Anne Dekas is a geomicrobiologist interested in how microbial life affects the chemistry and climate of our planet today and throughout time. She obtained her PhD in Geobiology at the California Institute of Technology in 2013, and her AB in Earth and Planetary Sciences at Harvard University in 2004. She joined the Earth System Science Department at Stanford University as an assistant professor in September 2015.
Assistant Professor, Earth System Science, Stanford University (2015 - Present)
Lawrence Postdoctoral Fellow, Lawrence Livermore National Laboratory (2013 - 2015)
NSF Graduate Research Fellow, Geological and Planetary Sciences, California Institute of Technology (2006 - 2009)
Technical Assistant, Planetary Protection Group, Jet Propulsion Laboratory (2005 - 2006)
Honors & Awards
Early Career Investigator in Marine Microbial Ecology and Evolution Award, Simons Foundation (2017-2020)
Outstanding Poster Presentation Award, Lawrence Livermore National Laboratory Poster Symposium (2014)
Lawrence Postdoctoral Fellowship, Lawrence Livermore National Laboratory (2013-2015)
Outstanding Oral Presentation, Southern California Geobiology Symposium (2009)
Tech Brief Award, NASA Jet Propulsion Laboratory (2009)
Graduate Research Fellowship, National Science Foundation (2006-2009)
Harvard College Award, Harvard University (2002, 2003)
John Harvard Award, Harvard University (2001)
Boards, Advisory Committees, Professional Organizations
Member, American Geophysical Union (2008 - Present)
Member, International Society for Microbial Ecology (2008 - Present)
PhD, California Institute of Technology, Geological and Planetary Sciences, Geobiology (2013)
AB, Harvard University, Earth and Planetary Sciences, Biogeochemistry (2004)
Current Research and Scholarly Interests
Environmental microbiology, deep-sea microbial ecology, marine biogeochemistry
- Fundamentals of Geobiology
EARTHSYS 205A, EPS 205, ESS 205 (Aut)
- Life at the Extremes: From the Deep Sea to Deep Space
EARTHSYS 36N (Win)
Independent Studies (1)
- Graduate Research
ESS 400 (Aut, Win, Spr, Sum)
- Graduate Research
Prior Year Courses
- Microbes and Climate
BIO 105, BIO 205, ESS 122, ESS 222 (Aut)
- Life at the Extremes: From the Deep Sea to Deep Space
EARTHSYS 36N (Win)
- Techniques in Environmental Microbiology
BIO 201, ESS 210 (Aut)
- Microbes and Climate
Doctoral Dissertation Reader (AC)
Stephanie Lim, Hanon McShea
Postdoctoral Faculty Sponsor
Chloe Baumas, Steffen Buessecker, Alex Jaffe
Doctoral Dissertation Advisor (AC)
Master's Program Advisor
Kate Bradley, Mikaela Salvador
Emily Paris, Rebecca Salcedo, Amanda Semler
Autochthonous carbon loading of macroalgae stimulates benthic biological nitrogen fixation rates in shallow coastal marine sediments
Frontiers in Microbiology
View details for DOI 10.3389/fmicb.2023.1312843
Single-cell analysis in hypersaline brines predicts a water-activity limit of microbial anabolic activity.
2023; 9 (51): eadj3594
Hypersaline brines provide excellent opportunities to study extreme microbial life. Here, we investigated anabolic activity in nearly 6000 individual cells from solar saltern sites with water activities (aw) ranging from 0.982 to 0.409 (seawater to extreme brine). Average anabolic activity decreased exponentially with aw, with nuanced trends evident at the single-cell level: The proportion of active cells remained high (>50%) even after NaCl saturation, and subsets of cells spiked in activity as aw decreased. Intracommunity heterogeneity in activity increased as seawater transitioned to brine, suggesting increased phenotypic heterogeneity with increased physiological stress. No microbial activity was detected in the 0.409-aw brine (an MgCl2-dominated site) despite the presence of cell-like structures. Extrapolating our data, we predict an aw limit for detectable anabolic activity of 0.540, which is beyond the currently accepted limit of life based on cell division. This work demonstrates the utility of single-cell, metabolism-based techniques for detecting active life and expands the potential habitable space on Earth and beyond.
View details for DOI 10.1126/sciadv.adj3594
View details for PubMedID 38134283
View details for PubMedCentralID PMC10745694
Microbially induced precipitation of silica by anaerobic methane-oxidizing consortia and implications for microbial fossil preservation.
Proceedings of the National Academy of Sciences of the United States of America
2023; 120 (51): e2302156120
Authigenic carbonate minerals can preserve biosignatures of microbial anaerobic oxidation of methane (AOM) in the rock record. It is not currently known whether the microorganisms that mediate sulfate-coupled AOM-often occurring as multicelled consortia of anaerobic methanotrophic archaea (ANME) and sulfate-reducing bacteria (SRB)-are preserved as microfossils. Electron microscopy of ANME-SRB consortia in methane seep sediments has shown that these microorganisms can be associated with silicate minerals such as clays [Chen et al., Sci. Rep. 4, 1-9 (2014)], but the biogenicity of these phases, their geochemical composition, and their potential preservation in the rock record is poorly constrained. Long-term laboratory AOM enrichment cultures in sediment-free artificial seawater [Yu et al., Appl. Environ. Microbiol. 88, e02109-21 (2022)] resulted in precipitation of amorphous silicate particles (~200 nm) within clusters of exopolymer-rich AOM consortia from media undersaturated with respect to silica, suggestive of a microbially mediated process. The use of techniques like correlative fluorescence in situ hybridization (FISH), scanning electron microscopy with energy dispersive X-ray spectroscopy (SEM-EDS), and nanoscale secondary ion mass spectrometry (nanoSIMS) on AOM consortia from methane seep authigenic carbonates and sediments further revealed that they are enveloped in a silica-rich phase similar to the mineral phase on ANME-SRB consortia in enrichment cultures. Like in cyanobacteria [Moore et al., Geology 48, 862-866 (2020)], the Si-rich phases on ANME-SRB consortia identified here may enhance their preservation as microfossils. The morphology of these silica-rich precipitates, consistent with amorphous-type clay-like spheroids formed within organic assemblages, provides an additional mineralogical signature that may assist in the search for structural remnants of microbial consortia in rocks which formed in methane-rich environments from Earth and other planetary bodies.
View details for DOI 10.1073/pnas.2302156120
View details for PubMedID 38079551
Mcr-dependent methanogenesis in Archaeoglobaceae enriched from a terrestrial hot spring.
