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Abstract

Permafrost and ice are important components of cryogenic planets and other bodies, which are abundant in the universe. Earth is one of many planets of the cold type. Earth's permafrost and ice provide an opportunity to test hypotheses that could be applied in the search for possible ecosystems and potential life on extraterrestrial cryogenic planets. Permafrost sediments in polar and alpine regions are natural ecosystems with a unique feature of low‐temperature preservation of the biological material and its genetic information. Therefore, permafrost studies allow reconstruction of the events that occurred in the Cenozoic and the prediction of possible life that might have been preserved before the effect of anthropogenic factors on these ecosystems, and could be found within ice or permafrost on other cryogenic planets.
RESEARCH ARTICLE
Earth's perennially frozen environments as a model of
cryogenic planet ecosystems
Elizaveta Rivkina
1
|Andrey Abramov
1
|Elena Spirina
1
|Lada Petrovskaya
2
|
Anastasia Shatilovich
1
|Lyubov Shmakova
1
|Viktoria Scherbakova
3
|
Tatiana Vishnivetskaya
1,4
1
Russian Academy of Sciences, Institute of
Physicochemical and Biological Problems in
Soil Science, Pushchino, Russia
2
Russian Academy of Sciences, Shemyakin
and Ovchinnikov Institute of Bioorganic
Chemistry, Moscow, Russia
3
Russian Academy of Sciences, Skryabin
Institute of Biochemistry and Physiology of
Microorganisms, Pushchino, Russia
4
University of Tennessee, Knoxville,
Tennessee, USA
Correspondence
Elizaveta Rivkina, Institute of Physicochemical
and Biological Problems in Soil Science,
Russian Academy of Sciences, Pushchino,
142290, Russia.
Email: elizaveta.rivkina@gmail.com
Funding information
National Science Foundation, Grant/Award
Number: DEB1442262; Presidium of the
Russian Academy of Sciences Program 22;
Russian Government Assignment, Grant/
Award Number: # AAAAA18
1180131901816
Abstract
Permafrost and ice are important components of cryogenic planets and other bodies,
which are abundant in the universe. Earth is one of many planets of the cold type.
Earth's permafrost and ice provide an opportunity to test hypotheses that could be
applied in the search for possible ecosystems and potential life on extraterrestrial
cryogenic planets. Permafrost sediments in polar and alpine regions are natural eco-
systems with a unique feature of lowtemperature preservation of the biological
material and its genetic information. Therefore, permafrost studies allow reconstruc-
tion of the events that occurred in the Cenozoic and the prediction of possible life
that might have been preserved before the effect of anthropogenic factors on these
ecosystems, and could be found within ice or permafrost on other cryogenic planets.
KEYWORDS
permafrost, cryopegs, microorganisms, astrobiology, review
1|INTRODUCTION
Perennially frozen environments are widely distributed on Earth with
permafrost occupying about 25% of the land surface.
1
The Phanero-
zoic terrestrial climate record, based on δ
18
O and δ
13
C measurements
for lowmagnesium calcite shells of marine fossils, shows prominent
temperature oscillations in Earth's history.
2
Climate changes affect
the expansion and subsequent shrinkage of polar and continental ice
sheets as well as repeated formation and thawing of the perennially
frozen ground. The permafrost history of the high northern latitudes
over the last two million years indicates that permafrost formed and
thawed repeatedly.
3
However, there is convincing evidence to suggest
that much of today's Siberian Arctic permafrost originated in cold con-
tinental climate conditions during the Late Pliocene and Pleistocene
epochs.
4
Permafrost is icecemented ground that sometimes also con-
tains ice wedges and ice veins. Under conditions of climate change
and apparent increases in ice melting and permafrost degradation,
Cenozoic microorganisms, their genetic material including RNA and
DNA fragments from dead cells, extracellular products of microbial
metabolism, pigments, proteins, and intracellular and extracellular
enzymes would be integrated into modern biomes affecting biodiver-
sity, biogeochemical processes, circulation of nutrients and green-
house gas formation.
5
The process of permafrost degradation by
Arctic rivers is ongoing and might be accelerated due to global
warming, although the ecological impact of the continual release of
ancient permafrost in aquatic systems has not been well studied.
Unknown life forms and genetic resources buried in permafrost will
be revealed as modern stateoftheart omics approaches emerge for
studying microbial communities. Besides providing practical knowl-
edge and having biotechnological applications, comprehensive study
of ancient permafrost sediments sheds light on some important funda-
mental questions. Studies from the oldest terrestrial permafrost have
shown preservation of microbial cells for a few million years.
6,7
The
metabolic state of microorganisms in permafrost under conditions of
Received: 10 April 2018 Revised: 3 July 2018 Accepted: 8 July 2018
DOI: 10.1002/ppp.1987
Permafrost and Periglac Process. 2018;111. © 2018 John Wiley & Sons, Ltd.wileyonlinelibrary.com/journal/ppp 1
constant low temperatures remains a mystery. Many studies are based
on the premise that samples from deep, permanently frozen sediments
represent a fossil archive of past surface communities.
8,9
However,
accumulating evidence is also consistent with the possibility that via-
ble cells in ancient sediments continue to function in ecologically sig-
nificant ways.
10-13
These microbial cells, which are metabolically
active and likely to perform biochemical reactions of DNA repair, pass
down to descendants any adaptive features and their genetic modifi-
cations. The latter scenario would reveal ample genetic differences
between modern and permafrost paleomicroorganisms. Another line
of research is concerned with quantitative and qualitative assessment
of organic matter confined within ancient permafrost sediments, per-
mafrost temperature monitoring, the impact of the underlying perma-
frost on the formation of modern landscapes, the rate of organic
matter transformation in modern permafrostaffected soils, and
changes in greenhouse gas emissions due to permafrost degradation.
Studies that focus on the astrobiological aspects of permafrost
research will provide help in the design of biological experiments in
the search for evidence of life on cryogenic planets.
2|CRYOSPHERE
The cryosphere consists of those sections of the Earth's surface where
water is in solid form and is subdivided into two parts: glaciosphere
(glaciers, ice sheets, ice and snow cover) and cryolithosphere (frozen
ground, which includes seasonally frozen soils and ancient
permafrost deposits) (Figure 1). Cryobiosphere refers to extremely
cold environments and the organisms living in it. Therefore, in addition
to snow, ice and permafrost, the cryobiosphere also includes
supercooled watersaturated clouds
14
and the upper atmosphere.
15
Thus, the cryobiosphere is a unique part of the biosphere occupying
a vast area from the upper atmosphere to the foot of the Antarctic
ice sheet.
16
The source of microorganisms in the cryolithosphere is
the overlying soil cover and coldwater ecosystems whereas in the
glaciosphere the source is cold water, the atmosphere and via atmo-
spheric transfer.
There are a number of environments in the biosphere that could
preserve viable microorganisms over geological time. Comparisons of
the number of viable microorganisms preserved in galite
17-19
and
amber
20
at temperatures above 0°Сwith those preserved in the
cryosphere at temperatures below 0°Сhave shown that the cryosphere
hosts a larger amount of extremely diverse viable microorganisms.
16
2.1 |Glaciosphere
The glaciosphere and its application to astrobiology have been
described previously.
21
A wide range of data on the microbial popula-
tions in modern and ancient ice can be found in the review Biosphere
ice
22
and in the monograph Life in Ancient Ice.
