Aina Maria Mas Calafell Genomics

METAGENOMICS

Introduction

Microbial communities Primary role in biogeochemical systems

Study of microbial communities

1.- Culture-based methodologies

Only isolated microbes

Phenotypic, biochemical and physiological tests Molecular techniques

o G+C content o DNA-DNA pairing o Targeted sequencing

(Rosselló-Mora & Amann, 2001)

Study of microbial communities 2.- Culture-independent

DNA sequence-based methodologies

Metagenomics

‘Culture-independent and genomic analysis of

microbial communities’

(Tringe & Rubin, 2005)

Low-throughput targeted sequencing

Well-conserved Large enough No horizontal transfer Molecular clock

Sequencing of 16S rRNA genes

2.1.- Sanger sequencing

(Janda & Abott, 2007)

Community structure

2.1.- Sanger sequencing

Low-throughput shotgun sequencing

(Breitbart et al., 2002)

A) Number of sequences from the uncultured shotgun libraries with a significant hit (E < 0.001) to GenBank

Functionallity

Community structure

2.1.- Sanger sequencing

Cost Prohibitive to sequence highly complex communities

Certain microbial and bacteriophague sequences are not carried

stably by E.coli

Study of microbial communities 2.- Culture-independent

DNA sequence-based methodologies

2.2.- SGS and TGS

Higher troughput Lower cost Faster In vitro amplification (SGS) or

lack of amplification (TGS)

(Mardis, 2008)

Whole-metagenome shotgun

Sequence of thousands of organisms at the

same time

Detection of very low abundance members

Increased taxonomic

resolution (Petrosino et al., 2009)

High-throughput shotgun sequencing

2.2.- SGS and TGS

High-throughput targeted

sequencing

Expansion of microbiome studies

(Segata et al., 2013)

2.2.- SGS and TGS

High-throughput shotgun sequencing

(Mardis, 2008)

Comparison of technologies (Luo et al., 2012)

(Barzon et al., 2011)

2.2.- SGS and TGS

High-throughput shotgun sequencing

(Bragg & Tyson 2014)

New view Metagenomics

Human microbiome

Soil

Sea

Deep mine

Honey bee colonies

(Fierer et al., 2007)

(Venter et al., 2004)

Human microbiome

Human microbiome

Humans ~ Superorganisms composed of human and microbial components

(Qin et al., 2010)

(Bäckhed et al., 2006)

Human Microbiome Project

‘To understand the microbial components of our genetic and metabolic landscape’ ‘To highlight how they contribute to our normal physiology and disease predisposition’

(NIH HMP Working group, 2009)

(Turnbaugh et al., 2007)

Human Microbiome Project

(The Human Microbiome Jumpstart Reference Strains Consortium, 2010)

(Cuomo et al., 2014)

(The Human Microbiome Project Consortium, 2012)

Human Microbiome

Future insights

Deeper coverage of the microbiome

Effects of age and

diet?

Effects of pathologic states?

Effect of different

living environments

Human Microbiome

New biomarkers for

defining our health

New ways for optimizing our personal nutrition

New ways to forecast our

predispositions to disorders

Future insights

Deeper coverage of the microbiome

Effects of age and

diet?

Effects of pathologic states?

Effect of different

living environments

A wide view of microbial communities

Meta’omics

(Segata et al., 2013)

Conclusions

The understanding of complex microbial communities can have several applications.

DNA sequence-based methods circumvented the obstacles of culture-based approaches.

Next-generation sequencing technologies have expanded the experimental tools available for studying microbiomes.

The current bottleneck is the downstream analysis of data.

Metagenomic studies are ongoing in several important ecosystems, specially the human body in health and disease.

Metagenomics is complemeted with different approaches to have an overall view of microbial communities, including their functionallity.

References Bäckhed et al. «Host-bacterial mutualism in the human intestine.» Science, 2006 Mar 25; 307(5717):1915-20.

Barzon et al. «Applications of Next-Generation Sequencing Technologies to.» Int. J. Mol. Sci., 2011; 12, 7861-7884.

Bragg andTyson. «Metagenomics using Next-Generation Sequencing.» Methods Mol Biol, 2014; 1096:183-201.

Breitbart et al. «Genomic analysis of uncultured marine viral communities.» Proceedings of the National Academy of Science,

2002 Oct 29; 99(22):14250-5. Cuomo, y otros. «Genome Sequence of the Pathogenic Fungus Sporothrix schenckii (ATCC 58251).» Genome Announc., 201;

May 22;2(3). Fierrer, y otros. «Metagenomic and small-subunit rRNA analyses of the genetic diversity of bacteria, archaea, fungi and

viruses in soil.» Appl Environ Microbiol, 2007; Nov; 73(21):7059-66. Janda, y Abbott. «16S rRNA Gene Sequencing for Bacterial Identification in the Diagnostic Laboratory: Pluses, Perils, and

Pitfalls.» J Clin Microbiol, 2007; Sep 45(9): 2761–2764.

Luo, Tsementzi, Kyrpides, Read, y Konstantinidis. «Direct Comparisons of Illumina vs. Roche 454 Sequencing Technologies on

the Same Microbial Community DNA Sample.» PLoS One, 2012; Feb 7(2).

Mardis. «The impact of next-generation sequencing technology on genetics.» Trends Genet, 2008; Mar; 24(3):133-41.

Petrosino, Highlander, Luna, Gibbs, y Versalovic. «Metagenomic Pyrosequencing and Microbial Identification.» Clin Chem, 2009: May 55(5):856-66.

Qin et. al «A human gut microbial gene catalogue established by metagenomic sequencing.» Nature, 2010; 464, 59-65.

Rosselló-Mora, y Amann. «The species concept for prokaryotes.» FEMS Microbiology reviews 25, 2001: 39-67.

Segata, Boernigen, Tickle, Morgan, Garrett, y Hutternhower. «Computational meta'omics for microbial community studies.»

Molecular Systems Biology, 2013: 9:666.

The Human Microbiome Consortium Jumpstart Reference Strains. «A catalog of reference genomes from the human

microbiome.» Science, 2010; May 21; 328(5981):994-9. Tringe, y Rubin. «Metagenomics: DNA sequencing of environmental samples.» Nature Reviews Genetics, 2005: Nov 6(11): 805-

14.

Turnbaugh, Ley, Hamady, Fraser-Liggett, y Knight. «The human microbiome project: exploring the microbial part of ourselves in a changing world.» Nature , 2007: Oct 449(7164): 804–810.

Venter, y otros. «Environmental Genome Shotgun Sequencing of the Sargasso Sea.» Science, 2004: April Vol. 304 no. 5667 pp. 66-74.