Antigen rapid tests, nasopharyngeal PCR and saliva PCR to detect SARS-CoV-2: A prospective comparative clinical trial.

BACKGROUND: Nasopharyngeal antigen Rapid Diagnostic Tests (RDTs), saliva RT-PCR and nasopharyngeal (NP) RT-PCR have shown different performance characteristics to detect patients infected by SARS-CoV-2, according to the viral load (VL)-and thus transmissibility. METHODS: In October 2020, we conducted a prospective trial involving patients presenting at testing centres with symptoms of COVID-19. We compared detection rates and performance of RDT, saliva PCR and nasopharyngeal (NP) PCR, according to VL and symptoms duration. RESULTS: Out of 949 patients enrolled, 928 patients had all three tests performed. Detection rates were 35.2% (95%CI 32.2-38.4%) by RDT, 39.8% (36.6-43.0%) by saliva PCR, 40.1% (36.9-43.3%) by NP PCR, and 41.5% (38.3-44.7%) by any test. For those with viral loads (VL) ≥106 copies/ml, detection rates were 30.3% (27.3-33.3), 31.4% (28.4-34.5), 31.5% (28.5-34.6), and 31.6% (28.6-34.7%) respectively. Sensitivity of RDT compared to NP PCR was 87.4% (83.6-90.6%) for all positive patients, 94.5% (91.5-96.7%) for those with VL≥105 and 96.5% (93.6-98.3%) for those with VL≥106. Sensitivity of STANDARD-Q®, Panbio™ and COVID-VIRO® Ag tests were 92.9% (86.4-96.9%), 86.1% (78.6-91.7%) and 84.1% (76.9-89.7%), respectively. For those with VL≥106, sensitivity was 96.6% (90.5-99.3%), 97.8% (92.1-99.7%) and 95.3% (89.4-98.5%) respectively. No patient with VL<104 was detected by RDT. Specificity of RDT was 100% (99.3-100%) compared to any PCR. RDT sensitivity was similar <4 days (87.8%, 83.5-91.3%) and ≥4 days (85.7%, 75.9-92.6%) after symptoms onset (p = 0.6). Sensitivity of saliva and NP PCR were 95.7% (93.1-97.5%) and 96.5% (94.1-98.1%), respectively, compared to the other PCR. CONCLUSIONS: RDT results allow rapid identification of COVID cases with immediate isolation of most contagious individuals. RDT can thus be a game changer both in ambulatory care and community testing aimed at stopping transmission chains, and even more so in resource-constrained settings thanks to its very low price. When PCR is performed, saliva could replace NP swabbing. TRIAL REGISTRATION: ClinicalTrial.gov Identifier: NCT04613310 (03/11/2020).

An Automated Dashboard to Improve Laboratory COVID-19 Diagnostics Management.

Background: In response to the COVID-19 pandemic, our microbial diagnostic laboratory located in a university hospital has implemented several distinct SARS-CoV-2 RT-PCR systems in a very short time. More than 148,000 tests have been performed over 12 months, which represents about 405 tests per day, with peaks to more than 1,500 tests per days during the second wave. This was only possible thanks to automation and digitalization, to allow high throughput, acceptable time to results and to maintain test reliability. An automated dashboard was developed to give access to Key Performance Indicators (KPIs) to improve laboratory operational management. Methods: RT-PCR data extraction of four respiratory viruses-SARS-CoV-2, influenza A and B and RSV-from our laboratory information system (LIS), was automated. This included age, gender, test result, RT-PCR instrument, sample type, reception time, requester, and hospitalization status etc. Important KPIs were identified and the visualization was achieved using an in-house dashboard based on the open-source language R (Shiny). Results: The dashboard is organized into three main parts. The "Filter" page presents all the KPIs, divided into five sections: (i) general and gender-related indicators, (ii) number of tests and positivity rate, (iii) cycle threshold and viral load, (iv) test durations, and (v) not valid results. Filtering allows to select a given period, a dedicated instrument, a given specimen, an age range or a requester. The "Comparison" page allows a custom charting of all the available variables, which represents more than 182 combination. The "Data" page, gives the user an access to the raw data in tables format, with possibility of filtering, allowing for a deeper analysis and data download. Informations are updated every 4 h. Conclusions: By giving a rapid access to a huge number of up-to-date information, represented using the most relevant visualization types, without the burden of timely data extraction and analysis, the dashboard represents a reliable and user-friendly tool for operational laboratory management. The dashboard represents a reliable and user-friendly tool improving the decision-making process, resource planning and quality management.

