Archaeal ammonia oxidisers are abundant in acidic, coarse-textured Australian soils

Soil Research ◽  
2011 ◽  
Vol 49 (8) ◽  
pp. 715 ◽  
Author(s):  
Cathryn A. O'Sullivan ◽  
Steven A. Wakelin ◽  
Ian R. P. Fillery ◽  
Adrienne L. Gregg ◽  
Margaret M. Roper
Keyword(s):  
Denaturing Gradient Gel Electrophoresis ◽  
Agricultural Soils ◽  
Amoa Gene ◽  
Banding Patterns ◽  
Soil Dna ◽  
Potential Nitrification ◽  
Gene Copies ◽  
Gradient Gel Electrophoresis ◽  
Key Driver ◽  
Ammonia Oxidising Archaea

The abundances of ammonia-oxidising archaea (AOA) and ammonia-oxidising bacteria (AOB) in soils underlying pastures in the south-west of Western Australia (WA) were investigated. Samples were collected from irrigated pastures and one unmanaged (driveway) area during December 2009. Archaeal and bacterial ammonia monooxygenase (amoA) genes were quantified using real-time PCR, and the diversity of the archaeal amoA genes was investigated using denaturing gradient gel electrophoresis (PCR-DGGE). AOA amoA gene copies outnumbered AOB in all samples. Numbers of archaeal amoA genes ranged from 4.1E+01 to 1.34E+05 gene copies/ng soil DNA. Bacterial amoA genes were below detection limits at three of the four sample sites and ranged from 8.9E+01 to 6.7E+02 gene copies/ng soil DNA at the remaining site. Potential nitrification rates (PNR) were not correlated with AOA or AOB gene abundance, but high PNR only occurred at the site with measureable numbers of AOB. The DGGE analysis revealed that the AOA community was diverse and variability in banding patterns was significantly affected by both site and depth (P < 0.05). Statistical analysis matching biological variation (AOA amoA genotypes) to environmental variables (BEST analysis) revealed that pH was the key driver of AOA community structure (ρ = 0.72; P = 0.005). Soil pH was also inversely correlated to abundance of AOA amoA genes in soil (ρ = 0.8; P = 0.003). This study has shown that AOA are important members of the nitrogen-cycling community in acidic WA pasture soils, and likely in the wider agricultural soils of WA.

scholarly journals Effect of DNA Extraction Method on the Apparent Microbial Diversity of Soil

2010 ◽  
Vol 76 (10) ◽  
pp. 3378-3382 ◽  
Author(s):  
Özgül İnceoǧlu ◽  
Eelco F. Hoogwout ◽  
Patrick Hill ◽  
Jan Dirk van Elsas
Keyword(s):  
Extraction Method ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Agricultural Soils ◽  
Extraction Methods ◽  
The Novel ◽  
Soil Dna ◽  
Dna Extraction Method ◽  
Gradient Gel Electrophoresis ◽  
Novel Method ◽  
Microbial Groups

ABSTRACT Four extraction methods, including a novel one, were compared for their efficiencies in producing DNA from three contrasting agricultural soils. Molecular analyses (PCR-denaturing gradient gel electrophoresis [DGGE] and clone libraries) focusing on different microbial groups were used as assessment criteria. Per soil, the DNA yields differed between extraction methods. Clear effects of method on apparent richness and community structure were found. Actinobacterial diversity based on soil DNA produced by two divergent methods revealed that a hitherto-undescribed group was obtained by the novel method.

Methanotrophic diversity in high arctic wetlands on the islands of Svalbard (Norway) - denaturing gradient gel electrophoresis analysis of soil DNA and enrichment cultures

2003 ◽  
Vol 49 (10) ◽  
pp. 602-612 ◽  
Author(s):  
Ingvild Wartiainen ◽  
Anne Grethe Hestnes ◽  
Mette M Svenning
Keyword(s):  
Methane Emission ◽  
Gel Electrophoresis ◽  
Denaturing Gradient Gel Electrophoresis ◽  
The Arctic ◽  
Rrna Gene ◽  
Climatic Warming ◽  
Soil Dna ◽  
Enrichment Cultures ◽  
Arctic Soil ◽  
Gradient Gel Electrophoresis

The methanotrophic community in arctic soil from the islands of Svalbard, Norway (78°N) was analysed by combining group-specific PCR with PCR of the highly variable V3 region of the 16S rRNA gene and then by denaturing gradient gel electrophoresis (DGGE). Selected bands were sequenced for identification. The analyses were performed with DNA extracted directly from soil and from enrichment cultures at 10 and 20 °C. The two genera Methylobacter and Methylosinus were found in all localities studied. The DGGE band patterns were simple, and DNA fragments with single base differences were separated. The arctic tundra is a potential source of extensive methane emission due to climatic warming because of its large reservoirs of stored organic carbon. Higher temperatures due to climatic warming can cause increased methane production, and the abundance and activity of methane-oxidizing bacteria in the arctic soil may be important regulators for methane emission to the atmosphere.Key words: methanotrophic diversity, Svalbard, arctic wetland, denaturing gradient gel electrophoresis.

