Potential Applications of Human Viral Metagenomics and Reference Materials: Considerations for Current and Future Viruses.
28 Oct 2020-Applied and Environmental Microbiology (American Society for Microbiology)-Vol. 86, Iss: 22
TL;DR: This work states that implementation of appropriate viral reference materials can aid in the benchmarking of current and development of novel assays for virus identification, discovery, and surveillance, and results will continue to translate into diverse applications.
Abstract: Viruses are ubiquitous particles comprising genetic material that can infect bacteria, archaea, and fungi, as well as human and other animal cells. Given that determining virus composition and function in association with states of human health and disease is of increasing interest, we anticipate that the field of viral metagenomics will continue to expand and be applied in a variety of areas ranging from surveillance to discovery and will rely heavily upon the continued development of reference materials and databases. Information regarding viral composition and function readily translates into biological and clinical applications, including the rapid sequence identification of pathogenic viruses in various sample types. However, viral metagenomic approaches often lack appropriate standards and reference materials to enable cross-study comparisons and assess potential biases which can be introduced at the various stages of collection, storage, processing, and sequence analysis. In addition, implementation of appropriate viral reference materials can aid in the benchmarking of current and development of novel assays for virus identification, discovery, and surveillance. As the field of viral metagenomics expands and standardizes, results will continue to translate into diverse applications.
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TL;DR: An overview of viral metagenomics is presented, with updates regarding the relations between alterations in the human virome and the pathogenesis of human diseases, recent findings related to COVID-19, and therapeutic applications related to thehuman virome.
Abstract: The human body is colonized by a wide range of microorganisms. The field of viromics has expanded since the first reports on the detection of viruses via metagenomic sequencing in 2002. With the continued development of reference materials and databases, viral metagenomic approaches have been used to explore known components of the virome and discover new viruses from various types of samples. The virome has attracted substantial interest since the outbreak of the coronavirus disease 2019 (COVID-19) pandemic. Increasing numbers of studies and review articles have documented the diverse virome in various sites in the human body, as well as interactions between the human host and the virome with regard to health and disease. However, there have been few studies of direct causal relationships. Viral metagenomic analyses often lack standard references and are potentially subject to bias. Moreover, most virome-related review articles have focused on the gut virome and did not investigate the roles of the virome in other sites of the body in human disease. This review presents an overview of viral metagenomics, with updates regarding the relations between alterations in the human virome and the pathogenesis of human diseases, recent findings related to COVID-19, and therapeutic applications related to the human virome.
22 citations
TL;DR:
Abstract: Motivation The growing number of microbial reference genomes enables the improvement of metagenomic profiling accuracy but also imposes greater requirements on the indexing efficiency, database size, and runtime of taxonomic profilers. Additionally, most profilers focus mainly on bacterial, archaeal, and fungal populations, while less attention is paid to viral communities. Results We present KMCP, a novel k-mer-based metagenomic profiling tool that utilizes genome coverage information by splitting the reference genomes into chunks and then stores k-mers in a modified and optimized COBS index for fast alignment-free sequence searching. KMCP combines k-mer similarity and genome coverage information to reduce the false positive rate of k-mer-based taxonomic classification and profiling methods. Benchmarking results based on simulated and real data demonstrate that KMCP, despite a longer running time than all other methods, not only allows the accurate taxonomic profiling of prokaryotic and viral populations but also provides confident pathogen detection in clinical samples of low depth. Availability and Implementation The software is open-source under the MIT license and available at https://github.com/shenwei356/kmcp.
5 citations
TL;DR: Increased understanding of the viral dark matter will continue with a combination of cultivation, microscopy, sequencing, and bioinformatic efforts, which are discussed in the present review.
Abstract: Viruses are part of the microbiome and have essential roles in immunology, evolution, biogeochemical cycles, health, and disease progression. Viruses influence a wide variety of systems and processes, and the continued discovery of novel viruses is anticipated to reveal new mechanisms influencing the biology of diverse environments. While the identity and roles of viruses continue to be discovered and understood through viral metagenomics, most of the sequences in virome datasets cannot be attributed to known viruses or may be only distantly related to species already described in public sequence databases, at best. Such viruses are known as the viral dark matter. Ongoing discoveries from the viral dark matter have provided insights into novel viruses from a variety of environments, as well as their potential in immunological processes, virus evolution, health, disease, therapeutics, and surveillance. Increased understanding of the viral dark matter will continue with a combination of cultivation, microscopy, sequencing, and bioinformatic efforts, which are discussed in the present review.
5 citations
TL;DR: In this paper, a review of the application of multi-omics techniques including genomics, transcriptomics, meta-transcriptomics, proteomics/meta-proteomics, and metabolomics to generate multiple datasets from a single sample have facilitated hypothesis generation leading to the identification of biological, molecular and ecological functions and mechanisms.
