Coronavirus testing
The virus that broke the planet
Alpina Non-fiction Publishing House presents Irina Yakutenko's book "The Virus that Broke the Planet. Why is SARS-CoV-2 so special and what should we do with it."
The coronavirus appeared as an unexpected gift for the new year 2020, and in a few months the world turned into a disaster series. The incredible efforts of the state stopped the spread of the virus, but in the fall the epidemic began to gain momentum again. What we know about SARS-CoV-2, why it kills some and passes asymptomatically in others, whether the vaccine is safe and when a cure will be found, how we treat COVID-19 without it, whether it is possible to fight the pathogen without closing the planet – the book answers these and many other questions. Although the pandemic is not over yet and we are constantly receiving new data about the virus, the fundamentals outlined in the text will not change: they serve as a framework on which the reader can string new knowledge.
We suggest reading a fragment of the chapter devoted to testing for coronavirus infection.
How to perceive RNA tests
RNA is a very capricious molecule and tends to fall apart at any opportunity. If the laboratory assistant who takes a smear, conducts a test or takes samples to the laboratory slightly violates the protocol, by the time he adds reagents to the test tube for RT-PCR, only fragments of viral genomes will remain there. In addition, not everyone can correctly take a smear from the nasopharynx – for this reason, home RNA tests that require this procedure seem to be a dubious idea. In one of the papers, the authors identified the virus in different types of samples, and a pharyngeal smear gave a truly positive result only in 32% of cases. The most accurate – 93% – was the analysis of fluid obtained from the lungs (bronchoalveolar lavage), but in order to get it, it is necessary to carry out an extremely unpleasant procedure under local anesthesia. For mass use, this method is obviously not suitable.
Finally, the time of sampling is critically important: we have repeatedly mentioned that with a prolonged course of the disease, the virus descends into the lungs, and a swab from the pharynx in this case often shows a negative response. As well as immediately after infection. PCR diagnostics, in principle, gives results with at least a decent degree of reliability only in the days around the appearance of symptoms. In the first days after infection, while the pathogen has not infected very many cells yet, it is easy to miss it. As the number of infected cells increases, the concentration of the virus begins to increase dramatically – it grows exponentially, which is well known to us.
In the works we talked about earlier, it was shown that a person is as contagious as possible the day before the onset of symptoms, that is, the concentration of viral particles in the upper respiratory tract is maximum these days and it is very difficult not to notice SARS-CoV-2 in the peak days of its reproduction in the pharynx.
Plus or minus 13 thousand
The imperfection of RNA tests caused another coronavirus scandal in February 2020. Then the epidemic developed only in China, and in one day (February 13) the number of registered patients with COVID-19 suddenly increased by 13 thousand. Citizens inclined to conspiracy thinking immediately declared that this was the truth, and before that "the authorities were hiding". In fact, on this day, the Chinese officially changed the method of accounting for the sick and began to record all those who have a characteristic clinical picture as infected. Prior to this, the result of RNA testing was required to confirm the diagnosis, and it not only took many days, but also often turned out to be false negative.
The situation with PCR tests described above looks rather bleak: low accuracy, a lot of false negative results, infinitely long logistics, lack of tests for mass inspections, high price (RT-PCR is expensive, since it requires not only devices and reagents, but also trained personnel). It seems that genetic testing is in principle unsuitable for mass screening and rapid identification of potential carriers, especially in situations where not two dozen, but 10,000 new carriers are detected in a day in the city, as it was, for example, at the peak in New York. But in order for schools, kindergartens, universities, offices and factories to be able to return to full-fledged work, testing must be mass.
Theoretically, it is possible to bring the PCR testing procedure to mind, hire new laboratory assistants, retrain the old ones properly and assign them to most large institutions, having previously purchased equipment there, reorganize sampling and analysis systems to reduce the time of test processing, and so on. But such a strategy radically increases the financial burden on the states, and they are not in the best economic situation right now.
