Publications by authors named "David Gilichinsky"

28 Publications

  • Page 1 of 1

Celerinatantimonas yamalensis sp. nov., a cold-adapted diazotrophic bacterium from a cold permafrost brine.

Int J Syst Evol Microbiol 2013 Dec 12;63(Pt 12):4421-4427. Epub 2013 Jul 12.

Institute of Physicochemical and Biological Problems in Soil Sciences, Russian Academy of Sciences, Pushchino, Moscow Region, Russian Federation, 142290.

A facultatively anaerobic nitrogen-fixing bacterium, strain C7(T), was isolated from a permafrost cryopeg on the Yamal Peninsula, Russia. Comparative analysis of 16S rRNA gene sequences revealed that this bacterium was closely related to Celerinatantimonas diazotrophica S-G2-2(T) with a similarity of 95.5 %. Strain C7(T) differed from Celerinatantimonas diazotrophica in its ability to hydrolyse gelatin and inability to use d-mannose, melibiose, l-rhamnose, myo-inositol, lactose, lactulose, d-mannitol, trehalose, dl-lactate, glycogen or l-proline as sole carbon sources. In addition, strain C7(T) grew over a temperature range of 0-34 °C with optimum growth at 18-22 °C. The whole-cell fatty acid profile included C16 : 0, C16 : 1ω7, C18 : 1ω7, C17 cyclo and summed feature 2 [comprising C12 : 0 aldehyde and/or unknown fatty acid 10.913 (MIDI designation) and/or iso-C16 : 1/C14 : 0 3-OH]. The DNA G+C content was 44.7 mol%. Strain C7(T) is thus considered to represent a novel species, for which the name Celerinatantimonas yamalensis sp. nov. is proposed. The type strain is C7(T) ( = VKM B-2511(T) = DSM 21888(T)).
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http://dx.doi.org/10.1099/ijs.0.045997-0DOI Listing
December 2013

Growth of Carnobacterium spp. from permafrost under low pressure, temperature, and anoxic atmosphere has implications for Earth microbes on Mars.

Proc Natl Acad Sci U S A 2013 Jan 24;110(2):666-71. Epub 2012 Dec 24.

Department of Microbiology and Cell Science, Space Life Sciences Laboratory, University of Florida, Merritt Island, FL 32953, USA.

The ability of terrestrial microorganisms to grow in the near-surface environment of Mars is of importance to the search for life and protection of that planet from forward contamination by human and robotic exploration. Because most water on present-day Mars is frozen in the regolith, permafrosts are considered to be terrestrial analogs of the martian subsurface environment. Six bacterial isolates were obtained from a permafrost borehole in northeastern Siberia capable of growth under conditions of low temperature (0 °C), low pressure (7 mbar), and a CO(2)-enriched anoxic atmosphere. By 16S ribosomal DNA analysis, all six permafrost isolates were identified as species of the genus Carnobacterium, most closely related to C. inhibens (five isolates) and C. viridans (one isolate). Quantitative growth assays demonstrated that the six permafrost isolates, as well as nine type species of Carnobacterium (C. alterfunditum, C. divergens, C. funditum, C. gallinarum, C. inhibens, C. maltaromaticum, C. mobile, C. pleistocenium, and C. viridans) were all capable of growth under cold, low-pressure, anoxic conditions, thus extending the low-pressure extreme at which life can function.
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http://dx.doi.org/10.1073/pnas.1209793110DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3545801PMC
January 2013

Structure of the O-specific polysaccharide from the lipopolysaccharide of Psychrobacter cryohalolentis K5(T) containing a 2,3,4-triacetamido-2,3,4-trideoxy-L-arabinose moiety.

J Nat Prod 2012 Dec 29;75(12):2236-40. Epub 2012 Nov 29.

Zelinsky Institute of Organic Chemistry, Russian Academy of Sciences, 119991 Moscow, Russia.

A novel constituent of bacterial polysaccharides, 2,3,4-triacetamido-2,3,4-trideoxy-L-arabinose, was found in the O-specific polysaccharide from the lipopolysaccharide of Psychrobacter cryohalolentis K5(T) and identified by 1D and 2D (1)H and (13)C NMR studies of the polysaccharide and a disaccharide obtained by solvolysis of the polysaccharide with triflic acid. The following structure of the branched polysaccharide was established by sugar analysis, triflic acid solvolysis, Smith degradation, and 2D NMR spectroscopy.
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http://dx.doi.org/10.1021/np300484mDOI Listing
December 2012

Structure of an acidic polysaccharide isolated from Psychrobacter maritimus 3pS containing a bacillosamine derivative.

Carbohydr Res 2012 Oct 21;359:7-10. Epub 2012 Jul 21.

Zelinsky Institute of Organic Chemistry, Russian Academy of Sciences, 119991 Moscow, Russia.

