Wayne H. Pollard

Microbial ecology and biodiversity in permafrost

Permafrost represents 26% of terrestrial soil ecosystems; yet its biology, essentially microbiology, remains relatively unexplored. The permafrost environment is considered extreme because indigenous microorganisms must survive prolonged exposure to subzero temperatures and background radiation for geological time scales in a habitat with low water activity and extremely low rates of nutrient and metabolite transfer. Yet considerable numbers and biodiversity of bacteria exist in permafrost, some of which may be among the most ancient viable life on Earth. This review describes the permafrost environment as a microbial habitat and reviews recent studies examining microbial biodiversity found in permafrost as well as microbial growth and activity at ambient in situ subzero temperatures. These investigations suggest that functional microbial ecosystems exist within the permafrost environment and may have important implications on global biogeochemical processes as well as the search for past or extant life in permafrost presumably present on Mars and other bodies in our solar system.

Microbial diversity and activity through a permafrost/ground ice core profile from the Canadian high Arctic

Culture-dependent and culture-independent methods were used in an investigation of the microbial diversity in a permafrost/massive ground ice core from the Canadian high Arctic. Denaturing gradient gel electrophoresis as well as Bacteria and Archaea 16S rRNA gene clone libraries showed differences in the composition of the microbial communities in the distinct core horizons. Microbial diversity was similar in the active layer (surface) soil, permafrost table and permafrost horizons while the ground ice microbial community showed low diversity. Bacteria and Archaea sequences related to the Actinobacteria (54%) and Crenarchaeota (100%) respectively were predominant in the active layer while the majority of sequences in the permafrost were related to the Proteobacteria (57%) and Euryarchaeota (76%). The most abundant phyla in the ground ice clone libraries were the Firmicutes (59%) and Crenarchaeota (82%). Isolates from the permafrost were both less abundant and diverse than in the active layer soil, while no culturable cells were recovered from the ground ice. Mineralization of [1-(14)C] acetic acid and [2-(14)C] glucose was used to detect microbial activity in the different horizons in the core. Mineralization was detected at near ambient permafrost temperatures (-15 degrees C), indicating that permafrost may harbour an active microbial population, while the low microbial diversity, abundance and activity in ground ice suggests a less hospitable microbial habitat.

Formation of Martian gullies by the action of liquid water flowing under current Martian environmental conditions

[1] Geomorphic evidence suggests that recent gullies on Mars were formed by fluvial activity. The Martian gully features are significant because their existence implies the presence of liquid water near the surface on Mars in geologically recent times. Irrespective of the ultimate source of the fluid carving the gullies, we seek to understand the behavior of this fluid after it reaches the Martian surface. We find that contrary to popular belief, the fluvially carved Martian gullies are consistent with formation conditions such as now occur on Mars, outside of the temperature-pressure stability regime of liquid water. Our model of the action of flowing pure liquid water produces the observed gully length distribution only at surface pressures and temperatures below the triple point where liquid water simultaneously boils and freezes and thus suggests that gullies were formed under conditions similar to present-day Mars. Our results suggest a typical flow rate of 30 m 3 /s to carve the gully channels. At least 0.15 km 3 has flowed across the surface of Mars to carvethegullysystemsobservedtoday,andthiswouldrequireanaquifer recharge rate of � 10 � 13 –10 � 12 m/yr. The absence of gullies on Mars that are long enough to have been created above the triple point pressure argues that the atmospheric pressure has not been significantly larger than it is now since the origin of the gullies. This result may imply that Mars does not possess a significant reservoir of condensed CO2.

Characterization of the Prokaryotic Diversity in Cold Saline Perennial Springs of the Canadian High Arctic

