key: cord-0812466-ek27vihk
authors: Xu, Yuzhong; Cheng, Minggang; Chen, Xinchun; Zhu, Jialou
title: Current approach in laboratory testing for SARS-CoV-2
date: 2020-08-21
journal: Int J Infect Dis
DOI: 10.1016/j.ijid.2020.08.041
sha: 541d088366175ea9e1cf03b9bb2aa99cdf685257
doc_id: 812466
cord_uid: ek27vihk

An outbreak of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) that began in Wuhan, Hubei Province of China, has rapidly spread to produce a global pandemic. It is now clear that person-to-person transmission of SARS-CoV-2 has been occurring and that the epidemic has been dramatically growing in recent months. Early, rapid and accurate diagnosis is of great significance for curtailing the spread of SARS-CoV-2. At present, several diagnostic techniques, e.g., viral culture, nucleic acid amplification test, have been used to detect the virus. However, the sensitivity and specificity of these methods are quite different, with the sample source and detection limit varying greatly. Here, we review all the types and characteristics of the current laboratory diagnostic assays available for detecting SARS-CoV-2 infection and summarize the selection strategies of testing and sampling sites at different disease stages to improve the diagnostic accuracy of Coronavirus Disease 2019 (COVID-19).

significance for curtailing the spread of SARS-CoV-2. At present, several diagnostic techniques, 23 e.g., viral culture, nucleic acid amplification test, have been used to detect the virus. However, the 24 sensitivity and specificity of these methods are quite different, with the sample source and detection importance. Here, we summarize advances made in technologies for rapid diagnosis and 40 confirmation of respiratory infections caused by SARS-CoV-2, as well as the selection strategies of 41 testing and sampling sites in SARS-CoV-2 detection. 42

Since the initial cases with pneumonia of unknown cause reported, viral culture and genetic 43 sequencing of isolates obtained from patients with pneumonia identified a novel coronavirus as the 44 etiology within 10 days in January 2020, benefiting our understanding of disease occurrence and 45 transmission, as well as diagnostic test development (1) . Although viral culture is relatively time-46 consuming and labor-intensive, it's much more useful in the initial phase of emerging epidemics 47 before other diagnostic assays are clinically available. Besides, unbiased, high-throughput 48 sequencing has been proved as a powerful tool for the discovery of pathogens (Table 1) . A BGI's 49 detection assay, based on next-generation sequencing, was approved emergency use authorization 50 (EUA) by National Medical Products Administration (NMPA) in China (Table S1 ). However, whole 51 genome sequencing is time-consuming and requires specialized instruments with high technical 52 thresholds, and thus is not recommended for widespread use in clinical. 53

In acute respiratory infection, real-time RT-PCR is routinely used to detect causative viruses from 54 respiratory specimens. WHO recommends that patients who meet the case definition for suspected 55 SARS-CoV-2 should be screened for the virus nucleic acid amplification test (Table 1) . Various real-56 time RT-PCR assays for the detection of SARS-CoV-2 RNA have been developed worldwide, with 57 different viral genes or regions targeted (Table S1 ). To date, 13 and 52 commercial SARS-CoV-2 58 real-time RT-PCR diagnostic panels have been issued emergency use authorization (EUA) by China 59 and the US, respectively, with the limit of detection (LoD) varying from 100 to 1000 copies/ml 60 (Table S1 ). Although its relatively high sensitivity of RT-PCR, there have been reports of multiple 61 false negative tests for the same patients infected with SARS-CoV-2 in China (2, 3), suggesting that fluctuating RT-PCR results were observed in several patients that the clinical specimens tested 64 positive for SARS-CoV-2 at first, then turned negative in the following test. But in the final, the 65 results returned to be positive (4). False negative results may be due to the selection of sampling 66 locations, poor sample quality, low viral load of the specimen, incorrect storage, and transportation, 67 as well as laboratory testing conditions and personnel operations. If a highly suspected patient was 68 detected negative for the virus, repeat the nucleic acid amplification test, or consider collecting a 69 more suitable sample. 70

Isothermal amplification techniques offer a substitutable choice to real-time RT-PCR with 71 comparable performance (Table 1) (Table S1) . 82

Serological assays provide an alternative diagnosis approach for the current rapidly growing 83 demand for rapid diagnosis of suspected patients and asymptomatic infections that the entire test 84 could be completed in a short time, even independent of specific equipment or places. They are 85 suggested to be used either in combination with molecular testing or for additional testing in 86 suspected cases with negative nucleic acid results to improve the detection accuracy of COVID-19. 87

In a study of 397 real-time RT-PCR confirmed COVID-19 patients and 128 virus-negative patients, 88

IgM/IgG assays showed a sensitivity and specificity of 88.66% and 90.63% in blood samples, 89 respectively (7). Combined IgM-IgG tests provided better sensitivity than tests for only IgM or IgG. 90

However, cross-reactivity of the serological assay to other coronaviruses has been observed (8). 91

Besides, serological testing is critically useful in disease surveillance and epidemiologic research. 92 SARS-CoV-2 was 4.65% in Los Angeles County (9). 367,000 people were estimated to be infected 94 with SARS-CoV-2, which is 43.53 times higher than the cumulative number ( Lastly, the selection of specimens for molecular assays is crucial in the laboratory diagnosis of 106 SARS-CoV-2 (Table 2) . To prevent misdiagnosis caused by insufficient viral load, BALF is the most 107 preferred specimen as the viral loads of respiratory tract specimens are highest in BALF, followed 108 

The New England journal of medicine

Chest CT for Typical 2019-nCoV Pneumonia: 136 Relationship to Negative RT-PCR Testing

False-negative of RT-PCR and prolonged nucleic acid conversion in 139 COVID-19: Rather than recurrence

Stability issues of RT-PCR testing of SARS-CoV-2 for 142 hospitalized patients clinically diagnosed with COVID-19

Rapid and visual detection of 2019 novel 145 coronavirus (SARS-CoV-2) by a reverse transcription loop-mediated isothermal amplification assay

Development of a reverse transcription-149 loop-mediated isothermal amplification as a rapid early-detection method for novel SARS-CoV-2

Development and clinical application of a rapid IgM-IgG 152 combined antibody test for SARS-CoV-2 infection diagnosis

Profiling Early Humoral Response to Diagnose 155

Clinical infectious diseases : an official publication of the 156

Infectious Diseases Society of America

Seroprevalence of SARS-CoV-2-PMC5527409

Detection of SARS-CoV-2 in Different Types of Clinical 165

Evaluating the accuracy of different respiratory 168 specimens in the laboratory diagnosis and monitoring the viral shedding of 2019-nCoV infections

Prolonged presence of SARS-CoV-2 viral RNA in 171 faecal samples

J o u r n a l P r e -p r o o f 9 Fecal testing for SARS-CoV-2 is highly recommended after viral clearance in the respiratory samples. * All patients were confirmed by SARS-CoV-2 detection (11, 12)