Abstract
Metagenomic next-generation sequencing has transformed the discovery and diagnosis of infectious disease, with the power to characterise the complete ‘infectome’ (bacteria, viruses, fungi, parasites) of an individual host organism. However, the identification of novel pathogens has been complicated by widespread microbial contamination in commonly used laboratory reagents. Using total RNA sequencing (“metatranscriptomics”) we documented the presence of contaminant viral sequences in multiple ‘blank’ negative control sequencing libraries that comprise a sterile water and reagent mix. Accordingly, we identified 14 viral sequences in 7 negative control sequencing libraries. As in previous studies, several circular replication-associated protein encoding (CRESS) DNA virus-like sequences were recovered in the blank control libraries, as well as contaminating sequences from the Totiviridae, Tombusviridae and Lentiviridae families of RNA virus. These data suggest that viral contamination of common laboratory reagents is likely commonplace and can comprise a wide variety of viruses.
Highlights
Culture-independent methods, metagenomic next-generation sequencing, have revolutionised pathogen discovery, streamlined pathways of clinical diagnosis, and have enhanced our ability to track infectious disease outbreaks [1], including the current COVID-19 pandemic [2,3]
The data generated by metagenomic next-generation sequencing (mNGS) are bountiful and cost-effective, they come with several inherent limitations, central of which is the possibility of reagent contamination [9]
There have been many new members of Circoviridae characterised in recent years, cycloviruses [5,43,44], we suggest that current and future characterisations of novel circovirusand circular replication-associated protein encoding (CRESS)-like genomes should be completed cautiously with additional confirmation steps
Summary
Culture-independent methods, metagenomic next-generation sequencing (mNGS), have revolutionised pathogen discovery, streamlined pathways of clinical diagnosis, and have enhanced our ability to track infectious disease outbreaks [1], including the current COVID-19 pandemic [2,3]. These methods can reveal the complete profile of pathogenic and commensal microorganisms within a host, comprising viruses, bacteria, fungi and eukaryotic parasites. The experimental preparation of samples for sequencing necessarily involves treatment with a variety of reagents, many of which have been shown to carry contaminating nucleic acids, including viral sequences [10,11,12,13,14,15]. Metagenomic analysis of human plasma samples revealed the presence of sequences of Kadipiro virus, a double-stranded positive-sense
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
