De novo metagenomic assembly reveals abundant novel major lineage of Archaea in hypersaline microbial communities

Abstract Background The intestinal microbiota plays a crucial role in protecting the host from pathogenic microbes, modulating immunity and regulating metabolic processes. We studied the simplified human intestinal microbiota (SIHUMIx) consisting of eight bacterial species with a particular focus on the discovery of novel small proteins with less than 100 amino acids (= sProteins), some of which may contribute to shape the simplified human intestinal microbiota. Although sProteins carry out a wide range of important functions, they are still often missed in genome annotations, and little is known about their structure and function in individual microbes and especially in microbial communities. Results We created a multi-species integrated proteogenomics search database (iPtgxDB) to enable a comprehensive identification of novel sProteins. Six of the eight SIHUMIx species, for which no complete genomes were available, were sequenced and de novo assembled. Several proteomics approaches including two earlier optimized sProtein enrichment strategies were applied to specifically increase the chances for novel sProtein discovery. The search of tandem mass spectrometry (MS/MS) data against the multi-species iPtgxDB enabled the identification of 31 novel sProteins, of which the expression of 30 was supported by metatranscriptomics data. Using synthetic peptides, we were able to validate the expression of 25 novel sProteins. The comparison of sProtein expression in each single strain versus a multi-species community cultivation showed that six of these sProteins were only identified in the SIHUMIx community indicating a potentially important role of sProteins in the organization of microbial communities. Two of these novel sProteins have a potential antimicrobial function. Metabolic modelling revealed that a third sProtein is located in a genomic region encoding several enzymes relevant for the community metabolism within SIHUMIx. Conclusions We outline an integrated experimental and bioinformatics workflow for the discovery of novel sProteins in a simplified intestinal model system that can be generically applied to other microbial communities. The further analysis of novel sProteins uniquely expressed in the SIHUMIx multi-species community is expected to enable new insights into the role of sProteins on the functionality of bacterial communities such as those of the human intestinal tract.

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Oral Spirochetes Implicated in Dental Diseases Are Widespread in Normal Human Subjects and Carry Extremely Diverse Integron Gene Cassettes

Applied and Environmental Microbiology ◽

10.1128/aem.00564-12 ◽

2012 ◽

Vol 78 (15) ◽

pp. 5288-5296 ◽

Cited By ~ 17

Author(s):

Yu-Wei Wu ◽

Mina Rho ◽

Thomas G. Doak ◽

Yuzhen Ye

Keyword(s):

Microbial Communities ◽

De Novo ◽

Human Subjects ◽

Human Microbiome ◽

Metagenomic Data ◽

De Bruijn Graph ◽

Content Type ◽

Gene Cassettes ◽

Dental Diseases ◽

Normal Human

ABSTRACTThe NIH Human Microbiome Project (HMP) has produced several hundred metagenomic data sets, allowing studies of the many functional elements in human-associated microbial communities. Here, we survey the distribution of oral spirochetes implicated in dental diseases in normal human individuals, using recombination sites associated with the chromosomal integron inTreponemagenomes, taking advantage of the multiple copies of the integron recombination sites (repeats) in the genomes, and using a targeted assembly approach that we have developed. We find that integron-containingTreponemaspecies are present in ∼80% of the normal human subjects included in the HMP. Further, we are able tode novoassemble the integron gene cassettes using our constrained assembly approach, which employs a unique application of the de Bruijn graph assembly information; most of these cassette genes were not assembled in whole-metagenome assemblies and could not be identified by mapping sequencing reads onto the known referenceTreponemagenomes due to the dynamic nature of integron gene cassettes. Our study significantly enriches the gene pool known to be carried byTreponemachromosomal integrons, totaling 826 (598 97% nonredundant) genes. We characterize the functions of these gene cassettes: many of these genes have unknown functions. The integron gene cassette arrays found in the human microbiome are extraordinarily dynamic, with different microbial communities sharing only a small number of common genes.

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MetaCompass: Reference-guided Assembly of Metagenomes

10.1101/212506 ◽

2017 ◽

Cited By ~ 7

Author(s):

Victoria Cepeda ◽

Bo Liu ◽

Mathieu Almeida ◽

Christopher M. Hill ◽

Sergey Koren ◽

...

Keyword(s):

De Novo ◽

Human Microbiome ◽

Human Microbiome Project ◽

Bacterial Genomes ◽

Guided Assembly ◽

Metagenomic Assembly

ABSTRACTMetagenomic studies have primarily relied on de novo approaches for reconstructing genes and genomes from microbial mixtures. While database driven approaches have been employed in certain analyses, they have not been used in the assembly of metagenomes. Here we describe the first effective approach for reference-guided metagenomic assembly of low-abundance bacterial genomes that can complement and improve upon de novo metagenomic assembly methods. When combined with de novo assembly approaches, we show that MetaCompass can generate more complete assemblies than can be obtained by de novo assembly alone, and improve on assemblies from the Human Microbiome Project (over 2,000 samples).

