Transitory microbial habitat in the hyperarid
Atacama Desert
Dirk Schulze-Makucha,b,1, Dirk Wagnerc,d, Samuel P. Kounavese,f, Kai Mangelsdorfg, Kevin G. Devineh,
Jean-Pierre de Verai, Philippe Schmitt-Kopplinj,k, Hans-Peter Grossartl,m, Victor Parron, Martin Kaupenjohanno,
Albert Galyp, Beate Schneidera,c, Alessandro Airoa, Jan Fr ¨
oslerq, Alfonso F. Davilar, Felix L. Arenss, Luis C´
acerest,
Francisco Sol´
ıs Cornejot, Daniel Carrizon, Lewis Dartnellu, Jocelyne DiRuggierov, Markus Fluryw, Lars Ganzertl,
Mark O. Gessnerl,x, Peter Grathwohly, Lisa Guanz, Jacob Heinza, Matthias Hessaa, Frank Kepplerbb, Deborah Mausa,
Christopher P. McKayr, Rainer U. Meckenstockq, Wren Montgomeryf, Elizabeth A. Oberline, Alexander J. Probstq,
Johan S. S´
aenzz, Tobias Sattlerbb, Janosch Schirmacka, Mark A. Sephtonf, Michael Schloterz,cc, Jenny Uhlk,
Bernardita Valenzuelat, Gisle Vestergaardz, Lars W ¨
ormerdd, and Pedro Zamoranot
aCenter of Astronomy & Astrophysics, Technical University Berlin, 10623 Berlin, Germany; bSchool of the Environment, Washington State University,
Pullman, WA 99164; cSection Geomicrobiology, GFZ German Research Centre for Geosciences, 14473 Potsdam, Germany; dInstitute of Earth and
Environmental Science, University of Potsdam, 14476 Potsdam, Germany; eDepartment of Chemistry, Tufts University, Medford, MA 02153; fDepartment of
Earth Science & Engineering, Imperial College London, London SW72AZ, United Kingdom; gSection Organic Geochemistry, GFZ German Research Centre for
Geosciences, 14473 Potsdam, Germany; hSchool of Human Sciences, London Metropolitan University, London N7 8BD, United Kingdom; iAstrobiological
Laboratories, Management and Infrastructure, Institute for Planetary Research, German Aerospace Center, 12489 Berlin, Germany; jAnalytical Food
Chemistry, Technical University M ¨
unchen, 85354 Freising-Weihenstephan, Germany; kAnalytical BioGeoChemistry, Helmholtz Zentrum M ¨
unchen, 85764
Oberschleissheim, Germany; lDepartment of Experimental Limnology, Leibniz Institute of Freshwater Ecology and Inland Fisheries, 16775 Stechlin,
Germany; mInstitute of Biochemistry & Biology, University of Potsdam, 14476 Potsdam, Germany; nMolecular Evolution Department, Centro de
Astrobiolog´
ıa, Instituto Nacional de T´
ecnica Aeroespacial-Consejo Superior de Investigaciones Cient´
ıficas (INTA-CSIC), 28850 Madrid, Spain; oFachgebiet
Bodenkunde, Technical University Berlin, 10623 Berlin, Germany; pCentre de Recherches P ´
etrographiques et G´
eochimiques, CNRS, Universit´
e de Lorraine,
54500 Vandoeuvre les Nancy, France; qBiofilm Centre, University of Duisburg-Essen, 45141 Essen, Germany; rPlanetary Systems Branch (Code SST), NASA
Ames Research Center, Moffett Field, CA 94035; sInstitute for Geological Sciences, Freie University Berlin, 12249 Berlin, Germany; tLaboratorio de
Microorganismos Extrem ´
ofilos, University of Antofagasta, Antofagasta 02800, Chile; uDepartment of Life Sciences, University of Westminster, London W1W
6UW, United Kingdom; vDepartment of Biology, The John Hopkins University, Baltimore, MD 21218; wDepartment of Crop & Soil Sciences, Washington
State University, Pullman, WA 99164; xDepartment of Ecology, Technical University Berlin, 10587 Berlin, Germany; yCenter for Applied Geosciences,
University of T ¨
ubingen, 72074 T ¨
ubingen, Germany; zComparative Microbiome Analysis, Helmholtz Zentrum M ¨
unchen, 85764 Oberschleissheim, Germany;
aaSystems Microbiology & Natural Products Laboratory, University of California, Davis, CA 95616; bbInstitute of Earth Sciences, Heidelberg University, 69120
Heidelberg, Germany; ccSoil Science, Technical University M ¨
unchen, 85354 Freising-Weihenstephan, Germany; and ddCenter for Marine Environmental
Sciences (MARUM), University of Bremen, 28359 Bremen, Germany
Edited by Mary K. Firestone, University of California, Berkeley, CA, and approved January 25, 2018 (received for review August 17, 2017)
Traces of life are nearly ubiquitous on Earth. However, a cen-
tral unresolved question is whether these traces always indicate
an active microbial community or whether, in extreme environ-
ments, such as hyperarid deserts, they instead reflect just dormant
or dead cells. Although microbial biomass and diversity decrease
with increasing aridity in the Atacama Desert, we provide multi-
ple lines of evidence for the presence of an at times metabolically
active, microbial community in one of the driest places on Earth.