Paradoxically, the path to the solution can run exactly in the opposite direction. There is no need to chase ultra-precise tests that are extremely sensitive to methodological flaws and give the result after a few days due to complex logistics. Such tests are good if our goal is to give each test subject a reliable answer whether he is a carrier of the virus right now. But if our task is to prevent new outbreaks, such individual accuracy is not needed. It is enough to identify most of the carriers at the stage when they are actively spreading the virus and can infect others. For this purpose, tests that "see" RNA only when there is a lot of it – because there is a lot of it just at the stage of active spread of the virus - will also work perfectly. Much more important is the accuracy of the speed with which the test gives the answer. If we use blind, but fast tests often enough, say, once every few days, we will still identify most of the most contagious carriers. And by tracking and isolating their contacts, we will stop a potential outbreak.
Rapid tests that are not based on real-time RT–PCR exist, but are not widely used - not least because the supervisory authorities have claims to their sensitivity. But in this case, such vigilance is rather harmful: in screening tests and tests for individual diagnostics, different qualities are important, and trying to use one instead of the other is at least unjustified, and at most harmful.
On one, two, three
In movies about medicine, doctors usually conduct tests like this: they take samples from a patient, drip a mysterious liquid into a test tube with them – and immediately see the result. In the most realistic films, a test tube can be placed in some device. Real coronavirus testing is not so cinematic: this is a long and tedious process that takes place in several stages and requires complex devices and qualified laboratory technicians. More precisely, this is what the gold standard looks like – real-time RT-PCR. But there are cinematic tests for SARS-CoV-2, too. For example, Abbott has developed a test system based on the so-called loop isothermal amplification reaction (LAMP – loopmediated isothermal amplification) with reverse transcription. It allows you to carry out all the necessary steps in the same test tube where the samples were placed. Real-time RT-PCR requires the transfer of solutions from one test tube to another, and this, firstly, is a risk factor for an incorrect result, and secondly, such protocols can only be performed by a trained specialist. In short, the meaning of the reaction is that, thanks to the clever selection of primers in the first cycles of the reaction, additional edge pieces are attached to the target "multiplied" DNA fragment, which are folded into loops. DNA polymerase sits on these loops and copies the fragment contained between them, and since the copy also contains self-rotating terminal superstructures, the process goes on as long as there are nucleotides in the test tube for the synthesis of new chains. Quite quickly, a large number of multiplied pieces accumulate in the test tube, which can be seen, for example, with the help of a special dye attached to DNA strands.
Another option for a quick test – for coronavirus antigens – is most similar to a pregnancy test. It is a strip on which two tracks are applied – from antibodies to coronavirus proteins and antibodies to these antibodies. In addition, antibodies to these proteins are also applied to the edge of the strip itself, but they recognize another fragment of the antigen (protein). Scrape or saliva diluted in conventional saline solution is dripped onto the strip and wait until capillary forces pull the liquid to the tracks. If there are enough viral particles in the sample, the antibodies on the edge of the strip will cling to the target protein and "arrive" to the first track. The antibodies contained in it will also recognize the viral antigen and grab it. In this case, the two types of antibodies will be physically close to each other. Special molecules are fixed on the "first" and "second" antibodies, which, when combined with each other, will give a color signal. The second track contains antibodies to the first antibodies and a "half" of the dye substance. If there are no viral particles in the sample, the first antibodies will only stain the second track. If there is a virus, then both stripes will be colored. (This is just one of the existing principles of test strips, others are possible).
Both types of rapid test systems allow you to get an answer whether there is viral RNA or proteins in a sample in minutes. However, such tests have a high percentage of errors due to insufficiently high sensitivity - accordingly, only a few of them are approved for mass use. But, as we discuss in the main text, with the competent application of fast tests, hyper-precision is not required.
As for individual decisions, the most responsible behavior in conditions where tests are highly likely to give an incorrect result will be to consider a negative answer as possibly erroneous, and a positive one as true. And if you have a "plus" – you do not need to go to retake the test and go to visit your grandmother, you need to stay at home, and in two or three weeks pass an antibody test (to be sure). If you do the test "just out of interest", in case of a negative answer, continue to take precautions (wear a mask, keep a distance). If you took care of the test because you were in close contact with someone who later got sick, it's better to stay at home just in case, and if this is not possible, then as much as possible observe all the precautions required for two weeks. In other words, if there are suspicions that you may be a carrier of the virus, it is better to be safe.
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