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NASA NTRS2005

Microbiological Methodology in Astrobiology

Searching for life in astromaterials to be delivered from the future missions to extraterrestrial bodies is undoubtedly related to studies of the properties and signatures of living microbial cells and microfossils on Earth. As model terrestrial analogs of Martian polar subsurface layers are often regarded the Antarctic glacier and Earth permafrost habitats where alive microbial cells preserved viability for millennia years due to entering the anabiotic state. For the future findings of viable microorganisms in samples from extraterrestrial objects, it is important to use a combined methodology that includes classical microbiological methods, plating onto nutrient media, direct epifluorescence and electron microscopy examinations, detection of the elemental composition of cells, radiolabeling techniques, PCR and FISH methods. Of great importance is to ensure authenticity of microorganisms (if any in studied samples) and to standardize the protocols used to minimize a risk of external contamination. Although the convincing evidence of extraterrestrial microbial life will may come from the discovery of living cells in astromaterials, biomorphs and microfossils must also be regarded as a target in search of life evidence bearing in mind a scenario that alive microorganisms had not be preserved and underwent mineralization. Under the laboratory conditions, processes that accompanied fossilization of cyanobacteria were reconstructed, and artificially produced cyanobacterial stromatolites resembles by their morphological properties those found in natural Earth habitats. Regarding the vital importance of distinguishing between biogenic and abiogenic signatures and between living and fossil microorganisms in analyzed samples, it is worthwhile to use some previously developed approaches based on electron microscopy examinations and analysis of elemental composition of biomorphs in situ and comparison with the analogous data obtained for laboratory microbial cultures and fossilized microorganisms. This communication will be focused on the analysis of our experience in working with ancient microorganisms and fossils and discussion of some issues that are crucial for development of the program for future finding of extraterrestrial life and its evidence.

Abyzov, S. S.↗

NASA NTRS2007

Astrobiological Significance of Microbial Extremophiles

The microflora of the cryosphere of planet Earth provides the best analogs for life forms that might be found in the permafrost or polar ice caps of Mars, near the surface of the cometary nuclei, or in the liquid water beneath and the ice crusts of icy moons of Jupiter and Saturn. The importance of study alkaliphilic microorganisms for astrobiology was enhanced by the findings of abundant carbonates and carbonate globules rimmed with possibly biogenic magnetites in association with the putative microfossils in the ALH84001 meteorite. Although the ALH84001 "nanofossils" were to small and simple to be unambiguously recognized as biogenic, they stimulated Astrobiology research and studies of microbial extremophiles and biomarkers in ancient rocks and meteorites. Recent studies of CI and CM carbonaceous meteorites have resulted in the detection of the well-preserved mineralized remains of coccoidal and filamentous microorganisms in cyanobacterial mats. Energy Dispersive X-ray Analysis has shown anomalous biogenic element ratios clearly indicating they are not recent biological contaminants. This paper reviews microbial extremophiles in context of their significance to Astrobiology. The study of halophilic microorganisms was started from work with saline soils and lakes, and one of the record of good growth for Haloferax mediterranei was shown at 30 percent NaC1. Although alkali-tolerant nitrifying bacteria had previously been reported, the first described alkaliphilic microorganism was the bacterium Streptococcus faecalis. Halophilic and alkaliphilic forms are relevant to conditions that might be found in closed impact basins and craters on Mars filled with evaporite deposits. The first obligately acidophilic bacterium described was Acidithiobacillus ferrooxydans (formally Thiobacillus ferrooxidans). Later thermophilic lithotrophic acidophiles were found, and the hyperacidophilic moderately thermophilic species of the genus Picrophilus were found to grow at negative pH. The epoch of study of thermophilic microorganisms starts with the discovery of Thermus aquaticus, and presently the maximum temperature for growth at 113 C was found for Pyrolobus fumarii. The microorganisms capable of growth at high temperatures and in hyperacidic environments on Earth are good analogs for life that might be able to survive in hot acidic droplets in the upper regimes of the atmosphere of Venus. The study of barophiles was made possible by engineering achievements leading to the development of the submersible crafts used to study the Black Smokers of the Deep-sea Hydrothermal vents. The first described radioresistant bacterium Deinococcus radiodurans can survive ionizing irradiation and other DNA-damaging assaults at doses that are lethal to all other organisms. These microbes are models for life that might endure high radiation environments in the ice near the surface of comets or on the icy moons of Jupiter and Saturn and in the seafloor deep beneath icy crusts Europa and Enceladus. This paper presents ESEM and FESEM images showing intact microbes preserved in the deep ice cores extracted from just above Lake Vostok, Antarctica that are considered analogs for life forms that might survive on comets and icy moons.

