Essays

The Quest for Extraterrestrial Life:What About the Viruses?

Dale Warren Griffin

Abstract

Recently, viruses have been recognized as the most numerous entities and the primary drivers of evolution onEarth. Historically, viruses have been mostly ignored in the field of astrobiology due to the view that they are notalive in the classical sense and if encountered would not present risk due to their host-specific nature. What wecurrently know of viruses is that we are most likely to encounter them on other life-bearing planets; that whilesome are exquisitely host-specific, many viruses can utilize hundreds of different host species; that viruses areknown to exist in our planet’s most extreme environments; and that while many do not survive long outsidetheir hosts, some can survive for extended periods, especially in the cold. In our quest for extraterrestrial life, weshould be looking for viruses; and while any encountered may pose no risk, the possibility of an encounter witha virus capable of accessing multiple cell types exists, and any prospective contact with such an organism shouldbe treated accordingly. Key Words: Astrobiology—Microbiology—Origin of life—Pathogens—Virus. Astro-biology 13, 774–783.

The study of viruses has not been applied in Astrobiology to the extent of other disciplines. This is despite viruses outnumbering allother organisms on earth by at least an order of magnitude. (Stedman and Blumberg, 2008)

Viruses: What Are They?

Viruses are the most numerous entities on Earth, with anestimated *1030 residing in our oceanic water column

and *1031 for the total planetary population (Breitbart andRohwer, 2005; Suttle, 2005; Youle et al., 2012). Suttle (2005)estimated that if you put the population of viruses that existin the water column of our oceans next to each other, oneafter the other in a straight line, and you anchored one endof that line to the surface of Earth, the resulting viral stringwould extend out into space *10 million light years. Thetypical number of viruses, most of which are bacteriophages(viruses whose hosts are bacteria), ranges from *106 to 108

per milliliter of seawater, and in near-shore marine sedi-ments and topsoils the number of viruses is approximately108 per cubic centimeter or gram (Breitbart and Rohwer,2005; Suttle, 2005; Srinivasiah et al., 2008). Lower concen-trations of viruses are known to exist in subsurface envi-ronments. In regard to diversity, the number of viralgenotypes in a kilogram of marine sediment has been esti-mated to be as high as 106 (Breitbart and Rohwer, 2005;Edwards and Rohwer, 2005). Viruses are found wherevercellular life exists, to include those that reside in the most

inhospitable environments (i.e., deep sea hydrothermalvents, hot springs, cold water, snow, and ice) (Kaminskyyand Zhivotovsky, 2010).

Viruses are composed of a genome surrounded by aprotective genome-encoded protein shell known as a capsid.In addition to the capsid, some viruses contain a lipid en-velope that contains viral and/or host-derived proteins.There are also ‘‘naked viruses’’ or viroids and the recentlydiscovered virophage (La Scola et al., 2008).

Most viruses range in size from 20 to 300 nm in contrast tobacteria that typically range from 500 to 1500 nm (Fig. 1)(Nayak, 2011). For example, Poliovirus, one of the members ofthe common human enterovirus group, is typically in the sizerange of 30 nm, but some viruses are bacterial in size. TheMimivirus has a diameter of *400 nm, and the filamentousEbola virus ranges from *900 to 14,000 nm in length (Geisbertand Jahrling, 1995; La Scola et al., 2003). Viruses grouped bytypes of genomic nucleic acid are listed in Table 1 and includegenomes composed of RNA or DNA variants. Currently in thefield of viral taxonomy, there are 6 recognized orders com-prising 22 families, 109 genera, and 549 species. In addition,there are 65 viral families that are not currently classified intoorders that contain 265 genera and 1734 species (ICTV, 2009).

U.S. Geological Survey, St. Petersburg, Florida.