An acidic polysaccharide was obtained from Psychrobacter maritimus 3pS isolated from a Siberian cryopeg sample (Kolyma lowland). The following structure of the tetrasaccharide repeating unit of the polysaccharide was established by sugar analysis along with (1)H and (13)C NMR spectroscopy: →2)-α-L-Rhap-(1→4)-α-D-GalpNAcA-(1→3)-α-D-QuipNAc4NHb-(1→3)-β-D-QuipNAc4NHb-(1→ where D-GalNAcA indicates 2-acetamido-2-deoxy-D-galacturonic acid and d-QuiNAc4NHb indicates 2-acetamido-2,4,6-trideoxy-4-[(S)-3-hydroxybutanoyl]amino-D-glucose.
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http://dx.doi.org/10.1016/j.carres.2012.07.007DOI Listing
October 2012

Ancient fungi in Antarctic permafrost environments.

FEMS Microbiol Ecol 2012 Nov 23;82(2):501-9. Epub 2012 Jul 23.

All-Russian Collection of Microorganisms (VKM), G K Skryabin Institute of Biochemistry and Physiology of Microorganisms, Russian Academy of Sciences, Pushchino, Moscow Region, Russia.

Filamentous fungi in 36 samples of Antarctic permafrost sediments were studied. The samples collected during the Russian Antarctic expedition of 2007-2009 within the framework of the Antarctic Permafrost Age Project (ANTPAGE) were recovered from different depths in ice-free oases located along the perimeter of the continent. Fungal diversity was determined by conventional microbiological techniques combined with a culture-independent method based on the analysis of internal transcribed spacer (ITS2) sequences in total DNA of the samples. The study revealed a rather low fungal population density in permafrost, although the diversity found was appreciable, representing more than 26 genera. Comparison of the data obtained by different techniques showed that the culture-independent method enabled the detection of ascomycetous and basidiomycetous fungi not found by culturing. The molecular method failed to detect members of the genera Penicillium and Cladosporium that possess small-sized spores known to have a high resistance to environmental changes.
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http://dx.doi.org/10.1111/j.1574-6941.2012.01442.xDOI Listing
November 2012

Cloning, purification, and characterization of a cold-adapted esterase produced by Psychrobacter cryohalolentis K5T from Siberian cryopeg.

FEMS Microbiol Ecol 2012 Nov 30;82(2):367-75. Epub 2012 Apr 30.

Institute of Physicochemical and Biological Problems in Soil Science, Russian Academy of Sciences, Puschino, Moscow Region, Russia.

A psychrotrophic gram-negative bacterium Psychrobacter cryohalolentis K5(T) was previously isolated from a cryopeg within Siberian permafrost and its genome has been completely sequenced. To clone and characterize potential cold-active lipases/esterases produced by P. cryohalolentis K5(T) , we have identified their potential genes by alignment with amino acid sequences of lipases/esterases from related bacteria. One of the targets, EstPc, was cloned and overexpressed in Escherichia coli BL21 (DE3) cells. The recombinant protein was produced with a 6x histidine tag at its C-terminus and purified by nickel affinity chromatography. Purified recombinant protein displayed maximum esterolytic activity with p-nitrophenyl butyrate (C4) as a substrate at 35 °C and pH 8.5. Activity assay conducted at different temperatures revealed that EstPc is a cold-adapted esterase which displayed more than 90% of its maximum activity at 0-5 °C. In contrast to many known cold-active enzymes, it possesses relatively high thermostability, preserving more than 60% of activity after incubation for 1 h at 80 °C. It was activated by Ca(2+) , Mn(2+) , and EDTA whereas Zn(+2) , Cu(+2) , Co(+2) , Ni(+2) , and Mg(+2) inhibited it. Various organic solvents (ethanol, methanol and others) inhibited the enzyme. Most non-ionic detergents, such as Triton X-100 and Tween 20 increased the lipase activity while SDS completely inhibited it.
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http://dx.doi.org/10.1111/j.1574-6941.2012.01385.xDOI Listing
November 2012

Regeneration of whole fertile plants from 30,000-y-old fruit tissue buried in Siberian permafrost.

Proc Natl Acad Sci U S A 2012 Mar 21;109(10):4008-13. Epub 2012 Feb 21.

Institute of Cell Biophysics, Russian Academy of Sciences, Pushchino 142290, Russia.

