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Soil Biology & Biochemistry 111 (2017) 66e77

Contents lists available at ScienceDirect

Soil Biology & Biochemistry

journal homepage: www.elsevier.com/locate/soilbio

In-depth analysis of core methanogenic communities from high elevation permafrost-affected wetlands

,

b

  • ,
  • ,

Sizhong Yang a 1, Susanne Liebner a 1, Matthias Winkel a, Mashal Alawi a, Fabian Horn a,

  • c
  • d

e,

Corina Dorfer , Julien Ollivier , Jin-sheng He f, Huijun Jin b, Peter Kühn c,

€

Michael Schloter d, Thomas Scholten c, Dirk Wagner a

,

*

a GFZ German Research Centre for Geosciences, Section 5.3 Geomicrobiology, Telegrafenberg, 14473 Potsdam, Germany b State Key Laboratory of Frozen Soils Engineering, Northwest Institute of Eco-Environment and Resources, Chinese Academy of Sciences, 730000, Lanzhou, China c University of Tübingen, Department of Geosciences, 72074 Tübingen, Germany d Helmholtz Zentrum München, Research Unit Comparative Microbiome Analysis, 85764 Neuherberg, Germany e Key Laboratory of Adaptation and Evolution of Plateau Biota, Northwest Institute of Plateau Biology, Chinese Academy of Sciences, 810008, Xi'ning, China f Peking University, Department of Ecology, College of Urban and Environmental Sciences, 100871, Beijing, China

  • a r t i c l e i n f o
  • a b s t r a c t

Article history:

Received 21 December 2016 Accepted 15 March 2017

The organic carbon of permafrost affected soils is receiving particular attention with respect to its fate and potential feedback to global warming. The structural and activity changes of methanogenic communities in the degrading permafrost-affected wetlands on the Tibetan Plateau can serve as fundamental elements for modelling feedback interaction of ecosystems to climate change. Hence, we aimed at anticipating if and how the rapid environmental changes occurring especially on the high altitude Tibetan platform will affect methanogenic communities. We identified methanogenic community composition, activity and abundance in wetland soils with different hydrological settings, permafrost extent and soil properties and pinpoint the environmental controls. We show that despite a pronounced natural gradient, the Tibetan high elevation wetland soils host a large methanogenic core microbiome. Hydrogenotrophic methanogens, in particular Methanoregula, and H2-dependent methanogenesis were overall dominant although acetoclastic methanogens in addition to hydrogenotrophs were among the dominating taxa in a minerotrophic fen. Tracing the Methanoregula community of the Tibetan Plateau using public databases revealed its global relevance in natural terrestrial habitats. Unlike the composition, the activity and abundance of methanogens varied strongly in the studied soils with higher values in alpine swamps than in alpine meadows. This study indicates that in the course of current wetland and permafrost degradation and the loss in soil moisture, a decrease in the methane production potential is expected on the high Tibetan Plateau but it will not lead to pronounced changes within the methanogenic community structure.

Keywords:

Methanogenic archaea Wetland

mcrA gene

Frozen ground Tibetan Plateau Biogeography

Methanoregula

© 2017 The Authors. Published by Elsevier Ltd. This is an open access article under the CC BY-NC-ND

license (http://creativecommons.org/licenses/by-nc-nd/4.0/).

1. Introduction

The plateau is warming at a rate approximately two times higher than the global average since the 1950s (IPCC, 2007), and even
The Qinghai-Tibet Plateau (QTP), also known as Tibetan Plateau, is the largest altitudinal permafrost unit on Earth. The lower elevation areas (<4000 m) are influenced by seasonally frozen ground, while on the high altitude plateau permafrost is more developed in depth, continuity and coverage (Zhou et al., 2000). faster at higher elevations (Wei and Fang, 2013). Permafrost degradation has occurred on the plateau during the last few decades, manifested by areal decrease of continuous and discontinuous permafrost, thinning of permafrost, shrinkage of isolated patches of permafrost and changes into seasonally frozen ground

(Cheng and Wu, 2007; Jin et al., 2009; Wu et al., 2015). Around

18.6% of its permafrost has degraded in the past 30 years and up to 46% permafrost will disappear in 100 years (Cheng and Wu, 2007; Cheng and Jin, 2013). Degradation of permafrost has led to a

* Corresponding author.