Pikuta, Elena V.↗

NASA NTRS2015

Inflight Microbial Monitoring- An Alternative Method to Culture Based Detection Currently Used on the International Space Station

Previous research has shown that potentially destructive microorganisms and human pathogens have been detected on the International Space Station (ISS). The likelihood of introducing new microorganisms occurs with every exchange of crew or addition of equipment or supplies. Microorganisms introduced to the ISS are readily transferred between crew and subsystems (i.e. ECLSS, environmental control and life support systems). Current microbial characterization methods require enrichment of microorganisms and at least a 48-hour incubation time. This increases the microbial load while detecting only a limited number of the total microorganisms. The culture based method detects approximately 1-10% of the total organisms present and provides no identification. To identify and enumerate ISS microbes requires that samples be returned to Earth for complete analysis. Therefore, a more expedient, low-cost, in-flight method of microbial detection, identification, and enumeration is warranted. The RAZOR EX, a ruggedized, commercial off the shelf, real-time PCR field instrument was tested for its ability to detect microorganisms at low concentrations within one hour. Escherichia coli, Salmonella enterica Typhimurium, and Pseudomonas aeruginosa were detected at low levels using real-time DNA amplification. Total heterotrophic counts could also be detected using a 16S gene marker that can identify up to 98% of all bacteria. To reflect viable cells found in the samples, RNA was also detectable using a modified, single-step reverse transcription reaction.

Inflight Microbial↗

NASA NTRS2015

Inflight Microbial Monitoring-An Alternative Method to Culture Based Detection Currently Used on International Space Station

Previous research has shown that microorganisms and potential human pathogens have been detected on the International Space Station (ISS). The potential to introduce new microorganisms occurs with every exchange of crew or addition of equipment or supplies. Previous research has shown that microorganisms introduced to the ISS are readily transferred between crew and subsystems and back (i.e. ECLSS, environmental control and life support systems). Current microbial characterization methods require enrichment of microorganisms and a 48-hour incubation time. This increases the microbial load while detecting a limited number of microorganisms. The culture based method detects approximately 1-10% of the total organisms present and provides no identification, To identify and enumerate ISS samples requires that samples to be returned to Earth for complete analysis. Therefore, a more expedient, low-cost, in-flight method of microbial detection, identification, and enumeration is warranted. The RAZOR EX, a ruggedized, commercial off the shelf, real-time PCR field instrument was tested for its ability to detect microorganism at low concentrations within one hour. Escherichia coli, Salmonella enterica Typhimurium, and Pseudomonas aeruginosa were detected at low levels using real-time DNA amplification. Total heterotrophic counts could also be detected using a 16S gene marker that can identify up to 98% of all bacteria. To reflect viable cells found in the samples, RNA was also detectable using a modified, single-step reverse transcription reaction.

Inflight Microbial↗

NASA NTRS1982

Apparatus and process for microbial detection and enumeration

An apparatus and process for detecting and enumerating specific microorganisms from large volume samples containing small numbers of the microorganisms is presented. The large volume samples are filtered through a membrane filter to concentrate the microorganisms. The filter is positioned between two absorbent pads and previously moistened with a growth medium for the microorganisms. A pair of electrodes are disposed against the filter and the pad electrode filter assembly is retained within a petri dish by retainer ring. The cover is positioned on base of petri dish and sealed at the edges by a parafilm seal prior to being electrically connected via connectors to a strip chart recorder for detecting and enumerating the microorganisms collected on filter.

Wilkins, J. R.↗

NASA NTRS2001

Trace level detection of analytes using artificial olfactometry

The present invention provides a device for detecting the presence of an analyte, wherein said analyte is a microorganism marker gas. The device comprises a sample chamber having a fluid inlet port for the influx of the microorganism marker gas; a fluid concentrator in flow communication with the sample chamber, wherein the fluid concentrator has an absorbent material capable of absorbing the microorganism marker gas and thereafter releasing a concentrated microorganism marker gas; and an array of sensors in fluid communication with the concentrated microorganism marker gas. The sensor array detects and identifies the marker gas upon its release from fluid concentrate.