We base this observation on four major lines of evidence: (i) a
physico-chemical characterization of the soil habitability after an
exceptional rain event, (ii) identified biomolecules indicative of
potentially active cells [e.g., presence of ATP, phospholipid fatty
acids (PLFAs), metabolites, and enzymatic activity], (iii) measure-
ments of in situ replication rates of genomes of uncultivated
bacteria reconstructed from selected samples, and (iv) microbial
community patterns specific to soil parameters and depths. We
infer that the microbial populations have undergone selection and
adaptation in response to their specific soil microenvironment and
in particular to the degree of aridity. Collectively, our results high-
light that even the hyperarid Atacama Desert can provide a habit-
able environment for microorganisms that allows them to become
metabolically active following an episodic increase in moisture
and that once it decreases, so does the activity of the microbiota.
These results have implications for the prospect of life on other
planets such as Mars, which has transitioned from an earlier wet-
ter environment to today’s extreme hyperaridity.
habitat |aridity |microbial activity |biomarker |Mars
The core region of the Atacama Desert is the most arid mid-
latitude desert on Earth and in the past has been devoid of
precipitation for decades. A mean annual precipitation of <20
mm reduces weathering rates and leaching losses to levels below
the accumulation rates of atmospheric salts and dust (1). Hence,
Significance
It has remained an unresolved question whether microorgan-
isms recovered from the most arid environments on Earth are
thriving under such extreme conditions or are just dead or
dying vestiges of viable cells fortuitously deposited by atmo-
spheric processes. Based on multiple lines of evidence, we show
that indigenous microbial communities are present and tempo-
rally active even in the hyperarid soils of the Atacama Desert
(Chile). Following extremely rare precipitation events in the dri-
est parts of this desert, where rainfall often occurs only once per
decade, we were able to detect episodic incidences of biologi-
cal activity. Our findings expand the range of hyperarid envi-
ronments temporarily habitable for terrestrial life, which by
extension also applies to other planetary bodies like Mars.
Author contributions: D.S.-M. designed research; D.S.-M., D.W., S.P.K., K.M., K.G.D.,
J.-P.d.V., P.S.-K., H.-P.G., V.P., A.G., B.S., A.A., J.F., A.F.D., F.L.A., L.C., F.S.C., D.C., L.D., J.D.,
M.F., L. Ganzert, M.O.G., P.G., L. Guan, J.H., M.H., F.K., D.M., R.U.M., W.M., E.A.O., A.J.P.,
J.S.S., T.S., J.S., M.A.S., M.S., J.U., B.V., G.V., L.W., and P.Z. performed research; D.W., S.P.K.,
K.M., K.G.D., P.S.-K., H.-P.G., V.P., M.K., A.G., M.O.G., P.G., F.K., R.U.M., A.J.P., M.S., G.V.,
and L.W. contributed new reagents/analytic tools; D.S.-M., D.W., S.P.K., K.M., K.G.D.,
J.-P.d.V., P.S.-K., H.-P.G., V.P., M.K., A.G., B.S., A.A., J.F., A.F.D., F.L.A., L.D., J.D., M.F.,
L. Ganzert, M.O.G., P.G., M.H., F.K., C.P.M., R.U.M., A.J.P., J.S., M.S., J.U., B.V., G.V., and
L.W. analyzed data; and D.S.-M., D.W., S.P.K., H.-P.G., and M.S. wrote the paper.
The authors declare no conflict of interest.
This article is a PNAS Direct Submission.
This open access article is distributed under Creative Commons Attribution-
NonCommercial-NoDerivatives License 4.0 (CC BY-NC-ND).
Data deposition: The metagenome sequences reported in this paper have been deposited
in the EMBL-EBI database (accession no. PRJEB20402 with the sample IDs ERS1666624–
ERS1666714) and in the GenBank/EMBL database (BioProject ID PRJNA395196).
1To whom correspondence should be addressed. Email: [email protected].
This article contains supporting information online at www.pnas.org/lookup/suppl/doi:10.
1073/pnas.1714341115/-/DCSupplemental.
Published online February 26, 2018.
