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Search indexed NASA NTRS and DOE OSTI research on propulsion, heat transfer, battery materials and energy systems. Follow report and document links to the original sources.

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Direct profiling of environmental microbial populations by thermal dissociation analysis of native rRNAs hybridized to oligonucleotide microarrays

Oligonucleotide microarrays were used to profile directly extracted rRNA from environmental microbial populations without PCR amplification. In our initial inspection of two distinct estuarine study sites, the hybridization patterns were reproducible and varied between estuarine sediments of differing salinities. The determination of a thermal dissociation curve (i.e., melting profile) for each probe-target duplex provided information on hybridization specificity, which is essential for confirming adequate discrimination between target and nontarget sequences.

Non-NASA Center↗

The origin and evolution of model organisms

The phylogeny and timescale of life are becoming better understood as the analysis of genomic data from model organisms continues to grow. As a result, discoveries are being made about the early history of life and the origin and development of complex multicellular life. This emerging comparative framework and the emphasis on historical patterns is helping to bridge barriers among organism-based research communities.

Review, Academic↗

Archaeal translation initiation revisited: the initiation factor 2 and eukaryotic initiation factor 2B alpha-beta-delta subunit families

As the amount of available sequence data increases, it becomes apparent that our understanding of translation initiation is far from comprehensive and that prior conclusions concerning the origin of the process are wrong. Contrary to earlier conclusions, key elements of translation initiation originated at the Universal Ancestor stage, for homologous counterparts exist in all three primary taxa. Herein, we explore the evolutionary relationships among the components of bacterial initiation factor 2 (IF-2) and eukaryotic IF-2 (eIF-2)/eIF-2B, i.e., the initiation factors involved in introducing the initiator tRNA into the translation mechanism and performing the first step in the peptide chain elongation cycle. All Archaea appear to posses a fully functional eIF-2 molecule, but they lack the associated GTP recycling function, eIF-2B (a five-subunit molecule). Yet, the Archaea do posses members of the gene family defined by the (related) eIF-2B subunits alpha, beta, and delta, although these are not specifically related to any of the three eukaryotic subunits. Additional members of this family also occur in some (but by no means all) Bacteria and even in some eukaryotes. The functional significance of the other members of this family is unclear and requires experimental resolution. Similarly, the occurrence of bacterial IF-2-like molecules in all Archaea and in some eukaryotes further complicates the picture of translation initiation. Overall, these data lend further support to the suggestion that the rudiments of translation initiation were present at the Universal Ancestor stage.

NASA Discipline Exobiology↗

Determining divergence times with a protein clock: update and reevaluation

A recent study of the divergence times of the major groups of organisms as gauged by amino acid sequence comparison has been expanded and the data have been reanalyzed with a distance measure that corrects for both constraints on amino acid interchange and variation in substitution rate at different sites. Beyond that, the availability of complete genome sequences for several eubacteria and an archaebacterium has had a great impact on the interpretation of certain aspects of the data. Thus, the majority of the archaebacterial sequences are not consistent with currently accepted views of the Tree of Life which cluster the archaebacteria with eukaryotes. Instead, they are either outliers or mixed in with eubacterial orthologs. The simplest resolution of the problem is to postulate that many of these sequences were carried into eukaryotes by early eubacterial endosymbionts about 2 billion years ago, only very shortly after or even coincident with the divergence of eukaryotes and archaebacteria. The strong resemblances of these same enzymes among the major eubacterial groups suggest that the cyanobacteria and Gram-positive and Gram-negative eubacteria also diverged at about this same time, whereas the much greater differences between archaebacterial and eubacterial sequences indicate these two groups may have diverged between 3 and 4 billion years ago.

NASA Discipline Exobiology↗

Whither microbiology? Phylogenetic trees

The direct cloning and sequencing of genes from uncultured microorganisms in a hot spring suggests that the diversity of life on Earth may be far in excess of that estimated from culturable species.