Lewis, Nathan S.↗

NASA NTRS2012

Next Generation Microbiology Requirements

As humans continue to explore deep into space, microorganisms will travel with them. The primary means to mitigate the risk of infectious disease are a combination of prudent spacecraft design and rigorous operational controls. The effectiveness of these methods are evaluated by microbiological monitoring of spacecraft, food, water, and the crew that is performed preflight, in-flight, and post-flight. Current NASA requirements associated with microbiological monitoring are based on culture-based methodology where microorganisms are grown on a semi-solid growth medium and enumerated. Subsequent identification of the organisms requires specialized labor and large equipment, which historically has been performed on Earth. Requirements that rely strictly on culture-based units limit the use of non-culture based monitoring technology. Specifically, the culture-based "measurement criteria" are Colony Forming Units (CFU, representing the growth of one microorganism at a single location on the agar medium) per a given volume, area, or sample size. As the CFU unit by definition is culture-based, these requirements limit alternative technologies for spaceflight applications. As spaceflight missions such as those to Mars extend further into space, culture-based technology will become difficult to implement due to the (a) limited shelf life of the culture media, (b) mass/volume necessary to carry these consumables, and (c) problems associated with the production of biohazardous material in the habitable volume of the spacecraft. In addition, an extensive amount of new knowledge has been obtained during the Space Shuttle, NASA-Mir, and International Space Station Programs, which gave direction for new or modified microbial control requirements for vehicle design and mission operations. The goal of this task is to develop and recommend a new set of requirements for vehicle design and mission operations, including microbiological monitoring, based upon "lessons learned" and new technology. During 2011, this study focused on evaluating potable water requirements by assembling a forum of internal and external experts from NASA, other federal agencies, and academia. Key findings from this forum included: (1) Preventive design and operational strategies should be stringent and the primary focus of NASA's mitigation efforts, as they are cost effective and can be attained with conventional technology. (2) Microbial monitoring hardware should be simple and must be able to measure the viability of microorganisms in a sample. Multiple monitoring technologies can be utilized as long as at the microorganisms being identified can also be confirmed as viable. (3) Evidence showing alterations in the crew immune function and microbial virulence complicates risk assessments and creates the need for very conservative requirements. (4) One key source of infectious agents will always be the crew, and appropriate preventative measures should be taken preflight. (5) Water systems should be thoroughly disinfected (sterilized if possible) preflight and retain a residual biocide throughout the mission. Future forums will cover requirements for other types of samples, specifically spaceflight food and environmental samples, such as vehicle air and vehicle and cargo surfaces. An interim report on the potable water forum has been delivered to the Human Research Program with a final report on the recommendations for all sample types being delivered in September 2013.

Ott, C. M.↗

NASA NTRS2015

In-Flight Microbial Monitor

Previous research has shown that potential human pathogens have been detected on the International Space Station (ISS). New microorganisms are introduced with every exchange of crew and cargo. Microorganisms introduced to the ISS are readily transferred between crew and subsystems (i.e., ECLSS, environmental control and life support systems). Current microbial characterization methods require a culture-based enrichment of microorganisms and at least a 48-hour incubation time. This increases the microbial load while detecting only a limited number of microorganisms. The culture-based method detects approximately 1-10% of the total organisms present and provides no identification. To identify and enumerate ISS samples requires that the microbes be returned to Earth for complete analysis. Therefore, a more expedient, low-cost, inflight method of microbial detection, identification, and enumeration is needed. The RAZOR EX, a ruggedized, commercial off the shelf, real-time PCR field instrument was tested for its ability to detect microorganisms at low concentrations within one hour. Escherichia coli, Salmonella enterica Typhimurium, and Pseudomonas aeruginosa were detected at low levels using real-time DNA amplification. Total heterotrophic counts could also be detected using a 16S gene marker that can identify up to 98% of all bacteria. To reflect viable cells found in the samples, RNA was also detectable using a modified, single-step reverse transcription reaction.

