For many years various medium solutions have been suggested and used in order to stabilize specimens. Diagnostic investigations are performed using those solutions to detect bacteria and viruses. They are based on balanced salt and saline solutions to maintain an almost neutral pH. Multiple laboratories have prepared an ‘in-house’ viral transport media, while others use commercial preparations that come along with DNA Extraction Kit with sterile swabs. There are cultural methods as well but the studies were originally focused on the capacity of a preparation where the infectivity of the virus was maintained at different temperatures prior to and during the transport and during the time it’s being stored at the laboratory. Minimum attention has been given to the other uses of VTM or the future impact of endogenous components. For example, a commercially available product is specially designed to detect and disable viruses and bacteria, along with inhibiting the nucleus activity that might have been present in the sample. Apart from this, there are many other products that are available for use in molecular detection.

Large Scale Pandemics

There is immense pressure on manufacturers in large-scale disease epidemics to provide viral transport media but at the same time, the pressure can become prohibitive. An acute shortage of VTM can be seen in Australia during the Sars-CoV-2 pandemic since the demand has been both local and international. The main reason for the shortage has been due to the lack of local manufacturers and reduced international flights. However, the demand generated by diagnostic and surveillance testing is being met by different VTMs and other solutions. The variation in results has caused some concern and initiation of a study that compared the stability of SARS-CoV-2 RNA in VTMs manufactured commercially and in-house. The commercial products were UTM-RT (Italy), Citoswab (China), and CP VTF (Australia), and were supplemented with bovine serum albumin and gelatin. While the in-house product was, VTM-1, based on phosphate-buffered saline supplemented with 0.5% gelatin. After the 30-45 minutes of preparations, extraction from each VTM of the RNA dilutions was tested by semi-quantitative real-time PCR. a series of tests were then conducted with dilutions from different samples.

Results Of The Investigation

After the dilution in different commercial VTMs, there was a more profound effect on the detection of SARS-CoV-2 RNA. The commercial product (VTM-2) did not detect any SARS-CoV-2 in the prepared dilution. VTM-3 and VTM-4, also commercial, had very weak reactivity with the samples with the highest concentration of RNA. however, the results of samples diluted in PBS and VTM-1 were almost identical. The variation in results of VTM 2-4 and PBS/VTM-1 shows a decrease in analytical sensitivity.

The experiment showed that some of the examined VTM solutions had significantly affected the purified viral RNA. The fake sample (XIPC RNA) was not immediately affected since it was prepared in tRNA solution and included the sample buffer that included inhibitors of nuclease activity. The tests were held at room temperature for 1 to 48 hours. In the first hour, one could see the adverse effects of each commercial VTM, and after 48 hours, no RNA was detected. Nevertheless, there was no change in the results for XIPC held in PBS or VTM-1.

Furthermore, in order to demonstrate that the adverse effects were not only restricted to SARS-Cov-2 but a different experiment was also designed with RNA for the Type A Influenza Virus. Similar results were found as no viral RNA was detected in commercial VTMs and in-house PBS and VTM.

The study clearly indicates that the commercial VTM solutions had an adverse impact on being able to detect RNA of both SARS-CoV-2 and influenza virus. The RNAs diluted in commercial VTMs show that the components of these VTMs have caused prevention in the detection of RNAs in different samples.

In-Depth Reflections

Upon profoundly reflecting on the results, we can see that the observations are an outcome of contact between viral nucleic acid and VTM for at least one hour every time. The destroyed nucleic acid levels in VTMs were significant as they were being perished with each addition. The adverse impact on the whole virus seemed insignificant, although along with the nucleic acid, perhaps the whole virus was compromised at levels that could hold diagnostic relevance.

Although further tests of patient samples were not possible since each submitted sample had already been exposed to VTM for many hours. During this time, it is possible that harmful effects may have already taken place. A genuine comparison would come from the immersion of different samples in a ‘neutral solution,’ like PBS, for a short period and then divide the sample for each of the VTM under study.

Citation

M.J., Frost, and Kirkland P.D. The impact of viral transport media on PCR assay results for the detection of nucleic acid from SARS-CoV-2. Royal College of Pathologists of Australasia. 2020. Retrieved August 30th, 2021 from https://www.pathologyjournal.rcpa.edu.au/article/S0031-3025(20)30940-5/fulltext#relatedArticles