Isolation and purification of microbial community DNA from soil naturally enriched for chitin

Biologia ◽  
2012 ◽  
Vol 67 (4) ◽  
Author(s):  
Pullabhotla Sarma ◽  
Vadlamudi Srinivas ◽  
Kondreddy Anil ◽  
Appa Podile
Keyword(s):  
16S Rdna ◽  
Gel Electrophoresis ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Sodium Borate ◽  
Metagenomic Dna ◽  
Soil Dna ◽  
Isolation And Purification ◽  
Gradient Gel Electrophoresis ◽  
Chitinase Genes ◽  
Sodium Borate Buffer

AbstractWe made an attempt to isolate and purify metagenomic DNA from chitin enriched soil. In this communication we report a modified direct lysis method for soil DNA extraction including initial pre-lysis washing of sample, followed by a rapid polyvinylpyrrolidone-agarose-based purification and electroelution of DNA using Gene-capsule™ assembly. Rapidity was achieved using low molarity conducting media (sodium-borate buffer) for electrophoresis by reducing run time for both the gel electrophoresis and electroelution. Extracted DNA was sufficiently pure and of high quality, evidenced by amplification of 16S rDNA and chitinase genes by PCR. Metagenomic nature of the DNA was confirmed by running V3 (16S rDNA) region amplicons using denaturing gradient gel electrophoresis. This method requires 30 min for purification, and less than 2 h for complete execution of protocol and becomes the first report on the isolation of metagenomic DNA from soil naturally enriched for chitin.

scholarly journals Simultaneous nitrification and phosphate removal by bioaugmented aerobic granules treating a fluoroorganic compound

2021 ◽  
Author(s):  
Anouk F. Duque ◽  
Vânia S. Bessa ◽  
Udo van Dongen ◽  
Merle K. de Kreuk ◽  
Raquel B. R. Mesquita ◽  
...  
Keyword(s):  
Denaturing Gradient Gel Electrophoresis ◽  
Amoa Gene ◽  
Granular Sludge ◽  
Phosphate Removal ◽  
Aerobic Granular Sludge ◽  
Ammonia Oxidizing Bacteria ◽  
Aerobic Granules ◽  
N Removal ◽  
Nitrite Oxidizing Bacteria ◽  
Gradient Gel Electrophoresis

Abstract The presence of toxic compounds in wastewater can cause problems for organic matter and nutrient removal. In this study, the long term effect of a model xenobiotic, 2-fluorophenol (2-FP), on ammonia oxidizing bacteria (AOB), nitrite oxidizing bacteria (NOB) and phosphate accumulating organisms (PAO) in aerobic granular sludge was investigated. Phosphate (P) and ammonium (N) removal efficiencies were high (&gt;93%) and, after bioaugmentation with 2-FP degrading strain FP1, 2-FP was completely degraded. Neither N nor P removal were affected by 50 mg L−1 of 2-FP in the feed stream. Changes in the aerobic granule bacterial communities were followed. Numerical analysis of the denaturing gradient gel electrophoresis (DGGE) profiles showed low diversity for the amoA gene with an even distribution of species. PAOs, including denitrifying PAO (dPAO), and AOB were present in the 2-FP degrading granules, although dPAO population decreased throughout the 444 days reactor operation. The results demonstrated that the aerobic granules bioaugmented with FP1 strain successfully removed N, P and 2-FP simultaneously.

scholarly journals Numerical Analysis of Grassland Bacterial Community Structure under Different Land Management Regimens by Using 16S Ribosomal DNA Sequence Data and Denaturing Gradient Gel Electrophoresis Banding Patterns

2001 ◽  
Vol 67 (10) ◽  
pp. 4554-4559 ◽  
Author(s):  
Allison E. McCaig ◽  
L. Anne Glover ◽  
James I. Prosser
Keyword(s):  
Ribosomal Dna ◽  
Gel Electrophoresis ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Sequence Data ◽  
Canonical Variate ◽  
Similarity Coefficients ◽  
Grassland Soils ◽  
Soil Dna ◽  
16S Ribosomal Dna ◽  
Gradient Gel Electrophoresis