ABSTRACT The springs at Gypsum Hill and Colour Peak on Axel Heiberg Island in the Canadian Arctic originate from deep salt aquifers and are among the few known examples of cold springs in thick permafrost on Earth. The springs discharge cold anoxic brines (7.5 to 15.8% salts), with a mean oxidoreduction potential of −325 mV, and contain high concentrations of sulfate and sulfide. We surveyed the microbial diversity in the sediments of seven springs by denaturing gradient gel electrophoresis (DGGE) and analyzing clone libraries of 16S rRNA genes amplified with Bacteria and Archaea-specific primers. Dendrogram analysis of the DGGE banding patterns divided the springs into two clusters based on their geographic origin. Bacterial 16S rRNA clone sequences from the Gypsum Hill library (spring GH-4) were classified into seven phyla (Actinobacteria, Bacteroidetes, Firmicutes, Gemmatimonadetes, Proteobacteria, Spirochaetes, and Verrucomicrobia); Deltaproteobacteria and Gammaproteobacteria sequences represented half of the clone library. Sequences related to Proteobacteria (82%), Firmicutes (9%), and Bacteroidetes (6%) constituted 97% of the bacterial clone library from Colour Peak (spring CP-1). Most GH-4 archaeal clone sequences (79%) were related to the Crenarchaeota while half of the CP-1 sequences were related to orders Halobacteriales and Methanosarcinales of the Euryarchaeota. Sequences related to the sulfur-oxidizing bacterium Thiomicrospira psychrophila dominated both the GH-4 (19%) and CP-1 (45%) bacterial libraries, and 56 to 76% of the bacterial sequences were from potential sulfur-metabolizing bacteria. These results suggest that the utilization and cycling of sulfur compounds may play a major role in the energy production and maintenance of microbial communities in these unique, cold environments.

Gradient analysis of cryoconite ecosystems from two polar glaciers

The cylindrical meltholes present in the ablation zones of many glaciers (termed cryoconite holes) contain complex microbial communities. A canonical correspondence analysis (CCA) of community structure and environmental gradients for cryoconite holes on two glaciers was undertaken. The Canada Glacier (77°37′S, 162°55′E) is located in the McMurdo Dry Valleys of Antarctica. The White Glacier (79°27′N, 90°40′W) is located on Axel Heiberg Island, Nunavut Territory, Canada. These glaciers are at similar, yet antipodal latitudes, are roughly the same size and endure approximately the same mean annual temperature. The Canada Glacier cryoconite communities were found to be significantly (P=0.001) associated with six environmental variables, which together explained 55% of the biological variation. The White Glacier cryoconite communities were not significantly associated with environmental variables. The differences in CCA results were attributed to the relative amount of disturbance and isolation between each glacier’s cryoconite holes. Canada Glacier cryoconite holes were mostly ice-covered and undisturbed by meltwater flow, whereas high meltwater production and open cryoconite holes on the White Glacier may continually reset the community structure and habitat variability due to inter-hole mixing.

Utilization of fluorescent microspheres and a green fluorescent protein-marked strain for assessment of microbiological contamination of permafrost and ground ice core samples from the Canadian High Arctic.

ABSTRACT Fluorescent microspheres were applied in a novel fashion during subsurface drilling of permafrost and ground ice in the Canadian High Arctic to monitor the exogenous microbiological contamination of core samples obtained during the drilling process. Prior to each drill run, a concentrated fluorescent microsphere (0.5-μm diameter) solution was applied to the interior surfaces of the drill bit, core catcher, and core tube and allowed to dry. Macroscopic examination in the field demonstrated reliable transfer of the microspheres to core samples, while detailed microscopic examination revealed penetration levels of less than 1 cm from the core exterior. To monitor for microbial contamination during downstream processing of the permafrost and ground ice cores, a Pseudomonas strain expressing the green fluorescent protein (GFP) was painted on the core exterior prior to processing. Contamination of the processed core interiors with the GFP-expressing strain was not detected by culturing the samples or by PCR to detect the gfp marker gene. These methodologies were quick, were easy to apply, and should help to monitor the exogenous microbiological contamination of pristine permafrost and ground ice samples for downstream culture-dependent and culture-independent microbial analyses.

