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COVID-19 testing involves analyzing samples to assess the current or past presence of SARS-CoV-2. The two main branches detect either the presence of the virus or of antibodies produced in response to infection. Tests for viral presence are used to diagnose individual cases and to allow public health authorities to trace and contain outbreaks. Antibody tests instead show whether someone once had the disease. They are less useful for diagnosing current infections because antibodies may not develop for weeks after infection. It is used to assess disease prevalence, which aids the estimation of the infection fatality rate.
Individual jurisdictions have adopted varied testing protocols, including whom to test, how often to test, analysis protocols, sample collection and the uses of test results. This variation has likely significantly impacted reported statistics, including case and test numbers, case fatality rates and case demographics. Because SARS-CoV-2 transmission occurs days after exposure (and before onset of symptoms) there is an urgent need for frequent surveillance and rapid availability of results.
Test analysis is often performed in automated, high-throughput, medical laboratories by medical laboratory scientists. Alternatively, point-of-care testing can be done in physician's offices and parking lots, workplaces, institutional settings or transit hubs.
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.
Detection of the virus
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.
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. The Minimum Information for Publication of Quantitative Real-Time PCR Experiments (MIQE) guidelines propose the term RT-qPCR, but not all authors adhere to this.
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%) between test results of different company brands and sampling methods.
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."
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.
Collecting saliva may be as effective as nasal and throat swabs, although this is not certain. Sampling saliva may reduce the risk for health care professionals by eliminating close physical interaction. It is also more comfortable for the patient. Quarantined people can collect their own samples. A saliva test's diagnostic value depends on sample site (deep throat, oral cavity, or salivary glands). Some studies have found that saliva yielded greater sensitivity and consistency when compared with swab samples.
On 4 January 2021, the US FDA issued an alert about the risk of false results, particularly false negative results, with the Curative SARS-Cov-2 Assay real-time RT-PCR test.
Demonstration of a nasopharyngeal swab for COVID-19 testing
Demonstration of a throat swab for COVID-19 testing
Isothermal amplification assays
Isothermal nucleic acid amplification tests also amplify the virus's genome. They are faster than PCR because they don't involve repeated heating and cooling cycles. These tests typically detect DNA using fluorescent tags, which are read out with specialized machines. CRISPR gene editing technology was modified to perform the detection: if the CRISPR enzyme attaches to the sequence, it colors a paper strip. The researchers expect the resulting test to be cheap and easy to use in point-of-care settings. The test amplifies RNA directly, without the RNA-to-DNA conversion step of RT-PCR.
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," Deborah Birx, head of the White House Coronavirus Task Force, said on 17 April 2020. "But there might be with the antigen test."
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%. "Based on this information, half or more of COVID-19 infected patients might be missed by such tests, depending on the group of patients tested," the WHO said. While some scientists doubt whether an antigen test can be useful against COVID-19, others have argued that antigen tests are highly sensitive when viral load is high and people are contagious, making them suitable for public health screening. Routine antigen tests can quickly identify when asymptomatic people are contagious, while follow-up PCR can be used if confirmatory diagnosis is needed.
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.
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.
SARS-CoV-2 antibodies' potency and protective period have not been established. Therefore, a positive antibody test may not imply immunity to a future infection. Further, whether mild or asymptomatic infections produce sufficient antibodies for a test to detect has not been established. Antibodies for some diseases persist in the bloodstream for many years, while others fade away.
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.
Average specificity of antigen tests is 99.5%, and average sensitivity is 56.8%, but there is extreme variation in sensitivity results (ranging from 0 to 94%) between test results of different company brands.
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.
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.
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 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 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.
Neutralizing vis-à-vis binding antibodies
Most if not all large scale COVID-19 antibody testing looks for binding antibodies only and does not measure the more important neutralizing antibodies (NAb). A NAb is an antibody that defends a cell from an infectious particle by neutralizing its biological effects. Neutralization renders the particle no longer infectious or pathogenic. A binding antibody binds to the pathogen but the pathogen remains infective; the purpose can be to flag the pathogen for destruction by the immune system. It may even enhance infectivity by interacting with receptors on macrophages. Since most COVID-19 antibody tests return a positive result if they find only binding antibodies, these tests cannot indicate that the subject has generated protective NAbs that protect against re-infection.
