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Global warming-induced sea ice loss in the Canadian Northwest Passage (NWP) will result in more shipping traffic, increasing the risk of oil spills. Microorganisms inhabiting NWP beach sediments may degrade hydrocarbons, offering a potential bioremediation strategy. In this study, the characterization and genomic analyses of 22 hydrocarbon-biodegradative bacterial isolates revealed that they contained a diverse range of key alkane and aromatic hydrocarbon-degradative genes, as well as cold and salt tolerance genes indicating they are highly adapted to the extreme Arctic environment. Some isolates successfully degraded Ultra Low Sulfur Fuel Oil (ULSFO) at temperatures as low as −5 °C and high salinities (3%–10%). Three isolates were grown in liquid medium containing ULSFO as sole carbon source over 3 months and variation of hydrocarbon concentration was measured at three time points to determine their rate of hydrocarbon biodegradation. Our results demonstrate that two isolates (Rhodococcus sp. R1B_2T and Pseudarthrobacter sp. R2D_1T) possess complete degradation pathways and can grow on alkane and aromatic components of ULSFO under Arctic conditions. Overall, these results demonstrate that diverse hydrocarbon-degrading microorganisms exist in the NWP beach sediments, offering a potential bioremediation strategy in the events of a marine fuel spill reaching the shores of the NWP. [ABSTRACT FROM AUTHOR]

Greenhouse gas (GHG) emissions from Arctic permafrost soils create a positive feedback loop of climate warming and further GHG emissions. Active methane uptake in these soils can reduce the impact of GHG on future Arctic warming potential. Aerobic methane oxidizers are thought to be responsible for this apparent methane sink, though Arctic representatives of these organisms have resisted culturing efforts. Here, we first used in situ gas flux measurements and qPCR to identify relative methane sink hotspots at a high Arctic cytosol site, we then labeled the active microbiome in situ using DNA Stable Isotope Probing (SIP) with heavy 13CH4 (at 100 ppm and 1000 ppm). This was followed by amplicon and metagenome sequencing to identify active organisms involved in CH4 metabolism in these high Arctic cryosols. Sequencing of 13C-labeled pmoA genes demonstrated that type II methanotrophs (Methylocapsa) were overall the dominant active methane oxidizers in these mineral cryosols, while type I methanotrophs (Methylomarinovum) were only detected in the 100 ppm SIP treatment. From the SIP-13C-labeled DNA, we retrieved nine high to intermediate quality metagenome-assembled genomes (MAGs) belonging to the Proteobacteria, Gemmatimonadetes, and Chloroflexi, with three of these MAGs containing genes associated with methanotrophy. A novel Chloroflexi MAG contained a mmoX gene along with other methane oxidation pathway genes, identifying it as a potential uncultured methane oxidizer. This MAG also contained genes for copper import, synthesis of biopolymers, mercury detoxification, and ammonia uptake, indicating that this bacterium is strongly adapted to conditions in active layer permafrost and providing new insights into methane biogeochemical cycling. In addition, Betaproteobacterial MAGs were also identified as potential cross-feeders with methanotrophs in these Arctic cryosols. Overall, in situ SIP labeling combined with metagenomics and genome binning demonstrated to be a useful tool for discovering and characterizing novel organisms related to specific microbial functions or biogeochemical cycles of interest. Our findings reveal a unique and active Arctic cryosol microbial community potentially involved in CH4 cycling. [ABSTRACT FROM AUTHOR]

The survey conducted about the microbial diversity in the sediments of seven springs in the Canadian Arctic region by denaturing gradient gel electrophoresis (DGGE) and analyzing clone libraries of 16S rRNA genes amplified with Bacteria and Archaea-specific primers is presented. Results showed utilization and cycling of sulfur compounds responsible for energy production and maintenance of microbial communities in Canadian cold environments.

