E.A. Vorobyova

Long-term preservation of microbial ecosystems in permafrost.

It has been established that significant numbers (up to 10 million cells per gram of sample) of living microorganisms of various ecological and morphological groups have been preserved under permafrost conditions, at temperatures ranging from -9 to -13 degrees C and depths of up to 100 m, for thousands and sometimes millions of years. Preserved since the formation of permafrost in sand-clay sediments of the Pliocene-Quaternary period and in paleosols and peats buried among them, these cells art the only living organisms that have survived for a geologically significant period of time. The complexity of the microbial community preserved varies with the age of the permafrost. Eukaryotes are found only in Holocene sediments; while prokaryotes are found to greater ages, i.e., Pliocene and Pleistocene. The diversity of microorganisms decreases with increasing age of sediments, and as a result cocci and corynebacteria are predominant. Enzyme activity (catalase and hydrolytic enzymes) and photosynthetic pigments (chlorophyll and pheophytin have also been detected in permafrost sediments. These results permit us to outline some approaches to the search for traces of life in the permafrost of Martian sediments by borehole core sampling. It is in the deep horizons (and not on the planet surface), isolated by permafrost from the external conditions, that results similar to those obtained on Earth can be expected.

Microorganisms and enzyme activity in permafrost after removal of long-term cold stress

Abstract Associations of immobilized microbial cells and organic-mineral complexes containing active enzymes are resistant to long-term (from tens of thousands to millions of years) effects of extremely low temperatures. This association enables the cells to restore their metabolic activity during permafrost thawing, because interactions with the heterogenous medium is made possible by the availability of active immobilized enzymes. The long-term effect of the cold probably favors an adaptational change of microbial metabolism that activates enzymes and cells during thawing.

The resistance of viable permafrost algae to simulated environmental stresses: implications for astrobiology

54 strains of viable green algae and 26 strains of viable cyanobacteria were recovered from 128 and 56 samples collected from Siberian and Antarctic permafrost, respectively, with ages from modern to a few million years old. Although species of unicellular green algae belonged to Chlorococcales were subdominant inside permafrost, green algae Pedinomonas sp . were observed in Antarctic permafrost. Filamentous cyanobacteria of Oscillatoriales , Nostocales were just found in Siberian permafrost. Algal biomass in the permanently frozen sediments, expressed as concentration of chlorophyll a , was 0.06–0.46 μg g −1 . The number of viable algal cells varied between 2 and 9×10 3 cfu g −1 , but the number of viable bacterial cells was usually higher from 10 2 to 9.2×10 5 cfu g −1 . Frozen but viable permafrost algae have preserved their morphological characteristics and photosynthetic apparatus in the dark permafrost. In the laboratory, they restored their photosynthetic activity, growth and development in favourable conditions at positive temperatures and with the availability of water and light. The discovery of ancient viable algae within permafrost reflects their ability to tolerate long-term freezing. In this study, the tolerance of algae and cyanobacteria to freezing, thawing and freezing–drying stresses was evaluated by short-term (days to months) low-temperature experiments. Results indicate that viable permafrost microorganisms demonstrate resistance to such stresses. Apart from their ecological importance, the bacterial and algal species found in permafrost have become the focus for novel biotechnology, as well as being considered proxies for possible life forms on cryogenic extraterrestrial bodies.

Microbial communities of ancient seeds derived from permanently frozen Pleistocene deposits

Microbial communities from the surface of ancient seeds of higher plants and embedding frozen material dated to the late Pleistocene (formed about 30 thousand years ago) were studied by various methods: scanning electron microscopy, epifluorescence microscopy, and inoculation of nutrient media, followed by identification of isolated cultures. Both prokaryotic and eukaryotic microorganisms were found on the surface of ancient seeds. The total quantity of bacterial cells determined by direct counting and dilution plating (CFU) for the samples of ancient seeds exceeded the value in the embedding frozen material by one to two orders of magnitude. This pattern was not maintained for mycelial fungi; their quantity in the embedding material was also rather high. A significant difference was revealed between the microbial communities of ancient seeds and embedding frozen material. These findings suggest that ancient plant seeds are a particular ecological niche for microorganisms existing in permafrost and require individual detailed study.

Micro-organisms and biomarkers in permafrost

Studies of microorganisms populating subsoil layers skipped the latent period (individual communications in the first half of this century), passed through the lagphase of accumulation of reliable information and overcoming of sound scepsis and reached eventually the stage of exponential growth. Today, we can state with certainty the ever-increasing interest shown by not only biologists but also experts from various natural sciences in the progress of studies in this field. This interest is related to changes in our concepts of extreme habitats and extremophilia of microorganisms under conditions of a heterophase medium. It becomes obvious that the overwhelming majority of microorganisms exist under the ‘extreme’ conditions of lowered and often negative temperatures, high pressure, low concentrations of accessible nutrients, oxygen, physiological dryness, etc. It becomes clear that the reserves of biomass in the interior of the earth abounding in ‘extreme’ habitats are many times the overall biomass of the surface and subsurface soil layers. The subject of this discussion is metabolic activity and, naturally, the possibility of cell adaptation to such a medium, i.e. in essence, the advisability of the ecological term ‘extreme medium’ in its application to viable microorganisms populating these biotopes under long-term stability of medium parameters.