23
The glaciosphere is
the totality of snowice formations on Earth including frozen water
present in permafrost in the form of ice veins and ice wedges.
Ice sheets and their biota have been extensively studied at depths
of up to 3 km in Greenland and even deeper in the Antarctic.
24-28
The
age of the oldest ice as well as the age of the oldest bacteria it con-
tains are subjects of current scientific debate. The maximum age of
pure ice in Greenland was estimated to be about 200 000 years
29
and the silty ice at the bottom 400850 000 years.
30
Ice older than
500 000 years was found at the Gulia ice cap on the Tibetan pla-
teau.
31,32
Ice with an estimated age of 2 million years was detected
at the bottom of the Antarctic ice sheet around Lake Vostok.
33
The
oldest ice with overlying ash is 8.1 million years old and was found
in Antarctica's Beacon Valley.
34,35
It was shown that the main path of microorganism entry into ice is
via wind transfer. The rate of microbial wind transfer could be inferred
from the number of microorganisms in seasonal snow and ice cover,
which is usually less than 10
2
cells g
1
. Approximately the same num-
ber of viable microorganisms was observed in the cores of ancient ice
sheets from different locations. Nevertheless, the number of microbial
cells in the drilling well in the ice sheet on Lake Vostok decreases in
FIGURE 1 Different parts of the cryosphere: the glaciosphere and the cryolithosphere
2RIVKINA ET AL.
the deeper layers as the age of the ice increases.
36
Small cells less than
3μm in size dominate the microbial population of glacial ice.
27
How-
ever, the number of microbial cells increases drastically with increasing
ice dustiness and in silty ice.
26,36
Permafrost ice wedge samples with
0.2% mineral components contained a relatively abundant number of
culturable cells compared to samples from other ice habitats.
37
Bacte-
rial and plant viruses can survive long periods of time in ice and were
isolated from 500and 100 000yearold Greenland ice.
23
Unfortu-
nately, this is also true of human viruses. It has been shown that
RNA of the influenza A virus was preserved at high concentration in
seasonal lake ice in the Arctic.
38,39
Veins of liquid water within the
ice as well as layers with a high content of mineral particles and, con-
sequently, thin films of unfrozen water, are habitats occupied by
microorganisms in this ecosystem.
40
DNA fragments have been isolated from young Greenland ice
with an age of 20004000 years
41
and from the silty ice of the lower
part of the Greenland ice sheet with an age of 450800 000 years.
30
2.2 |Cryolithosphere
The most populated part of the cryosphere is the cryolithosphere, or
permafrost, which occupies onequarter of the Earth's land surface.
Permafrost is an integral part of the Arctic, including the northern part
of North America and Eurasia as well as icefree areas of Greenland,
and Antarctica. Alpine, or mountain, permafrost is widespread at high
altitudes in mountains of all continents: Europe, Asia (Western China),
and North and South America. In addition, coastal (shelf) permafrost is
found in the Arctic Ocean and around Antarctica.
In the Siberian Arctic, formation of the permafrost started about
23 million years ago during the transition of vegetation from forest
to tundra, based on pollen and other records of terrestrial flora, marine
fauna, and ice activity.
42
The first cooling period was superseded by a
warm climate followed by a second cooling period, which began 1.7
million years ago. The climate of the second cooling period fluctuated
with a return of warmer epochs about every 100 000300 000
years.
43
The most significant warm periods marked in the Lake
El'gygytgyn borehole were 1.1, 0.4 and 0.01 million years ago.
44
Based
on these data the ages of the most ancient perennially frozen deposits
in the Northern Hemisphere are estimated to be from 1.1 to 2.8 mil-
lion years old. A slow rate freezing or numerous repetitive warming
cooling cycles resulted in the formation of the cryolithozone in its
present form, where layers that have not thawed over at least the last
2 million years can be found.
There are two main types of permafrost formationsyngenetic,
where the freezing of sediments occurred simultaneously with sedi-
mentation, and epigenetic, where freezing followed sedimentation.
Therefore, the age of syngenetic permafrost is equal to the age of the
sediments it contains; however, in epigenetic sediments the age of per-
mafrost usually reflects the last cooling period. The best indicator of
syngenetic permafrost is the presence of ice wedges, while epigenetic
permafrost has a low ice content. In both types of permafrost, lateral
and vertical migration are negligible, and thus bacteria in those deposits
are as old as the last freezing. The oldest syngenetic deposits known in
North America are in the Yukon River Valley. Tephrochronology studies
have dated these frozen deposits to be 740 000 ± 60 000 years old.
45,46
In Antarctica, the transition to tundra occurred around 1520 mil-
lion years ago,
47
followed by midMiocene cooling and continental
glaciation.
48
2.2.1 |Permafrost as an ecosystem
The primary characteristics used in the description of permanently fro-
zen sediments are subzero temperatures, measurements of ice con-
tent, and estimation of the time period during which deposits stay
frozen. Ancient permafrost deposits are several hundred meters thick
and underlie seasonally frozen cryosols. Permafrost and permafrost
affected soils contain ancient and modern viable microorganisms,
respectively. In permafrost sediments microbial cells of different forms
are adsorbed on organic and mineral particles. The number of microor-
ganisms reaches several million cells per gram. The temperature range
of the frozen deposits where viable microorganisms were detected is
quite wide: from deposits with a temperature of 1/2°C near the
southern permafrost boundary in Siberia and in some icefree oases
of the Antarctic coast, for example the Bellingshausen art area
49
to
the lowest temperature of 17/18°C on Ellesmere island, Canada
(80°N)
50
and 27°C in Antarctica's Dry Valleys (78°S).
7
Viable microorganisms have been found in permafrost up to
depths of 400 m in the delta of the Mackenzie River, Canada,
51,52
and at the altitude of 4700 m on the T ibetan Plateau.
53,54
The age
of microbial cells embedded in permafrost corresponds to the duration
of the depositsfrozenstate, which varies from a few thousand to mil-
lion years in northeastern Siberia, and up to 5.8 million years in Ant-
arctica.
7,34
Viable microorganisms, a huge mass of live material, are a
unique feature of permafrost sediments. Taking into account the
thickness of the cryolithosphere from 50 to 1000 m, it may be con-
cluded that permafrost contains many more microorganisms adapted
to low temperatures as compared with modern soils.
The total number of microorganisms as determined by
epifluorescence microscopy is 10
7
10
8
cells g
1
in Siberian and Cana-
dian Arctic permafrost.
55
In the permafrost deposits of the Antarctic
Dry Valleys, the total number of microbial cells is 10
5
10
6
cells g
1
,
two orders of magnitude less than in the Arctic.
7
Overall, the Antarctic
permafrost exhibits much less diversity and a lower total number of
viable microorganisms grown on nutrient media compared to that of
the Arctic permafrost. Microbiological analysis of permafrost samples
from various regions has shown that the number of viable bacteria
able to grow on nutrient media varies from 10
3
to 10
6
cells g
1
, equiv-
alent to 0.011.5% of the total number of microorganisms as deter-
mined by fluorescence microscopy.
6
Note that most microorganisms
from permafrost deposits show optimal growth at room temperature
and, according to the Morita classification,
56,57
have been considered
to belong to psychrotrophs or to be psychroactive. Nevertheless, the
microorganisms indigenous to permanently frozen environments could
be described as psychrophiles based on their existence at subzero
temperatures and adaptation to cold.