E-mail address: [email protected] (D. Wagner).

Authors contributed equally to the manuscript.

1

http://dx.doi.org/10.1016/j.soilbio.2017.03.007

0038-0717/© 2017 The Authors. Published by Elsevier Ltd. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).

S. Yang et al. / Soil Biology & Biochemistry 111 (2017) 66e77

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lowering of ground water levels, shrinking lakes and wetlands, and changes of grassland ecosystems from alpine meadows to steppes

(Jin et al., 2009; Cheng and Jin, 2013).

abundances, and significantly different composition in methanogenic communities. The objective of this research is to identify methanogenic community composition, activity and abundance in wetland soils with different hydrological settings (swamp and meadow), permafrost extent and soil properties, and to pinpoint the underlain environmental controls. Thereby, this study is the first to assess methanogenic communities on the Tibetan Plateau via deep sequencing on the functional gene of methanogenesis (methyl coenzyme M reductase A, mcrA) with a specific attention on the direct and indirect interactions between microbial taxa coexisting in the environment.
Besides rivers and lakes, wetlands such as alpine swamps and meadows are fed mainly through precipitation and occur together with frozen ground. Alpine swamps and meadows occupy a large

area of the eastern plateau (Zhou et al., 2000; Wang et al., 2016).

While water in alpine meadows flows slowly through shallow flooded zones, the water in swamps is typically stagnant (Wang et al., 2016). The Tibetan wetlands altogether occupy 50% of the total wetland area of China (Ding and Cai, 2007), with the majority (85%) distributed in the headwater regions of the Yangtze and Yellow Rivers at higher elevations (>4000 m) and the Zoige (Ruoergai) Peat Plateau at lower altitudes (<4000 m; Zhang et al., 2011). It was estimated that 7.4 Pg C was stored at the top one meter of the whole alpine grasslands (Yang et al., 2008). Although there is no specific carbon estimate for the Tibetan wetlands, the soil organic density of wetlands was in general far higher than that of the alpine steppe and of the plateau's average, highlighting the alpine wetlands as important carbon pool with the potential for

positive climate feedback (Ding and Cai, 2007; Yang et al., 2008;

Ding et al., 2016). As a result of climate warming and permafrost degradation, the area of Tibetan wetlands reduced by about 8% from 1970 to 2006 (Zhao et al., 2015). Due to hydrological deterioration, degeneration from wetlands to meadows or from meadows to steppes have been observed at local scales (Jin et al., 2009; Brierley et al., 2016), which has subsequently impaired their roles in regulating the flow of rivers and carbon stores (Cheng and Jin,

2013).

The greenhouse gas methane is a major end product of the microbial degradation of organic matter under anaerobic conditions. Wetlands contribute 70% to the global total emission of methane and therefore are a major research focus (Bridgham et al., 2013). Moreover, ice core records showed that the variations in the atmospheric CH4 content were consistent with the areal change of wetlands (Blunier et al., 1995). The methane fluxes on the Tibetan wetlands range from 9.6 to 214 mg CH4 mÀ2 dÀ1 (Jin et al., 1999;

Hirota et al., 2004; Cao et al., 2008; Chen et al., 2013) and are

generally comparable to Arctic permafrost regions (Wille et al., 2008; Sachs et al., 2010; also see Fig. S1). Both alpine swamps and meadows are thus potential hotspot of methane emission on the plateau.
Methanogenic archaea (methanogens) are responsible for the biological production of methane (methanogenesis). They commonly use H2/CO2 (hydrogenotrophic) and acetate (acetoclastic) as substrates under anaerobic conditions (Wagner and Liebner, 2009). Methanogens on the Tibetan Plateau were studied

in lakes (Liu et al., 2013) and wetland soils (Zhang et al., 2008; Deng et al., 2014; Cui et al., 2015; Tian et al., 2015). Most previous studies

on wetlands were confined to the Zoige Plateau on the eastern margin of the plateau at lower elevation (3400e3600 m a.s.l.). In this region methanogenic community structure, in contrast to methane production, is little responsive to temperature increase and seasonal change (Cui et al., 2015). Methanogenic communities on swamps and meadows at the high plateau platform (>4000 m), however, remain poorly investigated despite their supposedly large relevance for the greenhouse gas (GHG) budget of the entire Tibetan Plateau. Also, anticipating if and how the rapid environmental changes presently occurring on the high Tibetan altitude platform will propagate into methanogenic communities is important.

