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Metagenomics Study Tracks Microbial Community During Permafrost Thaw

By Andrea Anderson

NEW YORK (GenomeWeb News) – In a study appearing online yesterday in Nature, an American research team used metagenomic sequencing to not only look at the microbes, genes, and pathways found in Alaskan permafrost samples, but also to track the changes that occur in these communities once the permafrost thaws.

Because the site tested is predicted to experience warmer temperatures and increased permafrost thawing, those involved in the study hope to learn more about the interplay between the microbes in the long-frozen soil and organic carbon cycling both before and after thawing — information that may help produce more complete climate models.

"Currently in climate models, it's not really taken into account adequately what the microorganisms are doing," senior author Janet Jansson, a researcher affiliated with the Lawrence Berkeley National Laboratory and the US Department of Energy's Joint Genome Institute, told GenomeWeb Daily News. "The hope is to get enough information at the microscopic level that we'll have enough understanding to be able to inform climate models."

For the current study, the team also was able to use sequences from metagenomic data to stitch together the draft genome sequence of an uncharacterized microbe suspected of producing much of the methane released by the thawing permafrost samples.

"We consider this a very likely candidate for the methane production that we observed in our samples," Jansson said.

Relatively little is known about the microbial communities found in permafrost and other soils, since most of the microbes involved cannot be cultured in the lab. The microbes found in permafrost are particularly interesting, Jansson explained, since this soil has been frozen for so long and typically contains a great deal of organic carbon.

"In addition to the microbes, there's a lot of organic carbon that is kept in permafrost," she noted. "So, what do [the microbes] do when that carbon becomes accessible for degradation?"

To try to learn more about such interactions, the team tested three core samples collected from a black spruce forest site in Hess Creek, Alaska. The cores were about one meter (3.3 feet) deep. The upper portion of each core was comprised of soil from the so-called active layer, Jansson explained, which freezes and thaws seasonally. Below that is a permafrost layer that remains frozen year round.

In an effort to characterize the composition and functional capabilities of the microbial communities in the active layer and permafrost when these soils thaw, the researchers incubated parts of each core in the lab at 5°C (41°F) while gauging the levels of methane and carbon dioxide released by the samples.

Overall, they found that permafrost thawing corresponded to a burst of methane release, apparently stemming from methane that had been produced by so-called methogen microbes over time in the frozen soil.

Those methane levels declined after a week at the warmer temperature as other microbes slurped up the newly released methane — consumption that appears to have been accomplished by methane oxidizing methanotrophic bacteria, based on the team's experiments.

While the general patterns were similar in the different samples, though, there were also many differences between the two permafrost samples tested by sequencing.

Once the samples had thawed for two days and seven days, the researchers extracted DNA from the active and permafrost layers of two of the three core samples before and after thawing. They then used Roche 454 GS FLX sequencing of 16S rRNA to get an idea of the phylogeny of the bacteria and archaea present in the samples, along with metagenome sequencing with the Illumina GAII to look at the genes and pathways present.

Although the team found that a stress and starvation resistant group of bugs called Actinobacteria were abundant in both samples, the broader microbial communities differed in their phylogeny and gene repertoires.

"When we looked at the two permafrost samples, even though they were from the same location just a few meters apart from each other, they were very different," Jansson said. "The microorganisms were different and the pathways were very different."

Though this was not the pattern researchers initially expected, she added, such between-sample differences may not be that surprising given that the permafrost has been frozen so long, with each sampling site likely having had access to slightly different carbon sources over that time.

On the other hand, the team found that the metagenomic profiles in the permafrost samples became far more similar to one another and to the active layer samples after just two days of thawing.

"These two cores were very different, but when they thawed they started to converge," Jansson said. "The pathways and the microorganisms started to become more similar to each other and much more similar to the active layer — and it happened very quickly."

In contrast, the microbial communities detected in the active layers didn't change all that much once the samples thawed out, she explained, perhaps owing to the seasonal freezing and thawing that this soil had experienced.

"Every year they experience a thaw and a freeze and so they're adapted to that sort of a process," Jansson said, "whereas the microorganisms that were frozen for so long [in the permafrost] were not used to being thawed, so they had a much bigger shift."

Along with their analyses of the microbial changes during permafrost thawing, sequences from a combined assembly of metagenomic sequences from the permafrost gave the researchers the opportunity to put together the 1.9 million base draft genome sequence of a previously uncharacterized methanogen.

"A lot of the larger contigs corresponded to this methanogen. And when we binned the contigs we found that we had basically a draft genome of the methanogen," Jansson explained. "We didn't expect that. We weren't looking for it."

Though there appear to be other methanogens in the samples, the newly sequenced methanogen is quite different from any species characterized in the past, sharing just 65 percent identity with the most closely related known methanogen.

To follow up on their current findings, the researchers are now doing RNA and protein analyses to try to determine which of the genes in the permafrost metagenome are actually expressed while permafrost samples thaw and/or are incubated with different compounds.

"Since we did find this difference in carbon processing potential in the different permafrost cores, we're adding substrates to seeing how those are turned over and then looking at the expression of RNA and protein," Jansson said.

They are also planning to look at samples from other sites in Alaska that are known to have different levels of mineral and organic matter compared with the samples tested so far, including samples from a peat-like bog that has thawed naturally.

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