Geomicrobiology of subglacial ice above Lake Vostok, Antarctica

Science

Volumn 286, Issue 5447, 1999, Pages 2141-2144

  Priscu, John C ^a   Adams, Edward E ^b   Lyons, W Berry ^c   Voytek, Mary A ^d   Mogk, David W ^a   Brown, Robert L ^b   McKay, Christopher P ^e   Takacs, Cristina D ^a   Welch, Kathy A ^c   Wolf, Craig F ^a   Kirshtein, Julie D ^d   Avci, Recep ^a  

^a MONTANA STATE UNIVERSITY   (United States)

^b Montana State University   (United States)

^c UNIVERSITY OF ALABAMA   (United States)

^d U S GEOLOGICAL SURVEY   (United States)

^e NASA AMES RESEARCH CENTER   (United States)

Author keywords

[No Author keywords available]

Indexed keywords

ICE; RNA 16S;

ICE CORE; ICE THICKNESS; LACUSTRINE ENVIRONMENT; LAKE ECOSYSTEM; MICROBIOLOGY;

ACTINOBACTERIA; ANTARCTICA; ARCTIC CLIMATE; ARTICLE; COLONY FORMING CELL; GEOLOGY; LAKE; MICROBIOLOGY; PHYLOGENY; PRIORITY JOURNAL;

NON-PROGRAMMATIC;

ANTARCTIC REGIONS; BACTERIA; BACTERIAL PHYSIOLOGY; DNA, BACTERIAL; DNA, RIBOSOMAL; FRESH WATER; GENES, RRNA; ICE; MICROSCOPY, ELECTRON, SCANNING; MICROSCOPY, FLUORESCENCE; MINERALS; PRESSURE; PROTEOBACTERIA; RNA, RIBOSOMAL, 16S; TEMPERATURE; WATER MICROBIOLOGY;

ANTARCTICA;

EID: 0032763909     PISSN: 00368075     EISSN: None     Source Type: Journal    
DOI: 10.1126/science.286.5447.2141     Document Type: Article

References (36)

