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Jo Handelsman - One of the best experts on this subject based on the ideXlab platform.
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A statistical toolbox for Metagenomics: assessing functional diversity in microbial communities
BMC bioinformatics, 2008Co-Authors: Patrick D Schloss, Jo HandelsmanAbstract:The 99% of bacteria in the environment that are recalcitrant to culturing have spurred the development of Metagenomics, a culture-independent approach to sample and characterize microbial genomes. Massive datasets of metagenomic sequences have been accumulated, but analysis of these sequences has focused primarily on the descriptive comparison of the relative abundance of proteins that belong to specific functional categories. More robust statistical methods are needed to make inferences from metagenomic data. In this study, we developed and applied a suite of tools to describe and compare the richness, membership, and structure of microbial communities using peptide fragment sequences extracted from metagenomic sequence data. Application of these tools to acid mine drainage, soil, and whale fall metagenomic sequence collections revealed groups of peptide fragments with a relatively high abundance and no known function. When combined with analysis of 16S rRNA gene fragments from the same communities these tools enabled us to demonstrate that although there was no overlap in the types of 16S rRNA gene sequence observed, there was a core collection of operational protein families that was shared among the three environments. The results of comparisons between the three habitats were surprising considering the relatively low overlap of membership and the distinctively different characteristics of the three habitats. These tools will facilitate the use of Metagenomics to pursue statistically sound genome-based ecological analyses.
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Metagenomics or Megagenomics
Nature Reviews Microbiology, 2005Co-Authors: Jo HandelsmanAbstract:Metagenomics is an emerging and exciting new field that integrates biology and technology. Here, Jo Handelsman previews the selection of articles in this Focus issue.
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Biotechnological prospects from Metagenomics
Current Opinion in Biotechnology, 2003Co-Authors: Patrick D Schloss, Jo HandelsmanAbstract:The recognition that most microorganisms in the environment cannot be cultured by standard methods stimulated the development of Metagenomics, which is the genomic analysis of uncultured microorganisms. Two types of analysis have been used to obtain information from metagenomic libraries: a function-driven approach, in which metagenomic libraries are initially screened for an expressed trait, and a sequence-driven approach, in which libraries are initially screened for particular DNA sequences. New antibiotics and enzymes are among the early discoveries from Metagenomics. Future refinement of methods that enrich for genes of particular function will accelerate the rate of discovery of useful molecules.
Sean F Brady - One of the best experts on this subject based on the ideXlab platform.
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identification of the colicin v bacteriocin gene cluster by functional screening of a human microbiome metagenomic library
ACS Infectious Diseases, 2018Co-Authors: Sun M Han, Louis J Cohen, Yunhan Huang, Sean F BradyAbstract:The forces that shape human microbial ecology are complex. It is likely that human microbiota, similarly to other microbiomes, use antibiotics as one way to establish an ecological niche. In this study, we use functional Metagenomics to identify human microbial gene clusters that encode for antibiotic functions. Screening of a metagenomic library prepared from a healthy patient stool sample led to the identification of a family of clones with inserts that are 99% identical to a region of a virulence plasmid found in avian pathogenic Escherichia coli. Characterization of the metagenomic DNA sequence identified a colicin V biosynthetic cluster as being responsible for the observed antibiotic effect of the metagenomic clone against E. coli. This study presents a scalable method to recover antibiotic gene clusters from humans using functional Metagenomics and highlights a strategy to study bacteriocins in the human microbiome which can provide a resource for therapeutic discovery.
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expanding small molecule functional Metagenomics through parallel screening of broad host range cosmid environmental dna libraries in diverse proteobacteria
Applied and Environmental Microbiology, 2010Co-Authors: Jeffrey W Craig, Fangyuan Chang, Steven C Obiajulu, Sean F BradyAbstract:The small-molecule biosynthetic diversity encoded within the genomes of uncultured bacteria is an attractive target for the discovery of natural products using functional Metagenomics. Phenotypes commonly associated with the production of small molecules, such as antibiosis, altered pigmentation, or altered colony morphology, are easily identified from screens of arrayed metagenomic library clones. However, functional metagenomic screening methods are limited by their intrinsic dependence on a heterologous expression host. Toward the goal of increasing the small-molecule biosynthetic diversity found in functional metagenomic studies, we report the phenotypic screening of broad-host-range environmental DNA libraries in six different proteobacteria: Agrobacterium tumefaciens, Burkholderia graminis, Caulobacter vibrioides, Escherichia coli, Pseudomonas putida, and Ralstonia metallidurans. Clone-specific small molecules found in culture broth extracts from pigmented and antibacterially active clones, as well as the genetic elements responsible for the biosynthesis of these metabolites, are described. The host strains used in this investigation provided access to unique sets of clones showing minimal overlap, thus demonstrating the potential advantage conferred on functional Metagenomics through the use of multiple diverse host species.