ABSTRACT Bacterial diversity in unimproved and improved grassland soils was assessed by PCR amplification of bacterial 16S ribosomal DNA (rDNA) from directly extracted soil DNA, followed by sequencing of ∼45 16S rDNA clones from each of three unimproved and three improved grassland samples (A. E. McCaig, L. A. Glover, and J. I. Prosser, Appl. Environ. Microbiol. 65:1721–1730, 1999) or by denaturing gradient gel electrophoresis (DGGE) of total amplification products. Semi-improved grassland soils were analyzed only by DGGE. No differences between communities were detected by calculation of diversity indices and similarity coefficients for clone data (possibly due to poor coverage). Differences were not observed between the diversities of individual unimproved and improved grassland DGGE profiles, although considerable spatial variation was observed among triplicate samples. Semi-improved grassland samples, however, were less diverse than the other grassland samples and had much lower within-group variation. DGGE banding profiles obtained from triplicate samples pooled prior to analysis indicated that there was less evenness in improved soils, suggesting that selection for specific bacterial groups occurred. Analysis of DGGE profiles by canonical variate analysis but not by principal-coordinate analysis, using unweighted data (considering only the presence and absence of bands) and weighted data (considering the relative intensity of each band), demonstrated that there were clear differences between grasslands, and the results were not affected by weighting of data. This study demonstrated that quantitative analysis of data obtained by community profiling methods, such as DGGE, can reveal differences between complex microbial communities.

scholarly journals Nutrient Amendments in Soil DNA Stable Isotope Probing Experiments Reduce the Observed Methanotroph Diversity

2006 ◽  
Vol 73 (3) ◽  
pp. 798-807 ◽  
Author(s):  
Aur�lie C�bron ◽  
Levente Bodrossy ◽  
Nancy Stralis-Pavese ◽  
Andrew C. Singer ◽  
Ian P. Thompson ◽  
...  
Keyword(s):  
Stable Isotope ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Stable Isotope Probing ◽  
Population Diversity ◽  
Type I ◽  
Mineral Salts ◽  
Bacterial Populations ◽  
Soil Dna ◽  
Gradient Gel Electrophoresis ◽  
Soil Water Conditions

ABSTRACT Stable isotope probing (SIP) can be used to analyze the active bacterial populations involved in a process by incorporating 13C-labeled substrate into cellular components such as DNA. Relatively long incubation times are often used with laboratory microcosms in order to incorporate sufficient 13C into the DNA of the target organisms. Addition of nutrients can be used to accelerate the processes. However, unnatural concentrations of nutrients may artificially change bacterial diversity and activity. In this study, methanotroph activity and diversity in soil was examined during the consumption of 13CH4 with three DNA-SIP experiments, using microcosms with natural field soil water conditions, the addition of water, and the addition of mineral salts solution. Methanotroph population diversity was studied by targeting 16S rRNA and pmoA genes. Clone library analyses, denaturing gradient gel electrophoresis fingerprinting, and pmoA microarray hybridization analyses were carried out. Most methanotroph diversity (type I and type II methanotrophs) was observed in nonamended SIP microcosms. Although this treatment probably best reflected the in situ environmental conditions, one major disadvantage of this incubation was that the incorporation of 13CH4 was slow and some cross-feeding of 13C occurred, thereby leading to labeling of nonmethanotroph microorganisms. Conversely, microcosms supplemented with mineral salts medium exhibited rapid consumption of 13CH4, resulting in the labeling of a less diverse population of only type I methanotrophs. DNA-SIP incubations using water-amended microcosms yielded faster incorporation of 13C into active methanotrophs while avoiding the cross-feeding of 13C.

scholarly journals Analysis of Bacterial Community Composition by Oligonucleotide Fingerprinting of rRNA Genes

2002 ◽  
Vol 68 (7) ◽  
pp. 3243-3250 ◽  
Author(s):  
Lea Valinsky ◽  
Gianluca Della Vedova ◽  
Alexandra J. Scupham ◽  
Sam Alvey ◽  
Andres Figueroa ◽  
...  
Keyword(s):  
Denaturing Gradient Gel Electrophoresis ◽  
Bacterial Community Composition ◽  
Effective Means ◽  
Cost Effective ◽  
Rrna Genes ◽  
Soil Dna ◽  
Specific Pcr ◽  
Gradient Gel Electrophoresis ◽  
Oligonucleotide Fingerprinting ◽  
Methodological Advance

ABSTRACT One of the first steps in characterizing an ecosystem is to describe the organisms inhabiting it. For microbial studies, experimental limitations have hindered the ability to depict diverse communities. Here we describe oligonucleotide fingerprinting of rRNA genes (OFRG), a method that permits identification of arrayed rRNA genes (rDNA) through a series of hybridization experiments using small DNA probes. To demonstrate this strategy, we examined the bacteria inhabiting two different soils. Analysis of 1,536 rDNA clones revealed 766 clusters grouped into five major taxa: Bacillus, Actinobacteria, Proteobacteria, and two undefined assemblages. Soil-specific taxa were identified and then independently confirmed through cluster-specific PCR of the original soil DNA. Near-species-level resolution was obtained by this analysis as clones with average sequence identities of 97% were grouped in the same cluster. A comparison of these OFRG results with the results obtained in a denaturing gradient gel electrophoresis analysis of the same two soils demonstrated the significance of this methodological advance. OFRG provides a cost-effective means to extensively analyze microbial communities and should have applications in medicine, biotechnology, and ecosystem studies.

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