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FACE Soil Metagenome Comparisons in IMG. Melissa Dsouza , Peter Hallin , Craig Herbold, Rima Upchurch , & Paul Wilkinson . FACE Site Overview. Free-air and Carbon Dioxide Enrichment Elevated levels of CO2 expected in future decades Metagenome data in IMG/M Forested sites

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face soil metagenome comparisons in img

FACE Soil Metagenome Comparisons in IMG

Melissa Dsouza, Peter Hallin, Craig Herbold,

Rima Upchurch, & Paul Wilkinson

face site overview
FACE Site Overview
  • Free-air and Carbon Dioxide Enrichment
    • Elevated levels of CO2 expected in future decades
  • Metagenome data in IMG/M
    • Forested sites
      • Duke (Sweetgum forest)
      • Oak Ridge (Loblolly pine)
    • Nevada sites
      • Soil crust
      • Cresote
    • Maryland Estuary
metagenomes overview
Metagenomes Overview
  • Soil samples
  • Standardized extraction across all sites
    • Cheryl Kuske (LANL)
    • FastDNA Spin kit
    • 2007-2008 sampling
  • Soil samples
  • Extracted using FastDNA Spin kit
  • 454 Titanium sequencing at JGI
metagenomes overview1
Metagenomes Overview
  • Similar GC
  • Similar sequencing effort
  • Scaffold data
    • 65-75% - 1 gene
    • Length of scaffolds varied low as 100 to 630
      • 450-550 median
    • Basically unassembled reads
questions about face
Questions about FACE
  • Question 1: Are there differences in the taxonomic and functional distributions across all sites?
  • Question 2: Are there are any differences between taxonomy within each elevated and ambient sites?
  • Question 3: Are there any functional differences within each elevated and ambient sites?
taxonomy across sites
Taxonomy across sites
  • Similar phyla present overall, but there were a few differences between sites
  • Actinobacteria and Proteobacteria top phyla
    • Swapped top or second between sites
  • Other top groups shifted between sites
    • Firmicutes, Bacteroidetes, Cyanobacteria, Acidobacteria and Verrucomicrobia
nitrogen fixation
Nitrogen fixation
  • Added KEGG pathway (ko00910) to “Function Card” and compared in Metagenome samples
  • Output is matrix containing gene counts
  • Using R (Sorry IMG team!) we created a 2-dimensional clustering.
  • Clearly, samples cluster according to site, rather than CO2 level.
clustering of sites
Clustering of sites
  • PCA of all COGs in all sites
  • Axes 1 and 2 explain ~90% of the variation
    • Largest variation in COGs appeared related to site rather than treatment
  • Remaining analyses done by site

to examine differences

by treatment

Desert

Forests

Estuary

site function summary
Site Function Summary
  • Did not see significant differences in 3 of 5 sites
  • Maryland Estuary
    • Membrane biogenesis-related functions
  • Soil crust
    • Did not see a pattern
  • Was there overlap in COG functions
    • One: a transposase in both
site function summary1
Site Function Summary
  • Did not see significant differences in 3 of 5 sites
  • Maryland Estuary
    • Membrane biogenesis-related functions
  • Soil crust
    • Did not see a pattern
  • Was there overlap in COG functions
    • One: a transposase in both
transposase
Transposase
  • Maryland Estuary
    • Dominant group was Alphaproteobacteria
  • Nevada soil crust
    • Dominant group was Cyanobacteria
  • Reflective of dominant members of each site
maryland estuary
Maryland Estuary
  • COGs that were significant
    • Uncharacterized conserved protein
    • Transposase
    • Glycoltransferase
    • Serine/Theronine protein kinase
    • Membrane-fusion protein
maryland estuary1
Maryland Estuary
  • COGs that were significant
    • Uncharacterized conserved protein
    • Transposase
    • Glycoltransferase
    • Serine/Theronine protein kinase
    • Membrane-fusion protein
  • Cell membrane biogenesis
    • Studied phylogenetic distribution of the genes within these COGs revealed…
maryland estuary2
Maryland Estuary
  • Deltaproteobacteria were the dominant group for the phylogenetic distribution
  • This group might be the driving force between ambient and elevated within this site
  • Conclusion
nevada desert soil crust
Nevada Desert Soil Crust
  • COGs that were significant
    • Acetyltransferases
    • Predicted hydrolases or acyltransferases
    • Transposase
    • Cytochrome b subunit of the bc complex
    • Predicted ATPase
    • FOG: PAS/PAC domain
    • Molybdenum cofactor biosynthesis enzyme
  • Pfams
    • COX1
    • SBP_bac_5
    • HAMP
    • Hydrolase
    • MoeA_C
    • DUF349
    • Radical_SAM
    • SH3_3
  • TIGRfam
    • YD repeat (two copies)
    • delta-60 repeat domain
nevada desert soil crust1
Nevada Desert Soil Crust
  • COGs that were significant
    • Acetyltransferases
    • Predicted hydrolases or acyltransferases
    • Transposase
    • Cytochrome b subunit of the bc complex
    • Predicted ATPase
    • FOG: PAS/PAC domain
    • Molybdenum cofactor biosynthesis enzyme
  • Pfams
    • COX1
    • SBP_bac_5
    • HAMP
    • Hydrolase
    • MoeA_C
    • DUF349
    • Radical_SAM
    • SH3_3
  • TIGRfam
    • YD repeat (two copies)
    • delta-60 repeat domain
nevada desert soil crust2
Nevada Desert Soil Crust

Predicted ATPase

  • Actinobacteria, Alphaproteobacteria and Cyanobacteria were the most dominate in the distribution of functional categories
  • In one case Chloroflexi (shown right) were most dominate
  • Conclusions
final conclusions
Final Conclusions
  • Impact of elevated carbon dioxide
    • Did not see any specific function change
    • Taxon played a major part in our analyses for determining significance in functions
  • Sequence quality
    • Our datasets were mainly unassembled sequences
    • Low quality (shorter reads) affects annotation
    • Frame shift questions for 454 sequencing
    • Not seeing the “real” picture of these sites