ASTROBIOLOGYVolume 13, Number 8, 2013ª Mary Ann Liebert, Inc.DOI: 10.1089/ast.2012.0959

774

The question of whether viruses can be classified as aliveis one that has been debated since their discovery (Beijerinck,1899; Villarreal, 2004; Moreira and Lopez-Garcia, 2009). InVillarreal’s article titled ‘‘Are Viruses Alive?’’ he points outhow they have been viewed as ‘‘inert chemicals’’ or, due theirdependence on host cells for replication, living ‘‘a kind ofborrowed life’’ (Villarreal, 2004). Viruses dock with a hostcell membrane and use membrane fusion to ‘‘trick’’ the hostcell to internalize it or inject their genomes through theirhost’s cell wall/membrane. Once the virus has gained entryinto its host, it can enter a state of coexistence or pirate hostsystems for the purposes of replication (parasitic or lethalinfections). The production of more copies of oneself orprodigy is a hallmark of life, but the requirement of an ob-ligate host confounds the debate of whether viruses are alive(Moreira and Lopez-Garcia, 2009). While most regard viru-ses as pathogenic in nature, many virus-host relationshipsare asymptomatic in nature. In fact, viruses can shuttlebeneficial genes (i.e., antibiotic resistance, virulence, photo-synthetic, etc.) to a host that provide the host with enhancedability to compete in its environment (Mann et al., 2003;Roossinck, 2011). Regardless of one’s view as to whether avirus is alive or not, their ability to move genes betweenhosts has driven the evolution of life on this planet (Villar-real, 2004; Forterre, 2010).

Viral Host Relationships

Speculation on the origin of viruses has included the hy-pothesis that viruses occurred before the more complex pro-karyotes based on the existence of widespread genes that areonly found within viral genomes (virus hallmark genes, i.e.,reverse transcriptase, RNA-dependent RNA polymerase,jelly-roll capsid protein, and rolling-circle replication endo-nuclease) (Koonin et al., 2006; Forterre, 2013). Otherhypotheses have included that viruses are escaped genesor degenerative unicellular organisms that lost genetic capa-bility to replicate on their own (Bubanovic et al., 2005; Villar-real, 2005). While the origin of viruses is still an unresolvedissue [to date there is no viral fossil record in comparison tothat observed with cellular life (Laidler and Stedman, 2010;Orange et al., 2011)], the most commonly agreed point of viewis that viruses are ancient and polyphyletic, and evolved in-dependently of host replication systems (Villarreal, 2005;Moreira and Lopez-Garcia, 2009; Forterre, 2010).

In nature, viral-host relationships can be detrimental orbeneficial to the host. An example of a beneficial transfer ofgenetic material to a host by viruses is the bacteriophagegenes that encode Shiga-like exotoxin (Imamovic et al., 2009).This bacteriophage-mediated genetic transfer is believed tobenefit the bacterial host by enabling compromised host cells

FIG. 1. Comparative sizes of select microorgan-isms and genomic DNA. [Figure courtesy of Rey-nolds, K.A. and Pepper, I.L. (2000) Microorganismsin the environment. Chapter 2, Figure 2.2, p 11,published in Environmental Microbiology, edited byR.M. Maier, I.L. Pepper, and C.P. Gerba, ISBN 0-12-497570-4, copyright Elsevier B.V.] Color graphicsavailable online at www.liebertonline.com/ast

ASTROVIROLOGY 775

to release a toxin that is inhibitory to predators (protestgrazers) or the bacteria’s host (triggered by immune re-sponse) (Imamovic et al., 2009; Lainhart et al., 2009). Otherexamples of bacteriophage-mediated virulence conversionare bacteriophages that impart the capability of certain pro-karyote species to produce toxins (i.e., cholera, diphtheria,and staphylococcal) and antibiotics (Freeman, 1951; Betleyand Mekalanos, 1985; Waldor and Mekalanos, 1996). Inaquatic environments, bacteriophages have been shown tomove photosynthetic genes and thus sustain photosyntheticcapability in phototrophs (Mann et al., 2003).