58
2.2.2 |Unfrozen water in permafrost and cryopegs
Water in permafrost exists mostly in solid phase or as ice, which
accounts for 9398% of all the water. While free water is not available
in permanently frozen sediments, soсalled unfrozen water occurs that
RIVKINA ET AL.3
accounts for 1.57% of all the water. The number and thickness of
unfrozen brine water films depend on temperature and particle size
distribution in the permafrost deposits.
59-61
These thin films of unfro-
zen water enveloping the mineral and organic particles prevent micro-
bial cells from mechanical destruction by ice crystals, provide
conditions for metabolic reactions and possibility allow removal of
the end products.
40
Therefore, the unfrozen water films protect
microbial cells from both mechanical and biochemical death, thereby
maintaining microbial viability over geological time, from a few thou-
sand to millions of years. Thus, the unfrozen water films are character-
ized as a major ecological niche for microorganisms.
62
Because these
water films are not thicker than several nanometers
61
and are consid-
erably smaller than the size of the microbial cells, cell growth, division
(ie, reproduction) and movement within the films are unlikely.
16
The
amount of unfrozen water and the thickness of the water films are
independent of ice content. At the same time, they are affected by
the temperature of permafrost deposits, decreasing in amount and
size with a decrease in temperature.
59-61
Besides biotacontaining per-
manently frozen sediments of various origin, cryopegs represent
another important permafrostembedded closed ecosystem. Cryopegs
are the only freewatercontaining formations within permafrost char-
acterized by subzero temperatures, high salinity and complete isola-
tion from external factors. These unique formations of supercooled
free water lenses with mineralization up to 250 g L
1
(see ref
63
)
may be found in frozen deposits of marine origins in the Arctic
64
and continental Antarctica.
65
The physical state of water is extremely important for microor-
ganisms, because the availability of water determines whether cell
division and growth are possible. Therefore, in permafrost where
water is present only in the form of ice and thin films of unfrozen
water, the microbial cells are considered to have been unable to divide
since the sediments became frozen. Thus, the age of the microorgan-
isms is assumed to be the age of the permafrost. In the case of
cryopegs, where free water is present, cell division cannot be ruled
out, and therefore microorganisms from the overcooled brines may
be younger than the cryopegs themselves.
2.2.3 |Volcanic permafrost
In the Klyuchevskaya volcano group in central Kamchatka, volcanic
deposits found at altitudes greater than 1000 m above sea level are
permanently frozen and can reach thicknesses of more than 500 m
at summits higher than 3000 m above sea level. In polar regions, the
lower boundaries of permafrost distribution on volcanic slopes are
found at much lower elevations.
66-68
Permafrost deposits in the region
of the Klyuchevskaya volcano group are extremely fresh (no salts),
with temperatures around 3°C to 9°C, neutral pH (in a range from
5.8 to 8.1), lower than 0.2% total organic carbon, and an ice content
of 3540%. Microorganisms from the geothermal niches of volcanoes
are subject to natural cryoconservation when the temperature does
not rise above 0°C during formation of the volcanic permafrost.
68
During a recent study of the permafrost volcanic rocks collected on
Deception Island, South Shetland Islands, thermophilic bacteria grow-
ing at 5560°Сwere isolated. The number of colonyforming units was
~50 g
1
, and they belonged to the genus Geobacillus.
68
3|DIVERSITY OF ORGANISMS IN
PERMAFROST
Permafrost, including cryopegs, is a repository of diverse viable micro-
organisms represented by Grampositive and Gramnegative, aerobic,
anaerobic, sporeforming and asporogenous bacteria.
69-76
In numer-
ous permafrost deposits, primarily of lacustrine and marine origin, via-
ble methaneproducing archaea have been found.
77-80
Permafrost has been shown to contain other groups of microor-
ganisms, for example yeasts,
81,82
and mycelial fungi.
83
Photosynthetic
microorganisms, such as anaerobic purple nonsulfur bacteria,
84
fila-
mentous and unicellular cyanobacteria and green algae,
85,86
have been
isolated from permafrost deposits and described. A number of micro-
organisms isolated from permafrost and cryopegs have been taxonom-
ically identified as novel species (Table 1) and some of them have been
designated as type strains. Proteins and pigments that could serve as
biomarkers, for example intraand extracellular metabolic enzymes
such as invertase, catalase, protease and amylase,
100
pigments (chloro-
phyll aand b), pheophytin,
101,102
bacteriorhodopsin
103
and biogenic
methane,
78
have been detected in permafrost sediments.
Higher level biological materialmosses
104
and seeds
105
have
also been found in permafrost deposits. Some seeds have been suc-
cessfully grown.
106
Heterotrophic protistsamoebas, ciliates and fla-
gellates
89,107,108
have also been found in permafrost. Recently, new
giant viruses (parasites of acanthamoebae), which are representatives
of previously unknown families, Pythovirus sibericum
90
and Mollivirus
sibericum
91
(Table 1), were isolated from Arctic permafrost.
Permafrost can be considered a natural underground repository,
where physicochemical and temperature conditions remain constant
for long periods. It is the permanency of these conditions that allows
us to consider them as stable rather than extreme. At low tempera-
tures, the rate of biochemical reactions and biological processes slows
down, ensuring the preservation of biological material.
4|LIFE AT TEMPERATURES BELOW 0°C
A subzero temperature by itself is not a limiting factor to microbial
growth. In the laboratory, microorganisms are able to reproduce at
10°C when cultivated in nutrient media with added glycerol,
109
and
67% of bacterial strains isolated from permafrost grew at 2.5°C and
were able to form visible colonies within a 3week incubation period.
6
Planococcus halocryophilus strain Or1, isolated from high Arctic perma-
frost, grows and divides at 15°C, the lowest temperature demon-
strated to date, and is metabolically active at 25°C in frozen
permafrost microcosms.
110
Production of CO
2
and
14
CO
2
was
recorded at temperatures as low as 39°C from intact and
14
Cglu-
coseamended tundra soils (Barrow, Alaska) incubated for 1 year.
111
This is consistent with the lower temperature limit for microbial
growth established by Russell and Hamamoto.
112
Temperature condi-
tions of 10°C and a salinity of 200 g L
1
were found in cryopegs from
permafrost marine sediments. Metabolic activity in cryopegs was dem-
onstrated in an experiment using resazurin as a respiration indica-
tor.
113
The incorporation of D[
14
C] glucose into the cryopegs
bacterial biomass was later observed at 12°Сin laboratory
4RIVKINA ET AL.
experiments,
64,114
confirming the ability of cryopeg microorganisms to
carry out metabolic reactions.
Most researchers believe that at least some portion of the perma-
frost microbial communityabout 20%, according to Steven
115
can
grow at temperatures from 2to10°C.
71,73,113,116-119
At the same
time, 95% of the bacteria isolated from permafrost, do not grow at
all or grow very poorly at temperatures above 30°C.
62
The survival of biota in frozen and supercooled saline Cenozoic
ecosystems, as well as in PermoTriassic salt deposits
120
are examples
of the unique adaptation abilities of bacteria. It is important to under-
stand whether the salt resistance of these cells is linked to their
tolerance to low temperatures. Experiments have shown that halo-
philic bacteria survive better at low temperatures than nonhalophilic
bacteria, and that they remain viable at a temperature of 80°C in
the presence of 25% NaCl.
