Putative indigenous carbon-bearing alteration features in martian meteorite Yamato 000593.

We report the first observation of indigenous carbonaceous matter in the martian meteorite Yamato 000593. The carbonaceous phases are heterogeneously distributed within secondary iddingsite alteration veins and present in a range of morphologies including areas composed of carbon-rich spheroidal assemblages encased in multiple layers of iddingsite. We also observed microtubular features emanating from iddingsite veins penetrating into the host olivine comparable in shape to those interpreted to have formed by bioerosion in terrestrial basalts.

Toxicity of lunar dust assessed in inhalation-exposed rats.

Humans will again set foot on the moon. The moon is covered by a layer of fine dust, which can pose a respiratory hazard. We investigated the pulmonary toxicity of lunar dust in rats exposed to 0, 2.1, 6.8, 20.8 and 60.6 mg/m(3) of respirable-size lunar dust for 4 weeks (6 h/day, 5 days/week); the aerosols in the nose-only exposure chambers were generated from a jet-mill ground preparation of a lunar soil collected during the Apollo 14 mission. After 4 weeks of exposure to air or lunar dust, groups of five rats were euthanized 1 day, 1 week, 4 weeks or 13 weeks after the last exposure for assessment of pulmonary toxicity. Biomarkers of toxicity assessed in bronchoalveolar fluids showed concentration-dependent changes; biomarkers that showed treatment effects were total cell and neutrophil counts, total protein concentrations and cellular enzymes (lactate dehydrogenase, glutamyl transferase and aspartate transaminase). No statistically significant differences in these biomarkers were detected between rats exposed to air and those exposed to the two low concentrations of lunar dust. Dose-dependent histopathology, including inflammation, septal thickening, fibrosis and granulomas, in the lung was observed at the two higher exposure concentrations. No lesions were detected in rats exposed to ≤6.8 mg/m(3). This 4-week exposure study in rats showed that 6.8 mg/m(3) was the highest no-observable-adverse-effect level (NOAEL). These results will be useful for assessing the health risk to humans of exposure to lunar dust, establishing human exposure limits and guiding the design of dust mitigation systems in lunar landers or habitats.

Induction and construct UV protective yeast plasmid.

In this study, we apply concepts of synthetic biology in combination with conventional methods to assemble different genetic components to construct yeast resistant to UV radiation, and to induce production of anti-UV proteins. This work combines sequences of different promoters, STRESS-proteins, heat shock protein (HSP), kinase proteins, alcohol dehydrogenase protein (ADH), ribosomal binding sites, fluorescent reporter proteins, terminators, and a synthetic ribosomal switch. The aim of this investigation was to induce an anti-UV proteins, and to construct an anti-UV yeast plasmid to be used for protection of skin cells against UV radiation. This investigation demonstrates induction and construction of anti-UV genes and production of their corresponding proteins. Cultures of Saccharomyces cerevisiae (ATCC # 66348) were exposed to short-wave UV radiation and were then subjected to time-PCR to assess specific gene expression. Proteins were identified using two dimensional difference gel electrophoresis (2D DIGE) and LC-MS/MS. Different up-regulated and down-regulated proteins were identified. Highly expressed identified proteins were cloned into S. cerevisiae using a synthetic biology approach. Extracts from UV-induced genetically transformed yeasts were used to protect skin cell cultures (ATCC #2522-CRL) in vitro. Both microscopic analysis and an apoptosis assay showed protection of the skin cell cultures against UV radiation.

Direct detection of projectile relics from the end of the lunar basin-forming epoch.

The lunar surface, a key proxy for the early Earth, contains relics of asteroids and comets that have pummeled terrestrial planetary surfaces. Surviving fragments of projectiles in the lunar regolith provide a direct measure of the types and thus the sources of exogenous material delivered to the Earth-Moon system. In ancient [>3.4 billion years ago (Ga)] regolith breccias from the Apollo 16 landing site, we located mineral and lithologic relics of magnesian chondrules from chondritic impactors. These ancient impactor fragments are not nearly as diverse as those found in younger (3.4 Ga to today) regolith breccias and soils from the Moon or that presently fall as meteorites to Earth. This suggests that primitive chondritic asteroids, originating from a similar source region, were common Earth-Moon-crossing impactors during the latter stages of the basin-forming epoch.