NASA Discipline Exobiology↗

Origins of species: acquired genomes and individuality

Entire genomes with their accompanying protein synthetic systems are transferred throughout the biosphere primarily as bacteria and protists which become symbionts as they irreversibly integrate into pre-existing organisms to form more complex individuals. Individualization is stabilized by simultaneous transmission of once-separate heterologous genetic systems. The origin of new species is hypothesized to correlate with the acquisition, integration and subsequent inheritance of such acquired microbial genomes. These processes were recognized by Mereschkovsky ("Symbiogenesis" in Russian, 1909) and by Wallin ("Symbionticism", see p. 181, this issue).

NASA Discipline Exobiology↗

Chromosome map of the thermophilic archaebacterium Thermococcus celer

A physical map for the chromosome of the thermophilic archaebacterium Thermococcus celer Vu13 has been constructed. Thirty-four restriction endonucleases were tested for their ability to generate large restriction fragments from the chromosome of T. celer. Of these, the enzymes NheI, SpeI, and XbaI yielded the fewest fragments when analyzed by pulsed-field electrophoresis. NheI and SpeI each gave 5 fragments, while XbaI gave 12. The size of the T. celer chromosome was determined from the sum of the apparent sizes of restriction fragments derived from single and double digests by using these enzymes and was found to be 1,890 +/- 27 kilobase pairs. Partial and complete digests allowed the order of all but three small (less than 15 kilobase pairs) fragments to be deduced. These three fragments were assigned positions by using hybridization probes derived from these restriction fragments. The positions of the other fragments were confirmed by using hybridization probes derived in the same manner. The positions of the 5S, 16S, and 23S rRNA genes as well as the 7S RNA gene were located on this map by using cloned portions of these genes as hybridization probes. The 5S rRNA gene was localized 48 to 196 kilobases from the 5' end of the 16S gene. The 7S RNA gene was localized 190 to 504 kilobases from the 3' end of the 23S gene. These analyses demonstrated that the chromosome of T. celer is a single, circular DNA molecule. This is the first such demonstration of the structure of an archaebacterial chromosome.

Non-NASA Center↗

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↗

Microbial Characterization During the Early Habitation of the International Space Station

An evaluation of the microbiota from air, water, and surface samples provided a baseline of microbial characterization onboard the International Space Station (ISS) to gain insight into bacterial and fungal contamination during the initial stages of construction and habitation. Using 16S genetic sequencing and rep-PCR, 63 bacterial strains were isolated for identification and fingerprinted for microbial tracking. Of the bacterial strains that were isolated and fingerprinted, 19 displayed similarity to each other. The use of these molecular tools allowed for the identification of bacteria not previously identified using automated biochemical analysis and provided a clear indication of the source of several ISS contaminants. Strains of Bradyrhizobium and Sphingomonas unable to be identified using sequencing were identified by comparison of rep-PCR DNA fingerprints. Distinct DNA fingerprints for several strains of Methylobacterium provided a clear indication of the source of an ISS water supply contaminant. Fungal and bacterial data acquired during monitoring do not suggest there is a current microbial hazard to the spacecraft, nor does any trend indicate a potential health risk. Previous spacecraft environmental analysis indicated that microbial contamination will increase with time and will require continued surveillance. Copyright 2004 Springer-Verlag.

Fungi/genetics↗

Microbial monitoring of spacecraft and associated environments

Rapid microbial monitoring technologies are invaluable in assessing contamination of spacecraft and associated environments. Universal and widespread elements of microbial structure and chemistry are logical targets for assessing microbial burden. Several biomarkers such as ATP, LPS, and DNA (ribosomal or spore-specific), were targeted to quantify either total bioburden or specific types of microbial contamination. The findings of these assays were compared with conventional, culture-dependent methods. This review evaluates the applicability and efficacy of some of these methods in monitoring the microbial burden of spacecraft and associated environments. Samples were collected from the surfaces of spacecraft, from surfaces of assembly facilities, and from drinking water reservoirs aboard the International Space Station (ISS). Culture-dependent techniques found species of Bacillus to be dominant on these surfaces. In contrast, rapid, culture-independent techniques revealed the presence of many Gram-positive and Gram-negative microorganisms, as well as actinomycetes and fungi. These included both cultivable and noncultivable microbes, findings further confirmed by DNA-based microbial detection techniques. Although the ISS drinking water was devoid of cultivable microbes, molecular-based techniques retrieved DNA sequences of numerous opportunistic pathogens. Each of the methods tested in this study has its advantages, and by coupling two or more of these techniques even more reliable information as to microbial burden is rapidly obtained. Copyright 2004 Springer-Verlag.