Whole, fertile plants of Silene stenophylla Ledeb. (Caryophyllaceae) have been uniquely regenerated from maternal, immature fruit tissue of Late Pleistocene age using in vitro tissue culture and clonal micropropagation. The fruits were excavated in northeastern Siberia from fossil squirrel burrows buried at a depth of 38 m in undisturbed and never thawed Late Pleistocene permafrost sediments with a temperature of -7 °C. Accelerator mass spectrometry (AMS) radiocarbon dating showed fruits to be 31,800 ± 300 y old. The total γ-radiation dose accumulated by the fruits during this time was calculated as 0.07 kGy; this is the maximal reported dose after which tissues remain viable and seeds still germinate. Regenerated plants were brought to flowering and fruiting and they set viable seeds. At present, plants of S. stenophylla are the most ancient, viable, multicellular, living organisms. Morphophysiological studies comparing regenerated and extant plants obtained from modern seeds of the same species in the same region revealed that they were distinct phenotypes of S. stenophylla. The first generation cultivated from seeds obtained from regenerated plants progressed through all developmental stages and had the same morphological features as parent plants. The investigation showed high cryoresistance of plant placental tissue in permafrost. This natural cryopreservation of plant tissue over many thousands of years demonstrates a role for permafrost as a depository for an ancient gene pool, i.e., preexisting life, which hypothetically has long since vanished from the earth's surface, a potential source of ancient germplasm, and a laboratory for the study of rates of microevolution.
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http://dx.doi.org/10.1073/pnas.1118386109DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3309767PMC
March 2012

Cryptic diversity within the choanoflagellate morphospecies complex Codosiga botrytis - phylogeny and morphology of ancient and modern isolates.

Eur J Protistol 2012 Nov 17;48(4):263-73. Epub 2012 Feb 17.

Biocenter Cologne, Zoological Institute, Department of General Ecology, University of Cologne, Germany.

Choanoflagellates are closely related to metazoans and fungi according to recent phylogenetic studies; therefore the systematics of these organisms is of particular interest. The choanoflagellate morphospecies Codosiga botrytis is the first described choanoflagellate, and is one of the most frequently reported choanoflagellate species. In this study we present phylogenetic and morphological data on eight different strains of Codosiga botrytis. Among these there are five ancient strains; these cultures have been established from up to 43,000 years old cysts from Siberian permafrost. We found that based on the variable V4 region of the small subunit (SSU) of the rDNA, all the investigated freshwater isolates of Codosiga botrytis, together with Sphaeroeca volvox, form a cluster at the base of all other choanoflagellate species. Moreover, the morphospecies described classically as Codosiga botrytis contains at least four different genotypes separated by considerably high genetic distance. All these 'cryptic species' have identical general morphology and cell structure. Strains have a similar life cycle with several different life forms and large morphological plasticity. For the first time we were able to establish cultures from cryo-conserved cysts of choanoflagellates. The ancient strains did not differ significantly in partial SSU rDNA from the modern ones. Besides, no biogeographically pattern could be established. This fact and the low genetic distances of some strains from remote locations support the distribution of dormant stages via air.
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http://dx.doi.org/10.1016/j.ejop.2012.01.004DOI Listing
November 2012

Structure of the O-polysaccharide chain of the lipopolysaccharide of Psychrobacter muricolla 2pS(T) isolated from overcooled water brines within permafrost.

Carbohydr Res 2012 Feb 6;349:78-81. Epub 2011 Dec 6.

Zelinsky Institute of Organic Chemistry, Russian Academy of Sciences, 119991 Moscow, Russia.

Psychrotrophic bacteria of the genus Psychrobacter have not been studied in respect to lipopolysaccharide structure. In this work, we determined the structure of the O-specific polysaccharide of the lipopolysaccharide of Psychrobacter muricolla 2pS(T) isolated from overcooled (-9°C) water brines within permafrost. The polysaccharide was found to be acidic due to the presence of an amide of 2-acetamido-2-deoxy-l-guluronic acid with glycine (l-GulNAcA6Gly), which has not been hitherto found in nature. The following structure of the disaccharide repeating unit of the polysaccharide was established using composition analysis along with 1D and 2D (1)H and (13)C NMR spectroscopy: →4)-α-l-GulpNAcA6Gly-(1→3)-β-d-GlcpNAc-(1→
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http://dx.doi.org/10.1016/j.carres.2011.11.027DOI Listing
February 2012

Characterization of a bacterial community from a Northeast Siberian seacoast permafrost sample.

FEMS Microbiol Ecol 2010 Oct 2;74(1):103-13. Epub 2010 Aug 2.

Center for Microbial Ecology, Michigan State University, East Lansing, MI, USA.

