Planetary protection



Planetary protection is a guiding principle in the design of an interplanetary mission, aiming to prevent biological contamination of both the target celestial body and the Earth in the case of sample-return missions. Planetary protection reflects both the unknown nature of the space environment and the desire of the scientific community to preserve the pristine nature of celestial bodies until they can be studied in detail.

There are two types of interplanetary contamination. Forward contamination is the transfer of viable organisms from Earth to another celestial body. Back contamination is the transfer of extraterrestrial organisms, if such exist, back to the Earth's biosphere.

History
The potential problem of lunar and planetary contamination was first raised at the International Astronautical Federation VIIth Congress in Rome in 1956.

In 1958 the U.S. National Academy of Sciences (NAS) passed a resolution stating, “The National Academy of Sciences of the United States of America urges that scientists plan lunar and planetary studies with great care and deep concern so that initial operations do not compromise and make impossible forever after critical scientific experiments.” This led to creation of the ad hoc Committee on Contamination by Extraterrestrial Exploration (CETEX), which met for a year and recommended that interplanetary spacecraft be sterilized, and stated, “The need for sterilization is only temporary. Mars and possibly Venus need to remain uncontaminated only until study by manned ships becomes possible”.

In 1959, planetary protection was transferred to the newly formed Committee on Space Research (COSPAR). COSPAR in 1964 issued Resolution 26 affirming that:

In 1967, the US, USSR, and UK ratified the United Nations Outer Space Treaty. The legal basis for planetary protection lies in Article IX of this treaty:

This treaty has since been signed and ratified by 104 nation states. Another 24 have signed but not ratified. All the current space-faring nation states have both signed and ratified it. Amongst nations with space faring aspirations, some have not yet ratified: the United Arab Emirates, Syria and North Korea have signed but not yet ratified.

The Outer Space Treaty has consistent and widespread international support, and as a result of this, together with the fact that it is based on the 1963 declaration which was adopted by consensus in the UN National Assembly, it has taken on the status of customary international law. The provisions of the Outer Space Treaty are therefore binding on all states, even those who have neither signed nor ratified it. For forward contamination, the phrase to be interpreted is "harmful contamination". Two legal reviews came to differing interpretations of this clause (both reviews were unofficial). However the currently accepted interpretation is that “any contamination which would result in harm to a state’s experiments or programs is to be avoided”. NASA policy states explicitly that “the conduct of scientific investigations of possible extraterrestrial life forms, precursors, and remnants must not be jeopardized”.

COSPAR recommendations and categories
The Committee on Space Research (COSPAR) meets every two years, in a gathering of 2000 to 3000 scientists, and one of its tasks is to develop recommendations for avoiding interplanetary contamination. Its legal basis is Article IX of the Outer Space Treaty (see history below for details).

Its recommendations depend on the type of space mission and the celestial body explored. COSPAR categorizes the missions into 5 groups:
 * Category I: Any mission to locations not of direct interest for chemical evolution or the origin of life, such as the  Sun or Mercury. No planetary protection requirements.
 * Category II: Any mission to locations of significant interest for chemical evolution and the origin of life, but only a remote chance that spacecraft-borne contamination could compromise investigations. Examples include the Moon, Venus, and comets. Requires simple documentation only, primarily to outline intended or potential impact targets, and an end of mission report of any inadvertent impact site if such occurred.
 * Category III: Flyby and orbiter missions to locations of significant interest for chemical evolution or the origin of life, and with a significant chance that contamination could compromise investigations e.g., Mars, Europa, Enceladus. Requires more involved documentation than Category II. Other requirements, depending on the mission, may include trajectory biasing, clean room assembly, bioburden reduction, and if impact is a possibility, inventory of organics.
 * Category IV: Lander or probe missions to the same locations as Category III. Measures to be applied depend on the target body and the planned operations. "Sterilization of the entire spacecraft may be required for landers and rovers with life-detection experiments, and for those landing in or moving to a region where terrestrial microorganisms may survive and grow, or where indigenous life may be present. For other landers and rovers, the requirements would be for decontamination and partial sterilization of the landed hardware."
 * Missions to Mars in category IV are subclassified further:
 * Category IVa. Landers that do not search for Martian life - uses the Viking lander pre-sterilization requirements, a maximum of 300,000 spores per spacecraft and 300 spores per square meter.
 * Category IVb. Landers that search for Martian life. Adds stringent extra requirements to prevent contamination of samples.
 * Category IVc. Any component that accesses a Martian special region where terrestrial organisms are likely to propagate, or interpreted to have a high potential for existence of extamt Martian life forms. (see below) must be sterilized to at least to the Viking post-sterilization biological burden levels  of 30 spores total per spacecraft.


 * Category V: This is further divided into unrestricted and restricted sample return.
 * Unrestricted Category V: samples from locations judged by scientific opinion to have no indigenous lifeforms. No special requirements.
 * Restricted Category V: (where scientific opinion is unsure) the requirements include: absolute prohibition of destructive impact upon return, containment of all returned hardware which directly contacted the target body, and containment of any unsterilized sample returned to Earth.

For Category IV missions, a certain level of biological burden is allowed for the mission. In general this is expressed as a 'probability of contamination', required to be less than one chance in 10,000 of forward contamination per mission, but in the case of Mars Category IV missions (above) the requirement has been translated into a count of Bacillus spores per surface area, as an easy to use assay method.