It is expected that binding antibodies imply the presence of NAbs and for many viral diseases total antibody responses correlate somewhat with NAb responses but this is not established for COVID-19. A study of 175 recovered patients in China who experienced mild symptoms reported that 10 individuals had no detectable NAbs at discharge, or thereafter. How these patients recovered without the help of NAbs and whether they were at risk of re-infection was not addressed. An additional source of uncertainty is that even if NAbs are present, viruses such as HIV can evade NAb responses.
Studies have indicated that NAbs to the original SARS virus (the predecessor to the current SARS-CoV-2) can remain active for two years and are gone after six years. Nevertheless, memory cells including Memory B cells and Memory T cells can last much longer and may have the ability to reduce reinfection severity.
A Point of Care Test in Peru. A blood droplet is collected by a pipette.
Blood from pipette is then placed onto a COVID-19 rapid diagnostic test device.
Following recovery, many patients no longer have detectable viral RNA in upper respiratory specimens. Among those who do, RNA concentrations three days following recovery are generally below the range in which replication-competent virus has been reliably isolated.
No clear correlation has been described between length of illness and duration of post-recovery shedding of viral RNA in upper respiratory specimens.
Infectivity is indicated by the basic reproduction number (R0, pronounced "R naught") of the disease. SARS-CoV-2 is estimated to have an R0 of 2.2 to 2.5. This means that in a population where all individuals are susceptible to infection, each infected person is expected to infect 2.2 to 2.5 others in the absence of interventions. R0 can vary according factors such as geography, population demographics and density. In New York state R0 was estimated to be 3.4 to 3.8.
On average, an infected person begins showing symptoms five days after infection (the "incubation period") and can infect others beginning two to three days before that. One study reported that 44% of viral transmissions occur within this period. According to the CDC, a significant number of infected people who never show symptoms are nevertheless contagious. In vitro studies have not found replication-competent virus after 9 days from infection. The statistically estimated likelihood of recovering replication-competent virus approaches zero by 10 days.
Infectious virus has not been cultured from urine or reliably cultured from feces; these potential sources pose minimal if any risk of transmitting infection and any risk can be sufficiently mitigated by good hand hygiene.
Patterns and duration of illness and infectivity have not been fully described. However, available data indicate that SARS-CoV-2 RNA shedding in upper respiratory specimens declines after symptom onset. At 10 days recovery of replication-competent virus in viral culture (as a proxy of the presence of infectious virus) approaches zero. Although patients may produce PCR-positive specimens for up to six weeks, it remains unknown whether these samples hold infectious virus. After clinical recovery, many patients do not continue to shed. Among recovered patients with detectable RNA in upper respiratory specimens, concentrations after three days are generally below levels where virus has been reliably cultured. These data were generated from adults across a variety of age groups and with varying severity of illness. Data from children and infants were not available.
Public Health England announced a test on the 10th, using a real-time RT-PCR (RdRp gene) assay based on oral swabs. The test detected the presence of any type of coronavirus, including specifically identifying SARS-CoV-2. It was rolled out to twelve laboratories across the United Kingdom on 10 February.
Scientists from China first released information on the viral genome on 11 January 2020, sending multiple genomic sequences to GISAID, an indispensable mechanism for sharing influenza genetic sequence data. That day the Malaysian Institute for Medical Research (IMR) produced "primers and probes" specific to a SARS-CoV-2 RT-PCR test. The IMR's materials were used to diagnose Malaysia's first patient on 24 January. BGI Group was one of the first companies to receive emergency use approval from China's National Medical Products Administration for a nucleic acid test.
The German nucleic acid testing protocol was published on the 17th. Another early PCR test was developed by Charité University hospital in Berlin, working with academic collaborators in Europe and Hong Kong, and published on the 23rd. It used rtRT-PCR, and formed the basis of 250,000 kits distributed by the World Health Organization (WHO).
The first case in South Korea was confirmed on 19 January.
In Russia, the first COVID‑19 test was developed by the State Research Center of Virology and Biotechnology VECTOR. Production began on 24 January.
In the US, the Centers for Disease Control and Prevention (CDC) developed its SARS-CoV-2 Real Time PCR Diagnostic Panel. The protocol became available on the 28th. One of three tests in early kits failed due to faulty reagents.
South Korean company Kogenebiotech's clinical grade, nucleic acid test (PowerChek Coronavirus) was approved by Korea Centers for Disease Control and Prevention (KCDC) on 4 February.
