A novel strategy for SARS-CoV-2 mass screening with quantitative antigen testing of saliva: a diagnostic accuracy study

Abstract

Background Quantitative RT-PCR (RT-qPCR) of nasopharyngeal swab (NPS) samples for SARS-CoV-2 detection requires medical personnel and is time consuming, and thus is poorly suited to mass screening. In June, 2020, a chemiluminescent enzyme immunoassay (CLEIA; Lumipulse G SARS-CoV-2 Ag kit, Fujirebio, Tokyo, Japan) was developed that can detect SARS-CoV-2 nucleoproteins in NPS or saliva samples within 35 min. In this study, we assessed the utility of CLEIA in mass SARS-CoV-2 screening. MethodsWe did a diagnostic accuracy study to develop a mass-screening strategy for salivary detection of SARS-CoV-2 by CLEIA, enrolling hospitalised patients with clinically confirmed COVID-19, close contacts identified at community health centres, and asymptomatic international arrivals at two airports, all based in Japan. All test participants were enrolled consecutively. We assessed the diagnostic accuracy of CLEIA compared with RT-qPCR, estimated according to concordance (Kendall's coefficient of concordance, W), and sensitivity (probability of CLEIA positivity given RT-qPCR positivity) and specificity (probability of CLEIA negativity given RT-qPCR negativity) for different antigen concentration cutoffs (0•19 pg/mL, 0•67 pg/mL, and 4•00 pg/mL; with samples considered positive if the antigen concentration was equal to or more than the cutoff and negative if it was less than the cutoff). We also assessed a two-step testing strategy post hoc with CLEIA as an initial test, using separate antigen cutoff values for test negativity and positivity from the predefined cutoff values. The proportion of intermediate results requiring secondary RT-qPCR was then quantified assuming prevalence values of RT-qPCR positivity in the overall tested population of 10%, 30%, and 50%. Findings Self-collected saliva was obtained from 2056 participants between June 12 and Aug 6, 2020. Results of CLEIA and RT-qPCR were concordant in 2020 (98•2%) samples (Kendall's W=0•99). Test sensitivity was 85•4% (76 of 89 positive samples; 90% credible interval [CrI] 78•0-90•3) at the cutoff of 0•19 pg/mL; 76•4% (68 of 89; 68•2-82•8) at the cutoff of 0•67 pg/mL; and 52•8% (47 of 89; 44•1-61•3) at the cutoff of 4•0 pg/mL. Test specificity was 91•3% (1796 of 1967 negative samples; 90% CrI 90•2-92•3) at the cutoff of 0•19 pg/mL, 99•2% (1952 of 1967; 98•8-99•5) at the cutoff of 0•67 pg/mL, and 100•0% (1967 of 1967; 99•8-100•0) at the cutoff of 4•00 pg/mL. Using a two-step testing strategy with a CLEIA negativity cutoff of 0•19 pg/mL (to maximise sensitivity) and a CLEIA positivity cutoff of 4•00 pg/mL (to maximise specificity), the proportions of indeterminate results (ie, samples requiring secondary RT-qPCR) would be approximately 11% assuming a prevalence of RT-qPCR positivity of 10%, 16% assuming a prevalence of RT-qPCR positivity of 30%, and 21% assuming a prevalence of RT-qPCR positivity of 50%.Interpretation CLEIA testing of self-collected saliva is simple and provides results quickly, and is thus suitable for mass testing. To improve accuracy, we propose a two-step ...

Reduced Relative Sensitivity of the Elecsys SARS-CoV-2 Antigen Assay in Saliva Compared to Nasopharyngeal Swabs

“…However, its collection requires trained personnel, which are exposed to a potential risk of nosocomial transmission and may create discomfort for the patient. Saliva circumvents these problems, but its sensitivity and thus suitability has been differentially reported [ 2 , 3 , 4 , 5 , 6 , 7 , 8 , 9 , 10 , 11 , 12 , 13 , 14 , 15 , 16 , 17 , 18 , 19 , 20 , 22 , 23 , 24 , 25 , 26 , 27 , 28 , 29 , 30 , 31 , 32 , 33 , 34 , 35 , 36 , 37 , 38 , 39 , 40 , 41 ]. We recently conducted a large-scale head-to-head comparison of SARS-CoV-2 detection by RT-PCR in NPS and saliva and could show that saliva is a valid alternate specimen for SARS-CoV-2 detection by RT-PCR with particular advantages for testing children [ 21 ].…”

“…Our study shows that lower sensitivity can occur in antigen tests using saliva as a medium and highlights the need for developers to establish appropriate protocols for saliva testing. It must be considered that a reduced sensitivity of SARS-CoV-2 antigen testing in saliva is likely not restricted to the assay evaluated here, as several studies noted lower sensitivity in saliva [ 11 , 12 , 13 , 14 , 15 , 20 , 22 , 36 , 37 , 38 , 39 , 40 , 41 , 42 , 43 , 44 , 45 ]. Alternate saliva sampling strategies that allow concentrated saliva processing may thus be of advantage in combination with antigen tests to overcome the sensitivity restrictions.…”

“…As we showed, conducting head-to-head comparisons of NPS and saliva for both PCR and antigen test is critical to dissect if matrix effects in saliva exist. As only few studies have assessed saliva in the context of SARS-CoV-2 antigen tests, and those that have, recorded variable results [ 35 , 41 , 42 , 43 , 44 , 45 , 46 , 47 , 48 , 49 , 50 , 51 , 52 , 53 , 54 , 55 , 56 ], it can currently not be excluded that antigen tests generally show a reduced sensitivity across diverse tests systems. Alternatively, these effects may be test-dependent and can be modest enough to allow application in mass testing as a recent study suggests [ 41 ].…”

Comparison of four commercial, automated antigen tests to detect SARS-CoV-2 variants of concern

“…between 91 and 100% [5][6][7][8][9][10][11][12][13][14][15][16]. While assay specificities were frequently found to be relatively high [5][6][7][8][9][10][11][12][13][14][15][16], assay sensitivities according to international and national guidelines for rapid Ag tests in general, requiring positive rate percentages ≥ 80, have frequently not been met [17,18].…”

“…between 91 and 100% [5][6][7][8][9][10][11][12][13][14][15][16]. While assay specificities were frequently found to be relatively high [5][6][7][8][9][10][11][12][13][14][15][16], assay sensitivities according to international and national guidelines for rapid Ag tests in general, requiring positive rate percentages ≥ 80, have frequently not been met [17,18]. Further, the studies published so far did not compare different automated SARS-CoV-2 Ag assays among each other, which will be critical for diagnostic laboratories seeking to implement such assays.…”