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atmospheric deposition over millions of years has resulted in high
salt concentrations in the soils of the hyperarid area (2). The only
documented microhabitats in the core region of the Atacama
Desert are colonized by microbial communities thriving in sur-
ficial salt crusts, where microbial activity is enabled through del-
iquescence (3, 4). Even though there are traces of microbial life
in the subsurface of the Atacama Desert (5), it remains unclear
whether these environments support active microbial growth or
whether the observed cells are sporadically introduced by atmo-
spheric transport and continuously inactivated and degraded. To
answer this question, in April of 2015 we sampled soils from
the surface and near subsurface at six locations along a decreas-
ing moisture gradient [coastal soil (CS), alluvial fan (AL), red
sands (RS), Maria Elena (ME), Yungay (YU), and Lomas Bayas
(LB)] (SI Appendix, Fig. S1) and characterized them and their
microbial communities by using a broad suite of complemen-
tary methods. Since this sampling occurred shortly after an unex-
pected rain event, we repeated sampling in February 2016 and
January 2017 to determine whether the detected microbial activ-
ity in 2015 was ongoing or episodic and related to the temporary
increased availability of moisture.
Results
Environmental Setting. The selected CS site has been occasion-
ally subject to fog and rain, while sites further inland (ME, YU,
and LB) are located in hyperarid areas (6, 7), where water con-
tent of surface soils is generally <1% by weight (SI Appendix,
Fig. S2). Water activity is often below the threshold of ∼0.6
required to sustain metabolic activity (8). Relative humidity lev-
els are generally below 30% and daily UV irradiation doses
were ca. 30 J·m−2. Except for LB, where total organic carbon
(TOC) reached 0.25% (wt/wt), TOC at all other sites was less
than 0.1%. A prerequisite of our study was that the sampled
sites are relatively pristine and little affected by human contam-
ination, which was the case based on measured polycyclic aro-
matic hydrocarbon concentrations which are extremely low and
generally in the microgram per kilogram range or lower. Soil
minerals at all sites are dominated by alkali feldspar and pla-
gioclase with minor amounts of quartz, chlorite, and amphibole,
with some sites displaying a significant amount of anhydrite, bas-
sanite, gypsum, and carbonates. Sites subject to higher levels
of moisture (CS, AL, RS) contain large amounts of chlorides
(e.g., halite), while soils obtained from the hyperarid areas (YU,
ME, LB) mostly contain sulfates (e.g., gypsum or anhydrite),
perchlorates (ClO−
4), and chlorates (ClO−
3) (SI Appendix, Fig.
S3). The first set of field samples was taken in April 2015 1 mo
after a major El Ni˜
no triggered one of the rare rainfall events in
the Atacama Desert (9). Eight millimeters of precipitation was
recorded at Baquedana and 33 mm at Antofagasta, which was
the highest amount of precipitation since the beginning of the
official recording in 1978 (SI Appendix, Fig. S2C) and affected
all study sites. The second and third sampling campaigns were
conducted in February 2016 and January 2017, respectively, with
only two minor rain events in between (each 6.7 mm, recorded at
Antofagasta).
Microbial Diversity. Metagenomic analyses of the DNA pool from
topsoils revealed a high bacterial diversity at CS (nonpareil diver-
sity index of 21.2 ±0.5), similar to sandy soils, but considerably
higher compared with the drier areas of YU and ME (nonpareil
indexes of 19.5 ±0.6 and 18.9 ±0.1, respectively; SI Appendix,
Fig. S4A). Phylogenetic profiles (SI Appendix, Fig. S4B) indi-
cate that soils at YU were associated with microbiomes typical
for sandy environments and desert soils, mainly consisting of
Actinobacteria (5) with Corynebacteriales,Streptomycetales, and
Micrococcales being the dominant suborders and a proportional
decline of Rubrobacterales from the surface to the subsurface. In
contrast, ME was dominated by Geodermatophilaceae, known to
colonize hyperarid habitats (10) and tolerate high levels of oxida-
tive stress, desiccation, salts, and metals (11). The same general
trend of decreased biomass and diversity with increasing aridity
was found for Archaea and Fungi, even though their proportion
was lower than that of Bacteria. In all soil samples ∼200 fun-
gal marker genes were detected, almost exclusively belonging to
Ascomycota and Basidiomycota (SI Appendix, Fig. S5 Aand B).