Constructed molecular sensor to enhance metal detection by bacterial ribosomal switch-ion channel protein interaction.

Molecular biosensors are useful tools that detect metal ions or other potentially toxic chemicals. However, the efficiency of conventional sensors is limited in mixed metals substrates, which is the common way they are found in nature. The use of biosensors constructed from genetically modified living microbial systems has the potential of providing sensitive detection systems for specific toxic targets. Consequently, our investigation was aimed at assembling different genetic building blocks to produce a focused microbial biosensor with the ability to detect specific metals. This objective was achieved by using a synthetic biology approach. Our genetic building blocks, including a synchronized ribosomal switch-iron ion channel, along with sequences of promoters, metal-binding proteins (Fe, Pb), ribosomal binding sites, yellow fluorescence reporter protein (YFRP), and terminators, were constructed within the same biobrick in Escherichia coli. We used an rpoS ribosomal switch containing an aptamer, which responds to the specific metal ligands, in synchronization with an iron ion channel, TonB. This switch significantly stimulates translation, as expressed by higher fluorescence, number of colonies, and concentration of RNA in E. coli. The positive results show the effectiveness of using genetically tailored synchronized ribosomal switch-ion channels to construct microbial biosensors to detect specific metals, as tested in iron solutions.

Polyphasic characterization of a thermotolerant siderophilic filamentous cyanobacterium that produces intracellular iron deposits.

Despite the high potential for oxidative stress stimulated by reduced iron, contemporary iron-depositing hot springs with circum-neutral pH are intensively populated with cyanobacteria. Therefore, studies of the physiology, diversity, and phylogeny of cyanobacteria inhabiting iron-depositing hot springs may provide insights into the contribution of cyanobacteria to iron redox cycling in these environments and new mechanisms of oxidative stress mitigation. In this study the morphology, ultrastructure, physiology, and phylogeny of a novel cyanobacterial taxon, JSC-1, isolated from an iron-depositing hot spring, were determined. The JSC-1 strain has been deposited in ATCC under the name Marsacia ferruginose, accession number BAA-2121. Strain JSC-1 represents a new operational taxonomical unit (OTU) within Leptolyngbya sensu lato. Strain JSC-1 exhibited an unusually high ratio between photosystem (PS) I and PS II, was capable of complementary chromatic adaptation, and is apparently capable of nitrogen fixation. Furthermore, it synthesized a unique set of carotenoids, but only chlorophyll a. Strain JSC-1 not only required high levels of Fe for growth (≥40 µM), but it also accumulated large amounts of extracellular iron in the form of ferrihydrite and intracellular iron in the form of ferric phosphates. Collectively, these observations provide insights into the physiological strategies that might have allowed cyanobacteria to develop and proliferate in Fe-rich, circum-neutral environments.

Pathological calcification and replicating calcifying-nanoparticles: general approach and correlation.

Calcification, a phenomenon often regarded by pathologists little more than evidence of cell death, is becoming recognized to be important in the dynamics of a variety of diseases from which millions of beings suffer in all ages. In calcification, all that is needed for crystal formation to start is nidi (nuclei) and an environment of available dissolved components at or near saturation concentrations, along with the absence of inhibitors for crystal formation. Calcifying nanoparticles (CNP) are the first calcium phosphate mineral containing particles isolated from human blood and were detected in numerous pathologic calcification related diseases. Controversy and critical role of CNP as nidi and triggering factor in human pathologic calcification are discussed.

Do blood-borne calcifying nanoparticles self-propagate?

