Appendixes



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The Quarantine and Certification of Martian Samples Appendixes

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The Quarantine and Certification of Martian Samples This page in the original is blank.

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The Quarantine and Certification of Martian Samples Appendix A Deinococcus radiodurans as an Analogue to Extremophile Organisms That May Have Survived on Mars Michael J. Daly Department of Pathology, Uniformed Services University of the Health Sciences, Bethesda, Maryland The only genetic systems within which the equivalent of millions of years’ worth of background radiation accumulated as genetic damage have been studied in a living organism are those developed by the terrestrial bacterium Deinococcus radiodurans.1-7 This bacterium is capable of repairing massive genetic damage without lethality or increasing mutation frequency. As such, D. radiodurans is an excellent organism in which to consider the potential for survival and biological evolution beyond its planet of origin, as well as the ability of life to survive extremely long periods of metabolic dormancy in high-radiation environments. Ultimately, the survival of any organism in such environments will be determined by its ability to repair, and recover from, accumulated genetic damage. D. radiodurans could likely survive extended periods of metabolic dormancy near the surface of Mars, periods of time in which the equivalent of Mrads of radiation damage would be accumulated in any genetic material. It is likely that during the first 500 million years (0.5 Ga) of its existence, Mars had a warmer and wetter climate than it has now, and an active hydrologic cycle. This may slightly predate the time at which Earth first began to support water and life; evidence of chemically evolved life on Earth dates to ~0.6 Ga after its formation. 1   Battista, J.R., Earl, A.M., and Park, M.-J. 1999. Why is Deinococcus radiodurans so resistant to ionizing radiation? Trends Microbiol. 7:362. 2   Minton, K.W. 1996. Repair of ionizing-radiation damage in the radiation resistant bacterium Deinococcus radiodurans. Mutat. Res. DNA Repair 362:1. 3   Mattimore, V., and Battista, J.R. 1996. Radioresistance of Deinococcus radiodurans: Functions necessary to survive ionizing radiation are also necessary to survive prolonged desiccation. J. Bacteriol. 177:5232. 4   Lange, C.C., Wackett, L.P., Minton, K.W., and Daly, M.J. 1998. Engineering a recombinant Deinococcus radiodurans for organopollutant degradation in radioactive mixed waste environments. Nat. Biotechnol. 16:929. 5   Brim, H., McFarlan, S.C., Fredrickson, J.K., Minton, K.W., Zhai, M., Wackett, L.P., and Daly, M.J. 2000. Engineering Deinococcus radiodurans for metal remediation in radioactive mixed waste environments. Nat. Biotechnol. 18:85-90. 6   Daly, M.J., Ouyang, L., Fuchs, P., and Minton, K.W. 1994. In vivo damage and recA-dependent repair of plasmid and chromosomal DNA in the radioresistant bacterium Deinococcus radiodurans. J. Bacteriol. 176:3508. 7   White, O., Eisen, J.A., Heidelberg, J.F., Hickey, E.K., Peterson, J.D., Dodson, R.J., Haft, D.H., Gwinn, M.L., Nelson, W.C., Richardson, D.L., Moffat, K.S., Qin, H., Jiang, L., Pamphile, W., Crosby, M., Shen, M., Vamathevan, J.J., Lam, P., McDonald, L., Utterback, T., Zalewski, C., Makarova, K.S., Aravind, L., Daly, M.J., Minton, K.W., Fleischmann, R.D., Ketchum, K.A., Nelson, K.E., Salzberg, S., Smith, H.O., Venter, J.C., and Fraser, Claire M. 1999. Genome sequence of the radioresistant bacterium Deinococcus radiodurans R1. Science 286:1571-1577.