More extensive documentation is also required for Category IV. Other procedures required, depending on the mission, may include trajectory biasing, the use of clean rooms during spacecraft assembly and testing, bioload reduction, partial sterilization of the hardware having direct contact with the target body, a bioshield for that hardware, and, in rare cases, complete sterilization of the entire spacecraft.

For restricted Category V missions, the current recommendation is that no uncontained samples should be returned unless sterilized. Since sterilization of the returned samples would destroy much of their science value, current proposals involve containment and quarantine procedures. For details, see Containment and quarantine below. Category V missions also have to fulfill the requirements of Category IV to protect the target body from forward contamination.

Category I

 * Io, Sun, Mercury, undifferentiated metamorphosed asteroids

Category II

 * Callisto, comets, asteroids of category P, D, and C, Venus, Kuiper belt objects (KBO) < 1/2 size of Pluto.

Provisional Category II
These are likely to be Category II but may be Category III.


 * Ganymede, Titan, Triton, the Pluto-Charon system, and other large KBOs (> 1/2 size of Pluto), Ceres,

For Pluto and Charon, they state that more research is needed, because there is a remote possibility that the tidal interactions could maintain some water reservoir below the surface.

Similar considerations to the ones for Pluto and Charon apply to other larger KBOs.

Triton is insufficiently well understood at present to say it is definitely devoid of liquid water. The only close up observations to date are those of Voyager 2.

In a detailed discussion of Titan, scientists concluded that there was no danger of contamination of its surface, except short term adding of negligible amounts of organics.

However, Titan could have a below surface water reservoir that communicates with the surface, and if so, this could be at risk of contamination.

In the case of Ganymede, it has strong evidence for a subsurface ocean. The question is, given that its surface shows pervasive signs of resurfacing, is there any communication with its subsurface ocean?

They found no known mechanism by which this could happen, and the Galileo spacecraft found no evidence of cryovolcanism. Initially, they assigned it as Priority B minus, meaning that precursor missions are needed to assess its category before any surface missions. However, after further discussion they provisionally assigned it to Category II. This means that no precursor missions are required, though this could still be revised again, depending on future research.

If there is cryovolcanism on Ganymede or Titan, the undersurface reservoir is thought to be 50 – 150 km below the surface. They were unable to find a process that could transfer the surface melted water back down through 50 km of ice to the under surface sea.

This is why both Ganymede and Titan were assigned a reasonably firm provisional Category II, but pending results of future research.

Icy bodies that show signs of recent resurfacing need further discussion and might need to be assigned to a new category depending on future research. This approach has been applied, for instance, to missions to Ceres. The planetary protection Category is subject for review during the mission of the Ceres orbiter ('Dawn') depending on the results found.

Category III / IV

 * Mars because of possible surface habitats.
 * Europa because of its subsurface ocean.
 * Enceladus because of evidence of water plumes.

Category V
In the category V for sample return the conclusions so far are:


 * Restricted Category V: Mars, Europa, Enceladus.
 * Unrestricted Category V: Venus, the Moon.

with others to be decided.

Other objects
If there has been no activity for 3 billion years, it will not be possible to destroy the surface by terrestrial contamination, so can be treated as Category I. Otherwise, the category may need to be reassessed.

Mars special regions
A special region on Mars is a region classified by COSPAR where terrestrial organisms are likely to propagate, or interpreted to have a high potential for existence of extamt Martian life forms. . Based on current understanding, this includes any region with a high enough temperatures for Earth organisms to propagate (above -18°C), and with water in a form accessible to them (water activity higher than 0.6), both requirements to be satisfied simultaneously. Adding a margin, a special region is defined as one with water activity higher than 0.5 and temperature above -23°C

Other environmental factors such as the chemical makeup (such as perchlorates), the ionizing radiation, the UV radiation, and low atmospheric pressure are not used to restrict special regions, as there are Earth microbes that can tolerate them,. Also the presence of multiple factors simultaneously is not used to restrict special regions either because of the existence of polyextremophiles.

In principle native Martian life could have additional capabilities, and so, be able to propagate at lower temperatures or with lower water activity. However, since these capabilities are unknown they are not used to determine special regions.

There are currently no confirmed special regions. However there are many uncertain regions such as the recurring slope lineae. These are treated as special regions for the purposes of planetary protection, until more is known.

Catherine Conley, NASA's Planetary protection officer at the time, explains why the spacecraft we send to Mars are sterilized, 33 seconds into this video

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 * Third video on overview page of the NASA Office of Planetary Protection.

"“So we have to do all of our search for life activities, we have to look for the Mars organisms, without the background, without the noise of having released Earth organisms into the Mars environment”"

Limits of water activity 0.6 and temperature -18°C
The term "Special region" is understood to apply to any region on Mars where liquid and not too salty water can occur in a temperature range suitable for terrestrial life. The water activity for terrestrial life has to be above 0.6. As an example, honey in dry air has a water activity level of 0.6, and no terrestrial microbes can grow in honey.

The temperature also has to be above -18°C and both of these have to be satisfied simultaneously (see section 7.1.1, page 940 of ). Water that is too salty but warm enough or too cold with enough water activity does not count as a special region.

Adding a margin, a special region is defined as one with water activity higher than 0.5 and temperature above -23°C

The report remarks that the water activity limit of 0.6 is well determined with no terrestrial life known that can reproduce below that, though it is no.

However the lower temperature limit of Earth life is not so well understood, because of practical difficulties measuring extremely low rates of metabolism and cell division. However they were asked to check only for reproduction of Earth life on Mars 500 years into the future. They give the example of cryptoendolithic microbial communities in the Antarctic Dry Valleys which successfully invade sandstone over time periods of 1,000 to 10,000 years. The -18°C limit they give is sufficient to protect Mars from Earth life over a 500 year future timeframe (section 3.1, page 894 of ).

