The search for life in the solar system is acomplex process. More than 20 yearshave passed since the last direct search

for extraterrestrial life. The planet of choicewas Mars and the reasons for looking for lifewere based upon centuries of speculation and alittle scientific data about the planet (Taylor1999). Two identical spacecraft called Vikingwere sent to Mars, each composed of an orbiterand a lander. One of the functions of the lan-ders was to look for microbial life at two sepa-rate locations on the surface. The three Vikinglander experiments and the gas chromato-graph–mass spectrometer (GCMS) demonstrat-ed that no terrestrial-type life was found adja-cent to the landers and by inference, the rest ofthe planet’s surface (Klein 1978, 1979).

Yet now the possibility of finding both extinctand living (extant) life on Mars is being givenserious consideration again. We have a betterunderstanding of life on Earth than when theViking experiments were conceived in the 60s,both at the physical and molecular level. Anyhabitat suitable for the growth of higher organ-isms will also permit microbial growth but, inaddition, there are many habitats unfavourableto higher organisms where microorganismsexist and flourish. Microbes can live in envi-ronments ranging from the deserts of Antarcti-ca (figure 1) – the coldest driest places on Earth(Wynn-Williams 1999), to hot springs – thehottest, wettest places on Earth (Cowan 1999).This gives us an idea as to where we can specu-late that life might reside on Mars or Europa.

Planetary habitabilityScientists have recognized that there is a closecoupling between life and geological and possi-

ble atmospheric conditions (McKay 1991).Logically, one of the key criteria identified wasthe stability of liquid water over geologicallysignificant periods of time. Liquid water is afundamental requirement for life, not onlybecause reactions occur in solution, but alsobecause it plays a fundamental role in life’schemistry. The other most important atoms arecarbon, nitrogen, sulphur and phosphorus.

Together with the elements in water (hydrogenand oxygen), they form the CHNOPS ele-ments. Two bodies that may satisfy these crite-ria in the solar system are ancient Mars(McKay 1997) and possibly one of Jupiter’sGalilean satellites, Europa (Hiscox 1999).

From the data provided by the Viking lan-ders, Mars Pathfinder and analysis of 14 mete-orites from Mars, we are reasonably confidentthat the CHNOPS elements are or were presenton Mars (Banin and Mancinelli 1995,Mancinelli 1996). Analysis of images takenfrom orbit by Mariner 9, Viking Orbiters 1 and2 and Mars Surveyor provide almost definitiveevidence that liquid water once flowed freelyon the Martian surface (Baker 1982, Carr1996). This implies that the ancient climatewas warmer and wetter, i.e. a thicker carbondioxide atmosphere (McKay and Stoker 1989).Both Voyager 2 and more recently Galileo havereturned abundant images which indicate thatEuropa may have a liquid-water interior sur-rounded by a frozen exterior (Squyres et al.1983, Belton et al. 1996, Anderson et al. 1997).However, no detailed analysis has been con-ducted for the CHNOPS elements on Europa,although some may be inferred (Hiscox 1999).

Given that the surface topography of Marsindicates abundant liquid water in the past, anorigin-of-life event may have occurred and ledto the evolution of cellular life (Hiscox 1999).Since the Martian surface is extremely hostilefor life, one place to look for living Martianorganisms would be below the surface, per-haps deep underground where liquid watermay still be in abundance (Boston et al. 1992).Based upon hydrological models that take intoaccount global Martian topography (Clifford

5.28 October 2000 Vol 41

Several emerging technologies

are being investigated for

detecting potential extraterrestrial

microbial life, either extinct and/or

extant (living). These technologies

are particularly relevant for future

exobiological searches on Mars and

Europa, the two most promising

candidates in this solar system for

harbouring extraterrestrial life. Two

systems are discussed: ATP

measurement and Raman

spectroscopy. In particular,

experiments are described in which

samples from Antarctica (an

analogue environment for ancient

Mars) have been analysed using

Raman spectroscopy. This

technology is perhaps the most

versatile for searching for both

extinct and extant organisms.

Exobiological prospectingEmma Newton, Howell G M Edwards, David Wynn-Williams and Julian A Hiscox review the combination of

biology, geology and physics necessary to search for life off Earth.