Computer-Aided Medical Microbiology Monitoring Tool: A Strategy to Adapt to the SARS-CoV-2 Epidemic and That Highlights RT-PCR Consistency.

Since the beginning of the COVID-19 pandemic, important health and regulatory decisions relied on SARS-CoV-2 reverse transcription polymerase chain reaction (RT-PCR) results. Our diagnostic laboratory faced a rapid increase in the number of SARS-CoV-2 RT-PCR. To maintain a rapid turnaround time, we moved from a case-by-case validation of RT-PCR results to an automated validation and immediate results transmission to clinicians. A quality-monitoring tool based on a homemade algorithm coded in R was developed, to preserve high quality and to track aberrant results. We present the results of this quality-monitoring tool applied to 35,137 RT-PCR results. Patients tested several times led to 4,939 pairwise comparisons: 88% concordant and 12% discrepant. The algorithm automatically solved 428 out of 573 discrepancies. The most likely explanation for these 573 discrepancies was related for 44.9% of the situations to the clinical evolution of the disease, 27.9% to preanalytical factors, and 25.3% to stochasticity of the assay. Finally, 11 discrepant results could not be explained, including 8 for which clinical data was not available. For patients repeatedly tested on the same day, the second result confirmed a first negative or positive result in 99.2% or 88.9% of cases, respectively. The implemented quality-monitoring strategy allowed to: i) assist the investigation of discrepant results ii) focus the attention of medical microbiologists onto results requiring a specific expertise and iii) maintain an acceptable turnaround time. This work highlights the high RT-PCR consistency for the detection of SARS-CoV-2 and the necessity for automated processes to handle a huge number of microbiological results while preserving quality.

Performance of RT-PCR on Saliva Specimens Compared With Nasopharyngeal Swabs for the Detection of SARS-CoV-2 in Children: A Prospective Comparative Clinical Trial.

BACKGROUND: Saliva reverse transcriptase-Polymerase chain reaction (RT-PCR) is an attractive alternative for the detection of severe acute respiratory syndrome coronavirus 2 in adults with less known in children. METHODS: Children with coronavirus disease 2019 symptoms were prospectively enrolled in a 1-month comparative clinical trial of saliva and nasopharyngeal (NP) RT-PCR. Detection rates and sensitivities of saliva and NP RT-PCR were compared as well as discordant NP and saliva RT-PCR findings including viral loads (VLs). RESULTS: Of 405 patients enrolled, 397 patients had 2 tests performed. Mean age was 12.7 years (range, 1.2-17.9). Sensitivity of saliva was 85.2% (95% confidence interval: 78.2%-92.1%) when using NP as the standard; sensitivity of NP was 94.5% (89.8%-99.2%) when saliva was considered as the standard. For a NP RT-PCR VL threshold of ≥103 and ≥104 copies/mL, sensitivity of saliva increases to 88.7% and 95.2%, respectively. Sensitivity of saliva and NP swabs was, respectively, 89.5% and 95.3% in patient with symptoms less than 4 days (P = 0.249) and 70.0% and 95.0% in those with symptoms ≥4-7 days (P = 0.096). The 15 patients who had an isolated positive NP RT-PCR were younger (P = 0.034), had lower NP VL (median 5.6 × 103 vs. 3.9 × 107, P < 0.001), and could not drool saliva at the end of the sampling (P = 0.002). VLs were lower with saliva than with NP RT-PCR (median 8.7 cp/mL × 104; interquartile range 1.2 × 104-5.2 × 105; vs. median 4.0 × 107 cp/mL; interquartile range, 8.6 × 105-1 × 108; P < 0.001). CONCLUSIONS: While RT-PCR testing on saliva performed more poorly in younger children and likely after longer duration of symptoms, saliva remains an attractive alternative to NP swabs in children.

Encephalopathies Associated With Severe COVID-19 Present Neurovascular Unit Alterations Without Evidence for Strong Neuroinflammation.