Technology Portfolio System↗

NASA NTRS1974

Microbiological and experimental-histological investigations of lunar samples returned by the Lunar 16 automatic station

Lunar surface material was studied for its content of viable microorganisms (aerobic and anaerobic, fungi, and viruses); the effect of the lunar surface material on the growth of microorganisms and its interaction with somatic cells of mammals was also observed. No viable microorganisms were detected; the samples exhibited neither stimulant or inhibitory action on the growth of microorganisms, and also showed no cytopathogenic action on tissue cultures. A suspension of lunar surface material particles was not toxic when parenterally administered to certain laboratory animals. The particles were subjected to intense phagocytosis by connective tissue cells in vivo and in vitro.

Kaulen, D. R.↗

NASA NTRS1978

Microbial Check Valve for Shuttle

The Microbial Check Valve (MCV) is a device developed for the Space Shuttle that prevents the transfer of viable microorganisms within water systems. The device is essentially a bed of resin material, impregnated with iodine, that kills microorganisms on contact. It prevents the cross-contamination of microorganisms from a nonpotable system into the potable water system when these systems are interconnected. In this regard, the function of the device is similar to that of the 'air gap' found in conventional one-gravity systems. Basic design data are presented including pressure drop, scaling factors, sizing criteria, and the results of challenging the device with suspensions of seven microorganisms including aerobes, anaerobes and spore formers.

Colombo, G. V.↗

NASA NTRS1985

Report of the Microbial Development Working Group

In formulating ideas on the relationship of gravity to the development, growth, and reproduction of microorganisms, a rather liberal definition of microorganisms is used which includes bacteria, yeasts, protists, filamentous fungi, and single cells in culture. A principal advantage of microorganisms as experimental subjects is the rigor with which they can be defined and controlled. As single cells, each cell may be regarded as identical to the others in the population. This property applies to the morphology, physiology, and genetic parameters of the cells. The growth and development of the population is subject to precise manipulation as the nutritional requirements are known and minimal media formulations have been developed. Growth and differentiation can be manipulated in a variety of ways, such as alteration of the culture temperature and food supply, or by use of mutants. Finally, the short generation times of microorganisms provide the opportunity to conduct multigenerational studies within practical time limits and, in a similar vein, cellular responses to various stimuli or stresses are conveniently monitored because of the rapid response times of single cells.

Nelson, G.↗

NASA NTRS1989

Growth of a mat-forming photograph in the presence of UV radiation

Knowledge of the survival and growth of microorganisms in the presence of ultraviolet radiation is important for understanding the potential for life to exist in environments exposed to high fluxes of UV radiation. The growth of a mat-forming phototrophic prokaryote, Chloroflexus aurantiacus, was examined in the presence of continuous high UV irradiation under otherwise optimal growth conditions. Evidence was sought for an intrinsic ability to grow in the presence of UV radiation in a carefully chosen organism known to be unusually resistant to UV radiation, of ancient lineage among the phototrophs, to resemble ancient microfossils from the Precambrian, and to be a mat-former. It was assumed that even a high intrinsic UV resistance would be inadequate for survival and growth in the presence of very high UV fluxes, and iron (Fe3+) was selected as a common, abundant UV-absorbing substance that might protest microorganisms growing in or under iron-bearing sediments. The effectiveness of Fe(3+) was tested as a UV protective agent at low concentrations in thin layers. It was concluded that intrinsic UV resistance in some organisms may account for growth, not just survival, of these organisms when exposed to high UV fluxes under otherwise optimal growth conditions in an anoxic environment. It was also concluded that Fe(3+) bearing sediments of 1 mm or less in thickness may provide an adequate shield against high UV fluxes permitting the growth of microorganisms just below their surface. As long as growth conditions were met, then the evolution and development of microorganisms would not be hampered by high UV fluxes impinging upon the surface of iron-bearing sediments.

Pierson, Beverly K.↗

NASA NTRS2002

Environmental Regulation of Microbial Community Structure

Most naturally occurring microbes live in complex microbial communities consisting of thousands of phylotypes of microorganisms living in close proximity. Each of these draws nutrients from the environment and releases metabolic waste products, which may in turn serve as substrates for other microbial groups. Gross environmental changes, such as irradiance level, hydrodynamic flow regime, temperature or water chemistry can directly affect the productivity of some community members, which in turn will affect other dependent microbial populations and rate processes. As a first step towards the development of "standard" natural communities of microorganisms for a variety of potential NASA applications, we are measuring biogeochemical cycling in artificially structured communities of microorganisms, created using natural microbial mat communities as inoculum. The responses of these artificially assembled communities of microorganisms to controlled shifts in ecosystem incubation conditions is being determined. This research requires close linking of environmental monitoring, with community composition in a closed and controlled incubation setting. We are developing new incubation chamber designs to allow for this integrated approach to examine the interplay between environmental conditions, microbial community composition and biogeochemical processes.