The ISME journal
The preeminent source of biological methane on Earth is methyl coenzyme M reductase (Mcr)-dependent archaeal methanogenesis. A growing body of evidence suggests a diversity of archaea possess Mcr, although experimental validation of hypothesized methane metabolisms has been missing. Here, we provide evidence of a functional Mcr-based methanogenesis pathway in a novel member of the family Archaeoglobaceae, designated Methanoglobus nevadensis, which we enriched from a terrestrial hot spring on the polysaccharide xyloglucan. Our incubation assays demonstrate methane production that is highly sensitive to the Mcr inhibitor bromoethanesulfonate, stimulated by xyloglucan and xyloglucan-derived sugars, concomitant with the consumption of molecular hydrogen, and causing a deuterium fractionation in methane characteristic of hydrogenotrophic and methylotrophic methanogens. Combined with the recovery and analysis of a high-quality M. nevadensis metagenome-assembled genome encoding a divergent Mcr and diverse potential electron and carbon transfer pathways, our observations suggest methanogenesis in M. nevadensis occurs via Mcr and is fueled by the consumption of cross-fed byproducts of xyloglucan fermentation mediated by other community members. Phylogenetic analysis shows close affiliation of the M. nevadensis Mcr with those from Korarchaeota, Nezhaarchaeota, Verstraetearchaeota, and other Archaeoglobales that are divergent from well-characterized Mcr. We propose these archaea likely also use functional Mcr complexes to generate methane on the basis of our experimental validation in M. nevadensis. Thus, divergent Mcr-encoding archaea may be underestimated sources of biological methane in terrestrial and marine hydrothermal environments.
View details for DOI 10.1038/s41396-023-01472-3
View details for PubMedID 37452096
View details for PubMedCentralID 5543129
Constraining the composition and quantity of organic matter used by abundant marine Thaumarchaeota.
Marine Group I (MGI) Thaumarchaeota were originally described as chemoautotrophic nitrifiers, but molecular and isotopic evidence suggests heterotrophic and/or mixotrophic capabilities. Here, we investigated the quantity and composition of organic matter assimilated by individual, uncultured MGI cells from the Pacific Ocean to constrain their potential for mixotrophy and heterotrophy. We observed that most MGI cells did not assimilate carbon from any organic substrate provided (glucose, pyruvate, oxaloacetate, protein, urea, and amino acids). The minority of MGI cells that did assimilate it did so exclusively from nitrogenous substrates (urea, 15% of MGI; amino acids, 36% of MGI), and only as an auxiliary carbon source (<20% of that subset's total cellular carbon was derived from those substrates). At the population level, MGI assimilation of organic carbon comprised just 0.5-11% of total carbon. We observed extensive assimilation of inorganic carbon and urea- and amino acid-derived nitrogen (equal to that from ammonium), consistent with metagenomic and metatranscriptomic analyses performed here and previously showing a widespread potential for MGI to perform autotrophy and transport and degrade organic nitrogen. Our results constrain the quantity and composition of organic matter used by MGI and suggest they use it primarily to meet nitrogen demands for anabolism and nitrification. This article is protected by copyright. All rights reserved.
View details for DOI 10.1111/1462-2920.16299
View details for PubMedID 36478085
- Contributions of anoxic microsites to soil carbon protection across soil textures GEODERMA 2022; 425
Single-cell view of deep-sea microbial activity and intracommunity heterogeneity
Microbial activity in the deep sea is cumulatively important for global elemental cycling yet is difficult to quantify and characterize due to low cell density and slow growth. Here, we investigated microbial activity off the California coast, 50-4000 m water depth, using sensitive single-cell measurements of stable-isotope uptake and nucleic acid sequencing. We observed the highest yet reported proportion of active cells in the bathypelagic (up to 78%) and calculated that deep-sea cells (200-4000 m) are responsible for up to 34% of total microbial biomass synthesis in the water column. More cells assimilated nitrogen derived from amino acids than ammonium, and at higher rates. Nitrogen was assimilated preferentially to carbon from amino acids in surface waters, while the reverse was true at depth. We introduce and apply the Gini coefficient, an established equality metric in economics, to quantify intracommunity heterogeneity in microbial anabolic activity. We found that heterogeneity increased with water depth, suggesting a minority of cells contribute disproportionately to total activity in the deep sea. This observation was supported by higher RNA/DNA ratios for low abundance taxa at depth. Intracommunity activity heterogeneity is a fundamental and rarely measured ecosystem parameter and may have implications for community function and resilience.
View details for DOI 10.1038/s41396-022-01324-6
View details for Web of Science ID 000864589800001
View details for PubMedID 36202927
- Comparison of Microbial Profiling and Tracer Testing for the Characterization of Injector-Producer Interwell Connectivities WATER 2022; 14 (18)
Rates and physicochemical drivers of microbial anabolic activity in deep-sea sediments and implications for deep time.
Sediment microorganisms influence global climate and redox by altering rates of organic carbon burial. However, the activity and ecology of benthic microorganisms are poorly characterized, especially in the deep sea. Here, we conducted nearly 300 stable isotope tracer experiments in sediments from the Pacific and Atlantic Oceans (100-4500 m water depth) to determine the rates, spatial distribution, and physicochemical controls on microbial total anabolic activity, nitrogen fixation, and inorganic/organic carbon uptake. Using correlative and manipulative approaches, we find that total activity is limited primarily by organic carbon and/or energy. Activity correlates significantly with distance from shore, sediment depth, C:N ratios, and overlying chlorophyll concentrations, and is stimulated by carbon but not nitrogen additions. Consistent with this, nitrogen fixation was undetected despite relatively low concentrations of porewater ammonium and the previous detection of nifH genes. Inorganic carbon uptake accounted for 7-55% of carbon assimilation per sample (median 21%), suggesting chemoautotrophy is an important and unappreciated source of labile carbon in deep-sea sediments. Community 16S rRNA was dominated by Bacteria (<2% Archaea), primarily Desulfobacterales of the Deltaproteobacteria. Leveraging our findings, we modeled global benthic microbial activity through geologic time and find the potential for significant shifts in total activity with supercontinental cycles. This article is protected by copyright. All rights reserved.
View details for DOI 10.1111/1462-2920.16183
View details for PubMedID 36054699
An essential role for tungsten in the ecology and evolution of a previously uncultivated lineage of anaerobic, thermophilic Archaea.
