key: cord-0292887-leuz6ouy authors: Suzuki, H.; Akashi, Y.; Kato, D.; Takeuchi, Y.; Kiyasu, Y.; Terada, N.; Kurihara, Y.; Kuwahara, M.; Muramatsu, S.; Ueda, A.; Notake, S.; Nakamura, K. title: Analytical performance of rapid antigen tests for the detection of SARS-CoV-2 during widespread circulation of the Omicron variant date: 2022-05-19 journal: nan DOI: 10.1101/2022.05.17.22275034 sha: e49cd7b4aaca684f34ae5c6fbb81ffd7361d9961 doc_id: 292887 cord_uid: leuz6ouy Introduction: Antigen testing is essential in the clinical management of COVID-19. However, most evaluations of antigen tests have been performed before the emergence of the Omicron variant. Thus, an assessment of the diagnostic performance of antigen tests for the detection of SARS-CoV-2 during the circulation of Omicron variant is required. Methods: This prospective observational study evaluated QuickNavi-COVID19 Ag, a rapid antigen detection test between December 2021 and February 2022 in Japan, using real-time reverse transcription (RT)-PCR as a reference. Two nasopharyngeal samples were simultaneously collected for antigen testing and for RT-PCR. Variant analysis of the SARS-CoV-2 genomic sequencing was also performed. Results: In total, nasopharyngeal samples were collected from 1,073 participants (417 positive; 919 symptomatic; 154 asymptomatic) for analysis. Compared with those of RT-PCR, the sensitivity, specificity, positive predictive value, and negative predictive value were 94.2% (95% CI: 91.6%-96.3%), 99.5% (95% CI: 98.7%-99.9%), 99.2% (95% CI: 97.8%-99.8%), and 96.5% (95% CI: 94.8%-97.7%), respectively. The sensitivity among symptomatic individuals was 94.3% (95% CI: 91.5%-96.4%). Overall, 85.9% of sequences were classified as Omicron sublineage BA.1, 12.4% were Omicron sublineage BA.2, and 1.6% were Delta B.1.617.2. (Delta variant). Most of the samples (87.1%) had Ct values <25. Conclusions: The QuickNavi-COVID19 Ag test showed high diagnostic performance for the detection of the SARS-CoV-2 Omicron sublineages BA.1 and BA.2 from nasopharyngeal samples. . CC-BY-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. The copyright holder for this preprint this version posted https://doi.org/10.1101/2022.05.17.22275034 doi: medRxiv preprint . CC-BY-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. The copyright holder for this preprint this version posted https://doi.org/10.1101/2022.05.17.22275034 doi: medRxiv preprint The emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) B.1.1.529, i.e., the Omicron variant, dramatically increased the clinical cases of coronavirus disease 2019 . Owing to its high transmissibility, short incubation period [1] [2] [3] [4] and reduced vaccine efficacy [5, 6] , the Omicron variant sublineages BA. 1 [11, 12] . . CC-BY-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (which was not certified by peer review) Herein, we prospectively evaluated the diagnostic performance of a qualitative antigen test (QuickNavi-COVID19 Ag, Denka Co., Ltd., Tokyo, Japan) using nasopharyngeal swab samples. We also conducted a genomic sequencing analysis to identify SARS-CoV-2 variants. This study was conducted between December 28, 2021 and February 16, 2022. Sample collection and antigen testing were performed at a drive-through sample collection point at Tsukuba Medical Center Hospital (TMCH), and PCR was performed in the TMCH microbiology department. TMCH provides SARS-CoV-2 testing for the Tsukuba district in Japan. People with and without symptoms were referred from 65 clinics and a local public health center. All asymptomatic individuals had a history of contact with a confirmed or suspected COVID-19 cases. Informed consent was verbally obtained from all participants and was documented in their electronic medical record to prevent infection transmission, written informed consent was not obtained. The ethics board of the University of Tsukuba Hospital approved the study (approval number: R03-042), including the method of obtaining informed consent. . CC-BY-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (which was not certified by peer review) Two nasopharyngeal samples were separately collected by medical professionals one for RT-PCR and the other for antigen testing, as previously described [13] [14] [15] [16] [17] [18] [19] [20] . A nasopharyngeal sample was obtained from each nasal cavity. All antigen tests were immediately performed on site after sample collection. A swab was inserted into a specimen buffer tube, and three drops of the prepared specimen were added on the test device. The sample processing time was 8 minutes, and the result was analyzed visually by the personnel who collected the sample. For RT-PCR, a swab was diluted in 3 mL of Universal Transport Medium (Copan Italia S.p.A., Brescia, Italy) on site, and the sample was transferred to the TMCH microbiology department for in-house RT-PCR testing [13, 20] . A 200 μL aliquot of each nasopharyngeal sample was extracted with a magLEAD 6gC (Precision System Science Co., Ltd., Chiba, Japan), and 100 μL of purified sample was eluted. The eluted samples were transferred to Denka Co., Ltd. For reference real-time RT-PCR testing to identify SARS-CoV-2, we used a method developed by the National Institute of Infectious Diseases (NIID), Japan. This method used an Applied Biosystems QuantStudio 3 (Thermo Fisher Scientific Inc., Waltham, MA, USA) with a QuantiTect probe RT-PCR kit (Qiagen Inc., Germantown, MD, USA) and a primer/probe N and N2 set [22] . Until the evaluation, all samples were preserved at −80 °C. . CC-BY-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. The copyright holder for this preprint this version posted May 19, 2022. ; https://doi.org/10.1101/2022.05. 