2. Material and methods

2.1. Study site and sample collection

The study sites on the northeastern Tibetan Plateau were selected along the air mass trajectory of the eastern Asian monsoon, by taking account of frozen ground, elevation and hydrology. Four sites were sampled in August 2012 when the active layer reached its maximum thaw depth: Huashixia (HUA), Donggi Cona Lake (DCL), Gande (GAN), and Haibei Station (HAI; Fig. 1). The variability of soil properties between each site was analyzed through a principal component analysis (PCA). Three of the sites are located above 4000 m while HAI is located at an elevation of approximately 3200 m similar to the Zoige wetland. HUA and DCL are located in the discontinuous permafrost region, whereas GAN and HAI are underlain by seasonally frozen ground (SFG). The sites DCL and HAI are swamp wetlands, while GAN and HUA are alpine meadows. For each research site, a series of soil profiles were described along an elevation gradient. The description of the HUA catena has been

€

published recently (Dorfer et al., 2013). Of each catena, the profile which was most relevant for methanogenesis in terms of water level and soil moisture was selected for this study.
The DCL profile is located on a lakeshore wetland which derived from aeolian sediments. The sampling site at HUA is located on a patch of an alpine meadow near the Huashixia Permafrost Station. Laminated sandy or clayish sediments in yellowish and greyish were visible at this site where the discontinuous permafrost starts approximately at 80 cm below the surface. According to the Maduo climate station (98ꢀ120 E, 34ꢀ540 N, 4300 m a.s.l.) monitoring between 1953 and 2010, the mean annual air temperature (MAAT) near DCL and HUA is À4.1 ꢀC with a mean annual precipitation (MAP) of 326 mm and most precipitation occurring in summer (280 mm, May to October). The GAN site is close to a small creek in a valley which is affected by deep seasonally frozen ground. The soil texture mainly consists of silt and clay. The MAAT at the GAN site is À2.2 ꢀC, and the monthly average air temperature ranges from À15.2 ꢀC (January) to 8.6 ꢀC (July), with a MAP of around 550 mm in the period 1994 to 2010. Site HAI is a swamp affected by seasonally frozen ground where peat layer is visible in the entire profile. The MAAT at the Haibei weather station (37ꢀ29' - 37ꢀ450N, 101ꢀ12' - 101ꢀ230E, 2900e3500 m a.s.l.) is À1.7 ꢀC with a maximum of 27.6 ꢀC and a minimum of À37.1 ꢀC in the period from 1989 to 2010. The MAP is around 500 mm with 80% within the growing season from May to September. The vegetation is generally dominated by sedges (Kobresia and Carex) with different coverage and density over different study sites.
At all study sites, soil profiles were excavated down to the permafrost table (approx. 50e70 cm) or to the water table (approx. up to 200 cm). After soil profile description, soil samples were taken in parallel for DNA extraction, methane production incubation and soil property analysis from the middle of each horizon. The samples were labeled with starting site name followed by increasing numbers indicating horizons from top to bottom, e.g.
This study is based on the assumption that methanogenic archaea respond to permafrost degradation, shrinkage of wetlands and extension of meadows. We expect that meadows differ from wetlands in displaying lower methanogenic activities and

68

S. Yang et al. / Soil Biology & Biochemistry 111 (2017) 66e77

Fig. 1. Research sites in Qinghai province on the northeastern Qinghai-Tibet Plateau. The inset figure shows the frozen ground distribution on the plateau. The data is provided by Environmental and Ecological Science Data Center for West China, National Natural Science Foundation of China (http://westdc.westgis.ac.cn). The codes of SFG, SPF, DPF and CPF stand for seasonally frozen ground, sporadic permafrost, discontinuous permafrost and continuous permafrost, respectively. The red rectangle specifies the area in the enlarged map. Donggi Cona and Huashixia are located in discontinuous permafrost area, while Gande and Haibei Station are underlain by seasonally frozen ground. (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.)