In Wuhan, BGI opened a makeshift 2000-sq-meter emergency detection laboratory named "Huo-Yan" (Chinese: 火眼, "Fire Eye") on the 5th. It processed more than 10,000 samples/day. Construction required 5 days. The Wuhan Laboratory was followed by Huo-Yan labs in Shenzhen, Tianjin, Beijing, and Shanghai, in a total of 12 cities across China.
On 11 February, the test was approved by the Federal Service for Surveillance in Healthcare in Russia.
In the United States, the CDC refused to let other labs process tests that month, allowing an average of fewer than 100 samples/day to be processed. Tests using two components were not determined to be reliable until the 28th, and only then were state and local laboratories permitted to begin testing. The test was approved by the FDA under an EUA.
Due to limited testing, no countries had reliable data on the prevalence of the virus in their population. Testing variability distorts reported case fatality rates, which were probably overestimated in many countries due to sampling bias. Shortages of reagent and other supplies became a bottleneck for mass testing in the EU and UK and the US.
By 4 March, China reached 50,000 tests per day. Early in March, China reported accuracy problems with its PCR tests. A study examined 1070 samples from 205 Wuhan patients and reported varied sensitivity according to the methods and location of sample collection. Samples from bronchoalveolar lavage fluid specimens returned the highest sensitivity. The authors argued that CT scans showed even higher sensitivity.
US commercial labs began testing in early March. As of the 5th, LabCorp announced nationwide availability of COVID‑19 testing based on RT-PCR. Quest Diagnostics made nationwide testing available as of 9 March. US testing demand grew rapidly, causing backlogs of hundreds of thousands of tests at private US labs. Supplies of swabs and chemical reagents continued strained. On 25 May, the US required each state to take responsibility for meeting its testing needs. In March, the FDA issued EUAs for nucleic acid tests to Hologic (3/16), Abbott Laboratories (3/18), Thermo Fisher Scientific (3/19) Cepheid (3/21) and LabCorp (4/30).
On 16 March, the WHO called for ramping up testing programmes as the best way to slow the spread. Several European countries initially conducted more tests than the US. By 19 March, drive-in tests were offered in several large cities.
As of 22 March, according to the president of the Robert Koch Institute, Germany had capacity for 160,000 tests per week. As of 26 March, German Health Minister Jens Spahn estimated that Germany was conducting 200,000 tests per week. Germany has a large medical diagnostics industry, with more than a hundred testing labs that provided the technology and infrastructure to enable rapid increases in testing. Costs are borne by insurance when the test is ordered by a physician. As of the end of March at least 483,295 samples were tested and 33,491 (6.9%) had tested positive.
On 26 March, it was reported that 80% of test kits that Czechia purchased from China gave inaccurate results. Slovakia purchased 1.2 million antibody-based test kits from China that were found to be inaccurate. China accused Czechia and Slovakia of incorrect use of those tests. Ateş Kara of the Turkish Health Ministry said the test kits Turkey purchased from China had a "high error rate".
Spain purchased test kits from Chinese firm Shenzhen Bioeasy Biotechnology Co Ltd, but found that results were unacceptable. The maker explained that the incorrect results may stem from failure to collect samples or use the kits correctly. On 27 March, the Spanish ministry switched to another vendor, Shenzhen Bioeasy.
By 31 March, the United Arab Emirates was testing more of its population per head than any other country. UAE implemented a combination of drive-through sample collection, and a mass-throughput laboratory from Group 42 and BGI. The lab conduced tens of thousands RT-PCR tests per day and was the first to be operational at that scale other than China.
By the month's end, testing had surpassed 200k/week.
On 5 April, the U.S. subsidiary of China's BGI Group sent a proposal to the state of California offering to build in California, at cost ($10 million), the world's largest COVID-19 testing site, in two weeks, and train Americans to operate it. California's consultants recommended against it, because of the risk of security and commercial competition.
On 13 April, Health Canada approved a nucleic acid test from Spartan Bioscience. Institutions may "test patients" with a handheld DNA analyzer "and receive results without having to send samples away to a [central] lab".
By the start of April, the United Kingdom was delivering around 10,000 swab tests per day. The British NHS announced that it was piloting a scheme to test suspected cases at home, to remove the risk of one patient infecting others at a hospital or disinfecting an ambulance used to transport a patient.
The UK purchased 3.5 million antibody test kits from China, but in early April 2020 announced these were not usable. On 21 April 2020, the Indian Council of Medical Research (ICMR) advised Indian states to stop using test kits purchased from China after receiving complaints from one state. Rajasthan health minister Raghu Sharma on 21 April said the kits gave only 5.4 percent accurate results.
