Surface characteristics of spacecraft components affect the aggregation of microorganisms and may lead to different survival rates of bacteria on Mars landers

Layers of dormant endospores of Bacillus subtilis HA101 were applied to eight different spacecraft materials and exposed to martian conditions of low pressure (8.5 mbar), low temperature (-10 degrees C), and high CO(2) gas composition and irradiated with a Mars-normal ultraviolet (UV-visible- near-infrared spectrum. Bacterial layers were exposed to either 1 min or 1 h of Mars-normal UV irradiation, which simulated clear-sky conditions on equatorial Mars (0.1 tau). When exposed to 1 min of Mars UV irradiation, the numbers of viable endospores of B. subtilis were reduced three to four orders of magnitude for two brands of aluminum (Al), stainless steel, chemfilm-treated Al, clear-anodized Al, and black-anodized Al coupons. In contrast, bacterial survival was reduced only one to two orders of magnitude for endospores on the non-metal materials astroquartz and graphite composite when bacterial endospores were exposed to 1 min of Mars UV irradiation. When bacterial monolayers were exposed to 1 h of Mars UV irradiation, no viable bacteria were recovered from the six metal coupons listed above. In contrast, bacterial survival was reduced only two to three orders of magnitude for spore layers on astroquartz and graphite composite exposed to 1 h of Mars UV irradiation. Scanning electron microscopy images of the bacterial monolayers on all eight spacecraft materials revealed that endospores of B. subtilis formed large aggregates of multilayered spores on astroquartz and graphite composite, but not on the other six spacecraft materials. It is likely that the formation of multilayered aggregates of endospores on astroquartz and graphite composite is responsible for the enhanced survival of bacterial cells on these materials.     

Survival of spores of the UV-resistant Bacillus subtilis strain MW01 after exposure to low-earth orbit and simulated martian conditions: data from the space experiment ADAPT on EXPOSE-E

In the space experiment "Molecular adaptation strategies of microorganisms to different space and planetary UV climate conditions" (ADAPT), bacterial endospores of the highly UV-resistant Bacillus subtilis strain MW01 were exposed to low-Earth orbit (LEO) and simulated martian surface conditions for 559 days on board the European Space Agency's exposure facility EXPOSE-E, mounted outside the International Space Station. The survival of B. subtilis MW01 spores from both assays (LEO and simulated martian conditions) was determined by a colony-formation assay after retrieval. It was clearly shown that solar extraterrestrial UV radiation (λ≥110?nm) as well as the martian UV spectrum (λ≥200?nm) was the most deleterious factor applied; in some samples only a few spore survivors were recovered from B. subtilis MW01 spores exposed in monolayers. However, if shielded from solar irradiation, about 8% of MW01 spores survived in LEO conditions, and 100% survived in simulated martian conditions, compared to the laboratory controls. The results demonstrate the effect of shielding against the high inactivation potential of extraterrestrial solar UV radiation, which limits the chances of survival of even the highly UV-resistant strain of B. subtilis MW01 in the harsh environments of outer space and the martian surface.     

Resistance of bacterial endospores to outer space for planetary protection purposes--experiment PROTECT of the EXPOSE-E mission

Spore-forming bacteria are of particular concern in the context of planetary protection because their tough endospores may withstand certain sterilization procedures as well as the harsh environments of outer space or planetary surfaces. To test their hardiness on a hypothetical mission to Mars, spores of Bacillus subtilis 168 and Bacillus pumilus SAFR-032 were exposed for 1.5 years to selected parameters of space in the experiment PROTECT during the EXPOSE-E mission on board the International Space Station. Mounted as dry layers on spacecraft-qualified aluminum coupons, the "trip to Mars" spores experienced space vacuum, cosmic and extraterrestrial solar radiation, and temperature fluctuations, whereas the "stay on Mars" spores were subjected to a simulated martian environment that included atmospheric pressure and composition, and UV and cosmic radiation. The survival of spores from both assays was determined after retrieval. It was clearly shown that solar extraterrestrial UV radiation (λ≥110?nm) as well as the martian UV spectrum (λ≥200?nm) was the most deleterious factor applied; in some samples only a few survivors were recovered from spores exposed in monolayers. Spores in multilayers survived better by several orders of magnitude. All other environmental parameters encountered by the "trip to Mars" or "stay on Mars" spores did little harm to the spores, which showed about 50% survival or more. The data demonstrate the high chance of survival of spores on a Mars mission, if protected against solar irradiation. These results will have implications for planetary protection considerations.     

