Jacqueline Goordial

Functional characterization of bacteria isolated from ancient Arctic soil exposes diverse resistance mechanisms to modern antibiotics

Using functional metagenomics to study the resistomes of bacterial communities isolated from different layers of the Canadian high Arctic permafrost, we show that microbial communities harbored diverse resistance mechanisms at least 5,000 years ago. Among bacteria sampled from the ancient layers of a permafrost core, we isolated eight genes conferring clinical levels of resistance against aminoglycoside, β-lactam and tetracycline antibiotics that are naturally produced by microorganisms. Among these resistance genes, four also conferred resistance against amikacin, a modern semi-synthetic antibiotic that does not naturally occur in microorganisms. In bacteria sampled from the overlaying active layer, we isolated ten different genes conferring resistance to all six antibiotics tested in this study, including aminoglycoside, β-lactam and tetracycline variants that are naturally produced by microorganisms as well as semi-synthetic variants produced in the laboratory. On average, we found that resistance genes found in permafrost bacteria conferred lower levels of resistance against clinically relevant antibiotics than resistance genes sampled from the active layer. Our results demonstrate that antibiotic resistance genes were functionally diverse prior to the anthropogenic use of antibiotics, contributing to the evolution of natural reservoirs of resistance genes.

Nearing the cold-arid limits of microbial life in permafrost of an upper dry valley, Antarctica

Some of the coldest and driest permafrost soils on Earth are located in the high-elevation McMurdo Dry Valleys (MDVs) of Antarctica, but little is known about the permafrost microbial communities other than that microorganisms are present in these valleys. Here, we describe the microbiology and habitable conditions of highly unique dry and ice-cemented permafrost in University Valley, one of the coldest and driest regions in the MDVs (1700 m above sea level; mean temperature −23 °C; no degree days above freezing), where the ice in permafrost originates from vapour deposition rather than liquid water. We found that culturable and total microbial biomass in University Valley was extremely low, and microbial activity under ambient conditions was undetectable. Our results contrast with reports from the lower-elevation Dry Valleys and Arctic permafrost soils where active microbial populations are found, suggesting that the combination of severe cold, aridity, oligotrophy of University Valley permafrost soils severely limit microbial activity and survival.

Cold adaptive traits revealed by comparative genomic analysis of the eurypsychrophile Rhodococcus sp. JG3 isolated from high elevation McMurdo Dry Valley permafrost, Antarctica

The permafrost soils of the high elevation McMurdo Dry Valleys are the most cold, desiccating and oligotrophic on Earth. Rhodococcus sp. JG3 is one of very few bacterial isolates from Antarctic Dry Valley permafrost, and displays subzero growth down to -5°C. To understand how Rhodococcus sp. JG3 is able to survive extreme permafrost conditions and be metabolically active at subzero temperatures, we sequenced its genome and compared it to the genomes of 14 mesophilic rhodococci. Rhodococcus sp. JG3 possessed a higher copy number of genes for general stress response, UV protection and protection from cold shock, osmotic stress and oxidative stress. We characterized genome wide molecular adaptations to cold, and identified genes that had amino acid compositions favourable for increased flexibility and functionality at low temperatures. Rhodococcus sp. JG3 possesses multiple complimentary strategies which may enable its survival in some of the harshest permafrost on Earth.

Comparative activity and functional ecology of permafrost soils and lithic niches in a hyper-arid polar desert.

Permafrost in the high elevation McMurdo Dry Valleys of Antarctica ranks among the driest and coldest on Earth. Permafrost soils appear to be largely inhospitable to active microbial life, but sandstone lithic microhabitats contain a trophically simple but functional cryptoendolithic community. We used metagenomic sequencing and activity assays to examine the functional capacity of permafrost soils and cryptoendolithic communities in University Valley, one of the most extreme regions in the Dry Valleys. We found metagenomic evidence that cryptoendolithic microorganisms are adapted to the harsh environment and capable of metabolic activity at in situ temperatures, possessing a suite of stress response and nutrient cycling genes to fix carbon under the fluctuating conditions that the sandstone rock would experience during the summer months. We additionally identified genes involved in microbial competition and cooperation within the cryptoendolithic habitat. In contrast, permafrost soils have a lower richness of stress response genes, and instead the metagenome is enriched in genes involved with dormancy and sporulation. The permafrost soils also have a large presence of phage genes and genes involved in the recycling of cellular material. Our results underlie two different habitability conditions under extreme cold and dryness: the permafrost soil which is enriched in traits which emphasize survival and dormancy, rather than growth and activity; and the cryptoendolithic environment that selects for organisms capable of growth under extremely oligotrophic, arid and cold conditions. This study represents the first metagenomic interrogation of Antarctic permafrost and polar cryptoendolithic microbial communities.