OBJECTIVE: Coronavirus disease (COVID-19) has been associated with a large variety of neurologic disorders. However, the mechanisms underlying these neurologic complications remain elusive. In this study, we aimed at determining whether neurologic symptoms were caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) direct infection or by either systemic or local proinflammatory mediators. METHODS: In this cross-sectional study, we checked for SARS-CoV-2 RNA by quantitative reverse transcription PCR, SARS-CoV-2-specific antibodies, and 49 cytokines/chemokines/growth factors (by Luminex) in the CSF +/- sera of a cohort of 22 COVID-19 patients with neurologic presentation and 55 neurologic control patients (inflammatory neurologic disorder [IND], noninflammatory neurologic disorder, and MS). RESULTS: We detected anti-SARS-CoV-2 immunoglobulin G in patients with severe COVID-19 with signs of intrathecal synthesis for some of them. Of the 4 categories of tested patients, the CSF of IND exhibited the highest level of cytokines, chemokines, and growth factors. By contrast, patients with COVID-19 did not present overall upregulation of inflammatory mediators in the CSF. However, patients with severe COVID-19 (intensive care unit patients) exhibited higher concentrations of CCL2, CXCL8, and vascular endothelium growth factor A (VEGF-A) in the CSF than patients with a milder form of COVID-19. In addition, we could show that intrathecal CXCL8 synthesis was linked to an elevated albumin ratio and correlated with the increase of peripheral inflammation (serum hepatocyte growth factor [HGF] and CXCL10). CONCLUSIONS: Our results do not indicate active replication of SARS-CoV-2 in the CSF or signs of massive inflammation in the CSF compartment but highlight a specific impairment of the neurovascular unit linked to intrathecal production of CXCL8.

Case Report: Stepwise Anti-Inflammatory and Anti-SARS-CoV-2 Effects Following Convalescent Plasma Therapy With Full Clinical Recovery.

In these times of COVID-19 pandemic, concern has been raised about the potential effects of SARS-CoV-2 infection on immunocompromised patients, particularly on those receiving B-cell depleting agents and having therefore a severely depressed humoral response. Convalescent plasma can be a therapeutic option for these patients. Understanding the underlying mechanisms of convalescent plasma is crucial to optimize such therapeutic approach. Here, we describe a COVID-19 patient who was deeply immunosuppressed following rituximab (anti-CD20 monoclonal antibody) and concomitant chemotherapy for chronic lymphoid leukemia. His long-term severe T and B cell lymphopenia allowed to evaluate the treatment effects of convalescent plasma. Therapeutic outcome was monitored at the clinical, biological and radiological level. Moreover, anti-SARS-CoV-2 antibody titers (IgM, IgG and IgA) and neutralizing activity were assessed over time before and after plasma transfusions, alongside to SARS-CoV-2 RNA quantification and virus isolation from the upper respiratory tract. Already after the first cycle of plasma transfusion, the patient experienced rapid improvement of pneumonia, inflammation and blood cell counts, which may be related to the immunomodulatory properties of plasma. Subsequently, the cumulative increase in anti-SARS-CoV-2 neutralizing antibodies due to the three additional plasma transfusions was associated with progressive and finally complete viral clearance, resulting in full clinical recovery. In this case-report, administration of convalescent plasma revealed a stepwise effect with an initial and rapid anti-inflammatory activity followed by the progressive SARS-CoV-2 clearance. These data have potential implications for a more extended use of convalescent plasma and future monoclonal antibodies in the treatment of immunosuppressed COVID-19 patients.

Impact of different SARS-CoV-2 assays on laboratory turnaround time.