1: McKelvey Valley, McMurdo Dry Valleys region,Antarctica, seen from Shapeless Mountains. Thisice-free cold desert region was used to evaluatethe Mars Viking instrumentation and is still usedfor exobiological research.

Exobiological prospectingEmma Newton, Howell G M Edwards, David Wynn-Williams and Julian A Hiscox review the combination of

biology, geology and physics necessary to search for life off Earth.

Astrobiology

1993), Fogg (1999) predicted the existence ofartesian basins on Mars where pressurizedgroundwater may exist at relatively shallowdepths. Fogg (1999) suggested the possibilitythat such basins are extensive and couldinvolve Hellas and much of the northernplains. The implication of this is that the liq-uid-water resource on Mars might be easier toprobe and exploit than commonly assumed.Carr (1996) argued that the geothermal gradi-ent is such that Mars is likely to have liquidwater near the equator at depths as shallow asabout two kilometres.

Locating lifeBy studying different types of microbes fromextreme environments on Earth and in differ-ent habitats, we can paint a picture of whatand where life might be found on Mars andperhaps also Europa. Because microorganismsare usually invisible to the naked eye, theirphysical existence in an environment is oftenunsuspected – especially on other planets.Often it is not the microorganisms themselvesthat we observe in a natural environment, butinstead, like the Viking landers, we look forchemical evidence of their existence. Althoughphotosynthetic systems are energetically themost efficient way of obtaining energy formetabolism, there are anaerobic (oxygen-free)microbial systems that can obtain energy fromchemical transformations of iron, sulphur oreven hydrogen. The chemolithotrophic organ-isms that can do this do not require sunlightand can therefore evolve in the dark, geother-mally warmed ground-water of the deep sub-surface of Earth and hence possibly Mars(Boston et al. 1992). Anaerobic bacteria couldbe extant on present-day Mars, perhaps twokilometres beneath the surface, unfortunatelyout of the reach of current lander technology.

Another potential chemolithotrophic micro-bial community supported by anaerobic geo-thermal conditions could theoretically existbeneath the ice-crust of Jupiter’s moon Europa(Reynolds et al. 1983). Its eccentric orbit resultsin tidal friction, which may provide sufficientheat to permit the stability of liquid water. Asample return mission to Europa is even morecomplex than one to Mars, due to its increaseddistance from Earth. A favourable option is tosend several sophisticated robots that are able toexplore a Europan ocean. Currently, planetaryscientists are evaluating such techniques inAntarctica. An underground lake the size ofLake Ontario has been discovered 4.2 kmbeneath the central Antarctic ice sheet. The lake,called Vostok, has been isolated from the rest ofthe planet for over a million years. Scientists andengineers are developing an instrument called aCryobot – which can melt ice – and a Hydrobotsubmersible which will not only image theunderside of the ice and sediments, but will also

look for potentially life-supporting hot spots,analogous to those predicted for Europa.

Several procedures are available for lookingfor microbial activity. One of the most widelyused is measurement of respiration, as eitheroxygen uptake or carbon dioxide production. Asample of soil or water is incubated in a closedchamber under simulated natural conditionsand changes in one of these gases are measured.A similar principal was used by the Viking lan-ders – but with far more sensitivity. Here wereview three alternative techniques that holdpromise for prospecting for extant and extinctorganisms on Mars (and possibly Europa).

Adenosine triphosphateOne of the most important molecules for life onthis planet is adenosine triphosphate (ATP).ATP is the universal energy currency of a cell.ATP turns over rapidly in cells that are growing,i.e. metabolizing. Under starvation conditions,cellular ATP levels dip to a low value. Since ATPis lost very rapidly from dead or dormant organ-isms, ATP measurements provide a measure-ment of living biomass. In ecological studies, asample of water or soil is treated to remove theATP from the microbial cells, and the ATP levelof the extract can be easily measured by biolu-minescence (LaFerla et al. 1995). ATP measure-ments have been made most frequently in theoceans, where microbial numbers are low andwhere very sensitive methods to detect microbialactivity are needed. If we are considering thistechnique for a Mars lander, then the assump-tion would have to be made that ATP is used bya Martian biota. However, other nucleotidescan enhance turnover of ATP-dependent lumi-nescence (Ford et al. 1996), so alternatively anATP-based bioluminescence experiment couldbe carried in an experimental package on a lan-der to provide an assay for nucleotides presentin an extraterrestrial environment.

