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Review
. 2012 Feb;2(1):63-77.
doi: 10.1016/j.coviro.2011.12.004. Epub 2012 Jan 20.

Metagenomics and future perspectives in virus discovery

Affiliations
Review

Metagenomics and future perspectives in virus discovery

John L Mokili et al. Curr Opin Virol. 2012 Feb.

Abstract

Monitoring the emergence and re-emergence of viral diseases with the goal of containing the spread of viral agents requires both adequate preparedness and quick response. Identifying the causative agent of a new epidemic is one of the most important steps for effective response to disease outbreaks. Traditionally, virus discovery required propagation of the virus in cell culture, a proven technique responsible for the identification of the vast majority of viruses known to date. However, many viruses cannot be easily propagated in cell culture, thus limiting our knowledge of viruses. Viral metagenomic analyses of environmental samples suggest that the field of virology has explored less than 1% of the extant viral diversity. In the last decade, the culture-independent and sequence-independent metagenomic approach has permitted the discovery of many viruses in a wide range of samples. Phylogenetically, some of these viruses are distantly related to previously discovered viruses. In addition, 60-99% of the sequences generated in different viral metagenomic studies are not homologous to known viruses. In this review, we discuss the advances in the area of viral metagenomics during the last decade and their relevance to virus discovery, clinical microbiology and public health. We discuss the potential of metagenomics for characterization of the normal viral population in a healthy community and identification of viruses that could pose a threat to humans through zoonosis. In addition, we propose a new model of the Koch's postulates named the 'Metagenomic Koch's Postulates'. Unlike the original Koch's postulates and the Molecular Koch's postulates as formulated by Falkow, the metagenomic Koch's postulates focus on the identification of metagenomic traits in disease cases. The metagenomic traits that can be traced after healthy individuals have been exposed to the source of the suspected pathogen.

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Figures

Figure 1
Figure 1
Overview of viral metagenomic studies between 2002 and 2011. (a) The number of published papers on metagenomics (2003–2010), as determined by Pubmed searches using the keywords ‘metagenome OR metagenomics’. (b) Timeline of important landmarks and virus discoveries achieved with the metagenomic approach between 2002 and 2011. The following studies were used to generate the figure: [17••, 18, 20•, 62, 66, 70, 71, 74, 84, 85, 86, 88, 104, 105, 106, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 117, 118, 119, 120, 121, 122, 123, 148, 161, 162]. M: Main characterization method used: 454NGS, 454 high-throughput sequencing using GS FLX or GS titanium platform; sg-Sanger, shotgun library with Sanger sequencing method. S: sample, Symbols used for sample type: ip, insect pool; sb, skunk brain; int: intestine; panc: pancreas; hf: human feces; se, sewer effluent; ms: marine sediment; nasopharyngeal aspirates (NPA).
Figure 2
Figure 2
Flow chart for the generation of a viral metagenome using high-throughput sequencing.
Figure 3
Figure 3
The unknowns: sequences with no detectable homologs in Genbank. (a) Proportion of the unknowns reported in viral metagenomic studies of diverse environments. (b) Diagram illustrating the abundance of unknown and known sequences in the environment. The distinction between known and unknown depends on the thresholds used.
Figure 4
Figure 4
Metagenomic Koch's postulates. Comparison between a diseased and healthy control animal shows a significant difference between the metagenomic libraries (depicted by the histograms of relative abundance reads). In order to fulfill the metagenomic Koch's postulates: (1) The metagenomic traits in diseased subject must be significantly different from healthy subject. For example traits A, D, E and J found in the disease animal that are not present in the healthy control; (2) Inoculation of samples from the disease animal into the healthy control must lead to the induction of the disease state. Comparison of the metagenomes before and after inoculation should suggest the acquisition or increase of new metagenomic traits (A, E and P). New traits can be purified by methods such as serial dilution or time-point sampling of specimens from a disease animal. (3) Inoculation of the suspected purified traits into a healthy animal will induce disease if the traits form the etiology of the disease.
Figure 5
Figure 5
Monitoring of emerging infectious diseases using a metagenomic approach. A hypothetical example of the potential use of the Public Health Viral Metagenomics Surveillance (PHVMS) approach for virus discovery based on comparison of viromes sampled before (I) and during (II) an epidemic. Depicted here are the rank abundance curves for viral species (a–h), where g represents a newly introduced, highly pathogenic species and h a less virulent virus.

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