Universal Transport Medium Research Articles

Comparison of Abbot ID Now Method with Eurobioplex RT-PCR SARS-Cov-2 Multiplex Method for Detection of SARS-Cov-2 from Nasopharyngeal and Oropharyngeal Samples

Background: COVID 19 created an urgent demand for rapid diagnosis to encircle this pandemic and improve patient management. In this context, we evaluated the concordance of the ID NOW test compared to the Eurobioplex RT-PCR test in the rapid diagnosis of SARS-CoV-2. Methods: To evaluate the concordance of the assay at different viral loads, 154 positive samples were selected to represent the full range of Ct values observed on the Eurobioplex RT-PCR assay, ranging from 14 to 38 cycles. Positive concordance for the ID Now assay was calculating dusing Eurobioplex RT-PCR as the reference test. An additional 70 negative samples were selected to assess negative concordance. Results: Compared to Eurobioplex RT-PCR, the overall positive agreement was 79% with ID Now. Negative agreement was 100% for ID Now. ID Now showed 100% positive agreement for medium and high viral concentrations (Ct value <30). However, for Ct values >30, the positive agreement was 33.3% for ID Now. Conclusions: This study shows a major limitation of ID Now for specimens collected in universal transport media with lower viral concentrations. Further studies are necessary to evaluate the performance of ID Now for dry nasopharyngeal swabs (manufacturer's recommended method).

Culturing of SARS-CoV-2 from patient samples: Protocol for optimal virus recovery and assessment of infectious viral load

Optimal sampling, preservation, and culturing of SARS-CoV-2 from COVID-19 patients are critical for successful recovery of virus isolates and to accurately estimate contagiousness of the patient. In this study, we investigated the influence of the type of sampling media, storage time, freezing conditions, sterile filtration, and combinations of these to determine the optimal pre-analytic conditions for virus recovery and estimation of infectious viral load in COVID-19 patients. Further, we investigated the viral shedding kinetics and mucosal antibody response in 38 COVID-19 hospitalized patients. We found Universal Transport Medium (Copan) to be the most optimal medium for preservation of SARS-CoV-2 infectivity. Our data showed that the probability of a positive viral culture was strongly correlated to Ct values, however some samples did not follow the general trend. We found a significant correlation between plaque forming units and levels of mucosal antibodies and found that high levels of mucosal antibodies correlated with reduced chance of isolating the virus. Our data reveals essential parameters to consider from specimen collection over storage to culturing technique for optimal chance of isolating SARS-CoV-2 and accurately estimating patient contagiousness.

Analytical performance of the Abbott ID NOW 2.0 assay for SARS-CoV-2 detection in clinical samples from symptomatic patients

We evaluated the analytical performance of ID NOW™ COVID-19 2.0 assay versus conventional real-time reverse transcription-polymerase chain reaction (RT-PCR) using a total of 792 clinical samples from nasopharyngeal and oropharyngeal swabs, stored in frozen universal transport medium samples. Positive percent agreement (PPA) and negative percent agreement of ID NOW were 97.6 % and 100 %, respectively. The overall percent agreement between ID NOW and RT-PCR was 99.5 %. The PPA of ID NOW in detecting SARS-CoV-2 in 164 RT-PCR positive patients, all of whom had symptoms related COVID-19, was 97.1 % within 8 days since symptom onset, 97.9 % from 8 to 14 days since symptom onset, and 97.6 % after 14 days since symptom onset, with no significant difference between the days since symptom onset. The ID NOW assay demonstrated good performance, providing a rapid and randomly accessible alternative to conventional RT-PCR for timely SARS-CoV-2 detection, particularly in situations requiring rapid results for patient care.