An infected eukaryotic cell committing apoptosis (a primeexample of cellular altruism) is an example of a detrimentalrelationship. Some detrimental infections may be asymp-tomatic and not produce symptoms. Asymptomatic infec-tions may result in the assimilation of the viral genomewithin its host (termed lysogeny for bacteriophage infectionsof this type) followed by a latent or dormant phase. After aperiod of dormancy, latent or asymptomatic infections canenter a lytic stage that is typically triggered by some form ofhost cell stress ( Jiang and Paul, 1996; Williamson et al., 2002).The lytic stage results in rapid reproduction of the virusending with the release of the replicated viruses and oftenthe destruction of the host cell. The number of releasedviruses (known as burst size) can range from a few tothousands (Ellis and Delbruck, 1939; Bailey et al., 2004; Choiet al., 2010; Minor, 2011).

In regard to host specificity, viruses can be grouped intotwo classes: those that only infect a given host or host tissue

type (specific viruses) and those that can infect more thanone host (generalized viruses). Polio, mumps, and the den-gue viruses are examples that only cause disease in primates.Bacteriophages like the F-specific bacteriophage f1 only in-fect hosts that produce pili, which are bacterial cell surfaceprotein appendages used for conjugation ( Jacobson, 1972).Bacteriophages with broad host ranges have been shown toinfect many species within a given genus and in some in-stances to be able to infect multiple genera ( Jensen et al.,1998; Evans et al., 2010). Multi-host eukaryote viruses includemembers of the caliciviruses that are known to infect bothmarine and terrestrial mammals. Researchers have estimatedthat an infected 35-ton gray whale could shed *1013 calici-viruses daily (which could survive > 2 weeks at 15�C) andhypothesized that these types of oceanic reservoirs couldserve as inoculum for terrestrial hosts (Smith et al., 1998). TheAvian influenza virus is capable of infecting a number ofvertebrate hosts, which to date have included humans, pigs,horses, and birds (Naffakh et al., 2008). The cucumber mosaicviruses (vector-borne by aphids) have a host range of over800 species of plants (Palukaitis et al., 1992).

The benefit of host specificity is ‘‘finding a niche’’ whereadaptation to evolving host defense systems is limited to afew or one coevolving genome (Elena et al., 2009). The dis-advantage of host specificity is the probability that a newlyreplicated virus can find a host upon cellular release. Thebenefit of host generalization is the availability of manyhosts, which thus increases the likelihood that a newly rep-licated virus can find a receptive cell to infect. The

Table 1. Viral Groups Based on Genome Nucleic Acid Type

Nucleic acid Host Examples, Family—common name

RNA virusesSingle negative stranded Plants Bunyaviridae—Tomato spotted wilt virus

Invertebrates Rhabdoviridae—Y-organ virus (crab virus)Vertebrates Filoviridae—Ebola virus

Single positive stranded Bacteria Leviviridae—Enterobacteriophage MS2Fungi Barnaviridae—Mushroom virus XPlants Bromoviridae—Tobacco streak virusInvertebrates Tetraviridae—Nudaurelia beta virus (moths and butterflies)Vertebrates Picornaviridae—Poliovirus

Double stranded Bacteria Cystoviridae—Pseudomonas phage F6Fungi Chrysoviridae—Penicillium brevicompactum virusPlants Partitiviridae—White clover crytic virus 1Invertebrates Birnaviridae—Drosophila X virusVertebrates Reoviridae—Rotavirus

Single positive stranded—reverse transcriptase

Fungi Pseudoviridae—Saccharomyces cerevisiae Ty1 virusInvertebrates Metaviridae—Ascaris lumbricoides Tas virusVertebrates Retroviridae—HIV

DNA virusesSingle stranded Bacteria Inoviridae—Vibrio phage CTX

Plants Geminiviridae—Beet curly top virusInvertebrates Parvoviridae—Aedes aegypti densovirusVertebrates

Double stranded Bacteria Myoviridae—Enterobactriophage T4Amoeba, Algae, Fungi Mimiviridae—Acanthamoeba polyphaga mimivirusInvertebrates Iridoviridae—Tiger frog virusVertebrates Adenoviridae—Human Adenovirus species A–G

Double stranded—reversetranscriptase

Plants Caulimoviridae—Cauliflower mosaic virusVertebrates Hepadnaviridae—Hepatitis B virus

776 GRIFFIN

disadvantage is intracellular competition with co-infectingviruses and adapting to the evolution of multiple host de-fense systems (Elena et al., 2009). While experiments utilizingbacteria, plant, and mammalian viruses that typically infect asingle host demonstrated lost fitness (a lowered ability toefficiently infect) when forced to adapt to an alternate host,viruses such as the Foot and Mouth Disease Virus havedemonstrated the ability to acquire new hosts without losingoriginal host fitness (Elena et al., 2009).