1. M. J. Siegert, J. A. Dowdeswell, M. R. Gorman, H. F. McIntyre, Antarct. Sci. 8, 281 (1996).
2. A. P. Kapista, J. K. Ridley, D. G. de Q Robin, M. J. Siegert, I. A. Zotikov, Nature 381, 684 (1996).
3. J. R. Petit et al., Nature 399, 429 (1999).
4. J. Jouzel et al., Science 286, 2138 (1999).
5. Core 3590 was a longitudinal cut from a 10-cm-diameter, 44-cm-long core obtained from Vostok hole 5G. The core had minor cracking propagating <3 cm into the sidewall and no exsolved clathrates. Nonaggregated sediments were dispersed throughout the core, and five sediment inclusions ranging from 0.5 to 1 mm in diameter were present. Core sections were cut for analyses with a saw sterilized with ethanol Samples were processed under a sterile, positive-pressure laminar flow hood; sterile gloves, clean laboratory clothing, and hair covering were worn during handling. All core handling was conducted in a laboratory that had never been used to grow or process biological samples. Ion and trace chemical samples were rinsed thoroughly with 0.2-μm-filtered Barnstead-nanopure water until 4 to 10 mm of the outer surface had melted. The samples were then completely melted at room temperature in clean, sterile high-density polyethylene (HDPE) jars. Ions in filtered (0.2 μm) and unfiltered samples were analyzed by ion chromatography [K. A. Welch et al., J. Chromatogr. 793, 257 (1996)]; trace elements in unfiltered melt were determined by inductively coupled plasma mass spectroscopy (ICP-MS). Stable isotope samples were melted without rinsing and analyzed by mass spectrometry. SEM samples were rinsed and melted as for ion chemistry. Melted SEM samples were filtered onto sterile 0.2-μm filters with cleaned and sterilized equipment. A SEM control was prepared with 0.2-μm-filtered nanopure water frozen in a clean, sterile polycarbonate tube. The control core was melted, filtered, and analyzed by SEM with methods identical to those of the sample. Mineral and biological particles from the sample were unique with respect to that observed in the control core, indicating that the sample portions we analyzed were free from particulate matter contamination. A sample for epifluorescence microscopy of acridine orange-stained cells was melted in an acid-washed autoclaved bottle at 4°C. DNA staining and counting protocols are described in (31).
6. Crystal orientation was determined with a Rigsby stage [C. C. Langway, SIPRE Tech. Rep. No. 62 (U.S. Army Cold Regions Research and Engineering Laboratory, Hanover, NH, 1959)].
7. K. H. Wedepohl, Geochim. Cosmochim. Acta 59, 1217 (1995).
8. J. A. Killawee, I. J. Fairchild, J.-L. Tison, L. Janssens, R. Lorrain, Geochim. Cosmochim. Acta 62, 3637 (1998).
9. Concentrations of ions, DOC, and bacteria were predicted in Lake Vostok by applying the partitioning observed between these constituents in the surface waters and in accretion ice from Lakes Hoare (ions) and Bonney (DOC and bacteria) (16, 31).
10. J. I. Drever, The Geochemistry of Natural Waters (Prentice-Hall, Englewood Cliffs, NJ, 1988).
11. std -1] × 1000 where std is the standard mean ocean water reference.
12. J. R. O'Neil, J. Phys. Chem. 72, 3683 (1958).
13. Cryogenic SEM (JEOL-6100 SEM) and energy dispersive spectrornetry (EDS) were used to image and analyze particles captured by 0.2-μm filtration of melted ice (EDS could only be used on particles > 1 μm in diameter). Atomic force microscopy used a Digital Instrument's Dimension 3100 system in tapping mode.
14. D. W. Hyndman, Petrology of Igneous and Metamorphic Rocks (McGraw-Hill, New York, ed. 2, 1985).
15. D. M. Karl et al., Science 286, 2144 (1999).
16. J. C. Priscu et al., Science 280, 2095 (1998).
17. Particulates from 250 ml of melted ice core were collected on a sterile 0.2-μm polycarbonate filter. Genomic DNA was extracted from the filter with Chelex 100 resin [P. S. Walsh, D. A. Metzger, R. Higuchi, Biotechniques 10, 506 (1991) ]. PCR primers corresponding to conserved sequences within the 5′ and 3′ regions of the 16S ribosomal DNAs of bacteria and archae were used. Primary amplification was performed with B1 (E. coli position 8 to 27) and B2 (E. coli position 1492 to 1510) [W. Liesack, H. Weyland, E. Stackenbrandt, Microb. Ecol. 21, 191 (1991)] and Archael 1 (E. coli position 3 to 20) and B2 [C. F. Brunk, E. Avaniss-Aghajani, C. A. Brunk, Appl. Environ. Microbiol. 62, 872 (1996)]. A second amplification was performed with 1 μl from the first amplification and nested primers B3 (E. coli position 46 to 65) and B4 (E. coli position 536 to 519). PCR amplifications were performed following optimized protocols described by M. A. Voytek and B. B. Ward [Appl. Environ. Microbiol. 56, 2430 (1995)].