Nunataks are permanent ice-free rocky peaks that project above ice caps in polar regions, thus being exposed to extreme climatic conditions throughout the year. They undergo extremely low temperatures and scarcity of liquid water in winter, while receiving high incident and reflected (albedo) UVA-B radiation in summer. Here, we investigate the geomicrobiology of the permanently exposed lithic substrates of nunataks from Livingston Island (South Shetlands, Antarctic Peninsula), with focus on prokaryotic community structure and their main metabolic traits. Contrarily to first hypothesis, an extensive sampling based on different gradients and multianalytical approaches demonstrated significant differences for most geomicrobiological parameters between the bedrock, soil, and loose rock substrates, which overlapped any other regional variation. Brevibacillus genus dominated on bedrock and soil substrates, while loose rocks contained a diverse microbial community, including Actinobacteria, Alphaproteobacteria and abundant Cyanobacteria inhabiting the milder and diverse microhabitats within. Archaea, a domain never described before in similar Antarctic environments, were also consistently found in the three substrates, but being more abundant and potentially more active in soils. Stable isotopic ratios of total carbon (δ 13C) and nitrogen (δ 15N), soluble anions concentrations, and the detection of proteins involved in key metabolisms via the Life Detector Chip (LDChip), suggest that microbial primary production has a pivotal role in nutrient cycling at these exposed areas with limited deposition of nutrients. Detection of stress-resistance proteins, such as molecular chaperons, suggests microbial molecular adaptation mechanisms to cope with these harsh conditions. Since early Mars may have encompassed analogous environmental conditions as the ones found in these Antarctic nunataks, our study also contributes to the understanding of the metabolic features and biomarker profiles of a potential Martian microbiota, as well as the use of LDChip in future life detection missions. [ABSTRACT FROM AUTHOR]

Summary: The Gypsum Hill (GH) springs on Axel Heiberg Island in the Canadian high Arctic are host to chemolithoautotrophic, sulfur‐oxidizing streamers that flourish in the high Arctic winter in water temperatures from −1.3 to 7°C with ~8% salinity in a high Arctic winter environment with air temperatures commonly less than −40°C and an average annual air temperature of −15°C. Metagenome sequencing and binning of streamer samples produced a 96% complete Thiomicrorhabdus sp. metagenome‐assembled genome representing a possible new species or subspecies. This is the most cold‐ and salt‐extreme source environment for a Thiomicrorhabdus genome yet described. Metaproteomic and metatranscriptomic analysis attributed nearly all gene expression in the streamers to the Thiomicrorhabdus sp. and suggested that it is active in CO2 fixation and oxidation of sulfide to elemental sulfur. In situ geochemical and isotopic analyses of the fractionation of multiple sulfur isotopes determined the biogeochemical transformation of sulfur from its source in Carboniferous evaporites to biotic processes occurring in the sediment and streamers. These complementary molecular tools provided a functional link between the geochemical substrates and the collective traits and activity that define the microbial community's interactions within a unique polar saline habitat where Thiomicrorhabdus‐dominated streamers form and flourish. [ABSTRACT FROM AUTHOR]

The extent of fractionation of sulfur isotopes by sulfate‐reducing microbes is dictated by genomic and environmental factors. A greater understanding of species‐specific fractionations may better inform interpretation of sulfur isotopes preserved in the rock record. To examine whether gene diversity influences net isotopic fractionation in situ, we assessed environmental chemistry, sulfate reduction rates, diversity of putative sulfur‐metabolizing organisms by 16S rRNA and dissimilatory sulfite reductase (dsrB) gene amplicon sequencing, and net fractionation of sulfur isotopes along a sediment transect of a hypersaline Arctic spring. In situ sulfate reduction rates yielded minimum cell‐specific sulfate reduction rates < 0.3 × 10−15 moles cell−1 day−1. Neither 16S rRNA nor dsrB diversity indices correlated with relatively constant (38‰–45‰) net isotope fractionation (ε34Ssulfide‐sulfate). Measured ε34S values could be reproduced in a mechanistic fractionation model if 1%–2% of the microbial community (10%–60% of Deltaproteobacteria) were engaged in sulfate respiration, indicating heterogeneous respiratory activity within sulfate‐reducing populations. This model indicated enzymatic kinetic diversity of Apr was more likely to correlate with sulfur fractionation than DsrB. We propose that, above a threshold Shannon diversity value of 0.8 for dsrB, the influence of the specific composition of the microbial community responsible for generating an isotope signal is overprinted by the control exerted by environmental variables on microbial physiology. [ABSTRACT FROM AUTHOR]