Earth’s viruses, whether they have lytic or lysogenic hostrelationships, must survive the extracellular environment tofind new host cells. Based on our understanding of cellularlife on Earth and the driving role that viruses have played inevolution, it should be expected that, if viable extraterrestrialcellular life were discovered, viruses or virus-like entitieswould also be present. If we discover a planet that has suf-fered cellular extinction, it is possible that the last viableentity on the planet was viral or viral-like. If this is the caseand viruses are shed by the last remaining cellular hosts intovarious extracellular environments, what is the likelihoodthat these viral entities would remain viable for any ex-tended period of time? What do we know about extracellularviral survival on Earth?

Virus Survival and Life in Extreme Environments

Table 2 lists the results of virus survival experiments thatwere obtained for various extracellular environments thatinclude water, soil, and space. Unlike their metabolicallyactive hosts, viruses are not metabolically active outside thehost cell, do not require osmotic maintenance to remain vi-able, and thus are ideally suited for long-term survival. Inmarine surface water, UV-induced nucleic acid damage wasidentified as a primary contributor to viral inactivation inaddition to losses driven by particulates and heat-labilesubstances in the water column (Noble and Fuhrman, 1997).Gerba and Schaiberger (1975) had previously reported a re-duction in viral infectivity due to the association of viruseswith marine particulate matter but noted that this associationresulted in an increase in virus survival. While UV radiationcan kill viruses and host cells, viruses carrying matched orcompatible genes to those damaged in their prospective hostcan infect and thus restore host function (Villarreal, 2004;Mann et al., 2005). UV-damaged viruses are capable of in-jecting their genomes into host cells, and replication of thevirus can be restored if infection by multiple viruses resultsin an equivalent undamaged viral genome. Villarreal (2004)stated that ‘‘Viruses are the only known biological entitywith this kind of ‘phoenix phenotype’—the capacity to risefrom their own ashes.’’

While elevated temperature can be lethal to many viruses,there are linages that evolved to tolerate extreme heat envi-ronments such as those found in hot springs and aroundmarine geothermal vents (Anderson et al., 2011; Yoshida-Takashima et al., 2012). In two Yellowstone National Parkhot springs, where temperatures and pH ranged from 75�Cto 93�C and 1.5 to 6.5, respectively, 12 different morphotypesof viable archaeal viruses were reported (Rachel et al., 2002).In a study conducted in Italian volcanic acidic (pH 1.5) hotsprings where temperatures ranged from 87�C to 93�C, fivedifferent morphotypes of viable archaeal viruses (determinedvia enrichment assays and transmission electron microscopy)

were reported (Haring et al., 2005). In addition to thoseviruses that have evolved to thrive in extreme heat envi-ronments, likewise there are those known to have evolved totolerate extreme cold environments.

At an internal depth of 0.5–1.5 cm in Artic sea ice, wherehost activity was greatest, virus concentrations in melt ran-ged from 9.0 · 106 to 1.3 · 108 mL - 1,which was up to 100-foldhigher than that observed in the underlying water column(Maranger et al., 1994). Viable viruses were also observed ata temperature of - 12�C in sea-ice brine (Wells and Deming,2006). In Antarctica, viruses were observed associated withmoss that typically infect dicotyledonous plants, which rai-ses questions of host specificity and range in this cold-weather environment (Polischuk et al., 2007). Extreme cold orcryogenic storage is widely used to preserve microorgan-isms. Stocks of Influenza viruses as old as 40 years that werecryogenically stored ( - 70�C) demonstrated little loss of vi-ability (Merrill et al., 2008). With regard to long-term virussurvival, Hollings and Stone (1970) demonstrated that atroom-temperature storage 57 lyophilized stocks of plantviruses out of 74 remained viable after 1 year, and 19 of 74remained viable after 10 years. Priscu et al. (2006) argued foran improved understanding of life in ice on Earth to enhanceour search for life on other icy worlds.

