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Viewpoint
May 6, 2020

Interpreting Diagnostic Tests for SARS-CoV-2

Author Affiliations
  • 1Department of Microbiology, Apollo Hospitals, Chennai, India
  • 2Department of Microbiology, Yokohama City University, Yokohama, Japan
JAMA. 2020;323(22):2249-2251. doi:10.1001/jama.2020.8259

The pandemic of coronavirus disease 2019 (COVID-19) continues to affect much of the world. Knowledge of diagnostic tests for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is still evolving, and a clear understanding of the nature of the tests and interpretation of their findings is important. This Viewpoint describes how to interpret 2 types of diagnostic tests commonly in use for SARS-CoV-2 infections—reverse transcriptase–polymerase chain reaction (RT-PCR) and IgM and IgG enzyme-linked immunosorbent assay (ELISA)—and how the results may vary over time (Figure).

Thus far, the most commonly used and reliable test for diagnosis of COVID-19 has been the RT-PCR test performed using nasopharyngeal swabs or other upper respiratory tract specimens, including throat swab or, more recently, saliva. A variety of RNA gene targets are used by different manufacturers, with most tests targeting 1 or more of the envelope (env), nucleocapsid (N), spike (S), RNA-dependent RNA polymerase (RdRp), and ORF1 genes. The sensitivities of the tests to individual genes are comparable according to comparison studies except the RdRp-SARSr (Charité) primer probe, which has a slightly lower sensitivity likely due to a mismatch in the reverse primer.1

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    6 Comments for this article
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    Interpreting a Variety of Diagnostic Tests for SARS-CoV-2 and COVID-19
    Michael McAleer, PhD (Econometrics), Queen's | Asia University, Taiwan
    The detailed and informative paper on interpreting diagnostic tests for the SARS-CoV-2 virus that causes the COVID-19 disease is based on two types of diagnostics in common use, namely :

    (1) Reverse transcriptase–polymerase chain reaction (RT-PCR); 

    (2) IgM and IgG enzyme-linked immunosorbent assay (ELISA).

    It is striking that the results of the diagnostic tests can and do vary over time, which lends itself to the possibility of rigorous time series analysis, if appropriate data were to be made available. 

    The null hypothesis of any diagnostic test of the virus and the associated disease, as given in (1), is that
    a negative test is presumed, namely:

    Null Hypothesis (H0): No confirmation of the virus.

    The critical issue in using any diagnostic test, or combination of a variety of tests, is how to achieve a correct diagnosis of a positive outcome.  

    For a detailed discussion of diagnostic testing of COVID-19, see (2), which suggests a number of diagnostics of the viral infection, none of which seems to have been used in the invaluable Viewpoint.

    It is well known that the purpose of any diagnostic test is to achieve high power, which involves the rejection of a false null hypothesis, leading to a true positive diagnosis.

    In cases where several diagnostic tests are available, with none being paramount, the use of a variety of tests will lead to a more accurate diagnosis by increasing the number of tests.

    Based on available data for adult populations without being immunocompromised, the estimated variation over time in the outcomes of the diagnostic tests is considerable, in terms of amplitude, shape, and the length of time since the onset of the disease symptoms.

    As the estimated time intervals and rates of detection are based on separate reports, each of which has different assumptions and distinct methodologies, the data are approximations, such that the probability of detection of the disease, based on maximizing the power of the various test procedures, is presented qualitatively rather than quantitatively.

    The collection and availability of time series data from different studies, according to the age of patients, existing comorbidities, types of preexisting conditions, including the various stages such as for cancer patients, their current treatments, and genetic markers, would lead to an invaulable unbalanced panel data set.

    Such a rich set of data could be used to predict the probability of a more accurate detection of infections through a range of diagnostic tests.

    References

    1. McAleer, M. 2020. Is one diagnostic test for COVID-19 enough? Journal of Risk and Financial Management, 13(4:77), 1-3.

