Culture Independent Bacterial and Archaeal Diversity in Permafrost

Culture-independent methodologies have recently been applied to the study of microbial diversity in permafrost. These studies, which use molecular-based tools to analyze DNA extracted directly from permafrost (Spiegelman et al. 2005 and references therein), bypass the need for culturing and have increased the number of phylogenetic groups of Bacteria and Archaea associated with permafrost (Table 5.3). For example, the culturable microbial community in a Canadian high Arctic permafrost sample was dominated by Firmicutes-related isolates, whereas Actinobacteria-and Proteobacteria-related sequences were predominant in a culture-independent analysis, with the phyla Gemmatimonadetes, CFB and Planctomyces identified in the culture-independent survey but not among the isolates (Steven et al. 2007a). A diverse Bacteria community, comprised of 13 Bacteria phyla (Table 5.3), including three candidate phyla (phyla that have no cultured representatives), was detected in Spitsbergen Island permafrost 16S rRNA gene clone libraries (Hansen et al. 2007), while only four phyla (Actinobacteria, CFB, Firmicutes and Proteobacteria) were represented by cultured isolates (Hansen et al. 2007). 16S rRNA gene clone libraries constructed from Siberian permafrost DNA (Table 5.3) were dominated by sequences related to the Proteobacteria, Actinobacteria and Firmicutes, with Arthrobacter being abundant in both the culture-dependent and culture-independent surveys of microbial diversity (Vishnivetskaya et al. 2006). The proportion of 16S rRNA sequences related to the high G + C Gram-positive Bacteria was also found to increase with increasing age of Siberian permafrost (Willerslev et al. 2004a). Antarctic Dry Valley permafrost 16S rRNA gene clone libraries were composed of the phylogenetic groups Proteobacteria and Actinobacteria, with Arthrobacter, Bacillus, and Pseudomonas detected in all of the Antarctic permafrost clone libraries (Gilichinsky et al. 2007).

To date, very few studies have described the Archaea communities in permafrost using culture-independent methodologies. Other than a report of the detection of 16S rRNA genes related to the Crenarchaeota (affiliated to environmental group 1.1.b) in Chinese alpine permafrost (Ochsenreiter et al. 2003), all of the culture-independent characterizations of Archaea diversity in permafrost are from the Canadian high

Table 5.3 Phylogenetic groups of Bacteria and Archaea detected by culture-independent methods in various permafrost environments

Canadian high Canadian

Table 5.3 Phylogenetic groups of Bacteria and Archaea detected by culture-independent methods in various permafrost environments

Canadian high Canadian

Arctic

Kolyma

Spitsb-

Dry

Alaskan

high Arctic

Phylogenetic

perma-

lowlands

ergen

Valleys

ice

massive

group

frosta,b

Siberia0

Islandd

Antarcticae

wedgef

ground iceb

Bacteria

Acidobacteria

+

+

+

Actinobacteria

+

+

+

+

+

+

CFB

+

+

+

+

Firmicutes

+

+

+

+

+

+

Gemmatimonadetes

+

Planctomyces

+

+

Proteobacteria

+

+

+

+

+

+

Spirochaetes

+

Thermomicrobia

+

Verrucomicrobiae

+

OD1g

+

0P10g

+

TM7g

+

Unclassified

+

+

Archaea

Environmental

+

+

Crenarchaeota

Environmental

+

+

Euryarchaeota

Halophilic Archaea

+

+

Methanogenic

Archaea aSteven et al. (2007a) ┬╗Steven et al. (2007b) cVishnivetskaya et al. (2006) dHansen et al. (2007) eGilichinsky et al. (2007) fKatayama et al. (2007)

gCandidate divisions for which there are no cultured representatives

Arctic. Our studies have revealed that both of the major Archaea phyla (Euryarchaeota and Crenarchaeota) are present in Canadian permafrost, with sequences belonging to the Euryarchaeota being numerically dominant (Steven et al. 2007a, 2008a). Although methanogens have been isolated from Antarctic and Siberian permafrost (Rivkina et al. 1998; Gilichinsky et al. 2007), 16S rRNA gene sequences related to methanogenic Archaea were not detected in Canadian high Arctic permafrost, with the exception of a single sequence detected in a massive ground ice deposit (Steven et al. 2008a). An interesting result of the culture-independent characterization of Archaea communities in Canadian high Arctic permafrost was the detection of a significant number of sequences related to the halophilic Archaea, although the salinity in the permafrost was only moderate (Steven et al. 2007a, 2008a). The detection of halophilic organisms in only moderately saline permafrost provides circumstantial evidence that the primary microbial habitat in permafrost exists as thin saline liquid water veins surrounding soil particles (Price 2007).

It should be noted that the detection of a DNA sequence is not conclusive evidence that the phylogenetically related organism is active or even viable in permafrost, as the constant subzero temperatures are ideal for DNA preservation (Willerslev et al. 2003, 2004a; see Chap. 4). Thus, developing novel methods will be essential to determine if microorganisms identified in culture-independent surveys exist as viable cells or are the microbial equivalent of mammoths, frozen in time in the permafrost environment.

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