121,122
The main microbial community members in permafrost are
psychrohalotolerant. This may be described as a community of survi-
vors
123
characterized by their ability to enter a physiological state
known as starvationsurvival.
56,57,124
Thus, the permafrost microbial
community can be considered as an elite clubof the bacteria that
were able to adapt to the specific conditions in permafrost deposits
via an efficient repair mechanism. The latter allowed the permafrost
TABLE 1 Microorganisms in permafrost and cryopegs
Location Description
Age
(10
3
years)
Temperature
(°C)
Microorganisms
Total count
(cells g
1
)
Viable count
(cfu
a
g
1
)
Cultured microorganisms
taxonomically identified as a new
species
Northern Hemisphere
KolymaIndigirka Lowland,
Russia
Permafrost 51000 9/12 10
6
10
8
10
3
10
6
Methanogenic Archaea
Methanobacterim veterum sp. nov.
77
Methanobacterium arcticum sp. nov.
79
Prokaryotes
Carnobacterium inhibens subsp.
gilichinskyi
87
Exiguobacterium sibircum sp. nov.
71
Sphaerochaeta associata sp. nov.
88
Psychrobacter arcticus sp. nov
72
Eukaryotes: Protista
Flamella pleistocenica
89
Flamella beringiania
89
Giant viruses
Pythovirus sibericum
90
Mollivirus sibericum
91
Gydan Peninsula, Russia Permafrost 834 ––Prokaryotes
Microbacterium sp. Gd 413
92
Eukaryotes: Protista
Phalansterium arcticum
93
Cape Chukochii, Сoast of the
Kolyma Gulf, Russia
Cryopeg 100120 10 10
7
10
2
10
3
Prokaryotes
Psychrobacter cryohalolentis sp. nov.
72
Psychrobacter muriicola sp. nov.
94
Clostridium algoriphilum sp. nov.
73
Yamal Peninsula, Russia Cryopeg 100120 6–– Prokaryotes
Celerinatantimonas yamalensis sp. nov.
74
Desulfovibrio gilichinskyi sp. nov.
b
Varandey Peninsula, coast
of the Barents Sea, Russia
Cryopeg ~5 5–– Prokaryotes
Desulfovibrio arcticus sp. nov.
70
Lena River Delta, Russia Permafrost
affected soil
Modern 25 to +5 –– Methanogenic Archaea
Methanosarcina soligelidi sp. nov.
95
Alaska, Fox Tunnel, USA Permafrost 32 3–– Prokaryotes
Carnobacterium pleistocenium sp. nov.
69
High Arctic, Canada Permafrost 5716 10
7
10
3
Prokaryotes
Planococcus halocryophilus sp. nov.
96
Tumebacillus permanentifrigoris sp.
nov.
97
Clostridium tagluense sp. nov.
52
Tibetan Plateau, Western
China
Alpine permafrost 2030 ––Prokaryotes
Hymenobacter psychrotolerans sp. nov.
98
Southern Hemisphere
Dry Valleys, Antarctica Permafrost 15 27 10
5
10
6
10
1
Methanogenic Archaea
Methanosarcina sp. Ant1
99
Deception Island, Antarctica Permanently frozen
volcanic deposits
0.3 310
4
10
2
10
3
Prokaryotes
Geobacillus sp. D4455, D2355
68
a
Colonyforming units.
b
Paper is under review.
RIVKINA ET AL.5
microbial community to preserve its viability in permafrost, in contrast
to many microorganisms from modern tundra soils.
117
This may why
the bacteria found in permafrost deposits can also be found in Arctic
tundra soils
125
whereas many types of bacteria isolated from modern
Arctic soils are not found in permafrost.
126
The viability of microorganisms at low temperatures over geolog-
ical time suggests the possibility of metabolic reactions in permafrost
sediments. Indirect evidence confirms that ancient microbes can par-
ticipate in biogeochemical reactions at subzero temperatures and
nutrient deficiency in situ: the ability of immobilized enzymes in the
permafrost to be quickly activated
100
; the presence of metastable iron
sulfides
127
and nitrite
128
; and the ability to grow on nutrient media
after background radiation exposure for millions of years. Survival of
microorganisms in permafrost suggests the possibility of DNA repair,
that is, the existence of repair mechanisms that operate at a rate com-
parable to that of the damage accumulation in the genome.
The process of adaptation to permafrost conditions includes cold
protein synthesis and metabolic reactions at low temperatures. Stud-
ies have shown that Antarctic lichen can be metabolically active at
17°С.
129
There is evidence of DNA and protein synthesis at temper-
atures from 12 to 17°С, suggesting the possibility of cell division in
the snow cover of the South Pole.
130
Experiments with radioactively
labeled substrate have shown that ancient microorganisms in perma-
frost deposits from 600 000 to 1 million years old are not in a state
of absolute rest. Incorporation of labeled carbon CH
314
CO
2
in bacte-
rial lipids at temperatures up to 20°C shows that anabolic metabo-
lism is possible in ancient permafrost sediments.
12
Using
14
Clabeled
substrates (bicarbonate, H
14
CO
3
;acetate,
14
CH
3
CO
2
) it was demon-
strated that the natural population of methanogenic archaea of the
Late PlioceneEarly Pleistocene and Holocene permafrost deposits
carried out methane formation at temperatures up to 16.5°C (mini-
mum temperature of permafrost in the Arctic) and even at 28°C (per-
mafrost temperature in Antarctica's Beacon Valley). This revealed the
ability of microorganisms for energy metabolism, that is, redox reac-
tions after long periods in the permafrost.
7,78,131,132
From a biological point of view, the dosage of the natural radia-
tion, 12 mGy y
1
, emitted from the minerals that are dispersed in Arc-
tic permafrost deposits, is not sufficient for complete sterilization of
microbial complexes. On the other hand, the background radiation,
while not lethal, can promote selection and cause significant damage
to the DNA of ancient cells. However, the growth of permafrost
embedded microorganisms on nutrient media suggests their ability
for DNA reparation, indicating that the rate of DNA damage in perma-
frost is lower than its rate of repair.
133
Johnson and coauthors
13
came to a similar conclusion. Using
molecular biological approaches and direct measurement of perma-
frost sediment CO
2
production, they found evidence linking the
longterm viability of microorganisms with metabolic activity, and
DNA repair as a mechanism to maintain this activity.
Analysis of the complete genomes of permafrostisolated
microbes identified the genes responsible for the production of lipo-
lytic enzymes. Subsequent studies allowed analysis of the obtained
proteins. Both esterase and lipase, produced by a psychrotolerant
Psychrobacter cryohalolentis K5
T
isolated from Kolyma cryopegs, were
shown to be active and stable over a wide temperature range from 0
to 3035°С. For the esterase EstPc, the level of relative activity, 85
90%, remained essentially the same in the range between 30°Сand
С.
134-136
In the last 15 years permafrost microbiology has adopted modern
molecular techniques, such as metagenomics, PCR amplification and
phylogenetic identification of bacterial, archaeal and eukaryotic micro-
organisms. Longterm preservation of bacterial, archaeal and fungal
DNA has been observed in relatively young sediments of Late Pleisto-
cene and Holocene permafrost in the Canadian Arctic
55
and
Tibet,
54,137,138
as well as in older Late Pliocene and Early Pliocene sed-
iments in Siberia
119,126,139
and Antarctica.