A combination of culture-dependent and -independent techniques was used to characterize a bacterial community, examine cold adaptation of isocitrate lyase (icl) genes, and detect genes with important ecological functions in a permafrost sample from the Bykovsky Peninsula on the Laptev Sea coast of northeast Siberia. According to the 16S rRNA gene sequence, 47 of the cultured isolates were members of the phyla Firmicutes, Proteobacteria, and Actinobacteria, with 85% of the isolates belonging to the genera Arthrobacter and Planococcus. The 16S rRNA gene clone library derived from DNA from the same permafrost sample contained sequences from the same phyla plus a few from Acidobacteria, but favored the Firmicutes at the cost of the Actinobacteria. A partial sequence of the icl gene, a proposed marker for cold adaptation, was determined for 25 isolates that grew at 0 °C. Two Psychrobacter isolates contained two of the four residues shown to be important for low-temperature activity in Colwellia maris or Colwellia psychrerythreaea. The presence in the permafrost DNA of genes with ecosystem functions was determined using geochip 2.0. The highest number of genes identified was from the categories of aromatic and natural polymer degradation genes, perhaps reflecting selection for the use of tundra vegetation-produced carbon.
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http://dx.doi.org/10.1111/j.1574-6941.2010.00945.xDOI Listing
October 2010

Methanobacterium arcticum sp. nov., a methanogenic archaeon from Holocene Arctic permafrost.

Int J Syst Evol Microbiol 2011 Jan 19;61(Pt 1):144-147. Epub 2010 Feb 19.

Institute of Physicochemical and Biological Problems in Soil Sciences, Russian Academy of Sciences, 142290, Pushchino, Moscow Region, Russian Federation.

A mesophilic, non-motile, hydrogenotrophic, rod-shaped methanogen, designated M2(T), was isolated from Holocene permafrost sediments of the Kolyma lowland in the Russian Arctic. Cells were 3-6 μm long and 0.45-0.5 μm wide. Strain M2(T) grew on H(2)/CO(2) and formate. Optimum conditions for growth were 37°C, pH 6.8-7.2 and 0.1 M NaCl. The DNA G+C content was 38.1 mol%. On the basis of 16S rRNA gene sequence comparison with known methanogens, strain M2(T) was affiliated with the genus Methanobacterium and was most closely related to Methanobacterium veterum MK4(T) and Methanobacterium bryantii DSM 863(T) (both 99 % 16S rRNA gene sequence similarity). However, no significant DNA-DNA relatedness was observed between strain M2(T) and these type strains. We propose that strain M2(T) represents a novel species, with the name Methanobacterium arcticum sp. nov., with type strain M2(T) (=DSM 19844(T) =VKM B-2371(T)).
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http://dx.doi.org/10.1099/ijs.0.021311-0DOI Listing
January 2011

Clostridium tagluense sp. nov., a psychrotolerant, anaerobic, spore-forming bacterium from permafrost.

Int J Syst Evol Microbiol 2009 Jun;59(Pt 6):1421-6

Skryabin Institute of Biochemistry and Physiology of Microorganisms, Russian Academy of Sciences, Pushchino 142290, Moscow Region, Russian Federation.

A strictly anaerobic, Gram-positive, psychrotolerant, endospore-forming bacterium (strain A121(T)) was isolated from a permafrost sample collected in the Canadian High Arctic. Phylogenetic analysis of the 16S rRNA gene sequence of strain A121(T) showed its affiliation with the group of psychrophilic and psychrotolerant members of cluster I of the genus Clostridium, Clostridium bowmanii DSM 14206(T) being the closest relative (sequence similarity 98.5 %). Levels of DNA-DNA relatedness between strain A121(T) and the type strains of phylogenetically related species ranged from 33 to 52 %. Strain A121(T) grew in PY broth at temperatures between 4 and 28 degrees C (optimum 15-20 degrees C), at pH 6.0-8.0 (optimum pH 6.5-7.2) and in NaCl concentrations of 0-10.0 % (optimum 0-2.0 %). The strain utilized a narrow range of carbohydrates as sources of carbon and energy, including glucose, fructose, trehalose, maltose and starch; it also hydrolysed gelatin. Predominant fatty acids were C(16 : 1) cis9, C(16 : 1) cis9 DMA, C(16 : 0) and C(14 : 0). The DNA G+C content was 31.5 mol%. On the basis of its overall genotypic and phenotypic characteristics, strain A121(T) is classified within a novel species of the genus Clostridium, Clostridium tagluense sp. nov. The type strain is A121(T) (=VKM B-2369(T) =DSM 17763(T)).
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http://dx.doi.org/10.1099/ijs.0.002295-0DOI Listing
June 2009

Biogeography of two cold-adapted genera: Psychrobacter and Exiguobacterium.

ISME J 2009 Jun 26;3(6):658-65. Epub 2009 Mar 26.

Center for Microbial Ecology, Michigan State University, East Lansing, MI 48824, USA.