    2. Sharfstein, Joshua M., Becker. S, and Michelle M. Mello. 2020. Diagnostic testing for the novel coronavirus. JAMA. Published online March 9, 2020. doi:10.1001/jama.2020.3864.
    CONFLICT OF INTEREST: None Reported
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    Role of IgA in the Anti-SARS-CoV-2 Antibody Response
    Ger Rijkers, PhD | Laboratory for Medical Microbiology and Immunology, St. Elisabeth Hospital, Tilburg, The Netherlands
    In their Viewpoint on Interpreting Diagnostic Tests for SARS-CoV-2 Sethuraman and colleagues may have left out a major player of the humoral immune response against respiratory viruses, and that is IgA. By now there are a number of publications which demonstrate a clear-cut IgA anti-SARS-CoV-2 response (1-3). In our own series of over 35 patients, close analysis (3 serum samples per week) showed a robust IgA response with a kinetic pattern similar to that of the IgG response: on average 10 days after onset of symptoms, 50% of patients were positive for IgA and IgG antibodies and by week 3 plateau levels for IgA and IgG were reached. In the Figure of this paper the legend thus could be adapted by labeling of the green dashed curve as representing IgG and IgA antibodies.

    References

    1. Padoan A, Sciacovelli L, Basso D, et al. IgA-Ab response to spike glycoprotein of SARS-CoV-2 in patients with COVID-19: A longitudinal study. Clin Chim Acta. 2020; 507:164-166. doi: 10.1016/j.cca.2020.04.026.

    2. Béné MC, de Carvalho M, Eveillard M, Lebri Y. Good IgA bad IgG in SARS-CoV-2 infection? Clin Infect Dis. 2020. pii: ciaa426. doi: 10.1093/cid/ciaa426.

    3. Okba NMA, Müller MA, Li W, Wang C, et al. Severe Acute Respiratory Syndrome Coronavirus 2- Specific Antibody Responses in Coronavirus Disease 2019 Patients. Emerg Infect Dis. 2020;26(7).
    CONFLICT OF INTEREST: None Reported
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    Looking Forward to Updating This With Results of Newly Emerging Antigen Test
    William Prendergast, MD | Retired Ophthalmologist, former Clin. Asst. Prof. @ OHSU
    Excellent summary of testing available up to now. Brings together a lot of useful information. I sincerely hope that we can look forward to the authors updating this survey with information about antigen testing, which, I am reading in today's newspaper, was just given emergency approval by the FDA.
    CONFLICT OF INTEREST: None Reported
    Prognostic Value of Antibody-Relevance
    Liz Jenny-Avital, MD | Jacobi Medical Center
    I would love to see an analysis of Ab (even neutralizing Ab) among those who survive and those who die. Neutralizing Ab may not be protective, at least not in patients already sick enough to be hospitalized. Such an analysis might better inform our current belief in the value of convalescent plasma (CP). I believe that checking Ab in sick patients admitted to hospital might temper the enthusiasm for use of CP.

    That said, I think Ab may be useful for diagnosing some later COVID-associated morbidity (eg clot) when nasopharyngeal PCR is negative.
    CONFLICT OF INTEREST: None Reported
    Synchronous Seroconversion of IgG and IgM
    Vedat Aslan, MD | Antalya Education and Research Hospital
    If antigen on the surface T cell receptor matches the B cells' MHC receptor, it triggers proliferation and antibody production.

    Some B cells that are approved by T cells turn to short-life plasma cells that produce only IgM antibodies. This takes a short time. IgM antibodies are less specific and have low affinity. They can not pass through normal tissues.

    Some of these B cells go to germinal centers of lymphoid tissues. After somatic hypermutation and class switch, they turn to long-lived plasma cells and memory cells that produce IgA, IgG or IgE. This takes a longer
    time.

    Synchronous seroconversion of IgG and IgM means SARS-CoV-2 is not a new virus for the patient because the IgG response needs about two weeks in response to new antigens.

    There are two possibilities:

    1: SARS or another coronaviruses' surface antigens are the same as SARS-CoV-2 and so the antibody response is the same. Most Chinese people are immune to most of the coronaviruses. They can carry the virus but do not develop serious pneumonia more than European and American people.

    2: The antibody test used for diagnosis has cross reactivity to other coronaviruses.
    CONFLICT OF INTEREST: None Reported
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    Humoral Response in Asymptomatic Patients
    Bader Alwadaany, MSN | Kuwait PAAET, college of Nursing
    As it is still empirically baffling to pinpoint the factors influencing display of COVID 19 symptoms it would be informative to examine seroconversion for IgG, A and M and their titre levels in asymptomatic patients.
    CONFLICT OF INTEREST: None Reported
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