There has been a lot of talk about tests for COVID-19 (or the virus that causes it, SARS-CoV-2). The purpose of this piece is to try to clarify the nature of the tests that are usually referred to, and some other possible tests.
In general, most diagnostic tests for an infectious disease fall under the following headings:
1.Tests for the presence of the whole (‘live’) pathogen
2. Tests for the presence of some part of the pathogen. These can be divided into
a) tests for the nucleic acid of the pathogen (RNA in this case).
b) tests for an antigen of the pathogen
3. Tests for antibodies produced by the body in response to the presence of the pathogen.
1. Live pathogen tests.
For most bacterial diseases, this is the ‘gold standard’. Bacterial pathogens can (mostly) be grown easily in the lab, so this is a good way of detecting them.
It is not so easy for viruses. They don’t grow on their own – they have to get into a cell and use that cell to make more copies of the virus. (This makes it very debatable as to whether we consider a virus as ‘alive’; ‘viable’ is a better term). So you need to start with a culture of a human or other mammalian cell line that you can try to infect with the virus. There is a wide variety of such cell lines in common use in virology laboratories.
Viruses are often fussy – they will grow on some cell lines but not on others. Also, you want to be able to see the effect the virus has on the cells – a ‘cytopathic effect’. Fortunately, previous work with the related virus that caused SARS indicated which cell lines were most likely to be successful. So, quite soon after the first samples became available, several laboratories around the world had successfully cultivated SARS-CoV-2.
But this is not suitable as a diagnostic test. It is too slow (it takes several days or more) and needs high levels of containment. However, being able to grow the virus is important for several reasons, including:
– it is the only way of confirming that the virus is viable (and hence presumably infectious), which is necessary for investigating whether someone is shedding infectious virus, or for testing survival in the environment, for example.
– for lab-based testing of the effects of potential antiviral agents
– providing reasonably large amounts of the virus, especially for the development of some types of vaccines (especially those based on inactivated virus).
2. Tests for the presence of part of the virus.
This is the mainstay of current tests, using a technique known as RT-PCR (Reverse transcriptase-polymerase chain reaction). The RT part copies the viral RNA into DNA, and the PCR amplifies the DNA. It works through a series of cycles, each cycle doubling the amount of DNA. So after 10 cycles, you have 1,000 copies, after 20 cycles a million (106) copies, and after 30 cycles 109 copies. This can all be automated, so you put it into the machine which then tells you when you have a detectable product.
A key element in this process is that it requires the binding of specific ‘primers’ – short lengths of nucleic acid derived from the sequence of the DNA copy of the viral RNA. If the right DNA is present, the primers will bind and you get amplification. If it is not there, there’s no binding, and no amplification. So it is not only exquisitely sensitive, it is also highly specific.
You need to know the sequence of the viral RNA before you can do this. This sequence was made available very shortly after the virus was first identified.
It is important to remember that this test only detects the presence of viral RNA, not the whole virus. So a positive test does not necessarily mean that you are still infectious; it depends on how long the RNA remains after the virus has been inactivated by your immune response.
Some pathogens are detected or identified through the presence of other parts of their structure, mainly those that invoke an immune response – i.e., antigens. I have seen references to an ‘antigen test’ for this virus, but these are probably erroneous references to the RT-PCR test (to contrast it to the antibody tests, see below). There seems no point in an antigen test, which would be slower and less sensitive the RT-PCR.
3. Antibody tests.
These are highly controversial.
The principle is that during infection you produce antibodies (immunoglobulins) that are capable of reacting with a viral antigen (such as the proteins that form the ‘spikes’ on the surface of the virus). So you take a blood sample, mix it with a sample of the specific antigen, and the test detects the binding of the antibody to the antigen. So far, so good.
The problems are of two sorts. Firstly, sensitivity. During the early stages of infection, there may be only low levels of antibody. So you may not detect infection at that stage. Later on, after you have recovered, the antibody levels may drop off quite quickly – so it may not be a good test for immunity (see below).
Secondly, specificity. SARS-CoV-2 is closely related to the SARS virus, and (less closely) to a variety of other coronaviruses that cause a proportion of common colds. While it is easy to make the RT-PCR test specific (by choosing suitable primers), it is not so easy to get a highly specific antibody test.
These problems may be overcome in a lab setting, with experienced technicians, and larger samples of blood – and some laboratories are already conducting surveys using antibody tests. But we are not yet at the stage where you could reliably use a ‘pin-prick’ test in your own home.
This leads on to a related point. If a reliable antibody test says you have antibodies to the virus, what does that mean in practical terms? Especially, will you be resistant to further infection? I don’t want to get bogged down in the direct evidence, which is inconclusive – there are anecdotal reports of a second infection in individuals who have converted to RT-PCR negative (but there are other explanations for that), and on the other hand some animal experiments indicate that re-infection does not happen. The question I want to address is whether the detection of antibodies (assuming a reliable test) means you are now resistant to infection. A full answer, based on general concepts with other viruses, would be far too long and complex to be covered here -and I’m not an immunologist, so I’m not equipped to tackle it, but there are a few points worth making.
– With some viruses, you get ‘neutralising antibodies’ – i.e., binding of the antibodies to the virus is by itself enough to prevent the virus infecting cells (this can be detected in the lab). If that does not happen, the body has other ways of disposing of the virus-antibody complexes, including the activity of various cells such as macrophages and T cells. So in that case your resistance to the disease may depend on this cell-mediated immunity rather than, or as well as, the production of antibodies.
– As described above, antibody levels may fall off considerably after you recover from infection. But it’s worth noting that you can be effectively immune even if antibody levels are low – your immune system may ‘remember’ the previous exposure and produce antibodies very quickly in response to further exposure.
– Antibodies can be damaging as well as protective. The classic example is dengue, where a second infection, with a different strain, causes a much more serious disease – a phenomenon known as antibody-dependent enhancement. Although it is thought unlikely that this will happen with SARS-CoV-2, the prospect is enough to make the vaccine developers twitchy.
24 April 2020