The Mars surface is abundant in chaotropic agents that disrupt hydrogen bonding, such as  MgCl2,CaCl2, FeCl3, FeCl2, FeCl, LiCl, and perchlorate salts. These can disrupt biological processes at higher temperatures but for numerous species they depress the lowest temperature for microbes to propagate at temperatures below 10°C, by up to 10°C to 20°C. They found no research into whether this effect also occurs at -18°C or below. They note the possibility that this could depress the lower temperature limit, but experiments haven't been conducted on long enough timescales, with a potential doubling time of between several months and years

Perchlorates, ionizing radiation, UV, low pressure and multiple stressors not used to limit potential special regions
COSPAR considered many other environmental factors on Mars in addition to the temperature range and water activity. None of these were used to limit potential special regions.


 * The chemical environment of the Mars surface, including the perchlorates, are considered favourable for life, in particular, perchlorates can be an oxidant for hydrogen and carbon monoxide oxidizing organisms
 * The low pressures of the Mars atmosphere can be tolerated by some terrestrial bacteria
 * The UV flux is blocked by less than 1 mm of regolith or other organisms
 * From the MSL RAD measurements, ionizing radiation levels from cosmic radiation are so low as to be negligible. The intermittent solar storms increase the dose only for a few days and the Martian surface provides enough shielding so that the total dose from solar storms is less than double that from cosmic radiation/ Over 500 years the Mars surface would receive a cummulative dose of less than 50 Gy, far less than the dose where 90% of even a radiation senstiive bacterium such as e-coli would die (LD90 of ~200 - 400 Gy). These  facts  are  not  used  to  distinguish  Special Regions on Mars.
 * On Mars multiple stressors are present simultaneously, but polyextromphiles can often cope with multiple simultaneous stresses either using the same mechanism for them all or multiple mechanisms

Extant life with additional capabilities are not used to deliminate uncertain or special regions
The report briefly discusses the phrase:

"‘‘any region which is interpreted to have a high potential for the existence of extant martian life forms  is  also  defined  as  a  Special  Region’’"

There has been speculation that Martian life might be able to tolerate conditions that are outside of the range of terrestrial life, for instance, able to reproduce at much lower temperatures, and perhaps also in conditions with less water activity than is possible for terrestrial life.

The report doesn't give examples, but one such is Dirk Schulze Makuch's proposal that life on Mars may be using a mixture of water and hydrogen peroxide as its solvent. A 61.2% (by weight) mix of water and hydrogen peroxide has a freezing point of −56.5 °C, and also tends to super-cool rather than crystallize. It is also hygroscopic, an advantage in a water-scarce environment.

However the capabilities of any Martian life are currently unknown. The review committees on the Martian special regions have decided that since we lack any data on their capabilities, this requirement can't be used to determine any additional special regions. So they use the capabilities of Earth life exclusively

No known special regions
So far there are no known special regions. Most of them would require verification of existence of microhabitats that are impossible to see directly from orbit. However, there are many uncertain regions where it is possible that as we advance our knowledge of Mars, some of them turn out to be special regions. Amongst the top candidates are the Warm Seasonal flows on Mars (Recurrent Slope Lineae).

Another top candidate is based on the droplet like features that formed on the landing legs of the Phoenix lander. It landed in what is thought to be an ancient ocean bed near the north pole, the first and so far the only spacecraft to land successfully in polar regions. In December 2013, Nilton Renno and his team using the Michigan Mars Environmental Chamber were able to simulate the conditions at its landing site and the droplets. They formed salty brines within minutes when salt overlaid ice, with the salt, especially perchlorates, acting as an "antifreeze". The team concluded that suitable conditions for brine droplets may be widespread in the polar regions. Nilton Renno talks about their results in this video

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For more on these and other candidates, see Possible present day habitats for life on Mars.

Special regions caused by spacecraft in nominal landings or impact scenarios
The requirements also apply to regions that may be made into a special region by a spacecraft, e.g. an impact melting ice, or the sources of thermal energy foreign to Mars in the spacecraft itself. If a hard landing risks biological contamination of a special region, it has to sterilized to COSPAR category IVc, which is also required for any mission that may contact such a region.

Missions need to study their potential to create Spacecraft-Induced special regions during the planning phase and take action to make sure they are not inadvertently created, and spacecraft also need to avoid special regions if they are not clean enough to avoid contaminating them. The risk of spacecraft induced special regions needs to be evaluated separately for each mission, taking account of the spacecraft and the landing ellipse.

Special regions could be caused during a nominal landing through the warming effects of rocket exhausts, and temporary special regions could also be created when the hot aeroshell, heatshield, backshell or skycrane compnents land on icy ground in a nominal landing. The spacecraft is normally well insulated, so thermal sources within do not leak much radiation. However, special regions can also be caused in normal use of the lander or rover, by vibration or drilling. Examples include the vibrating sieve on the Viking sampler, the Rock Abrasion Tool (RAT) on the Mars Exploration Rover, the drills on Curiosity or ExoMars, and wheel slip or scuff if the rover gets temporarily "stuck". The lifetime of the special region would depend on the volume melted and whether or not it is heated enough for boiling to be initiated, which can happen at low temperatures with the low Martian atmospheric pressures (over much of the surface, pure water boils at 0°C). Once boiling starts, it may self deplete rapidly and so self-destruct.

