rRNA sequences and shotgun metagenomic analyses (enabled by anticipated improvements in DNA sequencing efficiencies) of low-biomass samples would provide a cultivation-independent assessment of microbial community composition within spacecraft assembly facilities and within and on a spacecraft.
Decision Point 1—Liquid Water
The absolute and unambiguous requirement of liquid water for the propagation of terrestrial organisms on Earth or on icy bodies in the outer solar system constrains the possibility of forward contamination to a relatively small number of objects in the outer solar system. See Chapter 4 for a detailed discussion of this point.
Decision Point 2—Key Elements
The origin and development of life are intimately linked to the periodic table of elements. The most important elements required by living systems for catalysis, organization of macromolecular structure, or energy transduction include, but are not limited to, carbon, hydrogen, nitrogen, oxygen, phosphorus, sulfur, potassium, magnesium, calcium, and iron. Sometimes other elements such as boron participate in chemical signaling between bacteria,2 or elements like selenium can incorporate into specific proteins as selanocysteine, now referred to as the twenty-first essential amino acid.3
Because of its pervasive role in many biological processes, phosphorus plays an indispensible role in living systems. In some marine organisms, arsenic can incorporate into lipids in place of phosphorus,4 but this generally toxic compound does not replace phosphorus in nucleic acids, in protein structures, or in catalytic functions. There is no consensus as to the minimum concentration of phosphorus required for growth by microorganisms. The cultivation of different bacterial taxa in phosphate at varying concentrations reveals a complex set of genes that mediate stress-response to phosphate limitation. In general, phosphorus limitation triggers adaption, including the up-regulation of specific genes involved in phosphate response stress that shift cells from utilization of inorganic phosphate to scavenging of organo-phosphate and polyphosphate from the environment,5,6 substituting sulfur for phosphate in membrane lipids in marine photosynthetic bacteria,7 changes in cell morphology to increase surface-to-volume ratio of the cell,8 and shutting down cell metabolism to survive in a dormant stage.9,10,11 In most cases, these studies were preformed with bacteria that grow in high concentrations of organic nutrients, and the concentrations of phosphorus at which the stress response is stimulated are considerably higher than those measured in oligiotrophic oceans (0.2 to 1 nM inorganic phosphate). Even in studies with the freshwater oligiotrophic bacterium Caulobacter crescentus, 30 μM of phosphate induced an adaptive response.12 In contrast, various strains of Rhizobium species, a nitrogen-fixing soil microbe, grew as rapidly at 0.05 μM phosphate as at 2 mM.13 The data on the effects of phosphate limitation on spore-forming microorganisms includes studies only with the mesophiles Bacillus subtilis and Clostridium perfringens. In B. subtilis, concentrations of phosphorus below 0.1 mM led to reduced growth rates and entry into stationary growth phase,14,15 whereas in C. perfringens, sporulation did not occur when phosphorus concentrations were less than 3 mM.16 There were no studies found that looked at the effects of phosphorus limitations on psychrophilic or psychrotolerant spore-forming bacteria.
The studies on phosphorus limitation using pure cultures of microorganisms have revealed the exquisite complexity of physiological responses and survival mechanisms. However, complex ecosystems exist in oligiotrophic environments where the concentration of phosphate is lower than the lowest concentration that either prevents growth or induces stress responses in most isolated microbes tested. Oligiotrophic oceans such as the Sargasso Sea in the northwestern Atlantic, the North Pacific subtropical gyre, and eastern part of the Mediterranean Sea have extremely low levels of dissolved phosphate. For example, the concentrations of phosphate in surface waters of the subtropical Sargasso Sea are from 0.2 to 1.0 nM.17,18,19 The canonical “Redfield ratio” used in biogeochemical models of the ocean is 106C:16N:1P and does not apply to oligiotrophic oceans where the N:P ratio can be higher than 30.20 Although these environments show limited phosphate stress, they have active ecosystems anchored by Prochlorococcus spp. as the primary producers. These photosynthetic bacteria are highly adapted to low levels of