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See detailRNA in circulation: sources and functions of extracellular exogenous RNA in blood
Galas, David J. UL; Wilmes, Paul UL; Wang, Kai

in Nelson, Karen (Ed.) Encyclopedia of Metagenomics (2014)

Molecules of many kinds are abundant in circulating blood and play a wide range of important roles, both known and unknown. These include macromolecules like proteins and nucleic acids and a wide range of ... [more ▼]

Molecules of many kinds are abundant in circulating blood and play a wide range of important roles, both known and unknown. These include macromolecules like proteins and nucleic acids and a wide range of smaller molecules. A number of questions are raised by recent findings of stable RNA molecules in plasma that is circulating RNA outside of cells. Among the issues that need to be addressed are: what are the origins of these RNA molecules; what are the mechanisms by which they enter and are stabilized in the blood; what are their possible biological functions; and finally, what are the potential applications of these extracellular RNA molecules in diagnostic and therapeutic medicine? While the precise biological functions remain to be pinned down, extracellular RNA has been proposed as a vehicle for a previously unknown cell-cell communication system. Recent reports of the detection of foreign, exogenous sources of some of the extracellular RNA have thus intensified the need to investigate and understand these processes. This overview summarizes the findings, some recent developments, and the current state of research in the circulating RNA field, and some of the key open questions in the field are specifically addressed. [less ▲]

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See detailThe sequential isolation of metabolites, RNA, DNA, and proteins from a single, undivided mixed microbial community sample
Muller, Emilie UL; Buschart, Anna UL; Roume, Hugo UL et al

in Protocol Exchange (2014)

Integrated omics of microbial consortia, comprising systematized metagenomic, metatranscriptomic, metaproteomic and meta-metabolomic analyses, allows in-depth characterization of organismal and functional ... [more ▼]

Integrated omics of microbial consortia, comprising systematized metagenomic, metatranscriptomic, metaproteomic and meta-metabolomic analyses, allows in-depth characterization of organismal and functional diversity in situ. To allow meaningful meta-omic data integration, truly systematic measurements of the typically heterogeneous sample biomass is required. Therefore, there is a need for analyzing biomolecular fractions obtained from single, undivided samples. Here, we share a methodological workflow for the reproducible isolation of concomitant polar and non-polar metabolites, RNA, DNA and proteins from samples obtained from a biological wastewater treatment plant. The methodological framework is applicable to other biological samples [1,2], is compatible with different kits for biomacromolecular isolation [1,2] with minimal tailoring, and represents an important first step in standardization for the emerging field of Molecular Eco-Systems Biology. [less ▲]

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See detailA 16S rRNA gene Illumina–based barcoded assay design for high throughput characterisation of microbial communities from anaerobic digesters
Calusinska, Magdalena; Goux, Xavier; Muller, Emilie UL et al

Poster (2014)

High throughput sequencing of 16S/18S rRNA gene is becoming an indispensable tool to explore microbial community ecology. To date, most of the studies using next generation amplicon sequencing of ... [more ▼]

High throughput sequencing of 16S/18S rRNA gene is becoming an indispensable tool to explore microbial community ecology. To date, most of the studies using next generation amplicon sequencing of microorganisms involved in the anaerobic digestion process (AD) are based on the 454 pyrosequencing. However, the cost per read obtained with the Illumina technology is currently less than 1/100 of that for the 454 pyrosequencing, thus enabling throughout sequencing and larger number of samples to be analysed per study (e.g. given the current Miseq output, around 100 000 reads per sample can be expected for a pool of 96 libraries). Moreover, the Illumina technology is less biased by the GC content of the template and currently allows for a relatively long sequence read of 600 bp (2 x 300bp). This consideration is particularly important, since longer sequences permit for more accurate assignment to a taxonomic group. Here, we designed and optimized an Illumina–based 16S rRNA amplicon approach for a high throughput characterization of microbial communities from different AD. [less ▲]

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See detailSystematic Design of 18S rRNA Gene Primers for Determining Eukaryotic Diversity in Microbial Consortia
Hugerth, Luisa; Muller, Emilie UL; Hu, Yue et al

in PLoS ONE (2014)

High-throughput sequencing of ribosomal RNA gene (rDNA) amplicons has opened up the door to large-scale comparative studies of microbial community structures. The short reads currently produced by ... [more ▼]