The nanotechnology industry is currently in the process of producing new nanoparticles. The biological activity of nanoparticles including adverse as well as beneficial effects tends to increase as their size decreases. The smaller the particles are, the greater their bioactivity and toxicity. Thus, one can easily conjecture the impact ofa nanoparticle if it could also self-replicate. This in vitro study reveals the self-propagating ability of unique calcifying nanoparticles (CNP) that can be as small as 50 nm in size and found in blood, blood products, and calcified soft tissues. Although specific detection techniques, morphological characteristics and biomineralizing properties of CNP are well established, their genomic information and self-propagating capability have always been challenged. The objective of this study is to document the propagation of CNP under physiological conditions, using inverted light microscopy (LM) and the Biostation IM time-lapse imaging system. Their detailed morphological structure was examined using scanning (SEM) and transmission (TEM) electron microscopy. This present study, in conjunction with previous findings of metabolic activity, antibiotic sensitivity, antibody specificity, morphological aspects and infectivity, validates CNP as self-replicators. Therefore these sterile-filterable, blood-borne nanoparticles should be of concern to the nanomedicine industry.

Association between Randall's plaque and calcifying nanoparticles.

OBJECTIVES: Randall initially described calcified subepithelial papillary plaques, which he hypothesized as nidi for urinary calculi. The discovery of calcifying nanoparticles (CNP), also referred to as nanobacteria, in calcified soft tissues has raised another hypothesis about their possible involvement in urinary stone formation. This research is the first attempt to investigate the potential association of these two hypotheses. METHODS: We collected renal papilla and blood samples from 17 human patients who had undergone laparoscopic nephrectomy. Immunohistochemical staining (IHS) was applied using monoclonal antibody (mAb) against CNP. Homogenized papillary tissues and serum samples were cultured for CNP. Scanning electron microscopy (SEM) and energy dispersive X-ray spectroscopy (EDS) were performed on papillary samples. Serum samples were tested for CNP antigen and antibody with enzyme-linked immunosorbent assay (ELISA). RESULTS: Randall's plaques (RP) were visible on gross inspection in 11 out of 17 samples. IHS was positive for CNP antigen in 8 of the visually positive samples, but in only 1 of the remaining samples. SEM revealed spherical apatite-formations in 14 samples confirmed by EDS analysis. In cultures, all serum samples and 13 tissue homogenates grew CNP. In ELISA, 14 samples were positive for CNP-antigen and 11 samples were positive for CNP-antibody. CONCLUSION: There was evidence of a link between detection of CNP and presence of RP. Although causality was not demonstrated, these results suggest that further studies with negative control samples should be made to explore the etiology of RP formation, thus leading to a better understanding of the pathogenesis of stone formation.

Biohazard potential of putative Martian organisms during missions to Mars.

Exploration Class missions to Mars will require precautions against potential contamination by any native microorganisms that may be incidentally pathogenic to humans. While the results of NASA's Viking biology experiments of the 1970s have been generally interpreted as inconclusive for surface organisms, and attributed to active but nonbiological chemistries, the possibility of native surface life has never been ruled out completely. It is possible that, prior to the first human landing on Mars, robotic craft and sample return missions will provide enough data to know with certainty whether future human landing sites harbor extant life forms. If native life were found to exist, it would be problematic to determine whether any of its species might present a medical danger to astronauts. Therefore, it will become necessary to assess empirically the risk that the planet contains pathogens based on terrestrial examples of pathogenicity and to take a reasonably cautious approach to biohazard protection. A survey of terrestrial pathogens was conducted with special emphasis on those whose evolution has not depended on the presence of animal hosts. The history of the development and implementation of Apollo anti-contamination protocol and recommendations of the National Research Council's Space Studies Board regarding Mars were reviewed. Organisms can emerge in Nature in the absence of indigenous animal hosts and both infectious and non-infectious human pathogens are therefore theoretically possible on Mars. Although remote, the prospect of Martian surface life, together with the existence of a diversity of routes by which pathogenicity has emerged on Earth, suggests that the probability of human pathogens on Mars, while low, is not zero. Still, since the discovery and study of Martian life can have long-term benefits for humanity, the risk that Martian life might include pathogens should not be an obstacle to human exploration. As a precaution, it is recommended that EVA (extravehicular activity) suits be decontaminated when astronauts enter surface habitats upon returning from field activity and that biosafety protocols approximating laboratory BSL 2 be developed for astronauts working in laboratories on the Martian surface. Quarantine of astronauts and Martian materials arriving on Earth should also be part of a human mission to Mars, and this and the surface biosafety program should be integral to human expeditions from the earliest stages of the mission planning.

Chemical mapping of proterozoic organic matter at submicron spatial resolution.