Raman spectroscopyWe are applying Raman spectroscopy to theanalysis and characterization of biologicalmaterials in geological environments. The tech-nique involves illumination of the sample witha laser beam and subsequent measurement ofscattered light. The Raman spectrum provides aunique fingerprint of the molecular groups ofinterest. Characteristic peaks from the spec-trum can be used to identify a target compoundamong other mixed samples. In addition,Raman spectroscopy is able to distinguishmaterial of both biological and non-biological(geological) origin. The focused laser beamallows analysis of discrete biological layerswithin mineral substrata and this can be furtherenhanced by using Raman microscopy; typicalfootprints for sample illumination are in theregion 1–100 mm. Visible lasers can be used toanalyse biomolecules and inorganic materials

of the mineral habitat (rock or soil). However,pigments of the photosynthetic microbes,which use sunlight to drive surface communi-ties, fluoresce under blue–green illumination. Inorder to study such organisms, which wereamong the earliest colonists of Earth over3.5 billion years ago, an infrared laser has beenused, so minimizing the effect of fluorescence.As cyanobacteria may have evolved on Marsduring the “warm wet” period at the same geo-logical time as on Earth (Davis and McKay1996), this approach could be used to detecttheir fossil biomolecules in Martian sediments.

Using a laboratory-based infrared Ramanspectrometer, we have examined translucentsandstone samples containing cryptoendolithicmicroorganisms from the Antarctic dry valleys.Figure 2 shows the Raman spectrum of theprofile of an endolithic community containingan algal zone (figure 3). The exposed outercrustal material showed a strong silica signal ofthe sandstone matrix at wave numbers 264and 464 cm–1, similar to the bedrock; noorganic material was detected. The black pig-mented layer of varying thickness in the lichenzone contained only calcium oxalate dihydrate(1472 cm–1), but no calcium monohydrate wasdetected, possibly due to a low signal-to-noiseratio. This may have been caused by absorp-tion of laser energy by the black pigment. Thewhite layer of the lichen zone consisted ofmainly hyaline fungal hyphae covered in whitecrystals of calcium oxylate monohydrate (1462and 1488 cm–1), but no calcium oxylate dihy-drate or calcium carbonate. The zone also con-tained some microalgae as evidenced by broadfeatures at 1320 cm–1, indicative of chloro-phyll, and a weak band at 1525 cm–1, indica-tive of the n(C=C) of carotenoid pigments.There were also several peaks in the2700–3200 cm–1 region indicative of organiccomponents of the microbial cell walls. Thegreen microalgal zone gave a variety of peaks,including chlorophyll and carotenoids similarto those of the white lichen zone. In this zone,additional peaks at 1659 and 1260 cm–1 areindicative of amides in cellular components.However, this algal zone contained 90% lesscalcium oxalate (1462 cm–1 band) than thewhite lichen zone containing abundant fungalmycelium. Spectra from the accumulation (ironrich) and the inner zones yielded very similarbackground spectra of abiotic sandstonedevoid of organic material.

Endolithic communities, up to 8 mm insidethe fabric of rock, might have been the finalsurvivors on Mars (Friedmann and Koriem1989) as the surface water was either frozeninto the permafrost or was lost from the surfacealtogether. So the Antarctic habitat providesone of the best models for life on ancient Mars(Wynn-Williams 1999). The biological signa-tures, or bio-markers, left behind by Martian

Astrobiology

5.29October 2000 Vol 41

endolithic microorganisms could be comparedwith an Antarctic database of the fingerprintsof similar biomolecules for interpreting futureexobiological surveys of Martian near-surfacelayers (Wynn-Williams and Edwards 2000).

Laboratory-based infrared Raman spectrome-ters weigh tens of kilograms and require highpower consumption. If they are to be incorpo-rated onto Mars landers then their weight andpower consumption must be reduced (Edwardsand Newton 1999). These instruments are deli-cate, too, and the shock of a hard landing onMars is a significant consideration, as well as afluctuation in surface temperatures of up to150 °C. Potential improvements to be consid-ered are CCD detectors, fibre optics, solid-statediode lasers and holographic filters. Theseimprovements would need to reduce the remoteRaman spectrometer to 1 kg or less, suitablefor biological studies and inclusion on a lander.One such instrument is being developed jointlyfor Antarctic evaluation and future Mars mis-sions by the University of Montana (USA), theBritish Antarctic Survey and the University ofBradford (Dickensheets et al. in press).