Left Out in the Cold: Life in Cryoenvironments

Cryoenvironments are generally defined as environments that exist continuously and predominately at subzero temperatures. They exist primarily in polar and alpine regions and consist of large-scale geomorphological features such as permafrost, glaciers, ice caps, and sea ice. Cryoenvironments also include relatively rare subzero habitats such as cold lakes and ponds, which can be permanently ice covered, and subzero saline springs, which flow throughout the year, warmed by geothermal gradients, and maintained liquid due to their high salinity (Andersen et al., 2002; Doyle et al., 2012). The primary constraint to life in cryoenvironments is the availability of liquid water; life needs liquid water to survive, mediate biochemical reactions, provide transport of molecules, and act as a solvent. It is not necessarily subzero temperatures that constrain life in cryoenvironments but rather the conditions that are typically found associated with subzero temperatures, which include freezing, desiccation, or high osmolarity. Microorganisms in subzero environments must, however, be able to cope with the thermodynamic effects of low temperatures including lower reaction rates, increased molecule stability, and conformational changes of proteins (Bakermans, 2008). Because the presence of liquid water in cryoenvironments is often facilitated through the freezing-point depression properties of various solutes, microorganisms must also be able to tolerate osmotic stress, usually in the form of high salinity. Despite these harsh environmental conditions, there is a recent and growing body of evidence that cryophilic microorganisms (those able to reproduce at <0 °C) exist and are metabolically active in these cryoenvironments at ambient temperatures.

Improved High-Quality Draft Genome Sequence of the Eurypsychrophile Rhodotorula sp. JG1b, Isolated from Permafrost in the Hyperarid Upper-Elevation McMurdo Dry Valleys, Antarctica

ABSTRACT Here, we report the draft genome sequence of Rhodotorula sp. strain JG1b, a yeast that was isolated from ice-cemented permafrost in the upper-elevation McMurdo Dry Valleys, Antarctica. The sequenced genome size is 19.39 Mb, consisting of 156 scaffolds and containing a total of 5,625 predicted genes. This is the first known cold-adapted Rhodotorula sp. sequenced to date.

Microbial Life in Antarctic Permafrost Environments

Permafrost is extensively distributed throughout terrestrial Antarctica, and contains soils that are the oldest, coldest, and driest on Earth. Currently, relatively little is known about the microbial life in Antarctic permafrost compared to its Arctic and Alpine permafrost counterparts. However, a growing body of evidence indicates that the Antarctic permafrost microbial community can be as diverse as those found in the Arctic, but soils have several orders of magnitude less biomass. Similar phyla of bacteria (Firmicutes, Actinobacteria, Bacteriodetes) are found and isolated in permafrost from both poles, and evidence to date from the Arctic indicate that permafrost is a habitable environment, in spite of the extreme cold, oligotrophy and low water activity associated with the soils. Of special interest in the Antarctic is the presence of dry permafrost soil that remains below 08 C and contains negligible amounts of water; such permafrost environments are found in the upper elevations of the McMurdo Dry Valleys, the only place on Earth where dry permafrost is known to occur. It is currently not known if active microbial communities can exist in such hyper-arid dry permafrost soils, and Antarctic permafrost in general. Questions that remain to be answered include whether permafrost microbial communities are active or viable in situ, or whether they represent cryopreserved fossils. If viable, what are the adaptations that allow life to be sustained over geologic timescales in permanently subzero environ- ments? And finally, are the cold and arid limits of microbial life found in the Dry Valley permafrost soils of the Antarctic?

Conserved genomic and amino acid traits of cold adaptation in subzero-growing Arctic permafrost bacteria

Abstract 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). Figure. No Caption available.

In Situ Field Sequencing and Life Detection in Remote (79°26′N) Canadian High Arctic Permafrost Ice Wedge Microbial Communities

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 the MinION. The MAM plate 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.

Microbial Life in Permafrost

Permafrost is a hostile environment that harbors a diverse and active microbial community. Next generation sequencing studies have demonstrated a wide diversity of microorganisms present in Arctic, Antarctic and high altitude permafrost soils. In situ activity of these microorganisms has been demonstrated through multiple lines of evidence. Radiolabeled studies and stable isotope probing have established that active respiration and DNA replication occur in permafrost soils under frozen conditions. Furthermore, microorganisms capable of subzero growth have been isolated from permafrost samples. These isolates have adapted to the permafrost environment through a multitude of molecular changes, such as increased expression of cold shock and metabolite transport proteins, reduced fatty acid saturation in the membrane, and presence of temperature specific isozymes. Recent studies have focused on permafrost thaw due to anthropogenic climate change. The subsequent thaw of frozen organic carbon stores in permafrost is thought to increase microbial activity and emissions of greenhouse gases to the atmosphere. As the permafrost thaws, the microbial community changes in terms of diversity and functional potential in response to warmer temperatures, and increased carbon and water availability.