Bebout, Leslie↗

NASA NTRS2001

Microbial contamination of spacecraft

Spacecraft and space habitats supporting human exploration contain a diverse population of microorganisms. Microorganisms may threaten human habitation in many ways that directly or indirectly impact the health, safety, or performance of astronauts. The ability to produce and maintain spacecraft and space stations with environments suitable for human habitation has been established over 40 years of human space flight. An extensive database of environmental microbiological parameters has been provided for short-term (< 20 days) space flight by more than 100 missions aboard the Space Shuttle. The NASA Mir Program provided similar data for long-duration missions. Interestingly, the major bacterial and fungal species found in the Space Shuttle are similar to those encountered in the nearly 15-year-old Mir. Lessons learned from both the US and Russian space programs have been incorporated into the habitability plan for the International Space Station. The focus is on preventive measures developed for spacecraft, cargo, and crews. On-orbit regular housekeeping practices complete with visual inspections are essential, along with microbiological monitoring. Risks associated with extended stays on the Moon or a Mars exploration mission will be much greater than previous experiences because of additional unknown variables. The current knowledge base is insufficient for exploration missions, and research is essential to understand the effects of space flight on biological functions and population dynamics of microorganisms in spacecraft. Equally important is a better understanding of the immune response and of human-microorganism-environment interactions during long-term space habitation.

Mir Project↗

NASA NTRS1986

Organically preserved microbial endoliths from the late Proterozoic of East Greenland

Diverse microorganisms ranging from cyanobacteria to eukaryotic algae and fungi live endolithically within ooids, hardgrounds and invertebrate shells on the present-day sea floor. These organisms are involved in the mechanical destruction of carbonates, and are useful ecological indicators of water depth and pollution. The Phanerozoic history of microbial endoliths has been elucidated through the study of microborings (the trace fossils of endolithic microorganisms) and rare cellularly preserved individuals, but nothing was known of the possible Precambrian evolution of comparable microorganisms until Campbell documented the occurrence of microborings in late Proterozoic ooids from central East Greenland. We now report the discovery of large populations of organically preserved endolithic microorganisms in silicified pisolites from 700-800-Myr-old Limestone-Dolomite Series of East Greenland. This fossil assemblage is significant for three reasons: (1) It confirms the prediction that oolites, pisolites and hardgrounds--the substrates for pre-Phanerozoic endoliths--provide a hitherto poorly explored but rewarding set of environments into which the search for early microfossils must be broadened; (2) the assemblage is diverse, containing about 12 taxa of morphologically distinct and previously unknown endolithic cyanobacteria, plus associated epilithic and interstitial populations; and (3) at least six of the fossil populations are indistinguishable in morphology, pattern of development, reproductive biology and inferred ecology from distinctive cyanobacterial species that bore ooids today in the Bahama Banks.

Non-NASA Center↗

NASA NTRS2004

Microbial astronauts: assembling microbial communities for advanced life support systems

Extension of human habitation into space requires that humans carry with them many of the microorganisms with which they coexist on Earth. The ubiquity of microorganisms in close association with all living things and biogeochemical processes on Earth predicates that they must also play a critical role in maintaining the viability of human life in space. Even though bacterial populations exist as locally adapted ecotypes, the abundance of individuals in microbial species is so large that dispersal is unlikely to be limited by geographical barriers on Earth (i.e., for most environments "everything is everywhere" given enough time). This will not be true for microbial communities in space where local species richness will be relatively low because of sterilization protocols prior to launch and physical barriers between Earth and spacecraft after launch. Although community diversity will be sufficient to sustain ecosystem function at the onset, richness and evenness may decline over time such that biological systems either lose functional potential (e.g., bioreactors may fail to reduce BOD or nitrogen load) or become susceptible to invasion by human-associated microorganisms (pathogens) over time. Research at the John F. Kennedy Space Center has evaluated fundamental properties of microbial diversity and community assembly in prototype bioregenerative systems for NASA Advanced Life Support. Successional trends related to increased niche specialization, including an apparent increase in the proportion of nonculturable types of organisms, have been consistently observed. In addition, the stability of the microbial communities, as defined by their resistance to invasion by human-associated microorganisms, has been correlated to their diversity. Overall, these results reflect the significant challenges ahead for the assembly of stable, functional communities using gnotobiotic approaches, and the need to better define the basic biological principles that define ecosystem processes in the space environment. Copyright 2004 Springer-Verlag.

