Optimising nucleic acid recovery from rapid antigen tests for whole genome sequencing of respiratory viruses

•RNALater has a more pronounced effect on nucleic acid preservation compared to DNA/RNA shield for virus genome recovery from rapid antigen tests.•Viral genomes were generally stable and recoverable after 72 h and 1 weeks when stored at 4 °C, especially in RNALater.•Effective genome recovery could b...

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Bibliographic Details
Published in:Journal of clinical virology 2024-10, Vol.174, p.105714, Article 105714
Main Authors: Butel-Simoes, G, Steinig, E, Savic, I, Zhanduisenov, M, Papadakis, G, Tran, T, Moselen, J, Caly, L, Williamson, DA, Lim, CK
Format: Article
Language:English
Subjects:
Animals
Antigens, Viral - genetics
Community surveillance
Detection
Diagnosis
Genome, Viral
Genomic epidemiology
Genomic Instability
Probe hybridization capture
Rapid antigen tests
Rats
Respiratory Tract Infections - diagnosis
Respiratory Tract Infections - virology
Respiratory viruses
RNA, Viral - genetics
Specimen Handling - methods
Subtyping
Viruses - classification
Viruses - genetics
Viruses - isolation & purification
Whole genome sequencing
Whole Genome Sequencing - methods
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Summary:•RNALater has a more pronounced effect on nucleic acid preservation compared to DNA/RNA shield for virus genome recovery from rapid antigen tests.•Viral genomes were generally stable and recoverable after 72 h and 1 weeks when stored at 4 °C, especially in RNALater.•Effective genome recovery could be achieved with a sequencing depth of 500,000 reads per sample in RNALater. Whole genome sequencing (WGS) of respiratory viruses from rapid antigen tests (RAT-WGS) is a novel approach to expanding genomic surveillance of respiratory infections. To date however, there are limited data on the genomic stability of these viruses on RATs. In this study, we investigated the effect of storage conditions and nucleic acid preservatives on the ability to enhance stability and improve recovery of respiratory virus genomes from RATs. A mixture of common respiratory viruses was used to inoculate RATs at different environmental temperatures (4°C, 20°C and 36°C), with two preservative reagents (RNALater and DNA/RNA shield) Nucleic acid was extracted from RATs at two different timepoints (72 h and seven days) and subject to real-time multiplex respiratory PCR to detect a range of respiratory viruses. WGS was performed using target-enrichment with the TWIST Comprehensive Viral Research Panel. Defined metrics from an automated in-house bioinformatic pipeline were used to assess and compare viral genome recovery under different conditions. Nucleic acid degradation (indicated by relative change in PCR cycle threshold and WGS-based metrics) was most notable at 20 °C and 36 °C. Storage in either RNALater or DNA / RNA shield improved genome recovery for respiratory viruses across all temperature conditions, although this was most pronounced for RNALater. Subtyping of Influenza viruses demonstrated the applicability of RAT-WGS in downstream genomic epidemiological surveillance. Under simulated conditions, RAT-WGS demonstrated that (i) viral genomes were generally stable at 4°C at 72 h and 1 week, (ii) RNALater has a more significant preservation of nucleic acids compared to DNA/RNA Shield and (iii) genome recovery can be achieved using a sequencing depth of 500,000 reads per sample in RNALater, across all respiratory viruses and conditions.
ISSN:1386-6532
1873-5967
1873-5967
DOI:10.1016/j.jcv.2024.105714