At CS, Archaea (mainly halobacteria) reached a maximum and
dominated the microbial community at a depth of 20–30 cm,
while everywhere else they accounted for <4% of sequence
reads (SI Appendix, Fig. S5C). Our DNA-based data were cor-
roborated by phospholipid fatty acid profiles (Fig. 1 Band D),
serving as biomarkers for living bacteria (12), and cultivation-
based approaches both identifying Actinobacteria (e.g., Acti-
nobacterium lienomycini,Kocuria sp.,Pseudonocardina sp.,Strep-
tomyces sp.), Proteobacteria (e.g., Pseudomonas sp.,Paracoccus
marinus), and Firmicutes (e.g., Bacillus litoralis,Bacillus simplex,
Halobacillus sp.).
Abundance and Identification of Dead and Living Microorganisms.
A unique cell-separation technique (13) was used to differen-
tiate between intracellular DNA (iDNA) indicating physically
intact and potentially viable cells and extracellular DNA (eDNA)
mainly representing preserved DNA from dead cells.
Quantitative PCR (qPCR) using universal bacterial primers
was performed for both DNA pools, for all soil samples taken
along the moisture gradients in 2015 and 2017, and at YU in
2016. After the rain event in 2015, the abundance of 16S rRNA
genes in the iDNA pool (proxy of living bacterial biomass) was
generally higher than in 2016/2017 (between ∼103and ∼106
gene copies per gram soil; SI Appendix, Table S1), increasing with
moisture (SI Appendix, Fig. S6). The copy numbers in the eDNA
pool showed larger variations of between ∼101and ∼107gene
copies per gram soil, and only at LB was no eDNA detectable
with the primers applied in the qPCR. In contrast to the rel-
atively high gene copy numbers in 2015, soil samples from the
same sites, but taken 2 y after the rain event, revealed a drastic
decrease in living cells (iDNA; equal to or less than 102gene
Fig. 1. (Aand B) Concentrations of intracellular ATP (iATP) and extracel-
lular ATP (eATP) (both n= 3) (A) and PLFAs (B) in Atacama Desert soils. A
decrease in the number of identified PLFAs indicates a decrease in diver-
sity, which is related to increasing aridity. (Cand D) Average cell-based ATP
concentrations were obtained by relating iATP concentrations (C) to total
biomass levels (D) measured at specific locations, which were obtained from
PLFA analysis.
Schulze-Makuch et al. PNAS |March 13, 2018 |vol. 115 |no. 11 |2671
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copies per gram soil). The same trend was also visible in the
eDNA pool although gene copy numbers were somewhat higher
than in the iDNA pool (SI Appendix, Table S1).
To provide a detailed characterization of living and dead
microorganisms, high-throughput sequencing of 16S rRNA gene
amplicons of both DNA pools was performed. In environments
supporting an active microbial community, the eDNA pool is
continuously replenished through biomass turnover of living cells
(14). As indicated by the large number of shared operational tax-
onomic units (OTUs), this was likely the case at CS where rel-
ative humidity (RH) and nutrients were highest, constantly sup-
plied by coastal fog. In contrast, OTUs identified at the hyperarid
locations showed less overlap between eDNA and iDNA com-
pared with CS, suggesting the dominance of autochthonous
microbial taxa rather than of inactive transitory microorgan-
isms periodically introduced by wind (Fig. 2A). Key organisms
of these communities consisted of unclassified Acidimicrobiales,
Actinobacteria,Alicyclobacillus,Burkholderia,Comamonadaceae,
and Xanthomonadaceae. Although abundances differed among
sites and soil depths, these characteristic taxa identified from
iDNA were found in all samples from all sites and depths, sug-
gesting a native and metabolically active desert core community.
Fig. 2. Microbial community structure and relationship between iDNA and
eDNA pools at six soil sampling sites in the Atacama Desert: CS, AL, RS, ME,
YU, and LB. (A) Venn diagrams of iDNA and eDNA OTU intersections for
samples collected at 0–5 cm and 20–30 cm depth. Numbers indicate the
numbers of different OTUs, and percentages refer to relative abundances
of reads unique to iDNA or eDNA. Bars to the left of the Venn diagrams
show relative abundances of bacterial orders in the subsets unique to the
iDNA and eDNA pools of the indicated sampling depth. (B) Classification of
iDNA pools from soil surface samples (0–5 cm) collected at the six sampling
sites in comparison with the iDNA and eDNA pools in subsurface soil layers.
(C) Classification of the subsurface iDNA pools (20–30 cm, 50 cm, 100 cm) in
comparison with the iDNA and eDNA pools in the surface soils (0–5 cm). The
bars show the percentages of OTU reads in the corresponding subsets, and
numbers indicate the numbers of different OTUs.
In general, as dryness increased, microbial diversity decreased,
analogous to previous observations (15, 16).