Impact of Omicron variant sublineage BA.2.75 on the OnSite COVID-19 Ag Rapid Test: the applicability of rapid antigen test with universal transport media

Background Rapid antigen testing (RAT) is one of the most powerful tools for SARS-CoV-2 detection. The OnSite COVID-19 Ag Rapid Test is an antigen-based, point-of-care test approved by the WHO for Emergency Use Listing. The Nucleocapsid (N) gene mutations found in the emerging Omicron sublineages lead to the question of RAT performance. Objective To ensure the diagnostic performance of the study RAT during rapidly mutated Omicron variants. Results We independently evaluated the performance of this assay in 1098 archived samples collected in Thailand during October 2022–February 2023, which were 798 and 300 COVID-19 real-time RT-PCR positive and negative, respectively. The assay performed with 100% sensitivity and 100% specificity using a cycle threshold (Ct) of <20 for the RT-PCR. The sensitivity decreased to 88% when using Ct <30. Most of the SARS-CoV-2 found were Omicron BA.2 (99%), harboring six known N mutations (P13L, E31del, S33del, R203K, G204R and S413R). Eight samples containing hybrid variants (XBB.1*, XBB.2 and XBJ) were detected by the study RAT. This RAT detects all Omicron sublineages known to be circulating in Thailand. Conclusions These results confirmed the good performance of the study RAT for detecting Omicron variants and its appropriateness for individual diagnosis and for genomic surveillance.

A-297 Usefulness of Non-Skin Samples in the PCR Diagnosis of Monkeypox

Abstract Background During 2022, human mpox cases were reported in several European countries. Until November 2022, in Spain, 7404 cases were detected, most of them linked to sexual intercourse. In that context, fast identification of cases and sources of infection were essential for an optimal management. Polymerase chain reaction (PCR) was the preferred laboratory test. Our objective was to evaluate the usefulness of sera and nasopharyngeal samples for MPXV detection, and its application in contact studies without skin or mucosal injuries. Methods In the Hospital Clínico Universitario of Zaragoza (Spain), 106 samples from 50 patients, 38 male and 12 female, aged between 11 to 62 years, with skin lesions suspected MPXV infection were analysed. Samples were collected between May and September 2022 in inactivating universal viral transport medium (Biocomma). Samples were obtained between 1 and 19 days of evolution. To perform the real-time PCR, the VIASURE Monkeypox Virus Real Time PCR Detection Kit and the VIASURE 48—Real Time PCR System were used. Nucleic acid extraction was performed with the automated nucleic acid extraction system MagLEAD12gC using the MagDEA®Dx SV reagents. Results Table 1 shows the results expressed in Ct values. Skin, anogenital and serum samples were the most efficient for MPXV diagnosis. Nevertheless, these data are influenced by the sample collection date. Conclusion Real-time PCR was positive in samples from all locations in 71,4% of the infected patients. Respiratory samples (pharyngeal and nasopharyngeal) were the ones that showed the highest number of false negative results, whereas there was only one skin sample and one serum sample with a negative result. This study shows that sera samples can be used in the diagnosis of MPXV infection because of ease of collection, applicability to contacts without lesions and the option of serology testing for other STI agents (HBV, HCV, HIV, Treponema pallidum…).

Development and Validation of Three Automated High-Throughput Molecular Tests to Detect Monkeypox Virus Infections.

The 2022 outbreak of the clade IIb monkeypox virus and subsequent global spread lead to an urgent need for the development of high-throughput, sensitive, and reproducible diagnostic tests. We developed 3 assays to detect monkeypox virus, 2 (MPXV+ and MPXV) for m2000 RealTime and 1 (MPXV) for Alinity m platforms. Dual targets in E9L and B6R (MPXV+) and J2L and B7R (MPXV) increased mutation resistance. In silico prediction indicates MPXV+ cross-reactivity with orthopox viruses and specific monkeypox virus detection with MPXV. m2000 RealTime MPXV+ and MPXV assay sensitivity was determined to be 3.2 plaque-forming units/mL using a reference virus culture diluted into universal transport medium (UTM). Alinity m MPXV lower limit of detection was 200 copies/mL using monkeypox virus plasmids in pooled UTM matrix. m2000 RealTime MPXV+ and MPXV assays were validated with lesion swabs in UTM and 1:1 saliva to UTM mixtures. Commercially available and remnant clinical lesion specimens in UTM were tested with RealTime MPXV+, RealTime MPXV and Alinity m MPXV assays and demonstrated high agreement to known mpox (MPX)-positive specimens. RealTime MPXV+, RealTime MPXV, and Alinity MPXV are high throughput and sensitive assays used for the detection of monkeypox virus. These assays maybe useful during MPX outbreaks.