The genera Exiguobacterium and Psychrobacter have been frequently detected in and isolated from polar permafrost and ice. These two genera have members that can grow at temperatures as low as -5 and -10 degrees C, respectively. We used quantitative PCR (Q-PCR) to quantify members of these genera in 54 soil or sediment samples from polar, temperate and tropical environments to determine to what extent they are selected by cold environments. These results were further analyzed by multiple linear regression to identify the most relevant environmental factors corresponding to their distribution. Exiguobacterium was detected in all three climatic zones at similar densities, but was patchier in the temperate and tropical samples. Psychrobacter was present in almost all polar samples, was at highest densities in Antarctica sediment samples, but was in very low densities and infrequently detected in temperate and tropical soils. Clone libraries, specific for the 16S rRNA gene for each genus, were constructed from a sample from each climatic region. The clone libraries were analyzed for alpha and beta diversities, as well as for variation in population structure by using analysis of molecular variance. Results confirm that both genera were found in all three climatic zones; however, Psychrobacter populations seemed to be much more diverse than Exiguobacterium in all three climatic zones. Furthermore, Psychrobacter populations from Antarctica are different from those in Michigan and Puerto Rico, which are similar to each other.
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http://dx.doi.org/10.1038/ismej.2009.25DOI Listing
June 2009

Ancient bacteria show evidence of DNA repair.

Proc Natl Acad Sci U S A 2007 Sep 29;104(36):14401-5. Epub 2007 Aug 29.

Department of Earth, Atmospheric, and Planetary Sciences, Massachusetts Institute of Technology, 77 Massachusetts Avenue, 54-810, Cambridge, MA 02139, USA.

Recent claims of cultivable ancient bacteria within sealed environments highlight our limited understanding of the mechanisms behind long-term cell survival. It remains unclear how dormancy, a favored explanation for extended cellular persistence, can cope with spontaneous genomic decay over geological timescales. There has been no direct evidence in ancient microbes for the most likely mechanism, active DNA repair, or for the metabolic activity necessary to sustain it. In this paper, we couple PCR and enzymatic treatment of DNA with direct respiration measurements to investigate long-term survival of bacteria sealed in frozen conditions for up to one million years. Our results show evidence of bacterial survival in samples up to half a million years in age, making this the oldest independently authenticated DNA to date obtained from viable cells. Additionally, we find strong evidence that this long-term survival is closely tied to cellular metabolic activity and DNA repair that over time proves to be superior to dormancy as a mechanism in sustaining bacteria viability.
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http://dx.doi.org/10.1073/pnas.0706787104DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1958816PMC
September 2007

Biogeochemistry of methane and methanogenic archaea in permafrost.

FEMS Microbiol Ecol 2007 Jul 11;61(1):1-15. Epub 2007 Apr 11.

Institute of Physicochemical and Biological Problems in Soil Sciences, Russian Academy of Sciences, Moscow, Russian Federation.

This study summarizes the findings of our research on the genesis of methane, its content and distribution in permafrost horizons of different age and origin. Supported by reliable data from a broad geographical sweep, these findings confirm the presence of methane in permanently frozen fine-grained sediments. In contrast to the omnipresence of carbon dioxide in permafrost, methane-containing horizons (up to 40.0 mL kg(-1)) alternate with strata free of methane. Discrete methane-containing horizons representing over tens of thousands of years are indicative of the absence of methane diffusion through the frozen layers. Along with the isotopic composition of CH(4) carbon (delta(13)C -64 per thousand to -99 per thousand), this confirms its biological origin and points to in situ formation of this biogenic gas. Using (14)C-labeled substrates, the possibility of methane formation within permafrost was experimentally shown, as confirmed by delta(13)C values. Extremely low values (near -99 per thousand) indicate that the process of CH(4) formation is accompanied by the substantial fractionation of carbon isotopes. For the first time, cultures of methane-forming archaea, Methanosarcina mazei strain JL01 VKM B-2370, Methanobacterium sp. strain M2 VKM B-2371 and Methanobacterium sp. strain MK4 VKM B-2440 from permafrost, were isolated and described.
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http://dx.doi.org/10.1111/j.1574-6941.2007.00315.xDOI Listing
July 2007

Evidence of influenza a virus RNA in siberian lake ice.

J Virol 2006 Dec 11;80(24):12229-35. Epub 2006 Oct 11.

Department of Biological Sciences, 217 Life Sciences Building, Bowling Green State University, Bowling Green, OH 43403, USA.

Influenza A virus infects a large proportion of the human population annually, sometimes leading to the deaths of millions. The biotic cycles of infection are well characterized in the literature, including in studies of populations of humans, poultry, swine, and migratory waterfowl. However, there are few studies of abiotic reservoirs for this virus. Here, we report the preservation of influenza A virus genes in ice and water from high-latitude lakes that are visited by large numbers of migratory birds. The lakes are along the migratory flight paths of birds flying into Asia, North America, Europe, and Africa. The data suggest that influenza A virus, deposited as the birds begin their autumn migration, can be preserved in lake ice. As birds return in the spring, the ice melts, releasing the viruses. Therefore, temporal gene flow is facilitated between the viruses shed during the previous year and the viruses newly acquired by birds during winter months spent in the south. Above the Arctic Circle, the cycles of entrapment in the ice and release by melting can be variable in length, because some ice persists for several years, decades, or longer. This type of temporal gene flow might be a feature common to viruses that can survive entrapment in environmental ice and snow.
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http://dx.doi.org/10.1128/JVI.00986-06DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1676296PMC
December 2006

Bacterial community in ancient Siberian permafrost as characterized by culture and culture-independent methods.

