Use of the genomic data

o Reconstruction of metabolic properties

o Nature’s Microbiome

o NGS in Population Genetics

Why are the diatoms so successful?

canonical urea metabolism in animals

duplication of CPS in secondary algae!

Diatoms have somehow reversed urea cycle, which server as a hub for the metabolism of the nitrogen and (to some extent) also the carbon!

Nature’s Microbiome

hologenome

Non specific in situ culture dependent approaches vs Metagenomics

Community composition16S rDNA marker95%, 97%, or 99% for

OTUs/phylotypeOperational Taxonomic Unitbinning

set of abundance histograms or set of binary presence/absence

vectors

Nature’s Microbiome

Community function

Decomposition methods: Principal Component AnalysesCanonical Correlation Analyses

Nature’s Microbiome

the number (richness) and distribution (evenness) of taxa expected within a single populationAlpha Diversity

Beta Diversity

‘‘How much do I need to sequence to completely characterize my microbiome?’’

absolute or relative overlap describe how many taxa are shared between samples

Nature’s Microbiome

Community function

Community composition16S rDNA marker95%, 97%, or 99% for

OTUs/phylotypeOperational Taxonomic Unitbinning

set of abundance histograms or set of binary presence/absence

vectorsDecomposition methods:

Principal Component AnalysesCanonical Correlation Analyses

How many microbial strains and species colonize eukaryotic hosts?

Alpha diversity inflated due to PCR and sequencing errorsX

Diversity involving closely related microbes omitted in NGS data sets (OTUs cutt-off)

Nature’s Microbiome: Chalanges

Raw data „denoising“ Detecting hidden strain diversity

AnalysesSequencing tools (e.g.LEA-Seq)

How many microbial strains and species colonize eukaryotic hosts?Low Error Amplicon Sequencing, LEA-Seq

Nature’s Microbiome:Chalanges

Dilute barcoded primer solution

20x coverage

How to detect sequence libraries plagued by contamination?

Nature’s Microbiome: Chalanges

Results from the source-tracking analysis using a previously published set of mammal gut samples, as well as soil samples, as potential sources to determine which of the 92 samples were potentially contaminated with soil.

Representative source data sets

>10% soil assignment<0.01 mammalian gut

92/27(5)

Negative controlsGeneration of source libraries

marine/fresh watercommon lab microbes

Number of biological replicates

Nature’s Microbiome: Outcomes

NGS and Population Genetics

Problem of genotyping uncertainty on NGS data

machine learning to separate true segregating variation from artifacts

• Sequencing errors• Misalignment errors• Low coverage• Random sampling of

homologous base pairs in heterozygotes

initial read mappinglocal realignment around indelsbase quality score recalibration

SNP discovery and genotyping to find all potential variants

NGS and Population Genetics

Problem of genotyping uncertainty on NGS datainitial read mappinglocal realignment around indelsbase quality score recalibration

Gap enabled alignment with Burrows-Wheeler transform: >15% reads spanning homozygous indels misalgned

NGS and Population Genetics

Problem of genotyping uncertainty on NGS data• Sequencing errors• Misalignment errors• Low coverage• Random sampling of

homologous base pairs in heterozygotes

NGS and Population Genetics: Outcomes