There are other instances (prophage in dormant or pre-served prokaryotes) yet unexplored where viruses mightsurvive considerable periods of time. Recently, viable bac-teria and fungi were cultured from desert dust samples(Saharan desert dust that fell onto ships sailing in the At-lantic) collected in 1838 and given to Charles Darwin (Gor-bushina et al., 2007). Could these bacteria and fungi that weredormant for over 160 years harbor inducible viruses (Gor-bushina et al., 2007)? Likewise, what of the potential presenceof prophage in a bacterium reported to have been revivedfrom a 25- to 40-million-year-old amber sample (Cano andBorucki, 1995)? It should be emphasized that our currentlevel of understanding of bacterial survival is limited by ourinability to culture the majority of these organisms, and thefield of virology is immature in comparison. Issues such aslong-term virus survival are hampered by our current lack ofculturable hosts. Does our understanding of virus survivaland viral host specificity translate well to what we may en-counter on other planets?

Cave Exiguus Creatura (Beware of the Tiny Creature)

As viruses are the most numerous entities on Earth, it islikely that we will encounter them in extraterrestrial habitatsthat harbor cellular life. Whether cellular life is sparse or not,the use of viruses or viral constituents as biomarkers may aidour efforts to identify life or understand how life originatedon extraterrestrial bodies ( Jalasvuori et al., 2009).

From our current state of knowledge of viruses and cel-lular specificity, the potential threat of extraterrestrial virusesto life on our planet via sample return missions or to astro-nauts is viewed with questionable risk. In regard to plannedsample return missions, the current debate on an acceptableparticle escape threshold that is based on the estimated lowersize limits of ‘‘living’’ cells (*50 nm) should be oriented tothe potential threat that may come from the lower size rangeof our known viruses, many of which exist in the range of< 50 nm (ESF/ESSC, 2012). One of the arguments in support

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of an astronaut low-risk scenario (or return via astronautcontamination) is that life may have been transported andseeded onto habitable planets and moons from a biologicallyrich planet (like Earth) via impact event ejecta (lithopan-spermia) (Nicholson et al., 2005) and thus when encounteredwill not truly be extraterrestrial or foreign in nature. Al-though this type of extraterrestrial encounter may indeedpose minimal risk, it could be argued that there is riskthrough long-term separation of populations and/or lack ofimmunity (even if restricted to prokaryote populations)much like what is seen on the human scale with travelers’diarrhea, the spread of smallpox by Europeans into NativeAmerican populations, and the introduction of the Myxomavirus into rabbit populations in Australia (Fenner, 1959;Jones, 2006).

Another possibility is that life we may encounter on otherplanets or moons evolved independently of life on Earth. Thelow-risk argument here is that any viruses that infect or-ganisms we encounter have specificity to their hosts and thuswill not be able to infect organisms of Earth origin. Althoughhost specificity is a trademark of Earth’s viruses, the exis-tence of generalized viruses like the cucumber mosaic virus,which can infect many different hosts within the kingdomPlantae, is well documented. Further, there is genetic evi-dence that some vertebrate viruses may have evolved froman ancient incidental infection of a vertebrate by a plant virus(Gibbs and Weiller, 1999). It may be that life on other planetsevolved so differently from the manner in which it evolvedon Earth that an extraterrestrial virus could not gain access toa terran cell, but it is also as likely that life evolved in a verysimilar manner as it evolved on Earth and that viral pene-tration into a terran cell is not insurmountable.