By month end, testing had surpassed 750k/week.
In May antibody tests were conducted on 5,603 major league baseball employees and 0.7% tested positive, showing that they had been infected. 70% of those who tested positive had had no symptoms. The US was conducting an average of 2.5 million tests per week for the week ending 17 May. This grew to 3.2 million by 14 June.
On 1 May, Quotient Limited announced the CE Mark for its MosaiQ COVID-19 antibody test, designed as a serological disease screen specific to the Coronavirus. The test has a 100% sensitivity and 99,8% specificity claim.
The FDA announced on 14 May a review of 15 adverse event reports about the Abbott ID Now device for low sensitivity.
On 21 May, researchers at Ben-Gurion University in Israel reported a one-minute coronavirus test with 90% accuracy, based on the "change in the resonance in the THz spectral range" shown by the coronavirus through THz spectroscopy.
As of 24 May, countries that publicised their testing data had typically performed tests equal to 2.6 percent of their population, although no country had tested more than 17.3%.
On 29 May Siemens received an EUA for its anti-spike RBD-targeting serology test that it believes detects neutralizing antibodies.
By month end, testing had surpassed 1.4m/week.
In June, researchers announced a nucleic acid diagnostic test using reverse transcription-loop-mediated isothermal amplification (RT-LAMP), an existing technology used in pathogenic microorganism identification, genetically modified ingredients, tumor detection, and embryo sex identification. The test identified virus in samples of serum, urine, saliva, oropharyngeal swabs and nasopharyngeal swabs. Once commercialized the test has the potential to provide rapid (30-45 minute) diagnosis at point of care. The test was 100% selective and highly sensitive, detecting virus at a concentration of .06 fg/ml.
On 6 November, the U.S. Food and Drug Administration (FDA) authorized the first serology test that detects neutralizing antibodies from recent or prior SARS-CoV-2 infection, which are antibodies that bind to a specific part of a pathogen and have been observed in a laboratory setting to decrease SARS-CoV-2 viral infection of cells. The FDA issued an emergency use authorization (EUA) for the cPass SARS-CoV-2 Neutralization Antibody Detection Kit, which specifically detects this type of antibody. The FDA granted Lucira Health emergency use authorization for the first US at-home rapid molecular diagnostic test. With a prescription from a healthcare provider, consumers can use the test kit to take a nasal swab then perform a 30-minute SARS-CoV-2 detection test at home.
On 15 December, the U.S. Food and Drug Administration (FDA) issued an emergency use authorization (EUA) for the first over-the-counter (OTC) fully at-home diagnostic test for COVID-19. The Ellume COVID-19 Home Test is a rapid, lateral flow antigen test, a type of test that runs a liquid sample along a surface with reactive molecules. The test detects fragments of proteins of the SARS-CoV-2 virus from a nasal swab sample from any individual two years of age or older. The Ellume COVID-19 Home Test uses a mid-turbinate nasal swab (sample is collected further back than the usual nasal swab, but not as far back as nasopharyngeal swabs, which are only appropriate for use by a trained health care provider) to detect certain proteins of the virus known as antigens. The Ellume COVID-19 Home Test uses an analyzer that connects with a software application on a smartphone to help users perform the test and interpret results. Results are delivered in as little as 20 minutes to individuals via their smartphone.
In drive-through testing, the person undergoing testing remains in a vehicle while a healthcare professional approaches the vehicle and obtains a sample, all while taking appropriate precautions such as wearing personal protective equipment (PPE). Drive-through centers helped South Korea accelerate its testing program.
In Hong Kong test subjects can stay home and receive a specimen tube. They spit into it, return it and later get the result.
In Israel, researchers at Technion and Rambam Hospital developed a method for testing samples from 64 patients simultaneously, by pooling the samples and only testing further if the combined sample was positive. Pool testing was then adopted in Israel, Germany, Ghana South Korea, Nebraska, China and the Indian states of Uttar Pradesh, West Bengal, Punjab, Chhattisgarh and Maharashtra.
Open source, multiplexed designs released by Origami Assays can test as many as 1122 patient samples using only 93 assays. These balanced designs can be run in small laboratories without robotic liquid handlers.
One study proposed a rapid immune response assay as a screening test, with a confirmatory nucleic acid test for diagnosis, followed by a rapid antibody test to determine course of action and assess population exposure/herd immunity.
