Diagnostic Methods of COVID-19

 

Positive viral tests indicate a current infection, while positive antibody tests indicate a prior infection. Other techniques include a CT scan, checking for elevated body temperature, checking for low blood oxygen level, and the deployment of detection dogs at airports.

Reverse transcription polymerase chain reaction

Polymerase chain reaction (PCR) is a process that amplifies (replicates) a small, well-defined segment of DNA many hundreds of thousands of times, creating enough of it for analysis. Test samples are treated with certain chemicals that allow DNA to be extracted. Reverse transcription converts RNA into DNA.

Reverse transcription polymerase chain reaction (RT-PCR) first uses reverse transcription to obtain DNA, followed by PCR to amplify that DNA, creating enough to be analyzed. RT-PCR can thereby detect SARS-CoV-2, which contains only RNA. The RT-PCR process generally requires a few hours.

Real-time PCR (qPCR) provides advantages including automation, higher-throughput and more reliable instrumentation. It has become the preferred method.

The combined technique has been described as real-time RT-PCR or quantitative RT-PCR and is sometimes abbreviated qRT-PCR, rRT-PCR or RT-qPCR, although sometimes RT-PCR or PCR are used.

Average sensitivity for rapid molecular tests were 95.2% (ranging from 68% to 100%) and average specificity was 98.9% (ranging from 92% to 100%).

Samples can be obtained by various methods, including a nasopharyngeal swab, sputum (coughed up material), throat swabs, deep airway material collected via suction catheter or saliva. Drosten et al. remarked that for 2003 SARS, "from a diagnostic point of view, it is important to note that nasal and throat swabs seem less suitable for diagnosis, since these materials contain considerably less viral RNA than sputum, and the virus may escape detection if only these materials are tested."

Sensitivity of clinical samples by RT-PCR is 63% for nasal swab, 32% for pharyngeal swab, 48% for feces, 72–75% for sputum, and 93–95% for bronchoalveolar lavage.

The likelihood of detecting the virus depends on collection method and how much time has passed since infection. According to Drosten tests performed with throat swabs are reliable only in the first week. Thereafter the virus may abandon the throat and multiply in the lungs. In the second week, sputum or deep airways collection is preferred.

Antigen

An antigen is the part of a pathogen that elicits an immune response. Antigen tests look for antigen proteins from the viral surface. In the case of a coronavirus, these are usually proteins from the surface spikes. SARS-CoV-2 antigens can be detected before onset of COVID-19 symptoms (as soon as SARS-CoV-2 virus particles) with more rapid test results, but with less sensitivity than PCR tests for the virus.

Antigen tests may be one way to scale up testing to much greater levels. Isothermal nucleic acid amplification tests can process only one sample at a time per machine. RT-PCR tests are accurate but require too much time, energy and trained personnel to run the tests."There will never be the ability on a [PCR] test to do 300 million tests a day or to test everybody before they go to work or to school.

Samples may be collected via nasopharyngeal swab, a swab of the anterior nares, or from saliva. The sample is then exposed to paper strips containing artificial antibodies designed to bind to coronavirus antigens. Antigens bind to the strips and give a visual readout. The process takes less than 30 minutes, can deliver results at point of care, and does not require expensive equipment or extensive training.

Swabs of respiratory viruses often lack enough antigen material to be detectable. This is especially true for asymptomatic patients who have little if any nasal discharge. Viral proteins are not amplified in an antigen test. According to the WHO the sensitivity of similar antigen tests for respiratory diseases like the flu ranges between 34% and 80%.

 

Imaging

Typical visible features on CT initially include bilateral multilobar ground-glass opacities with a peripheral or posterior distribution. COVID-19 can be identified with higher precision using CT than with RT-PCR.

Subpleural dominance, crazy paving, and consolidation may develop as the disease evolves. Chest CT scans and chest x-rays are not recommended for diagnosing COVID-19. Radiologic findings in COVID-19 lack specificity.

Antibody tests

Automated analyzer for immunoassays, used to find SARS-CoV-2 antibodies and quantitative results for SARS-CoV-2 antibody test. 

The body responds to a viral infection by producing antibodies that help neutralize the virus. Blood tests (serology tests) can detect the presence of such antibodies. Antibody tests can be used to assess what fraction of a population has once been infected, which can then be used to calculate the disease's mortality rate.

The most notable antibodies are IgM and IgG. IgM antibodies are generally detectable several days after initial infection, although levels over the course of infection and beyond are not well characterized. IgG antibodies generally become detectable 10–14 days after infection and normally peak around 28 days after infection.This pattern of antibody development seen with other infections, often does not apply to SARS-CoV-2, however, with IgM sometimes occurring after IgG, together with IgG or not occurring at all. Generally, however, median IgM detection occurs 5 days after symptom onset, whereas IgG is detected a median 14 days after symptom onset. IgG levels significantly decline after two or three months.

Genetic tests verify infection earlier than antibody tests. Only 30% of those with a positive genetic test produced a positive antibody test on day 7 of their infection.

 

 

 

Types

Rapid diagnostic test (RDT)

RDTs typically use a small, portable, positive/negative lateral flow assay that can be executed at point of care. RDTs may process blood samples, saliva samples, or nasal swab fluids. RDTs produce colored lines to indicate positive or negative results.

Enzyme-linked immunosorbent assay (ELISA)

ELISAs can be qualitative or quantitative and generally require a lab. These tests usually use whole blood, plasma, or serum samples. A plate is coated with a viral protein, such as a SARS-CoV-2 spike protein. Samples are incubated with the protein, allowing any antibodies to bind to it. The antibody-protein complex can then be detected with another wash of antibodies that produce a color/fluorescent readout.

Neutralization assay

Neutralization assays assess whether sample antibodies prevent viral infection in test cells. These tests sample blood, plasma or serum. The test cultures cells that allow viral reproduction (e.g., VeroE6 cells). By varying antibody concentrations, researchers can visualize and quantify how many test antibodies block virus replication.

Chemiluminescent immunoassay

Chemiluminescent immunoassays are quantitative lab tests. They sample blood, plasma, or serum. Samples are mixed with a known viral protein, buffer reagents and specific, enzyme-labeled antibodies. The result is luminescent. A chemiluminescent microparticle immunoassay uses magnetic, protein-coated microparticles. Antibodies react to the viral protein, forming a complex. Secondary enzyme-labeled antibodies are added and bind to these complexes. The resulting chemical reaction produces light. The radiance is used to calculate the number of antibodies. This test can identify multiple types of antibodies, including IgG, IgM, and IgA.

Reference – Wikipedia