High-throughput sequencing of ribosomal RNA gene (rDNA) amplicons has opened up the door to large-scale comparative studies of microbial community structures. The short reads currently produced by massively parallel sequencing technologies make the choice of sequencing region crucial for accurate phylogenetic assignments. While for 16S rDNA, relevant regions have been well described, no truly systematic design of 18S rDNA primers aimed at resolving eukaryotic diversity has yet been reported. Here we used 31,862 18S rDNA sequences to design a set of broad-taxonomic range degenerate PCR primers. We simulated the phylogenetic information that each candidate primer pair would retrieve using paired- or single-end reads of various lengths, representing different sequencing technologies. Primer pairs targeting the V4 region performed best, allowing discrimination with paired-end reads as short as 150 bp (with 75% accuracy at genus level). The conditions for PCR amplification were optimised for one of these primer pairs and this was used to amplify 18S rDNA sequences from isolates as well as from a range of environmental samples which were then Illumina sequenced and analysed, revealing good concordance between expected and observed results. In summary, the reported primer sets will allow minimally biased assessment of eukaryotic diversity in different microbial ecosystems. [less ▲]

Detailed reference viewed: 192 (7 UL)
See detailCommunity integrated omics links the dominance of a microbial generalist to fine-tuned resource usage
Muller, Emilie UL; Pinel, Nicolás; Laczny, Cedric Christian UL et al

Poster (2014)

Microbial communities are complex and dynamic systems that are influenced by stochastic-neutral processes but are mainly structured by resource availability and usage. High-resolution “meta-omics” offer ... [more ▼]

Microbial communities are complex and dynamic systems that are influenced by stochastic-neutral processes but are mainly structured by resource availability and usage. High-resolution “meta-omics” offer exciting prospects to investigate microbial populations in their native environment. In particular, integrated meta-omics, by allowing simultaneous resolution of fundamental niches (genomics) and realised niches (transcriptomics, proteomics and metabolomics), can resolve microbial lifestyles strategies (generalist versus specialist) in situ. We have recently developed the necessary wet- and dry-lab methodologies to carry out systematic molecular measurements of microbial consortia over space and time, and to integrate and analyse the resulting data at the population-level. We applied these methods to oleaginous mixed microbial communities located on the surface of anoxic biological wastewater treatment tanks to investigate how niche breadth (generalist versus specialist strategies) relates to community-level phenotypes and ecological success (i.e. population size). Coupled metabolomics and 16S rRNA gene-based deep sequencing demonstrate that the community-wide lipid accumulation phenotype is associated with the dominance of Candidatus Microthrix parvicella. By integrating population-level genomic reconstructions with transcriptomic and proteomic data, we found that the dominance of this microbial generalist population results from finely tuned resource usage and optimal foraging behaviour. Moreover, the fluctuating environmental conditions constrain the accumulation of variations, leading to a genetically homogeneous population likely due to fitness trade-offs. By integrating metagenomic, metatranscriptomic, metaproteomic and metabolomic information, we demonstrate that natural microbial population sizes and structures are intricately linked to resource usage and that differing microbial lifestyle strategies may explain the varying degrees of within-population genetic heterogeneity observed in metagenomic datasets. Elucidating the exact mechanism driving fitness trade-offs, e.g., antagonistic pleiotropy or others, will require additional integrated omic datasets to be generated from samples taken over space and time. Based on our observations, niche breadth and lifestyle strategies (generalists versus specialists) have to be considered as important factors for understanding the evolutionary processes governing microbial population sizes and structures in situ. [less ▲]

Detailed reference viewed: 123 (12 UL)
See detailCommunity integrated omics links the dominance of a microbial generalist to fine-tuned resource usage
Muller, Emilie UL; Pinel, Nicolás; Laczny, Cedric Christian UL et al

Scientific Conference (2014)

Microbial communities are complex and dynamic systems that are influenced by stochastic-neutral processes but are mainly structured by resource availability and usage. High-resolution “meta-omics” offer ... [more ▼]