The analysis must also look at the outcomes of an off nominal "Breakup and Burnup" scenario. Ice could be heated up by direct transfer from a potentially hot structure from atmosphereic heating as well as the kinetic energy of the impact converted to heat. Any radioisotope heating unit and the RTG's would generate heat energy for several decades. If it lands within a dry capping layer of regolith then it's likely to be separated from the wet layer it causes, but if fully buried in ice or icy soil it could cause pooling and persistence of water near to the RTG The report looked at distributions of ice and concluded that ice in the tropics is buried too deep to be a consideration

However the 2014/5 review corrected this due to evidence of ice present at depths of <1 meter in pole-facing slopes

The report also considered the droplets that formed on the Phoenix legs as well as a reported stickiness of its excavated soil samples that was reduced on exposure to the atmosphere, both of which suggested deliquescence, and concluded that these also need to be taken into consideration

The 2014 report provided a map of regions of Mars where there may be ice below the surface as well as potential RSL's. The 2015 review however said that such maps are most useful if accompanied by cautionary remarks on their limitations, as they are subject to change with new discoveries. See below.

Revisions of the definition of special regions in 2014/5 to include more potential surface habitats
The definition of a special region has been revised several times. In the 2006 study it was implicit that a special region must be defined by warm enough temperatures combined with sufficient water activity. If the Mars surface is mapped using those requirements alone and requiring them to overlap, the map would be blank. (see page 941 of )

""Were their intersection to have been mapped out on the martian surface, the 2006 study’s expectation for warm temperatures and high-enough water activity would have been a blank  map, as  the  posited  martian subsurface equilibrium conditions did not allow warm-enough temperatures and sufficient water activity to coexist. As such, no natural Special Region would exist outside of (possibly) gully systems or other non-understood features, or in the deep subsurface

.... By specifying a water activity value, one was automatically faced with a temperature that was too low to allow terrestrial organisms to replicate, and if the temperature was high enough for that, the relative humidity at that site would be excruciatingly low, allowing no replication on its part.""

The only place where habitats could occur in the 2006 report were deep below the surface, or perhaps places like the gully systems where water could be exposed to the surface dynamically from the subsurface.

However the 2014 study finds that though both are not present simultaneously, often they are present on the surface within a 24 hour period of each other due to the extreme day - night cycles on Mars. That then makes it possible that terrestrial biology could bridge the gap (e.g. retain the water through to warmer temperatures in the same day). Referring to the Viking and Curiosity (MSL) landing sites they say (page 941 of )

"... where in some  seasons  the  temperature  required for microbial replication was regularly reached during the driest part of the day, whereas at night, when the temperature was too low for replication, the relative humidity at the site was above 0.6 and nearly always close to 1.0. The non-overlap of the required values for a Special Region is reflected in Finding 4-12, but the fact that both could be reached within the same 24 h period, regularly, suggests that there may be a way for organisms to connect the favorable aspects of those periods across a bridge of biotic adaptation."

They go on to say that they didn't have any evidence yet that terrestrial organisms could bridge that gap, and they had some evidence that suggested it might be unbridgeable. However where the atmospheric pressure is above the triple point of water, precipitation that reaches the ground could melt and provide a temporary habitat. Also some materials such as clays, and the organisms themselves could retain more water than the soil itself.

Before the 2014 report was published, both NASA and ESA took steps to have it reviewed independently. This 2015 review concurred with 29 findings of the original report. They did not support one of them and proposed modifications to 15 others. The 2015 review comittee in their report said that they believe that some important aspects were not covered in the previous report. For instance:


 * The possibility of terrestrial contamination translocation (blown in the Martian dust)
 * The ability of multi-species microbial communities to alter dispersed small-scale habitats to make them more habitable. They could synthesize substances and adjust environmental parameters to make them more habitable.

This makes the definition of a special region more problematic. They give an example in chapter 2:

"There are many examples of small-scale and microscale environments on Earth that can host microbial communities, including biofilms, which may only be a few cell layers thick. The biofilm mode of growth, as noted previously, can provide affordable conditions for microbial propagation despite adverse and extreme conditions in the surroundings. On Earth, the heterogeneity of microbial colonization in extreme environments has become more obvious in recent years. To identify Special Regions across the full range of spatial scales relevant to microorganisms, a better understanding of the temperature and water activity of potential microenvironments on Mars is necessary. For instance, the interior of the crater Lyot in the northern mid-latitude has been described as an optimal microenvironment with pressure and temperature conditions that could lead to the formation of liquid water solutions during periods of high obliquity (Dickson and Head 2009). Craters, and even microenvironments underneath and on the underside of rocks, could potentially provide favorable conditions for the establishment of life on Mars, potentially leading to the recognition of Special Regions where landscape-scale temperature and humidity conditions would not enable it."

On dust transport and other forms of translocation they just say that the research so far is not sufficient to answer the question:

"A potential problem with designating Special Regions on Mars is that viable microorganisms that survive the trip to Mars could be transported into a distant Special Region by atmospheric processes, landslides, avalanches (although this risk is considered minimal), meteorite impact ejecta, and lander impact ejecta. In addition to dilution effects, the flux of ultraviolet radiation within the martian atmosphere would be deleterious to most airborne microbes and spores. However, dust could attenuate this radiation and enhance microbial viability. In addition, for microbes growing not as single cells but as tetrades or larger cell chains, clusters, or aggregates, the inner cells are protected against ultraviolet radiation. Examples are methanogenic archaea like Methanosarcina, halophilic archaea like Halococcus, or cyanobacteria like Gloeocapsa. This is certainly something that could be studied and confirmed or rejected in terrestrial Mars simulation chambers where such transport processes for microbes (e.g., by dust storms) are investigated. The SR-SAG2 report does not adequately discuss the transport of material in the martian atmosphere."