1 citations
08 Mar 2022
TL;DR: Li et al. as discussed by the authors presented KMCP, a k-mer-based metagenomic profiling tool that utilizes genome coverage information by splitting the reference genomes into chunks and then stores k-mers in a modified and optimized COBS index for fast alignment-free sequence searching.
Abstract: Abstract Motivation The growing number of microbial reference genomes enables the improvement of metagenomic profiling accuracy but also imposes greater requirements on the indexing efficiency, database size, and runtime of taxonomic profilers. Additionally, most profilers focus mainly on bacterial, archaeal, and fungal populations, while less attention is paid to viral communities. Results We present KMCP, a novel k -mer-based metagenomic profiling tool that utilizes genome coverage information by splitting the reference genomes into chunks and then stores k -mers in a modified and optimized COBS index for fast alignment-free sequence searching. KMCP combines k -mer similarity and genome coverage information to reduce the false positive rate of k -mer-based taxonomic classification and profiling methods. Benchmarking results based on simulated and real data demonstrate that KMCP, despite a longer running time than all other methods, not only allows the accurate taxonomic profiling of prokaryotic and viral populations but also provides confident pathogen detection in clinical samples of low depth. Availability and Implementation The software is open-source under the MIT license and available at https://github.com/shenwei356/kmcp .
1 citations
References
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TL;DR: This work presents an improved method for sequencing variable regions within the 16S rRNA gene using Illumina's MiSeq platform, which is currently capable of producing paired 250-nucleotide reads and demonstrates that it can provide data that are at least as good as that generated by the 454 platform while providing considerably higher sequencing coverage for a fraction of the cost.
Abstract: Rapid advances in sequencing technology have changed the experimental landscape of microbial ecology. In the last 10 years, the field has moved from sequencing hundreds of 16S rRNA gene fragments per study using clone libraries to the sequencing of millions of fragments per study using next-generation sequencing technologies from 454 and Illumina. As these technologies advance, it is critical to assess the strengths, weaknesses, and overall suitability of these platforms for the interrogation of microbial communities. Here, we present an improved method for sequencing variable regions within the 16S rRNA gene using Illumina's MiSeq platform, which is currently capable of producing paired 250-nucleotide reads. We evaluated three overlapping regions of the 16S rRNA gene that vary in length (i.e., V34, V4, and V45) by resequencing a mock community and natural samples from human feces, mouse feces, and soil. By titrating the concentration of 16S rRNA gene amplicons applied to the flow cell and using a quality score-based approach to correct discrepancies between reads used to construct contigs, we were able to reduce error rates by as much as two orders of magnitude. Finally, we reprocessed samples from a previous study to demonstrate that large numbers of samples could be multiplexed and sequenced in parallel with shotgun metagenomes. These analyses demonstrate that our approach can provide data that are at least as good as that generated by the 454 platform while providing considerably higher sequencing coverage for a fraction of the cost.
5,417 citations
"Potential Applications of Human Vir..." refers background in this paper
...However, random amplification methods may result in the overrepresentation of certain viruses, including ssDNA viruses....
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...Development of novel viral reference materials may aid to determine biases that can be introduced when testing current and novel reagents for viral nucleic acid extraction or conversion of RNA to cDNA, for instance....
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...Specifically, the term “mock community” arose to refer to a mix of bacterial DNA (52) and is currently used to refer to a mix of bacterial or fungal cells or DNA, as well as viral particles or nucleic acids in specific concentrations....
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...For instance, a study assessing the effect of MDA found the overrepresentation of bacteriophage M13 in a mock community composed of the seven DNA viruses bacteriophage lambda, vaccinia virus, phage phi29, adenovirus, bacteriophage M13, mouse minute virus p, and a porcine circovirus (31)....
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...For instance, a study evaluating the nucleic acid extraction efficiency from HeLa cells spiked with four viruses, including the double-stranded DNA Epstein-Barr virus, doublestranded RNA reovirus 3, single-stranded RNA feline leukemia virus, and respiratory syncytial virus, using 11 commercially available extraction kits based on silica membrane column, magnetic bead, and precipitation-based extractions found that dual extraction methods may provide improved sensitivity for recovering nucleic acids from viruses with specific biochemical and biophysical characteristics (41)....
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TL;DR: Genomic and evolutionary evidence of the occurrence of a SARS-CoV-2-like CoV (named Pangolin-Cov) in dead Malayan pangolins is found and suggests that pangolin species are a natural reservoir of SARS
1,213 citations
"Potential Applications of Human Vir..." refers background in this paper
...Although it has been demonstrated that bats may have been the original reservoirs of the novel coronavirus, genomic evidence has also shown that the pangolin may have served as an intermediate host (60)....