Bacillus subtilis spore survival and expression of germination-induced bioluminescence after prolonged incubation under simulated Mars atmospheric pressure and composition: implications for planetary protection and lithopanspermia

Bacterial endospores in the genus Bacillus are considered good models for studying interplanetary transfer of microbes by natural or human processes. Although spore survival during transfer itself has been the subject of considerable study, the fate of spores in extraterrestrial environments has received less attention. In this report we subjected spores of a strain of Bacillus subtilis, containing luciferase resulting from expression of an sspB-luxAB gene fusion, to simulated martian atmospheric pressure (7-18 mbar) and composition (100% CO(2)) for up to 19 days in a Mars simulation chamber. We report here that survival was similar between spores exposed to Earth conditions and spores exposed up to 19 days to simulated martian conditions. However, germination-induced bioluminescence was lower in spores exposed to simulated martian atmosphere, which suggests sublethal impairment of some endogenous spore germination processes.     

Effects of a simulated martian UV flux on the cyanobacterium, Chroococcidiopsis sp. 029

Dried monolayers of Chroococcidiopsis sp. 029, a desiccation-tolerant, endolithic cyanobacterium, were exposed to a simulated martian-surface UV and visible light flux, which may also approximate to the worst-case scenario for the Archean Earth. After 5 min, there was a 99% loss of cell viability, and there were no survivors after 30 min. However, this survival was approximately 10 times higher than that previously reported for Bacillus subtilis. We show that under 1 mm of rock, Chroococcidiopsis sp. could survive (and potentially grow) under the high martian UV flux if water and nutrient requirements for growth were met. In isolated cells, phycobilisomes and esterases remained intact hours after viability was lost. Esterase activity was reduced by 99% after a 1-h exposure, while 99% loss of autofluorescence required a 4-h exposure. However, cell morphology was not changed, and DNA was still detectable by 4',6-diamidino-2-phenylindole staining after an 8-h exposure (equivalent to approximately 1 day on Mars at the equator). Under 1 mm of simulant martian soil or gneiss, the effect of UV radiation could not be detected on esterase activity or autofluorescence after 4 h. These results show that under the intense martian UV flux the morphological signatures of life can persist even after viability, enzymatic activity, and pigmentation have been destroyed. Finally, the global dispersal of viable, isolated cells of even this desiccation-tolerant, ionizing-radiation-resistant microorganism on Mars is unlikely as they are killed quickly by unattenuated UV radiation when in a desiccated state. These findings have implications for the survival of diverse microbial contaminants dispersed during the course of human exploratory class missions on the surface of Mars.     

Mutagenesis in bacterial spores exposed to space and simulated martian conditions: data from the EXPOSE-E spaceflight experiment PROTECT

As part of the PROTECT experiment of the EXPOSE-E mission on board the International Space Station (ISS), the mutagenic efficiency of space was studied in spores of Bacillus subtilis 168. After 1.5 years' exposure to selected parameters of outer space or simulated martian conditions, the rates of induced mutations to rifampicin resistance (Rif(R)) and sporulation deficiency (Spo(-)) were quantified. In all flight samples, both mutations, Rif(R) and Spo(-), were induced and their rates increased by several orders of magnitude. Extraterrestrial solar UV radiation (>110?nm) as well as simulated martian UV radiation (>200?nm) led to the most pronounced increase (up to nearly 4 orders of magnitude); however, mutations were also induced in flight samples shielded from insolation, which were exposed to the same conditions except solar irradiation. Nucleotide sequencing located the Rif(R) mutations in the rpoB gene encoding the β-subunit of RNA polymerase. Mutations isolated from flight and parallel mission ground reference (MGR) samples were exclusively localized to Cluster I. The 21 Rif(R) mutations isolated from the flight experiment showed all a C to T transition and were all localized to one hotspot: H482Y. In mutants isolated from the MGR, the spectrum was wider with predicted amino acid changes at residues Q469K/L/R, H482D/P/R/Y, and S487L. The data show the unique mutagenic power of space and martian surface conditions as a consequence of DNA injuries induced by solar UV radiation and space vacuum or the low pressure of Mars.     

