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Most microbes in the biosphere remain unculturable. Whole genome shotgun (WGS) sequencing of environmental DNA (metagenomics) can be used to study the genetic and metabolic properties of natural microbial communities. However, in communities of high complexity, metagenomics fails to link specific microbes to specific ecological functions. To overcome this limitation, we developed a method to target microbial subpopulations by labeling DNA through stable isotope probing (SIP), followed by WGS sequencing. Metagenome analysis of microbes from Lake Washington in Seattle that oxidize single-carbon (C1) compounds shows specific sequence enrichments in response to different C1 substrates, revealing the ecological roles of individual phylotypes. We also demonstrate the utility of our approach by extracting a nearly complete genome of a novel methylotroph, Methylotenera mobilis, reconstructing its metabolism and conducting genome-wide analyses. This high-resolution, targeted metagenomics approach may be applicable to a wide variety of ecosystems. [ABSTRACT FROM AUTHOR]

Xylose is a major constituent of plant lignocellulose, and its fermentation is important for the bioconversion of plant biomass to fuels and chemicals. Pichia stipitis is a well-studied, native xylose-fermenting yeast. The mechanism and regulation of xylose metabolism in P. stipitis have been characterized and genes from P. stipitis have been used to engineer xylose metabolism in Saccharomyces cerevisiae. We have sequenced and assembled the complete genome of P. stipitis. The sequence data have revealed unusual aspects of genome organization, numerous genes for bioconversion, a preliminary insight into regulation of central metabolic pathways and several examples of colocalized genes with related functions. The genome sequence provides insight into how P. stipitis regulates its redox balance while very efficiently fermenting xylose under microaerobic conditions. [ABSTRACT FROM AUTHOR]

Microbial communities are vital in the functioning of all ecosystems; however, most microorganisms are uncultivated, and their roles in natural systems are unclear. Here, using random shotgun sequencing of DNA from a natural acidophilic biofilm, we report reconstruction of near-complete genomes of Leptospirillum group II and Ferroplasma type II, and partial recovery of three other genomes. This was possible because the biofilm was dominated by a small number of species populations and the frequency of genomic rearrangements and gene insertions or deletions was relatively low. Because each sequence read came from a different individual, we could determine that single-nucleotide polymorphisms are the predominant form of heterogeneity at the strain level. The Leptospirillum group II genome had remarkably few nucleotide polymorphisms, despite the existence of low-abundance variants. The Ferroplasma type II genome seems to be a composite from three ancestral strains that have undergone homologous recombination to form a large population of mosaic genomes. Analysis of the gene complement for each organism revealed the pathways for carbon and nitrogen fixation and energy generation, and provided insights into survival strategies in an extreme environment. [ABSTRACT FROM AUTHOR]