Caution on use of maps
The 2014/5 revisions caution against the use of maps in chapter 5

"In general, the review committee contends that the use of maps to delineate regions with a lower or higher probability to host Special Regions is most useful if the maps are accompanied by cautionary remarks on their limitations. Maps that illustrate the distribution of specific relevant landforms or other surface features can only represent the current (and incomplete) state of knowledge for a specific time—knowledge that will certainly be subject to change or be updated as new information is obtained."

Planetary categorizations of solar system objects
Some targets are easily categorized. Others are assigned provisional categories by COSPAR, pending future discoveries and research.

The 2009 COSPAR Workshop on Planetary Protection for Outer Planet Satellites and Small Solar System Bodies covered this in some detail. Most of these assessments are from that report, with some later refinements. This workshop also gave more precise definitions for some of the categories:

The Coleman-Sagan equation
The aim of the current regulations is to keep the number of microorganisms low enough so that the probability of contamination of Mars (and other targets) is acceptable. It is not an objective to make the probability of contamination zero.

The aim is to keep the probability of contamination of 1 chance in 10,000 of contamination per mission flown. This figure is obtained typically by multiplying together the number of microorganisms on the spacecraft, the probability of growth on the target body, and a series of bioload reduction factors.

In detail the method used is the Coleman-Sagan equation.

$$P_c = N_0 R P_S P_t P_R P_g$$.

where
 * $$N_0$$ = the number of microorganisms on the spacecraft initially
 * $$R$$ = Reduction due to conditions on spacecraft before and after launch
 * $$P_S$$ = Probability that microorganisms on the spacecraft reach the surface of the planet
 * $$P_t$$ = Probability that spacecraft will hit the planet - this is 1 for a lander
 * $$P_R$$ = Probability of microorganism to be released in the environment when on the ground, usually set to 1 for crashlanding.
 * $$P_g$$ = Probability of growth. For targets with liquid water this is set to 1 for sake of the calculation.

Then the requirement is $$P_c < 10^{-4}$$

The $$10^{-4}$$ is a number chosen by Sagan et al., somewhat arbitrarily. Sagan and Coleman assumed that about 60 missions to the Mars surface would occur before the exobiology of Mars is thoroughly understood, 54 of those successful, and 30 flybys or orbiters, and the number was chosen to endure a probability to keep the planet free from contamination of at least 99.9% over the duration of the exploration period.

Critiques
The Coleman Sagan equation has been criticised because the individual parameters are often not known to better than a magnitude or so. For example, the thickness of the surface ice of Europa is unknown, and may be thin in places, which can give rise to a high level of uncertainty in the equation. It has also been criticised because of the inherent assumption made of an end to the protection period and future human exploration. In the case of Europa, this would only protect it with reasonable probability for the duration of the period of exploration.

Greenberg has suggested an alternative, to use the natural contamination standard - that our missions to Europa should not have a higher chance of contaminating it than the chance of contamination by meteorites from Earth.

Another approach for Europa is the use of binary decision trees which is favoured by the Committee on Planetary Protection Standards for Icy Bodies in the Outer Solar System under the auspices of the Space Studies Board. This goes through a series of seven steps, leading to a final decision on whether to go ahead with the mission or not.

Containment and quarantine for restricted Category V sample return
In the case of restricted Category V missions, Earth is protected through quarantine of sample and astronauts, and sample containment.

In the case of a Mars sample return, missions would be designed so that no part of the capsule that encounters the Mars surface is exposed to the Earth environment.

One way to do that is to enclose the sample container within a larger outer container from Earth, in the vacuum of space. The integrity of any seals is essential and the system must also be monitored to check for the possibility of micro-meteorite damage during return to Earth.

The recommendation of the ESF report is that

"“No uncontained Mars materials, including space craft surfaces that have been exposed to the Mars environment should be returned to Earth unless sterilised" ..."For unsterilised samples returned to Earth, a programme of life detection and biohazard testing, or a proven sterilisation process, shall be undertaken as an absolute precondition for the controlled distribution of any portion of the sample.”"

No restricted category V returns have been carried out in recent times.

During the Apollo program the sample returns were regulated through the Extra-Terrestrial Exposure Law. This was rescinded in 1991, so new legislation would need to be enacted. The Apollo era quarantine procedures are of interest as the only attempt to date of a return to Earth of a sample that, at the time, was thought to have a remote possibility of including extraterrestrial life.

Samples and astronauts were quarantined in the Lunar Receiving Laboratory. The methods used would be considered inadequate for containment by modern standards. Also the lunar receiving laboratory would be judged a failure by its own design criteria as the sample return didn't contain the lunar material, with two failure points during the Apollo 11 return mission, at the splashdown and at the facility itself.

However the Lunar Receiving Laboratory was built quickly with only two years from start to finish, a time period now considered inadequate. Lessons learned from it can help with design of any Mars sample return receiving facility.

Design criteria for a proposed Mars Sample Return Facility, and for the return mission, have been developed by the American National Research Council, and the European Space Foundation. They concluded that it could be based on biohazard 4 containment but with more stringent requirements to contain unknown microorganisms possibly as small as or smaller than the smallest Earth microorganisms known, the ultramicrobacteria. The ESF study also recommended that it should be designed to contain the smaller gene transfer agents if possible, as these could potentially transfer DNA from martian microorganisms to terrestrial microorganisms if they have a shared evolutionary ancestry. It also needs to double as a clean room facility to protect the samples from terrestrial contamination that could confuse the sensitive life detection tests that would be used on the samples.