The PROCESS experiment: amino and carboxylic acids under Mars-like surface UV radiation conditions in low-earth orbit

The search for organic molecules at the surface of Mars is a top priority of the next Mars exploration space missions: Mars Science Laboratory (NASA) and ExoMars (ESA). The detection of organic matter could provide information about the presence of a prebiotic chemistry or even biological activity on this planet. Therefore, a key step in interpretation of future data collected by these missions is to understand the preservation of organic matter in the martian environment. Several laboratory experiments have been devoted to quantifying and qualifying the evolution of organic molecules under simulated environmental conditions of Mars. However, these laboratory simulations are limited, and one major constraint is the reproduction of the UV spectrum that reaches the surface of Mars. As part of the PROCESS experiment of the European EXPOSE-E mission on board the International Space Station, a study was performed on the photodegradation of organics under filtered extraterrestrial solar electromagnetic radiation that mimics Mars-like surface UV radiation conditions. Glycine, serine, phthalic acid, phthalic acid in the presence of a mineral phase, and mellitic acid were exposed to these conditions for 1.5 years, and their evolution was determined by Fourier transform infrared spectroscopy after their retrieval. The results were compared with data from laboratory experiments. A 1.5-year exposure to Mars-like surface UV radiation conditions in space resulted in complete degradation of the organic compounds. Half-lives between 50 and 150?h for martian surface conditions were calculated from both laboratory and low-Earth orbit experiments. The results highlight that none of those organics are stable under low-Earth orbit solar UV radiation conditions.     

美国火星表面探测使命述评（上）

从1975年发射（1976年着陆火星）的海盗-1探测器以来,美国已成功执行了6次火星表面探测使命,即海盗-1与海盗-2轨道器/着陆器,＂火星探路者＂（MPF）着陆器/巡游车,＂勇气＂与＂机遇＂火星探测巡游车（MER）,以及＂凤凰＂着陆器;而推迟到2011年发射的＂好奇心＂火星科学实验室（MSL）将火星着陆技术与表面巡游车技术推向一个新的高度。从＂海盗＂着陆器到＂好奇心＂巡游车、美国历经三种火星着陆系统与三代火星表面巡游车技术的发展。三种着陆系统为着陆腿着陆系统（＂海盗＂与＂凤凰＂）,气囊着陆系统（MPF与MER）,以及空中吊机着陆系统（MSL）。三代巡游车为MPF＂旅居者＂巡游车、MER＂勇气＂与＂机遇＂巡游车,以及MSL＂好奇心＂巡游车。现在,美国在火星进入、降落与着陆（EDL）运作与表面避障移动方面,已达到技术成熟与先进的水平,满足安全着陆与表面移动探测的要求。文章阐述美国上述七项火星表面探测使命的立项背景、科学目标与有效载荷、飞行系统组成,以及飞行运作程序;分析美国火星着陆技术与表面巡游车技术的发展。     