Hunting for mpox (monkeypox) mimickers: Use of the Biofire meningitis/encephalitis panel on lesion swabs to support alternative viral diagnoses

Mpox (formerly monkeypox) is an emerging zoonotic disease of public health concern that presents as a rash mimicking other common viral exanthems. Unlike traditional testing algorithms relying on several assays, the BioFire FilmArray meningitis/encephalitis (ME) panel simultaneously detects common viruses causing rashes; however, Biofire ME is only licensed for testing on cerebral spinal fluid. This study evaluated use of the Biofire ME panel for detection and discrimination of herpes simplex virus types 1 and 2 (HSV-1 and HSV-2), varicella zoster virus (VZV), human herpesviruses type 6 (HHV-6), enteroviruses (EVs), and human paraechoviruses (HPeVs) from a dermal or mucocutaneous swabs collected in universal transport media (UTM). Results of the BioFire ME panel were compared against methods used during clinical testing. Ten-fold serial dilutions in UTM of cultured viruses were used to compare analytical sensitivity, and analytical specificity was assessed using panels of microorganisms in UTM. Clinical sensitivity and specificity were assessed using 20 positive specimens each for HHV-1, HHV-2, HHV-6, VZV, EVs, and HPeV, as well as 35 known negative specimens that included 15 mpox-positive specimens. Biofire ME was as sensitive as comparator methods, and correctly discriminated all HSV-1, HSV-2, VZV, HHV-6, EVs, and HPeVs from mpox and mpox-mimickers. Cross-reaction between EV and rhinoviruses A, B, and C were noted in the specificity panel. Swabs in UTM collected for mpox testing are suitable for use on the Biofire ME panel, allowing more streamlined diagnostic testing for viral exanthems in patients under investigation for mpox infection.

Prospective Evaluation of SARS-CoV-2 Rapid Antigen Test for Screening of Asymptomatic Caregivers.

This study aimed to assess clinical performance of a rapid antigen test (RAT) for screening asymptomatic patients during the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) omicron outbreak. RAT with the routine real-time reverse transcription-polymerase chain reaction (rRT-PCR) using the same nasopharyngeal swab in universal transport medium was performed for rapid screening of asymptomatic caregivers of emergent patients from March to April 2022 in a tertiary-care hospital in Korea. Clinical performance of RAT compared to that analyzed by rRT-PCR was evaluated. A total of 900 caregivers were enrolled in this study, of which 14 (1.6%) were RAT-positive and 44 (5.0%) were positive for rRT-PCR. Overall sensitivity and specificity of RAT were 31.8% and 100.0%, respectively. Caution must be taken when using RAT as a screening test for asymptomatic caregivers as this may lead to outbreaks among high-risk patients.

314. Performance of Saliva Compared to Nasopharyngeal Swab in Detection of SARS-CoV-2 at a Field Site in Rural Guatemala Using the Roche Cobas® Liat® Point-of-Care Assay

Abstract Background Saliva samples are less invasive but not considered the gold standard for detecting SARS-CoV-2, and they are not validated for the Roche Cobas® Liat® platform. We aimed to evaluate the performance of a saliva sample compared to nasopharyngeal (NP) swab in the detection of SARS-CoV-2 on the Roche Cobas® Liat® platform at a research site in rural Guatemala. Methods Adults in an existing cohort study with influenza-like illness (ILI) provided clinical data and underwent NP swab (Copan) collection by trained nurses. Swabs were immediately placed into universal transport media (UTM), stored at 2-8 °C for less than 30 minutes and then tested on the Cobas Liat platform for SARS-CoV-2. Consenting subjects who had not eaten or drank in the last 2 hours were asked to provide a 5-mL saliva sample directly into an RNAse free container. The saliva sample remained at 2-8 °C for 24 hours, then diluted 1:2 with 0.85% saline (to reduce viscosity) and run on the same assay. We used descriptive statistics to compare the performance of saliva to NP swabs. Results Of the 28 subjects screened (1/10/22 - 4/26/22), 23 (82%) were consented and enrolled. The majority of subjects were male (78.3%) with a mean age of 31 years (range: 18-59 years). Of the 23 subjects enrolled, 14 (30.4%) reported fever and cough, 16 (34.8%) reported cough only, 14 (30.4%) reported fever only, and 2 (4.3%) reported cough and nasal congestion. The median symptom duration was 3.0 days (IQR: 1.5-5 days). Of the 23 subjects, 5 (23.7%) tested positive for SARS-CoV-2 in both sample types (NP swab and saliva); 3 (13%) were discordant, including 1 (4.3%) saliva-positive only and 2 (8.7%) NP-positive only. Compared to NP swab, preliminary performance of saliva in detection of SARS-CoV-2 included a sensitivity, specificity, accuracy, positive predictive value, and negative predictive value of 71.4%, 93.7%, 87%, 83.3%, and 88.2%, respectively. The Fisher’s exact test p-value (0.003) shows concordance between both tests. Conclusion Our preliminary results show good precision between NP and saliva samples in detection of SARS-CoV-2 on the Roche Cobas® Liat® platform. Ongoing data collection will provide greater insight on the discordant results, but our findings support the continued use of saliva in the detection of SARS-CoV-2 in this population. Disclosures Molly Lamb, PhD, Pfizer: Grant/Research Support Edwin J. Asturias, MD, Curevac: DSMB Member|Fundacion para la Salud Integral de los Guatemaltecos: Board Member|Inovio: DSMB Member|Merck: Honoraria|Pfizer: Grant/Research Support Diva Calvimontes, n/a, Pfizer: Grant/Research Support Daniel Olson, MD, Fundacion para la Salud Integral de los Guatemaltecos: Board Member|Pfizer: Grant/Research Support|Roche: Grant/Research Support.