7,140,141
Studies have shown that in the majority of ecosystems, DNA
released from a cell into the environment is usually destroyed rela-
tively quickly. Current research results
9,13,142,143
set the DNA preser-
vation limit to 400600 000 years in permafrost. PCR amplifications of
isolated DNA fragments in more ancient formations were not
observed. Nevertheless, the above limit is not definite and further
research aimed at isolation and analysis of total DNA is needed to
realistically assess the possibility of searching for DNA and microbial
cells in Martian permafrost.
5|RELEVANCE OF TERRESTRIAL
PERMAFROST STUDIES TO ASTROBIOLOGY
The fact that permafrost deposits occupy about 25% of the Earth's
surface unarguably defines the Earth as a cryogenic planet just like
all the other planets of the Solar System except Mercury and
Venus.
144,145
There is now growing evidence that the physical and
chemical surface properties of early Earth and Mars were very simi-
lar,
146
and, by analogy with terrestrial extremophilic microbial commu-
nities, potential life on Mars is suggested.
147
Although the presence of
toxic ions, perchlorate, conditions of extreme desiccation, high levels
of UV and ionizing radiation, etc., have been recorded on the surface
of Mars, traces of ancient ecosystems may be found below the surface
ground layer in permafrost, which is present on cryogenic
planets.
148,149
Furthermore, Earth's ice and permafrost may be
regarded as a model for astrobiology in determining potentially prom-
ising areas for the search for life on other cryogenic planets. Firstly,
microorganisms and other traces of life in terrestrial ice are considered
a representative model for extraterrestrial ecosystems such as Jupi-
ter's icy moon Europa, or Saturn's icy moon Enceladus.
21
Secondly,
given that hypersaline supercooled brines are the only possible form
of free water in Martian permafrost, the halophilic and psychrophilic
microbial community of terrestrial cryopegs and subsurface brines
may serve as a unique model for microbial life on Mars.
64,65
Thirdly,
frozen volcanic deposits that highlight the possible preservation of
thermophilic life forms at subzero temperatures are an important
object of study for astrobiology.
66
It should be noted that one of the weaknesses of Earth's perma-
frost as a model for astrobiology is that the age of permafrost on Earth
and other cryogenic planets of the Solar System is not comparable.
The most ancient permafrost on Earth, in Antarctica's Dry Valleys, is
a few tens of millions of years old, while the age of Martian permafrost
is measured in billions of years.
149
To compensate for this difference
6RIVKINA ET AL.
in part, we should compare the most ancient permafrost on Earth with
the youngest permafrost on Mars. Analysis of Mars surface images
(DTM provided by Mars Express High Resolution Stereo Camera,
HRSC) indicates that the youngest permafrost on Mars is found near
its northern pole. The latter is likely to be a region of relatively young
volcanoes and products of their eruption, the age of which may be
comparable with that of Earth's permafrost.
150
Therefore, the closest
analogues could be found among permafrost of volcanic origin. The
most ancient permafrost volcanic sediments discovered on Earth are
415 million years old
151
and are comparable with the most recent
volcanic sediments on Mars, which are permafrost volcanic rocks with
an age of 110 million years.
149
It was shown that thermophilic micro-
organisms associated with volcanic activity remain viable in perma-
frost.
66,68
Members of the genus Geobacillus (Table 1) with optimum
growth temperature of 6265°Сhave been isolated from the Antarctic
permafrost of Deception Island.
68
This suggests the possibility of find-
ing traces of life in the frozen volcanic deposits on Mars.
From the perspective of astrobiology, especially after liquid brines
in the shallow subsurface were found on Mars,
152
an important step
was the isolation of live microorganisms from terrestrial cryopegs
64
and underground brines.
65
Diverse microorganisms were isolated from
Arctic cryopegs including heterotrophic aerobic bacteria (10
2
cells mL
1
), acetogenic methanogens (10
2
cells mL
1
) and halophilic sulfate
reducers (up to 10
6
cells mL
1
).
114
Abundant fungal communities dom-
inated by Basidiomycota were discovered in hypersaline unfrozen
brines found within the Antarctic permafrost in northern Victoria
Land.
153
A microbial community with the density of 10
6
cells mL
1
was discovered in the extreme cryogenic brine ecosystem (13°C;
salinity, 200 g L
1
) of Lake Vida (Antarctica).
154
Growth of permafrost bacteria at conditions close to those on
Mars provides important evidence that life could be found on cryo-
genic planets. A new subspecies of the genus Carnobacterium,
Carnobacterium inhibens subsp. gilichinskyi, showed optimal growth in
aCO
2
enriched atmosphere at a low pressure of 7 mbar and temper-
ature of 0°C.
155
Permafrost bacteria Exiguobacterium sibircum strains
25515
T
and 73 from the order Bacillales of the phylum Firmicutes,
and hydrogenutilizing methanogenic archaeon Methanobacterium
veterum strain MK4
T
showed survival and growth recovery after expo-
sure to simulated Martian surface conditions (surface temperature
40°C to +25°C; humidity 095%; UV flux 19.3 W m
2
; 49% CO
2
;
50% Ar) while embedded within regolith simulant for 40 days.
156
Per-
chlorate salts with concentrations of 3.3 mM Mg
2+
and 2.4 mM ClO
4
are prevalent on the surface of Mars as confirmed by the onboard
Wet Chemistry Lab during the Phoenix lander mission.
157
A cold
adapted halophilic Antarctic archaeon, Halorubrum lacusprofundi, was
able to grow anaerobically with 40 mM perchlorate; and methano-
genic archaea isolated from Siberian permafrost grew in the presence
of 4 mM sodium and 2 mM magnesium perchlorates with
Methanobacterium articum M2
T
showing growth with up to 9 mM per-
chlorate.
158
Survival of terrestrial organisms on the surface of Mars
may be limited by stresses associated with desiccation and highly oxi-
dizing conditions arising from interactions of water vapor and the
Martian regolith.
156
Evidence from microorganisms surviving and
growing in a variety of frozen terrestrial ecosystems has expanded
our understanding of how life permeates, functions and persists under
challenging conditions and has provided support for possible life on
planets of a cryogenic nature.
6|CONCLUSIONS
Cenozoic permafrost is a unique ecosystem not impacted by anthro-
pogenic factors. The genes and metabolic products of viable microbial
cellshave persisted over geological time. Microbiological and molecular
biological studies of paleobiological objects in permafrost extend our
knowledge of the biosphere's spatial and temporal boundaries and
set a new direction in Quaternary geology, geocryology, bacterial pale-
ontology and exobiology. Furthermore, the ability of viable microor-
ganisms in permafrost to carry out metabolic reactions at subzero
temperatures and produce coldactive enzymes makes the Earth's per-
mafrost one of only a few natural models for exobiology.
ACKNOWLEDGEMENTS
This work was supported by Russian Government Assignment #
AAAAA181180131901816 and the Presidium of the Russian Acad-
emy of Sciences Program 22, and the National Science Foundation
DEB1442262.
ORCID
Elizaveta Rivkina http://orcid.org/0000-0001-7949-8056
Tatiana Vishnivetskaya http://orcid.org/0000-0002-0660-023X
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How to cite this article: Rivkina E, Abramov A, Spirina E, et al.