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The Quarantine and Certification of Martian Samples Subsequent to this watery period martian life, if it existed, may have become subterranean and/or dormant, or it may have become extinct during the transition to a hostile environment caused by Mars’s loss of atmosphere and water. The time when conditions effectively became hostile to life are not known; the loss of atmosphere and water from Mars was gradual and is still occurring. It is not justified to assume a priori that all martian life, if it existed, has become extinct. If life existed on Mars, it likely evolved to survive the increasingly harsh conditions, and/or it found environments that are shielded from the planet’s present extreme climate. As our knowledge of Earth’s biosphere increases, extreme environments originally believed to be far too harsh to support life are regularly being found to support flourishing microbial communities. Organisms discovered in these extreme environments are called extremophiles, and collectively they demonstrate the remarkable ability of life to thrive over a wide range of extremely hostile environments, e.g., high temperature (these organisms are called thermophiles) or high pressure (barophiles). Similarly successful adaptations may have been made by microbes on Mars in environments at least equally hostile. D. radiodurans is an excellent example of an extremophile. It can grow continuously, without induced mutation or any effect on its growth rate, in the presence of 6,000 rad of radiation per hour.8,9 Further, it demonstrates astounding survival: it can withstand an acute dose of radiation of 1.5 Mrad (at 0 ºC) or 4.0 Mrad (at –70 ºC) without lethality. Even more extraordinary is the fact that following such massive irradiation, these cells can repair their genomes within 12 to 24 hours.10 Unpublished work by J. Battista has shown that desiccated D. radiodurans can survive exposure at higher doses, as much as 15 Mrad (Figure 5.3 in Chapter 5). These qualities are key to answering the question of whether life is extinct or extant on Mars, where the commonly accepted “post-transition period” from a life-supporting to a life-challenging biosphere might subject a dormant microbe to up to 2 billion years’ worth of accumulating background radiation, along with freezing and/or desiccation. The background cosmic radiation on Mars is similar to that on Earth, ~0.5 rad/year. Thus on the order of 1,000 Mrad of accumulated radiation dose could have been delivered to a microbe locked in a cryptobiotic state at the end of the transition period. As a single acute dose, this amount of radiation damage would overwhelm D. radiodurans and any other known organism. However, fluctuations in the Mars surface environment, and other events (such as burial by crater ejecta), could present the microbial population with periodic opportunities to become active, repair damage, and replenish their numbers while the radiation occurred. In the context of current technological and financial constraints, the question of where life on Mars is most likely to be metabolically active is distinctly separate from the question of where preserved life is most likely to be detected on Mars by missions to the planet in the near future. By analogy to the extremophiles on Earth, it is likely that metabolically active life on Mars would occur in geothermally heated subsurface water tables or spaces where it would be shielded from the harsh surface conditions. Such Mars subterranean environments will, however, be beyond human exploration for many years. In the foreseeable future, exploration of Mars will be limited to surface and near-surface environments. Although these accessible environments are exposed to harsh atmospheric and solar stress, such conditions do not preclude dormant life. It is possible that periodic melts of subsurface water caused by geothermal events and/or orbital variations bring subsurface microorganisms into surface and near-surface environments where they become frozen and cryopreserved in ground ice. If a martian microorganism had a proficient genetic repair system, it could survive frozen in such a site for millions or even hundreds of millions of years. If intermittently, though rarely, the local surface environments also underwent thawing for the same reasons, it would allow revival of organisms, repair of accumulated genetic damage, and repopulation of their environments. D. radiodurans provides a good 8   Lange, C.C., Wackett, L.P., Minton, K.W., and Daly, M.J. 1998. Engineering a recombinant Deinococcus radiodurans for organopollutant degradation in radioactive mixed waste environments. Nat. Biotechnol. 16:929. 9   Brim, H., McFarlan, S.C., Fredrickson, J.K., Minton, K.W., Zhai, M., Wackett, L.P., and Daly, M.J. 2000. Engineering Deinococcus radiodurans for metal remediation in radioactive mixed waste environments. Nat. Biotechnol. 18:85-90. 10   Daly, M.J., Ouyang, L., Fuchs, P., and Minton, K.W. 1994. In vivo damage and recA-dependent repair of plasmid and chromosomal DNA in the radioresistant bacterium Deinococcus radiodurans. J. Bacteriol. 176:3508.

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The Quarantine and Certification of Martian Samples model system in which to consider preservation and irradiation on the accessible surface and in near-surface environments of Mars. The following factors are known to dramatically affect an organism’s radiation resistance: Genome size. The smaller the genome, the more resistant it is. Ambient nutrient conditions. The presence of nutrients supports DNA repair. Temperature. Typically, the colder an organism is, the more resistant to radiation it is. Freezing increases radiation resistance. Hydration. Organisms are much more resistant to radiation when desiccated.