A Biomolecular Isolation Framework for Molecular Eco-Systems Biology

Poster (2012)

With the advent of high-throughput omic technologies, powerful and sensitive methods are available for the analysis of nucleic acid, protein and small molecule complements obtained from biological samples ... [more ▼]

With the advent of high-throughput omic technologies, powerful and sensitive methods are available for the analysis of nucleic acid, protein and small molecule complements obtained from biological samples. Molecular eco-systems biology studies based on the integration of genomic, transcriptomic, proteomic and metabolomic data are faced with major challenges arising from the complexity, dynamics and heterogeneity of mixed microbial consortia. In order to facilitate meaningful data integration, analysis and modeling, it is essential that biomolecular fractions obtained for high-throughput omic analyses are representative of single unique samples. We have developed a new methodological framework for the reproducible isolation of high-quality genomic DNA, large and small RNA, proteins, and polar and non-polar metabolites from single unique mixed microbial community samples. The methodology is based around reproducible cryogenic sample preservation and cell lysis. Metabolites are extracted first using organic solvents, followed by the sequential isolation of nucleic acids and proteins using chromatographic spin column technology. The methodology was validated by comparison to traditional dedicated and simultaneous biomolecular isolation methods. To prove the broad applicability of the methodology, we applied it to microbial consortia of biotechnological, environmental and medical interest. Importantly, the developed methodology will allow exploitation of the inherent heterogeneity and dynamics within microbial consortia through spatial and temporal sampling of biological systems to allow later deconvolution of community-wide, population-wide and individual-level processes using the generated omic data. This approach has the potential to identify associations between distinct biomolecules and which may provide pointers towards unravelling previously unknown metabolic processes. Finally, by providing a standardized workflow, the methodology lays the foundation for comparative eco-systematic studies of different natural microbial consortia in the future. [less ▲]

Deuterium-exchange metabolomics identifies N-methyl lyso phosphatidylethanolamines as abundant lipids in acidophilic mixed microbial communities

in Metabolomics : Official journal of the Metabolomic Society (2012), 8(4), 566-578

Natural microbial communities are extremely diverse and contain uncharacterized but functionally important small molecules. By coupling a deuterium (D) labeling technique to high mass accuracy untargeted ... [more ▼]

Natural microbial communities are extremely diverse and contain uncharacterized but functionally important small molecules. By coupling a deuterium (D) labeling technique to high mass accuracy untargeted liquid chromatography-electrospray ionization-mass spec- trometry (LC–ESI–MS) metabolomic analysis, we found that natural acidophilic microbial biofilms dominated by bacteria of the genus Leptospirillum contained unusual lyso phosphatidylethanolamine (PE) lipids in high abundance (more than 10 nmol/mg of dry biomass). The unusual polar head group structure of these lipids is similar to lipids found in phylogenetically unrelated acidophilic chemo- autolithotrophs and may be related to the affinity of these lipids for iron and calcium ions. Correlations of lyso phospholipid and proteome abundance patterns suggest a link between the lyso phospholipids and the UBA-type substrain of Leptospirillum group II. By combining untar- geted metabolomics with D exchange we demonstrate the ability to identify cryptic but biologically functional small molecules in mixed microbial communities. [less ▲]

Genome Sequence of "Candidatus Microthrix parvicella" Bio17-1, a Long-Chain-Fatty-Acid-Accumulating Filamentous Actinobacterium from a Biological Wastewater Treatment Plant

in Journal of Bacteriology (2012), 194(23), 6670-6671

Candidatus Microthrix bacteria are deeply branching filamentous actinobacteria which occur at the water-air interface of biological wastewater treatment plants, where they are often responsible for ... [more ▼]

Candidatus Microthrix bacteria are deeply branching filamentous actinobacteria which occur at the water-air interface of biological wastewater treatment plants, where they are often responsible for foaming and bulking. Here, we report the first draft genome sequence of a strain from this genus: "Candidatus Microthrix parvicella" strain Bio17-1. [less ▲]

Architecture reflects function: high-resolution microscopic and molecular insights into acid mine drainage biofilms

Scientific Conference (2011, August)

Microbial Eco-Systems Biology: from molecules to ecosystems and back

Presentation (2011, April)

Microbial Eco-Systems Biology: from molecules to ecosystems and back

Presentation (2011, January)

Correlative microscopy for phylogenetic and ultrastructural characterization of microbial communities

in Environmental Microbiology Reports (2011)

Transmission electron microscopy (TEM) can provide ultrastructural information for cells in microbial community samples and phylogenetic information can be recovered via molecular surveys. Here we report ... [more ▼]

Transmission electron microscopy (TEM) can provide ultrastructural information for cells in microbial community samples and phylogenetic information can be recovered via molecular surveys. Here we report an approach to link these data sets by coupling fluorescence in situ hybridization (FISH) with either conventional biological or cryogenic TEM. The method could fundamentally improve our understanding of the organization and functioning of microbial communities in natural systems. [less ▲]

From molecules to ecosystems and back: bioinformatic approaches for Eco-Systems Biology

Scientific Conference (2010, November)

Tandem proteomics and metabolomics for elucidating cryptic metabolic transformations within mixed microbial communities

Scientific Conference (2010, October)

Characterization of Extracellular Polymeric Substances from Acidophilic Microbial Biofilms

in Applied and Environmental Microbiology (2010), 76(9), 2916-2922

We examined the chemical composition of extracellular polymeric substances (EPS) extracted from two natural microbial pellicle biofilms growing on acid mine drainage (AMD) solutions. The EPS obtained from ... [more ▼]

We examined the chemical composition of extracellular polymeric substances (EPS) extracted from two natural microbial pellicle biofilms growing on acid mine drainage (AMD) solutions. The EPS obtained from a mid-developmental-stage biofilm (DS1) and a mature biofilm (DS2) were qualitatively and quantitatively compared. More than twice as much EPS was derived from DS2 as from DS1 (approximately 340 and 150 mg of EPS per g [dry weight] for DS2 and DS1, respectively). Composition analyses indicated the presence of carbohydrates, metals, proteins, and minor quantities of DNA and lipids, although the relative concentrations of these components were different for the two EPS samples. EPS from DS2 contained higher concentrations of metals and carbohydrates than EPS from DS1. Fe was the most abundant metal in both samples, accounting for about 73% of the total metal content, followed by Al, Mg, and Zn. The relative concentration profile for these metals resembled that for the AMD solution in which the biofilms grew, except for Si, Mn, and Co. Glycosyl composition analysis indicated that both EPS samples were composed primarily of galactose, glucose, heptose, rhamnose, and mannose, while the relative amounts of individual sugars were substantially different in DS1 and DS2. Additionally, carbohydrate linkage analysis revealed multiply linked heptose, galactose, glucose, mannose, and rhamnose, with some of the glucose in a 4-linked form. These results indicate that the biochemical composition of the EPS from these acidic biofilms is dependent on maturity and is controlled by the microbial communities, as well as the local geochemical environment. [less ▲]