Before a sample return, new quarantine laws would be required. Environmental assessment would also be required, and various other domestic and international laws not present during the Apollo era would need to be negotiated.

Decontamination procedures
For all spacecraft missions requiring decontamination, the starting point is clean room assembly in US federal standard class 100 cleanrooms. These are rooms with fewer than 100 particles of size 0.5 µm or larger per cubic foot. Engineers wear cleanroom suits with only their eyes exposed. Components are sterilized individually before assembly, as far as possible, and they clean surfaces frequently with alcohol wipes during assembly. Spores of Bacillus subtilis was chosen for not only its ability to readily generate spores, but its well-established use as a model species. It is a useful tracker of UV irradiation effects because of its high resilience to a variety of extreme conditions. As such it is an important indicator species for forward contamination in the context of planetary protection.

For Category IVa missions (Mars landers that do not search for Martian life), the aim is to reduce the bioburden to 300,000 bacterial spores on any surface from which the spores could get into the Martian environment. Any heat tolerant components are heat sterilized to 114 °C. Sensitive electronics such as the core box of the rover including the computer, are sealed and vented through high-efficiency filters to keep any microbes inside.

For more sensitive missions such as Category IVc (to Mars special regions), a far higher level of sterilization is required. These need to be similar to levels implemented on the Viking landers, which were sterilized for a surface which, at the time, was thought to be potentially hospitable to life similar to special regions on Mars today.

In microbiology, it is usually impossible to prove that there are no microorganisms left viable, since many microorganisms are either not yet studied, or not cultivable. Instead, sterilization is done using a series of tenfold reductions of the numbers of microorganisms present. After a sufficient number of tenfold reductions, the chance that there any microorganisms left will be extremely low.

Dry heat sterilization
The two Viking Mars landers were sterilized using dry heat sterilization. After preliminary cleaning to reduce the bioburden to levels similar to present day Category IVa spacecraft, the Viking spacecraft were heat-treated for 30 hours at 112 °C, nominal 125 °C (five hours at 112 °C was considered enough to reduce the population tenfold even for enclosed parts of the spacecraft, so this was enough for a million-fold reduction of the originally low population).

Revised figures as used in 2017 extend the range up to 200 C using various formulae, see table 1 of the 2017 survey article by Shirey et al, "An Overview of Surface Heat Microbial Reduction as a Viable Microbial Reduction Modality for Spacecraft Surfaces." . For encapsulated surfaces the formula for 2-3 decimal reductions are:


 * Range 116 to 125 °C, $$5*0.5* 10^{((125 –T) / 15)}$$ hours. Figures are not given for temperatures below 116 °C, but if the formla is extrapolated down to the Viking temperatures, that makes it 18 hours for a 2-3 decimal reductions and just short of ten hours at 116 °C
 * Range 125 to 140 °C, $$5*0.5* 10^{((125 –T) / 21)}$$ hours. Example, at 140 °C it's = 0.48 hours
 * Range 140 to 200 °C, $$5*0.5* 10^{((140 –T) / (23*T/140))}$$ hours. Example, at 200 °C it's = 0.037 hours or about 2 minutes 14 seconds.

A million fold reduction is achieved with 350 °C for  1 hr.  and  500  °C  for  0.5  secs

Challenge of modern materials
Modern materials however are often not designed to handle such temperatures, especially since modern spacecraft often use "commercial off the shelf" components. Problems encountered include nanoscale features only a few atoms thick, plastic packaging, and conductive epoxy attachment methods. Also many instrument sensors cannot be exposed to high temperature, and high temperature can interfere with critical alignments of instruments.

New methods
As a result, new methods are needed to sterilize a modern spacecraft to the higher categories such as Category IVc for Mars, similar to Viking. Methods under evaluation, or already approved, include :


 * Vapour phase hydrogen peroxide - effective, but can affect finishes, lubricants and materials that use aromatic rings and sulfur bonds. This has been established, reviewed, and a NASA/ESA specification for use of VHP has been approved by the Planetary Protection Officer, but it has not yet been formally published.
 * Ethylene oxide - this is widely used in the medical industry, and can be used for materials not compatible with hydrogen peroxide. It is under consideration for missions such as ExoMars.
 * Gamma radiation and electron beams have been suggested as a method of sterilization, as they are used extensively in the medical industry. They need to be tested for compatibility with spacecraft materials and hardware geometries, and are not yet ready for review.
 * Supercritical carbon dioxide snow (Mars) - is most effective against traces of organic compounds rather than whole microorganisms. Has the advantage though that it eliminates the organic traces - while other methods kill the microorganisms, they leave organic traces that can confuse life detection instruments. Is under study by JPL and ESA.

Some other methods are of interest as they can sterilize the spacecraft after arrival on the planet.
 * Supercritical carbon dioxide snow (Mars).
 * Passive sterilization through UV radiation (Mars). Highly effective, most microbes are killed in minutes, some that are particularly resistant to UV radiation survive for hours. However this does not sterilize microbes in shadows or cracks.
 * Passive sterilization through particle fluxes (Europa). Plans for missions to Europa take credit for reductions due to this.