Abundance of Endospores. The dormant component of the bac-
terial communities was specifically assessed for the 2015 sam-
pling period by quantifying endospores, which are characteris-
tic of the phylum Firmicutes and stand out by their exceptional
resistance to environmental stresses. Dipicolinic acid (DPA),
a specific biomarker of intact endospores, was detected at all
sites. Endospore concentrations in surface layers, however, de-
creased with increasing aridity by almost two orders of magni-
tude (7.7 ×105to 1.5 ×104spores per gram soil; SI Appendix,
Table S2). The large size of this community suggests that an
extensive and persistent Firmicutes seed bank remains available
in the Atacama Desert, which is in agreement with the domi-
nance of isolates from this phylum in our cultivation experiments.
Importantly, the contribution of endospores to the iDNA pool
was likely minor, because the conventional extraction methods
that were used to extract iDNA from intact cells do not usu-
ally extract DNA from spores (17). Therefore, it is not surprising
that the phylogenetic composition derived from iDNA sequenc-
ing does not reflect the abundance of endospores from the phy-
lum Firmicutes (SI Appendix, Fig. S7).
Metabolic Activity. As many of the cultivated bacteria were
spore formers, we used independent complementary analytical
approaches to obtain conclusive evidence for living microorgan-
isms and their potential activity in hyperarid habitats.
First, evidence for microbial activity was obtained by a fluores-
cein diacetate hydrolysis assay to determine enzymatic activity
(SI Appendix, Table S3). Enzymatic activity was highest at the
surface of CS, ME, and LB sites, but still detectable at all other
sites and shallow depths (0–5 cm and 20–30 cm, respectively),
except for LB 20–30 cm, where it was below the detection limit
of 10−3nmol·g−1·h−1.
Second, our hypothesis of active, native microbial communi-
ties was supported by ATP analyses. These analyses allow the
separation of intracellular ATP (iATP), indicative of viable cells,
and extracellular ATP (eATP), indicative of ATP remaining in
the soil after cell lysis. iATP levels were up to three orders of
magnitude higher (3 ×10−12 mol·g−1) at CS compared with
the sampling sites located in the driest desert core (e.g., 2 ×
10−15 mol·g−1at YU 100 cm depth; Fig. 1). Overall, ATP anal-
yses supported the general trend of decreasing microbial activity
with increasing aridity, both along the studied moisture transect
(2015) and in YU surface soils from 2015 to 2017 (SI Appendix,
Table S1). The ATP analyses provide evidence that even in the
most arid sites of the Atacama core region native microorgan-
isms can be at times metabolically active.
Third, the presence of metabolically active microorganisms
was supported by the analyses of water-extractable metabolites
via direct injection electrospray ionization Fourier-transform ion
cyclotron resonance mass spectrometry [ESI(-) FT-ICR-MS],
which allowed the accurate calculation of elemental formulas.
On average, a rich signature of ∼1,600 elemental compositions
(CHNOS) was detected in all samples, indicating a geochemi-
cal footprint (18) typical of natural organic matter superimposed
by a biological footprint of fresh organic material (19) involving
amino acids, small peptides, and fatty acids (SI Appendix, Fig. S8).
Results suggest that water-soluble organic compounds consisted
mainly of aliphatic carbohydrates and fatty acids (CHO), while
nitrogen- and sulfur-containing compounds (CHNO and CHOS)
were less abundant. The relative abundance of compounds, pre-
dominantly reflecting metabolic activity, decreased significantly
along the aridity gradient from the coast to inland with con-
stant low amounts of organic compounds at the hyperarid loca-
tions of ME, YU, and LB. This trend again suggests a marked
decline in metabolic activity from moist to hyperarid soil habitats
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(SI Appendix, Fig. S9). Nevertheless, there was clear conserva-
tion of biosignatures even in the hyperarid locations, showing evi-
dence of past and, most likely, recent metabolic activity, especially
at a depth of 20–30 cm in YU (2015), where different types of
metabolites were distinguishable (SI Appendix, Fig. S9F).
Fourth, the metagenomic analysis from the soil samples ob-
tained in April 2015 indicated that microorganisms were active
even in the driest soil samples. Sequence abundance of mycobac-
teriophages, gordoniaphages, and streptomycophages correlated
positively with that of their respective hosts found in the differ-
ent samples [Spearman coefficient correlations of ρ= 0.69 at CS,
ρ= 0.88 at ME, and ρ= 0.67 at YU (SI Appendix, Fig. S4 C–E)].
The detected virus–host relationships seem to be consistent with
the observations that microbial blooms are followed by a bloom
of phages specific for the microbes that dominated the micro-
bial bloom (20, 21). However, while most identified viruses are
phages persistent in the environment that have also been iden-
tified by other desert studies (22), the vast majority of the iden-
tified viruses (95+%) belong to the family Caudovirales, which
includes both virulent and temperate members. Thus, we cannot
exclude the presence of a large number of undetected prophages
that might stay dormant for long periods of time.
