key: cord-264261-98h1bmb2
authors: Caruana, Giorgia; Croxatto, Antony; Coste, Alix T.; Opota, Onya; Lamoth, Frederic; Jaton, Katia; Greub, Gilbert
title: Diagnostic strategies for SARS-CoV-2 infection and interpretation of microbiological results
date: 2020-06-25
journal: Clin Microbiol Infect
DOI: 10.1016/j.cmi.2020.06.019
sha: 
doc_id: 264261
cord_uid: 98h1bmb2

BACKGROUND: To face the current COVID-19 pandemic, diagnostic tools are essential. It is recommended to use real-time RT-PCR for RNA viruses in order (i) to perform a rapid and accurate diagnostic, (ii) to guide patient care and management and (iii) to guide epidemiological strategies. Further studies are warranted to define the role of serological diagnosis and a possible correlation between serological response and prognosis. OBJECTIVES: To guide clinical microbiologists in the use of these diagnostic tests and clinicians in the interpretation of their results. SOURCES: A research of literature was performed through PubMed and Google Scholar using the keywords SARS-CoV-2, SARS-CoV-2 molecular diagnosis, SARS-CoV-2 immune response, SARS-CoV-2 serology/antibody testing, coronavirus diagnosis. CONTENT: The present review discusses performances, limitations and use of current and future diagnostic tests for SARS-CoV-2. IMPLICATIONS: Real-time RT-PCR remains the reference method for diagnosis of SARS-CoV-2 infection. On the other hand, notwithstanding its varying sensitivity according to the time of infection, serology represents a valid asset (i) to try to solve possible discrepancies between a highly suggestive clinical and radiological presentation and negative RT-PCR, (ii) to solve discrepancies between different PCR assays, and (iii) for epidemiological purposes.

In December 2019, numerous cases of pneumonia of unknown etiology were reported in 46 Wuhan (China) [1] . In January, the novel causative virus named SARS-CoV-2 was identified, 47 which spread to other Chinese regions and to other countries, causing a world pandemic [2, 3] . 48 The clinical presentation of this disease, named "coronavirus disease 2019 (COVID-19)", 49 varied from asymptomatic or mild flu-like symptoms to severe bilateral pneumonia with acute 50 respiratory distress and death. A rapid replication of the virus within the first 24 hours from 51 the infection and the relatively high (about 3) reproduction number were described [4] . 52 The available viral genome sequences allowed to soon recognize the close relationship (recombination), thus donating them some genomic plasticity [5] . Furthermore, RNA 59 biosynthesis seems to use a virus-specific template switch, which results in transcription of 60 sub-genomic mRNAs and eventually leading to homologous RNA recombination [5] . 61 Nevertheless, by encoding a 3'-5'exoribonuclease within nonstructural protein 14 (nsp14-62 ExoN), which is required for high-fidelity replication, the mutation capacity of SARS-CoV-2 63 is debated [6] . In the end, somehow the plasticity allowed Coronaviridae to acquire a rich 64 strains biodiversity and the ability to jump species, which already caused previous zoonotic 65 outbreaks, as for MERS-CoV and SARS-CoV [7-9]. 66 Starting from observed similarities in a short region of RdRp gene between SARS-CoV-2 and 67 a bat coronavirus (BatCoVRaTG13), further sequences were identified to be 96% identical at 68 the whole-genome level, corroborating the hypothesis of animals to humans spill-over [10].

As of June 3, more than 6 million cases of COVID-19 have been declared, including more 

RNA extraction methods can generally be classified into i) one-step (with the RT step and the 106 PCR reaction in the same tube) and ii) two-steps RT-PCR (initial creation of DNA copies 107 with RT reaction followed by their addiction to the PCR reaction). Typically, one-step PCR Table 1 ), which can allow to improve sensitivity and 113 TAT but also sometimes increase the costs.

Despite the good performance of the validated NAATs, there is still a risk of false negative 115 results. Most of them concern the pre-analytic setting, such as the timing of the specimen 116 collection (too early or too late in the infection course, including the limit of detection due to 117 late infections with atypical manifestations), the quality of sampling (insufficient material) or 118 type of specimens (bronchoalveolar lavage -BAL exhibits the highest sensitivity, followed by 119 induced sputum, naso-pharyngeal -NP swab, oro-pharyngeal -OP swab, and feces), and finally 120 the sample transport (inappropriate container, exposure to extreme temperatures, etc.) [18] [19] . Sensitivity and specificity of serological assays can also be affected by the target antigen. As 146 highlighted by Meyer and colleagues, the S protein (produced in more advanced stage of 147 SARS-CoV-2 infection) showed lower levels of sensitivity and more specificity (especially 148 the S1 subunit) compared to the N protein, [38] . Noteworthy, we recently observed (A. Coste 149 et al., data not shown) that the antibodies directed against the N protein seems to decrease 150 earlier than the S protein; thus the sensitivity of assays targeting only the N protein may be 151 impaired according to the timing of infection (Figure 1 ). For this reason, we recommend to 152 systematically use two tests, one targeting the S protein and one targeting the N protein for 153 diagnostic purposes. For sero-epidmiological studies, a test targeting the S protein is 154 recommended. The added value to target the S protein is that the titers will likely better reflect 155 protection against reinfection. In Table 2 we summarized the interpretation of diagnostic assays with sensitivity greater than 95% and specificity superior or equal to 98% should be 165 used. As well as for NAATs, different platforms can be considered also for serological tests. Interestingly, the appearance of IgM occurs at the same time than IgG, thus, the main 183 advantage to also test IgM is to assess the timing of the infection. Table 3 . 206 While serological assays can represent a useful epidemiological asset, NAATs remains the 207 gold standard for diagnosis, due to their high sensitivity even at early stages of the disease. Samples like blood and urines were found to be weakly-to-none sensitive, while the virus was 231 also found in feces and perineal swabs of patients with gastro-intestinal symptoms [44] [45] , 

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