There are several hypothetical scenarios where an extra-terrestrial virus capable of infecting various cell types mayevolve. One hypothesis in regard to the evolution of lifeon Earth is the ‘‘clay hypothesis,’’ which asserts that self-replicating clays and associated organic molecules providedtemplates for primitive organic synthesis (Cairns-Smith,1966). In this hypothesis, biochemistry evolved from geo-chemistry due to natural selective environmental pressureson inorganics (Cairns-Smith, 2005). Clay structural environ-ments can provide a shielded cellular-like environment fornucleic acid (i.e., UV shielding) and cellular evolution, andthis potential mode of life evolution may have occurred onother planets or extraterrestrial bodies (Cairns-Smith andHartman, 1986; Holm et al., 1992; Scappini et al., 2004). It hasbeen hypothesized, based on the existence of widespreadgenes only found within viral genomes, that a ‘‘virus world’’may have evolved from inorganic precellular compartmentsand coevolved with emerging cellular life (Koonin et al.,2006). In this scenario, it is possible that extraterrestrialviruses at an early evolutionary stage of cellular coevolutionmay not have yet acquired host specificity and, if encoun-tered, are capable of infecting cellular life of Earth origin.

Another scenario that could result in an encounter withgeneralized viruses is a situation where viruses may evolvein an end-life phase of a dying world. Imagine if Earth, abiologically rich planet harboring diverse viral communities,was to slowly undergo a loss of cellular life to a point ofextinction. It is not unimaginable that viral evolution in thissetting would favor generalized viruses as available hostnumbers decreased. Currently on Earth, viral specificity

outside pathogenic relationships (and even here one can ar-gue host benefits) is a means by which advantageous genescan be quickly shuttled to related cells giving the communityadvantages in competitive environments. From the perspec-tive of a competing cellular linage, sharing advantageousgenes with potential adversaries could be detrimental; thusviral host specificity is a valuable asset. While host specificityis advantageous in a biologically prime environment, it wouldnot be on a dying world. Due to a dwindling of availableresources, the ability of previously adversarial or competitivehosts to enter a state of altruism and share genetic information(i.e., ability to metabolize different nutrients or adapt to po-tentially harmful physical stressors such as temperaturechange) would become paramount. Altruism in the microbialworld under low-nutrient conditions has been demonstratedas an advantageous strategy (Blower et al., 2012). Viruses andtheir ability to facilitate rapid genetic exchange can be viewedas the architects of altruism on our planet. An altruistic stateon a dying planet may result in the last viable entity on theplanet being an extremely versatile virus. In this scenario,which may be similar to what occurred on Mars if cellular lifewas at one time abundant and subsequently significantly re-duced in number or lost, it is possible that a generalized virusevolved and is currently extant or preserved in ice.

What may present an obstacle to identifying extraterres-trial microbial life and an understanding of its potential risksto Earth is our perceptions of life and its origins. It has beenonly recently that the profuse nature of viruses or the rolethat they have played in driving the evolution of all organ-isms on our planet has been recognized. Historically, viruseshave been dismissed as important entities in regard to evo-lution and planetary exploration due in part to the fact thatmost of the first identified and described viruses were host-specific pathogens. What we know of viruses at this point isthat we are most likely to encounter them on other life-bearing planets and that, yes, they tend to be host-specific,but the existence of multi-host viruses is well known. Ad-ditionally, viruses are known to have adapted to extremeenvironments, and while most do not survive long outsidethe host cell, prolonged survival in cold-temperature settingshas been documented. We should be looking for viruses inour quest for extraterrestrial life, and it may be that virusespose no risk to human planetary exploration. However, thepossibility of risk exists, and our potential contact with themshould be treated accordingly.

Acknowledgments

I would like to thank Dr. Hon Ip of the USGS NationalWildlife Health Center located in Madison, Wisconsin, forreviewing the manuscript and offering some creative insight.

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Address correspondence to:Dale Warren Griffin

Environmental/Public Health MicrobiologistU.S. Geological Survey

600 4th Street SouthSt. Petersburg, FL 33701

E-mail: dgriffin@usgs.gov

Submitted 21 December 2012Accepted 20 April 2013

ASTROVIROLOGY 783