SummaryAs our knowledge of the survival limits ofmicrobial life on Earth broadens with the dis-covery of organisms in increasingly hostilehabitats, so we must broaden our view of thelikelihood of finding evidence of former, oreven current, life on Mars and/or Europa. Thechallenge will be finding sites where bio-molecules are preserved for comparison withour increasing Earthly database of microbialfingerprints. �

Emma Newton and Howell G M Edwards, Chemi-cal and Forensic Sciences, University of Bradford,UK; David Wynn-Williams, British Antarctic Sur-vey, Cambridge; Julian A Hiscox, School of Animaland Microbial Sciences, University of Reading, UK.Correspondence to: j.a.hiscox@reading.ac.uk orSchool of Animal and Microbial Sciences, Universityof Reading, Whiteknights, PO Box 228, ReadingRG6 6AJ, UK.

ReferencesAnderson J D et al. 1997 Science 276 1236–1239.Baker V R 1982 The Channels of Mars University of Texas Press.Banin A and R L Mancinelli 1995 Adv. Space. Res. 15(3)(3)163–(163)170.Belton M J S et al. 1996 Science 274(5286) 377–385.Boston P J et al. 1992 Icarus 95 300–308.Carr M H 1996 Water on Mars Oxford University Press.Clifford S M 1993 J. Geophys. Res. 98 10 973–11 016.Cowan D A 1999 The Search for Life on Mars ed. J A Hiscox 37–48British Interplanetary Society.Davis W L and C P McKay 1996 Ori. Life Evol. Biosph. 26 61–73.Dickensheets D L et al. in press J. Raman Spectroscopy 31.Edwards H G M and E M Newton 1999 The Search for Life onMars ed. J A Hiscox 83–88 British Interplanetary Society.Fogg M J 1999 The Search for Life on Mars ed. J A Hiscox 66–72British Interplanetary Society.Ford S R et al. 1996 J. Biolum. Chemilum. 11(3) 149–167.

Friedmann E I and A M Koriem 1989 Adv. Space. Res. 9(6)167–(166)172.Hiscox J A 1999 Astronomy & Geophysics 40 2.22–2.26.Hiscox J A 1999 The Search for Life on Mars ed. J A Hiscox 18–25British Interplanetary Society.Klein H P 1978 Icarus 34 666–674.Klein H P 1979 Rev. Geophys. 17 1655–1662.LaFerla R et al. 1995 Marine Ecology 16(4) 307–315.Mancinelli R L 1996 Adv. Space. Res. 12 (12)241–(212)248.

McKay C P 1991 Icarus 91 93–100.McKay C P 1997 Ori. Life Evol. Biosph. 27 263–289.McKay C P and C R Stoker 1989 Rev. Geophys. 27(2) 189–214.Reynolds R T et al. 1983 Icarus 56 246–254.Squyres S W et al. 1983 Nature 301 225–226.Taylor R S 1999The Search for Life on Mars ed. J A Hiscox 3–17British Interplanetary Society.Wynn-Williams D D 1999The Search for Life on Mars ed. J A Hiscox49–57 British Interplanetary Society.

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3000 2500 2000 1500 1000 500

inte

nsity

(arb

itrar

y un

its)

wavenumber cm–1

iron-stained crust

black lichen

hyaline lichen

micro-algae

inorganic accumulation

bedrock

COx

CarChl

Car

Ami

Qua Sil

CWa

2: Compilation Raman spectrum for the profile of a visible endolithic community in sandstone fromEast Beacon, Antarctica (corresponding to the starat in figure 3). The baselines of spectra for layersdescribed in the text have been displaced to show significant vibrational peaks indicative of organicand inorganic compounds. Car = carotenoid; COx = calcium oxalate; Chl = chlorophyll; CWa = cellwall material; Ami = amide; Qua = quartz; Sil = silica.

3: A vertical profile of an endolithic community in sandstone from East Beacon, Antarctica, consistingof: an outer iron-stained crust; a black lichen zone; a white hyaline lichen zone; a green algae layerabout 8 mm inside the rock; a light coloured accumulation layer; inner bedrock. Scale bar = 5 mm.