Summary: Microbial metabolism of the thawing organic carbon stores in permafrost results in a positive feedback loop of greenhouse gas emissions. CO2 and CH4 fluxes and the associated microbial communities in Arctic cryosols are important in predicting future warming potential of the Arctic. We demonstrate that topography had an impact on CH4 and CO2 flux at a high Arctic ice‐wedge polygon terrain site, with higher CO2 emissions and lower CH4 uptake at troughs compared to polygon interior soils. The pmoA sequencing suggested that USCα cluster of uncultured methanotrophs is likely responsible for observed methane sink. Community profiling revealed distinct assemblages across the terrain at different depths. Deeper soils contained higher abundances of Verrucomicrobia and Gemmatimonadetes, whereas the polygon interior had higher Acidobacteria and lower Betaproteobacteria and Deltaproteobacteria abundances. Genome sequencing of isolates from the terrain revealed presence of carbon cycling genes including ones involved in serine and ribulose monophosphate pathways. A novel hybrid network analysis identified key members that had positive and negative impacts on other species. Operational Taxonomic Units (OTUs) with numerous positive interactions corresponded to Proteobacteria, Candidatus Rokubacteria and Actinobacteria phyla, while Verrucomicrobia and Acidobacteria members had negative impacts on other species. Results indicate that topography and microbial interactions impact community composition. [ABSTRACT FROM AUTHOR]

Quantitative PCR (q-PCR), stable isotope probing (SIP) of DNA, denaturing gradient gel electrophoresis (DGGE) fingerprinting and sequencing of the 16S rRNA and pmoA genes are used for analyzing the activity and diversity of methanotrophic bacteria in active-layer soils from Ellesmere Island in the Canadian high Arctic. DNA-SIP is used for identifying the active methanotrophs in high Arctic soil for the first time.

Microarrays targeting the 16S rRNA genes of bacteria found in cold environments and hydrocarbon degradation genes as well as quantitative reverse transcriptase PCR targeting key functional genes were used to assess the bacterial community structure and function. The findings from the study demonstrated the value of molecular tools to monitor the polar bacteria and their associated functions during bioremediation.

The heterotrophic and autotrophic compositions of bacterial isolates from the saline springs of Gypsum Hill (GH) are examined to identify their phylogenetic and physiological characteristics that enable microorganisms to inhabit and sustain Arctic ecosystem. The results revealed that the springs' microbial community is primarily sustained by chemolithoautotrophic primary production performed by sulfur-oxidizing bacteria, even in the period of continuous illumination in the Arctic summer.