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Choice of assembly software has a critical impact on virome characterisation

10.1101/479105 ◽

2018 ◽

Author(s):

Thomas D.S. Sutton ◽

Adam G. Clooney ◽

Feargal J. Ryan ◽

R. Paul Ross ◽

Colin Hill

Keyword(s):

De Novo ◽

Vital Role ◽

Well Performance ◽

Community Members ◽

Reference Databases ◽

Downstream Analysis ◽

Metagenomic Assembly ◽

Assembly Performance ◽

Genomic Repeats

AbstractBackgroundThe viral component of microbial communities play a vital role in driving bacterial diversity, facilitating nutrient turnover and shaping community composition. Despite their importance, the vast majority of viral sequences are poorly annotated and share little or no homology to reference databases. As a result, investigation of the viral metagenome (virome) relies heavily on de novo assembly of short sequencing reads to recover compositional and functional information. Metagenomic assembly is particularly challenging for virome data, often resulting in fragmented assemblies and poor recovery of viral community members. Despite the essential role of assembly in virome analysis and difficulties posed by these data, current assembly comparisons have been limited to subsections of virome studies or bacterial datasets.DesignThis study presents the most comprehensive virome assembly comparison to date, featuring 16 metagenomic assembly approaches which have featured in human virome studies. Assemblers were assessed using four independent virome datasets, namely; simulated reads, two mock communities, viromes spiked with a known phage and human gut viromes.ResultsAssembly performance varied significantly across all test datasets, with SPAdes (meta) performing consistently well. Performance of MIRA and VICUNA varied, highlighting the importance of using a range of datasets when comparing assembly programs. It was also found that while some assemblers addressed the challenges of virome data better than others, all assemblers had limitations. Low read coverage and genomic repeats resulted in assemblies with poor genome recovery, high degrees of fragmentation and low accuracy contigs across all assemblers. These limitations must be considered when setting thresholds for downstream analysis and when drawing conclusions from virome data.

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CAMISIM: Simulating metagenomes and microbial communities

10.1101/300970 ◽

2018 ◽

Cited By ~ 4

Author(s):

Adrian Fritz ◽

Peter Hofmann ◽

Stephan Majda ◽

Eik Dahms ◽

Johannes Dröge ◽

...

Keyword(s):

Microbial Communities ◽

De Novo ◽

Real Data ◽

Small Data ◽

Data Sets ◽

Sequencing Data ◽

Taxonomic Profiling ◽

Benchmark Data ◽

Sequencing Technologies ◽

Wide Range

Shotgun metagenome data sets of microbial communities are highly diverse, not only due to the natural variation of the underlying biological systems, but also due to differences in laboratory protocols, replicate numbers, and sequencing technologies. Accordingly, to effectively assess the performance of metagenomic analysis software, a wide range of benchmark data sets are required. Here, we describe the CAMISIM microbial community and metagenome simulator. The software can model different microbial abundance profiles, multi-sample time series and differential abundance studies, includes real and simulated strain-level diversity, and generates second and third generation sequencing data from taxonomic profiles or de novo. Gold standards are created for sequence assembly, genome binning, taxonomic binning, and taxonomic profiling. CAMSIM generated the benchmark data sets of the first CAMI challenge. For two simulated multi-sample data sets of the human and mouse gut microbiomes we observed high functional congruence to the real data. As further applications, we investigated the effect of varying evolutionary genome divergence, sequencing depth, and read error profiles on two popular metagenome assemblers, MEGAHIT and metaSPAdes, on several thousand small data sets generated with CAMISIM. CAMISIM can simulate a wide variety of microbial communities and metagenome data sets together with truth standards for method evaluation. All data sets and the software are freely available at: https://github.com/CAMI-challenge/CAMISIM

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Evaluating the Fidelity of De Novo Short Read Metagenomic Assembly Using Simulated Data

PLoS ONE ◽

10.1371/journal.pone.0019984 ◽

2011 ◽

Vol 6 (5) ◽

pp. e19984 ◽

Cited By ~ 54

Author(s):

Miguel Pignatelli ◽

Andrés Moya

Keyword(s):

De Novo ◽

Simulated Data ◽

Short Read ◽

Metagenomic Assembly

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A de novo metagenomic assembly program for shotgun DNA reads

Bioinformatics ◽

10.1093/bioinformatics/bts162 ◽

2012 ◽

Vol 28 (11) ◽

pp. 1455-1462 ◽

Cited By ~ 35

Author(s):

Binbin Lai ◽

Ruogu Ding ◽

Yang Li ◽

Liping Duan ◽

Huaiqiu Zhu

Keyword(s):

De Novo ◽

Metagenomic Assembly

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Elucidation of roles for vitamin B12 in regulation of folate, ubiquinone, and methionine metabolism

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1612360114 ◽

2017 ◽

Vol 114 (7) ◽

pp. E1205-E1214 ◽

Cited By ~ 37

Author(s):

Margaret F. Romine ◽

Dmitry A. Rodionov ◽

Yukari Maezato ◽

Lindsey N. Anderson ◽

Premchendar Nandhikonda ◽

...