2022; 13 (1): 3773
Trace metals have been an important ingredient for life throughout Earth's history. Here, we describe the genome-guided cultivation of a member of the elusive archaeal lineage Caldarchaeales (syn. Aigarchaeota), Wolframiiraptor gerlachensis, and its growth dependence on tungsten. A metagenome-assembled genome (MAG) of W. gerlachensis encodes putative tungsten membrane transport systems, as well as pathways for anaerobic oxidation of sugars probably mediated by tungsten-dependent ferredoxin oxidoreductases that are expressed during growth. Catalyzed reporter deposition-fluorescence in-situ hybridization (CARD-FISH) and nanoscale secondary ion mass spectrometry (nanoSIMS) show that W. gerlachensis preferentially assimilates xylose. Phylogenetic analyses of 78 high-quality Wolframiiraptoraceae MAGs from terrestrial and marine hydrothermal systems suggest that tungsten-associated enzymes were present in the last common ancestor of extant Wolframiiraptoraceae. Our observations imply a crucial role for tungsten-dependent metabolism in the origin and evolution of this lineage, and hint at a relic metabolic dependence on this trace metal in early anaerobic thermophiles.
View details for DOI 10.1038/s41467-022-31452-8
View details for PubMedID 35773279
Geological activity shapes the microbiome in deep-subsurface aquifers by advection.
Proceedings of the National Academy of Sciences of the United States of America
2022; 119 (25): e2113985119
Subsurface environments host diverse microorganisms in fluid-filled fractures; however, little is known about how geological and hydrological processes shape the subterranean biosphere. Here, we sampled three flowing boreholes weekly for 10 mo in a 1478-m-deep fractured rock aquifer to study the role of fracture activity (defined as seismically or aseismically induced fracture aperture change) and advection on fluid-associated microbial community composition. We found that despite a largely stable deep-subsurface fluid microbiome, drastic community-level shifts occurred after events signifying physical changes in the permeable fracture network. The community-level shifts include the emergence of microbial families from undetected to over 50% relative abundance, as well as the replacement of the community in one borehole by the earlier community from a different borehole. Null-model analysis indicates that the observed spatial and temporal community turnover was primarily driven by stochastic processes (as opposed to deterministic processes). We, therefore, conclude that the observed community-level shifts resulted from the physical transport of distinct microbial communities from other fracture(s) that outpaced environmental selection. Given that geological activity is a major cause of fracture activity and that geological activity is ubiquitous across space and time on Earth, our findings suggest that advection induced by geological activity is a general mechanism shaping the microbial biogeography and diversity in deep-subsurface habitats across the globe.
View details for DOI 10.1073/pnas.2113985119
View details for PubMedID 35696589
Cold Seeps on the Passive Northern U.S. Atlantic Margin Host Globally Representative Members of the Seep Microbiome with Locally Dominant Strains of Archaea.
Applied and environmental microbiology
Marine cold seeps are natural sites of methane emission and harbor distinct microbial communities capable of oxidizing methane. The majority of known cold seeps are on tectonically active continental margins, but recent discoveries have revealed abundant seeps on passive margins as well, including on the U.S. Atlantic Margin (USAM). We sampled in and around four USAM seeps and combined pore water geochemistry measurements with amplicon sequencing of 16S rRNA and mcrA (DNA and RNA) to investigate the microbial communities present, their assembly processes, and how they compare to communities at previously studied sites. We found that the USAM seeps contained communities consistent with the canonical seep microbiome at the class and order levels but differed markedly at the sequence variant level, especially within the anaerobic methanotrophic (ANME) archaea. The ANME populations were highly uneven, with just a few dominant mcrA sequence variants at each seep. Interestingly, the USAM seeps did not form a distinct phylogenetic cluster when compared with other previously described seeps around the world. Consistent with this, we found only a very weak (though statistically significant) distance-decay trend in seep community similarity across a global data set. Ecological assembly indices suggest that the USAM seep communities were assembled primarily deterministically, in contrast to the surrounding nonseep sediments, where stochastic processes dominated. Together, our results suggest that the primary driver of seep microbial community composition is local geochemistry-specifically methane, sulfide, nitrate, acetate, and ammonium concentrations-rather than the geologic context, the composition of nearby seeps, or random events of dispersal. IMPORTANCE Cold seeps are now known to be widespread features of passive continental margins, including the northern U.S. Atlantic Margin (USAM). Methane seepage is expected to intensify at these relatively shallow seeps as bottom waters warm and underlying methane hydrates dissociate. While methanotrophic microbial communities might reduce or prevent methane release, microbial communities on passive margins have rarely been characterized. In this study, we investigated the Bacteria and Archaea at four cold seeps on the northern USAM and found that despite being colocated on the same continental slope, the communities significantly differ by site at the sequence variant level, particularly methane-cycling community members. Differentiation by site was not observed in similarly spaced background sediments, raising interesting questions about the dispersal pathways of cold seep microorganisms. Understanding the genetic makeup of these discrete seafloor ecosystems and how their microbial communities develop will be increasingly important as the climate changes.
View details for DOI 10.1128/aem.00468-22
View details for PubMedID 35607968
Quantifying Microbial Activity In Situ: the Link between Cells and Cycles.
Metagenomic sequencing of environmental samples has dramatically expanded our knowledge of microbial taxonomic and metabolic diversity and suggests metabolic interdependence is widespread. However, translating these insights into knowledge of ecosystem function and, therefore, implications for local and global chemistry, remains a challenge. In this commentary, I argue that making direct measurements of microbial activity in situ is an essential step to confirm gene-based hypotheses of microbial physiology and bridge advances in microbial ecology with a predicative understanding of global chemistry and climate. Making these measurements across a range of spatial scales and experimentally manipulated conditions contributes to a process-based understanding and, therefore, more robust predictions of how activity will respond to changing environmental conditions. I discuss recent advancements in quantifying microbial activity in situ and highlight several lines of research in marine microbiology that leverage complementary genomic and isotopic methods to connect microbes and global chemistry.
View details for DOI 10.1128/mSystems.00758-21
View details for PubMedID 34463583
Characterizing the "fungal shunt": Parasitic fungi on diatoms affect carbon flow and bacterial communities in aquatic microbial food webs.