Heat sterilization at higher temperatures with modern components, 200 - 300°C, or 500°C
In the NASA Venus Technology Plan (2010), they explore ideas for a rover to operate at Venus surface temperatures for long periods of time. One option is to cool it to 200 - 300°C. Another is to design it to run at 500°C with no cooling. For the first option, they say (page 16)

"1) Medium-Temperature Semiconductor-Based Electronics: Medium-temperature (200–300°C) electronics are not only technically less difficult than electronics that operate at Venus surface temperatures but also have terrestrial commercial applications. A broad set of component options, including microprocessor and memory devices exist."

Sauders et al have done work on an "automation rover" that could function at up to 500 C using a mixture of mechanical components, as in the early mechanical ballistic computers, and electronics, and have suggested using this for sterile rovers. .

"An automaton rover  could  be  subjected  to  much  higher  contamination  control  procedures  than traditional rovers. It could be baked at extremely high temperatures, irradiated, and subjected to multiple  chemical  baths  to  kill  any  bacteria.  Thus,  automatons  would  be  valuable  in  highly contamination control sensitive environments, like collecting samples from the dark, water streaks on  Mars.  In  this  type  of  situation,  the  automaton  would  likely  be  working  in  tandem  with  a traditional Mars rover."

Brian Wilcox is working on a 100% sterile probe to descend into the Europan ocean. . It would be deployed from a less sterile lander and it would be sterilized by keeping it at a temperature of 500°C throughout the journey out to Europa. In his abstract he says

"A central thesis of this work is that we must start by addressing the Planetary Protection constraints, and not to try to add them on at the end. Specifically, all hardware in the probe would be designed to survive heat sterilization at 500 °C for extended periods, as required to meet the COSPAR 1-in-10,000 probability per mission of biological contamination of the ocean"

The NASA press release says

"To ensure no Earth microbes hitched a ride, the probe would heat itself to over 900 degrees Fahrenheit (482 degrees Celsius) during its cruise on a spacecraft. That would kill any residual organisms and decompose complex organic molecules that could affect science results."

Bioburden detection and assessment
The spore count is used as an indirect measure of the number of microorganisms present. Typically 99% of microorganisms by species will be non-spore forming and able to survive in dormant states, and so the actual number of viable dormant microorganisms remaining on the sterilized spacecraft is expected to be many times the number of spore-forming microorganisms.

One new spore method approved is the "Rapid Spore Assay". This is based on commercial rapid assay systems, detects spores directly and not just viable microorganisms and gives results in 5 hours instead of 72 hours.

Challenges
It is also long been recognized that spacecraft cleaning rooms harbour polyextremophiles as the only microbes able to survive in them. For example, in a recent study, microbes from swabs of the Curiosity rover were subjected to desiccation, UV exposure, cold and pH extremes. Nearly 11% of the 377 strains survived more than one of these severe conditions.

This does not mean that these microbes have contaminated Mars. This is just the first stage of the process of bioburden reduction. To contaminate Mars they also have to survive the low temperature, vacuum, UV and ionizing radiation during the months long journey to Mars, and then have to encounter a habitat on Mars and start reproducing there. Whether this has happened or not is a matter of probability. The aim of planetary protection is to make this probability as low as possible. The currently accepted target probability of contamination per mission is to reduce it to less than 0.01%, though in the special case of Mars, scientists also rely on the hostile conditions on Mars to take the place of the final stage of heat treatment decimal reduction used for Viking. But with current technology scientists cannot reduce probabilities to zero.

New methods
Two recent molecular methods have been approved for assessment of microbial contamination on spacecraft surfaces.


 * Adenosine triphosphate (ATP) detection - this is a key element in cellular metabolism. This method is able to detect non cultivable organisms. It can also be triggered by non viable biological material so can give a "false positive".
 * Limulus Amebocyte Lysate assay - detects lipopolysaccharides (LPS). This compound is only present in Gram-negative bacteria. The standard assay analyses spores from microbes that are primarily Gram-positive, making it difficult to relate the two methods.

Impact prevention
This particularly applies to orbital missions, Category III, as they are sterilized to a lower standard than missions to the surface. It is also relevant to landers, as an impact gives more opportunity for forward contamination, and impact could be on an unplanned target, such as a special region on Mars.

The requirement for an orbital mission is that it needs to remain in orbit for at least 20 years after arrival at Mars with probability of at least 99% and for 50 years with probability at least 95%. This requirement can be dropped if the mission is sterilized to Viking sterilization standard.

In the Viking era (1970s), the requirement was given as a single figure, that any orbital mission should have a probability of less than 0.003% probability of impact during the current exploratory phase of exploration of Mars.

For both landers and orbiters, the technique of trajectory biasing is used during approach to the target. The spacecraft trajectory is designed so that if communications are lost, it will miss the target.

Issues with impact prevention
Despite these measures there has been one notable failure of impact prevention. The Mars Climate Orbiter which was sterilized only to Category III, crashed on Mars in 1999 due to a mix-up of imperial and metric units. The office of planetary protection stated that it is likely that it burnt up in the atmosphere, but if it survived to the ground, then it could cause forward contamination.

Mars Observer is another Category III mission with potential planetary contamination. Communications were lost three days before its orbital insertion maneuver in 1993. It seems most likely it did not succeed in entering into orbit around Mars and simply continued past on a heliocentric orbit. If it did succeed in following its automatic programming, and attempted the manoeuvre, however, there is a chance it crashed on Mars.

Three landers have had hard landings on Mars. These are Schiaparelli EDM lander, the Mars Polar Lander, and Deep Space 2. These were all sterilized for surface missions but not for special regions (Viking pre-sterilization only). Mars Polar Lander, and Deep Space 2 crashed into the polar regions which are now treated as special regions because of the possibility of forming liquid brines.

