Canadian Society of Clinical Chemists (CSCC) consensus guidance for testing,selection and quality management of SARS-CoV-2 point-of-care tests

ObjectivesA consensus guidance is provided for testing, utility and verification of SARS-CoV-2 point-of-care test (POCT) performance and implementation of a quality management program, focusing on nucleic acid and antigen targeted technologies.Design and MethodsThe recommendations are based on current literature and expert opinion from the members of Canadian Society of Clinical Chemists (CSCC), and are intended for use inside or outside of healthcare settings that have varied levels of expertise and experience with POCT.Results and ConclusionsHere we discuss sampling requirements, biosafety, SARS-CoV-2 point-of-care testing methodologies (with focus on Health Canada approved tests), test performance and limitations, test selection, testing utility, development and implementation of quality management systems, quality improvement, and medical and scientific oversight.     

Viral load may impact the diagnostic performance of nasal swabs in nucleic acid amplification test and quantitative antigen test for SARS-CoV-2 detection

IntroductionCompared to nasopharyngeal swabs (NPS), there has been insufficient evaluation of the diagnostic performance of nasal swabs (NS) for the detection of severe acute respiratory coronavirus 2 (SARS-CoV-2) in the nucleic acid amplification test (NAAT) and quantitative SARS-CoV-2 antigen test (QAT).MethodsWe prospectively compared healthcare worker-collected and flocked NS within nine days after symptom onset to paired NPS to detect SARS-CoV-2 in NAAT and QAT on the fully automated Lumipulse system. The agreement between sample types was evaluated, and cycle threshold (Ct) values and antigen levels were used as surrogate viral load measures.ResultsSixty sets of NPS and NS samples were collected from 40 patients with COVID-19. The overall agreements between NAAT and QAT samples were 76.7% and 65.0%, respectively. In NAAT, the Ct value of NS was significantly higher, 5.9, than that of NPS. Thirty-nine (95.1%) NS tested positive in 41 positive-paired NPS with Ct ≤ 30. The negative correlation was observed between antigen levels of NS in QAT and Ct values of NS in NAAT (r = ?0.88). In QAT, the antigen level of NS was significantly lower than that of NPS. Thirty-six (90.0%) NS tested positive in 40 positive-paired NPS with antigen levels >100 pg/mL, which were collected significantly earlier than those with antigen levels ≤100 pg/mL.ConclusionsIn NAAT and QAT, NS had limited performance in detecting SARS-CoV-2 compared to NPS. However, NS may be helpful for patients with COVID-19 with high viral loads or those in the early stages of the illness.     

Indications for SARS-CoV-2 nucleic acid amplification test for areas with low endemicity

IntroductionThe optimal indication for the nucleic acid amplification test (NAAT) in areas with low endemicity for coronavirus disease 2019 (COVID-19) is unclear. This study aimed to identify patients who should undergo the NAAT for COVID-19 diagnosis.MethodsWe retrospectively analyzed the clinical data of patients with suspected COVID-19 who underwent NAAT between October 5, 2020, and May 31, 2021 in our institution.ResultsA total of 1238 patients were enrolled and NAAT positive results were observed in 40 patients (3.2%). The NAAT positivity rate was 34.3% (23/67) in patients with a history of close contact and 1.5% (17/1171) in patients without a history of close contact. Olfactory/gustatory dysfunction and a history of stay in other prefectures were independent risk factors of COVID-19 in patients without a history of close contact. On the other hand, the NAAT positivity rate was only 0.7% (8/1073) in patients without olfactory/gustatory dysfunction and a history of stay in other prefectures. Among them, the group without respiratory symptoms/sign had only one NAAT-positive case (0.1%: 1/1073).ConclusionsThis study revealed that a history of close contact, olfactory/gustatory dysfunction, and a history of stay in other prefectures are key eligibility criteria for NAAT in areas with relatively few patients with COVID-19. On the other hand, NAAT may not be necessary in cases without all of these factors and respiratory symptoms/sign.     

