RNA-stable isotope probing: from carbon flow within key microbiota to targeted transcriptomes

Lueders, Tillmann; Dumont, Marc G.; Bradford, Lauren; Manefield, Mike

doi:10.1016/j.copbio.2016.05.001

Cited by 45 publications

(29 citation statements)

References 51 publications

(66 reference statements)

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“…The SIP method produces a bias missing the species that are profoundly dependent on such conditions. Nevertheless, SIP presents an effective, molecular-based metabolism studying tool which is not dependent on growth as much as on cell-level reactions [35]. These reactions require the synthesis and transfer of labelled carbon to nucleic acids in adequate levels to allow the separation from un-labelled background nucleic acids [42,90].…”

Section: Discussionmentioning

confidence: 99%

“…In the Outokumpu subsurface sulfate reducers have been detected at various depths [21], and enrichment of acetate-consuming microbial communities from the depth of 967 m was observed with acetate amended microcosms [30].The great majority of deep biosphere microorganisms remain uncultivable in laboratory conditions, setting limits to research on their energy and carbon sources. Stable isotope probing (SIP), first introduced by Radajewski et al [34], is an advantageous method in the analysis of metabolism routes in the environment and characterization of species linked to assimilation of the studied substrate [35]. SIP has been successfully used on a broad scale in environmental microbiology, revealing the species linked to actively performed functions [36][37][38][39].Acetate can support methanogens, and thus, the compounds formed can activate further microbial groups.…”

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confidence: 99%

“…The great majority of deep biosphere microorganisms remain uncultivable in laboratory conditions, setting limits to research on their energy and carbon sources. Stable isotope probing (SIP), first introduced by Radajewski et al [34], is an advantageous method in the analysis of metabolism routes in the environment and characterization of species linked to assimilation of the studied substrate [35]. SIP has been successfully used on a broad scale in environmental microbiology, revealing the species linked to actively performed functions [36][37][38][39].…”

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Rare Biosphere Archaea Assimilate Acetate in Precambrian Terrestrial Subsurface at 2.2 km Depth

et al. 2018

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The deep biosphere contains a large portion of the total microbial communities on Earth, but little is known about the carbon sources that support deep life. In this study, we used Stable Isotope Probing (SIP) and high throughput amplicon sequencing to identify the acetate assimilating microbial communities at 2260 m depth in the bedrock of Outokumpu, Finland. The long-term and short-term effects of acetate on the microbial communities were assessed by DNA-targeted SIP and RNA targeted cell activation. The microbial communities reacted within hours to the amended acetate. Archaeal taxa representing the rare biosphere at 2260 m depth were identified and linked to the cycling of acetate, and were shown to have an impact on the functions and activity of the microbial communities in general through small key carbon compounds. The major archaeal lineages identified to assimilate acetate and metabolites derived from the labelled acetate were Methanosarcina spp., Methanococcus spp., Methanolobus spp., and unclassified Methanosarcinaceae. These archaea have previously been detected in the Outokumpu deep subsurface as minor groups. Nevertheless, their involvement in the assimilation of acetate and secretion of metabolites derived from acetate indicated an important role in the supporting of the whole community in the deep subsurface, where carbon sources are limited.Geosciences 2018, 8, 418 2 of 20 utilizing soluble gases, such as crustal methane or hydrogen from radiolytic decomposition of water, have been made since [5][6][7][8]. More specifically, the metabolic capabilities of deep terrestrial subsurface microbial communities include the pathways covering, e.g., methanogenesis [9][10][11], anaerobic methane oxidation [12], acetogenesis [4,13], sulfate reduction [14-16], oxidation of sulfur by denitrification [17], and reduction of iron [14].The Outokumpu deep scientific drill hole, located in Eastern Finland, hosts a unique environment piercing through crystalline Precambrian bedrock and its numerous saline fluid filled fracture zones. The composition of the microbial communities and the geochemical characteristics of the Outokumpu deep subsurface have been studied in detail [11,[18][19][20][21][22][23][24][25][26][27]. The metabolic strategies of the microbial communities at different depths in Outokumpu have been estimated through functional gene analysis and metagenomes [11,21]. Purkamo et al. [21,22] suggested that microbial communities commonly employ heterotrophic carbon assimilation in the Outokumpu deep subsurface, and that the communities have the capacity of both sulphate and nitrate reduction. At greater depths, genetic potential for autotrophic acetogenesis and hydrogenotrophic methanogenesis has been detected from Outokumpu metagenomes together with H 2 oxidation and CO 2 fixation coupling hydrogenases [11].The terrestrial deep subsurface represents an oligotrophic, nutrient-poor, and isolated habitat. The fact that heterotrophy appears to be common in the Outokumpu deep biosphere means that the carbon sub...

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Section: Discussionmentioning

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Rare Biosphere Archaea Assimilate Acetate in Precambrian Terrestrial Subsurface at 2.2 km Depth

et al. 2018

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“…Stable isotopes can be combined with molecular techniques through DNA-, RNA-, and protein stable isotope probing (SIP) to reveal information about the phylogeny and activity of specific organisms responsible for the transformation of a particular substrate. DNA-SIP requires cellular growth for labelled elements to be incorporated into the DNA and subsequently detected (Lueders et al, 2016). Due to variations in density based on G-C content, labelled samples must be compared to unlabeled controls (Youngblut and Buckley, 2014).…”

Section: Mapping Substrate Utilizationmentioning

confidence: 99%

“…In AD, DNA-SIP has been used to identify the acetoclastic methanogens that dominate under high ammonia conditions (Hao et al, 2015) and identify cellulose degraders (Li et al, 2009;Limam et al, 2014). RNA-SIP, in contrast to DNA-SIP, can be used to track labels in both rRNA and mRNA and does not require cellular replication or growth (Lueders et al, 2016). Applied to AD, RNA-SIP has been used to trace labelled glucose through glucose-, propionate-, and acetate-degrading bacteria and acetoclastic methanogens (Ito et al, 2012;Ito et al, 2011) and reveal the diversity of fatty acid degrading bacteria (Hatamoto et al, 2007).…”

Section: Mapping Substrate Utilizationmentioning

confidence: 99%