Methane‐cycling communities in a permafrost‐affected soil on Herschel Island, Western Canadian Arctic: active layer profiling of mcrA and pmoA genes

In Arctic wet tundra, microbial controls on organic matter decomposition are likely to be altered as a result of climatic disruption. Here, we present a study on the activity, diversity and vertical distribution of methane-cycling microbial communities in the active layer of wet polygonal tundra on Herschel Island. We recorded potential methane production rates from 5 to 40 nmol h(-1) g(-1) wet soil at 10 °C and significantly higher methane oxidation rates reaching values of 6-10 μmol h(-1) g(-1) wet soil. Terminal restriction fragment length polymorphism (T-RFLP) and cloning analyses of mcrA and pmoA genes demonstrated that both communities were stratified along the active layer vertical profile. Similar to other wet Arctic tundra, the methanogenic community hosted hydrogenotrophic (Methanobacterium) as well as acetoclastic (Methanosarcina and Methanosaeta) members. A pronounced shift toward a dominance of acetoclastic methanogens was observed in deeper soil layers. In contrast to related circum-Arctic studies, the methane-oxidizing (methanotrophic) community on Herschel Island was dominated by members of the type II group (Methylocystis, Methylosinus, and a cluster related to Methylocapsa). The present study represents the first on methane-cycling communities in the Canadian Western Arctic, thus advancing our understanding of these communities in a changing Arctic.

Microbial characterization of a subzero, hypersaline methane seep in the Canadian High Arctic.

We report the first microbiological characterization of a terrestrial methane seep in a cryo-environment in the form of an Arctic hypersaline (∼24% salinity), subzero (−5 °C), perennial spring, arising through thick permafrost in an area with an average annual air temperature of −15 °C. Bacterial and archaeal 16S rRNA gene clone libraries indicated a relatively low diversity of phylotypes within the spring sediment (Shannon index values of 1.65 and 1.39, respectively). Bacterial phylotypes were related to microorganisms such as Loktanella, Gillisia, Halomonas and Marinobacter spp. previously recovered from cold, saline habitats. A proportion of the bacterial phylotypes were cultured, including Marinobacter and Halomonas, with all isolates capable of growth at the in situ temperature (−5 °C). Archaeal phylotypes were related to signatures from hypersaline deep-sea methane-seep sediments and were dominated by the anaerobic methane group 1a (ANME-1a) clade of anaerobic methane oxidizing archaea. CARD-FISH analyses indicated that cells within the spring sediment consisted of ∼84.0% bacterial and 3.8% archaeal cells with ANME-1 cells accounting for most of the archaeal cells. The major gas discharging from the spring was methane (∼50%) with the low CH4/C2+ ratio and hydrogen and carbon isotope signatures consistent with a thermogenic origin of the methane. Overall, this hypersaline, subzero environment supports a viable microbial community capable of activity at in situ temperature and where methane may behave as an energy and carbon source for sustaining anaerobic oxidation of methane-based microbial metabolism. This site also provides a model of how a methane seep can form in a cryo-environment as well as a mechanism for the hypothesized Martian methane plumes.

The Icebreaker Life Mission to Mars: a search for biomolecular evidence for life.

The search for evidence of life on Mars is the primary motivation for the exploration of that planet. The results from previous missions, and the Phoenix mission in particular, indicate that the ice-cemented ground in the north polar plains is likely to be the most recently habitable place that is currently known on Mars. The near-surface ice likely provided adequate water activity during periods of high obliquity, ≈ 5 Myr ago. Carbon dioxide and nitrogen are present in the atmosphere, and nitrates may be present in the soil. Perchlorate in the soil together with iron in basaltic rock provides a possible energy source for life. Furthermore, the presence of organics must once again be considered, as the results of the Viking GCMS are now suspect given the discovery of the thermally reactive perchlorate. Ground ice may provide a way to preserve organic molecules for extended periods of time, especially organic biomarkers. The Mars Icebreaker Life mission focuses on the following science goals: (1) Search for specific biomolecules that would be conclusive evidence of life. (2) Perform a general search for organic molecules in the ground ice. (3) Determine the processes of ground ice formation and the role of liquid water. (4) Understand the mechanical properties of the martian polar ice-cemented soil. (5) Assess the recent habitability of the environment with respect to required elements to support life, energy sources, and possible toxic elements. (6) Compare the elemental composition of the northern plains with midlatitude sites. The Icebreaker Life payload has been designed around the Phoenix spacecraft and is targeted to a site near the Phoenix landing site. However, the Icebreaker payload could be supported on other Mars landing systems. Preliminary studies of the SpaceX Dragon lander show that it could support the Icebreaker payload for a landing either at the Phoenix site or at midlatitudes. Duplicate samples could be cached as a target for possible return by a Mars Sample Return mission. If the samples were shown to contain organic biomarkers, interest in returning them to Earth would be high.

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.