A study of quality assessment in SARS-CoV-2 pathogen nucleic acid amplification tests performance; from the results of external quality assessment survey of clinical laboratories in the Tokyo Metropolitan Government external quality assessment program in 2020

IntroductionThe Tokyo Metropolitan Government (TMG) conducted an external quality assessment (EQA) survey of pathogen nucleic acid amplification tests (NAATs) as a TMG EQA program for SARS-CoV-2 for clinical laboratories in Tokyo.MethodsWe diluted and prepared a standard product manufactured by Company A to about 2,500 copies/mL to make a positive control and distribute it with a negative control. The participants reported the use of the NAATs methods for SARS-CoV-2, the name of the real-time RT-PCR kit, the name of the detection device, the target gene(s), nucleic acid extraction kit, Threshold Cycle value in the case of RT-PCR and the Threshold time value and Differential calculation value in the case of Loop-Mediated Isothermal Amplification (LAMP) method.ResultsAs a result, 17 laboratories using fully automated equipment and 34 laboratories using the RT-PCR method reported generally appropriate results in this EQA survey. On the other hand, among the laboratories that adopted the LAMP method, there were a plurality of laboratories that judged positive samples to be negative.ConclusionThe false negative result is considered to be due to the fact that the amount of virus genome contained in the quality control reagent used this time was below the detection limit of the LAMP method combined with the rapid extraction reagent for influenza virus. On the other hand, false positive results are considered to be due to the non-specific reaction of the NAATs. The EQA program must be continued for the proper implementation of the pathogen NAATs.     

False positive results in severe acute respiratory coronavirus 2 (SARS-CoV-2) rapid antigen tests for inpatients

Severe acute respiratory syndrome coronavirus 2 rapid antigen detection (RAD) test kits are widely used as primary screening test in Japan because rapid diagnosis of coronavirus disease 2019 (COVID-19) is critical for infection control. We report cases with RAD test false-positive results in a ward for patients with disabilities. RAD tests potentially evoke hospital operational risk. It is desirable that performing PCR test appropriately when patients admitted to a medical treatment ward with COVID-19 symptoms instead of RAD test.     

Diagnostic performance of a novel digital immunoassay (RapidTesta SARS-CoV-2): A prospective observational study with nasopharyngeal samples

IntroductionDigital immunoassays are generally regarded as superior tests for the detection of infectious disease pathogens, but there have been insufficient data concerning SARS-CoV-2 immunoassays.MethodsWe prospectively evaluated a novel digital immunoassay (RapidTesta SARS-CoV-2). Two nasopharyngeal samples were simultaneously collected for antigen tests and Real-time RT-PCR.ResultsDuring the study period, 1127 nasopharyngeal samples (symptomatic patients: 802, asymptomatic patients: 325) were evaluated. For digital immunoassay antigen tests, the sensitivity was 78.3% (95% CI: 67.3%–87.1%) and the specificity was 97.6% (95% CI: 96.5%–98.5%). When technicians visually analyzed the antigen test results, the sensitivity was 71.6% (95% CI: 59.9%–81.5%) and the specificity was 99.2% (95% CI: 98.5%–99.7%). Among symptomatic patients, the sensitivity was 89.4% (95% CI; 76.9%–96.5%) with digital immunoassay antigen tests, and 85.1% (95% CI; 71.7%–93.8%) with visually analyzed the antigen test, respectively.ConclusionsThe sensitivity of digital immunoassay antigen tests was superior to that of visually analyzed antigen tests, but the rate of false-positive results increased with the introduction of a digital immunoassay device.     

Immunochromatographic test for the detection of SARS-CoV-2 in saliva

We evaluated the rapid immunochromatographic test for severe acute respiratory coronavirus 2 (SARS-CoV-2) antigen detection using 16 saliva specimens collected from 6 COVID-19 hospitalized patients, and detected N-antigen in 4 of 7 RT-PCR positive specimens. This POCT detected SARS-CoV-2 antigen in saliva and would be useful for COVID-19 diagnosis.     

Clinical relevance of nucleic acid amplification test for patients with urinary tuberculosis during antituberculosis treatment

The nucleic acid amplification test (NAAT) has been valuable in the diagnosis of urinary tuberculosis; however, no studies have attempted to determine the significance of NAAT post treatment. We encountered three patients with urinary tuberculosis who underwent sequential NAAT during antituberculosis chemotherapy and post treatment. All patients were diagnosed as having urinary tuberculosis by positive NAAT and specific renal deformity revealed by imaging. In two of the three patients, positive culture results were obtained and one was negative in standard culture. During antituberculosis chemotherapy, a negative NAAT was obtained from 3 to 5 months after the start of treatment and no positive culture results were obtained during the same period. At the end of chemotherapy, 6 months or more after the start of medication, all patients had negative NAAT results. These results suggest that NAAT for Mycobacterium tuberculosis provides an effective and rapid detection method for urinary tuberculosis both pre- and post-treatment.     