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Handbook of Molecular Microbial Ecology II: Metagenomics in Different Habitats - Expanding Small-Molecule Functional Metagenomics through Parallel Screening of Broad-Host-Range Cosmid Environmental DNA Libraries in Diverse Proteobacteria
Applied and Environmental Microbiology, 2010Co-Authors: Jeffrey W Craig, Fangyuan Chang, Steven C Obiajulu, Sean F BradyAbstract:The small-molecule biosynthetic diversity encoded within the genomes of uncultured bacteria is an attractive target for the discovery of natural products using functional Metagenomics. Phenotypes commonly associated with the production of small molecules, such as antibiosis, altered pigmentation, or altered colony morphology, are easily identified from screens of arrayed metagenomic library clones. However, functional metagenomic screening methods are limited by their intrinsic dependence on a heterologous expression host. Toward the goal of increasing the small-molecule biosynthetic diversity found in functional metagenomic studies, we report the phenotypic screening of broad-host-range environmental DNA libraries in six different proteobacteria: Agrobacterium tumefaciens, Burkholderia graminis, Caulobacter vibrioides, Escherichia coli, Pseudomonas putida, and Ralstonia metallidurans. Clone-specific small molecules found in culture broth extracts from pigmented and antibacterially active clones, as well as the genetic elements responsible for the biosynthesis of these metabolites, are described. The host strains used in this investigation provided access to unique sets of clones showing minimal overlap, thus demonstrating the potential advantage conferred on functional Metagenomics through the use of multiple diverse host species.
Robin Patel - One of the best experts on this subject based on the ideXlab platform.
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identification of prosthetic joint infection pathogens using a shotgun Metagenomics approach
Clinical Infectious Diseases, 2018Co-Authors: Matthew Thoendel, Patricio Jeraldo, Arlen D Hanssen, Matthew P Abdel, Nicholas Chia, Janet Yao, Kerryl E Greenwoodquaintance, Robin PatelAbstract:Background Metagenomic shotgun sequencing has the potential to change how many infections, particularly those caused by difficult-to-culture organisms, are diagnosed. Metagenomics was used to investigate prosthetic joint infections (PJIs), where pathogen detection can be challenging. Methods Four hundred eight sonicate fluid samples generated from resected hip and knee arthroplasties were tested, including 213 from subjects with infections and 195 from subjects without infection. Samples were enriched for microbial DNA using the MolYsis basic kit, whole-genome amplified, and sequenced using Illumina HiSeq 2500 instruments. A pipeline was designed to screen out human reads and analyze remaining sequences for microbial content using the Livermore Metagenomics Analysis Toolkit and MetaPhlAn2 tools. Results When compared to sonicate fluid culture, Metagenomics was able to identify known pathogens in 94.8% (109/115) of culture-positive PJIs, with additional potential pathogens detected in 9.6% (11/115). New potential pathogens were detected in 43.9% (43/98) of culture-negative PJIs, 21 of which had no other positive culture sources from which these microorganisms had been detected. Detection of microorganisms in samples from uninfected aseptic failure cases was conversely rare (7/195 [3.6%] cases). The presence of human and contaminant microbial DNA from reagents was a challenge, as previously reported. Conclusions Metagenomic shotgun sequencing is a powerful tool to identify a wide range of PJI pathogens, including difficult-to-detect pathogens in culture-negative infections.
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A Novel Prosthetic Joint Infection Pathogen, Mycoplasma salivarium, Identified by Metagenomic Shotgun Sequencing.