Required testing levels are a function of disease spread. The more the cases, the more tests are needed to manage the outbreak. COVID-19 tends to grow exponentially at the beginning of an outbreak, meaning that the number of required tests initially also grows exponentially. If properly targeted testing grows more rapidly than cases, it can be contained.
WHO recommends increasing testing until fewer than 10% are positive in any given jurisdiction.
Economist Paul Romer reported that the US has the technical capacity to scale up to 20 million tests per day, which is his estimate of the scale needed to fully remobilize the economy. The Edmond J. Safra Center for Ethics estimated on 4 April that this capacity could be available by late July. Romer pointed to single-molecule real-time sequencing equipment from Pacific Biosciences and to the Ion Torrent Next-Generation Sequencing equipment from ThermoFisher Scientific. According to Romer, "Recent research papers suggest that any one of these has the potential to scale up to millions of tests per day." This plan requires removing regulatory hurdles. Romer estimated that $100 billion would cover the costs.
Romer also claimed that high test accuracy is not required if tests are administered frequently enough. He ran model simulations in which 7% of the population is tested every day using a test with a 20% false negative rate and a 1% false positive rate. The average person would be tested roughly every two weeks. Those who tested positive would go into quarantine. Romer's simulation indicated that the fraction of the population that is infected at any given time (known as the attack rate) peaks reaches roughly 8% in about thirty days before gradually declining, in most runs reaching zero at 500 days, with cumulative prevalence remaining below 20%.
Countries around the world developed tests independently and in partnership with others.
Nucleic acid tests
Tests developed in China, France, Germany, Hong Kong, Japan, the United Kingdom, and the US targeted different parts of the viral genome. WHO adopted the German system for manufacturing kits sent to low-income countries without the resources to develop their own.
Abbott Laboratories' ID Now nucleic acid test uses isothermal amplification technology. The assay amplifies a unique region of the virus's RdRp gene; the resulting copies are then detected with "fluorescently-labeled molecular beacons". The test kit uses the company's "toaster-size" ID Now device, which is widely deployed in the US. The device can be used in laboratories or in point of care settings, and provides results in 13 minutes or less.
Primerdesign offers its Genesig Real-Time PCR Coronavirus (COVID‑19). Cobas SARS-CoV-2 Qualitative assay runs on the Cobas® 6800/8800 Systems by Roche Molecular Systems. They are offered by the United Nations and other procurement agencies.
Quidel's "Sofia 2 SARS Antigen FIA" is a lateral flow test that uses monoclonal antibodies to detect the virus's nucleocapsid (N) protein. The result is read out by the company's Sofia 2 device using immunofluorescence. The test is simpler and cheaper but less accurate than nucleic acid tests. It can be deployed in laboratories or at point of care and gives results in 15 minutes. A false negative result occurs if the sample's antigen level is positive but below the test's detection limit, requiring confirmation with a nucleic acid test.
Serology (antibody) tests
Antibodies are usually detectable 14 days after the onset of the infection. Multiple jurisdictions survey their populations using these tests. The test requires a blood draw.
A summary review in BMJ has noted that while some "serological tests … might be cheaper and easier to implement at the point of care [than RT-PCR]", and such testing can identify previously infected individuals, "caution is warranted … using serological tests for … epidemiological surveillance". The review called for higher quality studies assessing accuracy with reference to a standard of "RT-PCR performed on at least two consecutive specimens, and, when feasible, includ[ing] viral cultures." CEBM researchers have called for in-hospital 'case definition' to record "CT lung findings and associated blood tests" and for the WHO to produce a "protocol to standardise the use and interpretation of PCR" with continuous re-calibration.
|Samples source||Positive rate|
|Bronchoalveolar lavage fluid specimens||93% (14/15)|
|Nasal swabs||63% (5/8)|
|Fibrobronchoscope brush biopsy||46% (6/13)|
|Pharyngeal swabs||32% (126/398)|
Accuracy is measured in terms of specificity and selectivity. Test errors can be false positives (the test is positive, but the virus is not present) or false negatives, (the test is negative, but the virus is present).
Sensitivity and specificity
Sensitivity indicates whether the test accurately identifies whether the virus is present. Each test requires a minimum level of viral load in order to produce a positive result. A 90% sensitive test will correctly identify 90% of infections, missing the other 10% (a false negative). Even relatively high sensitivity rates can produce high rates of false negatives in populations with low incidence rates.
Specificity indicates how well-targeted the test is to the virus in question. Highly specific tests pick up only the virus in question. Non-selective tests pick up other viruses as well. A 90% specific test will correctly identify 90% of those who are uninfected, leaving 10% with a false positive result.