DCL1, DCL2, DCL3 (Table S1). The soil properties for each depth were analyzed at the University of Tübingen according to the different soil horizons was weighed into 100 ml glass jars in triplicates and closed with a rubber stopper. The samples were evacuated and flushed with ultrapure N2. The headspace CH4 concentration was determined over time by gas chromatography after adding 6 ml of sterile acetate solution (10 mM) or sterile and anoxic tap water in combination with H2/CO2 (80:20 v/v,150 kPa). A sample without substrate served as control. The slurries were incubated at 10 ꢀC according to the natural soil temperature regime. CH4 production rates were calculated from the linear increase in CH4 concentration over time.

€

methods described by Dorfer et al. (2013). All soil chemical and physical parameters used in this study are listed in Table S1. Among these measures, calcium concentrations, in association with pH, overall have a large effect on the type of wetland that arises. These two factors are particularly important for the distinction between bogs and fens (Keddy, 2010).

2.2. Methane production rates

The samples were transferred into serum bottles in the lab and incubated in triplicates. Potential methane production rates were determined by gas chromatography as previously reported (Wagner et al., 2005). Briefly, fresh soil material (ca. 20 g) from

2.3. DNA extraction and mcrA 454 amplicon sequencing

For logistic reasons, total soil DNA was immediately extracted in triplicates in the field using the FastDNA SPIN kit (MP Biomedicals,

S. Yang et al. / Soil Biology & Biochemistry 111 (2017) 66e77

69

Germany). The air-dried DNA was then stored on QIAsafe DNA 96-

2.5. Quantitative PCR

well filter plates (Qiagen, Germany) at ambient temperature according to the manufacturer instructions. In the lab, the DNA extract was dissolved from the filter and purified with the MiniElute PCR Purification Kit (Qiagen, Germany). In the course of a pilot shotgun sequencing of total DNA prior to this study, the DNA from the upper (top) and lower (down) horizons were pooled for HAI (HAI_t: 5 þ 20 cm, HAI_d: 40 þ 60 cm) and HUA (HUA_t: 5 þ 15 cm, HUA_d: 35 þ 55 cm) to obtain sufficient amounts of DNA. The mcrA fragment was amplified using the commonly-used primer set mlas and mcrA-rev. For multiplexing pyrosequencing, we used barcodetagged forward and reverse primers at the 50 end. PCR reactions
Quantitative PCR was performed on a Bio-Rad CFX instrument
(Bio-Rad, Munich, Germany) using the mlas/mcrA-rev primer pair. Different DNA template concentrations were tested to find the optimal dilutions avoiding inhibitions. The 25 tained 12.5 l of iTaq universal Sybr Green supermix (Bio-Rad, Munich, Germany), 0.25 M concentrations of the primers, and 5 of diluted DNA template. The serial dilution of DNA standard from a mcrA clone (M. barkeri) ranged from 101 to 105 copies À1. PCR was ml reactions con-

mm

ml ml run by an initial denaturation for 10 min at 95 ꢀC, followed by 40 cycles of denaturation at 95 ꢀC for 30 s, annealing at 55ꢀ for 30 s, and extension at 72 ꢀC for 45 s. Melt curve analysis was performed after the final extension by increasing the temperature from 50 to 95 ꢀC at a ramp of 1.0 ꢀC every 5 s. Each measurement was performed in triplicates, with R2 of 0.994 and efficiency of 95%. The amount was given as copies per gram fresh soil. were performed in triplicates in 50 DNA template (5e10 ng/ l) and 0.1

mm

L reactions containing 1.0 M of each primer using the mL

m

MangoMix PCR ready solution (Bioline GmbH, Berlin) according to the manufactor's manual. The PCR conditions were as follows: initial denaturation at 95 ꢀC for 3 min; 30 cycles at 94 ꢀC for 30 s, 55 ꢀC for 45 s, and 72 ꢀC for 45 s, with a final extension at 72 ꢀC for 5 min. Afterwards, parallel PCR products for each sample were pooled and subsequently purified with the MiniElute PCR purifi- cation kit (Qiagen, Hilden, Germany). Pyrosequencing of the equalized mixture was performed on a Roche GS-FLX þþ Titanium platform at Eurofins Genomics (Germany). Sequencing of technical replicates showed highly reproducible results (Fig. S2).