Potential benthic habitats of early Mars lakes, probably oligotrophic, could range from hydrothermal to cold sediments. Dynamic processes in the water column (such as turbidity or UV penetration) as well as in the benthic bed (temperature gradients, turbation, or sedimentation rate) contribute to supply nutrients to a potential microbial ecosystem. High altitude, oligotrophic, and deep Andean lakes with active deglaciation processes and recent or past volcanic activity are natural models to assess the feasibility of life in other planetary lake/ocean environments and to develop technology for their exploration. We sampled the benthic sediments (down to 269 m depth) of the oligotrophic lake Laguna Negra (Central Andes, Chile) to investigate its ecosystem through geochemical, biomarker profiling, and molecular ecology studies. The chemistry of the benthic water was similar to the rest of the water column, except for variable amounts of ammonium (up to 2.8 ppm) and nitrate (up to 0.13 ppm). A life detector chip with a 300-antibody microarray revealed the presence of biomass in the form of exopolysaccharides and other microbial markers associated to several phylogenetic groups and potential microaerobic and anaerobic metabolisms such as nitrate reduction. DNA analyses showed that 27% of the Archaea sequences corresponded to a group of ammonia-oxidizing archaea (AOA) similar (97%) to Nitrosopumilus spp. and Nitrosoarchaeum spp. (Thaumarchaeota), and 4% of Bacteria sequences to nitrite-oxidizing bacteria from the Nitrospira genus, suggesting a coupling between ammonia and nitrite oxidation. Mesocosm experiments with the specific AOA inhibitor 2-Phenyl-4,4,5,5-tetramethylimidazoline-1-oxyl 3-oxide (PTIO) demonstrated an AOA-associated ammonia oxidation activity with the simultaneous accumulation of nitrate and sulfate. The results showed a rich benthic microbial community dominated by microaerobic and anaerobic metabolisms thriving under aphotic, low temperature (4°C), and relatively high pressure, that might be a suitable terrestrial analog of other planetary settings. [ABSTRACT FROM AUTHOR]

Permafrost subzero environments harbor diverse, active communities of microorganisms. However, our understanding of the subzero growth, metabolisms, and adaptive properties of these microbes remains very limited. We performed transcriptomic analyses on two subzero-growing permafrost isolates with different growth profiles in order to characterize and compare their cold temperature growth and cold-adaptive strategies. The two organisms, Rhodococcus sp. JG3 (-5 to 30°C) and Polaromonas sp. Eur3 1.2.1 (-5 to 22°C), shared several common responses during low temperature growth, including induction of translation and ribosomal processes, upregulation of nutrient transport, increased oxidative and osmotic stress responses, and stimulation of polysaccharide capsule synthesis. Recombination appeared to be an important adaptive strategy for both isolates at low temperatures, likely as a mechanism to increase genetic diversity and the potential for survival in cold systems. While Rhodococcus sp. JG3 favored upregulating iron and amino acid transport, sustaining redox potential, and modulating fatty acid synthesis and composition during growth at -5°C compared to 25°C, Polaromonas sp. Eur3 1.2.1 increased the relative abundance of transcripts involved in primary energy metabolism and the electron transport chain, in addition to signal transduction and peptidoglycan synthesis at 0°C compared to 20°C. The increase in energy metabolism may explain why Polaromonas sp. Eur3 1.2.1 is able to sustain growth rates at 0°C comparable to those at higher temperatures. For Rhodococcus sp. JG3, flexibility in use of carbon sources, iron acquisition, control of membrane fatty acid composition, and modulating redox and co-factor potential may be ways in which this organism is able to sustain growth over a wider range of temperatures. Increasing our understanding of the microbes in these habitats helps us better understand active pathways and metabolisms in extreme environments. Identifying novel, thermolabile, and cold-active enzymes from studies such as this is also of great interest to the biotechnology and food industries. [ABSTRACT FROM AUTHOR]

Permafrost accounts for 27% of all soil ecosystems and harbors diverse microbial communities. Our understanding of microorganisms in permafrost, their activities and adaptations, remains limited. Using five subzero-growing (cryophilic) permafrost bacteria, we examined features of cold adaptation through comparative genomic analyses with mesophilic relatives. The cryophiles possess genes associated with cold adaptation, including cold shock proteins, RNA helicases, and oxidative stress and carotenoid synthesis enzymes. Higher abundances of genes associated with compatible solutes were observed, important for osmoregulation in permafrost brine veins. Most cryophiles in our study have higher transposase copy numbers than mesophiles. We investigated amino acid (AA) modifications in the cryophiles favoring increased protein flexibility at cold temperatures. Although overall there were few differences with the mesophiles, we found evidence of cold adaptation, with significant differences in proline, serine, glycine and aromaticity, in several cryophiles. The use of cold/hot AA ratios of >1, used in previous studies to indicate cold adaptation, was found to be inadequate on its own. Comparing the average of all cryophiles to all mesophiles, we found that overall cryophiles had a higher ratio of cold adapted proteins for serine (more serine), and to a lesser extent, proline and acidic residues (fewer prolines/acidic residues). [ABSTRACT FROM AUTHOR]

