-15686. (Diversa Corporation)
Zengler, K., Paradkar, A., & Keller, M. (2005). New methods to access microbial diversity for small molecule discovery. Natural Products , 275-293. (Diversa Corporation)