Thermal Shock Viral RNA Release in COVID-19 Testing

Abstract Introduction/Objective The public health emergency of the COVID-19 pandemic emphasized the crucial role of medical laboratory professionals and scientists in molecular diagnostics laboratories to ensure success in infection control strategies. The demand for laboratory testing using nucleic acid amplification tests to detect SARS-CoV-2 RNA imposed strains in laboratory supplies. Here, we explored an alternative cost-effective solution that will simplify the pre-PCR steps by using a simple heating method to release viral RNA. Methods/Case Report Samples tested using the reference automated extraction method were used:100 samples identified as positive for SARS-CoV-2 RNA and 500 samples tested negative for SARS-CoV-2 RNA were used for the study and sorted with equal distribution according to Ct values of (a) &amp;lt;20, (b) 20–30, and (c) &amp;gt;30.100 ul from swab preserved in Universal Transport Medium was treated with 30 μg of proteinase K, and another set was tested without proteinase K pre-treatment. All samples with or without proteinase K were diluted to minimize PCR inhibitors. The thermal shock protocol was set at (98°C, 5 minutes; 4°C, 2 minutes) and screened for purity. Performance and method verification studies were performed. Internal extraction, positive template, and no template controls were markers used for testing quality. The experimental study was performed by qualified testing personnel and all under the same experimental conditions. Results (if a Case Study enter NA) The Ct values from the thermal shock RNA release were compared to the automated extraction method and statistically analyzed.The criteria for acceptability for validation of this new RNA extraction proceeding were set to 100% concordance compared to the commercial kit using an automated extraction. PCR efficiency was at 98% and a slope of -3.3. Within run precision of 2% and limits of detection from 200 to 20,000 copies/uL The method 100% (50/50) concordance on samples previously identified as negative by automated methods and identified 86% (86/100) with a mean difference of 3 Ct. Conclusion Our findings suggest that the thermal shock treatment of nasopharyngeal swabs in viral transport media can successfully extract viral nucleic acid for nucleic acid amplification and is a reasonable alternative for chemical extraction methods when molecular diagnostic laboratories persistently encounter supply chain issues.

Comparison of two SARS-CoV-2 RT-PCR assays and implication of the instrument software on cycle threshold (Ct) value

The Cycle Threshold (Ct) value of SARS-CoV-2 RT-PCR are used as an indicator of viral load. Using a collection of 45 fresh nasopharyngeal samples collected on universal transport media, we compare the Ct value obtained with 2 RT-PCR assays, the Alinity M SARS-CoV-2 and the Alinity M RESP-4-Plex (Abbott Molecular, Des Plaines, Illinois, Etats-Unis) processed on an Alinity M device. The assays are highly correlated; however, the Ct values were in median lower of 4.54 with the Alinity M RESP-4-Plex. This difference could be attributed to earlier detection of positivity by the software of the Alinity M rather than a difference in RT-PCR performances. The Ct-value of SARS-CoV-2 RT-PCR should be interpreted with caution taking into account the clinical context, pre-analytical and analytical findings.