While many habitable niches on Earth are characterized by permanently cold conditions, little is known about the spatial structure of seasonal communities and the importance of substrate-cell associations in terrestrial cyroenvironments. Here we use the 16S rRNA gene as a marker for genetic diversity to compare two visually distinct but spatially integrated surface microbial mats on Axel Heiberg Island, Canadian high arctic, proximal to a perennial saline spring. This is the first study to describe the bacterial diversity in microbial mats on Axel Heiberg Island. The hypersaline springs on Axel Heiberg represent a unique analog to putative subsurface aquifers on Mars. The Martian subsurface represents the longest-lived potentially habitable environment on Mars and a better understanding of the microbial communities on Earth that thrive in analog conditions will help direct future life detection missions. The microbial mats sampled on Axel Heiberg are only visible during the summer months in seasonal flood plains formed by melt water and run-off from the proximal spring. Targeted-amplicon sequencing revealed that not only does the bacterial composition of the two mat communities differ substantially from the sediment community of the proximal cold spring, but that the mat communities are distinct from any other microbial community in proximity to the Arctic springs studied to date. All samples are dominated by Gammaproteobacteria: Thiotichales is dominant within the spring samples while Alteromonadales comprises a significant component of the mat communities. The two mat samples differ in their Thiotichales:Alteromonadales ratio and contribution of Bacteroidetes to overall diversity. The red mats have a greater proportion of Alteromonadales and Bacteroidetes reads. The distinct bacterial composition of the mat bacterial communities suggests that the spring communities are not sourced from the surface, and that seasonal melt events create ephemerally habitable niches with distinct microbial communities in the Canadian high arctic. The finding that these surficial complex microbial communities exist in close proximity to perennial springs demonstrates the existence of a transiently habitable niche in an important Mars analog site. [ABSTRACT FROM AUTHOR]

Significant progress is being made in the development of the next generation of low cost life detection instrumentation with much smaller size, mass and energy requirements. Here, we describe in situ life detection and sequencing in the field in soils over laying ice wedges in polygonal permafrost terrain on Axel Heiberg Island, located in the Canadian high Arctic (79°26'N), an analog to the polygonal permafrost terrain observed on Mars. The life detection methods used here include (1) the cryo-iPlate for culturing microorganisms using diffusion of in situ nutrients into semi-solid media (2) a Microbial Activity Microassay (MAM) plate (BIOLOG Ecoplate) for detecting viable extant microorganisms through a colourimetric assay, and (3) the Oxford Nanopore MinION for nucleic acid detection and sequencing of environmental samples and the products of MAM plate and cryo-iPlate. We obtained 39 microbial isolates using the cryo-iPlate, which included several putatively novel strains based on the 16S rRNA gene, including a Pedobacter sp. (96% closest similarity in GenBank) which we partially genome sequenced using theMinION. TheMAMplate successfully identified an active community capable of L-serine metabolism, which was used for metagenomic sequencing with the MinION to identify the active and enriched community. A metagenome on environmental ice wedge soil samples was completed, with base calling and uplink/downlink carried out via satellite internet. Validation of MinION sequencing using the Illumina MiSeq platform was consistent with the results obtained with the MinION. The instrumentation and technology utilized here is pre-existing, low cost, low mass, low volume, and offers the prospect of equipping micro-rovers and micro-penetrators with aggressive astrobiological capabilities. Since potentially habitable astrobiology targets have been identified (RSLs on Mars, near subsurface water ice on Mars, the plumes and oceans of Europa and Enceladus), future astrobiology missions will certainly target these areas and there is a need for direct life detection instrumentation. [ABSTRACT FROM AUTHOR]