Proceedings of the National Academy of Sciences of the United States of America
2021; 118 (23)
Microbial interactions in aquatic environments profoundly affect global biogeochemical cycles, but the role of microparasites has been largely overlooked. Using a model pathosystem, we studied hitherto cryptic interactions between microparasitic fungi (chytrid Rhizophydiales), their diatom host Asterionella, and cell-associated and free-living bacteria. We analyzed the effect of fungal infections on microbial abundances, bacterial taxonomy, cell-to-cell carbon transfer, and cell-specific nitrate-based growth using microscopy (e.g., fluorescence in situ hybridization), 16S rRNA gene amplicon sequencing, and secondary ion mass spectrometry. Bacterial abundances were 2 to 4 times higher on individual fungal-infected diatoms compared to healthy diatoms, particularly involving Burkholderiales. Furthermore, taxonomic compositions of both diatom-associated and free-living bacteria were significantly different between noninfected and fungal-infected cocultures. The fungal microparasite, including diatom-associated sporangia and free-swimming zoospores, derived 100% of their carbon content from the diatom. By comparison, transfer efficiencies of photosynthetic carbon were lower to diatom-associated bacteria (67 to 98%), with a high cell-to-cell variability, and even lower to free-living bacteria (32%). Likewise, nitrate-based growth for the diatom and fungi was synchronized and faster than for diatom-associated and free-living bacteria. In a natural lacustrine system, where infection prevalence reached 54%, we calculated that 20% of the total diatom-derived photosynthetic carbon was shunted to the parasitic fungi, which can be grazed by zooplankton, thereby accelerating carbon transfer to higher trophic levels and bypassing the microbial loop. The herein termed "fungal shunt" can thus significantly modify the fate of photosynthetic carbon and the nature of phytoplankton-bacteria interactions, with implications for diverse pelagic food webs and global biogeochemical cycles.
View details for DOI 10.1073/pnas.2102225118
View details for PubMedID 34074785
Microbial diversity and activity in Southern California salterns and bitterns: analogues for remnant ocean worlds.
Concurrent osmotic and chaotropic stress make MgCl2 -rich brines extremely inhospitable environments. Understanding the limits of life in these brines is essential to the search for extraterrestrial life on contemporary and relict ocean worlds, like Mars, which could host similar environments. We sequenced environmental 16S rRNA genes and quantified microbial activity across a broad range of salinity and chaotropicity at a Mars-analogue salt harvesting facility in Southern California, where seawater is evaporated in a series of ponds ranging from kosmotropic NaCl brines to highly chaotropic MgCl2 brines. Within NaCl brines, we observed a proliferation of specialized halophilic Euryarchaeota, which corresponded closely with the dominant taxa found in salterns around the world. These communities were characterized by very slow growth rates and high biomass accumulation. As salinity and chaotropicity increased, we found that the MgCl2 -rich brines eventually exceeded the limits of microbial activity. We found evidence that exogenous genetic material is preserved in these chaotropic brines, producing an unexpected increase in diversity in the presumably-sterile MgCl2 -saturated brines. Because of their high potential for biomarker preservation, chaotropic brines could therefore serve as repositories of genetic biomarkers from nearby environments-both on Earth and beyond-making them prime targets for future life detection missions. This article is protected by copyright. All rights reserved.
View details for DOI 10.1111/1462-2920.15440
View details for PubMedID 33621409
PPIT: an R package for inferring microbial taxonomy from nifH sequences.
Bioinformatics (Oxford, England)
MOTIVATION: Linking microbial community members to their ecological functions is a central goal of environmental microbiology. When assigned taxonomy, amplicon sequences of metabolic marker genes can suggest such links, thereby offering an overview of the phylogenetic structure underpinning particular ecosystem functions. However, inferring microbial taxonomy from metabolic marker gene sequences remains a challenge, particularly for the frequently sequenced nitrogen fixation marker gene, nitrogenase reductase (nifH). Horizontal gene transfer in recent nifH evolutionary history can confound taxonomic inferences drawn from the pairwise identity methods used in existing software. Other methods for inferring taxonomy are not standardized and require manual inspection that is difficult to scale.RESULTS: We present Phylogenetic Placement for Inferring Taxonomy (PPIT), an R package that infers microbial taxonomy from nifH amplicons using both phylogenetic and sequence identity approaches. After users place query sequences on a reference nifH gene tree provided by PPIT (n=6317 full-length nifH sequences), PPIT searches the phylogenetic neighborhood of each query sequence and attempts to infer microbial taxonomy. An inference is drawn only if references in the phylogenetic neighborhood are: (1) taxonomically consistent and (2) share sufficient pairwise identity with the query, thereby avoiding erroneous inferences due to known horizontal gene transfer events. We find that PPIT returns a higher proportion of correct taxonomic inferences than BLAST-based approaches at the cost of fewer total inferences. We demonstrate PPIT on deep-sea sediment and find that Deltaproteobacteria are the most abundant potential diazotrophs. Using this dataset we show that emending PPIT inferences based on visual inspection of query sequence placement can achieve taxonomic inferences for nearly all sequences in a query set. We additionally discuss how users can apply PPIT to the analysis of other marker genes.AVAILABILITY: PPIT is freely available to non-commercial users at https://github.com/bkapili/ppit. Installation includes a vignette that demonstrates package use and reproduces the nifH amplicon analysis discussed here. The raw nifH amplicon sequence data have been deposited in the GenBank, EMBL, and DDBJ databases under BioProject number PRJEB37167.SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.
View details for DOI 10.1093/bioinformatics/btab100
View details for PubMedID 33580675
- DNA Tracer Transport Through Porous Media-The Effect of DNA Length and Adsorption WATER RESOURCES RESEARCH 2021; 57 (2)
NanoSIMS sample preparation decreases isotope enrichment: magnitude, variability and implications for single-cell rates of microbial activity.
The activity of individual microorganisms can be measured within environmental samples by detecting uptake of isotope-labeled substrates using nano-scale secondary ion mass spectrometry (nanoSIMS). Recent studies have demonstrated that sample preparation can decrease 13 C and 15 N enrichment in bacterial cells, resulting in underestimates of activity. Here, we explore this effect with a variety of preparation types, microbial lineages, and isotope labels to determine its consistency and therefore potential for correction. Specifically, we investigated the impact of different protocols for fixation, nucleic acid staining, and catalyzed reporter deposition fluorescence in situ hybridization (CARD-FISH) on >14,500 archaeal and bacterial cells (Methanosarcina acetivorans, Sulfolobus acidocaldarius, and Pseudomonas putida) enriched in 13 C, 15 N, 18 O, 2 H, and/or 34 S. We found these methods decrease isotope enrichments by up to 80% - much more than previously reported - and that the effect varies by taxa, growth phase, isotope label, and applied protocol. We make recommendations for how to account for this effect experimentally and analytically. We also re-evaluate published nanoSIMS datasets, and revise estimated microbial turnover times in the marine subsurface and nitrogen fixation rates in pelagic unicellular cyanobacteria. When sample preparation is accounted for, cell-specific rates increase and are more consistent with modeled and bulk rates. This article is protected by copyright. All rights reserved.