Viable bacteria, methane and high ice content in antarctic permafrost: Relevance to Mars

1Florida State Univ., Tallahassee, FL 32306-2043, USA, FAX: 1-904-644-9829, email: friedm@bio.fsu.edu; 2Lab. Soil Cryol., Russian Acad. Sei., Puschino, Moscow Region 142292, RUSSIA, FAX: 7-096-779-0532, e-mail: gilichin@issp.sherpukov.su; aByrd Polar Res. Ctr., Ohio State Univ., Columbus, OH 43210-1002, USA, FAX: 1614-292-4697, e-mail: wilsongs@geology.ohio-state.edu, 4Dept. Soil Biol., Moscow State Univ., 119899 Moscow, RUSSIA, e-mail: rott@soina.msk.ru; 5Dept. Oceanogr., Florida State University, Tallahassee, FL 32306-3015, USA, FAX: 1-904-644-2581, e-mail: jchanton@mailer.fsu.edu; ~Department of Biology, Florida A. and M. Univ., Tallahassee, FL 32307, USA, FAX: 1-904-561-2996, e-mail: friedm@bio.fsu.edu; 7NASA-Ames Res. Ctr., Mail Stop 245-3, Moffett Field, CA 93035, USA, FAX: 1415-604-6776, e-mail: mckay@galileo.arc.nasa.gov.

Role of cell differentiation in high tolerance by prokaryotes of long-term preservation in permafrost

Abstract The effect of low temperature on the cell structure of bacteria isolated from permafrost results in structural changes leading to cell differentiation into types of resting cells rangign from spores showing a high endogenous dormancy to typical “dormant” cells of non-spore-forming bacteria showing exogenous dormancy, which is considered to be less highly resistant to extreme conditions in laboratory experiments. In permafrost, dormant cells of non-spore-forming bacteria may demonstrate considerable resistance to long-term freezing and as a result a higher survival level than spore-forming bacteria.

Detection of microbial cells and preliminary estimation of their physiological state by x-ray microanalysis

The method of X-ray microanalysis was used for the detection of microbial cells among cell-like particles immediately in the samples of 170 thousand year old ancient Antarctic permafrost sediments. The X-ray spectra and the quantitative parameters of P, S, Ca and K contents in cell-like particles, as well as ratios Ca/K and P/S, were compared with the corresponding data obtained for microbial cells of various physiological states: vegetative cells -- resting forms -- non-viable micromummies. The absence of P and S peaks in X-ray spectra in the most of cell-like particles allowed us to regard them as non-living objects. Among other investigated cell-like particles we were able to find the resting forms of microorganisms with the increased intracellular level of Ca, high Ca/K ratio and low P/S ratio. So, X-ray microanalysis is a promising tool for a primary detection of microbial cells in situ and determination of their physiological state.

Microbial activity in Martian analog soils after ionizing radiation: implications for the preservation of subsurface life on Mars

At present, the surface of Mars is affected by a set of factors that can prevent the survival of Earth-like life. However, the modern concept of the evolution of the planet assumes the existence more favorable for life climate in the past. If in the past on Mars had formed a biosphere, similar to the one that originated in the early Earth, it is supposed that it is preserved till now in anabiotic state in the bowels of the planet, like microbial communities inhabiting the ancient permafrost of Arctic and Antarctic. In the conditions of modern Martian regolith, this relic life seems to be deprived of the possibility of damage reparation (or these processes occur on a geological time scale), and ionizing radiation should be considered the main factor inhibiting such anabiotic life. In the present study, we studied soil samples, selected in two different extreme habitats of the Earth: ancient permafrost from the Dry Valleys of Antarctica and Xerosol soil from the mountain desert in Morocco, gamma-irradiated with 40 kGy dose at low pressure (1 Torr) and low temperature (−50 °C). Microbial communities inhabiting these samples showed in situ high resistance to the applied effects, retained high number of viable cells, metabolic activity, and high biodiversity. Based on the results, it is assumed that the putative biosphere could be preserved in the dormant state for at least 500 thousand years and 8 million years in the surface layer of Mars regolith and at 5 m depth, respectively, at the current level of ionizing radiation intensity.

Europa Lander Mission: A Challenge to Find Traces of Alien Life

Keldysh Institute of Applied Mathematics, Russiaemail: korab@iki.rssi.ruAbstract.An international effort dedicated to science exploration of Jupiter system plannedby ESA and NASA in the beginning of next decade includes in-depth science investigation ofEuropa. In parallel to EJSM (Europa-Jupiter System Mission) Russian Space Agency and theacademy of Science plan Laplace-Europa Lander mission, which will include the small telecom-munication and scienceorbiter and the surface element: Europa Lander. In-situ methods on thelander provide the only direct possibility to assess environmental conditions, and to performthe search for signatures of life. A critical advantage of such in situ analysis is the possibilityto enhance concentration and detection limits and to provide ground truth for orbital measure-ments. The science mission of the lander is biological, geophysical, chemical, and environmentalcharacterizations of the Europa surface. Remote investigations from the orbit around Europawould not be sufficient to address fully the astrobiology, geodesy, and geology goals. The scienceobjectives of the planned mission, the synergy between the Europa Lander and EJSM missionelements, and a brief description of the Laplace-Europa Lander mission are presented.Keywords.Europa; Jupiter; Galilean satellites; habitability; space research.