Clinical infectious diseases : an official publication of the Infectious Diseases Society of America, 2017Co-Authors: Matthew Thoendel, Patricio Jeraldo, Matthew P Abdel, Nicholas Chia, Kerryl E. Greenwood-quaintance, James M. Steckelberg, Douglas R. Osmon, Robin PatelAbstract:Defining the microbial etiology of culture-negative prosthetic joint infection (PJI) can be challenging. Metagenomic shotgun sequencing is a new tool to identify organisms undetected by conventional methods. We present a case where Metagenomics was used to identify Mycoplasma salivarium as a novel PJI pathogen in a patient with hypogammaglobulinemia.
Matthew Thoendel - One of the best experts on this subject based on the ideXlab platform.
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identification of prosthetic joint infection pathogens using a shotgun Metagenomics approach
Clinical Infectious Diseases, 2018Co-Authors: Matthew Thoendel, Patricio Jeraldo, Arlen D Hanssen, Matthew P Abdel, Nicholas Chia, Janet Yao, Kerryl E Greenwoodquaintance, Robin PatelAbstract:Background Metagenomic shotgun sequencing has the potential to change how many infections, particularly those caused by difficult-to-culture organisms, are diagnosed. Metagenomics was used to investigate prosthetic joint infections (PJIs), where pathogen detection can be challenging. Methods Four hundred eight sonicate fluid samples generated from resected hip and knee arthroplasties were tested, including 213 from subjects with infections and 195 from subjects without infection. Samples were enriched for microbial DNA using the MolYsis basic kit, whole-genome amplified, and sequenced using Illumina HiSeq 2500 instruments. A pipeline was designed to screen out human reads and analyze remaining sequences for microbial content using the Livermore Metagenomics Analysis Toolkit and MetaPhlAn2 tools. Results When compared to sonicate fluid culture, Metagenomics was able to identify known pathogens in 94.8% (109/115) of culture-positive PJIs, with additional potential pathogens detected in 9.6% (11/115). New potential pathogens were detected in 43.9% (43/98) of culture-negative PJIs, 21 of which had no other positive culture sources from which these microorganisms had been detected. Detection of microorganisms in samples from uninfected aseptic failure cases was conversely rare (7/195 [3.6%] cases). The presence of human and contaminant microbial DNA from reagents was a challenge, as previously reported. Conclusions Metagenomic shotgun sequencing is a powerful tool to identify a wide range of PJI pathogens, including difficult-to-detect pathogens in culture-negative infections.
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Direct Detection and Identification of Prosthetic Joint Pathogens in Synovial Fluid (SF) by Metagenomic Shotgun Sequencing
Open Forum Infectious Diseases, 2017Co-Authors: Morgan Ivy, Matthew Thoendel, Patricio Jeraldo, Arlen D Hanssen, Matthew P Abdel, Nicholas Chia, Aaron J Tande, Kerryl E. Greenwood-quaintance, Janet Yao, Jayawant N MandrekarAbstract:Abstract Background Detection and identification of microorganism(s) involved in periprosthetic joint infection (PJI) can inform surgical management and directed antibiotic therapy. Metagenomic shotgun sequencing is a powerful tool with the potential to change how many PJIs are diagnosed as it allows direct detection and identification of pathogens in clinical specimens. In the largest series to date, we utilized a Metagenomics-based approach applied to SF to define potential microbial etiologies of failed total knee arthroplasties (TKAs). Methods Synovial fluid was collected from 112 failed TKAs [74 PJI and 38 aseptic implant failure (AF)] via preoperative arthrocentesis. Cell count and differential, standardized culture and DNA-based metagenomic shotgun sequencing were performed. Human DNA was depleted using the MolYsis basic kit prior to DNA extraction, whole genome amplification, and sequencing. Taxonomic assignment of reads and pathogen identification was achieved using a pipeline incorporating k-mer- and marker gene-based classification software. A scheme for analysis and filtration of false-positives was created and applied, incorporating cut-offs for the number of reads, quality scores, and coverage across a reference genome. Patients were classified as having PJI using the IDSA criteria and expert review. Analyses were recorded as percent agreement, with 95% confidence intervals (CI), of Metagenomics to SF culture. Results Metagenomic analysis identified the known pathogen in 54 (90%) (CI, 79.5%–96.2%) of the 60 culture-positive PJIs analyzed and one (2%) (CI, 0.0%–8.9%) potential polymicrobial infection not detected by culture. For the 14 culture-negative PJIs tested, Metagenomics showed 79% (CI, 49.2%–95.3%) agreement