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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. 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

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  1. FACE Soil Metagenome Comparisons in IMG Melissa Dsouza, Peter Hallin, Craig Herbold, Rima Upchurch, & Paul Wilkinson

  2. 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

  3. 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

  4. 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

  5. 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?

  6. 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

  7. Functional overview across sites

  8. 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.

  9. Maryland Estuary CO +/- lights up

  10. 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

  11. 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

  12. 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

  13. Transposase • Maryland Estuary • Dominant group was Alphaproteobacteria • Nevada soil crust • Dominant group was Cyanobacteria • Reflective of dominant members of each site

  14. Maryland Estuary • COGs that were significant • Uncharacterized conserved protein • Transposase • Glycoltransferase • Serine/Theronine protein kinase • Membrane-fusion protein

  15. 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…

  16. 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

  17. 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

  18. 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

  19. 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

  20. 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

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