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{"project":"LitCovid-PD-GO-BP","denotations":[{"id":"T10","span":{"begin":84,"end":97},"obj":"http://purl.obolibrary.org/obo/GO_0003968"},{"id":"T11","span":{"begin":84,"end":97},"obj":"http://purl.obolibrary.org/obo/GO_0003899"},{"id":"T12","span":{"begin":521,"end":537},"obj":"http://purl.obolibrary.org/obo/GO_0016032"},{"id":"T13","span":{"begin":1068,"end":1082},"obj":"http://purl.obolibrary.org/obo/GO_0019076"},{"id":"T14","span":{"begin":2078,"end":2087},"obj":"http://purl.obolibrary.org/obo/GO_0006810"},{"id":"T15","span":{"begin":2767,"end":2774},"obj":"http://purl.obolibrary.org/obo/GO_0004526"},{"id":"T16","span":{"begin":2956,"end":2960},"obj":"http://purl.obolibrary.org/obo/GO_0003968"},{"id":"T17","span":{"begin":4858,"end":4874},"obj":"http://purl.obolibrary.org/obo/GO_0016032"},{"id":"T18","span":{"begin":6988,"end":7002},"obj":"http://purl.obolibrary.org/obo/GO_0019076"},{"id":"T19","span":{"begin":7632,"end":7646},"obj":"http://purl.obolibrary.org/obo/GO_0019076"}],"text":"How to Test: Diagnostic Tests in Use or Under Evaluation\nAlthough real-time reverse transcriptase polymerase chain reaction (RT-PCR)–based assays performed in the laboratory on respiratory specimens are the cornerstone of COVID-19 diagnostic testing, several novel or complementary diagnostic methods are being developed and evaluated (16). Figure 2 depicts the adequacy of the principal assay types used or proposed for COVID-19 for 4 key use cases. Among patients diagnosed with COVID-19, the occurrence of concomitant viral infections has been reported to range from below 6% (29) to greater than 60% (30). As a result, it is not possible to rule out SARS–CoV-2 infection merely by detecting another respiratory pathogen.\nFigure 2. Heat map showing the adequacy of principal assay types (rows) for 4 key use cases.\n* This assumes that assays in development or currently undergoing regulatory evaluation prove to be accurate.\n† The utility of antibody detection assays for diagnosing acute infections is probably very limited around the time of symptom onset, when viral shedding and transmission risk seem to be highest. Thus, although such tests may have a role among persons presenting late in the course of their infection, the potential for misuse is high.\n\nLaboratory-Based Molecular Testing\nThe current diagnostic strategy recommended by the CDC to identify patients with COVID-19 is to test samples taken from the respiratory tract to assess for the presence of 1 or several nucleic acid targets specific to SARS–CoV-2 (25). A nasopharyngeal specimen is the preferred choice for swab-based SARS–CoV-2 testing, but oropharyngeal, mid-turbinate, or anterior nares samples also are acceptable (31, 32). Samples should be obtained by using a flocked swab, if available, to enhance the collection and release of cellular material. Swabs with an aluminum or plastic shaft are preferred. Swabs that contain calcium alginate, wood, or cotton should be avoided, because they may contain substances that inhibit PCR testing. Ideally, swabs should be transferred into universal transport medium immediately after sample collection to preserve viral nucleic acid. Samples taken from sputum, endotracheal aspirates, and bronchoalveolar lavage also may be sent directly to the microbiology laboratory for processing, and may have greater sensitivity than upper respiratory tract specimens (33). Inadequate sample collection may result in a false-negative test. After specimen collection, samples undergo RNA extraction followed by qualitative RT-PCR for target detection.\nIn the United States, the CDC has developed the most widely used SARS–CoV-2 assay. The kit contains PCR primer–probe sets for 2 regions of the viral nucleocapsid gene (N1 and N2), and for the human RNase P gene to ensure the RNA extraction was successful. This assay differs from the World Health Organization primer–probe sets, which target the SARS–CoV-2 RNA-dependent RNA polymerase (RdRP) and envelope (E) genes (25). Both assays have high analytic sensitivity and specificity for SARS–CoV-2, with minimal cross-reactivity with other circulating strains of coronaviruses, and both use a cycle threshold of less than 40 as the criterion for positivity. The CDC kit may be used by state public health laboratories, other laboratories determined by the state to be qualified, and clinical laboratories that meet the regulatory requirements of the Clinical Laboratories Improvement Amendment (CLIA) to perform high-complexity testing (27). Dozens of laboratories have applied for Emergency Use Authorization (EUA) from the FDA for their own laboratory-developed assays (34). The FDA also has granted an EUA for several commercial assays (35), further expanding the ability of clinical laboratories to use these platforms (Table).