Astrobiology 2006 Jun;6(3):400-14

Center for Microbial Ecology, Michigan State University, East Lansing, Michigan, USA.

The microbial composition of ancient permafrost sediments from the Kolyma lowland of Northeast Eurasia was examined through culture and culture-independent approaches. These sediments have been continuously frozen for 5,000 to 2-3 million years. A total of 265 Bacteria 16S rRNA gene sequences were amplified from the permafrost total-community genomic DNA and screened by amplified ribosomal 16S rRNA restriction analysis. Members of three major lineages were found: gamma-Proteobacteria (mostly Xanthomonadaceae), Actinobacteria, and Firmicutes. We also determined partial 16S rRNA gene sequences of 49 isolates from a collection of 462 aerobes isolated from these sediments. The bacteria included Actinomycetales (Arthrobacter and Microbacteriaceae); followed by the Firmicutes (Exiguobacterium and Planomicrobium); the Bacteroidetes (Flavobacterium); the gamma-Proteobacteria (Psychrobacter); and the alpha-Proteobacteria (Sphingomonas). Both culture and culture-independent approaches showed the presence of high and low G+C Gram-positive bacteria and gamma-Proteobacteria. Some of the 16S rRNA gene sequences of environmental clones matched those of Arthrobacter isolates. Two-thirds of the isolates grew at -2.5 degrees C, indicating that they are psychroactive, and all are closely related to phylogenetic groups with strains from other cold environments, mostly commonly from Antarctica. The culturable and non-culturable microorganisms found in the terrestrial permafrost provide a prototype for possible life on the cryogenic planets of the Solar System.
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http://dx.doi.org/10.1089/ast.2006.6.400DOI Listing
June 2006

Psychrobacter cryohalolentis sp. nov. and Psychrobacter arcticus sp. nov., isolated from Siberian permafrost.

Int J Syst Evol Microbiol 2006 Jun;56(Pt 6):1285-1291

Center for Microbial Ecology and NASA Astrobiology Institute, Michigan State University, East Lansing, MI 48824, USA.

Three Gram-negative, non-motile, non-pigmented, oxidase-positive coccobacilli capable of growth at temperatures from -10 to 30 degrees C and salinities of 0 to 1.7 M NaCl were isolated from Siberian permafrost and characterized. Both 16S rRNA and gyrB gene sequencing studies placed the isolates in the Gammaproteobacteria within the genus Psychrobacter. However, with higher bootstrap values and reproducible tree topologies, gyrB represented a more reliable phylogenetic marker for the taxonomy of Psychrobacter species. DNA-DNA hybridization data supported gyrB tree topologies and established two relatedness groups within the three isolates; neither of these groups was related at the species level to any previously described Psychrobacter species. The two groups of isolates could be differentiated phenotypically from 13 previously described Psychrobacter species using API strips. These results support the existence of two novel species of Psychrobacter, for which we propose the names Psychrobacter cryohalolentis sp. nov. (type strain K5(T) = DSM 17306(T) = VKM B-2378(T)) and Psychrobacter arcticus sp. nov. (type strain 273-4(T) = DSM 17307(T) = VKM B-2377(T)).
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http://dx.doi.org/10.1099/ijs.0.64043-0DOI Listing
June 2006

Crosslinks rather than strand breaks determine access to ancient DNA sequences from frozen sediments.

Genetics 2006 Jun 2;173(2):1175-9. Epub 2006 Apr 2.

Diagenesis was studied in DNA obtained from Siberian permafrost (permanently frozen soil) ranging from 10,000 to 400,000 years in age. Despite optimal preservation conditions, we found the sedimentary DNA to be severely modified by interstrand crosslinks; single- and double-stranded breaks; and freely exposed sugar, phosphate, and hydroxyl groups. Intriguingly, interstrand crosslinks were found to accumulate approximately 100 times faster than single-stranded breaks, suggesting that crosslinking rather than depurination is the primary limiting factor for ancient DNA amplification under frozen conditions. The results question the reliability of the commonly used models relying on depurination kinetics for predicting the long-term survival of DNA under permafrost conditions and suggest that new strategies for repair of ancient DNA must be considered if the yield of amplifiable DNA from permafrost sediments is to be significantly increased. Using the obtained rate constant for interstrand crosslinks the maximal survival time of amplifiable 120-bp fragments of bacterial 16S ribosomal DNA was estimated to be approximately 400,000 years. Additionally, a clear relationship was found between DNA damage and sample age, contradicting previously raised concerns about the possible leaching of free DNA molecules between permafrost layers.
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http://dx.doi.org/10.1534/genetics.106.057349DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1526502PMC
June 2006

Characterization of Exiguobacterium isolates from the Siberian permafrost. Description of Exiguobacterium sibiricum sp. nov.