View details for DOI 10.1111/1462-2920.15264
View details for PubMedID 33000528
Evidence for phylogenetically and catabolically diverse active diazotrophs in deep-sea sediment.
The ISME journal
Diazotrophic microorganisms regulate marine productivity by alleviating nitrogen limitation. However, we know little about the identity and activity of diazotrophs in deep-sea sediments, a habitat covering nearly two-thirds of the planet. Here, we identify candidate diazotrophs from Pacific Ocean sediments collected at 2893m water depth using 15N-DNA stable isotope probing and a novel pipeline for nifH sequence analysis. Together, these approaches detect an unexpectedly diverse assemblage of active diazotrophs, including members of the Acidobacteria, Firmicutes, Nitrospirae, Gammaproteobacteria, and Deltaproteobacteria. Deltaproteobacteria, predominately members of the Desulfobacterales and Desulfuromonadales, are the most abundant diazotrophs detected, and display the most microdiversity of associated nifH sequences. Some of the detected lineages, including those within the Acidobacteria, have not previously been shown to fix nitrogen. The diazotrophs appear catabolically diverse, with the potential for using oxygen, nitrogen, iron, sulfur, and carbon as terminal electron acceptors. Therefore, benthic diazotrophy may persist throughout a range of geochemical conditions and provide a stable source of fixed nitrogen over geologic timescales. Our results suggest that nitrogen-fixing communities in deep-sea sediments are phylogenetically and catabolically diverse, and open a new line of inquiry into the ecology and biogeochemical impacts of deep-sea microorganisms.
View details for DOI 10.1038/s41396-019-0584-8
View details for PubMedID 31907368
Characterizing Chemoautotrophy and Heterotrophy in Marine Archaea and Bacteria With Single-Cell Multi-isotope NanoSIP.
Frontiers in microbiology
2019; 10: 2682
Characterizing and quantifying in situ metabolisms remains both a central goal and challenge for environmental microbiology. Here, we used a single-cell, multi-isotope approach to investigate the anabolic activity of marine microorganisms, with an emphasis on natural populations of Thaumarchaeota. After incubating coastal Pacific Ocean water with 13C-bicarbonate and 15N-amino acids, we used nanoscale secondary ion mass spectrometry (nanoSIMS) to isotopically screen 1,501 individual cells, and 16S rRNA amplicon sequencing to assess community composition. We established isotopic enrichment thresholds for activity and metabolic classification, and with these determined the percentage of anabolically active cells, the distribution of activity across the whole community, and the metabolic lifestyle-chemoautotrophic or heterotrophic-of each cell. Most cells (>90%) were anabolically active during the incubation, and 4-17% were chemoautotrophic. When we inhibited bacteria with antibiotics, the fraction of chemoautotrophic cells detected via nanoSIMS increased, suggesting archaea dominated chemoautotrophy. With fluorescence in situ hybridization coupled to nanoSIMS (FISH-nanoSIMS), we confirmed that most Thaumarchaeota were living chemoautotrophically, while bacteria were not. FISH-nanoSIMS analysis of cells incubated with dual-labeled (13C,15N-) amino acids revealed that most Thaumarchaeota cells assimilated amino-acid-derived nitrogen but not carbon, while bacteria assimilated both. This indicates that some Thaumarchaeota do not assimilate intact amino acids, suggesting intra-phylum heterogeneity in organic carbon utilization, and potentially their use of amino acids for nitrification. Together, our results demonstrate the utility of multi-isotope nanoSIMS analysis for high-throughput metabolic screening, and shed light on the activity and metabolism of uncultured marine archaea and bacteria.
View details for DOI 10.3389/fmicb.2019.02682
View details for PubMedID 31920997
View details for PubMedCentralID PMC6927911
Microbial Community Composition in Deep‐Subsurface Reservoir Fluids Reveals Natural Interwell Connectivity
Water Resources Research
View details for DOI 10.1029/2019WR025916
High-quality genome sequences of uncultured microbes by assembly of read clouds.
Although shotgun metagenomic sequencing of microbiome samples enables partial reconstruction of strain-level community structure, obtaining high-quality microbial genome drafts without isolation and culture remains difficult. Here, we present an application of read clouds, short-read sequences tagged with long-range information, to microbiome samples. We present Athena, a de novo assembler that uses read clouds to improve metagenomic assemblies. We applied this approach to sequence stool samples from two healthy individuals and compared it with existing short-read and synthetic long-read metagenomic sequencing techniques. Read-cloud metagenomic sequencing and Athena assembly produced the most comprehensive individual genome drafts with high contiguity (>200-kb N50, fewer than ten contigs), even for bacteria with relatively low (20*) raw short-read-sequence coverage. We also sequenced a complex marine-sediment sample and generated 24 intermediate-quality genome drafts (>70% complete, <10% contaminated), nine of which were complete (>90% complete, <5% contaminated). Our approach allows for culture-free generation of high-quality microbial genome drafts by using a single shotgun experiment.
View details for PubMedID 30320765
Widespread nitrogen fixation in sediments from diverse deep-sea sites of elevated carbon loading.
Nitrogen fixation, the biological conversion of N2 to NH3 , is critical to alleviating nitrogen limitation in many marine ecosystems. To date, few measurements exist of N2 fixation in deep-sea sediments. Here, we conducted >400 bottle incubations with sediments from methane seeps, whale falls, and background sites off the western coast of the United States from 600 to 2893 m water depth to investigate the potential rates, spatial distribution, and biological mediators of benthic N2 fixation. We found that N2 fixation was widespread, yet heterogeneously distributed with sediment depth at all sites. In some locations, rates exceeded previous measurements by >10X, and provided up to 31% of the community anabolic growth requirement for nitrogen. Diazotrophic activity appeared to be inhibited by pore water ammonium: N2 fixation was only observed if incubation ammonium concentrations were ≤25 muM, and experimental additions of ammonium reduced diazotrophy. In seep sediments, N2 fixation was dependent on CH4 and coincident with sulfate reduction, consistent with previous work showing diazotrophy by microorganisms mediating sulfate-coupled methane oxidation. However, the pattern of diazotrophy was different in whale fall and associated reference sediments, where it was largely unaffected by CH4 , suggesting catabolically different diazotrophs at these sites. This article is protected by copyright. All rights reserved.