\nTable. The 28 Commercial SARS–CoV-2 in Vitro Diagnostic Assays Given an EUA From the FDA as of 4 April 2020 The lack of an established reference standard, use of differing sample collection and preparation methods, and an incomplete understanding of viral dynamics across the time course of infection hamper rigorous assessment of the diagnostic accuracy of the many newly introduced SARS–CoV-2 assays (36). Serum and urine are usually negative for the presence of viral nucleic acid, regardless of illness severity (33). Of importance, the ability of RT-PCR assays to rule out COVID-19 on the basis of upper respiratory tract samples obtained at a single time point remains unclear. Conversely, after a patient has had a positive test result, several authorities have recommended obtaining at least 2 negative upper respiratory tract samples, collected at intervals of 24 hours or longer, to document SARS–CoV-2 clearance (37, 38).\n\nPoint-of-Care Molecular Diagnostics\nLow-complexity, rapid (results within 1 hour) molecular diagnostic tests for respiratory viral infections that are CLIA waived (FDA approved for use outside the laboratory by nonlaboratory personnel) include cartridge-based assays on platforms that include the Abbott ID NOW (Abbott Laboratories), BioFire FilmArray (bioMérieux), cobas Liat (Roche Diagnostics), and GeneXpert (Cepheid) (39).\nRapid point-of-care assays for SARS–CoV-2 on instruments such as these will be critical to expand point-of-care testing. The Xpert Xpress SARS–CoV-2 test (Cepheid) has received an FDA EUA and is performed on the GeneXpert platform, which is already widely used for tuberculosis and HIV testing, especially in low- and middle-income countries. This capacity might be useful to scale up testing across the world as well as in settings where rapid results at the point of care would enable clinical decisions, although testing throughput may be a limiting factor.\n\nAntigen Detection Tests\nTests that detect respiratory syncytial virus or influenza virus antigens by immunoassay directly from clinical specimens have been commercially available for decades, are of low complexity, and may provide results within minutes at the point of care (40). Current tools for influenza and respiratory syncytial virus suffer from suboptimal sensitivity to rule out disease (41, 42); the same challenge would probably exist for SARS–CoV-2, and tests would need to be implemented with clear guidance on correct interpretation. Prototypes of such tests for other novel coronaviruses have not received regulatory approval (43, 44) but are under development (45). Monoclonal antibodies against the nucleocapsid protein of SARS–CoV-2 have been generated, which might form the basis of a future rapid antigen detection test (20).\n\nSerology\nSerologic tests that identify antibodies (such as IgA, IgM, and IgG) to SARS–CoV-2 from clinical specimens (such as blood or saliva), such as enzyme-linked immunosorbent assays, may be less complex than molecular tests and have the potential to be used for diagnosis in certain situations (46). However, their utility for diagnosing acute infections is probably limited around the time of symptom onset, when viral shedding and transmission risk seem to be highest (32). Antibody responses to infection take days to weeks to be reliably detectable (46). Negative results would not exclude SARS–CoV-2 infection, particularly among those with recent exposure to the virus. Cross-reactivity of antibody to non–SARS–CoV-2 coronavirus proteins is also a potential problem, whereby positive results may be the result of past or present infection with other human coronaviruses (47). Serologic assays might be more relevant in scenarios in which patients present to medical care with late complications of disease, when RT-PCR may be falsely negative, because viral shedding drops over time (48).\nThe development of serologic assays that accurately assess prior infection and immunity to SARS–CoV-2 will be essential for epidemiologic studies, ongoing surveillance, vaccine studies, and potentially for risk assessment of health care workers. Immunoassays are already on the market in some countries, but their diagnostic accuracy and optimal use remain undefined."}

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