Performance Evaluation of the Microfluidic Antigen LumiraDx SARS-CoV-2 and FluA/B Test in Diagnosing COVID-19 and Influenza in Patients with Respiratory Symptoms.

IntroductionCoronavirus disease 2019 (COVID-19) and influenza share similar symptoms, which hampers diagnosis. Given that they require different containment and treatment strategies, fast and accurate distinction between the two infections is needed. This study evaluates the sensitivity and specificity of the microfluidic antigen LumiraDx SARS-CoV-2 and Flu A/B Test for simultaneous detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and influenza A/B from a single nasal swab.MethodsNasal samples were collected from patients as part of the ASPIRE (NCT04557046) and INSPIRE (NCT04288921) studies at point-of-care testing sites in the USA. ASPIRE study participants were included after developing COVID-19 symptoms in the last 14 days or following a positive SARS-CoV-2 test in the last 48 h. INSPIRE study participants were included after developing influenza symptoms in the last 4 days. Samples were extracted into proprietary buffer and analysed using the LumiraDx SARS-CoV-2 and Flu A/B Test. A reference sample was taken from each subject, placed into universal transport medium and tested using reference SARS-CoV-2 and influenza reverse transcription polymerase chain reaction (RT-PCR) tests. The test and reference samples were compared using the positive percent agreement (PPA) and negative percent agreement (NPA), together with their 95% confidence intervals (CIs).ResultsAnalysis of the data from the ASPIRE (N = 124) and INSPIRE (N = 159) studies revealed high levels of agreement between the LumiraDx SARS-CoV-2 and Flu A/B Test and the reference tests in detecting SARS-CoV-2 (PPA = 95.5% [95% CI 84.9%, 98.7%]; NPA = 96.0% [95% CI 90.9%, 98.3%]), influenza A (PPA = 83.3% [95% CI 66.4%, 92.7%]; NPA = 97.7% [95% CI 93.4%, 99.2%]) and influenza B (PPA = 80.0% [95% CI 62.7%, 90.5%]; NPA = 95.3% [95% CI 90.2%, 97.9%]).ConclusionsThe LumiraDx SARS-CoV-2 and Flu A/B Test shows a high agreement with the reference RT-PCR tests while simultaneously detecting and differentiating between SARS-CoV-2 and influenza A/B.Trial RegistrationClinicalTrials.gov identifiers NCT04557046 and NCT04288921.Supplementary InformationThe online version contains supplementary material available at 10.1007/s40121-022-00696-8.

Agile design and development of a high throughput cobas SARS-CoV-2 RT-PCR diagnostic test

Diagnostic testing is essential for management of the COVID-19 pandemic. An agile assay design methodology, optimized for the cobas® 6800/8800 system, was used to develop a dual-target, qualitative SARS-CoV-2 RT-PCR test using commercially available reagents and existing sample processing and thermocycling profiles. The limit of detection was 30–52 copies/mL for USA-WA1/2020. Assay sensitivity was confirmed for SARS-CoV-2 variants Alpha, Beta, Gamma, Delta and Kappa. The coefficients of variation of the cycle threshold number (Ct) were between 1.1 and 2.2%. There was no difference in Ct using nasopharyngeal compared to oropharyngeal swabs in universal transport medium (UTM). A small increase in Ct was observed with specimens collected in cobas PCR medium compared to UTM. In silico analysis indicated that the dual-target test is capable of detecting all >1,800,000 SARS-CoV-2 sequences in the GISAID database. Our agile assay design approach facilitated rapid development and deployment of this SARS-CoV-2 RT-PCR test.

Passengers screening for SARS-CoV-2 infection at Rome Fiumicino Airport: a strategy aimed at limiting the spreading of the infection through COVID-free air flights revealed the airport role as a formidable sentinel center for monitoring pandemic trends and viral variants circulation