View details for PubMedID 29968367
Characterization of benthic biogeochemistry and ecology at three methane seep sites on the Northern U.S. Atlantic margin
Deep Sea Research Part II: Topical Studies in Oceanography
2018; 150: 41-56
View details for DOI 10.1016/j.dsr2.2018.03.001
Early-career scientists explore newly discovered methane seeps
View details for DOI 10.1029/2017EO068011
Activity and interactions of methane seep microorganisms assessed by parallel transcription and FISH-NanoSIMS analyses
2016; 10 (3): 678-692
To characterize the activity and interactions of methanotrophic archaea (ANME) and Deltaproteobacteria at a methane-seeping mud volcano, we used two complimentary measures of microbial activity: a community-level analysis of the transcription of four genes (16S rRNA, methyl coenzyme M reductase A (mcrA), adenosine-5'-phosphosulfate reductase α-subunit (aprA), dinitrogenase reductase (nifH)), and a single-cell-level analysis of anabolic activity using fluorescence in situ hybridization coupled to nanoscale secondary ion mass spectrometry (FISH-NanoSIMS). Transcript analysis revealed that members of the deltaproteobacterial groups Desulfosarcina/Desulfococcus (DSS) and Desulfobulbaceae (DSB) exhibit increased rRNA expression in incubations with methane, suggestive of ANME-coupled activity. Direct analysis of anabolic activity in DSS cells in consortia with ANME by FISH-NanoSIMS confirmed their dependence on methanotrophy, with no (15)NH4(+) assimilation detected without methane. In contrast, DSS and DSB cells found physically independent of ANME (i.e., single cells) were anabolically active in incubations both with and without methane. These single cells therefore comprise an active 'free-living' population, and are not dependent on methane or ANME activity. We investigated the possibility of N2 fixation by seep Deltaproteobacteria and detected nifH transcripts closely related to those of cultured diazotrophic Deltaproteobacteria. However, nifH expression was methane-dependent. (15)N2 incorporation was not observed in single DSS cells, but was detected in single DSB cells. Interestingly, (15)N2 incorporation in single DSB cells was methane-dependent, raising the possibility that DSB cells acquired reduced (15)N products from diazotrophic ANME while spatially coupled, and then subsequently dissociated. With this combined data set we address several outstanding questions in methane seep microbial ecosystems and highlight the benefit of measuring microbial activity in the context of spatial associations.
View details for DOI 10.1038/ismej.2015.145
View details for Web of Science ID 000370472500013
View details for PubMedID 26394007
View details for PubMedCentralID PMC4817681
Global metagenomic survey reveals a new bacterial candidate phylum in geothermal springs
Analysis of the increasing wealth of metagenomic data collected from diverse environments can lead to the discovery of novel branches on the tree of life. Here we analyse 5.2 Tb of metagenomic data collected globally to discover a novel bacterial phylum ('Candidatus Kryptonia') found exclusively in high-temperature pH-neutral geothermal springs. This lineage had remained hidden as a taxonomic 'blind spot' because of mismatches in the primers commonly used for ribosomal gene surveys. Genome reconstruction from metagenomic data combined with single-cell genomics results in several high-quality genomes representing four genera from the new phylum. Metabolic reconstruction indicates a heterotrophic lifestyle with conspicuous nutritional deficiencies, suggesting the need for metabolic complementarity with other microbes. Co-occurrence patterns identifies a number of putative partners, including an uncultured Armatimonadetes lineage. The discovery of Kryptonia within previously studied geothermal springs underscores the importance of globally sampled metagenomic data in detection of microbial novelty, and highlights the extraordinary diversity of microbial life still awaiting discovery.
View details for DOI 10.1038/ncomms10476
View details for Web of Science ID 000369019300004
View details for PubMedID 26814032
Spatial distribution of nitrogen fixation in methane seep sediment and the role of the ANME archaea
2014; 16 (10): 3012-3029
Nitrogen (N2) fixation was investigated at Mound 12, Costa Rica, to determine its spatial distribution and biogeochemical controls in deep-sea methane seep sediment. Using (15)N2 tracer experiments and isotope ratio mass spectrometry analysis, we observed that seep N2 fixation is methane-dependent, and that N2 fixation rates peak in a narrow sediment depth horizon corresponding to increased abundance of aggregates of anaerobic methanotrophic archaea (ANME-2) and sulfate-reducing bacteria (SRB). Using fluorescence in situ hybridization coupled to nanoscale secondary ion mass spectrometry (FISH-NanoSIMS), we directly measured (15)N2 uptake by ANME-2/SRB aggregates (n = 26) and observed maximum (15)N incorporation within ANME-2-dominated areas of the aggregates, consistent with previous analyses. NanoSIMS analysis of single cells (n = 34) from the same microcosm experiment revealed no (15)N2 uptake. Together, these observations suggest that ANME-2, and possibly physically associated SRB, mediate the majority of new nitrogen production within the seep ecosystem. ANME-2 diazotrophy was observed while in association with members of two distinct orders of SRB: Desulfobacteraceae and Desulfobulbaceae. The rate of N2 fixation per unit volume biomass was independent of the identity of the associated SRB, aggregate size and morphology. Our results show that the distribution of seep N2 fixation is heterogeneous, laterally and with depth in the sediment, and is likely influenced by chemical gradients affecting the abundance and activity of ANME-2/SRB aggregates.
View details for DOI 10.1111/1462-2920.12247
View details for Web of Science ID 000343867700002
View details for PubMedID 24107237
Nitrate-based niche differentiation by distinct sulfate-reducing bacteria involved in the anaerobic oxidation of methane
2014; 8 (1): 150-163
Diverse associations between methanotrophic archaea (ANME) and sulfate-reducing bacterial groups (SRB) often co-occur in marine methane seeps; however, the ecophysiology of these different symbiotic associations has not been examined. Here, we applied a combination of molecular, geochemical and Fluorescence in situ hybridization (FISH) coupled to nanoscale secondary ion mass spectrometry (FISH-NanoSIMS) analyses of in situ seep sediments and methane-amended sediment incubations from diverse locations (Eel River Basin, Hydrate Ridge and Costa Rican Margin seeps) to investigate the distribution and physiology of a newly identified subgroup of the Desulfobulbaceae (seepDBB) found in consortia with ANME-2c archaea, and compared these with the more commonly observed associations between the same ANME partner and the Desulfobacteraceae (DSS). FISH analyses revealed aggregates of seepDBB cells in association with ANME-2 from both environmental samples and laboratory incubations that are distinct in their structure relative to co-occurring ANME/DSS consortia. ANME/seepDBB aggregates were most abundant in shallow sediment depths below sulfide-oxidizing microbial mats. Depth profiles of ANME/seepDBB aggregate abundance revealed a positive correlation with elevated porewater nitrate relative to ANME/DSS aggregates in all seep sites examined. This relationship with nitrate was supported by sediment microcosm experiments, in which the abundance of ANME/seepDBB was greater in nitrate-amended incubations relative to the unamended control. FISH-NanoSIMS additionally revealed significantly higher (15)N-nitrate incorporation levels in individual aggregates of ANME/seepDBB relative to ANME/DSS aggregates from the same incubation. These combined results suggest that nitrate is a geochemical effector of ANME/seepDBB aggregate distribution, and provides a unique niche for these consortia through their utilization of a greater range of nitrogen substrates than the ANME/DSS.
