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Project Progress

The overall goal of this project is to provide “Field Truth” about microbial biology in permafrost, including understanding the current composition of the Arctic and Antarctic permafrost microbial community, whether it is active in situ, and once genomic information suggests genes of importance to the cryo-environment, to test the importance of those genes or markers in situ. Two distinct environments were examined: Arctic (permafrost under typical subarctic tundra, near the East Siberian Sea coast, Russia) and Antarctic (permafrost of the Beacon Dry Valley, Antarctica). We determined both indigenous microbes and those which could grow aerobically and anaerobically at 10^ºC in their native soil by polymerase chain reaction (PCR) amplification, cloning and sequencing of 16SrRNA genes found in the soil DNA. To do this we employed high-throughput sequencing and wrote script to check for sequence quality, and automatically compared the sequence to rrn sequences in the Ribosomal Database Project (RDP). Sequences were grouped into categories from the RDP’s phylogenetic hierarchy based on the closest database match. Analysis of this library of over 2000 clones revealed over 380 bacterial species, although the Proteobacteria and Gram Positive Bacteria were the most common Divisions found. Importantly, spore-forming bacteria were minor members of the community, representing 10 to 23% of the total clones in all of the 1.8 to 8 million year old permafrost samples. Similar phylogenic groups appeared in both Polar Regions, and some of these phylotypes have members that are known to live in other cold or frozen environments. About 170 clones are novel (with high quality sequence of from 500 to 800 bp, and low similarity% with a closest RDP Data Base sequence).

To evaluate whether the permafrost microbes are active at the in situ temperatures of -9 to -11^ºC, we measured ¹⁴CH₄ production from ¹⁴C-bicarbonate and acetate and ¹⁴C assimilation into microbial biomass in permafrost samples incubated in the laboratory at temperatures down to -15^ºC. We found ¹⁴C methane production in a peaty sediment collected 80 cm below the permafrost surface that had been frozen for 3000 years. We also isolated a psychrophilic methanogen, a Methanosarcina spp., from this site. The results are of interest to astrobiology since they show a viable chemolithotrophic psychrotolerant anaerobic bacterial community that is a reasonable model for extraterrestrial life since they assimilate CO₂and other simple compounds that might be found in frozen subsurface environments on cryogenic planets without free oxygen, inaccessible organic matter and extremely limited water activity. The second evidence of microbial activity by indigenous microbes at in situ temperatures comes from lenses of overcooled brines (cryopeds, salinity 170-300 g/l) found at depths of 40 to 50 m within permafrost sandy soils of marine origin, dating back to 100-120 thousand years BP. These are the only hydrological systems on the Earth with average annual temperatures below zero, high salinity, and isolation from external factors throughout their geologic history. Our microbiological investigation showed that these samples contain psychrophilic-halotolerant microorganisms by assimilation of ¹⁴C-D-glucose into cells, a population density of 3.3×10⁷ to 6.7×10⁷ cells/ml brine water, enrichment at 5^ºC of the major anaerobic physiological groups (heterotrophs, methanogens, sulfate reducers, and acetogens) and yielded an isolate identified as a Psychrobacter, which can grow at subzero temperatures. These brines are considered good models for Martian life since Martian free water may occur as brine lenses in permafrost that were formed when Mars lost its atmosphere and became dry and cold.