Earth's perennially frozen environments as a model of cryo-
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2018;111. https://doi.org/10.1002/ppp.1987
RIVKINA ET AL.11
... This deep massive ice is surely older than 20,000 years BP (Baroni and Orombelli, 1987) and it is assumed to be in frozen form since that time. Therefore, this deep ice layer may represent a fossil archive of past microbial diversity because of the age of microorganisms may be assumed to coincide with the age of the ice (Willerslev et al., 2004;Willerslev and Cooper, 2005;Rivkina et al., 2018). ...
... The ecological question arising from the above speculation may be summarized as follows: does it make sense to assume the existence of an appreciable level of in situ fungal activity (at the low temperatures measured in cores: minimum variable between −16.4 and −31.1 Cmaximum variable between −1.9 and −11.7 C) or such activity can be considered negligible? The microbial activity at sub-zero temperatures, long considered minimal, has been re-valued by studies carried out since early 2000s that have detected the presence of a slow, but appreciable, metabolic activity of both prokaryotic and eukaryotic microbial communities successfully colonizing cold ecosystems (Price, 2000;Gunde-Cimermann et al., 2003;Price and Sowers, 2004;Su et al., 2016;Buzzini et al., 2018;Rivkina et al., 2018;da Silva et al., 2019). Murray and colleagues (2012) found bacterial communities metabolically active at −13 C in brines of an ice-sealed Antarctic lake and concluded that the discovery in this ecosystem of in situ biotic and abiotic processes occurring at sub-zero temperatures could provide a system to study the habitability of isolated terrestrial ecosystems. ...
... archaea, bacteria and algae among others) could be another factors putatively playing a crucial role in regulating the ecological balance of the fungal assemblage. Antarctic permafrost can host from 10 3 to 10 6 bacterial cells g −1 , followed by fungi and archaea which occur from 200 to 1000 times lesser (Rivkina et al., 2004(Rivkina et al., , 2018. Likewise, algae have been found in Antarctic brines (Baldi et al., 2011;McMinn et al., 2014;Choua et al., 2018). ...
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A perennially frozen lake at Boulder Clay site (Victoria Land, Antarctica), characterized by the presence of frost mounds, have been selected as an in‐situ model for ecological studies. Different samples of permafrost, glacier ice and brines have been studied as a unique habitat system. An additional sample of brines (collected in another frozen lake close to the previous one) was also considered. Alpha‐ and beta‐diversity of fungal communities showed both intra‐ and inter‐cores significant (p < 0.05) differences, which suggest the presence of interconnection among the habitats. Therefore, the layers of frost mound and the deep glacier could be interconnected while the brines could probably be considered as an open habitat system not interconnected with each other. Moreover the absence of similarity between the lake ice and the underlying permafrost suggested that the lake is perennially frozen based. The predominance of positive significant (p < 0.05) co‐occurrences among some fungal taxa allowed to postulate the existence of an ecological equilibrium in the habitats systems. The positive significant (p < 0.05) correlation between salt concentration, total organic carbon and pH, and some fungal taxa suggests that a few abiotic parameters could drive fungal diversity inside these ecological niches.
... Earth is a planet of cryogenic type where all four elements (snow, ice/sea ice, glaciers and permafrost) of the cryosphere are present (Rivkina et al. 2018). Based on data from the U.S. Geological Survey (Williams and Ferrigno 2012), ∼15.6 × 10 6 km 2 of Earth's surface is covered by glaciers; the extent of seasonal snow cover in the Northern Hemisphere varies from 46.9 × 10 6 to 3.5 × 10 6 km 2 , and floating sea ice covers vast areas of the polar oceans, ranging in extent from ∼5 × 10 6 to ∼15-18 × 10 6 km 2 in both the Northern and Southern Hemispheres. ...
... The number of isolates and their taxonomic diversity depended on the origin and age of the permafrost, as well as on the permafrost's properties, including total organic carbon concentration, salinity and pH (Gilichinsky, Wagener and Vishnivetskaya 1995;Vorobyova et al. 1997), and on nutrient media and laboratory conditions used for microorganisms recovery (Palumbo et al. 1996). Diverse microorganisms from three domains-Archaea, Bacteria and Eukarya-and viruses were isolated from permafrost; however, only a small number of isolates were taxonomically identified (Rivkina et al. 2018). Successful culturing of those microorganisms is direct evidence of life preservation in permafrost (Khlebnikova et al. 1990;Vishnivetskaya et al. 2000;Steven et al. 2007). ...
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Permafrost describes the condition of earth material (sand, ground, organic matter, etc.) cemented by ice when its temperature remains at or below 0°C continuously for longer than 2 years. Evidently, permafrost is as old as the time passed from freezing of the earth material. Permafrost is a unique phenomenon and may preserve life forms it encloses. Therefore, in order to talk confidently about the preservation of paleo-objects in permafrost, knowledge about the geological age of sediments, i.e. when the sediments were formed, and permafrost age, when those sediments became permanently frozen, is essential. There are two types of permafrost-syngenetic and epigenetic. The age of syngenetic permafrost corresponds to the geological age of its sediments, whereas the age of epigenetic permafrost is less than the geological age of its sediments. Both of these formations preserve microorganisms and their metabolic products; however, the interpretations of the microbiological and molecular-biological data are inconsistent. This paper reviews the current knowledge of time-temperature history and age of permafrost in relation to available microbiological and metagenomic data.
... 21,24,25 Moreover, microbial cells can persist within ice (e.g., 26,27 ) and permafrost up to a few million years and may still be capable of sustaining basic metabolic activities. [28][29][30] Therefore, we hypothesize that as permafrost warms, microbial communities may enhance silicate weathering in polar environments, even in xerous and ultraxerous settings. ...
... 41 Furthermore, cyanobacteria may have formed primordial soils of early Earth, and prospective extraterrestrial soil formation processes on Mars and other bodies may also be tied to bioweathering by polyextremophiles. 30,41 Traces of such biological surface alteration can be used as inorganic biosignatures, which are defined as the chemical, morphological, and mineralogical (biomineral) biproducts of microbemineral interactions. 42,43 Considering the compelling evidence of chemical and potentially biological weathering from previous studies in Antarctica (e.g., 35,41 ) and the role of microbes in accelerating silicate weathering (e.g., [44][45][46][47] ), we hypothesize that cyanobacterial mats in polar environments have the potential to enhance chemical weathering rates via both elevated pH in the solution and locally generated micro-acidic regions within their EPS on the grains via cell-surface attachment. ...
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... Microorganisms flourishing in tough cold environments have been reported from time to time; psychrophiles are habitants of environments such as the Siberian permafrost, which is considered a distinctive environment, having permanently icy grounds, limited availability of organic matter, low water activity, and additional factors [5][6][7][8][9]. Among psychrophilic microorganisms, bacteria and fungi have been isolated and investigated comprehensively as compared to yeasts, which constitute a minor part, but all forms have the potential of producing essential psychrozymes [10,11]. ...
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Citation: Hamid, B.; Bashir, Z.; Yatoo, A.M.; Mohiddin, F.; Majeed, N.; Bansal, M.; Poczai, P.; Almalki, W.H.; Sayyed, R.Z.; Shati, A.A.; et al. Cold-Active Enzymes and Their Potential Industrial Applications-A Review. Molecules 2022, 27, 5885.
