1. Optimal preparation of SARS-CoV-2 viral transport medium for culture
- Author
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Richard A. Strugnell, Nancy Wang, Claire Fraser, Damian F. J. Purcell, Elizabeth Trajcevska, Elena Paraskeva, Julie L. McAuley, Eric Bert, and Michelle Sait
- Subjects
0301 basic medicine ,VTM ,Coronavirus disease 2019 (COVID-19) ,Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) ,Culture ,Short Report ,Biology ,Validation testing ,Replication competent virus ,Cell Line ,Specimen Handling ,lcsh:Infectious and parasitic diseases ,03 medical and health sciences ,COVID-19 Testing ,0302 clinical medicine ,Virology ,Chlorocebus aethiops ,Diagnosis ,Vaccine escape ,Animals ,Humans ,lcsh:RC109-216 ,030212 general & internal medicine ,Vero Cells ,Viral culture ,SARS-CoV-2 ,COVID-19 ,Anti-Bacterial Agents ,Culture Media ,030104 developmental biology ,Infectious Diseases ,Viral replication ,Viral Transport medium ,RNA, Viral ,Transport media - Abstract
Introduction The sudden arrival of the COVID-19 pandemic placed significant stresses on supply chains including viral transport medium (VTM). The VTM that was urgently required needed to support viral replication, as well as other routine diagnostic approaches. We describe the preparation and validation testing of VTM for rapidly expanding diagnostic testing, where the capacity of the VTM to preserve viral integrity, for culture, isolation and full sequence analysis, was maintained. Methods VTM was prepared using different methods of sterilization then ‘spiked’ with virus. The VTM was investigated using viral culture in Vero cells, and for nucleic acid detection by quantitative PCR. Results The best results were obtained by filter and autoclave-based sterilization. The VTM proved robust for culture-based analyses provided the inoculated VTM was stored at 4 °C, and tested within 48 h. The filtered VTM also supported PCR-based diagnosis for at least 5 days when the mock inoculated VTM was held at room temperature. Discussion The manual handling of VTM production, including filling and sterilization, was optimized. SARS-CoV-2 was spiked into VTM to assess different sterilization methods and measure the effects of storage time and temperature upon VTM performance. While most diagnostic protocols will not require replication competent virus, the use of high quality VTM will allow for the next phase of laboratory analysis in the COVID-19 pandemic, including drug and antibody susceptibility analysis of re-isolated SARS-CoV-2, and for the testing of vaccine escape mutants.
- Published
- 2021