Background and aims: At the Fiumicino Rome Airport, passengers can undergo the antigen or molecular test for SARS-CoV-2 at the COVID-19 Test Area (CTA), in compliance with the boarding regulations on air flights and those for entering the destination countries. The aim of this study was to describe the adaptability and usefulness of using rapid molecular tests designed for point of care (POC) in a context such as the CTA of Rome Fiumicino airport, by describing the volume and activities performed in the last period of the state of emergency in Italy, July 10, 2021- March 31, 2022.&#x0D; &#x0D; Materials and methods: Rapid molecular tests for SARS-CoV-2 were carried out with ID NOWTM COVID-19 assay on the ID NOWTM platform using dry swabs collected from passengers (or airport staff) at the CTA. Swabs were stored at room temperature and analyzed within 1-2 hours of collection. Sequencing for SARS-CoV-2 variants of concern (VOC) identification was carried out on swabs dipped in universal transport medium (UTM), after new swab gathering from consenting passengers resulted positive with the rapid molecular assay. A proficiency panel prepared with the viral isolate 2019-nCoV/Italy INMI1 was used to evaluate analytical sensitivity; clinical sensitivity was tested by performing real-time polymerase chain reaction (RT-PCR) tests on 50 swabs obtained from passengers who previously resulted positive to ID NOWTM COVID-19 assay.&#x0D; &#x0D; Results: In total, 14632 rapid molecular tests for SARS-CoV-2 were performed at CTA and 5.6% resulted positive. Sequence analysis of samples with Ct value &lt;25 (61,2 %) with real-time RT-PCR test allowed detection of all the main VOCs: B.1.617.2 (Delta) in 18% of samples, BA.1 (Omicron-1) in 34% of positive swabs and BA.2 (Omicron-2) lineage in 9% of samples. The BA.2 variant was detected in the Lazio region for the first time at CTA, in one sample obtained from an Italian passenger coming from Doha, Qatar. A proficiency panel for SARS-CoV-2 detection was used to establish the limit of detection of ID NOWTM COVID-19 assay. For clinical sensitivity evaluation, 50 subjects with positive results by ID NOWTM COVID-19 assay were tested with routine real-time RT-PCR method in the laboratory and confirmed for SARS-CoV-2 detection.&#x0D; &#x0D; Conclusions: The great success among passengers, demonstrated by the large number of rapid molecular tests performed, confirmed the appreciation of this service by the users. Our experience at CTA showed that airports, railways and maritime stations should represent strategic surveillance centers for monitoring of circulation of respiratory pathogens, and may have a direct impact on the implementation of prevention and control strategies for public health against the spreading of infectious diseases. Indeed, SARS-CoV-2 data obtained at the CTA faithfully reflected the Italian epidemic curve trend of SARS-CoV-2 and its variants. The rapid molecular ID NOWTM COVID-19 assay used at CTA has proven to be suitable for use in a special context, such as the Rome Fiumicino Airport, though it was designed for use in POC. Due to the simplicity of use, robustness of method and technology that allows the return of results in a few minutes, this system revealed itself to be an optimal tool at the CTA, ensuring passengers boarding on COVID-free flights in a short time and security.

Simple Saliva Sample Collection for the Detection of SARS-CoV-2 Variants Compared With Nasopharyngeal Swab Sample.

The use of saliva samples for diagnosis of SARS-CoV-2 infection offers several advantages, including ease of sample collection, feasibility of self-collection, and minimization of medical staff exposure to infection. The emergence of new SARS-CoV-2 variants has had an impact on the viral load of specimens and the results of real-time reverse transcription-polymerase chain reaction (rRT-PCR). To compare nasopharyngeal swab and saliva samples for the diagnosis of SARS-CoV-2 using rRT-PCR. In this study, participants were recruited prospectively, and paired nasopharyngeal swab and saliva samples were collected simultaneously from each participant. After adding universal transport medium, RNA was extracted in an identical manner for both sample types, and samples were tested using rRT-PCR. In addition, samples with positive results were tested for SARS-CoV-2 variants. Of the 338 paired samples, 100 nasopharyngeal swab and 101 saliva samples tested positive for SARS-CoV-2. The rRT-PCR results of the saliva and nasopharyngeal swab samples showed a positive percent agreement of 95.0% (95% CI, 88.7%-98.4%), a negative percent agreement of 97.9% (95% CI, 95.2%-99.3%), and an overall percent agreement of 96.8% (95% CI, 94.3%-98.4%). SARS-CoV-2 was detected in the saliva samples of 6 participants with negative nasopharyngeal sample results. In addition, the sensitivity of saliva samples was similar to that of nasopharyngeal samples for detecting various SARS-CoV-2 variants, including the Omicron variant. Saliva samples can be used as an alternative to nasopharyngeal samples for convenient and effective detection of various SARS-CoV-2 variants.

