Unbiased Parallel Detection of Viral Pathogens in Clinical Samples by Use of a Metagenomic Approach

Abstract

Viral infectious diseases represent a major threat to public health and are among the greatest disease burdens worldwide. Rapid and accurate identification of viral agents is crucial for both outbreak control and estimating regional disease burdens. Recently developed metagenomic methods have proven to be powerful tools for simultaneous pathogen detection. Here, we performed a systematic study of the capability of the short-read-based metagenomic approach in the molecular detection of viral pathogens in nasopharyngeal aspirate samples from patients with acute lower respiratory tract infections (n ‫؍‬ 16). Using the highthroughput capacity of ultradeep sequencing and a dedicated data interpretation method, we successfully identified seven species of known respiratory viral agents from 15 samples, a result that was consistent with results of conventional PCR assays. We also detected a coinfected case that was missed by regular PCR testing. Using the metagenomic data, 11 draft genomes of the abundantly detected viruses in the samples were reconstructed with 21.84% to 98.53% coverage. Our results show the power of the short-read-based metagenomic approach for accurate and parallel screening of viral pathogens. Although there are some inherent difficulties in applying this approach to clinical samples, including a lack of controls, limited specimen quantity, and high contamination rate, our work will facilitate further application of this unprecedented high-throughput method to clinical samples.Despite great advances in modern medicine, infectious diseases are among the most common causes of human death and represent the greatest disease burdens worldwide, as estimated by the WHO (13). As we have experienced in the past decade, viral pathogens, especially respiratory viruses, pose significant threats to public health. Indeed, both the severe acute respiratory syndrome (SARS) outbreak and the H1N1 influenza A virus pandemic had serious impacts on public health as well as the global economy. Undoubtedly, the rapid identification of newly emerging pathogens using advanced molecular techniques has played a crucial role in controlling such infectious diseases worldwide. Moreover, accurate determination of regional agent-specific infectious diseases provides critical information for local authorities to optimize public health prevention strategies and maximize the limited available resources, particularly in developing countries.Molecular assays are becoming increasingly popular for laboratory diagnosis because they outperform traditional viral diagnostic methods, such as cell-culture-based testing and antigen detection, in efficiency, sensitivity, and specificity. In addition, several multiplex PCR assays and DNA microarray testing methods have been proposed to screen dozens to hundreds of pathogens simultaneously (10,11,23). Metagenomics is a powerful tool for viral and microbial community characterization and new pathogen identification because genetic materials are isolated directly from environmental/clinical samples ...

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