Microbial communities are complex and dynamic systems that are influenced by stochastic-neutral processes but are mainly structured by resource availability and usage. High-resolution “meta-omics” offer exciting prospects to investigate microbial populations in their native environment. In particular, integrated meta-omics, by allowing simultaneous resolution of fundamental niches (genomics) and realised niches (transcriptomics, proteomics and metabolomics), can resolve microbial lifestyles (generalist versus specialist lifestyle strategies) in situ. We have recently developed the necessary wet- and dry-lab methodologies to carry out systematic molecular measurements of microbial consortia over space and time, and to integrate and analyse the resulting data at the population-level. We applied these methods to oleaginous mixed microbial communities located on the surface of anoxic biological wastewater treatment tanks to investigate how niche breadth (generalist versus specialist lifestyle strategies) relates to community-level phenotypes and ecological success (i.e. population size). Coupled metabolomics and 16S rRNA gene-based deep sequencing demonstrate that the community-wide lipid accumulation phenotype is associated with the dominance of Candidatus Microthrix parvicella. By integrating population-level genomic reconstructions with transcriptomic and proteomic data, we found that the dominance of this microbial generalist population results from finely tuned resource usage and optimal foraging behaviour. Moreover, the fluctuating environmental conditions constrain the accumulation of variations, leading to a genetically homogeneous population likely due to fitness trade-offs. By integrating metagenomic, metatranscriptomic, metaproteomic and metabolomic information, we demonstrate that natural microbial population sizes and structures are intricately linked to resource usage and that differing microbial lifestyle strategies may explain the varying degrees of within-population genetic heterogeneity observed in metagenomic datasets. Elucidating the exact mechanism driving fitness trade-offs, e.g., antagonistic pleiotropy or others, will require additional integrated omic datasets to be generated from samples taken over space and time. Based on our observations, niche breadth and lifestyle strategies (generalists versus specialists) have to be considered as important factors for understanding the evolutionary processes governing microbial population sizes and structures in situ. [less ▲]

Detailed reference viewed: 145 (9 UL)
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See detailMicrobiome and type 1 diabetes
Wampach, Linda UL; Wilmes, Paul UL; De Beaufort, Carine UL

E-print/Working paper (2013)

The human microbiome (the collective of microorganisms, which inhabit the human body) and changes therein (often referred to as microbial dysbiosis) is emerging as a potential player in the development of ... [more ▼]

The human microbiome (the collective of microorganisms, which inhabit the human body) and changes therein (often referred to as microbial dysbiosis) is emerging as a potential player in the development of type 1 diabetes mellitus. This section discusses the human microbiome and its potential involvement in type 1 diabetes through its central roles in energy metabolism and modulation of the immune system. [less ▲]

Detailed reference viewed: 295 (21 UL)
See detailEco-Systems Biology of activated sludge microbial communities
Wilmes, Paul UL

Presentation (2013, October)

Detailed reference viewed: 9 (0 UL)
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See detailMEMBRANE ASSEMBLY
Shah, Pranjul UL; Wilmes, Paul UL

Patent (2013)

Detailed reference viewed: 98 (8 UL)
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See detailTreg induction by a rationally selected mixture of Clostridia strains from the human microbiota
Atarshi, Koji; Tanoue, Takeshi; Oshima, Kenshiro et al

in Nature (2013), 500

Manipulation of the gut microbiota holds great promise for the treatment of inflammatory and allergic diseases1, 2. Although numerous probiotic microorganisms have been identified3, there remains a ... [more ▼]

Manipulation of the gut microbiota holds great promise for the treatment of inflammatory and allergic diseases1, 2. Although numerous probiotic microorganisms have been identified3, there remains a compelling need to discover organisms that elicit more robust therapeutic responses, are compatible with the host, and can affect a specific arm of the host immune system in a well-controlled, physiological manner. Here we use a rational approach to isolate CD4+FOXP3+ regulatory T (Treg)-cell-inducing bacterial strains from the human indigenous microbiota. Starting with a healthy human faecal sample, a sequence of selection steps was applied to obtain mice colonized with human microbiota enriched in Treg-cell-inducing species. From these mice, we isolated and selected 17 strains of bacteria on the basis of their high potency in enhancing Treg cell abundance and inducing important anti-inflammatory molecules—including interleukin-10 (IL-) and inducible T-cell co-stimulator (ICOS)—in Treg cells upon inoculation into germ-free mice. Genome sequencing revealed that the 17 strains fall within clusters IV, XIVa and XVIII of Clostridia, which lack prominent toxins and virulence factors. The 17 strains act as a community to provide bacterial antigens and a TGF-β-rich environment to help expansion and differentiation of Treg cells. Oral administration of the combination of 17 strains to adult mice attenuated disease in models of colitis and allergic diarrhoea. Use of the isolated strains may allow for tailored therapeutic manipulation of human immune disorders. [less ▲]