18. Particulates from 250 ml of melted ice core were collected on a sterile 0.2-μm polycarbonate filter. Genomic DNA was extracted from the filter with Chelex 100 resin [P. S. Walsh, D. A. Metzger, R. Higuchi, Biotechniques 10, 506 (1991) ]. PCR primers corresponding to conserved sequences within the 5′ and 3′ regions of the 16S ribosomal DNAs of bacteria and archae were used. Primary amplification was performed with B1 (E. coli position 8 to 27) and B2 (E. coli position 1492 to 1510) [W. Liesack, H. Weyland, E. Stackenbrandt, Microb. Ecol. 21, 191 (1991)] and Archael 1 (E. coli position 3 to 20) and B2 [C. F. Brunk, E. Avaniss-Aghajani, C. A. Brunk, Appl. Environ. Microbiol. 62, 872 (1996)]. A second amplification was performed with 1 μl from the first amplification and nested primers B3 (E. coli position 46 to 65) and B4 (E. coli position 536 to 519). PCR amplifications were performed following optimized protocols described by M. A. Voytek and B. B. Ward [Appl. Environ. Microbiol. 56, 2430 (1995)].
19. Particulates from 250 ml of melted ice core were collected on a sterile 0.2-μm polycarbonate filter. Genomic DNA was extracted from the filter with Chelex 100 resin [P. S. Walsh, D. A. Metzger, R. Higuchi, Biotechniques 10, 506 (1991) ]. PCR primers corresponding to conserved sequences within the 5′ and 3′ regions of the 16S ribosomal DNAs of bacteria and archae were used. Primary amplification was performed with B1 (E. coli position 8 to 27) and B2 (E. coli position 1492 to 1510) [W. Liesack, H. Weyland, E. Stackenbrandt, Microb. Ecol. 21, 191 (1991)] and Archael 1 (E. coli position 3 to 20) and B2 [C. F. Brunk, E. Avaniss-Aghajani, C. A. Brunk, Appl. Environ. Microbiol. 62, 872 (1996)]. A second amplification was performed with 1 μl from the first amplification and nested primers B3 (E. coli position 46 to 65) and B4 (E. coli position 536 to 519). PCR amplifications were performed following optimized protocols described by M. A. Voytek and B. B. Ward [Appl. Environ. Microbiol. 56, 2430 (1995)].
20. Particulates from 250 ml of melted ice core were collected on a sterile 0.2-μm polycarbonate filter. Genomic DNA was extracted from the filter with Chelex 100 resin [P. S. Walsh, D. A. Metzger, R. Higuchi, Biotechniques 10, 506 (1991) ]. PCR primers corresponding to conserved sequences within the 5′ and 3′ regions of the 16S ribosomal DNAs of bacteria and archae were used. Primary amplification was performed with B1 (E. coli position 8 to 27) and B2 (E. coli position 1492 to 1510) [W. Liesack, H. Weyland, E. Stackenbrandt, Microb. Ecol. 21, 191 (1991)] and Archael 1 (E. coli position 3 to 20) and B2 [C. F. Brunk, E. Avaniss-Aghajani, C. A. Brunk, Appl. Environ. Microbiol. 62, 872 (1996)]. A second amplification was performed with 1 μl from the first amplification and nested primers B3 (E. coli position 46 to 65) and B4 (E. coli position 536 to 519). PCR amplifications were performed following optimized protocols described by M. A. Voytek and B. B. Ward [Appl. Environ. Microbiol. 56, 2430 (1995)].
21. PCR amplification for T-RFLP was performed with fluorescently labeled bacterial 16S ribosomal DNA [bacterial B3 and B4 (17)]. A 2-μl portion of the PCR product was digested with Sau3A following the protocol of Promega. Digestion fragments were analyzed by capillary electrophoresis on an ABI 310.
22. B. L. Maidak et al., Nucleic Acids Res. 27, 171 (1999). http://www.cme.msu.edu/RDP/html/index.html.
23. S. F. Altschul et al., Nucleic Acids Res. 25, 3389 (1997). Classified by comparison with the GenBank database with BLAST.
24. S. S. Abyzov, in Antarctic Microbiology, E. I. Friedmann, Ed. (Wiley-Liss, New York, 1993). pp. 265-295.
25. 2. Six replicate injections yielded a coefficient of variation of 2%.
26. M. S. Twickler, M. J. Spencer, W. B. Lyons, P. A. Mayewski, Nature 320, 156 (1986).
27. K. Kawamura, I. Suzuki, Y. Fujii, O. Watanabe, Geophys. Res. Lett. 23, 2665 (1996).
28. 3H-labeled mannitol incubations were terminated by filtration alone.
29. National Science Foundation Workshop-THe Lake Vostok Study: A Curiosity or a Focus for Interdisciplinary Investigations, Washington, DC, 7 to 8 September 1998 (see www.Ideo.columbia.edu/vostok/).
30. -1) plates prepared with the same media. The inoculated media were incubated for 4 months at 4°C in air at 1 atm in the dark (liquid media preparations were on a shaker during incubation).
31. M. H. Carr et al., Nature 391, 363 (1998).
32. R. T. Pappalardo et al., J. Geophys. Res. 104, 24015 (1999).
33. R. Greenberg et al., Icarus 141, 263 (1999).
34. C. D. Takacs and J. C. Priscu, Microb. Ecol. 36, 239 (1998).
35. M. Legrand and P. Mayewski, Rev. Geophys. 35, 219 (1997).
36. Discussions with D. Karl, J. R. Petit, P. Price, J. Palais, B. Vaughn, and M. Edens helped improve the manuscript. C. Wend, C. Colonero, and E. Graham performed the DOC, stable isotope, and ICP-MS analyses, respectively. We thank all involved with coring operations. This research was supported by T. McCoy, Vice President for Research, Montana State University, and NSF grants OPP94-19413, OPP92-11773, OPP98-15512, and OPP98-15998.