WWW.ABSTRACT.DISLIB.INFO
FREE ELECTRONIC LIBRARY - Abstracts, online materials
 
<< HOME
CONTACTS



Pages:   || 2 | 3 | 4 |

«Annu. Rev. Genet. 2004. 38:525–52 doi: 10.1146/annurev.genet.38.072902.091216 Copyright c 2004 by Annual Reviews. All rights reserved First ...»

-- [ Page 1 ] --

Annu. Rev. Genet. 2004. 38:525–52

doi: 10.1146/annurev.genet.38.072902.091216

Copyright c 2004 by Annual Reviews. All rights reserved

First published online as a Review in Advance on July 14, 2004

METAGENOMICS: Genomic Analysis

of Microbial Communities

Christian S. Riesenfeld,1,2 Patrick D. Schloss,1

and Jo Handelsman1,2

Department of Plant Pathology,1 Microbiology Doctoral Training Program,2 University of

Wisconsin-Madison, Madison, Wisconsin 53706; email: joh@plantpath.wisc.edu

Key Words microbial ecology, environmental genomics, community genomics, culture-independent, and unculturable bacteria I

Abstract

Uncultured microorganisms comprise the majority of the planet’s biological diversity. Microorganisms represent two of the three domains of life and contain vast diversity that is the product of an estimated 3.8 billion years of evolution. In many environments, as many as 99% of the microorganisms cannot be cultured by standard techniques, and the uncultured fraction includes diverse organisms that are only distantly related to the cultured ones. Therefore, culture-independent methods are essential to understand the genetic diversity, population structure, and ecological roles of the majority of microorganisms. Metagenomics, or the culture-independent genomic analysis of an assemblage of microorganisms, has potential to answer fundamental questions in microbial ecology. This review describes progress toward understanding the biology of uncultured Bacteria, Archaea, and viruses through metagenomic analyses.

CONTENTS INTRODUCTION..................................................... 526 METAGENOMICS DEFINED........................................... 527 LINKING PHYLOGENY AND FUNCTION WITHIN SPECIES............... 529 Phylogenetic Anchors................................................ 529 Function Then Phylogeny............................................. 529 Phylogeny Then Function............................................. 533 Acidobacterium Phylogeny and Function................................. 533 Archaeal Phylogeny and Function....................................

–  –  –

INTRODUCTION

Obtaining bacteria in pure culture is typically the first step in investigating bacterial processes. However, standard culturing techniques account for 1% or less of the bacterial diversity in most environmental samples (2). Although some significant breakthroughs have resulted from recent attempts to culture the as-yet-unculturable bacteria (56, 89, 99, 127), a suite of culture-independent techniques are needed to complement efforts to culture the thousands or millions of unknown species in the environment.

A new era of microbial ecology was initiated when sequencing of ribosomal RNAs and the genes encoding them was introduced to describe uncultured bacteria in the environment. The first approach was to sequence clones from a 5S rRNA cDNA library derived from the symbiotic community within the tubeworm Riftia pachyptila (109). Variations of this method generated a set of culture-independent techniques to (a) reconstruct phylogenies, (b) compare microbial distributions among samples using either nucleotide sequence or restriction fragment length polymorphisms (RFLPs), and (c) quantify the relative abundance of each taxonomic group using membrane hybridization or fluorescent in situ hybridization (2, 47, 57, 78–80).

The most startling result of the many microbial diversity studies that have employed 16S rRNA culture-independent methods is the richness of the uncultured microbial world. As of April 1, 2004, GenBank contained 21,466 16S rRNA genes from cultured prokaryotes and 54,655 from uncultured prokaryotes, according to the search terms described by Rapp´ & Giovannoni (90), and many of those from e uncultured organisms affiliate with phyla that contain no cultured members. When Woese (121) originally proposed a 16S rRNA-based phylogeny, 12 bacterial phyla were recognized, each with cultured representatives. Since then, 14 additional phyla with cultured representatives have been identified. In addition, 16S rRNA gene sequence analysis suggests 26 candidate phyla that have no known cultured representatives (90). Therefore, half of the known microbial phyla have no cultured representatives.