Low-specificity tests have a low positive predictive value (PPV) when prevalence is low. For example, suppose incidence is 5%. Testing 100 people at random using a test that has a specificity of 95% would yield on average 5 people who are actually negative who would incorrectly test positive. Since 5% of the subjects actually are positive, another five would also test positive correctly, totaling 10 positive results. Thus, the PPV is 50%, an outcome no different from a coin toss. In this situation retesting those with a positive result increases the PPV to 94.5%, meaning that only 4.5% of the second tests would return the incorrect result, on average less than 1 incorrect result.
Causes of test error
Improper sample collection, exemplified by failure to acquire enough sample and failure to insert a swab deep into the nose. This results in insufficient viral load, one cause of low clinical sensitivity.
The time course of infection also affects accuracy. Samples may be collected before the virus has had a chance to establish itself or after the body has stopped its progress and begun to eliminate it. A May 2020 review of PCR-RT testing found that the median probability of a false-negative result decreased from 100% on day 1, to 67% on day 4. On the day of symptom onset, the probability was 38%, which decreased to 20% 3 days later.
Improper storage for too long a time can cause RNA breakdown and lead to wrong results as viral particles disintegrate.
Improper design and manufacture can yield inaccurate results. Millions of tests made in China were rejected by various countries throughout the period of March 2020 through May 2020.
Test makers typically report the accuracy levels of their tests when seeking approval from authorities. In some jurisdictions, these results are cross-validated by additional assessments. Reported results may not be achieved in clinical settings due to such operational inconsistencies.
High sensitivity kits are recommended to assess people without symptoms, while lower sensitivity tests are adequate when diagnosing symptomatic patients.
The University of Oxford's Centre for Evidence-Based Medicine (CEBM) has pointed to mounting evidence that "a good proportion of 'new' mild cases and people re-testing positives via RT-PCR after quarantine or discharge from hospital are not infectious, but are simply clearing harmless virus particles which their immune system has efficiently dealt with", and have called for "an international effort to standardize and periodically calibrate testing". On 7 September, the UK government issued "guidance for procedures to be implemented in laboratories to provide assurance of positive SARS-CoV-2 RNA results during periods of low prevalence, when there is a reduction in the predictive value of positive test results".
On 4 January 2021, the US FDA issued an alert about the risk of false results, particularly false negative results, with the Curative SARS-Cov-2 Assay real-time RT-PCR test.
Isothermal nucleic amplification test
One study reported that the ID Now COVID-19 test showed sensitivity of 85.2%. Abbott responded that the issue could have been caused by analysis delays. Another study rejected the test in their clinical setting because of this low sensitivity.
The WHO recommends countries that do not have testing capacity and national laboratories with limited experience on COVID‑19 send their first five positives and the first ten negative COVID‑19 samples to one of the 16 WHO reference laboratories for confirmatory testing. Out of the sixteen reference laboratories, seven are in Asia, five in Europe, two in Africa, one in North America and one in Australia.
Iceland managed the pandemic with aggressive contact tracing, inbound travel restrictions, testing, and quarantining, but with less aggressive lock-downs.
Researchers tested the entire population of Vò, the site of Italy's first COVID‑19 death. They tested about 3,400 people twice, at an interval of ten days. About half the people testing positive had no symptoms. All discovered cases were quarantined. Along with restricting travel to the commune, new infections were completely eliminated.
Unlike other Asian countries, Japan did not experience a pandemic of SARS or MERS, so the country's PCR testing system was not well developed. Japan preferentially tested patients with severe illness and their close contacts at the beginning. Japan's Novel Coronavirus Expert Meeting chose cluster measures to identify infections clusters. The Expert Meeting analyzed the outbreak from Wuhan and identified conditions leading to clusters (closed spaces, crowded spaces and close-contact), and asked people to avoid them.
In January, contact tracers took action shortly after the first infection was found. Only administrative tests were carried out at first, until insurance began covering PCR tests on 6 March. Private companies began to test, and the test system gradually expanded.
On 3 April, those with positive tests were legally permitted to recuperate at home or in a hotel if they had asymptomatic or mild illness, ending the hospital bed shortage. The first wave (from China) was contained, but a second wave (caused by returnees from Europe and the US) in mid-March led to spreading infection in April. On 7 April, Japan declared a state of emergency, (less strict than a lockdown, because it did not block cities or restrict outings). On 13 May, antigen test kits became covered by insurance, and were combined with a PCR test for diagnosis.