False-positive for SARS-CoV-2 antigen test in a man with acute HIV infection

Although rapid antigen tests for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is convenient, some articles have demonstrated their low sensitivity indicating false-negative results should always be considered. Here, we raise the issue of false-positive on rapid antigen tests for SARS-CoV-2 with the first case of acute HIV infection who repeatedly positive for the rapid antigen test. A 39-year-old man was admitted to our hospital complaining of high-grade fever, dry cough, general fatigue, and anorexia. The rapid antigen test performed on a nasopharyngeal swab sample was positive, therefore the patient was separated in an isolated room apart from the COVID-19 ward while awaiting the confirmatory RT-PCR result. However, the RT-PCR for SARS-CoV-2 performed on nasopharyngeal swabs was repeatedly negative (three times), while the antigen test was repeatedly positive (three times in total). This patient was eventually diagnosed with acute human immunodeficiency virus (HIV) infection based on a high titer of HIV-RNA and absence of plasma HIV-1/2 antibodies. Physicians should consider the possibility of false-positive results in addition to false-negative results when using a rapid antigen test for SARS-CoV-2, and keep in mind that nucleic acid amplification tests are needed to confirm the diagnosis.     

新型冠状病毒核酸临床检测要点及经验

2019年12月以来,新型冠状病毒感染引起的肺炎病例数快速攀升。因患者体内病毒核酸的存在是目前唯一的确诊依据,而目前临床上普遍反映核酸检测阳性率不够理想,故对病毒核酸检测的实验室条件、检验人员及检测过程中各个环节都有较高要求。本文着重阐述了新型冠状病毒核酸检测各个环节中的检测要点、最新的行业规范及共识内容,同时探讨临床检测中的疑难问题,以期为新型冠状病毒所致疾病的准确诊断提供理论依据。     

A comparison of SARS-CoV-2 nucleocapsid and spike antibody detection using three commercially available automated immunoassays

IntroductionCommercially available serological assays for SARS-CoV-2 detect antibodies to either the nucleocapsid or spike protein. Here we compare the performance of the Beckman-Coulter SARS-CoV-2 spike IgG assay to that of the Abbott SARS-CoV-2 nucleocapsid IgG and Roche Anti-SARS-CoV-2 nucleocapsid total antibody assays. In addition, we document the trend in nucleocapsid and spike antibodies in sequential samples collected from convalescent plasma donors.MethodsPlasma or serum samples from 20 individual SARS-CoV-2 RT-PCR-positive inpatients (n = 172), 20 individual convalescent donors with a previous RT-PCR-confirmed SARS-CoV-2 infection (n = 20), were deemed positive SARS-CoV-2 samples. RT-PCR-negative inpatients (n = 24), and 109 pre-SARS-CoV-2 samples were determined to be SARS-CoV-2 negative. Samples were assayed by the Abbott, Roche, and Beckman assays.ResultsAll three assays demonstrated 100% specificity. Abbott, Beckman, and Roche platforms had sensitivities of 98%, 93%, and 90% respectively, with the difference in sensitivity attributed primarily to samples from immunocompromised patients. After the exclusion of samples immunocompromised patients, all assays exhibited ≥ 95% sensitivity. In sequential samples collected from the same individuals, the Roche nucleocapsid antibody assay demonstrated continually increasing signal intensity, with maximal values observed at the last time point examined. In contrast, the Beckman spike IgG antibody signal peaked between 14 and 28 days post positive SARS-CoV-2 PCR and steadily declined in subsequent samples. Subsequent collections 51–200 days (median of 139 days) post positive SARS-CoV-2 RT-PCR from five inpatients and five convalescent donors revealed that spike and nucleocapsid antibodies remained detectable for several months after confirmed infection.ConclusionsThe three assays are sensitive and specific for SARS-CoV-2 antibodies. Nucleocapsid and spike antibodies were detectable for up to 200 days post-positive SARS-CoV-2 PCR but demonstrated markedly different trends in signal intensity.