Introduction. Clinical microbiology laboratories have had to cope with an increase in the volume of tests due to the emergence of the SARS-CoV-2 virus. Short turnaround times (TATs) are important for case tracing and to help clinicians in patient management. In such a context, high-throughput systems are essential to process the bulk of the tests. Rapid tests are also required to ensure shorter TATs for urgent situations. In our laboratory, SARS-CoV-2 assays were initially implemented on our custom platform using a previously published method. The commercial cobas 6800 (Roche diagnostics) assay and the GeneXpert Xpress (Cepheid) SARS-CoV-2 assay were implemented on 24 March and 8 April 2020, respectively, as soon as available.Hypothesis/Gap Statement. Despite the abundant literature on SARS-CoV-2 assays, the articles focus mainly on the diagnostic performances. This is to our knowledge the first article that specifically studies the TAT of different assays.Aim. We aimed to describe the impact of various SARS-CoV-2 assays on the TAT at the beginning of the outbreak.Methodology. In this study, we retrospectively analysed the TAT of all SARS-CoV-2 assays performed in our centre between 24 February and 9 June, 2020.Results. We retrieved 33 900 analyses, with a median TAT of 6.25 h. TATs were highest (6.9 h) when only our custom platform was used (24 February to 24 March, 2020). They were reduced to 6.1 h when the cobas system was introduced (24 March to 8 April, 2020). The implementation of the GeneXpert further reduced the median TAT to 4.8 h (8 April to 9 June, 2020). The GeneXpert system had the shortest median TAT (1.9 h), followed by the cobas (5.5 h) and by our custom platform (6.9 h).Conclusion. This work shows that the combination of high-throughput systems and rapid tests allows the efficient processing of a large number of tests with a short TAT. In addition, the use of a custom platform allowed the quick implementation of an in-house test when commercial assays were not yet available.

Epidemiology, risk factors and clinical course of SARS-CoV-2 infected patients in a Swiss university hospital: An observational retrospective study.

BACKGROUND: This study aims to describe the epidemiology of COVID-19 patients in a Swiss university hospital. METHODS: This retrospective observational study included all adult patients hospitalized with a laboratory confirmed SARS-CoV-2 infection from March 1 to March 25, 2020. We extracted data from electronic health records. The primary outcome was the need to mechanical ventilation at day 14. We used multivariate logistic regression to identify risk factors for mechanical ventilation. Follow-up was of at least 14 days. RESULTS: 145 patients were included in the multivariate model, of whom 36 (24.8%) needed mechanical ventilation at 14 days. The median time from symptoms onset to mechanical ventilation was 9·5 days (IQR 7.00, 12.75). Multivariable regression showed increased odds of mechanical ventilation with age (OR 1.09 per year, 95% CI 1.03-1.16, p = 0.002), in males (OR 6.99, 95% CI 1.68-29.03, p = 0.007), in patients who presented with a qSOFA score ≥2 (OR 7.24, 95% CI 1.64-32.03, p = 0.009), with bilateral infiltrate (OR 18.92, 3.94-98.23, p<0.001) or with a CRP of 40 mg/l or greater (OR 5.44, 1.18-25.25; p = 0.030) on admission. Patients with more than seven days of symptoms on admission had decreased odds of mechanical ventilation (0.087, 95% CI 0.02-0.38, p = 0.001). CONCLUSIONS: This study gives some insight in the epidemiology and clinical course of patients admitted in a European tertiary hospital with SARS-CoV-2 infection. Age, male sex, high qSOFA score, CRP of 40 mg/l or greater and a bilateral radiological infiltrate could help clinicians identify patients at high risk for mechanical ventilation.

Comparison of SARS-CoV-2 serological tests with different antigen targets.

BACKGROUND: These last months, dozens of SARS-CoV-2 serological tests have become available with varying performances. A major effort was completed to compare 17 serological tests available in April 2020 in Switzerland. METHODS: In a preliminary phase, we compared 17 IgG, IgM, IgA and pan Ig serological tests including ELISA, LFA, CLIA and ECLIA on a panel of 182 sera, comprising 113 sera from hospitalized patients with a positive RT-PCR, and 69 sampled before 1st November 2019, expected to give a positive and negative results, respectively. In a second phase, the five best performing and most available tests were further evaluated on a total of 582 sera (178 and 404 expected positive and negative, respectively), allowing the assessment of 20 possible cross-reactions with other viruses. RESULTS: In the preliminary phase, among eight IgG/pan-Ig ELISA or CLIA/ECLIA tests, five had a sensitivity and specificity above 90 % and 98 % respectively, and on six IgM/IgA tests, only one was acceptable. Only one LFA test on three showed good performances for both IgG and IgM. For all the tests IgM and IgG aroused concomitantly. In the second phase, no test showed particular cross-reaction. We observed an important heterogeneity in the development of the antibody response. CONCLUSIONS: The majority of the evaluated tests exhibited high performances of IgG/pan-Ig sensitivity and specificity to detect the serological response of moderately to critically ill hospitalized patients. The IgM and IgA tests showed mostly insufficient performances with no added value for the early diagnostic on the cohort tested in this study.

