1. Microbial Diversity in Drain Hole Waters of the Henderson Mine John R. Spear Jason Sahl Division of Environmental Science and Engineering Colorado School of Mines, Golden, CO jspear@mines.edu

2. $125 x 10 6

3. Amount spent in the U.S. on marshmallows in 2005

4. What Is Life?

12. You are here Euryarchaeota Crenarchaeota A “Map” of Life Euryarchaeota Crenarchaeota N.R. Pace, Science, 1997

15. Molecular Method Env. Sample Bulk DNA/RNA Extraction Community rRNA/rDNA Clones PCR Cloning rRNA/rDNA Sequences Sequencing Phylogenetic Trees Databases

16. 7150’ DH-3 Data Fungus Sulfate Reducing Bacteria

17. 7025’ DH-1 Data Fungus Sulfate Reducing Bacteria

18. 7025’ DH-1-Post Packer Data Fungus Sulfate Reducing Bacteria

19. 7025’ DH-4-Post Packer Data Fungus Sulfate Reducing Bacteria

20. DH Chemistry

21. HM

22. Conclusions Potential for 3 new groups (phyla) of Bacteria!! Something is happening with N!! Community shift pre- verses packed- drill holes. Surprising diversity for a subsurface. Low abundance of both Archaea and Eucarya! Publish?

23. Thanks To: My Start-up money…. Team Bio--Tom K., Jeff M., Alexis T., Betsy S., Jeff W., Kevin M. HUSEP Folks Henderson Mine and Chip deWolfe

24. “If it wasn’t for the optimist, the pessimist would never know…. how happy he wasn’t.”

25. Henderson, New Drill Core Plan, to drill a core into the HUSEP facility area For some portion of that: –Take a segment for biological consideration Sterile?BEADS + Tracer Core collection--on site Core sample sections to Socorro, CSM, CU –Continue drilling to depth

26. Henderson Mine, 2 Nov 05

27. Molecular Method Env. Sample Contact Slide Bulk DNA/RNA Extraction Community rRNA/rDNA Clones PCR Cloning = Method= Result rRNA/rDNA Sequences / Databases Nucleic Acid Probes Sequencing Probe Design Whole-Cell Hybridization (FISH) Phylogenetic Trees Nucleic Acid Hybridization

30. DH-2, DH-3 + ???? To Do…. Water = 1.5’ PVC Pipe Inserted with Glass and/or Fe Growth Slides Anoxic H 2 O out

31. DH-1 Water Chemistry + Cultures SpeciesConcentration Si4+0.05 mM Ca2+8.7 mM Mg2+1.3 mM Al3+4.8 mM Na+7.9 mM Sr2+0.04 mM K+1.0 mM FeT1.5 mM Mn2+21 mM F-12 mM Cl-0.15 mM NO2-Not detected NO3-1.0 mM Br-Not detected PO43-Not detected SO42-48 mM DH-1 at 7150’, pH 5.8 Culture efforts: Swanner & Templeton, CU: DH-13 (Fe + Mn) DH-33 (Mn) DH-42 (Fe + Mn)