Part 1: Overview

Excerpts from a report written by Dr Kerry Mills, Chair of the International Virolens Research Consortium

Since the emergence of the SARS-CoV-2 delta strain and widespread vaccination, the landscape of viral detection has changed immensely. Firstly, the delta strain has a rapid onset of infection, with peak viremia (and therefore infectivity) before symptoms appear. Secondly, vaccination prevents severe disease in most people, but does not preclude infections in vaccinated people. Indeed, these people can, even when free of symptoms, pass the virus on to others.

Use-case for Virolens

The original and ongoing use for Virolens is as a rapid mass-screening device. It is not intended for diagnosis of COVID-19 disease, nor is it appropriate for epidemiological work. As such, the ideal candidate for a Virolens test is an asymptomatic person entering a place of work, an airport, a hospital, an arena or other place where comingling of people is necessary.

What does Virolens detect?

Virolens detects whole virions. It does not detect RNA, whether this is inside virions or shed by a previously infected person. It does not detect individual SARS-CoV-2 proteins, whether these are part of virions or the result of the lysis of infected cells. As such, the results of Virolens cannot be directly compared with RT-PCR or rapid antigen testing, as each of these techniques is measuring something different. Virolens is the only test that can screen for people with whole virions in their nasopharyngeal tract (Figure 1).

*Figure 1: comparison of Virolens with other testing modalities. Virolens is the only test that will not produce false positives from cell lysis, free viral RNA, or free viral proteins.*

What does RT-PCR detect?

RT-PCR detects viral RNA. Swabs taken from potentially infected people are put into a liquid medium and transported to the laboratory, where they are processed to remove all the proteins, fats, free nucleotides and carbohydrates in the sample, leaving only the RNA. This RNA is then incubated with a number of reagents to transcribe the RNA into DNA (reverse transcription - RT), and is then used in a traditional quantitative PCR to amplify specific regions of the SARS-CoV-2 genome. As seen in Figure 1, RT-PCR is not specific for infection, as it detects SARS-CoV-2 RNA in virions, in cell lysis debris, and as free RNA.

What does rapid antigen testing detect?

Rapid antigen testing (RAT) detects the presence of a single type of protein that is unique to the SARS-CoV-2 virion. This is usually the spike protein, as it is the most abundant protein on the surface of the virion. However, as seen in Figure 1, the test cannot differentiate between an active infection, lysed cell contents, or degraded, non-infectious virions. Furthermore, the limit of detection of RATs is quite high - around 10,000 to 100,000 times less sensitive than PCR (Table 1).

*Table 1: comparison of the targets, LODs, periods of detection and limitations of Virolens and current screening technologies*

Time course of testing capabilities

As described above, the different tests detect different inputs. An additional complexity in the situation is also the window of detection. In Figure 2, correlation of the infection course and the windows of detection of various testing modalities is given.

*Figure 2: comparison of SARS-CoV-2 detection across different technologies. Note that Virolens correlates tightly with the true infectious period of the virus.*

From this figure, it is clear that PCR detection starts at the onset of symptoms and continues long after viral clearance. In fact, several studies have shown that many people test positive by RT-PCR, even when cell culture has shown that they are not infectious.3,4 Antigen testing is more tightly correlated with infection, but is far less sensitive than either RT-PCR or Virolens (Table1). The most important thing to note in this figure is that the correlation between presence of infectious virions and illness hardly overlaps, especially in severe and critical illness. Therefore a person who is hospitalised with COVID-19 is very unlikely to be infectious to others, but may well test positive by PCR.