Environmental Monitoring/methods↗

Microbial Characterization of Free Floating Condensate Aboard the Mir Space Station

Three samples of humidity condensate that had accumulated behind panels aboard the Russian space station Mir were collected and returned to earth for analysis. As these floating masses of liquid come into contact with the astronauts and the engineering systems, they have the potential to affect both crew health and systems performance. Using a combination of culturing techniques, a wide variety of organisms were isolated included Escherichia coli, Serratia marcescens, and a presumed Legionella species. In addition, microscopic analysis indicated the presence of protozoa, dust mites, and spirochetes. These findings suggest the need for more comprehensive microbial analysis of the environment through the use of new methodologies to allow a more thorough risk assessment of spacecraft. Copyright 2004 Springer-Verlag.

Spacecraft↗

A genomic timescale of prokaryote evolution: insights into the origin of methanogenesis, phototrophy, and the colonization of land

BACKGROUND: The timescale of prokaryote evolution has been difficult to reconstruct because of a limited fossil record and complexities associated with molecular clocks and deep divergences. However, the relatively large number of genome sequences currently available has provided a better opportunity to control for potential biases such as horizontal gene transfer and rate differences among lineages. We assembled a data set of sequences from 32 proteins (approximately 7600 amino acids) common to 72 species and estimated phylogenetic relationships and divergence times with a local clock method. RESULTS: Our phylogenetic results support most of the currently recognized higher-level groupings of prokaryotes. Of particular interest is a well-supported group of three major lineages of eubacteria (Actinobacteria, Deinococcus, and Cyanobacteria) that we call Terrabacteria and associate with an early colonization of land. Divergence time estimates for the major groups of eubacteria are between 2.5-3.2 billion years ago (Ga) while those for archaebacteria are mostly between 3.1-4.1 Ga. The time estimates suggest a Hadean origin of life (prior to 4.1 Ga), an early origin of methanogenesis (3.8-4.1 Ga), an origin of anaerobic methanotrophy after 3.1 Ga, an origin of phototrophy prior to 3.2 Ga, an early colonization of land 2.8-3.1 Ga, and an origin of aerobic methanotrophy 2.5-2.8 Ga. CONCLUSIONS: Our early time estimates for methanogenesis support the consideration of methane, in addition to carbon dioxide, as a greenhouse gas responsible for the early warming of the Earths' surface. Our divergence times for the origin of anaerobic methanotrophy are compatible with highly depleted carbon isotopic values found in rocks dated 2.8-2.6 Ga. An early origin of phototrophy is consistent with the earliest bacterial mats and structures identified as stromatolites, but a 2.6 Ga origin of cyanobacteria suggests that those Archean structures, if biologically produced, were made by anoxygenic photosynthesizers. The resistance to desiccation of Terrabacteria and their elaboration of photoprotective compounds suggests that the common ancestor of this group inhabited land. If true, then oxygenic photosynthesis may owe its origin to terrestrial adaptations.

Methane/metabolism↗

Microbial structural diversity estimated by dilution-extinction of phenotypic traits and T-RFLP analysis along a land-use intensification gradient

The present work tested whether the relationship between functional traits and inoculum density reflected structural diversity in bacterial communities from a land-use intensification gradient applying a mathematical model. Terminal restriction fragment length polymorphism (T-RFLP) analysis was also performed to provide an independent assessment of species richness. Successive 10-fold dilutions of a soil suspension were inoculated onto Biolog GN(R) microplates. Soil bacterial density was determined by total cell and plate counts. The relationship between phenotypic traits and inoculum density fit the model, allowing the estimation of maximal phenotypic potential (Rmax) and inoculum density (KI) at which Rmax will be half-reduced. Though Rmax decreased with time elapsed since clearing of native vegetation, KI remained high in two of the disturbed sites. The genetic pool of bacterial community did not experience a significant reduction, but the active fraction responding in the Biolog assay was adversely affected, suggesting a reduction in the functional potential. c2004 Federation of European Microbiological Societies. Published by Elsevier B.V. All rights reserved.

NASA Center KSC↗