2.6. Network construction

OTUs with average relative abundances less than 0.01% of the total number of OTU phylotypes were removed (Ma et al., 2016). Afterwards, we calculated all pairwise Spearman's rank correlations via R basic package (https://www.r-project.org/). The cooccurrence network was inferred based on the OTUs with a Spearman correlation coefficient (rho) > 0.6 and statistically significant p value < 0.01 (Barberan et al., 2012). This filtering step removed poorly represented OTUs and reduced network complexity, facilitating the determination of the methanogenic core community. The network was then generated by igraph package (v 1.0.1) (Csardi and Nepusz, 2006). The nodes in this network represent individual OTUs and the edges that connect these nodes represent correlations between OTUs. Community subgroups were detected via a walktrap modularity algorithm according to the internal ties and the pattern of ties between different groups.

2.4. Meta-data processing, alpha- and beta-diversity analysis

Raw sequences were processed by the mothur software package
(v.1.31.2) (Schloss et al., 2009), following the standard operating

procedure (https://www.mothur.org/wiki/454_SOP) with an addi-

tional frameshift check with FrameBot (see Table S2 for details). After fasta sequences were extracted from the raw sff sequence file in mothur, these sequences were first subjected to a translation check using the FrameBot tool (http://fungene.cme.msu.edu). Sequences with frameshift errors were excluded from further analysis with custom python script. The subsequent filtering removed the sequences with errors in barcodes or primers, homopolymers longer than 6 nucleotides, ambiguous bases, or average quality scores less than 25. Sequences shorter than 300 nucleotides or longer than 600 nucleotides were also discarded. The unique sequences were then aligned in mothur by default setting with referring to the pre-aligned mcrA sequences provided by the Fungene Pipeline database (http://fungene.cme.msu.edu). Chimeras were detected to be 7.7% of all the reads with the UCHIME algorithm by using the mothur software platform. Using the furthest neighbor clustering method in mothur, the valid sequences were assigned into operational taxonomic units (OTUs) at 84% identity of mcrA gene sequences (Yang et al., 2014). The abundance-based coverage estimator, Chao1 and Shannon estimator of species diversity, rarefaction curves and beta diversity metrics were calculated with R packages of vegan (v.2.0e7) (Oksanen et al., 2013) and phyloseq (v.1.10.0) (McMurdie and Holmes, 2013). For alpha diversity analysis, we avoided subsampling for the reasons given by McMurdie and Holmes (2014). For the beta diversity measures, the proportional abundance was used. We compared different normalization methods during the processing and found that the proportional standardization kept data features as close as possible to the original dataset and avoided the risk of overfitting our analysis (data not shown). The abundance variation of the major taxa across samples was visualized by bubble plot by using package ggplot2 v.1.0.0 (Wickham, 2009). The principal component analysis (PCA) was performed with the vegan package v.2.0e7 (Oksanen et al., 2013) and Venn diagrams were implemented in the VennDiagram package (Chen, 2013).

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  • Anaerobic Methanotrophic Communities Thrive in Deep Submarine Permafrost Received: 12 September 2017 Matthias Winkel1, Julia Mitzscherling1, Pier P