美国火星表面探测使命述评（下）

从1975年8月发射（1976年7月着陆火星）的海盗-1探测器以来,美国已成功执行了6次火星表面探测使命,即海盗-1与海盗-2轨道器/着陆器,＂火星探路者＂（MPF）着陆器/巡游车,＂勇气＂与＂机遇＂火星探测巡游车（MER）,以及＂凤凰＂着陆器;而推迟到2011年发射的火星科学实验室（MSL）将火星着陆技术与表面巡游车技术推向一个新的高度。从＂海盗＂着陆器到MSL＂好奇心＂巡游车、美国历经三种火星着陆系统与三代火星表面巡游车技术的发展。三种着陆系统为着陆腿着陆系统（＂海盗＂与＂凤凰＂）,气囊着陆系统（MPF与MER）,以及空中吊机着陆系统（MSL）。三代巡游车为MPF＂旅居者＂巡游车、MER＂勇气＂与＂机遇＂巡游车,以及MSL＂好奇心＂巡游车。现在,美国在火星进入、降落与着陆（EDL）运作与表面避障移动方面,已达到技术成熟与先进的水平,满足安全着陆与表面移动探测的要求。文章阐述美国上述七项火星表面探测使命的立项背景、科学目标与有效载荷、飞行系统组成,以及飞行运作程序;分析美国火星着陆技术与表面巡游车技术的发展。     

一种三元结构的火星采样返回任务方案分析

介绍了目前国外提出的一种三元结构的火星采样返回任务方案,整个方案分3次发射,分别发射漫游车、着陆器和轨道器,每次发射间隔为4年,最终目的是将火星样品带回地球。该方案的优势在于,通过3次发射分别完成漫游车巡视勘察、着陆器现场探测、轨道器数据中继和在轨探测,最终综合完成火星采样返回,能够极大地缓解项目进度和资金压力,充分利用每次发射窗口分步骤完成探测任务。文章重点对方案涉及的关键技术进行了分析,包括样品获取与封装、行星保护、精准着陆、漫游车的危险规避能力和移动性、火星上升器、交会与样品捕获、地球再入器技术等;对方案的前景和优势进行了探讨,并给出几点启示,如精准着陆或成为今后行星探测着陆方式的新趋势,火星采样返回任务将是人类火星探测的里程碑,今后的深空探测任务趋向国际合作模式等。     

Survival and germinability of Bacillus subtilis spores exposed to simulated Mars solar radiation: implications for life detection and planetary protection

Bacterial spores have been considered as microbial life that could survive interplanetary transport by natural impact processes or human spaceflight activity. Deposition of terrestrial microbes or their biosignature molecules onto the surface of Mars could negatively impact life detection experiments and planetary protection measures. Simulated Mars solar radiation, particularly the ultraviolet component, has been shown to reduce spore viability, but its effect on spore germination and resulting production of biosignature molecules has not been explored. We examined the survival and germinability of Bacillus subtilis spores exposed to simulated martian conditions that include solar radiation. Spores of B. subtilis that contain luciferase resulting from expression of an sspB-luxAB gene fusion were deposited on aluminum coupons to simulate deposition on spacecraft surfaces and exposed to simulated Mars atmosphere and solar radiation. The equivalent of 42 min of simulated Mars solar radiation exposure reduced spore viability by nearly 3 logs, while germination-induced bioluminescence, a measure of germination metabolism, was reduced by less than 1 log. The data indicate that spores can retain the potential to initiate germination-associated metabolic processes and produce biological signature molecules after being rendered nonviable by exposure to Mars solar radiation.     

仿尺蠖型火星车设计与验证

针对传统被动悬架式火星车在火星复杂地形运动中可能存在的问题，设计了仿尺蠖型火星车，包括沉陷脱困设计、爬坡设计、防托底设计和抬轮设计；识别了仿尺蠖型火星车移动系统的关键参数，计算得到不同火星车抬升高度的关键参数值和一个尺蠖运动周期的位移值；计算结果表明仿尺蠖型火星车可以解决被动悬架式火星车的上述问题。对所设计的仿尺蠖型火星车开展了地面试验和飞行试验验证，地面试验验证了车轮沉陷脱困能力、越障能力和爬坡能力；作为仿尺蠖型火星车应用的实例，“祝融号”火星车成功经历了飞行试验，它在火星表面的顺利运行表明仿尺蠖型火星车设计措施有效。     

Immunological detection of small organic molecules in the presence of perchlorates: relevance to the life marker chip and life detection on Mars