Keyword(s):

Vitamin B12 ◽

Microbial Communities ◽

Binding Proteins ◽

De Novo ◽

Transcriptional Regulator ◽

Vitamin B ◽

Methionine Metabolism ◽

Community Metabolism ◽

Light Sensing ◽

Allosteric Effector

Only a small fraction of vitamin B12-requiring organisms are able to synthesize B12 de novo, making it a common commodity in microbial communities. Initially recognized as an enzyme cofactor of a few enzymes, recent studies have revealed additional B12-binding enzymes and regulatory roles for B12. Here we report the development and use of a B12-based chemical probe to identify B12-binding proteins in a nonphototrophic B12-producing bacterium. Two unexpected discoveries resulted from this study. First, we identified a light-sensing B12-binding transcriptional regulator and demonstrated that it controls folate and ubiquinone biosynthesis. Second, our probe captured proteins involved in folate, methionine, and ubiquinone metabolism, suggesting that it may play a role as an allosteric effector of these processes. These metabolic processes produce precursors for synthesis of DNA, RNA, and protein. Thereby, B12 likely modulates growth, and by limiting its availability to auxotrophs, B12-producing organisms may facilitate coordination of community metabolism.

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Diverse DNA modification in marine prokaryotic and viral communities

10.1101/2021.05.08.442635 ◽

2021 ◽

Author(s):

Satoshi Hiraoka ◽

Tomomi Sumida ◽

Miho Hirai ◽

Atsushi Toyoda ◽

Shinsuke Kawagucci ◽

...

Keyword(s):

Microbial Communities ◽

Single Molecule ◽

Evolutionary History ◽

Defense Mechanism ◽

Dna Methyltransferases ◽

Dna Modifications ◽

History Of ◽

Viral Communities ◽

Metagenomic Assembly ◽

Marine Microbial Communities

Chemical modifications of DNA, including methylation, play an important role in prokaryotes and viruses. However, our knowledge of the modification systems in environmental microbial communities, typically dominated by members not yet cultured, is limited. Here, we conducted 'metaepigenomic' analyses by single-molecule real-time sequencing of marine microbial communities. In total, 233 and 163 metagenomic assembly genomes (MAGs) were constructed from diverse prokaryotes and viruses, respectively, and 220 modified motifs and 276 DNA methyltransferases (MTases) were identified. Most of the MTases were not associated with the defense mechanism. The MTase-motif correspondence found in the MAGs revealed 10 novel pairs, and experimentally confirmed the catalytic specificities of the MTases. We revealed novel alternative motifs in the methylation system that are highly conserved in Alphaproteobacteria, illuminating the co-evolutionary history of the methylation system and host genome. Our findings highlight diverse unexplored DNA modifications that potentially affect the ecology and evolution of prokaryotes and viruses.

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Corynebacterium Comparative Genomics Reveals a Role for Cobamide Sharing in the Skin Microbiome

10.1101/2020.12.02.407395 ◽

2020 ◽

Author(s):

Mary Hannah Swaney ◽

Lindsay R Kalan

Keyword(s):

Comparative Genomics ◽

Microbial Communities ◽

Community Dynamics ◽

De Novo ◽

Skin Barrier ◽

Skin Microbiome ◽

Bacterial Phyla ◽

Host Interactions ◽

Complex Interactions ◽

Associated Species

ABSTRACTThe human skin microbiome is a key player in human health, with diverse functions ranging from defense against pathogens to education of the immune system. Recent studies have begun unraveling the complex interactions within skin microbial communities, shedding light on the invaluable role that skin microorganisms have in maintaining a healthy skin barrier. While the Corynebacterium genus is a dominant taxon of the skin microbiome, relatively little is known how skin-associated Corynebacteria contribute to microbe-microbe and microbe-host interactions on the skin. Here, we performed a comparative genomics analysis of 71 Corynebacterium species from diverse ecosystems, which revealed functional differences between host- and environment-associated species. In particular, host-associated species were enriched for de novo biosynthesis of cobamides, which are a class of cofactor essential for metabolism in organisms across the tree of life but are produced by a limited number of prokaryotes. Because cobamides have been hypothesized to mediate community dynamics within microbial communities, we analyzed skin metagenomes for Corynebacterium cobamide producers, which revealed a positive correlation between cobamide producer abundance and microbiome diversity, a trait associated with skin health. We also provide the first metagenome-based assessment of cobamide biosynthesis and utilization in the skin microbiome, showing that both dominant and low abundant skin taxa encode for the de novo biosynthesis pathway and that cobamide-dependent enzymes are encoded by phylogenetically diverse taxa across the major bacterial phyla on the skin. Taken together, our results support a role for cobamide sharing within skin microbial communities, which we hypothesize mediates community dynamics.

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