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For the diagnosis of SARS-CoV-2 infection they are commonly used two types of viral tests: nucleic acid amplification techniques (NAAT) and antigen detection test. Both allow the detection of the virus in biological samples.
The proof PCR amplifies DNA fragments of any origin: viral, bacterial, etc. to levels that are detectable. When the reverse transcription of RNA into DNA is carried out prior to PCR, it is known as RT-PCR (Reverse Transcription-Polymerase Chain Reaction).
RT-PCR is the most widely used NAAT test to determine if some SARS-CoV-2 virus genes are present in a clinical sample. The previous phase of DNA synthesis from RNA is necessary, since the genome of this virus is RNA.
Commonly, since the onset of the pandemic, RT-PCR has been referred to as PCR.
Nasopharyngeal, nasal, or oropharyngeal swab RT-PCR is the gold standard test for the molecular diagnosis of SARS-CoV-2 infection.
Too it can be done in saliva, but in this sample the viral load is lower, so the test may have less sensitivity. However, some studies suggest that saliva tests might perform better in cases of omicron infection. This test can also detect the virus in other types of samples such as those from the lower respiratory tract, blood, urine, and stool.
How does each screening test work?
TAAN tests detect genetic sequences of two or more of the structural proteins, S (spicule), E (envelope), and N (nucleocapsid) protein, and other non-structural proteins of SARS-CoV-2.
For their part, antigen tests are immunoassays that are designed, for the most part, to detect the presence of protein N. To achieve this, these tests carry antibodies that are developed in animals such as mice, rabbits, and even humans. These identify the protein if it is present and when interacting with it give a positive reaction. In antigen tests, the sample must be previously treated with a solution that releases the N protein from the rest of the viral structure, so that it can react with the antibodies that are present in the test.
Most of the antigen tests authorized in Europe detect viral antigen in nasal, nasopharyngeal and oropharyngeal swab samples, and some in saliva.
Are there false positives in antigen tests?
We speak of a false positive when the test result indicates the presence of the virus without the person being infected.
The RT-PCR technique is highly specific (99.5%), so the probability of a false positive is very low. It is also very sensitive, so much so that it can detect the virus between 3 and 5 days following acquiring the infection and up to weeks later. Although this viral RNA test does not always indicate current infection, it does not differentiate between active infection and resolved infection.
False positives in the RT-PCR test are usually due to sample contamination. Also the low viral load, especially in the beginning of the infection or during its resolution, can give a doubtful positive result.
Antigen tests, like NAATs, are highly specific. They are comparable to the RT-PCR technique in detecting SARS-CoV-2 infection in the first week following infection. In the nasopharynx, the highest viral load is observed mainly in the first 5 to 7 days and then decreases until its disappearance. After this period of time, RT-PCR has a higher diagnostic sensitivity than the antigen test.
False positives in antigen tests are more frequent in massive screenings in populations with a low prevalence of infection. They can also be due to sample contamination.
Factors that minimize the possibility of false positives in diagnostic tests are a high prevalence of infection in the community, the presence of symptoms suggestive of Covid-19 in the person, and a high specificity of the test.
Confirm the result of an antigen test
The result of the antigen test must be confirmed by NAAT depending on the type of case and the probability of infection. It is considered that there is a high probability of infection if there is contact with a confirmed or suspected positive case and the person is not fully vaccinated or has not suffered from SARS-CoV-2 infection in the last 3 months.
In the community setting, in a person with symptoms suggestive of Covid-19 of 5 days or less of evolution, a professional positive antigen test confirms the infection.
In general, the positive antigen test may need confirmation by RT-PCR in a situation with a low probability of infection, whether or not the person is symptomatic. Also in the event of an outbreak in a social health center to confirm the first cases and in people who are hospitalized. In addition, a positive result of a self-diagnostic antigen test must be confirmed to ensure that there is infection.
However, health authorities may consider the results of self-diagnostic tests to establish isolation and control measures.
In the case of a negative antigen test, if the person is symptomatic, it should be confirmed by RT-PCR, especially if the symptoms began more than 5 days before, due to the decrease in the sensitivity of the antigen test. Also in the case of a negative antigen test in an asymptomatic person if there is a high probability of infection.
Diagnosis of infected people is crucial for the control and reduction of transmission. It usually marks the start of contact tracing.
Despite the high efficacy of RT-PCR as a diagnostic confirmation test, sometimes a negative result may not completely rule out SARS-CoV-2 infection. If infection is suspected, the test results should be evaluated in the clinical-epidemiological context of the patient and in conjunction with other diagnostic tests.
María del Mar Tavío Pérez. Professor of Microbiology, University of Las Palmas de Gran Canaria.
This article was originally published on
The Conversation.
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