The proposed ExoMars mission, due to launch in 2018, aims to look for evidence of extant and extinct life in martian rocks and regolith. Previous attempts to detect organic molecules of biological or abiotic origin on Mars have been unsuccessful, which may be attributable to destruction of these molecules by perchlorate salts during pyrolysis sample extraction techniques. Organic molecules can also be extracted and measured with solvent-based systems. The ExoMars payload includes the Life Marker Chip (LMC) instrument, capable of detecting biomarker molecules of extant and extinct Earth-like life in liquid extracts of martian samples with an antibody microarray assay. The aim of the work reported here was to investigate whether the presence of perchlorate salts, at levels similar to those at the NASA Phoenix landing site, would compromise the LMC extraction and detection method. To test this, we implemented an LMC-representative sample extraction process with an LMC-representative antibody assay and used these to extract and analyze a model sample that consisted of a Mars analog sample matrix (JSC Mars-1) spiked with a representative organic molecular target (pyrene, an example of abiotic meteoritic infall targets) in the presence of perchlorate salts. We found no significant change in immunoassay function when using pyrene standards with added perchlorate salts. When model samples spiked with perchlorate salts were subjected to an LMC-representative liquid extraction, immunoassays functioned in a liquid extract and detected extracted pyrene. For the same model sample matrix without perchlorate salts, we observed anomalous assay signals that coincided with yellow coloration of the extracts. This unexpected observation is being studied further. This initial study indicates that the presence of perchlorate salts, at levels similar to those detected at the NASA Phoenix landing site, is unlikely to prevent the LMC from extracting and detecting organic molecules from martian samples.     

地面力学在火星壤力学参数估计研究中的 进展与展望

掌握火星土壤的力学参数是保障火星车顺利完成巡视探测的关键。通过对已成功开展的火星巡视探测任务进行汇总，探讨了复杂火星地貌对火星车移动性能的影响，分析了基于地面力学理论的火星土壤力学参数估计方法，包括：基于车轮的火星壤在轨力学参数估计方法和基于轮壤力学模型的力学参数辨识方法。最后，对未来关于地面力学在星壤力学参数评估进行了探讨。     

Vanguard--a European robotic astrobiology-focussed Mars sub-surface mission proposal

We present a new European Mars mission proposal to build on the UK-led Beagle2 Mars mission and continue its astrobiology-focussed investigation of Mars. The small surface element to be delivered to the Martian surface--Vanguard--is designed to be carried by a Mars Express-type spacecraft bus to Mars and adopts a similar entry, descent and landing system as Beagle2. The surface element comprises a triad of robotic devices--a lander, a micro-rover of the Sojourner class for surface mobility, and three ground-penetrating moles mounted onto the rover for sub-surface penetration to 5 m depth. The major onboard instruments on the rover include a Raman spectrometer/imager, a laser plasma spectrometer, an infrared spectrometer--these laser instruments provide the basis for in situ "remote" sensing of the sub-surface Martian environment within a powerful scientific package. The moles carry the instruments' sensor head array to the sub-surface. The moles are thus required to undergo a one-way trip down the boreholes without the need for recovery of moles or samples, eliminating much of the robotic complexity invoked by such operations.     

火星精确着陆制导问题分析与展望

美国火星科学实验室(MSL)任务成功将“好奇”号火星车着陆到火星表面,开创了火星精确着陆探测的新局面。以MSL着陆任务为典型代表,分析了目前火星着陆探测进入、下降和着陆（Entry, Descent and Landing, EDL）过程的制导方案及制导系统的发展趋势。以在火星高海拔、复杂地形区域定点着陆为潜在工程目标,归纳了火星EDL过程面临的制导主要问题。根据未来制导系统自主性和自适应性的技术需求及潜在工程任务制导面临的问题,提出了火星EDL制导方面需要解决的关键技术,并对其在未来工程中的应用潜力进行了展望。     

Survival of Bacillus pumilus spores for a prolonged period of time in real space conditions