Japan's PCR test count per capita remained far smaller than in some other countries even though its positive test rate was lower. Excess mortality was observed in March.[failed verification][failed verification] The Expert Meeting stated, "The Japanese health care system originally carries out pneumonia surveillance, allowing it to detect most of the severely ill patients who develop pneumonia. There are a large number of CT scanners in Japan and they have spread to small hospitals all over the country, so pneumonia patients are rarely missed. In that sense, it meets the same standards as other countries that mainly carry out PCR tests." The group recommended using CT scans data and doctor's findings for diagnosis. On the Diamond Princess cruise ship, many people who initially tested negative later tested positive. Half of coronavirus-positives there who remained mild or asymptomatic had pneumonia findings on CT scans and their CT image showed a frosted glass shadow that is characteristic of infection.
As of 18 July, Japan's daily PCR testing capacity was about 32,000, more than three times the 10,000 cases as of April. When the antigen test is added to it, the number is about 58,000. The number of tests per 1,000 people in the United States is about 27 times that of Japan, the UK is 20 times, Italy is 8 times, and South Korea is twice (as of 26 July).
The number of those infected with coronavirus and inpatients has increased in July, but the number of serious cases has not increased. This is thought to be due to the proper testing of those infected in July compared to those in April. In April, the number of tests could not catch up with the increase in the number of infected people, and the test standards were strict, so the test positive rate exceeded 30% at the peak. It means that there were quite a few cases where the those infected was not PCR tested. It is thought that the severe case was preferentially tested though there were a lot of mild cases and asymptomatic carriers mainly in the young during the first wave. In other words, it became possible to grasp the actual situation of infection much better than before by strengthening the testing system. At the end of July, accommodation facilities for mild and asymptomatic carriers became full, and the authorities requested hospitals to prepare beds for the mild. However, it became difficult to treat patients with other illnesses and to maintain the ICU system including the staff due to the occupation of hospital beds by patients with mild symptoms.
On 27 April, Russia tested 3 million people and had 183,000 positive results. On 28 April Anna Popova, head of Federal Service for Surveillance in Healthcare (Roszdravnadzor) stated that 506 laboratories were testing; that 45% of those who tested positive had no symptoms; that 5% of patients had a severe form; and 40% of infections were from family members. Illness improved from six days to one day after symptoms appeared. Antibody testing was carried out on 3,200 Moscow doctors, finding 20% immunity.
In late October 2020 Slovakia tested 3.62 million people in a weekend, from a population of 5.4m, representing 67% of the total (or 82% of the adult population), 38,359 tested positive, representing 1.06% of those tested. The government considered the mass test would significantly assist in controlling the virus and avoid a lockdown and may repeat the exercise at a later date.
South Korea's broad testing approach helped reduce spread. Testing capacity, largely in private sector labs, was built up over several years by the South Korean government in the early 2000s.
The government exploited the resident registration number (RRN) system. Authorities mobilized young men who were eligible for military service as social service agents, security and public health doctors. Public health doctors were mainly dispatched to public health centers and life treatment centers where mildly ill patients were accommodated. They performed PCR tests and managed mild patients. Social service agents worked in pharmacies to fill staff shortages. Korea's 10k PCR tests per million residents was the world's highest as of 13 April rising to 20k by mid-June. Twenty-seven Korean companies exported test kits worth $48.6 million in March, and were asked to provide test kits or humanitarian assistance by more than 120 countries. Korean authorities set up a treatment center to isolate and manage patients with asymptomatic and minor illnesses in one facility in order to vacate hospital beds for the more severely ill.
Centers were sited mainly at national facilities and corporate training centers. The failure of Korea's MERS quarantine in May 2015 left Korea more prepared for COVID-19 than countries that did not face that pandemic. Then President Park Geun-hye allowed Korean CDC-approved private sector testing for infectious diseases in 2016. Korea already had a system for isolating, testing and treating infectious disease patients separately from others. Patients with respiratory illness but no epidemiological relevance were treated at the National Hospital, and those with epidemiological relevance were treated at selected clinics.
Korea established a large scale drive-through/walk-through" test testing program. However, the most common method was "mobile examination". In Daegu City, 54% of samples were collected by 23 March in home or hospital. Collecting samples door-to-door of avoided the risk of travel by possibly infected patients, but required additional staff. Korea solved the problem by drafting more than 2,700 public insurance doctors.