    Anaerobic Methanotrophic Communities Thrive in Deep Submarine Permafrost Received: 12 September 2017 Matthias Winkel1, Julia Mitzscherling1, Pier P

    www.nature.com/scientificreports OPEN Anaerobic methanotrophic communities thrive in deep submarine permafrost Received: 12 September 2017 Matthias Winkel1, Julia Mitzscherling1, Pier P. Overduin2, Fabian Horn1, Maria Winterfeld3, Accepted: 22 December 2017 Ruud Rijkers1, Mikhail N. Grigoriev4, Christian Knoblauch 5, Kai Mangelsdorf6, Dirk Wagner1 Published: xx xx xxxx & Susanne Liebner1 Thawing submarine permafrost is a source of methane to the subsurface biosphere. Methane oxidation in submarine permafrost sediments has been proposed, but the responsible microorganisms remain uncharacterized. We analyzed archaeal communities and identifed distinct anaerobic methanotrophic assemblages of marine and terrestrial origin (ANME-2a/b, ANME-2d) both in frozen and completely thawed submarine permafrost sediments. Besides archaea potentially involved in anaerobic oxidation of methane (AOM) we found a large diversity of archaea mainly belonging to Bathyarchaeota, Thaumarchaeota, and Euryarchaeota. Methane concentrations and δ13C-methane signatures distinguish horizons of potential AOM coupled either to sulfate reduction in a sulfate- methane transition zone (SMTZ) or to the reduction of other electron acceptors, such as iron, manganese or nitrate. Analysis of functional marker genes (mcrA) and fuorescence in situ hybridization (FISH) corroborate potential activity of AOM communities in submarine permafrost sediments at low temperatures. Modeled potential AOM consumes 72–100% of submarine permafrost methane and up to 1.2 Tg of carbon per year for the total expected area of submarine permafrost. This is comparable with AOM habitats such as cold seeps. We thus propose that AOM is active where submarine permafrost thaws, which should be included in global methane budgets. Terrestrial permafrost landscapes, which developed during glacial cold periods, are known to be a large reservoir of organic carbon (~1300 Pg)1.

  • The Potential for Polyphosphate Metabolism in Archaea and Anaerobic Polyphosphate Formation in Methanosarcina Mazei Fabiana S

    www.nature.com/scientificreports OPEN The potential for polyphosphate metabolism in Archaea and anaerobic polyphosphate formation in Methanosarcina mazei Fabiana S. Paula1,2,6*, Jason P. Chin 3,6, Anna Schnürer2, Bettina Müller2, Panagiotis Manesiotis 4, Nicholas Waters 1,5, Katrina A. Macintosh 3, John P. Quinn3, Jasmine Connolly1, Florence Abram1, John W. McGrath3 & Vincent O’Flaherty 1* Inorganic polyphosphate (polyP) is ubiquitous across all forms of life, but the study of its metabolism has been mainly confned to bacteria and yeasts. Few reports detail the presence and accumulation of polyP in Archaea, and little information is available on its functions and regulation. Here, we report that homologs of bacterial polyP metabolism proteins are present across the major taxa in the Archaea, suggesting that archaeal populations may have a greater contribution to global phosphorus cycling than has previously been recognised. We also demonstrate that polyP accumulation can be induced under strictly anaerobic conditions, in response to changes in phosphate (Pi) availability, i.e. Pi starvation, followed by incubation in Pi replete media (overplus), in cells of the methanogenic archaeon Methanosarcina mazei. Pi-starved M. mazei cells increased transcript abundance of the alkaline phosphatase (phoA) gene and of the high-afnity phosphate transport (pstSCAB-phoU) operon: no increase in polyphosphate kinase 1 (ppk1) transcript abundance was observed. Subsequent incubation of Pi-starved M. mazei cells under Pi replete conditions, led to a 237% increase in intracellular polyphosphate content and a > 5.7-fold increase in ppk1 gene transcripts. Ppk1 expression in M. mazei thus appears not to be under classical phosphate starvation control.
  • Interactions Between Anaerobic Fungi and Methanogens in the Rumen and Their Biotechnological Potential in Biogas Production from Lignocellulosic Materials