View details for DOI 10.1038/ismej.2013.147
View details for Web of Science ID 000328605200015
View details for PubMedID 24008326
Polyphosphate Storage during Sporulation in the Gram-Negative Bacterium Acetonema longum
JOURNAL OF BACTERIOLOGY
2013; 195 (17): 3940-3946
Using electron cryotomography, we show that the Gram-negative sporulating bacterium Acetonema longum synthesizes high-density storage granules at the leading edges of engulfing membranes. The granules appear in the prespore and increase in size and number as engulfment proceeds. Typically, a cluster of 8 to 12 storage granules closely associates with the inner spore membrane and ultimately accounts for ∼7% of the total volume in mature spores. Energy-dispersive X-ray spectroscopy (EDX) analyses show that the granules contain high levels of phosphorus, oxygen, and magnesium and therefore are likely composed of polyphosphate (poly-P). Unlike the Gram-positive Bacilli and Clostridia, A. longum spores retain their outer spore membrane upon germination. To explore the possibility that the granules in A. longum may be involved in this unique process, we imaged purified Bacillus cereus, Bacillus thuringiensis, Bacillus subtilis, and Clostridium sporogenes spores. Even though B. cereus and B. thuringiensis contain the ppk and ppx genes, none of the spores from Gram-positive bacteria had granules. We speculate that poly-P in A. longum may provide either the energy or phosphate metabolites needed for outgrowth while retaining an outer membrane.
View details for DOI 10.1128/JB.00712-13
View details for Web of Science ID 000323047900016
View details for PubMedID 23813732
IDENTIFICATION OF DIAZOTROPHIC MICROORGANISMS IN MARINE SEDIMENT VIA FLUORESCENCE IN SITU HYBRIDIZATION COUPLED TO NANOSCALE SECONDARY ION MASS SPECTROMETRY (FISH-NANOSIMS)
METHODS IN ENZYMOLOGY: RESEARCH ON NITRIFICATION AND RELATED PROCESSES, VOL 486, PART A
2011; 486: 281-305
Growing appreciation for the biogeochemical significance of uncultured microorganisms is changing the focus of environmental microbiology. Techniques designed to investigate microbial metabolism in situ are increasingly popular, from mRNA-targeted fluorescence in situ hybridization (FISH) to the "-omics" revolution, including metagenomics, transcriptomics, and proteomics. Recently, the coupling of FISH with nanometer-scale secondary ion mass spectrometry (NanoSIMS) has taken this movement in a new direction, allowing single-cell metabolic analysis of uncultured microbial phylogenic groups. The main advantage of FISH-NanoSIMS over previous noncultivation-based techniques to probe metabolism is its ability to directly link 16S rRNA phylogenetic identity to metabolic function. In the following chapter, we describe the procedures necessary to identify nitrogen-fixing microbes within marine sediment via FISH-NanoSIMS, using our work on nitrogen fixation by uncultured deep-sea methane-consuming archaea as a case study.
View details for DOI 10.1016/B978-0-12-381294-0.00012-2
View details for Web of Science ID 000286404100012
View details for PubMedID 21185440
Deep-Sea Archaea Fix and Share Nitrogen in Methane-Consuming Microbial Consortia
2009; 326 (5951): 422-426
Nitrogen-fixing (diazotrophic) microorganisms regulate productivity in diverse ecosystems; however, the identities of diazotrophs are unknown in many oceanic environments. Using single-cell-resolution nanometer secondary ion mass spectrometry images of 15N incorporation, we showed that deep-sea anaerobic methane-oxidizing archaea fix N2, as well as structurally similar CN-, and share the products with sulfate-reducing bacterial symbionts. These archaeal/bacterial consortia are already recognized as the major sink of methane in benthic ecosystems, and we now identify them as a source of bioavailable nitrogen as well. The archaea maintain their methane oxidation rates while fixing N2 but reduce their growth, probably in compensation for the energetic burden of diazotrophy. This finding extends the demonstrated lower limits of respiratory energy capable of fueling N2 fixation and reveals a link between the global carbon, nitrogen, and sulfur cycles.
View details for DOI 10.1126/science.1178223
View details for Web of Science ID 000270818600052
View details for PubMedID 19833965
Bacillus canaveralius sp nov., an alkali-tolerant bacterium isolated from a spacecraft assembly facility
INTERNATIONAL JOURNAL OF SYSTEMATIC AND EVOLUTIONARY MICROBIOLOGY
2009; 59: 2015-2019
Two Gram-positive, rod-shaped, alkali-tolerant (pH 10.5), endospore-forming bacteria (strains KSC SF8bT and KSC SF10a) were isolated from surfaces within the Payload Hazardous Servicing Facility, where robotic spacecraft are assembled and tested before launch, at the Kennedy Space Center at Cape Canaveral. Based on 16S rRNA gene sequence similarities, these strains were shown to belong to the family Bacillaceae and the genus Bacillus. The highest 16S rRNA gene sequence similarity was approximately 97.5%, observed between the novel strains and Bacillus selenatarsenatis SF-1T. Several phenotypic characteristics, such as growth with 10% NaCl and assimilation of melibiose and lactose, were useful in the discrimination of this novel species from the closely related alkali-tolerant species Bacillus firmus and B. selenatarsenatis. DNA-DNA hybridization studies revealed reassociation values of less than 45% between strain KSC SF8bT and its closest genotypic neighbours. The combination of unique phenotypic and genotypic characteristics allowed the differentiation of these alkali- and halotolerant spore-forming strains from related Bacillus species, and a novel species, Bacillus canaveralius sp. nov., is proposed. The type strain is KSC SF8bT (=ATCC BAA-1493T=MTCC 8908T).