The government disclosed personal information to the public via KCDC without patient consent. The authorities used digital surveillance to trace possible spread.
New York State
New York State's control measures consisted of PCR tests, stay-at-home measures and strengthening the healthcare system. On 29 February before its first case, the state allowed testing at the Wordsworth Center. They managed to convince the CDC to approve tests at state laboratories and the FDA to approve a test kit. As of 13 March the state was conducting more than 1,000 daily tests, growing to 10,000/day on 19 March. In April, the number exceeded 20,000. Many people queued at hospitals to get tested. On 21 March New York City health officials directed medical providers to test only those entering the hospital, for lack of PPE.
USS Theodore Roosevelt
Following an outbreak, 94% of the 4,800 aircraft carrier crew were tested. Roughly 60 percent of the 600-plus sailors who tested positive were asymptomatic. Five infected sailors who completed quarantine subsequently developed flu-like symptoms and again tested positive.
A shortage of trained medical laboratory scientists, assay reagents, analyzers, transport medium, and PPE coupled with high demand had limited initially limited the availability of testing and led to significantly increased turnaround times.
Testing statistics by country
Testing strategies vary by country and over time, with some countries testing very widely, while others have at times focused narrowly on only testing the seriously ill. The country that tests only people showing symptoms will have a higher figure for "% (Confirmed cases as percentage of tested samples or tested cases)" than the country that also tests others. If two countries are alike in every respect, including which people they test, the one that tests more people will have a higher "Confirmed / million people". Studies have also found that countries that test more, relative to the number of deaths, have lower estimated case fatality rates and younger age distributions of cases.
|Antigua and Barbuda||15 January||8,198||samples||187||2.3||85,142||1,942|||
|Bosnia and Herzegovina||19 January||568,743||samples||118,383||20.8||166,225||34,600|||
|Burkina Faso||17 January||118,020||samples||9,188||7.8||5,646||440|||
|Costa Rica||15 January||546,784||samples||184,187||33.7||109,369||36,842|||
|Dominican Republic||15 January||947,423||samples||191,339||20.2||87,093||17,589|||
|DR Congo||13 January||102,460||20,478||20.0||1,144||229|||
|El Salvador||16 January||660,519||samples||50,784||7.7||101,834||7,830|||
|Equatorial Guinea||16 January||81,780||5,356||6.5||62,476||4,092|||
|Faroe Islands||20 January||216,677||samples||652||0.30||4,158,069||12,512|||
|Ivory Coast||15 January||293,382||samples||24,856||8.5||11,122||942|||
|New Caledonia||15 January||19,824||samples||41||0.21||73,042||151|||
|New Zealand||20 January||1,472,326||samples||1,911||0.13||295,435||383|||
|North Korea||19 June||922||cases||0||0||36||0|||
|North Macedonia||18 January||432,241||samples||88,992||20.6||208,095||42,844|||
|Northern Cyprus[i]||20 January||373,605||samples||1,971||0.53||1,146,028||6,046|||
|Papua New Guinea||17 January||40,697||cases||835||2.1||4,555||93|||
|Saint Kitts and Nevis||15 January||6,391||cases||34||0.53||121,870||648|||
|Saint Lucia||16 January||22,060||samples||611||2.8||121,283||3,359|||
|Saint Vincent||16 January||24,615||cases||478||1.9||223,344||4,337|||
|San Marino||20 January||31,303||samples||2,815||9.0||914,437||82,233|||
|Saudi Arabia||19 January||11,770,347||samples||365,325||3.1||338,094||10,494|||
|South Africa||16 January||7,558,774||cases||1,325,659||17.5||127,448||22,352|||
|South Korea||19 January||4,985,981||samples||73,115||1.5||96,424||1,414|||
|South Sudan||19 January||83,134||3,773||4.5||6,506||295|||
|Sri Lanka||16 January||1,459,691||samples||51,594||3.5||66,949||2,366|||
|The Gambia||15 January||34,280||samples||3,910||11.4||15,768||1,799|||
|Trinidad and Tobago||16 January||77,299||cases||7,370||9.5||56,671||5,403|||
|United Arab Emirates||19 January||23,706,842||samples||260,223||1.1||2,469,629||27,108|||
|United Kingdom||20 January||65,875,546||samples||3,505,754||5.3||975,273||51,902|||
|United States||20 January||287,519,743||samples||24,109,760||8.4||868,675||72,842|||
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