Finally, we investigated in situ genome replication rates of
organisms via a genome-resolved metagenomics approach (23)
of samples retrieved from YU and ME. For YU, we recon-
structed a draft genome of the most dominant organism (uncul-
tivated Actinobacteria) with an estimated completeness of 92%
A
BC
Fig. 3. Genome-resolved metagenomics analyses and results. (A) Work flow
and main results from genome-resolved metagenomics. For details please
see SI Appendix,Read-Based Metagenomics, Genome-Resolved Metage-
nomics and in Situ Replication Rates. Genome replication forks are symbolic.
(B) Overview of iRep values retrieved from the four genomes from the YU
and ME sites (color codes correspond to those in A). Dashed line at value 1
marks threshold at which no replication occurs. Dashed line at value 2 marks
where each genome of a population has on average bidirectional repli-
cation taking place (24). (C) Rank-abundance curves based on rpS3 genes.
Colored genomes correspond to those in Aand B. For the YU site (Left rank-
abundance curve) the most dominant organism was reconstructed, and all
other genomes were fragmented. For the ME site (Right rank-abundance
curve) three genomes were reconstructed. The three most abundant organ-
isms were Actinobacteria, which were similar in GC and abundance.
based on single-copy genes (Fig. 3). The iRep value (24) of
the genome was 1.57, which is indicative of its slow replication.
For the ME site, we retrieved three high-quality draft genomes
belonging to members of the phyla Chloroflexi,Actinobacteria,
and Saccharibacteria (completeness 86–98%; Fig. 3). The in situ
replication rates of these genomes varied between 1.86 and 3.31.
The lower replication rates compare with literature values of
a wide array of organisms across multiple phyla, but the iRep
value of 2.48 for the Chloroflexi indicates that each genome in the
population has on average one bidirectional replication ongo-
ing (24). The genome replication rate for the Actinobacteria was
extremely high, which indicates that each genome of this popu-
lation had several replication forks at the time of sampling, thus
providing strong evidence for microbial activity.
A Transitory Habitat? The continuous decline of nonstructural
water at 20–30 cm depth at YU, from 2.7% by weight in 2015
to 0.2% and 0.1% in 2016 and 2017, respectively, suggests
temporarily favorable conditions for the activity of specialized
microorganisms after the rare precipitation event until water
activity fell again beneath a critical threshold (Fig. 4A). Miner-
alogical data confirmed a desiccation process in the later years
as some of the gypsum at YU 20–30 cm dehydrated to anhy-
drite (Fig. 4B). The steep decline of recovered iDNA by three
to five orders of magnitudes (Fig. 4 Cand D) indicates that the
sampling campaign in 2015, shortly after a major and very rare
rainfall event, tapped into a temporary, time-constrained habi-
tat, rather than a permanent one. ATP analyses, indicative of
active organisms, also support that assessment. The iATP values
follow that trend, declining by more than three orders of magni-
tude in the YU surface soil (Fig. 4E), but remain constant in the
deeper soil layer at YU (Fig. 4F), pointing to a longer retention
of microbial activity. Possibly some water released by the desicca-
tion of gypsum to anhydrite remains accessible to a specific part
of the microbial community. This possibility is supported by iso-
topic evidence. The δD values for the waters in the hydrous sul-
fate minerals suggest that small amounts of water accessible to
microorganisms might be available even in these hyperarid soils
(SI Appendix, Table S4), e.g., in the form of thin H2O films at
mineral surfaces (25) or as a product of mineral–water exchange
reactions (26).
Two other rainfall events occurred in August 2015 and June
2016 (each 6.7 mm at Antofagasta), but it is unclear how much
rain fell at YU. No indication of that rainfall event was observed
in our sharply declining iDNA and iATP values for the surface
soils from April 2015 to January 2017. This suggests that both the
August 2015 and June 2016 events were either insufficient to trig-
ger temporarily habitable conditions and a microbial response
or, since they occurred several months before our next sampling
round, were too small and subsided by the time we resampled.
Davis et al. (27) estimated that 2 mm or more precipitation are
needed to provide free water for the support of biological activity
in the soil. In previous studies, Navarro-Gonzalez et al. (7) were
not able to recover any DNA from the YU site, and Warren-
Rhodes et al. (28) reported on the virtual absence of hypolithic
cyanobacteria. Certainly, the literature data can provide only a
qualitative assessment about the presence or activity of microor-
ganisms, because of both the methodologies used and the spatial
heterogeneity of the sites. In contrast to our study, it appears
that previous sampling campaigns did not tap into habitable
conditions.