Predicting Venous Thromboembolic Events in Patients with Coronavirus Disease 2019 Requiring Hospitalization: an Observational Retrospective Study by the COVIDIC Initiative in a Swiss University Hospital.

BACKGROUND: Coronavirus disease 2019 (COVID-19) can result in profound changes in blood coagulation. The aim of the study was to determine the incidence and predictors of venous thromboembolic events (VTE) among patients with COVID-19 requiring hospital admission. Subjects and Methods. We performed a retrospective study at the Lausanne University Hospital with patients admitted because of COVID-19 from February 28 to April 30, 2020. RESULTS: Among 443 patients with COVID-19, VTE was diagnosed in 41 patients (9.3%; 27 pulmonary embolisms, 12 deep vein thrombosis, one pulmonary embolism and deep vein thrombosis, one portal vein thrombosis). VTE was diagnosed already upon admission in 14 (34.1%) patients and 27 (65.9%) during hospital stay (18 in ICU and nine in wards outside the ICU). Multivariate analysis revealed D-dimer value > 3,120 ng/ml (P < 0.001; OR 15.8, 95% CI 4.7-52.9) and duration of 8 days or more from COVID-19 symptoms onset to presentation (P 0.020; OR 4.8, 95% CI 1.3-18.3) to be independently associated with VTE upon admission. D-dimer value ≥ 3,000 ng/l combined with a Wells score for PE ≥ 2 was highly specific (sensitivity 57.1%, specificity 91.6%) in detecting VTE upon admission. Development of VTE during hospitalization was independently associated with D-dimer value > 5,611 ng/ml (P < 0.001; OR 6.3, 95% CI 2.4-16.2) and mechanical ventilation (P < 0.001; OR 5.9, 95% CI 2.3-15.1). CONCLUSIONS: VTE seems to be a common COVID-19 complication upon admission and during hospitalization, especially in ICU. The combination of Wells ≥ 2 score and D - dimer ≥ 3,000 ng/l is a good predictor of VTE at admission.

Comparison of SARS-CoV-2 RT-PCR on a high-throughput molecular diagnostic platform and the cobas SARS-CoV-2 test for the diagnostic of COVID-19 on various clinical samples.

OBJECTIVES: In order to cope with the rapid spread of the COVID-19 pandemic, we introduced on our in-house high-throughput molecular diagnostic platform (MDx Platform) a real-time reverse transcriptase PCR (RT-PCR) to detect the SARS-CoV-2 from any clinical specimens. The aim of this study was to compare the RT-PCR results obtain with the MDx Platform and the commercial assay cobas SARS-CoV-2 (Roche) on nasopharyngeal swab and other clinical specimens including sputum, bronchial aspirate, bronchoalveolar lavage and anal swabs. METHODS: Samples received in our laboratory from patients suspected of COVID-19 (n = 262) were tested in parallel with our MDx platform SARS-CoV-2 PCR and with the cobas SARS-CoV-2 test. RESULTS: The overall agreement between the two tests for all samples tested was 99.24% (260/262), which corresponded to agreements of 100% (178/178) on nasopharyngeal swabs, 95.45% (42/44) on lower respiratory tract specimen with discordant resultS obtained for very high cycle threshold (Ct) value and 100% (40/40) on anorectal swabs. The Ct values for nasopharyngeal swabs displayed an excellent correlation (R2 > 96%) between both tests. CONCLUSIONS: The high agreements between the cobas SARS-CoV-2 test and the MDx platform supports the use of both methods for the diagnostic of COVID-19 on various clinical samples. Very few discrepant results may occur at very low viral load.

Viral load of SARS-CoV-2 across patients and compared to other respiratory viruses.

RT-PCRs to detect SARS-CoV-2 RNA is key to manage the COVID-19 pandemic. We analyzed SARS-CoV-2 viral loads from 22'323 RT-PCR results according to samples types, gender, age, and health units. Viral load did not show any difference across age and appears to be a poor predictor of disease outcome. SARS-CoV-2 viral load showed similar high viral loads than the one observed for RSV and influenza B. The importance of viral load to predict contagiousness and to assess disease progression is discussed.