Meteorite argument
Alberto G. Fairén and Dirk Schulze-Makuch published an article in Nature recommending that planetary protection measures need to be scaled down. They gave as their main reason for this, that exchange of meteorites between Earth and Mars means that any life on Earth that could survive on Mars has already got there and vice versa.

Robert Zubrin used similar arguments in favour of his view that the back contamination risk has no scientific validity.

Rebuttal by NRC
The meteorite argument was examined by the NRC in the context of back contamination. It is thought that all the Martian meteorites originate in relatively few impacts every few million years on Mars. The impactors would be kilometers in diameter and the craters they form on Mars tens of kilometers in diameter. Models of impacts on Mars are consistent with these findings.

Earth receives a steady stream of meteorites from Mars, but they come from relatively few original impactors, and transfer was more likely in the early Solar System. Also some life forms viable on both Mars and on Earth might be unable to survive transfer on a meteorite, and there is so far no direct evidence of any transfer of life from Mars to Earth in this way.

The NRC concluded that though transfer is possible, the evidence from meteorite exchange does not eliminate the need for back contamination protection methods.

Impacts on Earth able to send microorganisms to Mars are also infrequent. Impactors of 10 km across or larger can send debris to Mars through the Earth's atmosphere but these occur rarely, and were more common in the early Solar System.

Proposal to end planetary protection for Mars
In their 2013 paper "The Over Protection of Mars", Alberto Fairén and Dirk Schulze-Makuch suggested that space agencies no longer need to protect Mars, essentially using Zubrin's meteorite transfer argument. This was rebutted in a follow up article "Appropriate Protection of Mars", in Nature by the current and previous planetary protection officers Catherine Conley and John Rummel. . The debate continues in articles in Astrobiology journal in 2017, 2018  and 2019

Critique of Category V containment measures
The scientific consensus is that the potential for large-scale effects, either through pathogenesis or ecological disruption, is extremely small. Nevertheless, returned samples from Mars will be treated as potentially biohazardous until scientists can determine that the returned samples are safe. The goal is to reduce the probability of release of a Mars particle to less than one in a million.

The International Committee Against Mars Sample Return agree with the assessment of low probability of large-scale effects, but consider the proposed containment measures insufficient, given the possible severity of the worst-case scenario. They come to this conclusion partly as a result of considerations of human error and the novelty of the mission proposal. Consequently, they advocate much more in situ research before undertaking a Mars Sample Return (MSR).

Proposal for extension of protection to non-biological considerations
A COSPAR workshop in 2010, looked issues to do with protecting areas from non biological contamination. They recommended that COSPAR expand its remit to include such issues.

Recommendations of the workshop include:

Ideas for doing this suggested since the workshop include protected special regions, or "Planetary Parks" to keep regions of the Solar System pristine for future scientific investigation, and also for ethical reasons.

Proposal to extend time scale for contamination prevention
This is about whether the aim is to make sure that any contamination is contained (for instance on or in the spacecraft) for some fixed future time frame like a century, or whether exploration should be done in such a way that contamination is contained indefinitely.

Astrobiologist Christopher McKay has argued that until we have better understanding of Mars, our explorations should be biologically reversible. For instance if all the microorganisms introduced to Mars so far remain dormant within the spacecraft, they could in principle be removed in the future, leaving Mars completely free of contamination from modern Earth lifeforms.

In the 2010 workshop one of the recommendations for future consideration was to extend the period for contamination prevention to the maximum viable lifetime of dormant microorganisms introduced to the planet.

In the case of Europa, a similar idea has been suggested, that it is not enough to keep it free from contamination during our current exploration period. It might be that Europa is of sufficient scientific interest that the human race has a duty to keep it pristine for future generations to study as well. This was the majority view of the 2000 task force examining Europa, though there was a minority view of the same task force that such strong protection measures are not required.

Protecting objects beyond the Solar System
The proposal by the German physicist Claudius Gros, that the technology of the Breakthrough Starshot project may be utilized to establish a biosphere of unicellular organisms on otherwise only transiently habitable exoplanets, has sparked a discussion, to what extent planetary protection should be extended to exoplanets.

Claudius Gros he also considers the possiblity of young oxygen rich planets. M-dwarf planets may have oxygen levels, from the dissociation of water, of many bars. This might sterilize the planet of any emerging primitive life, but be suitable for oxygen using Earth life. If so, he argues that it might be ethically acceptable to seed it with microbes that initiate a new biosphere there in probes that would get there thousands of years from now. The need to preserve it for scientific study, he argues, is not as relevant as for solar system bodies, because of the long timescales to get there and likely development of computer simulations to study evolution at early stages like that. The missions should value higher life and abort if any are detected from orbit. If the planet already has prokaryotes then he suggests that this may be ethically acceptable.

"For terrestrial life it is customto attribute value nearly exclusively to complex life, viz to animals and plants. Killinga few billion bacteria while brushing teeth does not cause, to give an example, moralheadaches The situation changes however when it comes to extrasolar life, for which wemay attribute value also to future evolutionary pathways. This is a delicate situation.Is it admissible to bring eukaryotes to a planet in a prokaryotic state, superseding suchindigenous life with lifeforms having the potential to develop into complex ecologies ... Endowing a selected number of exoplanetswith the possibility to evolve higher life forms would in this case not interfere with theevolution of yet simple life forms on potentially billions of other planets."