Discussion
The Atacama Desert soil microbiome has evolved as a result of
the prevailing environmental conditions. While the soil surface
was dominated by desiccation and UV-resistant species (Geo-
dermatophilaceae and Rubrobacter), deeper layers with higher
salt content were dominated by halophilic bacteria such as
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Fig. 4. Comparison of sampling events of April 2015, February 2016, and
January 2017 at YU. (A) Available nonstructural water decreases significantly
from 2015 to 2017. (B) Some of the gypsum at YU 20–30 desiccated and
formed anhydrite. (Cand D) Intracellular DNA amounts indicative of living
organisms drop by several orders of magnitudes at 0–5 cm and 20–30 cm
depths. (Eand F) Intracellular ATP amounts indicative of active organisms
drop by several orders of magnitudes at 0–5 cm, but stay constant at 20–
30 cm depth.
Betaproteobacteria(Comamonadaceae) or Firmicutes (Bacillaceae,
Alicyclobacillaceae) (SI Appendix, Fig. S7) and halophilic Archaea
(i.e., Halobacteria) at CS 20–30 cm (SI Appendix, Fig. S5C).
Notably, microbiomes associated with the hyperarid soils were
dominated by Bacteria rather than Archaea, consistent with an
assessment of the global distribution of archaeal abundances in
soils (29). The amount of unique OTUs in the iDNA pool of
the surface and subsurface soils of the hyperarid localities ME
and YU was much lower than at the moister sites. OTUs shared
between iDNA recovered from the surface and eDNA from
deeper layers (Fig. 2B) and between iDNA from the subsurface
and eDNA from the surface (Fig. 2C) drastically increased com-
pared with those in CS, indicating distinct microbial populations
at different depths with increasing dryness. This indicates that
selection pressures for microorganisms were much higher in the
hyperarid surface soils than in the wetter coastal area, resulting
in species well adapted to the extreme dryness and UV radiation.
However, some salt-tolerant bacteria such as Acidimicrobiales,
Comamonadaceae, and Bacillaceae potentially survive in deeper
soil layers after being buried (SI Appendix, Fig. S7), e.g., by ongo-
ing atmospheric deposition of salts and sediments or by halo- and
thermoturbation of soils. Alternatively, microbial communities
might have persisted in the subsurface since the onset of deserti-
fication, or an initial community successively changed over geo-
logical time to cope with altered environmental conditions (5).
Our results suggest that incoming microbial “newcomers” have
at least been exposed to passive environmental selection, but
also maintain transient activity even in the deeper soil layers and
sustain viability in the Atacama Desert for very long time peri-
ods. However, it remains questionable whether the organisms
found in this environment are adapting to the harsh conditions
present. Bacteria reaching the Atacama Desert by atmospheric
processes have been exposed to desiccation and UV stress dur-
ing aerial transport, possibly for extended periods (30). This sug-
gests that environmental species filtering could be an impor-
tant factor contributing to shaping the indigenous microbial
communities. In line with this hypothesis, our shotgun metage-
nomics data revealed several genes associated with dehydra-
tion tolerance [e.g., groEL,dnaK,fadD,glgX-malZ,phaC (31)]
and radiation/desiccation tolerance [e.g., recQ (32)]. Immunoas-
says corroborated these metagenomic results by detecting ATP
synthase, GroEL, CspA, and DPS DNA-protecting proteins at
YU (50 cm), CS, and RS, and metaproteomic analyses of sam-
ples taken at YU also confirmed the presence of ATP synthase
and GroEL.
One additional challenge for microorganisms to persist at
both surface and subsurface locations is the low organic mat-
ter content characteristic of hyperarid soils. A higher TOC
and moisture content allowed a higher total microbial biomass
and diversity (Fig. 1 and SI Appendix, Fig. S6). For example,
chloromethane (CH3Cl) release during low-temperature ther-
molysis of surface soil samples, which is indicative of hetero-
bonded methyl groups of organic matter, was highest for CS
(∼300 ng·g−1) and much lower at the hyperarid sites (∼1–
5 ng·g−1) (SI Appendix, Fig. S10). Stable hydrogen isotope anal-
yses of the released chloromethane confirmed the biochemical
origin of the methyl group. The emission profiles of CH3Cl are
almost identical to observations made by the Curiosity rover on
Mars (33), where even harsher environmental conditions prevail
than in the hyperarid core of the Atacama Desert [lack of water,
scarcity of organic matter, high UV irradiation, and high salt con-
tent in the soils including bassanite and perchlorates (34)]. No
rain can fall from the Martian atmosphere today (35), but liq-
uid water can be present near the Martian surface in the form of
nightly snow storms/ice microbursts (36), fog (37), near-surface
groundwater (38), and possibly also from mineral dehydration
reactions (39). On Mars, the deeper soil layers with a higher
water activity and reduced exposure to environmental stresses
(e.g., UV irradiation, large daily temperature fluctuations) are
expected to be more suitable for supporting life. At YU this was
the case, with the gypsum-rich soil layer at a depth of 20–30 cm
containing a higher biomass and microbial diversity (Figs. 1 B
and Dand 2) and also retaining a similar level of activity beyond
2015 for at least 2 y more. Thus, we observe in the hyperarid core
of the Atacama Desert a transitory habitat with microorganisms
that are active for short periods of time and which can serve as a
reasonable working model for Mars.