Among the phyla that contain cultured members, a few contain many isolates and the rest contain too few to represent the full spectrum of diversity in the phylum. For example, Hugenholtz (53) found that 97% of prokaryotes deposited in the Australian Culture of Microorganisms in 2001 were members of just four phyla: the Proteobacteria (54%), Actinobacteria (23%), Firmicutes (14%), and

METAGENOMICS

Bacteroidetes (6%). Within GenBank, 76% of the 16S rRNA gene sequences of cultured prokaryotes are from these four groups. But other phyla may be more diverse, prevalent, and ecologically consequential in the environment. 16S rRNA gene sequences from the Acidobacterium phylum are among the most abundant in clone libraries obtained from soil and have been found in all soils examined, suggesting that the Acidobacteria play important roles in soil ecosystems. However, of the 684 Acidobacterium 16S rRNA gene sequences in GenBank, only 19 (2.8%) are from cultured isolates, providing an inadequate collection to describe the physiological diversity of the phylum. Other than 16S rRNA gene sequences, little is known about the bacteria within the 22 poorly cultured phyla and 26 candidate phyla. Many terms, such as unculturable, uncultivated, as yet uncultured, and not yet cultured, are used to refer to microorganisms that we know of only through culture-independent means. In this review, we refer to them as uncultured.





Describing the phylogenetic diversity of uncultured microorganisms is only the first step. A greater challenge is to assign ecological roles to them. The uncultured microbiota must play pivotal roles in natural environmental processes and are a large untapped resource for biotechnology applications. Exploiting the rich microbial biodiversity for enzyme and natural product discovery is an active research area that has been reviewed elsewhere (39, 45, 46, 65, 66, 77, 97, 104). This review discusses the application of culture-independent genomics-based approaches to understand the genetic diversity, population structure, and ecology of complex microbial assemblages (26, 93, 94).

METAGENOMICS DEFINED

“Metagenomics” describes the functional and sequence-based analysis of the collective microbial genomes contained in an environmental sample (Figure 1) (45).

Other terms have been used to describe the same method, including environmental DNA libraries (110), zoolibraries (55), soil DNA libraries (68), eDNA libraries (13), recombinant environmental libraries (22), whole genome treasures (77), community genome (114), whole genome shotgun sequencing (115), and probably others. In this review, we use metagenomics to describe work that has been presented with all of these names because it is the most commonly used term (15, 27, 35, 59–61, 65, 66, 82, 105, 107, 117, 118), was used for the title of the first international conference on the topic (“Metagenomics 2003” held in Darmstadt, Germany), and is the focus of an upcoming issue of the journal Environmental Microbiology. The definition applied here excludes studies that use PCR to amplify gene cassettes (52) or random PCR primers to access genes of interest (17, 32), since these methods do not provide genomic information beyond the genes that are amplified. Many environments have been the focus of metagenomics, including soil, the oral cavity, feces, and aquatic habitats, as well as the hospital metagenome, a term intended to encompass the genetic potential of organisms in hospitals that conribute to public health concerns such as antibiotic resistance and nosocomial infections (20).

528 RIESENFELD SCHLOSS HANDELSMAN

Figure 1 Metagenomics involves constructing a DNA library from an environment’s microbial population and then analyzing the functions and sequences in the library.

The concept of cloning DNA directly from an environment was initially suggested by Pace (79) and first implemented by Schmidt et al. (106), who constructed a λ phage library from a seawater sample and screened it for 16S rRNA genes.

Advances by the DeLong group in cloning DNA directly from seawater provided the landmark work that launched the field (110). Development of metagenomic analyses of soil was slower than with seawater because of the technical challenges of cloning DNA from the complex matrix of soil, which contains many compounds that bind to DNA or inhibit the enzymatic reactions required for cloning. Significant progress has been made, producing libraries that have substantially advanced understanding the functions in the soil community (96). The past eight years have witnessed an explosion of interest and activity in metagenomics, accompanied by advances in technology that have facilitated studies at a scale that was not feasible when the field began. For example, the seminal paper in 1996 by Stein et al. (110) reported the sequencing and reconstruction of a 40-kb fragment from an uncultured marine archaeon, which was a major undertaking at the time. In 2004, Venter et al.

(115) reported their attempt to sequence the entire metagenome of the Sargasso Sea by obtaining over 1 million kb of nonredundant sequence. The advances in sequencing technology have expanded the approaches and questions that can be

METAGENOMICS

considered with metagenomics, providing access to a staggering amount of genomic information. Metagenomic technology has been successful at all scales—it has been used to study single genes (e.g., cellulases, 48), pathways (e.g., antibiotic synthesis, 96), organisms (e.g., Archaea, 110), and communities (e.g., acid mine drainage biofilm, 114). Approaches that involve massive sequencing to capture entire communities will likely become more common with further advances in sequencing technology.

LINKING PHYLOGENY AND FUNCTION

WITHIN SPECIES

Phylogenetic Anchors The first metagenomic studies aimed to link a function with its phylogenetic source, providing information about one species within a community. One of the challenges with this approach is to link a phenotype with the identity of the original host. Three approaches have been taken: Screen a metagenomic library for a phenotype and then attempt to determine the phylogenetic origin of the cloned DNA (Table 1), screen clones for a specific phylogenetic anchor (e.g., 16S rRNA) or gene and then sequence the entire clone and search for genes of interest among the genes flanking the anchor (Table 2), or sequence the entire metagenome and identify interesting genes and phylogenetic anchors in the resulting reconstructed genomes (Table 3).