    Interactions Between Anaerobic Fungi and Methanogens in the Rumen and Their Biotechnological Potential in Biogas Production from Lignocellulosic Materials

    microorganisms Review Interactions between Anaerobic Fungi and Methanogens in the Rumen and Their Biotechnological Potential in Biogas Production from Lignocellulosic Materials Yuqi Li 1 , Zhenxiang Meng 1, Yao Xu 1, Qicheng Shi 1, Yuping Ma 1, Min Aung 1,2, Yanfen Cheng 1,* and Weiyun Zhu 1 1 Laboratory of Gastrointestinal Microbiology, National Center for International Research on Animal Gut Nutrition, Nanjing Agricultural University, Nanjing 210095, China; [email protected] (Y.L.); [email protected] (Z.M.); [email protected] (Y.X.); [email protected] (Q.S.); [email protected] (Y.M.); [email protected] (M.A.); [email protected] (W.Z.) 2 Department of Animal Nutrition, University of Veterinary Science, Nay Pyi Taw 15013, Myanmar * Correspondence: [email protected]; Tel.: +86-25-84395523 Abstract: Anaerobic fungi in the digestive tract of herbivores are one of the critical types of fiber- degrading microorganisms present in the rumen. They degrade lignocellulosic materials using unique rhizoid structures and a diverse range of fiber-degrading enzymes, producing metabolic products such as H2/CO2, formate, lactate, acetate, and ethanol. Methanogens in the rumen utilize some of these products (e.g., H2 and formate) to produce methane. An investigation of the interac- tions between anaerobic fungi and methanogens is helpful as it provides valuable insight into the microbial interactions within the rumen. During the last few decades, research has demonstrated that anaerobic fungi stimulate the growth of methanogens and maintain methanogenic diversity. Meanwhile, methanogens increase the fiber-degrading capability of anaerobic fungi and stimulate Citation: Li, Y.; Meng, Z.; Xu, Y.; Shi, metabolic pathways in the fungal hydrogenosome.
  • Prokaryotic Communities of Acidic Peatlands from the Southern Brazilian Atlantic Forest

    Prokaryotic Communities of Acidic Peatlands from the Southern Brazilian Atlantic Forest

    Brazilian Journal of Microbiology (2012): 661-674 ISSN 1517-8382 PROKARYOTIC COMMUNITIES OF ACIDIC PEATLANDS FROM THE SOUTHERN BRAZILIAN ATLANTIC FOREST R. M. Etto1, L. M. Cruz1, E. C. Jesus2, C. W. Galvão3, F. Galvão4, E. M. Souza1, F. O. Pedrosa1, M. B. R. Steffens1* 1Departamento de Bioquímica e Biologia Molecular, Universidade Federal do Paraná, Curitiba, PR, Brasil; 2Empresa Brasileira de Pesquisa Agropecuária - Agrobiologia, Seropédica, RJ, Brasil; 3Departamento de Biologia Estrutural, Molecular e Genética, Universidade Estadual de Ponta Grossa, Ponta Grossa, PR, Brasil; 4Departamento de Ciências Florestais, Universidade Federal do Paraná, Curitiba, PR, Brasil. Submitted: July 05, 2011; Approved: June 07, 2012. ABSTRACT The acidic peatlands of southern Brazil are ecosystems essential for the maintenance of the Atlantic Forest, one of the 25 hot-spots of biodiversity in the world. In this work, we investigated the composition of prokaryotic communities in four histosols of three acidic peatland regions by constructing small-subunit (SSU) rRNA gene libraries and sequencing. SSU rRNA gene sequence analysis showed the prevalence of Acidobacteria (38.8%) and Proteobacteria (27.4%) of the Bacteria domain and Miscellaneous (58%) and Terrestrial (24%) groups of Crenarchaeota of the Archaea domain. As observed in other ecosystems, archaeal communities showed lower richness than bacterial communities. We also found a limited number of Euryarchaeota and of known methanotrophic bacteria in the clone libraries. Key words: acidic peatland, prokaryotic communities, SSU rRNA gene, microbial ecology INTRODUCTION estimated that these environment cover approximately 611,883 ha, i.e., around 0.07 % of the national territory (55). Peatlands are ecosystems with high quantity of organic The acidic peatlands of the Paraná state in southern Brazil matter due to high water table levels and low temperatures.
  • Supplementary Materials