To prevent forward contamination and maintain the scientific integrity of future life-detection missions, it is important to characterize and attempt to eliminate terrestrial microorganisms associated with exploratory spacecraft and landing vehicles. Among the organisms isolated from spacecraft-associated surfaces, spores of Bacillus pumilus SAFR-032 exhibited unusually high resistance to decontamination techniques such as UV radiation and peroxide treatment. Subsequently, B. pumilus SAFR-032 was flown to the International Space Station (ISS) and exposed to a variety of space conditions via the European Technology Exposure Facility (EuTEF). After 18 months of exposure in the EXPOSE facility of the European Space Agency (ESA) on EuTEF under dark space conditions, SAFR-032 spores showed 10-40% survivability, whereas a survival rate of 85-100% was observed when these spores were kept aboard the ISS under dark simulated martian atmospheric conditions. In contrast, when UV (>110?nm) was applied on SAFR-032 spores for the same time period and under the same conditions used in EXPOSE, a ～7-log reduction in viability was observed. A parallel experiment was conducted on Earth with identical samples under simulated space conditions. Spores exposed to ground simulations showed less of a reduction in viability when compared with the "real space" exposed spores (～3-log reduction in viability for "UV-Mars," and ～4-log reduction in viability for "UV-Space"). A comparative proteomics analysis indicated that proteins conferring resistant traits (superoxide dismutase) were present in higher concentration in space-exposed spores when compared to controls. Also, the first-generation cells and spores derived from space-exposed samples exhibited elevated UVC resistance when compared with their ground control counterparts. The data generated are important for calculating the probability and mechanisms of microbial survival in space conditions and assessing microbial contaminants as risks for forward contamination and in situ life detection.     

Microbial diversity of Indian Ocean hydrothermal vent plumes: microbes tolerant of desiccation, peroxide exposure, and ultraviolet and gamma-irradiation

The microbial diversity of Kali chimney plumes, part of a hydrothermal vent field in the Rodriguez Triple Junction, Indian Ocean (depth approximately 2,240 m), was examined in an attempt to discover "extremotolerant" microorganisms that have evolved unique resistance capabilities to this harsh environment. Water and sediment samples were collected from the vent and from sediments located at various distances (2-20 m) away from and surrounding the chimney. Samples were screened for hypertolerant microbes that are able to withstand multiple stresses. A total of 46 isolates were selected for exposure to a number of perturbations, such as heat shock, desiccation, H(2)O(2), and ultraviolet (UV) and gamma-irradiation. The survival of Psychrobacter sp. L0S3S-03b following exposure to >1,000 J/m(2) UV(254) radiation was particularly intriguing amid a background of varying levels of resistance. Vegetative cells of this non-spore-forming microbe not only survived all of the treatments, but also exhibited a 90% lethal dose of 30 s when exposed to simulated martian UV radiation and a 100% lethal dose of 2 min when exposed to full spectrum UV, which is comparable to findings for bacterial endospores.     

The photochemical stability of carbonates on Mars

Carbonates, predominately MgCO3, have been spectroscopically identified at a level of 2-5% in martian dust. However, in spite of this observation, and a large number of climate studies that suggest 1 to several bars of CO2 should be sequestered in carbonate rocks, no outcrop-scale exposures of carbonate have been detected anywhere on Mars to date. To address one hypothesis for this long-standing puzzle, the effect of ultraviolet (UV) light on the stability of calcium carbonate in a simulated martian atmosphere was experimentally investigated. Using 13C-labeled calcite, we found no experimental evidence of the UV photodecomposition of calcium carbonate in a simulated martian atmosphere. Extrapolating the lower limit of detection of our experimental system to an upper limit of carbonate decomposition on Mars yields a quantum efficiency of 3.5 x 10(-8) molecules/photon over the wavelength interval of 190-390 nm and a maximum UV photodecomposition rate of 1.2 x 10(-13) kg m(-2) s(-1) from a calcite surface. The maximum loss of bulk calcite due to this process would be 2.5 nm year(-1) (Mars year). However, calcite is expected to be thermodynamically stable on the surface of Mars, and potential UV photodecomposition reaction mechanisms indicate that, though calcium carbonate may decompose under vacuum, it would be stable in a CO2 atmosphere. Given the expected stability of carbonate on Mars and our inability to detect carbonate decomposition, we conclude that it is unlikely that the apparent absence of extensive carbonate deposits on the martian surface is due to UV photodecomposition in the current environment.