Function Then Phylogeny Diverse activities have been discovered by functional analysis of metagenomic libraries. New antibiotics (11–14, 36, 68, 96, 119, 120), hydrolytic and degradative enzymes (21, 48–50, 59, 60, 91, 96, 117), biosynthetic functions (31, 61), antibiotic resistance enzymes (22, 92), and membrane proteins (69) have been identified. The diversity of functionally active clones discovered in metagenomic libraries validates the use of functional screens as one means to characterize the libraries. Antimicrobial screens have revealed new antibiotics such as terragine (119), turbomycin A and B (36), and acyl tyrosines (13), as well as previously described antibiotics such as indirubin (68) and violacein (12). Most of these compounds are structurally based on common cell substituents, such as amino acids, and none requires more than a few genes for its synthesis. The goal of identifying new polyketide, macrolide, and peptide antibiotics (45) may require different methods. Enhancing expression of genes in metagenomic libraries may lead to discovery of a wider array of natural products. This will be accomplished by moving the libraries into alternative hosts, such as Streptomyces, which was the basis for discovery of terragine (119). Alternative hosts may enhance gene expression or provide starting materials that Escherichia coli does not contain. E. coli can be engineered to express a wider range of functions by introducing genes encoding new sigma factors, rare tRNAs, or functions required to synthesize starting materials TABLE 1 Metagenomics discovery based on functional screens

–  –  –

for antibiotic biosynthesis that are deficient in E. coli. Alternatively, sequences that carry conserved regions of genes associated with antibiotic biosynthesis, such as the polyketide synthases and peptide synthetases, may be identified by sequencedbased screens that do not require heterologous gene expression. This approach successfully identified clones carrying a novel hybrid polyketide synthase-peptide synthetase gene cluster from an uncultured bacterial symbiont of a beetle (81).

Novel enzymes have been revealed in metagenomic libraries by screening clones directly for activity (49, 50, 96). Pigments have been identified by visual inspection (12, 36, 68). These methods require handling individual clones, usually in an array format. Because the frequency of active clones is low, high-throughput methods are essential for efficient screening. Selection for the ability to grow on hydroxyl-butyrate as the sole carbon and nitrogen source provided a powerful selection for clones carrying new degradative enzymes (49), and selection for antibiotic resistance identified new antibiotic resistance determinants from soil (22,

92) and from oral flora (27).



Pages:   || 2 | 3 | 4 |


Similar works:

«THÈSE En vue de l'obtention du DOCTORAT DE L’UNIVERSITÉ DE TOULOUSE Délivré par l'Université Toulouse III Paul Sabatier Discipline ou spécialité : Ecologie Comportementale Présentée et soutenue par Jérémie CORNUAU Le 04-12-12 Titre : Signaux multiples et choix du partenaire chez le triton palmé Lissotriton helveticus JURY Thierry Lengagne (Chargé de recherche, Rapporteur) Bruno Faivre (Professeur, Rapporteur) Claude Miaud (Directeur EPHE, Examinateur) Christophe Thébaud...»

«Working Paper WP 10-6 September 2010 Harnessing the Power of Technology to Enhance Financial Literacy Education and Personal Financial Well-Being: A Review of the Literature, Proposed Model, and Action Agenda Wendy L. Way and Nancy Wong Center for Financial Security WP 10-6 Harnessing the Power of Technology to Enhance Financial Literacy Education and Personal Financial Well-Being: A Review of the Literature, Proposed Model, and Action Agenda Wendy L. Way, Professor and Associate Dean School of...»

«Smart State smart fishing Ecological assessment of Queensland’s East Coast Pearl Fishery A report to the Australian Government Department of Environment and Heritage on the ecologically sustainable management of a highly selective dive fishery Brooke Young Department of Primary Industries and Fisheries With contributions from Phil Gaffney, Malcolm Dunning, Kerrod Beattie, Shannon Ryan and Jeff Bibby DPI&F September 2004 INTRODUCTION In 1992, Australian Commonwealth, State, Territory and local...»

«SPECIES ACTION PLANS SONG THRUSH (Turdus philomelos) DESCRIPTION The song thrush is slightly smaller than a blackbird, with characteristic speckles on the breast. It can be distinguished from the mistle thrush by its smaller size, warm brown back as opposed to grey brow n and the habit of singing from more concealed locations. The mistle thrush prefers the highest most prominent position available. General Ecology A bird of woodland, that has adapted to suburban gardens, parks and hedgerows....»