    Supplementary Materials

    SUPPLEMENTARY TABLE 1. Scientific names of free-living organisms sampled in this study along with taxonomic classification and GO coverage. # Organism Taxonomy Superkingdom GO coverage (%) 1 Aeropyrum pernix K1 Crenarchaeota-Desulfurococcales Archaea 56 2 Sulfolobus solfataricus P2 Crenarchaeota- Sulfolobales Archaea 52 3 Sulfolobus acidocaldarius DSM 639 Crenarchaeota-Sulfolobales Archaea 54 4 Thermofilum pendens Hrk 5 Crenarchaeota-Thermoproteales Archaea 53 5 Pyrobaculum islandicum DSM 4184 Crenarchaeota- Thermoproteales Archaea 52 6 Caldivirga maquilingensis IC-167 Crenarchaeota- Thermoproteales Archaea 58 7 Metallosphaera sedula DSM 5348 Crenarchaeota- Sulfolobales Archaea 55 8 Staphylothermus marinus F1 Crenarchaeota- Desulfurococcales Archaea 55 9 Pyrobaculum calidifontis JCM 11548 Crenarchaeota- Thermoproteales Archaea 52 10 Hyperthermus butylicus DSM 5456 Crenarchaeota- Desulfurococcales Archaea 54 11 Sulfolobus islandicus L.S.2.15 Crenarchaeota- Sulfolobales Archaea 54 12 Thermoproteus neutrophilus V24Sta Crenarchaeota- Thermoproteales Archaea 51 13 Desulfurococcus kamchatkensis 1221n Crenarchaeota- Desulfurococcales Archaea 55 14 Methanosphaerula palustris E1-9c Euryarchaeota- Methanomicrobia Archaea 57 15 Archaeoglobus fulgidus DSM 4304 Euryarchaeota-Archaeoglobi Archaea 56 16 Natronomonas pharaonis DSM 2160 Euryarchaeota-Halobacteria Archaea 52 17 Haloquadratum walsbyi DSM 16790 Euryarchaeota-Halobacteria Archaea 51 18 Halobacterium salinarum R1 Euryarchaeota-Halobacteria Archaea 51 19 Methanobacterium Euryarchaeota-Methanobacteria
  • Methanolinea Mesophila Sp. Nov., a Hydrogenotrophic Methanogen Isolated from Rice Field Soil, and Proposal of the Archaeal Family Methanoregulaceae Fam

    Methanolinea Mesophila Sp. Nov., a Hydrogenotrophic Methanogen Isolated from Rice Field Soil, and Proposal of the Archaeal Family Methanoregulaceae Fam

    Archived version from NCDOCKS Institutional Repository http://libres.uncg.edu/ir/asu/ Methanolinea Mesophila Sp. Nov., A Hydrogenotrophic Methanogen Isolated From Rice Field Soil, And Proposal Of The Archaeal Family Methanoregulaceae Fam. Nov. Within The Order Methanomicrobiales By: Sanae Sakai, Masayuki Ehara, I-Cheng Tseng, Takashi Yamaguchi, Suzanna L. Brauer, Hinsby Cadillo-Quiroz, Stephen H. Zinder and Hiroyuki Imachi Abstract A novel mesophilic, hydrogenotrophic methanogen, designated strain TNRT, was isolated from an anaerobic, propionate-degradation enrichment culture that was originally established from a rice field soil sample from Taiwan. Cells were non-motile rods, 2.0–6.5 mm long by 0.3 mm wide. Filamentous (up to about 100 mm) and coccoid (about 1 mm in diameter) cells were also observed in cultures in the late exponential phase of growth. Strain TNRT grew at 20–40 6C (optimally at 37 6C), at pH 6.5–7.4 (optimally at pH 7.0) and in the presence of 0– 25 g NaCl l”1 (optimally at 0 g NaCl l”1). The strain utilized H2/CO2 and formate for growth and produced methane. The G+C content of the genomic DNA was 56.4 mol%. Based on sequences of both the 16S rRNA gene and the methanogen-specific marker gene mcrA, strain TNRT was related most closely to Methanolinea tarda NOBI-1T; levels of sequence similarities were 94.8 and 86.4 %, respectively. The 16S rRNA gene sequence similarity indicates that strain TNRT and M. tarda NOBI-1T represent different species within the same genus. This is supported by shared phenotypic properties, including substrate usage and cell morphology, and differences in growth temperature.