Exploring the Potential of Next-Generation Sequencing in Detection of Respiratory Viruses

Abstract

dEfficient detection of human respiratory viral pathogens is crucial in the management of patients with acute respiratory tract infection. Sequence-independent amplification of nucleic acids combined with next-generation sequencing technology and bioinformatics analyses is a promising strategy for identifying pathogens in clinical and public health settings. It allows the characterization of hundreds of different known pathogens simultaneously and of novel pathogens that elude conventional testing. However, major hurdles for its routine use exist, including cost, turnaround time, and especially sensitivity of the assay, as the detection limit is dependent on viral load, host genetic material, and sequencing depth. To obtain insights into these aspects, we analyzed nasopharyngeal aspirates from a cohort of 81 Thai children with respiratory disease for the presence of respiratory viruses using a sequence-independent next-generation sequencing approach and routinely used diagnostic real- Laboratories nowadays largely perform viral species-specific assays for virus diagnosis in clinical samples to increase the sensitivity of detection and reduce the time needed for diagnosis. However, an etiological agent cannot be identified in many cases despite the use of a wide range of sensitive diagnostic assays (1-5). This can depend on the timing of sampling, performance of the individual assays, and also the involvement of divergent viruses that are not detected due to the high specificity of the assays. New perspectives for research and diagnostic applications of virus detection have opened up with recent advances in sequence-independent amplification techniques combined with next-generation sequencing platforms. These technologies are well known for their enormous output of sequence data at a relatively high but decreasing cost.Sequence-independent next-generation sequencing approaches have been applied successfully to various fields in virology, including virus discovery, whole-virus genome reconstruction, and minority variant analyses (6-10). Sequence-independent amplification of nucleic acids combined with next-generation sequencing technology and bioinformatic analyses is a promising strategy for the rapid identification of pathogens in clinical and public health settings. It allows the characterization of numerous known pathogens simultaneously and of novel pathogens that elude conventional testing. The general idea, however, is that it is unlikely that genomics-based tools will soon be used in a clinical diagnostic setting (11). Its major hurdles are cost-effectiveness; high-throughput formats for clinical settings; turnaround time; the requirement for investments in bioinformatics tools, databases, and data management; training of personnel; and the reporting and interpretation of guidelines upon the identification of viruses of which the clinical relevance is not clear (12). In addition, issues regarding patient privacy need to be resolved before the transition of genomics-based tools from a research setting to t...

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