Comparative Diagnostic Accuracy of the STANDARD M10 Assay for the Molecular Diagnosis of SARS-CoV-2 in the Point-of-Care and Critical Care Settings.

Accurate and rapid molecular diagnosis of COVID-19 is a crucial step to tackle the ongoing pandemic. The primary objective of this study was to estimate the real-world performance of the novel RT-PCR STANDARD M10 SARS-CoV-2 assay in a large number of nasopharyngeal (NP) specimens eluted in universal transport medium. The secondary objective was to evaluate the compatibility of this kit in testing NP samples eluted in an inactivated transport medium (essential for point-of-care testing) and lower respiratory tract (LRT) specimens, which are commonly collected in critical care. A total of 591 samples were analyzed. Compared with the standard extraction-based RT-PCR Allplex 2019-nCoV (time-to-result of 270 min), the sensitivities of the STANDARD M10 were 100% (95% CI: 98.1–100%), 95.5% (95% CI: 91.7–97.6%), and 99.5% (95% CI: 97.2–99.9%) for ≥1 gene, the ORF1ab gene, and the E gene, respectively, while the specificity was 100% (95% CI: 98.7–100%). The diagnostic accuracy was 100% in testing both NP samples eluted in an inactivated transport medium and LRT specimens. STANDARD M10 reliably detects SARS-CoV-2 in 60 min, may be used as a POC tool, and is suitable for testing LRT specimens in the critical care setting.

SARS-CoV-2 Transmission Control Measures in the Emergency Department: The Role of Rapid Antigenic Testing in Asymptomatic Subjects.

Limiting transmission of SARS-CoV-2 from asymptomatic people assumes the paramount importance of keeping fragile subjects protected. We evaluated the utility of rapid SARS-CoV-2 antigen testing in asymptomatic subjects attending emergency departments in non-COVID-19 areas, using a single nasopharyngeal swab specimen collected in universal transport medium to perform both rapid antigen testing and rRT-PCR (used as reference standard) in a cohort of 899 patients. In the overall sample, the rapid antigen test had 43.9% sensitivity, 100% specificity, 100% positive predictive value, 93.6% negative predictive value. Considering subjects with rRT-PCR cycle threshold ≤30, the test had 80.4% sensitivity, 100% specificity, 100% positive predictive value, 98.8% negative predictive value. Considering subjects with rRT-PCR cycle threshold ≤25, the test had 94.7% sensitivity, 100% specificity, 100% positive predictive value and 99.7% negative predictive value. Despite low sensitivity, routine application of rapid antigen testing in the emergency department can lead to isolation in less than 30 min of about a half of asymptomatic COVID-19 subjects assigned to non-COVID-19 areas by clinical triage. The rapid test correctly identified 94.7% of asymptomatic patients with cycle threshold ≤ 25 that are supposed to be more infective; thus, it could be a useful measure to contain viral transmission in non-COVID-19 areas.

Highly sensitive, scalable, and rapid SARS-CoV-2 biosensor based on In2O3 nanoribbon transistors and phosphatase

Developing convenient and accurate SARS-CoV-2 antigen test and serology test is crucial in curbing the global COVID-19 pandemic. In this work, we report an improved indium oxide (In2O3) nanoribbon field-effect transistor (FET) biosensor platform detecting both SARS-CoV-2 antigen and antibody. Our FET biosensors, which were fabricated using a scalable and cost-efficient lithography-free process utilizing shadow masks, consist of an In2O3 channel and a newly developed stable enzyme reporter. During the biosensing process, the phosphatase enzymatic reaction generated pH change of the solution, which was then detected and converted to electrical signal by our In2O3 FETs. The biosensors applied phosphatase as enzyme reporter, which has a much better stability than the widely used urease in FET based biosensors. As proof-of-principle studies, we demonstrate the detection of SARS-CoV-2 spike protein in both phosphate-buffered saline (PBS) buffer and universal transport medium (UTM) (limit of detection [LoD]: 100 fg/mL). Following the SARS-CoV-2 antigen tests, we developed and characterized additional sensors aimed at SARS-CoV-2 IgG antibodies, which is important to trace past infection and vaccination. Our spike protein IgG antibody tests exhibit excellent detection limits in both PBS and human whole blood ((LoD): 1 pg/mL). Our biosensors display similar detection performance in different mediums, demonstrating that our biosensor approach is not limited by Debye screening from salts and can selectively detect biomarkers in physiological fluids. The newly selected enzyme for our platform performs much better performance and longer shelf life which will lead our biosensor platform to be capable for real clinical diagnosis usage.Electronic Supplementary MaterialSupplementary material (materials and methods for device fabrication, functionalization of In2O3 devices, photographs of the liquid gate measurement setup, mobilities of the nine devices labeled in Fig. 1(b), family curves of IDS-VDS with the liquid gate setup and current change after bubbling the substrate solution (current vs. time curve for S1 antigen detection)) is available in the online version of this article at 10.1007/s12274-022-4190-0.