Detailed reference viewed: 209 (10 UL)
See detailFrom integrated omics to Eco-Systems Biology of mixed microbial communities
Wilmes, Paul UL

Presentation (2013, July)

Detailed reference viewed: 10 (1 UL)
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See detailReproducibility: In praise of open research measures
Kolker, Eugene; Altintas, Ilkay; Bourne, Philip et al

in Nature (2013), 498

Detailed reference viewed: 187 (14 UL)
See detailMicrobiome-derived small RNA in human circulation
Wilmes, Paul UL

Scientific Conference (2013, June)

Detailed reference viewed: 13 (0 UL)
See detailFrom sludge to gut: towards eco-systems biology of the human microbiome
Wilmes, Paul UL

Scientific Conference (2013, June)

Detailed reference viewed: 8 (0 UL)
See detailFrom integrated omics to Eco-Systems Biology of microbial community-driven processes
Wilmes, Paul UL

Scientific Conference (2013, May)

Detailed reference viewed: 9 (0 UL)
See detailEco-systems biology of microbial communities: integration of biomolecular information from unique samples
Muller, Emilie UL; Roume, Hugo UL; Wilmes, Paul UL

Poster (2013)

In microbial ecology, high-resolution molecular approaches are essential to characterize the vast organismal and functional diversity and to understand the interactions between environmental factors and ... [more ▼]

In microbial ecology, high-resolution molecular approaches are essential to characterize the vast organismal and functional diversity and to understand the interactions between environmental factors and microbial communities (MCs). Molecular eco-systems biology based on the integration of genomics and functional omics, allows conclusive links to be drawn between genetic potential and function. However, the field faced major challenges arising from the heterogeneity and dynamics of MCs. Hence, to facilitate meaningful data integration, analysis and modeling, it is crucial to obtain standardised, reproducible and simultaneous measurements of multiple features from a unique sample. We have developed a new methodological framework for the isolation of high-quality DNA, large and small RNA, proteins and metabolites fractions from undivided MC samples. The methodology is based on cryogenic sample preservation and cell lysis. Metabolites are first extracted using organic solvents, followed by sequential isolation of biomacromolecules using chromatographic spin column technology. The methodology was validated by comparison to commonly used dedicated methods and its broad applicability was demonstrated on MCs of biotechnological, environmental and medical interest. Applying this method to sewage plant MCs has allowed us to determine community-level keystone genes and to probe the functional relevance of the population-level composite genomes, leading to the identification of key players of the community. This methodological framework lays the foundation for standardized molecular eco-systems biology and offers exciting prospects for elucidating the genetic blueprints and the functional relevance of specific populations within MCs, particularly in relation to environment-driven demography changes leading to catastrophic bifurcations. [less ▲]

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See detailFrom meta-omics to causality: experimental models for human microbiome research
Fritz, Joëlle UL; Desai, Mahesh UL; Shah, Pranjul UL et al

in Microbiome (2013), 1(14),

Large-scale ‘meta-omic’ projects are greatly advancing our knowledge of the human microbiome and its specific role in governing health and disease states. A myriad of ongoing studies aim at identifying ... [more ▼]

Large-scale ‘meta-omic’ projects are greatly advancing our knowledge of the human microbiome and its specific role in governing health and disease states. A myriad of ongoing studies aim at identifying links between microbial community disequilibria (dysbiosis) and human diseases. However, due to the inherent complexity and heterogeneity of the human microbiome, cross-sectional, case–control and longitudinal studies may not have enough statistical power to allow causation to be deduced from patterns of association between variables in high-resolution omic datasets. Therefore, to move beyond reliance on the empirical method, experiments are critical. For these, robust experimental models are required that allow the systematic manipulation of variables to test the multitude of hypotheses, which arise from high-throughput molecular studies. Particularly promising in this respect are microfluidics-based in vitro co-culture systems, which allow high-throughput first-pass experiments aimed at proving cause-and-effect relationships prior to testing of hypotheses in animal models. This review focuses on widely used in vivo, in vitro, ex vivo and in silico approaches to study host-microbial community interactions. Such systems, either used in isolation or in a combinatory experimental approach, will allow systematic investigations of the impact of microbes on the health and disease of the human host. All the currently available models present pros and cons, which are described and discussed. Moreover, suggestions are made on how to develop future experimental models that not only allow the study of host-microbiota interactions but are also amenable to high-throughput experimentation. [less ▲]

Detailed reference viewed: 379 (22 UL)