17.22275034 doi: medRxiv preprint In case of discrepancy between the in-house PCR and the NIID method results for the presence or absence of SARS-CoV-2, additional examinations with the Xpert Xpress SARS-CoV-2 and GeneXpert system (Cepheid, Sunnyvale, CA, USA) [23] were performed, and those results were used as the final judgment. Sequencing results were then analyzed for SARS-CoV-2 variants using Illumina's DRAGEN COVIDSeq Test Pipeline. . CC-BY-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. The copyright holder for this preprint this version posted The sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) of the antigen tests were calculated with 95% confidence intervals (CIs). The sensitivity stratified by the cycle threshold (Ct) value based on the N2 set of the NIID method was also evaluated. All statistical analyses were conducted using R 4.1.2 software (R Foundation, Vienna, Austria) with the "readxl," "tidyverse," and "epiR" packages. . CC-BY-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (which was not certified by peer review) Nasopharyngeal samples were collected from 1,073 participants during the study period; 919 were from symptomatic individuals and 154 were from asymptomatic individuals. For symptomatic participants, the median duration from symptom onset to sample collection was 2 days (interquartile range: 1-3 days). Of the 1,073 samples, 411 were SARS-CoV-2 positive by RT-PCR with the NIID method. There were six discordances between the NIID and in-house RT-PCR results. Of the six discordant samples, all were SARS-CoV-2 positive when analyzed by the GeneXpert® system. In this study, more than 90% of individuals who were PCR positive for SARS-CoV-2 were correctly identified by rapid antigen testing, which included both symptomatic and asymptomatic individuals. However, the small number of asymptomatic participants might . CC-BY-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. The copyright holder for this preprint this version posted Bayart also showed that six antigen tests had significantly decreased sensitivities for samples with low viral loads of Omicron variant [11] . Meanwhile, the BinaxNOW (Abbott Diagnostics Scarborough Inc., ME, USA) rapid antigen test was shown to detect 95.2% (95% CI: 91%-98%) of samples with RT-PCR Ct values <30 and 82.1% (95% CI: 77%-87%) of those with Ct values <35 during an Omicron surge period [24] . In a 2020-2021 clinical study, we evaluated the QuickNavi-COVID19 Ag test using 1,934 samples. The test sensitivity was 89.3% (95% CI: 82%-94%) for symptomatic individuals and 67.1% (95% CI: 55%-78%) for asymptomatic individuals, and no false positives were observed [15] . A re-evaluation of the QuickNavi-COVID19 Ag test was performed in 1,510 cases during the Delta variant dominantly circulating period in 2021, and the test sensitivity was 88.3% (95% CI: 83%-93%) for symptomatic individuals and 69.4% (95% CI: 60%-78%) . CC-BY-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. The copyright holder for this preprint this version posted https://doi.org/10.1101/2022.05.17.22275034 doi: medRxiv preprint for asymptomatic individuals. Three false-positive tests (0.2%) were identified (20) . The current study found a slightly better clinical performance for the QuickNavi-COVID19 Ag test during the widespread Omicron circulating period. The improvement could be owing to the higher proportions of symptomatic individuals and people with high viral loads. The Ct values of people who are infected with the Omicron variant are considered to be nearly the same as for previous variants; thus, it is unclear why a high proportion of participants had a high viral load [25] . An increased false-positive rate has been reported for several lots of the QuickNavi-COVID19 Ag test due to inappropriate materials [26] ; this problem was identified several months after production of the test kits. In this study, we performed 1,200 tests (eight lots) including the lots that were within 3 months of the expiration date, and no false positives were observed. In addition, there were only three false-positive samples identified during the Omicron variant period evaluation. Swabs for the antigen test and RT-PCR were obtained separately, and the sample collection procedure for