A NanoSIMS ion microprobe was used to map the submicron-scale distributions of carbon, nitrogen, sulfur, silicon, and oxygen in organic microfossils and laminae in a thin section of the approximately 0.85 billion year old Bitter Springs Formation of Australia. The data provide clues about the original chemistry of the microfossils, the silicification process, and the biosignatures of specific microorganisms and microbial communities. Chemical maps of fossil unicells and filaments revealed distinct wall- and sheath-like structures enriched in C, N, and S, consistent with their accepted biological origin. Surprisingly, organic laminae, previously considered to be amorphous, also exhibited filamentous and apparently compressed spheroidal structures defined by strong enrichments in C, N, and S. By analogy to NanoSIMS data from the well-preserved microfossils, these structures were interpreted as being of biological origin, most likely representing densely packed remnants of microbial mats. Given that the preponderance of organic matter in Precambrian sediments is similarly "amorphous," our findings indicate that a re-evaluation of ancient specimens via in situ structural, chemical, and isotopic study is warranted. Our analyses have led us to propose new criteria for assessing the biogenicity of problematic kerogenous materials, and, thus, these criteria can be applied to assessments of poorly preserved or fragmentary organic residues in early Archean sediments and any that might occur in meteorites or other extraterrestrial samples.

A potential cause for kidney stone formation during space flights: enhanced growth of nanobacteria in microgravity.

BACKGROUND: Although some information is available regarding the cellular/molecular changes in immune system exposed to microgravity, little is known about the reasons of the increase in the kidney stone formation in astronauts during and/or after long duration missions at zero gravity (0 g). In our earlier studies, we have assessed a unique agent, nanobacteria (NB), in kidney stones and hypothesized that NB have an active role in calcium phosphate-carbonate deposition in kidney. In this research we studied effect of microgravity on multiplication and calcification of NB in vitro. METHODS: We examined NB cultures in High Aspect Rotating Vessels (HARVs) designed at the NASA's Johnson Space Center, which are designed to stimulate some aspects of microgravity. Multiplication rate and calcium phosphate composition of those NB were compared with NB cultured on stationary and shaker flasks. Collected aliquots of the cultures from different incubation periods were analyzed using spectrophotometer, SEM, TEM, EDX, and x-ray diffraction techniques. RESULTS: The results showed that NB multiplied 4.6x faster in HARVs compared to stationary cultures, and 3.2x faster than shaker flask conditions. X-ray diffraction and EDX analysis showed that the degree of apatite crystal formation and the properties of the apatite depend on the specific culture conditions used. CONCLUSION: We now report an increased multiplication rate of NB in microgravity-simulated conditions. Thus, NB infection may have a potential role in kidney stone formation in crew members during space flights. For further proof to this hypothesis, screening of the NB antigen and antibody level in flight crew before and after flight would be necessary.

Comparison of antibody--antigen interactions on collagen measured by conventional immunological techniques and atomic force microscopy.

We have developed a means of using atomic force microscopy (AFM) to repeatedly localize a small area of interest (4 x 4 microm(2)) within a 0.5-cm(2) area on a heterogeneous sample, to obtain and localize high-resolution images and force measurements on nonideal samples (i.e., samples that better reflect actual biological systems, not prepared on atomically flat surfaces). We demonstrate the repeated localization and measurement of unbinding forces associated with antibody--antigen (ab--ag) interactions, by applying AFM in air and in liquid to visualize and measure polyclonal ab--ag interactions, using chicken collagen as a model system. We demonstrate that molecular interactions, in the form of ab--ag complexes, can be visualized by AFM when secondary antibodies are conjugated to 20-nm colloidal gold particles. We then compare those results with established immunological techniques, to demonstrate broader application of AFM technology to other systems. Data from AFM studies are compared with results obtained using immunological methods traditionally employed to investigate ab--ag interactions, including enzyme-linked immunosorbent assay, immunoblotting, and in situ immunofluorescence. Finally, using functionalized AFM tips with a flexible tether [poly(ethylene glycol) 800] to which a derivatized antibody was attached, we analyzed force curve data to measure the unbinding force of collagen antibody from its antigen, obtaining a value of approximately 90 +/- 40 pN with a MatLab code written to automate the analyses of force curves obtained in force--volume mode. The methodology we developed for embedded collagen sections can be readily applied to the investigation of other receptor--ligand interactions.