Conclusions
Although both microbial biomass and diversity in the Atacama
Desert decrease with increasing aridity, our study shows that
even the lowest precipitation levels on Earth can sustain episodic
incidences of microbial activity. There is no single agreed-upon
method known to date reaching the bar of evidence for micro-
bial activity for such low-biomass environments. However, using
our complementary tool box of combining different method-
ologies, including unique genome-resolved metagenomics, we
have addressed the question of microbial activity and can answer
it positively for the sampling time after the major precipita-
tion event in 2015. Thereafter, the biomarkers for microbial
activity dropped dramatically, inferring that the transitory hab-
itable conditions have ended until the next major rain event may
occur, providing a sufficient amount of free water for the micro-
bial biota. The insights gained from the hyperarid core of the
Atacama Desert can serve as a working model for Mars, where
environmental stresses are even harsher. If life ever evolved
on Mars, the results presented here suggest that it could have
endured the transition from the early aquatic stage, through
increasing aridity cycles, and perhaps even found a subsurface
niche beneath today’s severely hyperarid surface.
Materials and Methods
Detailed methods, including a description of the sampling sites with numer-
ous figures and tables, are provided in SI Appendix. Two unique methods
were used. The e/i-DNA methodology is described in detail in an appropri-
ate subject journal (13), with some of the associated issues discussed else-
where (40). The validity of the e/i-DNA method is further supported by a
2674 |www.pnas.org/cgi/doi/10.1073/pnas.1714341115 Schulze-Makuch et al.
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EARTH, ATMOSPHERIC,
AND PLANETARY SCIENCES
ECOLOGY
positive correlation between microbial biomass and the Shannon index (SI)
calculated for iDNA (r=0.62), but not with either calcium or sulfate con-
centrations. Conversely, the SI for eDNA correlated with calcium and sulfate
soil concentrations (r=0.74 and 0.61, respectively), but not with biomass
(r=0.03). The other state-of-the-art method was used to measure in situ
replication rates of genomes by calculating the number of active replication
forks reconstructed from the metagenome sequences (15) (see SI Appendix,
Read-Based Metagenomics, Genome-Resolved Metagenomics and in Situ
Replication Rates for more details). All data reported in this paper are com-
piled in SI Appendix and have been archived at GenBank/EMBL under Bio-
Project ID PRJNA395196 and at EMBL-EBI under accession no. PRJEB20402
(sample IDs ERS1666624–ERS1666714).
ACKNOWLEDGMENTS. D.S.-M. acknowledges support by the European Re-
search Council Advanced Grant Habitability Of Martian Environments
(339231), which provided base funding for the study, including sample
collection. The fungal diversity assessment was supported by funding (to
H.-P.G.) through the Leibniz Senatsausschuss Wettbewerb Project MycoLink
and DFG Project Microprime (GR1540/23-1). F.K. received financial support
from the German Science Foundation (DFG KE 884/8-2). The 16S rRNA
gene amplicon (MiSeq) sequencing was financed through the Helmholtz
Research Program “Geosystem–The Changing Earth” and the data were
processed by Fabian Horn (GFZ German Research Center for Geosciences–
Helmholtz Center Potsdam). The stable isotopic composition of water was
carried out through the Europlanet 2020 Research Infrastructure supported
by the European Union’s Horizon 2020 research and innovation program
(654208). Endospore quantification benefited from funding by the Deep
Carbon Observatory through a Pilot Project (L.W.). Part of the organic geo-
chemical analyses were performed at Imperial College London, supported by
United Kingdom Space Agency Grant ST/N000560/1. V.P. and D.C.’s work was
supported by the Spanish Ministry of Economy and Competitiveness Grants
ESP2015-69540-R and RYC-2014-19446, respectively. G.V. was supported by
a Humboldt Research Fellowship for postdoctoral researchers. M.F. acknowl-
edges support from National Institute of Food and Agriculture Hatch Project
1014527. The Leibniz Institute of Freshwater Ecology & Inland Fisheries (IGB)
housed the workshop at which much of the presented work was coordi-
nated. We thank M. Degebrodt for technical assistance.
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