View details for DOI 10.1099/ijs.0.009167-0
View details for Web of Science ID 000271435400027
View details for PubMedID 19567559
Diverse syntrophic partnerships from-deep-sea methane vents revealed by direct cell capture and metagenomics
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
2008; 105 (19): 7052-7057
Microorganisms play a fundamental role in the cycling of nutrients and energy on our planet. A common strategy for many microorganisms mediating biogeochemical cycles in anoxic environments is syntrophy, frequently necessitating close spatial proximity between microbial partners. We are only now beginning to fully appreciate the diversity and pervasiveness of microbial partnerships in nature, the majority of which cannot be replicated in the laboratory. One notable example of such cooperation is the interspecies association between anaerobic methane oxidizing archaea (ANME) and sulfate-reducing bacteria. These consortia are globally distributed in the environment and provide a significant sink for methane by substantially reducing the export of this potent greenhouse gas into the atmosphere. The interdependence of these currently uncultured microbes renders them difficult to study, and our knowledge of their physiological capabilities in nature is limited. Here, we have developed a method to capture select microorganisms directly from the environment, using combined fluorescence in situ hybridization and immunomagnetic cell capture. We used this method to purify syntrophic anaerobic methane oxidizing ANME-2c archaea and physically associated microorganisms directly from deep-sea marine sediment. Metagenomics, PCR, and microscopy of these purified consortia revealed unexpected diversity of associated bacteria, including Betaproteobacteria and a second sulfate-reducing Deltaproteobacterial partner. The detection of nitrogenase genes within the metagenome and subsequent demonstration of (15)N(2) incorporation in the biomass of these methane-oxidizing consortia suggest a possible role in new nitrogen inputs by these syntrophic assemblages.
View details for DOI 10.1073/pnas.0711303105
View details for Web of Science ID 000255921200048
View details for PubMedID 18467493
Microbial burden and diversity of commercial airline cabin air during short and long durations of travel
2008; 2 (5): 482-497
Total microbial burden and diversity associated with commercial airliner cabin air was assessed by molecular methods in 125 air samples from the business-class sections of 16 domestic and international flights. Viable microbial burden within these cabin air parcels constituted only 1-10% of the total microbial population and ranged from below detection limits to 1.2 x 10(4) cells m(-3) as determined with a validated ATP-based technology. Cultivable bacterial diversity was almost entirely limited to Gram-positive bacteria such as Staphylococcus and Bacillus. In contrast, cloning and sequencing 16S rRNA gene directly from the samples without cultivation indicated a significantly broader diversity, as sequences representing more than 100 species, and encompassing 12 classes of bacteria, were retrieved in varying abundance. Sequences of proteobacterial and Gram-positive lineage were retrieved most frequently (58% and 31% of all clone sequences, respectively), with Gram-positive and alpha-proteobacterial sequences dominating international flight samples and beta- and gamma-proteobacterial sequences comprising the largest portion of those retrieved from domestic flights. Significant differences in bacterial load and diversity were noted between samples obtained on domestic and international flights. The disparities observed in microbial abundance and diversity further underscore the immense value of state-of-the art molecular assays in augmenting traditional culture-based techniques.
View details for DOI 10.1038/ismej.2008.11
View details for Web of Science ID 000255974500004
View details for PubMedID 18256704
Molecular bacterial community analysis of clean rooms where spacecraft are assembled
FEMS MICROBIOLOGY ECOLOGY
2007; 61 (3): 509-521
Molecular bacterial community composition was characterized from three geographically distinct spacecraft-associated clean rooms to determine whether such populations are influenced by the surrounding environment or the maintenance of the clean rooms. Samples were collected from facilities at the Jet Propulsion Laboratory (JPL), Kennedy Space Flight Center (KSC), and Johnson Space Center (JSC). Nine clone libraries representing different surfaces within the spacecraft facilities and three libraries from the surrounding air were created. Despite the highly desiccated, nutrient-bare conditions within these clean rooms, a broad diversity of bacteria was detected, covering all the main bacterial phyla. Furthermore, the bacterial communities were significantly different from each other, revealing only a small subset of microorganisms common to all locations (e.g. Sphingomonas, Staphylococcus). Samples from JSC assembly room surfaces showed the greatest diversity of bacteria, particularly within the Alpha- and Gammaproteobacteria and Actinobacteria. The bacterial community structure of KSC assembly surfaces revealed a high presence of proteobacterial groups, whereas the surface samples collected from the JPL assembly facility showed a predominance of Firmicutes. Our study presents the first extended molecular survey and comparison of NASA spacecraft assembly facilities, and provides new insights into the bacterial diversity of clean room environments .
View details for DOI 10.1111/j.1574-6941.2007.00360.x
View details for Web of Science ID 000248961900011
View details for PubMedID 17655710
Isolation and characterization of bacteria capable of tolerating the extreme conditions of clean room environments
APPLIED AND ENVIRONMENTAL MICROBIOLOGY
2007; 73 (8): 2600-2611
In assessing the bacterial populations present in spacecraft assembly, spacecraft test, and launch preparation facilities, extremophilic bacteria (requiring severe conditions for growth) and extremotolerant bacteria (tolerant to extreme conditions) were isolated. Several cultivation approaches were employed to select for and identify bacteria that not only survive the nutrient-limiting conditions of clean room environments but can also withstand even more inhospitable environmental stresses. Due to their proximity to spacefaring objects, these bacteria pose a considerable risk for forward contamination of extraterrestrial sites. Samples collected from four geographically distinct National Aeronautics and Space Administration clean rooms were challenged with UV-C irradiation, 5% hydrogen peroxide, heat shock, pH extremes (pH 3.0 and 11.0), temperature extremes (4 degrees C to 65 degrees C), and hypersalinity (25% NaCl) prior to and/or during cultivation as a means of selecting for extremotolerant bacteria. Culture-independent approaches were employed to measure viable microbial (ATP-based) and total bacterial (quantitative PCR-based) burdens. Intracellular ATP concentrations suggested a viable microbial presence ranging from below detection limits to 10(6) cells/m(2). However, only 0.1 to 55% of these viable cells were able to grow on defined culture medium. Isolated members of the Bacillaceae family were more physiologically diverse than those reported in previous studies, including thermophiles (Geobacillus), obligate anaerobes (Paenibacillus), and halotolerant, alkalophilic species (Oceanobacillus and Exiguobacterium). Non-spore-forming microbes (alpha- and beta-proteobacteria and actinobacteria) exhibiting tolerance to the selected stresses were also encountered. The multiassay cultivation approach employed herein enhances the current understanding of the physiological diversity of bacteria housed in these clean rooms and leads us to ponder the origin and means of translocation of thermophiles, anaerobes, and halotolerant alkalophiles into these environments.
View details for DOI 10.1128/AEM.03007-06
View details for Web of Science ID 000246542400023
View details for PubMedID 17308177
High-mass triple systems: The classical Cepheid Y Carinae
2005; 130 (2): 789-793
View details for Web of Science ID 000230721600034