the reference RT-PCR test may have been inappropriate. The flawed procedure may have caused false negatives, which was observed in a previous evaluation [20] . Clinicians should be aware that defective products are a possibility, especially if there are many positive results during a low-prevalence period. . CC-BY-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. The copyright holder for this preprint this version posted This study has some limitations. First, the samples were collected at one site in Japan, and most samples were collected soon after symptom onset. The sample size for asymptomatic individuals might have been insufficient. Second, the assessment of lateral flow device results can vary among examiners [27] . Third, the reference RT-PCR examinations were performed with frozen samples, and the storage and transportation processes may have affected the test results. In addition, study samples were collected from the nasopharyngeal tract, and anterior nasal samples were not analyzed. In conclusion, the current study showed that the QuickNavi-COVID19 Ag test had a high diagnostic performance for the detection of SARS-CoV-2 Omicron sublineages BA.1 and BA.2 in nasopharyngeal samples. . CC-BY-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. The copyright holder for this preprint this version posted . CC-BY-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. The copyright holder for this preprint this version posted https://doi.org/10.1101/2022.05.17.22275034 doi: medRxiv preprint . CC-BY-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. The copyright holder for this preprint this version posted https://www.who.int/publications/i/item/antigen-detection-in-the-diagnosis-of-sars-. CC-BY-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (which was not certified by peer review) . CC-BY-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (which was not certified by peer review) The copyright holder for this preprint this version posted Proportions of SARS-CoV-2 variants by genomic sequencing analysis . CC-BY-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (which was not certified by peer review) The copyright holder for this preprint this version posted May 19, 2022. ; https://doi.org/10.1101/2022.05.17.22275034 doi: medRxiv preprint Shorter serial intervals in SARS-CoV-2 cases with Omicron BA.1 variant compared with Delta variant, the Netherlands Rapid spread of SARS-CoV-2 Omicron subvariant BA.2 in a single-source community outbreak Time from Exposure to Diagnosis among Quarantined Close Contacts of SARS-CoV-2 Omicron Variant Index Case-Patients Rapid epidemic expansion of the SARS-CoV-2 Omicron variant in southern Africa Covid-19 Vaccine Effectiveness against the Omicron (B.1.1.529) Variant Analytical Sensitivity of Six SARS-CoV-2 Rapid Antigen Tests for Omicron versus Delta Variant Impaired detection of omicron by SARS-CoV-2 rapid antigen tests A Prospective Evaluation of the Analytical Performance of GENECUBE((R)) HQ SARS-CoV-2 and GENECUBE((R)) FLU A/B Clinical evaluation of the rapid nucleic acid amplification point-of-care test (Smart Gene SARS-CoV-2) in the analysis of nasopharyngeal and anterior nasal samples Prospective analytical performance evaluation of the QuickNavi-COVID19 Ag for asymptomatic individuals International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (which was not certified by peer review) The copyright holder for this preprint this version posted The evaluation of a novel digital immunochromatographic assay with silver amplification to detect SARS-CoV-2 How to Obtain a Nasopharyngeal Swab Specimen Diagnostic performance of a novel digital immunoassay (RapidTesta SARS-CoV-2): A prospective observational study with nasopharyngeal samples The evaluation of a newly developed antigen test (QuickNavi-COVID19 Ag) for SARS-CoV-2: A prospective observational study in Japan A prospective evaluation of diagnostic performance of a combo rapid antigen test QuickNavi-Flu+COVID19 Ag The evaluation of the utility of the GENECUBE HQ SARS-CoV-2 for anterior nasal . CC-BY-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (which was not certified by peer review) The copyright holder for this preprint this version posted Development of Genetic Diagnostic Methods for Detection for Novel Coronavirus Multicenter Evaluation of the Cepheid Xpert Xpress SARS-CoV-2 Test Comparison of SARS-CoV-2 Reverse Transcriptase Polymerase Chain Reaction and BinaxNOW Rapid Antigen Tests at a Community Site During an Omicron Surge : A Cross-Sectional Study The SARS-CoV-2 Omicron Variant Does Not Have Higher Nasal Viral Loads Compared to the Delta Variant in Symptomatic and Asymptomatic Individuals . CC-BY-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (which was not certified by peer review) . CC-BY-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. The copyright holder for this preprint this version posted May 19, 2022. ; https://doi.org/10.1101/2022.05.17.22275034 doi: medRxiv preprint