A preliminary investigation into light-modulated replication of nanobacteria and heart disease.

OBJECTIVE: The purpose of this preliminary study is to evaluate the effect of various wavelengths of light on nanobacteria (NB). BACKGROUND DATA: NB and mitochondria use light for biological processes. NB have been described as multifunctional primordial nanovesicles with the potential to utilize solar energy for replication. NB produce slime, a process common to living bacteria. Slime release is an evolutionary important stress-dependent phenomenon increasing the survival chance of individual bacteria in a colony. In the cardiovascular system, stress-induced bacterial colony formation may lead to a deposition of plaque. METHODS: Cultured NB were irradiated with NASA-LEDs at different wavelengths of light: 670, 728 and 880 nm. Light intensities were about 500k Wm(-2), and energy density was 1 x 10(4) J m(-2). RESULTS: Monochromatic light clearly affected replication of NB. Maximum replication was achieved at 670 nm. CONCLUSIONS: The results indicate that suitable wavelengths of light could be instrumental in elevating the vitality level of NB, preventing the production of NB-mediated slime, and simultaneously increasing the vitality level of mitochondria. The finding could stimulate the design of cooperative therapy concepts that could reduce death caused by myocardial infarcts.

Living nanovesicles--chemical and physical survival strategies of primordial biosystems.

Life on Earth and Mars could have started with self-assembled nanovesicles similar to the present nanobacteria (NB). To resist extreme environmental stress situations and periods of nutritional deprivation, nanovesicles would have had a chemical composition protected by a closed mineralized compartment, facilitating their development in a primordial soup, or other early wet environment. Their survivability would have been enhanced if they had mechanisms for metabolic communication, and an ability to collect primordially available energy forms. Here, we establish an irreducible model system for life formation starting with NB.

Earth's earliest biosphere-a proposal to develop a collection of curated archean geologic reference materials.

The discovery of evidence indicative of life in a Martian meteorite has led to an increase in interest in astrobiology. As a result of this discovery, and the ensuing controversy, it has become apparent that our knowledge of the early development of life on Earth is limited. Archean stratigraphic successions containing evidence of Earth's early biosphere are well preserved in the Pilbara Craton of Western Australia. The craton includes part of a protocontinent consisting of granitoid complexes that were emplaced into, and overlain by, a 3.51-2.94 Ga volcanigenic carapace - the Pilbara Supergroup. The craton is overlain by younger supracrustal basins that form a time series recording Earth history from approximately 2.8 Ga to approximately 1.9 Ga. It is proposed that a well-documented suite of these ancient rocks be collected as reference material for Archean and astrobiological research. All samples would be collected in a well-defined geological context in order to build a framework to test models for the early evolution of life on Earth and to develop protocols for the search for life on other planets.

Antibody binding in altered gravity: implications for immunosorbent assay during space flight.

A single antibody-incubation step of an indirect, enzyme-linked immunosorbent assay (ELISA) was performed during microgravity, Martian gravity (0.38 G) and hypergravity (1.8 G) phases of parabolic flight, onboard the NASA KC-135 aircraft. Antibody-antigen binding occurred within 15 seconds; the level of binding did not differ between microgravity, Martian gravity and 1 G (Earth's gravity) conditions. During hypergravity and 1 G, antibody binding was directly proportional to the fluid volume (per microtiter well) used for incubation; this pattern was not observed during microgravity. These effects in microgravity may be due to "fluid spread" within the chamber (observed during microgravity with digital photography), leading to greater fluid-surface contact and subsequently antibody-antigen contact. In summary, these results demonstrate that: i) ELISA antibody-incubation and washing steps can be successfully performed by human operators during microgravity, Martian gravity and hypergravity; ii) there is no significant difference in antibody binding between microgravity, Martian gravity and 1 G conditions; and iii) a smaller fluid volume/well (and therefore less antibody) was required for a given level of binding during microgravity. These conclusions indicate that reduced gravity would not present a barrier to successful operation of immunosorbent assays during spaceflight.