Extremophiles 2006 Aug 18;10(4):285-94. Epub 2006 Feb 18.

NASA Astrobiology Institute, Center for Microbial Ecology, Michigan State University, 540 Plant and Soil Science Building, East Lansing, MI, USA.

Three Gram-positive bacterial strains, 7-3, 255-15 and 190-11, previously isolated from Siberian permafrost, were characterized and taxonomically classified. These microorganisms are rod-shaped, facultative aerobic, motile with peritrichous flagella and their growth ranges are from -2.5 to 40 degrees C. The chemotaxonomic markers indicated that the three strains belong to the genus Exiguobacterium. Their peptidoglycan type was A3alpha L-Lys-Gly. The predominant menaquinone detected in all three strains was MK7. The polar lipids present were phosphatidyl-glycerol, diphosphatidyl-glycerol and phosphatidyl-ethanolamine. The major fatty acids were iso-C13:0, anteiso-C13:0, iso-C15:0, C16:0 and iso-C17:0. Phylogenetic analysis based on 16S rRNA and six diverse genes, gyrB (gyrase subunit B), rpoB (DNA-directed RNA polymerase beta subunit), recA (homologous recombination), csp (cold shock protein), hsp70 (ClassI-heat shock protein-chaperonin) and citC (isocitrate dehydrogenase), indicated that the strains were closely related to Exiguobacterium undae (DSM 14481(T)) and Exiguobacterium antarcticum (DSM 14480(T)). On the basis of the phenotypic characteristics, phylogenetic data and DNA-DNA reassociation data, strain 190-11 was classified as E. undae, while the other two isolates, 7-3 and 255-15, comprise a novel species, for which the name Exiguobacterium sibiricum sp. nov. is proposed.
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http://dx.doi.org/10.1007/s00792-005-0497-5DOI Listing
August 2006

Biodiversity of cryopegs in permafrost.

FEMS Microbiol Ecol 2005 Jun;53(1):117-28

Institute of Physicochemical and Biological Problems in Soil Science, Russian Academy of Sciences, 142290 Pushchino, Moscow Region, Russian Federation.

This study describes the biodiversity of the indigenous microbial community in the sodium-chloride water brines (cryopegs) derived from ancient marine sediments and sandwiched within permafrost 100-120,000 years ago after the Arctic Ocean regression. Cryopegs remain liquid at the in situ temperature of -9 to -11 degrees C and make up the only habitat on the Earth that is characterized by permanently subzero temperatures, high salinity, and the absence of external influence during geological time. From these cryopegs, anaerobic and aerobic, spore-less and spore-forming, halotolerant and halophilic, psychrophilic and psychrotrophic bacteria, mycelial fungi and yeast were isolated and their activity was detected below 0 degrees C.
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http://dx.doi.org/10.1016/j.femsec.2005.02.003DOI Listing
June 2005

Beringian paleoecology inferred from permafrost-preserved fungal DNA.

Appl Environ Microbiol 2005 Feb;71(2):1012-7

Microbial Discovery, Novozymes A/S, Smørmosevej 25 1B1, DK-2880 Bagsvaerd, Denmark.

The diversity of fungi in permanently frozen soil from northeastern Siberia was studied by culture-independent PCR amplification of diverse environmental 18S rRNA genes. Elaborate protocols to avoid contamination during drilling, sampling, and amplification were used. A broad diversity of eukaryotic DNA sequences that were 510 bp long, including sequences of various fungi, plants, and invertebrates, could be obtained reproducibly from samples that were up to 300,000 to 400,000 years old. The sequences revealed that ancient fungal communities included a diversity of cold-adapted yeasts, dark-pigmented fungi, plant-parasitic fungi, and lichen mycobionts. DNA traces of tree-associated macrofungi in a modern tundra sample indicated that there was a shift in fungal diversity following the last ice age and supported recent results showing that there was a severe change in the plant composition in northeastern Siberia during this period. Interestingly, DNA sequences with high homology to sequences of coprophilic and keratinophilic fungi indicated that feces, hair, skin, and nails could have been sources of ancient megafauna DNA recently reported to be present in small amounts of Siberian permafrost sediments.
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http://dx.doi.org/10.1128/AEM.71.2.1012-1017.2005DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC546757PMC
February 2005

Diverse plant and animal genetic records from Holocene and Pleistocene sediments.

