A simple boiling-based DNA extraction for RAPD profiling of landfarm soil to provide representative metagenomic content

Landfarm soils are employed in industrial and petrochemical residue bioremediation. This process induces selective pressure directed towards microorganisms capable of degrading toxic compounds. Detailed description of taxa in these environments is difficult due to a lack of knowledge of culture conditions required for unknown microorganisms. A metagenomic approach permits identification of organisms without the need for culture. However, a DNA extraction step is first required, which can bias taxonomic representativeness and interfere with cloning steps by extracting interference substances. We developed a simplified DNA extraction procedure coupled with metagenomic DNA amplification in an effort to overcome these limitations. The amplified sequences were used to generate a metagenomic data set and the taxonomic and functional representativeness were evaluated in comparison with a data set built with DNA extracted by conventional methods. The simplified and optimized method of RAPD to access metagenomic information provides better representativeness of the taxonomical and metabolic aspects of the environmental samples.

The Microbiome of Brazilian Mangrove Sediments as Revealed by Metagenomics

Here we embark in a deep metagenomic survey that revealed the taxonomic and potential metabolic pathways aspects of mangrove sediment microbiology. The extraction of DNA from sediment samples and the direct application of pyrosequencing resulted in approximately 215 Mb of data from four distinct mangrove areas (BrMgv01 to 04) in Brazil. The taxonomic approaches applied revealed the dominance of Deltaproteobacteria and Gammaproteobacteria in the samples. Paired statistical analysis showed higher proportions of specific taxonomic groups in each dataset. The metabolic reconstruction indicated the possible occurrence of processes modulated by the prevailing conditions found in mangrove sediments. In terms of carbon cycling, the sequences indicated the prevalence of genes involved in the metabolism of methane, formaldehyde, and carbon dioxide. With respect to the nitrogen cycle, evidence for sequences associated with dissimilatory reduction of nitrate, nitrogen immobilization, and denitrification was detected. Sequences related to the production of adenylsulfate, sulfite, and H2S were relevant to the sulphur cycle. These data indicate that the microbial core involved in methane, nitrogen, and sulphur metabolism consists mainly of Burkholderiaceae, Planctomycetaceae, Rhodobacteraceae, and Desulfobacteraceae. Comparison of our data to datasets from soil and sea samples resulted in the allotment of the mangrove sediments between those samples. The results of this study add valuable data about the composition of microbial communities in mangroves and also shed light on possible transformations promoted by microbial organisms in mangrove sediments.

Structural and Functional Insights from the Metagenome of an Acidic Hot Spring Microbial Planktonic Community in the Colombian Andes

A taxonomic and annotated functional description of microbial life was deduced from 53 Mb of metagenomic sequence retrieved from a planktonic fraction of the Neotropical high Andean (3,973 meters above sea level) acidic hot spring El Coquito (EC). A classification of unassembled metagenomic reads using different databases showed a high proportion of Gammaproteobacteria and Alphaproteobacteria (in total read affiliation), and through taxonomic affiliation of 16S rRNA gene fragments we observed the presence of Proteobacteria, micro-algae chloroplast and Firmicutes. Reads mapped against the genomes Acidiphilium cryptum JF-5, Legionella pneumophila str. Corby and Acidithiobacillus caldus revealed the presence of transposase-like sequences, potentially involved in horizontal gene transfer. Functional annotation and hierarchical comparison with different datasets obtained by pyrosequencing in different ecosystems showed that the microbial community also contained extensive DNA repair systems, possibly to cope with ultraviolet radiation at such high altitudes. Analysis of genes involved in the nitrogen cycle indicated the presence of dissimilatory nitrate reduction to N2 (narGHI, nirS, norBCDQ and nosZ), associated with Proteobacteria-like sequences. Genes involved in the sulfur cycle (cysDN, cysNC and aprA) indicated adenylsulfate and sulfite production that were affiliated to several bacterial species. In summary, metagenomic sequence data provided insight regarding the structure and possible functions of this hot spring microbial community, describing some groups potentially involved in the nitrogen and sulfur cycling in this environment. Citation: Jimenez DJ, Andreote FD, Chaves D, Montana JS, Osorio-Forero C, et al. (2012) Structural and Functional Insights from the Metagenome of an Acidic Hot Spring Microbial Planktonic Community in the Colombian Andes. PLoS ONE 7(12): e52069. doi:10.1371/journal.pone.0052069