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Veterans Health Administration1, University of California, San Diego2, University of Montana3, J. Craig Venter Institute4, Wellcome Trust Sanger Institute5, Texas A&M University6, University of Illinois at Urbana–Champaign7, University of Pittsburgh8, Universidad Autónoma de Nuevo León9, Columbia University10, University of Alberta11, Cornell University12, Autonomous University of Barcelona13, King's College London14, French Institute of Health and Medical Research15, University of Wisconsin-Madison16, Yale University17, Université catholique de Louvain18
TL;DR: It is shown that bacteriophages can specifically target cytolytic E. faecalis, which provides a method for precisely editing the intestinal microbiota, and is linked with more severe clinical outcomes and increased mortality in patients with alcoholic hepatitis.
Abstract: Chronic liver disease due to alcohol-use disorder contributes markedly to the global burden of disease and mortality1–3. Alcoholic hepatitis is a severe and life-threatening form of alcohol-associated liver disease. The gut microbiota promotes ethanol-induced liver disease in mice4, but little is known about the microbial factors that are responsible for this process. Here we identify cytolysin—a two-subunit exotoxin that is secreted by Enterococcus faecalis5,6—as a cause of hepatocyte death and liver injury. Compared with non-alcoholic individuals or patients with alcohol-use disorder, patients with alcoholic hepatitis have increased faecal numbers of E. faecalis. The presence of cytolysin-positive (cytolytic) E. faecalis correlated with the severity of liver disease and with mortality in patients with alcoholic hepatitis. Using humanized mice that were colonized with bacteria from the faeces of patients with alcoholic hepatitis, we investigated the therapeutic effects of bacteriophages that target cytolytic E. faecalis. We found that these bacteriophages decrease cytolysin in the liver and abolish ethanol-induced liver disease in humanized mice. Our findings link cytolytic E. faecalis with more severe clinical outcomes and increased mortality in patients with alcoholic hepatitis. We show that bacteriophages can specifically target cytolytic E. faecalis, which provides a method for precisely editing the intestinal microbiota. A clinical trial with a larger cohort is required to validate the relevance of our findings in humans, and to test whether this therapeutic approach is effective for patients with alcoholic hepatitis. In patients with alcoholic hepatitis, cytolysin-positive Enterococcus faecalis strains are correlated with liver disease severity and increased mortality, and in mouse models these strains can be specifically targeted by bacteriophages.
422 citations
"Potential Applications of Human Vir..." refers background in this paper
...This was demonstrated in a recent study where bacteriophages infecting cytolysin-producing Enterococcus faecalis strains ameliorated liver disease severity in patients with alcoholic hepatitis (13)....
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TL;DR: VizBin can be applied de novo for the visualization and subsequent binning of metagenomic datasets from single samples, and it can be used for the post hoc inspection and refinement of automatically generated bins.
Abstract: Background: Metagenomics is limited in its ability to link distinct microbial populations to genetic potential due to a current lack of representative isolate genome sequences. Reference-independent approaches, which exploit for example inherent genomic signatures for the clustering of metagenomic fragments (binning), offer the prospect to resolve and reconstruct population-level genomic complements without the need for prior knowledge. Results: We present VizBin, a Java-based application which offers efficient and intuitive reference-independent visualization of metagenomic datasets from single samples for subsequent human-in-the-loop inspection and binning. The method is based on nonlinear dimension reduction of genomic signatures and exploits the superior pattern recognition capabilities of the human eye-brain system for cluster identification and delineation. We demonstrate the general applicability of VizBin for the analysis of metagenomic sequence data by presenting results from two cellulolytic microbial communities and one human-borne microbial consortium. The superior performance of our application compared to other analogous metagenomic visualization and binning methods is also presented. Conclusions: VizBin can be applied de novo for the visualization and subsequent binning of metagenomic datasets from single samples, and it can be used for the post hoc inspection and refinement of automatically generated bins. Due to its computational efficiency, it can be run on common desktop machines and enables the analysis of complex metagenomic datasets in a matter of minutes. The software implementation is available at https://claczny.github.io/VizBin under the BSD License (four-clause) and runs under Microsoft Windows, Apple Mac OS X (10.7 to 10.10), and Linux.
408 citations
TL;DR: In this short review, key information from the literature pertaining to each processing step is described, and consequently, general recommendations for future 16S rRNA gene metabarcoding experiments are made.
Abstract: The development and continuous improvement of high-throughput sequencing platforms have stimulated interest in the study of complex microbial communities Currently, the most popular sequencing approach to study microbial community composition and dynamics is targeted 16S rRNA gene metabarcoding To prepare samples for sequencing, there are a variety of processing steps, each with the potential to introduce bias at the data analysis stage In this short review, key information from the literature pertaining to each processing step is described, and consequently, general recommendations for future 16S rRNA gene metabarcoding experiments are made
382 citations
"Potential Applications of Human Vir..." refers methods in this paper
...Most reference materials available are intended for microbiome analysis targeting the bacterial fraction, and these have been shown to be helpful in benchmarking nucleic acid extraction methods (54), sequencing platforms and sequencing kits (55), assemblers, and taxonomic and functional classification tools (56, 57)....
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