«ESD REPORT  SERIES No. 5 Pearl Oyster Fishery Ecologically Sustainable Development FRDC – Subprogram Authors: Fletcher, W., Friedman, K., Weir, V., McCrea, J. and Clark, R. Department of Fisheries Western Australian Fisheries and Marine Research Laboratories PO Box 20 North Beach WA 6920 Telephone (08) 9203 0111 Facsimile (08) 9203 0199 Website: http://www.fish.wa.gov.au Published by the Department of Fisheries, Western Australia ESD Report Series No. 5, January 2006 ISSN: 1448...»

«Theory in Action, Vol. 1, No. 4, October 2008 (© 2008) DOI:10.3798/tia.1937-0237.08021 Legitimizing Myth and the Search for Meaning Ali Shehzad Zaidi1 This paper examines the process of mystification that accompanies new paradigms, and discusses such contemporary legitimizing myths as The Myth of the Magic of the Market, The Myth of the Inefficient Public Sector, The Myth of the Objective Media, and The Myth of American Exceptionalism. While these legitimizing myths limit our understanding of...»

«Etnografia de uma caminhada ecológica em meio à paisagem híbrida da ilha1 Ethnography of an ecological walk amidst the hybrid landscape of the island Márcio Antonio Farias de Freitas*1 Palavras-chave: Resumo: Este texto é resultado de minha pesquisa de Áreas protegidas; mestrado, que discute o conflito socioambiental existente no Caminhada; Campinho da Fonte Grande, Vitória-ES, cenário de uma Conflitos ambientais; disputa profundamente desigual em termos de poder político Hibridismo...»

«ATTACHMENT 3 PERSONNEL TRAINING PROGRAM US ECOLOGY MICHIGAN MID 074 259 565 Personnel Training, Revision 10/8/12 EPA ID. No. MI D074259565 FORM EQP5111 ATTACHMENT TEMPLATE MODULE A10 GENERAL INFORMATION: PERSONNEL TRAINING The Administrative Rules for Part 111, Hazardous Waste Management, of the Michigan Natural Resources and Environmental Protection Act, 1994 PA 451, as amended (Act 451), Rule 299.9501 (R 299.9501), R 299.9605, and Title 40 Code of Federal Regulations (40 CFR) 264.16, and...»

«219 Geo-Environment and Landscape Evolution III “Pimp your landscape” – an interactive land-use planning support tool C. Fürst1, C. Davidsson1, K. Pietzsch2, M. Abiy1, M. Volk3, C. Lorz4 & F. Makeschin1 Institute for Soil Science and Site Ecology, TU Dresden, Germany Pietzsch-IT-Service, Markkleeberg, Germany Helmholtz Centre for Env. Research, UFZ, Leipzig, Germany Chair for Landscape Ecology, TU Dresden, Germany Abstract In the context of the INTERREG IIIA project IT-Reg-EU...»

«ECOLOGICAL TANKS, INC. AQUA AIRE ™ Concrete Class I Wastewater Treatment Plants OWNER’S MANUAL Alternate Aerator Locations Aerator Air Supply Line Inlet Outlet Aeration/Mixing Clarifier Chamber Air Drop Line Air Deflection Disc Patent Pending Model AA600 Model AA750 Model AA750T Model AA800 Model AA1200 Model AA1500 “Copyright Notice” No part of this publication may be reproduced, stored in any retrieval system, or transmitted in any form or by any means, electronic, mechanical,...»

«Integrated Pest Management of Quandong Moth control of quandong moth in quandong orchards A report for the Rural Industries Research and Development Corporation by Kaye Ferguson, Department of Applied and Molecular Ecology, University of Adelaide and Peter Bailey, Entomology Section, South Australian Research & Development Institute, Adelaide. December 2001 RIRDC Publication No 01/172 RIRDC Project No SAR 4-A © 2001 Rural Industries Research and Development Corporation. All rights reserved....»

«Instructions for use Long-form paper (Microbial ecology) Molecular monitoring and isolation of previously uncultured bacterial strains from sheep rumen Running title: Isolation of previously uncultured bacteria from rumen S. Koike1*, Y. Handa2, H. Goto1, K. Sakai1, E. Miyagawa2, H. Matsui1, S. Ito3 and Y. Kobayashi1 Research Faculty of Agriculture, Hokkaido University, Sapporo 060-8589 Japan Department of Dairy Science, Rakuno Gakuen University, Ebetsu 069-8501 Japan Creative Research...»





 
<<  HOME   |    CONTACTS
2017 www.abstract.dislib.info - Abstracts, online materials

Materials of this site are available for review, all rights belong to their respective owners.
If you do not agree with the fact that your material is placed on this site, please, email us, we will within 1-2 business days delete him.