Review↗

NASA NTRS2010

Psychrophilic and Psychrotolerant Microbial Extremophiles in Polar Environments

The microbial extremophiles that inhabit the polar regions of our planet are of tremendous significance. The psychrophilic and psychrotolerant microorganisms, which inhabit all of the cold environments on Earth have important applications to Bioremediation, Medicine, Pharmaceuticals, and many other areas of Biotechnology. Until recently, most of the research on polar microorganisms was confined to studies of polar diatoms, yeast, fungi and cyanobacteria. However, within the past three decades, extensive studies have been conducted to understand the bacteria and archaea that inhabit the Arctic and Antarctic sea-ice, glaciers, ice sheets, permafrost and the cryptoendolithic, cryoconite and ice-bubble environments. These investigations have resulted in the discovery of many new genera and species of anaerobic and aerobic microbial extremophiles. Exotic enzymes, cold-shock proteins and pigments produced by some of the extremophiles from polar environments have the potential to be of great benefit to Mankind. Knowledge about microbial life in the polar regions is crucial to understanding the limitations and biodiversity of life on Earth and may provide valuable clues to the Origin of Life on Earth. The discovery of viable microorganisms in ancient ice from the Fox Tunnel, Alaska and the deep Vostok Ice has shown that microorganisms can remain alive while cryopreserved in ancient ice. The psychrophilic lithoautotrophic homoacetogen isolated from the deep anoxic trough of Lake Untersee is an ideal candidate for life that might inhabit comets or the polar caps of Mars. The spontaneous release of gas from within the Anuchin Glacier above Lake Untersee may provide clues to the ice geysers that erupt from the tiger stripe regions of Saturn s moon Enceladus. The methane productivity in the lower regimes of Lake Untersee may also provide insights into possible mechanisms for the recently discovered methane releases on Mars. Since most of the other water bearing bodies of our Solar system are frozen worlds, microbial extremophiles from the Polar Regions of Earth are of great importance to Astrobiology in understanding where and how to search for evidence of life elsewhere in the Cosmos.

Hoover, Richard B.↗

NASA NTRS2011

Microbiological Lessons Learned from the Space Shuttle

After 30 years of being the centerpiece of NASA s human spacecraft, the Space Shuttle will retire. This highly successful program provided many valuable lessons for the International Space Station (ISS) and future spacecraft. Major microbiological risks to crewmembers include food, water, air, surfaces, payloads, animals, other crewmembers, and ground support personnel. Adverse effects of microorganisms are varied and can jeopardize crew health and safety, spacecraft systems, and mission objectives. Engineering practices and operational procedures can minimize the negative effects of microorganisms. To minimize problems associated with microorganisms, appropriate steps must begin in the design phase of new spacecraft or space habitats. Spacecraft design must include requirements to control accumulation of water including humidity, leaks, and condensate on surfaces. Materials used in habitable volumes must not contribute to microbial growth. Use of appropriate materials and the implementation of robust housekeeping that utilizes periodic cleaning and disinfection will prevent high levels of microbial growth on surfaces. Air filtration can ensure low levels of bioaerosols and particulates in the breathing air. The use of physical and chemical steps to disinfect drinking water coupled with filtration can provide safe drinking water. Thorough preflight examination of flight crews, consumables, and the environment can greatly reduce pathogens in spacecraft. The advances in knowledge of living and working onboard the Space Shuttle formed the foundation for environmental microbiology requirements and operations for the International Space Station (ISS) and future spacecraft. Research conducted during the Space Shuttle Program resulted in an improved understanding of the effects of spaceflight on human physiology, microbial properties, and specifically the host-microbe interactions. Host-microbe interactions are substantially affected by spaceflight. Astronaut immune functions were found to be altered. Selected microorganisms were found to become more virulent during spaceflight. The increased knowledge gained on the Space Shuttle resulted in further studies of the host-microbe interactions on the ISS to determine if countermeasures were necessary. Lessons learned from the Space Shuttle Program were integrated into the ISS resulting in the safest space habitat to date.

Pierson, Duane L.↗