... Microorganisms flourishing in tough cold environments have been reported from time to time; psychrophiles are habitants of environments such as the Siberian permafrost, which is considered a distinctive environment, having permanently icy grounds, limited availability of organic matter, low water activity, and additional factors [5][6][7][8][9]. Among psychrophilic microorganisms, bacteria and fungi have been isolated and investigated comprehensively as compared to yeasts, which constitute a minor part, but all forms have the potential of producing essential psychrozymes [10,11]. ...
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More than 70% of our planet is covered by extremely cold environments, nourishing a broad diversity of microbial life. Temperature is the most significant parameter that plays a key role in the distribution of microorganisms on our planet. Psychrophilic microorganisms are the most prominent inhabitants of the cold ecosystems, and they possess potential cold-active enzymes with diverse uses in the research and commercial sectors. Psychrophiles are modified to nurture, replicate, and retain their active metabolic activities in low temperatures. Their enzymes possess characteristics of maximal activity at low to adequate temperatures; this feature makes them more appealing and attractive in biotechnology. The high enzymatic activity of psychrozymes at low temperatures implies an important feature for energy saving. These enzymes have proven more advantageous than their mesophilic and thermophilic counterparts. Therefore, it is very important to explore the efficiency and utility of different psychrozymes in food processing, pharmaceuticals, brewing, bioremediation, and molecular biology. In this review, we focused on the properties of cold-active enzymes and their diverse uses in different industries and research areas. This review will provide insight into the areas and characteristics to be improved in cold-active enzymes so that potential and desired enzymes can be made available for commercial purposes.
... В обзоре "Earth's perennially frozen environments as a model of cryogenic planet ecosystems" (Rivkina et al., 2018) обобщены последние результаты исследований жизнеспособной биоты многолетнемерзлых отложений Земли. Самые последние сведения о фотосинтетических микроорганизмах, сохранивших жизнеспособность в вечной мерзлоте, приведены в обзоре "Insights into community of photosynthetic microorganisms from permafrost" (Vishnivetskaya et al., 2020). ...
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В монографии изложены результаты многолетних разноплановых исследований по бактериальной палеонтологии, которые касались роли бактерий в породои рудообразовании земных отложений от древнейших до современных, включая и систему вечной мерзлоты. Для удобства или в помощь читающим, в книге первая глава посвящена описанию современных микроорганизмов. Монография предназначена для широкого круга: палеонтологов, литологов, биологов, геологов. Кроме этого эта работа может служить прекрасным учебным пособием для студентов, магистрантов и аспирантов высших учебных заведений.
... Within the European Alps, the Stelvio area exhibits these effects in a particularly evident way, with important consequences on microbial ecosystems (Ponti et al., 2021). Despite their frozen status, many microbial species inhabiting permafrost cores are considered metabolically active, and this activity can obviously increase in the case of permafrost thawing (even temporary) (Bakermans et al., 2003;Rivkina et al., 2004Rivkina et al., , 2018Steven et al., 2006;Tuorto et al., 2014). Therefore, recent studies have included the monitoring of microbial parameters in order to gain a more complete picture of the interactions between biotic and abiotic factors occurring in these changing ecosystems (Wagner et al., 2007;Hollesen et al., 2011;Graham et al., 2012;Donhauser and Frey, 2018). ...
Article
The impact of climate change in the European Alps has been roughly twice the global average, dramatically reducing permafrost extent and thickening of its active layer. Therefore, the study of the abiotic factors (i.e. chemical/physical parameters) affecting the microbial diversity inhabiting Alpine permafrost appears to be of dramatic relevance. Within the European Alps, the Stelvio area exhibits these effects in a particularly evident way, with important consequences on microbial ecosystems. Therefore, microbial communities inhabiting a permafrost core collected in the Scorluzzo active rock glacier (Stelvio Pass, Italian Central Alps) were investigated along a depth gradient (410 to 524 cm from the surface). The taxonomic structures of bacterial and fungal communities were investigated via a next-generation sequencing (NGS) approach (Illumina MiSeq), targeting the bacterial V3-V4 regions of 16S rRNA and the fungal ITS2 region. Abiotic soil factors (grain size, electrical conductivity, ice/water content, pH, Loss-on-Ignition - LOI, total and organic carbon, nitrogen and phosphorous) were analysed. Richness and Shannon-H diversity indices were correlated to abiotic factors. Bacterial diversity was significantly (p < 0.05) correlated with LOI, while fungal diversity was significantly (p < 0.05) correlated with the depth gradient. The Constrained Analysis of Principal (CAP) coordinates were used to study the correlation between abiotic parameters and the taxonomic structure of bacterial and fungal communities. Among all tested variables, the depth gradient, water content, pH and LOI affected the taxonomic structure of bacterial communities (in particular, the abundance of bacterial amplicon sequence variants - ASVs - assigned to Afipia sp., Chloroflexi, Gaiella sp., Oryzihumus sp. and Serratia, sp.), while fungal communities (ASVs assigned to Naganishia sp., Rhodotorula sp., Sordariomycetes and Taphrinales) were affected by the depth gradient. Co-occurrences (calculated by Pearson correlation coefficient) among microbial taxa (i.e. bacteria vs bacteria, bacteria vs fungi, fungi vs fungi) were investigated: the prevalence of significant (p < 0.05) positive co-occurrences was found, suggesting that the coexistence of different microbial taxa could play a crucial role in maintaining the ecological and taxonomic balance of both bacterial and fungal communities inhabiting the Alpine permafrost ecosystem. These findings suggest that the bacterial and fungal diversity of Alpine permafrost are affected in different ways by some abiotic factors.
... Permafrost and arid polar deserts of Antarctica are considered to be most close in the totality of physicochemical conditions to the known characteristics of Martian regolith. This is the cause to carry out microbiological study of astrobiological character in the most extreme regions of Antarctica, and particularly in the region of Dry Valleys [13,16,32]. The dose of irradiation used in the experiments corresponded to the long-lasting effect of cosmic radiation on Martian regolith, and accumulation of radiationinduced defects by cells supposedly preserved in an anabiotic state in Martian ground. ...
Article
Ionizing radiation is an important environmental factor affecting the dynamics of biospheric processes in the past and present, as well as limiting the spread of life outside the Earth. The effect of radiation on microorganisms has been studied for decades, but studies of the response of natural microbial ecosystems are still scarce. We have studied the effect of 100 kGy gamma irradiation under low pressure (1 Torr) and low temperature (-50°C) on microbial community of the ancient Antarctic permafrost sedimentary rock. After irradiation, the total number of prokaryotic cells determined by epifluorescence microscopy, as well as the number of metabolically active bacterial and archaeal cells detected by fluorescence in situ hybridization remained at the control level, while the number of cultured heterotrophic bacteria decreased by an order of magnitude. Using the multisubstrate testing method, it has been found that the microbial complex retained a high potential metabolic activity and functional diversity after exposure to a combination of extreme physical factors. The resistance demonstrated by the microbial community significantly exceeded the generally accepted estimates of the prokaryotes' radioresistance and indicated an underestimation of the microorgan-isms' radioresistance in natural habitats and the important role of mineral heterophase environment and irradiation conditions (pressure, temperature). The study confirmed the potential for long-term cryopreservation of viable terrestrial-like microorganisms in the Martian regolith, as well as the possibility of transferring anabiotic life forms as a part of small bodies in the space environment.
... Being a habitat for diverse cold-adapted microorganisms, Siberian permafrost is a unique environment which is characterized by the presence of permanently frozen ground, limited accessibility of organic matter, low water activity and other factors [9][10][11]. Previously, we have described several lipolytic enzymes from the permafrost bacterium Psychrobacter cryohalolentis K5 T [12][13][14]. ...
