ISEEM:Paper DiversityMethods Computational methods for microbial diversity review
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Paper by Eisen et al for PLoS Computational Biology
Outline
Summary and introduction
Organized in part around the metagenomic diversity workflow See Here
A key point is to compare and contrast tools for analyzing rRNA with those for analyzing metagenomic data.
Generation of data
rRNA PCR
- Background
- PCR amplification with conserved primers
- Sequencing
- Standard
- 454
See Media:Margulies_etal_Nature_2005.pdf, Media:Mardis_et_al._Cell_2008.pdf
- Indirect characterization
- ARISA
- T-RFLP
- Array
- Indirect characterization
- Informatics issues
- Primer design
See "Review and re-analysis of domain-specific 16S primers."
- Chimera checking
- Assembly of sequences from paired ends
- rRNA databases
PCR of other genes
- Done sometimes to cover broad diversity
- RecA/RadA best example
- Frequently done to focus on narrower groups or functional classes
- Amanox
- Nitrogen fixation
Metagenomic Sequencing
- Background
- Identifying rRNA genes
- Identifying other genes
- Which genes should be used?
- Assembly
- Metagenomic databases
- CAMERA
- RAST
- IMG/M
General analysis of gene families (rRNA and protein)
rRNA vs. protein
Sequence quality issues
- Chimera checking
Alignment
Dividing into OTUs
- rRNA focused methods
Phylogenetic trees
- Building trees
- Phylogenetic classification
- Alignment independent method
- PhyloPythia - doesn't use tree, SVM at each node
- Phylogenetic structure
- Phylogenetic based OTUs
Web servers that do multiple things
Species Diversity Measurements Focused on Single Communities
Richness estimators
- Reviews
- Parametric
- Non parametric
Alpha diversity
Issues with different sequencing methods
- 454 Sequencing
- See recent paper by R. Knight on methods
Comparing between samples
General approaches and Introduction
- Multiple ways to compare communities and to think about the comparisons
- Good discussion of this on p559 in Lozupone and Knight review
- I (Eisen) view things a little differently
- Would be useful to at least briefly link the microbial diversity/phylodiversity literature to the non-microbial ecology literature on this subject
- Note parallel development of methods and tools to answer many of the same questions with different datasets. Emphasize need to unify.
- First key issue is whether to analyze communities one by one with some metric and then to compare the scores or to develop scores explicitly from comparing communities
- i.e. alpha vs. beta diversity
- Within community metric could be
- Richness
- Diversity
- Within community metric could be
- Lozupone and Knight outline a few other issues including
- Qualitative vs. quantitative
- Qualitative do not take into account species counts, just presence/absence
- Species based or tree based
- Qualitative vs. quantitative
OTU and phylotype focused
- Presence/absence of OTUs
- Presence/absence of phylotypes (at all levels)
- SONS
Coverage and Statistical Significance
- See f-Libshuff
- See Original libshuff
Phylogenetic structure of communities
- See Tree Climber
- See AP Martin paper on phylogenetic based comparison where FST and P test introduced
- See AP Martin paper #1
- Unifrac
- Weighted Unifrac
Metagenomic distances
Statistical issues
- Sampling
- Statistical concepts (e.g. experimental unit (species, community, environment, day, etc))
- Assessing reliability and repeatability of results
Case Studies and Science
Reviews on methods
- See New Review by Schloss
- See new Schloss paper
- http://www.nature.com/ismej/journal/v2/n7/abs/ismej200844a.html;jsessionid=12A1751090E8D55918D9AC086B76837F
- http://www.ncbi.nlm.nih.gov/pubmed/18435746?ordinalpos=18&itool=EntrezSystem2.PEntrez.Pubmed.Pubmed_ResultsPanel.Pubmed_RVDocSum
Metagenomics informatics
- Reviews
- Chen and Pachter. PLoS Comp. Biol 2005
- Schloss and Handelsman Statistical toolbox
- Binning
- Phylopythia by McHardy et al.
General diversity things
- See Curtis et al. review
- See Global patterns in microbial diversity paper
- See Beta diversity paper by Knight
- See Hughes rarefaction paper
Biogeography
- See Taxa Area Paper
- Also see Other island paper
- See Freir paper
- See Green et al on microbial euks
Tables
Data Types
- rRNA sequence
- Pros
- Cons
- rRNA tRFLP
- Pros
- Cons
- metagenomics
- Pros
- Cons
- rRNA sequence
Diversity Methods
- alpha diversity
- Phylogenetic diversity (PD)
- Pros
- Cons
- Applications
- References
- Phylogenetic diversity (PD)
- beta diversity