A method comparison study of the high throughput automated HISCL® SARS-CoV-2 antigen assay using nasopharyngeal swab samples from symptomatic and asymptomatic subjects against conventional RT-PCR.

Our study aim was to evaluate the performance of the automated Sysmex HISCL® severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2) antigen assay against reverse‐transcription polymerase chain reaction (RT‐PCR). We tested 277 remnant frozen nasopharyngeal swab samples, stored in universal transport medium (UTM), yielding a sensitivity of 94.9% against historical RT‐PCR results with cycle threshold (C t) < 30, and a sensitivity of 76.7% for C t < 35, and specificity of 100% (all C t values) confirming compatibility of UTM‐diluted samples with the assay system. Thereafter, we prospectively collected 141 nasopharyngeal swab samples in UTM from healthcare workers and 1369 paired swabs (400 UTM; 969 dry) from individuals at a public health testing center, with the first swab (UTM) reserved for RT‐PCR, yielding a positivity rate of 4.6%. HISCL assay performance using UTM swabs was superior to dry swabs, with a sensitivity of 100% (95% confidence interval [CI] 71.5%–100%) at C t < 30 versus 92.3% (95%CI 81.5%–97.9%), and a specificity of 99.3% (95% CI 98.1–99.89) against 83.3% (95%CI 80.7%–85.6%). We conclude that this antigen assay is suitable for high throughput facilities where the primary indication for testing is to rule out infection with low RT‐PCR C t values (proxy for high viral loads) to curb viral spread.

Clinical Comparative Evaluation of the LabTurboTM AIO® Reverse Transcription-Polymerase Chain Reaction and World Health Organization-Recommended Assays for the Detection of Emerging SARS-CoV-2 Variants of Concern.

PurposeSevere acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the causative agent behind coronavirus disease-2019 (COVID-19). Single-plex reverse transcription-polymerase chain reaction (RT-PCR)-based assays are widely used for COVID-19 detection but exhibit decreased sensitivity and specificity in detecting the rapidly spreading SARS-CoV-2 variants; in contrast, multiplex RT-PCR reportedly yields better results. Here, we aimed at comparatively analyzing the clinical performance of the LabTurboTM AIO COVID-19 RNA testing kit, a multiplex quantitative RT-PCR kit, including a three-target (E, N1, and RNase P), single-reaction, triplex assay used for SARS-CoV-2 detection, with that of the WHO-recommended RT-PCR assay.Materials and MethodsResidual, natural, nasopharyngeal swabs obtained from universal transport medium specimens at SARS-CoV-2 testing centers (n = 414) were collected from May to October 2021. For SARS-CoV-2 qRT-PCR, total viral nucleic acid was extracted. The limit of detection (LOD) and the comparative clinical performances of the LabTurboTM AIO COVID-19 RNA kit and the WHO-recommended RT-PCR assay were assessed. Statistical analysis of the correlation was performed and results with R2 values >0.9 were considered to be highly correlated.ResultsThe LOD of the LabTurboTM AIO COVID-19 RNA kit was 9.4 copies/reaction for the target genes N1 and E. The results obtained from 102 SARS-CoV-2-positive and 312 SARS-CoV-2-negative samples showed 100% correlation with previous WHO-recommended RT-PCR assay results.ConclusionMultiplex qRT-PCR is a critical tool for detecting unknown pathogens and employs multiple target genes. The LabTurboTM AIO COVID-19 RNA testing kit provides an effective and efficient assay for SARS-CoV-2 detection and is highly compatible with SARS-CoV-2 variants.