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The gene coding for a novel cold-active esterase PMGL3 was previously obtained from a Siberian permafrost metagenomic DNA library and expressed in Escherichia coli. We elucidated the 3D structure of the enzyme which belongs to the hormone-sensitive lipase (HSL) family. Similar to other bacterial HSLs, PMGL3 shares a canonical α/β hydrolase fold and is presumably a dimer in solution but, in addition to the dimer, it forms a tetrameric structure in a crystal and upon prolonged incubation at 4 °C. Detailed analysis demonstrated that the crystal tetramer of PMGL3 has a unique architecture compared to other known tetramers of the bacterial HSLs. To study the role of the specific residues comprising the tetramerization interface of PMGL3, several mutant variants were constructed. Size exclusion chromatography (SEC) analysis of D7N, E47Q, and K67A mutants demonstrated that they still contained a portion of tetrameric form after heat treatment, although its amount was significantly lower in D7N and K67A compared to the wild type. Moreover, the D7N and K67A mutants demonstrated a 40 and 60% increase in the half-life at 40 °C in comparison with the wild type protein. Km values of these mutants were similar to that of the wt PMGL3. However, the catalytic constants of the E47Q and K67A mutants were reduced by ~40%.
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The analysis of metagenomes from four Late Pleistocene permafrost samples allowed us to recognize nearly four hundred genera of protists and fungi, as well as nematodes, in the microeukaryotic assemblage. The sample of the ancient oxbow lake sediments is characterized by the highest taxonomic diversity. Heterotrophic protists and autotrophs dominated the deposits that formed under hydromorphic conditions. Fungi, in turn, prevailed in the Ice Complex deposits. In general, metagenomic analysis characterizes the assemblages from permafrost deposits more entirely than the standard methods of enrichment cultivation.
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Polar permafrost is at the forefront of climate change, yet only a few studies have enriched the native methane-producing microbes that might provide positive feedbacks to climate change. Samples Ant1 and Ant2, collected in Antarctic Miers Valley from permafrost sediments, with and without biogenic methane, respectively, were evaluated for methanogenic activity and presence of methanogens. After a one-year incubation of both samples under anaerobic conditions, methane production was observed only at room temperature in microcosm Ant1 with CO2/H2 (20/80) as carbon and energy sources, and was monitored during the subsequent 10 years. The concentration of methane in the headspace of microcosm Ant1 changed from 0.8% to a maximum of 45%. Archaeal 16S rRNA genes from microcosm Ant1 were related to psychrotolerant Methanosarcina lacustris. Repeated efforts at achieving a pure culture of this organism were unsuccessful. Metagenomic reads obtained for the methane-producing microcosm Ant1 were assembled and resulted in a 99.84% complete genome affiliated with the genus Methanosarcina. The metagenome assembled genome contained cold-adapted enzymes and pathways suggesting that the novel uncultured Methanosarcina sp. Ant1 is adapted to sub-freezing conditions in permafrost. This is the first methanogen genome reported from the 15,000 years old permafrost of the Antarctic Dry Valleys.
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Brines are hypersaline solutions which have been found within the Antarctic permafrost from the Tarn Flat area (Northern Victoria Land). Here, an investigation on the possible presence and diversity of fungal life within those peculiar ecosystems has been carried out for the first time. Brines samples were collected at 4- and 5-meter depths (TF1 and TF2, respectively), from two brines separated by a thin ice layer. The samples were analyzed via Illumina MiSeq targeting the ITS region specific for both yeasts and filamentous fungi. An unexpected high alpha diversity was found. Beta diversity analysis revealed that the two brines were inhabited by two phylogenetically diverse fungal communities (Unifrac value: 0.56, p value < 0.01; Martin's P-test p-value < 0.001) characterized by several specialist taxa. The most abundant fungal genera were Candida sp., Leucosporidium sp., Naganishia sp. and Sporobolomyces sp. in TF1, and Leucosporidium sp., Malassezia sp., Naganishia sp. and Sporobolomyces sp. in TF2. A few hypotheses on such differentiation have been done: i) the different chemical and physical composition of the brines; ii) the presence in situ of a thin layer of ice, acting as a physical barrier; and iii) the diverse geological origin of the brines.
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Cryopegs, lenses of hypersaline unfrozen soil or water within permafrost, are a model for astrobiology, since free water can only be present on cryogenic bodies and planets in the form of brine. In this paper the diversity of aerobic halophilic-psychrotrophic microorganisms from an Alaskan cryopeg (Barrow Cape) were studied and described for the first time. This cryopeg is characterized by a constant subzero temperature (–7°C), high salinity (total mineralization is about 120 g/L) and isolation from external influences for a geologically significant period of time. Our study has revealed a large number of microorganisms capable of growth at low temperature (4°C) in a wide range of salinities from 5 to 250 g/L of NaCl, the latter being 3 times higher than the natural salt concentration of the Alaskan cryopeg. The microorganisms identified are comprised of four major phyla: Actinobacteria (genera Brevibacterium, Citricoccus, Microbacterium), Firmicutes (genus Paenibacillus), Bacteroidetes (genus Sphingobacterium), and Proteobacteria (genus Ochrobactrum).
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Icy worlds in the solar system and beyond have attracted a remarkable attention as possible habitats for life. The current consideration about whether life exists beyond Earth is based on our knowledge of life in terrestrial cold environments. On Earth, glaciers and ice sheets have been considered uninhabited for a long time as they seemed too hostile to harbor life. However, these environments are unique biomes dominated by microbial communities which maintain active biochemical routes. Thanks to techniques such as microscopy and more recently DNA sequencing methods, a great biodiversity of prokaryote and eukaryote microorganisms have been discovered. These microorganisms are adapted to a harsh environment, in which the most extreme features are the lack of liquid water, extremely cold temperatures, high solar radiation and nutrient shortage. Here we compare the environmental characteristics of icy worlds, and the environmental characteristics of terrestrial glaciers and ice sheets in order to address some interesting questions: (i) which are the characteristics of habitability known for the frozen worlds, and which could be compatible with life, (ii) what are the environmental characteristics of terrestrial glaciers and ice sheets that can be life-limiting, (iii) What are the microbial communities of prokaryotic and eukaryotic microorganisms that can live in them, and (iv) taking into account these observations, could any of these planets or satellites meet the conditions of habitability? In this review, the icy worlds are considered from the point of view of astrobiological exploration. With the aim of determining whether icy worlds could be potentially habitable, they have been compared with the environmental features of glaciers and ice sheets on Earth. We also reviewed some field and laboratory investigations about microorganisms that live in analog environments of icy worlds, where they are not only viable but also metabolically active.
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This book describes the effects of cold climates on the surface of the earth. Using scientific principles, the authors describe the evolution of ground thermal conditions and the origin of natural features such as frost heave, solifluction, slope instabilities, patterned ground, pingos and ice wedges. The thermodynamic conditions accompanying the freezing of water in porous materials are examined and their fundamental role in the ice segregation and frost heave processes is demonstrated in a clear and simple manner. This book concentrates on the analysis of the causes and effects of frozen ground phenomena, rather than on the description of the natural features characteristic of freezing or thawing ground. Its scientific approach provides a basis for geotechnical analyses such as those essential to resource development.
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