for negative findings; potential pathogens were identified in three (21%) (CI, 4.7%–50.8%) culture-negative PJI cases, with one being polymicrobial. Of the 37 culture-negative AF cases, Metagenomics showed 97% (CI, 85.8%–99.9%) agreement with negative culture and identified one (3%) (CI, 0.0%–14.2%) potential pathogen. For the one culture-positive AF case, metagenomic results were negative, suggesting possible culture contamination. Conclusion Metagenomic shotgun sequencing performed on SF can be used to diagnose PJI and may be particularly useful for culture-negative PJI. Disclosures R. Patel, ASM: Board Member, None; CD Diagnostics, BioFire, Curetis, Merck, Hutchison Biofilm Medical Solutions, Accelerate Diagnostics, Allergan, and The Medicines Company: Grant Investigator, Grant recipient; Curetis: Consultant, Monies paid to my employer; A patent on Bordetella pertussis/parapertussis PCR issued, a patent on a device/method for sonication with royalties paid by Samsung to Mayo Clinic, and a patent on an anti-biofilm substance issued: Patents, Patents, any money is paid to my employer; Actelion: DSMB, Money paid to my employer; ASM and IDSA: Editor’s stipends, Editor’s stipends; NBME, Up-to-Date and the Infectious Diseases Board Review Course: NBME, Up-to-Date and the Infectious Diseases Board Review Course, Honoraria; Roche, ASM, and IDSA: Travel reimbursement, Travel reimbursement
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A Novel Prosthetic Joint Infection Pathogen, Mycoplasma salivarium, Identified by Metagenomic Shotgun Sequencing.
Clinical infectious diseases : an official publication of the Infectious Diseases Society of America, 2017Co-Authors: Matthew Thoendel, Patricio Jeraldo, Matthew P Abdel, Nicholas Chia, Kerryl E. Greenwood-quaintance, James M. Steckelberg, Douglas R. Osmon, Robin PatelAbstract:Defining the microbial etiology of culture-negative prosthetic joint infection (PJI) can be challenging. Metagenomic shotgun sequencing is a new tool to identify organisms undetected by conventional methods. We present a case where Metagenomics was used to identify Mycoplasma salivarium as a novel PJI pathogen in a patient with hypogammaglobulinemia.
Mya Breitbart - One of the best experts on this subject based on the ideXlab platform.
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A field guide to eukaryotic circular single-stranded DNA viruses: insights gained from Metagenomics
Archives of Virology, 2012Co-Authors: Karyna Rosario, Siobain Duffy, Mya BreitbartAbstract:Despite their small size and limited protein-coding capacity, the rapid evolution rates of single-stranded DNA (ssDNA) viruses have led to their emergence as serious plant and animal pathogens. Recently, Metagenomics has revealed an unprecedented diversity of ssDNA viruses, expanding their known environmental distributions and host ranges. This review summarizes and contrasts the basic characteristics of known circular ssDNA viral groups, providing a resource for analyzing the wealth of ssDNA viral sequences identified through Metagenomics. Since ssDNA viruses are largely identified based on conserved rolling circle replication proteins, this review highlights distinguishing motifs and catalytic residues important for replication. Genomes identified through Metagenomics have demonstrated unique ssDNA viral genome architectures and revealed characteristics that blur the boundaries between previously well-defined groups. Metagenomic discovery of ssDNA viruses has created both a challenge to current taxonomic classification schemes and an opportunity to revisit hypotheses regarding the evolutionary history of these viruses.
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Exploring the viral world through Metagenomics.
Current opinion in virology, 2011Co-Authors: Karyna Rosario, Mya BreitbartAbstract:Viral Metagenomics, or shotgun sequencing of purified viral particles, has revolutionized the field of environmental virology by allowing the exploration of viral communities in a variety of sample types throughout the biosphere. The introduction of viral Metagenomics has demonstrated that dominant viruses in environmental communities are not well-represented by the cultured viruses in existing sequence databases. Viral metagenomic studies have provided insights into viral ecology by elucidating the genetic potential, community structure, and biogeography of environmental viruses. In addition, viral Metagenomics has expanded current knowledge of virus-host interactions by uncovering genes that may allow viruses to manipulate their hosts in unexpected ways. The intrinsic potential for virus discovery through viral Metagenomics can help advance a wide array of disciplines including evolutionary biology, pathogen surveillance, and biotechnology.