Dataset Information

Accessing the soil metagenome for studies of microbial diversity.

ABSTRACT: Soil microbial communities contain the highest level of prokaryotic diversity of any environment, and metagenomic approaches involving the extraction of DNA from soil can improve our access to these communities. Most analyses of soil biodiversity and function assume that the DNA extracted represents the microbial community in the soil, but subsequent interpretations are limited by the DNA recovered from the soil. Unfortunately, extraction methods do not provide a uniform and unbiased subsample of metagenomic DNA, and as a consequence, accurate species distributions cannot be determined. Moreover, any bias will propagate errors in estimations of overall microbial diversity and may exclude some microbial classes from study and exploitation. To improve metagenomic approaches, investigate DNA extraction biases, and provide tools for assessing the relative abundances of different groups, we explored the biodiversity of the accessible community DNA by fractioning the metagenomic DNA as a function of (i) vertical soil sampling, (ii) density gradients (cell separation), (iii) cell lysis stringency, and (iv) DNA fragment size distribution. Each fraction had a unique genetic diversity, with different predominant and rare species (based on ribosomal intergenic spacer analysis [RISA] fingerprinting and phylochips). All fractions contributed to the number of bacterial groups uncovered in the metagenome, thus increasing the DNA pool for further applications. Indeed, we were able to access a more genetically diverse proportion of the metagenome (a gain of more than 80% compared to the best single extraction method), limit the predominance of a few genomes, and increase the species richness per sequencing effort. This work stresses the difference between extracted DNA pools and the currently inaccessible complete soil metagenome.

SUBMITTER: Delmont TO

PROVIDER: S-EPMC3067229 | biostudies-literature | 2011 Feb

REPOSITORIES: biostudies-literature

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Accessing the soil metagenome for studies of microbial diversity.

Delmont Tom O TO Robe Patrick P Cecillon Sébastien S Clark Ian M IM Constancias Florentin F Simonet Pascal P Hirsch Penny R PR Vogel Timothy M TM

Applied and environmental microbiology 20101223 4

Soil microbial communities contain the highest level of prokaryotic diversity of any environment, and metagenomic approaches involving the extraction of DNA from soil can improve our access to these communities. Most analyses of soil biodiversity and function assume that the DNA extracted represents the microbial community in the soil, but subsequent interpretations are limited by the DNA recovered from the soil. Unfortunately, extraction methods do not provide a uniform and unbiased subsample o ...[more]

PMID: 21183646

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Project description:In soil, the link between microbial diversity and carbon transformations is challenged by the concept of functional redundancy. Here, we hypothesized that functional redundancy may decrease with increasing carbon source recalcitrance and that coupling of diversity with C cycling may change accordingly. We manipulated microbial diversity to examine how diversity decrease affects the decomposition of easily degradable (i.e., allochthonous plant residues) versus recalcitrant (i.e., autochthonous organic matter) C sources. We found that a decrease in microbial diversity (i) affected the decomposition of both autochthonous and allochthonous carbon sources, thereby reducing global CO2 emission by up to 40%, and (ii) shaped the source of CO2 emission toward preferential decomposition of most degradable C sources. Our results also revealed that the significance of the diversity effect increases with nutrient availability. Altogether, these findings show that C cycling in soil may be more vulnerable to microbial diversity changes than expected from previous studies, particularly in ecosystems exposed to nutrient inputs. Thus, concern about the preservation of microbial diversity may be highly relevant in the current global-change context assumed to impact soil biodiversity and the pulse inputs of plant residues and rhizodeposits into the soil.IMPORTANCE With hundreds of thousands of taxa per gram of soil, microbial diversity dominates soil biodiversity. While numerous studies have established that microbial communities respond rapidly to environmental changes, the relationship between microbial diversity and soil functioning remains controversial. Using a well-controlled laboratory approach, we provide empirical evidence that microbial diversity may be of high significance for organic matter decomposition, a major process on which rely many of the ecosystem services provided by the soil ecosystem. These new findings should be taken into account in future studies aimed at understanding and predicting the functional consequences of changes in microbial diversity on soil ecosystem services and carbon storage in soil.

Project description:BackgroundCellulose, an abundant and renewable polysaccharides, constitutes the largest resource for bioconversion of biofuels. Plant polysaccharides hydrolysis is catalyzed by cellulases, which include endoglucanases, exoglucanases, and β-glucosidases. Converting cellulose and hemicellulose to short chains of oligosaccharides by endo-/exoglucanases is the key step for biofuel transformation. Intriguingly, β-glucanases with transglycosylation activity not only can relieve product inhibition of glucan hydrolysis but also has potential application as biocatalysts for functional materials.ResultsHere, a metagenomic fosmid library was constructed from a paddy soil for cellulase screening. One purified clone showing carboxymethylcellulase activity was isolated, and the complete β-glucanase gene (umcel9y-1) was cloned and overexpressed in Escherichia coli. Phylogenetic analysis indicated that β-glucanase Umcel9y-1 belonged to the theme C of glycoside hydrolase family 9. Amino acids sequence showed 58.4 % similarity between Umcel9y-1 and its closest characterized reference, cellulase Cel01. Biological characterization showed that Umcel9y-1 was an efficient endoglucanase and also exhibited high activities of exoglucanase and transglycosylation. The transglycosylation products of Umcel9y-1 including sophorose, laminaribiose, and gentiobiose, and transglycosylation was detected under all activated conditions. The order of catalytic efficiency for polysaccharides, cellooligosaccharides, and aryl-β-glycosides was p-nitrophenol-D-cellobioside, barley glucan, cellopentaose, cellotetraose, cellotriose, hydroxyethylcellulose, cellohexose, laminarin, and carboxymethylcellulose, respectively. The barley glucan was the optimal polysaccharides for Umcel9y-1 with K m and K cat/K m values of 13.700 mM and 239.152 s(-1) mM(-1), respectively.ConclusionBiological characterizations of recombinant Umcel9y-1 showed that the versatile β-glucanase had efficient endoglucanase activity to barley glucan and also exhibited high activities of exoglucanase and transglycosylation. The optimum conditions of recombinant Umcel9y-1 was pH 6.5-7.0 at 37 °C with predominant halotolerance and high-thermal stability. These results indicate that the novel metagenomic-derived β-glucanase may be a potent candidate for industrial applications.

Dataset Information

Accessing the soil metagenome for studies of microbial diversity.

Publications

Accessing the soil metagenome for studies of microbial diversity.

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