Science 2003 May 17;300(5620):791-5. Epub 2003 Apr 17.

Department of Evolutionary Biology, Zoological Institute, University of Copenhagen, Universitetsparken 15, Denmark DK-2100 Ø.

Genetic analyses of permafrost and temperate sediments reveal that plant and animal DNA may be preserved for long periods, even in the absence of obvious macrofossils. In Siberia, five permafrost cores ranging from 400,000 to 10,000 years old contained at least 19 different plant taxa, including the oldest authenticated ancient DNA sequences known, and megafaunal sequences including mammoth, bison, and horse. The genetic data record a number of dramatic changes in the taxonomic diversity and composition of Beringian vegetation and fauna. Temperate cave sediments in New Zealand also yielded DNA sequences of extinct biota, including two species of ratite moa, and 29 plant taxa characteristic of the prehuman environment. Therefore, many sedimentary deposits may contain unique, and widespread, genetic records of paleoenvironments.
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http://dx.doi.org/10.1126/science.1084114DOI Listing
May 2003

Reproduction and metabolism at - 10 degrees C of bacteria isolated from Siberian permafrost.

Environ Microbiol 2003 Apr;5(4):321-6

Department of Earth Sciences, University of Southern California, 3651 Trousdale Parkway, Los Angeles, CA 90089-0740, USA.

We report the isolation and properties of several species of bacteria from Siberian permafrost. Half of the isolates were spore-forming bacteria unable to grow or metabolize at subzero temperatures. Other Gram-positive isolates metabolized, but never exhibited any growth at - 10 degrees C. One Gram-negative isolate metabolized and grew at - 10 degrees C, with a measured doubling time of 39 days. Metabolic studies of several isolates suggested that as temperature decreased below + 4 degrees C, the partitioning of energy changes with much more energy being used for cell maintenance as the temperature decreases. In addition, cells grown at - 10 degrees C exhibited major morphological changes at the ultrastructural level.
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http://dx.doi.org/10.1046/j.1462-2920.2003.00419.xDOI Listing
April 2003

Aspartic acid racemization and age-depth relationships for organic carbon in Siberian permafrost.

Astrobiology 2002 ;2(1):77-82

Jet Propulsion Laboratory, California Institute of Technology, Pasadena, CA, USA.

We have analyzed the degree of racemization of aspartic acid in permafrost samples from Northern Siberia, an area from which microorganisms of apparent ages up to a few million years have previously been isolated and cultured. We find that the extent of aspartic acid racemization in permafrost cores increases very slowly up to an age of approximately 25,000 years (around 5 m in depth). The apparent temperature of racemization over the age range of 0-25,000 years, determined using measured aspartic acid racemization rate constants, is -19 degrees C. This apparent racemization temperature is significantly lower than the measured environmental temperature (-11 to -13 degrees C) and suggests active recycling of D-aspartic acid in Siberian permafrost up to an age of around 25,000 years. This indicates that permafrost organisms are capable of repairing some molecular damage incurred while in a "dormant" state over geologic time.
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http://dx.doi.org/10.1089/153110702753621358DOI Listing
December 2002

Phylogenetic diversity of a bacterial community determined from Siberian tundra soil DNA.

Microbiology (Reading) 1997 Dec;143 ( Pt 12):3913-3919

Center For Microbial Ecology, Michigan State University,East Lansing, MI 48824-1325,USA.

Genomic DNA was isolated from the active layer of tundra soil collected from the Kolyma lowland, Northeast Eurasia, near the Arctic Ocean coast. The SSU (small subunit) rRNA genes were amplified with eubacterial primers from the bulk genomic community DNA and cloned into plasmid vectors. Forty-three SSU rDNA clones were obtained, and all of them had different RFLP patterns. Phylogenetic analysis based on partial sequences (about 300 bp) established with the maximum likelihood method revealed the presence of three major and several minor groups that fell into 11 of the established lines of bacteria, and one sequence that could not be assigned to any of the described groups. Most of the clones belonged to the alpha (20.9%) and delta (25.6%) subdivisions of the Proteobacteria, with lesser proportions in the beta (9.3%) and gamma (4.7%) subdivisions, groups typically isolated from soil by culture methods. Fewer than 12% of the clones belonged to Gram-positive bacteria, and 16% of the clones were related to Fibrobacter. The majority of the clones (70%) had sequences that were 5-15% different from those in the current databases, and 7% of the clones had sequences that differed by more than 20% from those in the database. The results suggest that these tundra-derived clones are very diverse in phylogeny, and that many probably reflect new genera or families. Hence, most of the tundra soil bacterial community has never been isolated and thus the physiology and function of its dominant members appears to be unknown.
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http://dx.doi.org/10.1099/00221287-143-12-3913DOI Listing
December 1997
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