The impact of viral transport media on PCR assay results for the detection of nucleic acid from SARS-CoV-2 and other viruses

Abstract

During the 2020 SARS-Cov-2 pandemic, there has been an acute shortage of viral transport medium. Many different products have been used to meet the demands of large-scale diagnostic and surveillance testing. The stability of SARS-Cov-2 RNA was assessed in several commercially produced transport media and an in-house solution. Coronavirus RNA was rapidly destroyed in the commercial transport media though the deleterious effects on intact virus were limited. Similar results were obtained for a Type A influenza virus. There was reduced detection of both virus and nucleic acid when a herpesvirus sample and purified DNA were tested. Collectively these data showed that the commercial viral transport media contained nucleases or similar substances and may seriously compromise diagnostic and epidemiological investigations.Recommendations to include foetal bovine serum as a source of protein to enhance the stabilising properties of viral transport media are contraindicated. Almost all commercial batches of foetal bovine serum contain pestiviruses and at times other bovine viruses. In addition to the potential for there to be nucleases in the transport medium, the presence of these viruses and other extraneous nucleic acid in samples may compromise the interpretation of sequence data. The inclusion of foetal bovine serum presents a biosecurity risk for the movement of animal pathogens and renders these transport media unsuitable for animal disease diagnostic applications. While these transport media may be suitable for virus culture purposes, there could be misleading results if used for nucleic acid-based tests. Therefore, these products should be evaluated to ensure fitness for purpose.

Saliva Sampling and Its Direct Lysis, an Excellent Option To Increase the Number of SARS-CoV-2 Diagnostic Tests in Settings with Supply Shortages

“…However, we have found a great variability in the results obtained using the QE lysis protocol when applied to swab samples, most likely due to variations in the material of the swabs used and to variations in the preparation of the viral transport medium employed (data not shown). In this regard, it has recently been reported that the composition of viral transport media can affect the detection of RNA from SARS-CoV-2 and other viruses (16), and due to the scarcity of these media, several laboratories have started to prepare their own transport media, introducing an additional confusion factor. A similar situation occurs with swabs, since in view of the scarcity of suitable materials, other materials are being employed, despite the fact that some of them are known to inhibit RT-PCR (17).…”

The impact of viral transport media on PCR assay results for the detection of nucleic acid from SARS-CoV-2

“…The design of this study and the methods employed are described in full elsewhere. 10 A selection of samples was also tested using the CDC designed assay (2019-nCoV_N2) targeting the N gene. 11 This investigation showed that there was a profound impact on the detection of SARS-CoV-2 RNA after dilution in the commercial VTM products (VTM-2 to VTM-4).…”

“…The design of this study and the methods employed are described in full elsewhere. 10 A selection of samples was also tested using the CDC designed assay (2019-nCoV_N2) targeting the N gene. 11…”

Saliva sampling is an excellent option to increase the number of SARS CoV2 diagnostic tests in settings with supply shortages

“…However, in our experience we have found a great variability in the results obtained using the QE lysis protocol when applied to swab samples, most likely due to variations in the material of the swabs used and to variations in the preparation of the viral transport medium employed (data not shown). In this regard, it has recently been reported that the composition of viral transport media can affect the detection of viral RNA from SARS-CoV-2 and other viruses [17] and, due to the scarcity of it, several laboratories have started to prepare their own transport media introducing an additional confusion factor. A similar situation occurs with the swabs, since in view of the scarce suitable materials, other materials are being employed, despite the fact that some of them are known to inhibit the RT-PCR reactions [18].…”