key: cord-0713018-s0f9pwgy
authors: Poopalasingam, Nareshkumar; Korenkov, Michael; Ashurov, Artem; Strobel, Janina; Fish, Irina; Hellmich, Martin; Gruell, Henning; Lehmann, Clara; Heger, Eva; Klein, Florian
title: Determining the reliability of rapid SARS-CoV-2 antigen detection in fully vaccinated individuals
date: 2022-02-19
journal: J Clin Virol
DOI: 10.1016/j.jcv.2022.105119
sha: 12b4ee4ebd56711c32606b122e591f8ee3e6ac3f
doc_id: 713018
cord_uid: s0f9pwgy

BACKGROUND: Rapid antigen detection tests (RADT) are commonly used as SARS-CoV-2 diagnostic tests both by medical professionals and laypeople. However, the performance of RADT in vaccinated individuals has not been fully investigated. OBJECTIVES: RT-qPCR and rapid antigen detection testing were performed to evaluate the performance of the Standard Q COVID-19 Ag Test in detecting SARS-CoV-2 breakthrough infections in vaccinated individuals. STUDY DESIGN: Two swab specimens, one for RT-qPCR and one for RADT, were collected from vaccinated individuals in an outpatient clinic. For comparison of RADT performance in vaccinated and unvaccinated individuals, a dataset already published by this group was used as reference. RESULTS: A total of 696 samples were tested with both RT-qPCR and RADT that included 692 (99.4%) samples from vaccinated individuals. Of these, 76 (11.0%) samples were detected SARS-CoV-2 positive by RT-qPCR and 45 (6.5%) samples by the Standard Q COVID-19 Ag test. Stratified by Ct values, sensitivity of the RADT was 100.0%, 94.4% and 81.1% for Ct ≤ 20 (n=18), Ct ≤ 25 (n=36) and Ct ≤ 30 (n=53), respectively. Samples with Ct values ≥ 30 (n=23) were not detected. Overall RADT specificity was 99.7% and symptom status did not affect RADT performance. Notably, RADT detected 4 out of 4 samples of probable Omicron variant infection based on single nucleotide polymorphism analysis. CONCLUSION: Our results show that RADT testing remains a valuable tool in detecting breakthrough infections with high viral RNA loads.

In combating the SARS-CoV-2 pandemic, a variety of measures has been taken to break transmission chains [1] [2] [3] [4] . For example, rapid antigen detection tests (RADT) have been implemented as a public health strategy in numerous countries to screen for SARS-CoV-2 infected individuals [5] [6] [7] [8] . Since the introduction of RADT, multiple publications have shown that antigen tests can reliably detect individuals with high SARS-CoV-2-RNA loads, who pose an increased risk of SARS-CoV-2 transmission (> 6 log 10 copies/ml) [9] [10] [11] [12] .

Several COVID-19 vaccines with high efficacy against severe illness and death have been approved since the end of 2020 [13] [14] [15] [16] [17] [18] . Nevertheless, as vaccine-induced serum antibody levels decline over time [19] [20] [21] , an increasing incidence of breakthrough infections is reported amongst vaccinated individuals [22] [23] [24] .

Therefore, regular screening measures to identify potentially infectious vaccinated individuals are of critical importance. Given the positive impact of immunization on viral clearance [25, 26] , one might expect overall sensitivity of RADT to decline.

However, the question whether RADT performance is indeed affected in vaccinated individuals remains unanswered. Here, we set out to address this question testing 696 individuals in a side-by-side comparison using RADT and RT-qPCR at our infectious disease outpatient clinic.

Routine SARS-CoV-2 testing by RT-qPCR was performed between the 06 th of December 2021 and 13 th of December 2021 in the outpatient clinic of the University Hospital Cologne, Department I of Internal Medicine, Cologne, Germany. Individuals from the general population with symptoms suggestive for COVID-19, or exposure to SARS-CoV-2, as well as hospital staff for routine screening were eligible for testing.

For quality control of RADT performance in vaccinated breakthrough infections, two swab specimens, one from the oro-/nasopharynx for RT-qPCR and one from the nasopharynx for RADT testing, were collected by trained personnel after oral consent. Swabs for RT-qPCR were transferred into virus transport and preservation medium (biocomma ® , Shenzhen, China). The on-site RADT was performed using the second swab. Data on vaccination and symptom status were retrospectively October 2020-January 2021) was used [10] .

The Standard Q COVID-19 Ag Test (SD Biosensor Inc., Suwon-si, Republic of Korea/Hoffmann La Roche AG, Basel, Switzerland) was performed according to the manufacturer's instructions. RADT with any visible test lines were considered positive, if the control line was also present.

Different SARS-CoV-2 RT-qPCR protocols were used for detection of SARS-CoV-2.

In brief, the following methods were used: (i) cobas® SARS-CoV-2 test kit running be found in previous publications [10, 27] .

The presence or absence of spike gene mutations suggestive of the Delta or Omicron variants of SARS-CoV-2 was determined using the VirSNiP L452R and S371L/S373P Mutation Assays (TIB Molbiol). In this probe-based assay, single nucleotide mutations resulting in melting temperature differences are determined by melting curve analysis following PCR amplification. RNA for mutational analysis was This manuscript was submitted to the Institutional Review Board of the University of Cologne which stated that no separate ethics application to and statement of ethical approval by the local ethics committee is required for the retrospective evaluation. 

Since the approval of several COVID-19 vaccines by regulatory authorities [13] [14] [15] , more than 10 billion doses have been administered to date [28, 29] . Due to the efficacious immune response in vaccinated individuals [16] [17] [18] , discussions have emerged whether previously described performance data of RADT are still valid [9] [10] [11] [12] . Although vaccine-induced immunity has been associated with expedited viral clearance, analyses of viral kinetics have described peak viral RNA loads to be similar in vaccinated and unvaccinated individuals [25, [30] [31] [32] . This suggests the performance of the lateral flow immunochromatographic assay to be unaffected by vaccination status in high viral RNA load samples.

To investigate the reliability of rapid SARS-CoV-2 antigen detection in vaccinated individuals, we obtained paired samples for RT-qPCR and on-site RADT, of which 687 were eligible for analysis. As identified in prior evaluations, the performance of tested RADT seems to mainly be dependent on the viral RNA load of the investigated samples [9] [10] [11] [12] . The observed difference in RADT performance between symptomatic and asymptomatic cases can, therefore, be explained by higher viral RNA loads in symptomatic individuals, which is consistent with RADT performance in unvaccinated subjects. Comparing collected data from vaccinated participants with our previous findings in an unvaccinated cohort [10] , the Standard Q COVID-19

RADT shows a similar performance for samples with high viral RNA loads. The study is subject to some limitations including a limited sample size as stated above. For the evaluation of test performance in Omicron variant cases, larger investigations are needed. Additionally, for the comparison between vaccinated and unvaccinated subjects, data from unvaccinated individuals was used from a previous publication for which swabs were obtained from the oro-and nasopharynx. Retrieved data on symptoms are limited to symptom status and were not specified in terms of quality or duration. Also, this study was restricted to the Standard Q COVID-19 Ag test and may not be representative for other RADT kits.

In conclusion, rapid antigen testing reliably detected samples with high viral RNA loads including 4 samples of the newly emerged Omicron variant among vaccinated individuals. Although potentially less sensitive for samples with medium (20 -30) Ct values in vaccinated than unvaccinated cases, one might still suspect that subjects 

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NPV, 95.0% (95% CI = 93.1 to

NPV, 94.6% (95% CI = 90

☒ The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.☐The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: