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1.
PLoS One ; 16(10): e0257834, 2021.
Article in English | MEDLINE | ID: covidwho-1468159

ABSTRACT

COVID-19 testing is required before admission of a patient in the hospitals, invasive procedures, major and minor surgeries etc. Real Time Polymerase chain reaction is the gold standard test for the diagnosis, but requires well equipped biosafety laboratory along with trained manpower. In this study we have evaluated the diagnostic accuracy of novel TrueNat molecular assay for detecting SARS CoV-2. TrueNat is a chip-based real time PCR test and works on portable, light weight, battery powered equipment and can be used in remote areas with poor infrastructure. In this study 1807 patients samples were collected for both TrueNat and RTPCR COVID-19 testing during study period. Of these 174 (9.7%) and 174 (15%) were positive by RTPCR and TrueNat respectively and taking results of RTPCR as gold standard TrueNat test showed a sensitivity, specificity and diagnostic accuracy of 69.5, 90.9% and 89.2% respectively. It can be concluded that TrueNat is a simple, easy to use, good rapid molecular diagnostic test for diagnosis of COVID-19 especially in resource limited settings and will prove to be a game changer of molecular diagnostics in future.


Subject(s)
COVID-19/diagnosis , Reverse Transcriptase Polymerase Chain Reaction/methods , SARS-CoV-2/genetics , COVID-19/virology , COVID-19 Testing , Humans , Point-of-Care Systems , Real-Time Polymerase Chain Reaction/instrumentation , Retrospective Studies , Reverse Transcriptase Polymerase Chain Reaction/instrumentation , SARS-CoV-2/isolation & purification , Sensitivity and Specificity
2.
PLoS One ; 16(7): e0252509, 2021.
Article in English | MEDLINE | ID: covidwho-1388919

ABSTRACT

The current global pandemic due to the SARS-CoV-2 has pushed the limits of global health systems across all aspects of clinical care, including laboratory diagnostics. Supply chain disruptions and rapidly-shifting markets have resulted in flash-scarcity of commercial laboratory reagents; this has motivated health care providers to search for alternative workflows to cope with the international increase in demand for SARS-CoV-2 testing. The aim of this study is to present a reproducible workflow for real time RT-PCR SARS-CoV-2 testing using OT-2 open-source liquid-handling robots (Opentrons, NY). We have developed a framework that includes a code template which is helpful for building different stand-alone robotic stations, capable of performing specific protocols. Such stations can be combined together to create a complex multi-stage workflow, from sample setup to real time RT-PCR. Using our open-source code, it is easy to create new stations or workflows from scratch, adapt existing templates to update the experimental protocols, or to fine-tune the code to fit specific needs. Using this framework, we developed the code for two different workflows and evaluated them using external quality assessment (EQA) samples from the European Molecular Genetics Quality Network (EMQN). The affordability of this platform makes automated SARS-CoV-2 PCR testing accessible for most laboratories and hospitals with qualified bioinformatics personnel. This platform also allows for flexibility, as it is not dependent on any specific commercial kit, and thus it can be quickly adapted to protocol changes, reagent, consumable shortages, or any other temporary material constraints.


Subject(s)
COVID-19 Nucleic Acid Testing/instrumentation , SARS-CoV-2/isolation & purification , Clinical Coding , Early Diagnosis , Humans , RNA, Viral/genetics , Real-Time Polymerase Chain Reaction/instrumentation , Reverse Transcriptase Polymerase Chain Reaction/instrumentation , Robotics , SARS-CoV-2/genetics , Workflow
3.
Diagn Microbiol Infect Dis ; 100(4): 115381, 2021 Aug.
Article in English | MEDLINE | ID: covidwho-1269260

ABSTRACT

To compare the practicability (usability and satisfaction) and analytical performances of VitaPCR™ Flu A&B Assay (Credo Diagnostics Biomedical Pte. Ltd., Singapore, Republic of Singapore) and Xpert® Xpress Flu/RSV kit (Cepheid, Sunnyvale, USA), two rapid point-of-care (POC) nucleic acid amplification tests (NAATs) by reference to multiplex RT-PCR for respiratory viruses. Nasopharyngeal swabs (n=117) were collected from patients with influenza-like illness in Paris, France. Thawed specimens were further analyzed with both NAATs. The usability was comparable for both NAATs. Satisfaction questionnaire was better for the VitaPCR™ platform for the short time of test result in 20 minutes. Both NAATs showed comparable sensitivities (VitaPCRTM: 95.0%; Xpert® Xpress: 97.5%) and specificities (100%) for influenza A/B RNA detection, with excellent reliability and accuracy between both NAATs. Both VitaPCR™ and Xpert® Xpress NAATs can be implemented in hospital setting as POC NAATs to rapidly detect influenza A/B RNA in symptomatic patients.


Subject(s)
Molecular Diagnostic Techniques/instrumentation , Reagent Kits, Diagnostic/standards , Real-Time Polymerase Chain Reaction/instrumentation , Viruses/genetics , Humans , Influenza A virus/genetics , Influenza, Human/diagnosis , Influenza, Human/virology , Molecular Diagnostic Techniques/methods , Nasopharynx/virology , Point-of-Care Testing/standards , Real-Time Polymerase Chain Reaction/standards , Reproducibility of Results , Sensitivity and Specificity , Viruses/classification , Viruses/isolation & purification
4.
Mol Diagn Ther ; 25(4): 495-504, 2021 07.
Article in English | MEDLINE | ID: covidwho-1231951

ABSTRACT

BACKGROUND: Molecular tests are the mainstay of detecting severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). However, their accessibility can be limited by the long examination time and inability to evaluate multiple samples at once. OBJECTIVE: This study evaluated the analytical performance of the newly developed rapid molecular assays GENECUBE® HQ SARS-CoV-2 and GENECUBE® FLU A/B. METHOD: This prospective study was conducted between 14 December 2020 and 9 January 2021 at a polymerase chain reaction (PCR) center. Samples were collected from the nasopharynx with flocked swabs. Molecular tests were performed with the GENECUBE® system and reference reverse transcription (RT)-PCR, and the results of the two assays were compared. RESULT: Among 1065 samples, 81 (7.6%) were positive for SARS-CoV-2 on the reference RT-PCR. Three showed discordance between GENECUBE® HQ SARS-CoV-2 and the reference RT-PCR; the total, positive, and negative samples of concordance for the two assays were 99.7%, 100%, and 99.7%, respectively. All discordant cases were positive with GENECUBE® HQ SARS-CoV-2 and negative with the reference RT-PCR. SARS-CoV-2 was detected in all three samples using another molecular assay for SARS-CoV-2. For GENECUBE® FLU A/B, the total, positive, and negative samples of concordance for the two assays were 99.5%, 100%, and 99.1%. CONCLUSION: The GENECUBE® HQ SARS-CoV-2 and GENECUBE® FLU A/B demonstrated sufficient analytical performance to detect SARS-CoV-2 and influenza virus A/B.


Subject(s)
COVID-19 Nucleic Acid Testing/methods , COVID-19/diagnosis , Real-Time Polymerase Chain Reaction/methods , SARS-CoV-2/genetics , Benchmarking , COVID-19/virology , COVID-19 Nucleic Acid Testing/instrumentation , Humans , Nasopharynx/virology , Prospective Studies , Real-Time Polymerase Chain Reaction/instrumentation , SARS-CoV-2/isolation & purification , Sensitivity and Specificity
5.
J Appl Lab Med ; 6(2): 421-428, 2021 03 01.
Article in English | MEDLINE | ID: covidwho-1120035

ABSTRACT

BACKGROUND: Detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) by reverse transcription PCR is the primary method to diagnose coronavirus disease 2019 (COVID-19). However, the analytic sensitivity required is not well defined and it is unclear how available assays compare. METHODS: For the Abbott RealTime SARS-CoV-2 assay (m2000; Abbott Molecular), we determined that it could detect viral concentrations as low as 26 copies/mL, we defined the relationship between cycle number and viral concentrations, and we tested naso- and oropharyngeal swab specimens from 8538 consecutive individuals. Using the m2000 as a reference assay method, we described the distribution of viral concentrations in these patients. We then used selected clinical specimens to determine the positive percent agreement of 2 other assays with more rapid turnaround times [Cepheid Xpert Xpress (GeneXpert; Cepheid); n = 27] and a laboratory developed test on the Luminex ARIES system [ARIES LDT (Luminex); n = 50] as a function of virus concentrations, from which we projected their false-negative rates in our patient population. RESULTS: SARS-CoV-2 was detected in 27% (95% CI: 26%-28%) of all specimens. Estimated viral concentrations were widely distributed, and 17% (95% CI: 16%-19%) of positive individuals had viral concentrations <845 copies/mL. Positive percent agreement was strongly related to viral concentration, and reliable detection (i.e., ≥95%) was observed at concentrations >100 copies/mL for the GeneXpert but not the ARIES LDT, corresponding to projected false-negative rates of 4% (95% CI: 0%-21%) and 27% (95% CI: 11%-46%), respectively. CONCLUSIONS: Substantial proportions of clinical specimens have low to moderate viral concentrations and may be missed by methods with less analytic sensitivity.


Subject(s)
COVID-19 Nucleic Acid Testing/instrumentation , COVID-19/diagnosis , Reagent Kits, Diagnostic , Real-Time Polymerase Chain Reaction/instrumentation , SARS-CoV-2/isolation & purification , Adult , Aged , Aged, 80 and over , COVID-19/virology , False Negative Reactions , Female , Humans , Limit of Detection , Male , Middle Aged , RNA, Viral/isolation & purification , Reproducibility of Results , Retrospective Studies , SARS-CoV-2/genetics
6.
Front Cell Infect Microbiol ; 11: 613304, 2021.
Article in English | MEDLINE | ID: covidwho-1088903

ABSTRACT

Background: The emerging Coronavirus Disease-2019 (COVID-19) has challenged the public health globally. With the increasing requirement of detection for SARS-CoV-2 outside of the laboratory setting, a rapid and precise Point of Care Test (POCT) is urgently needed. Methods: Targeting the nucleocapsid (N) gene of SARS-CoV-2, specific primers, and probes for reverse transcription recombinase-aided amplification coupled with lateral flow dipstick (RT-RAA/LFD) platform were designed. For specificity evaluation, it was tested with human coronaviruses, human influenza A virus, influenza B viruses, respiratory syncytial virus, and hepatitis B virus, respectively. For sensitivity assay, it was estimated by templates of recombinant plasmid and pseudovirus of SARS-CoV-2 RNA. For clinical assessment, 100 clinical samples (13 positive and 87 negatives for SARS-CoV-2) were tested via quantitative reverse transcription PCR (RT-qPCR) and RT-RAA/LFD, respectively. Results: The limit of detection was 1 copies/µl in RT-RAA/LFD assay, which could be conducted within 30 min at 39°C, without any cross-reaction with other human coronaviruses and clinical respiratory pathogens. Compared with RT-qPCR, the established POCT assay offered 100% specificity and 100% sensitivity in the detection of clinical samples. Conclusion: This work provides a convenient POCT tool for rapid screening, diagnosis, and monitoring of suspected patients in SARS-CoV-2 endemic areas.


Subject(s)
COVID-19 Nucleic Acid Testing/methods , COVID-19/diagnosis , Real-Time Polymerase Chain Reaction/methods , SARS-CoV-2/genetics , COVID-19/virology , COVID-19 Nucleic Acid Testing/instrumentation , Coronavirus Nucleocapsid Proteins/genetics , DNA Primers/genetics , Humans , Phosphoproteins/genetics , Point-of-Care Testing , RNA, Viral/genetics , Real-Time Polymerase Chain Reaction/instrumentation , Recombinases/metabolism , Reverse Transcription , SARS-CoV-2/isolation & purification , Sensitivity and Specificity
7.
Jpn J Infect Dis ; 74(1): 29-34, 2021 Jan 22.
Article in English | MEDLINE | ID: covidwho-1049199

ABSTRACT

The disease caused by severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) in Wuhan, China, in December 2019, has rapidly spread worldwide. SARS-CoV-2 is usually detected via real-time reverse-transcription polymerase chain reaction (RT-PCR). However, the increase in specimen load in institutions/hospitals necessitates a simpler detection system. Here, we present an ultra-rapid, real-time RT-PCR assay for SARS-CoV-2 detection using PCR1100 device. Although PCR1100 tests only one specimen at a time, the amplification period is less than 20 min and the sensitivity and specificity match those of conventional real-time RT-PCR performed on large instruments. The method is potentially helpful when daily multiple SARS-CoV-2 testing is needed, for example to confirm virus-free status prior to patient discharge.


Subject(s)
COVID-19 Testing/instrumentation , COVID-19/virology , Real-Time Polymerase Chain Reaction/instrumentation , Reverse Transcriptase Polymerase Chain Reaction/instrumentation , SARS-CoV-2/isolation & purification , COVID-19 Testing/methods , Humans , Real-Time Polymerase Chain Reaction/methods , Reverse Transcriptase Polymerase Chain Reaction/methods , SARS-CoV-2/genetics , Sensitivity and Specificity
8.
Talanta ; 224: 121850, 2021 Mar 01.
Article in English | MEDLINE | ID: covidwho-919513

ABSTRACT

In detecting infectious diseases, such as coronavirus 2019 (COVID-19), real-time reverse-transcription polymerase chain reaction (RT-PCR) is one of the most important technologies for RNA detection and disease diagnosis. To achieve high quality assurance, appropriate positive and negative controls are critical for disease detection using RT-PCR kits. In this study, we have found that commercial kits often adopt DNAs instead of RNAs as the positive controls, which can't report the kit problems in reverse transcription, thereby increasing risk of the false negative results when testing patient samples. To face the challenge, we have proposed and developed the chemically modified RNAs, such as phosphoroselenaote and phosphorothioate RNAs (Se-RNA and S-RNA), as the controls. We have found that while demonstrating the high thermostability, biostability, chemostability and exclusivity (or specificity), both Se-RNA and S-RNA can be fine templates for reverse transcription, indicating their potentials as both positive and negative controls for RT-PCR kits.


Subject(s)
COVID-19 Nucleic Acid Testing/methods , COVID-19/diagnosis , RNA, Viral/analysis , Real-Time Polymerase Chain Reaction/methods , Reverse Transcriptase Polymerase Chain Reaction/methods , COVID-19 Nucleic Acid Testing/instrumentation , DNA, Viral/analysis , False Negative Reactions , Humans , RNA Stability , RNA, Viral/chemistry , Reagent Kits, Diagnostic , Real-Time Polymerase Chain Reaction/instrumentation , Reverse Transcriptase Polymerase Chain Reaction/instrumentation , SARS-CoV-2/genetics
9.
J Appl Lab Med ; 5(6): 1307-1312, 2020 11 01.
Article in English | MEDLINE | ID: covidwho-696741

ABSTRACT

BACKGROUND: Numerous nucleic acid amplification assays utilizing different target genes of the SARS-CoV-2 genome have received emergency use authorization (EUA) by the United States Food and Drug Administration (FDA). Limited data are available comparing the test performance characteristics of these assays. METHODS: A diagnostic comparison study was performed to evaluate the performance of the Cepheid Xpert Xpress SARS-CoV-2 assay compared to the Hologic Panther Fusion SARS-CoV-2 assay using clinical nasopharyngeal specimens. Agreement between the two assays was assessed by overall, positive, and negative percent agreement and Cohen's kappa coefficient. RESULTS: A total of 104 (54 positive and 50 negative) clinical nasopharyngeal samples were tested by both assays. Using the Panther Fusion as a reference standard, the Xpert demonstrated an overall agreement of 99.0% [95% confidence interval (CI): 94.8-100], positive percent agreement of 98.1% (95% CI: 90.1-100), and a negative percent agreement of 100% (95% CI: 94.2-100). The kappa coefficient was 0.98 (95% CI: 0.94-1.0). One sample positive by the Panther Fusion with a cycle threshold (Ct) of 38.6 was found to be reproducibly negative by the Xpert assay. CONCLUSIONS: The Cepheid Xpert Xpress SARS-CoV-2 assay provides test performance comparable to the Hologic Panther Fusion SARS-CoV-2 assay while offering laboratories rapid, on-demand testing capacity.


Subject(s)
Betacoronavirus/isolation & purification , Clinical Laboratory Techniques/instrumentation , Coronavirus Infections/diagnosis , Molecular Diagnostic Techniques/instrumentation , Pneumonia, Viral/diagnosis , RNA, Viral/isolation & purification , Automation, Laboratory/instrumentation , Betacoronavirus/genetics , COVID-19 , COVID-19 Testing , Clinical Laboratory Techniques/statistics & numerical data , Coronavirus Infections/epidemiology , Coronavirus Infections/virology , Humans , Molecular Diagnostic Techniques/statistics & numerical data , Multiplex Polymerase Chain Reaction/instrumentation , Pandemics , Pneumonia, Viral/epidemiology , Pneumonia, Viral/virology , Point-of-Care Systems/statistics & numerical data , Reagent Kits, Diagnostic/statistics & numerical data , Real-Time Polymerase Chain Reaction/instrumentation , Reproducibility of Results , SARS-CoV-2 , Time Factors , United States/epidemiology
10.
Virulence ; 11(1): 964-967, 2020 12.
Article in English | MEDLINE | ID: covidwho-690823

ABSTRACT

Currently, testing for coronavirus is performed with time and personnel consuming PCR assays. The aim of this study was to evaluate the sensitivity, specificity and capacity of a fully automated, random access high-throughput real-time PCR-based diagnostic platform for the detection of SARS-CoV-2. The NeuMoDx N96 system displayed an equal or better detection rate for SARS-CoV-2 compared with the LightCycler 480II system and showed a specificity of 100%. The median PCR run time for all 28 PCR runs was 91 (IQR 84-97) minutes. The capacity of the NeuMoDx N96 could easily surpass the capacity of most currently used molecular test systems and significantly reduce the turn-around time.


Subject(s)
Betacoronavirus/isolation & purification , High-Throughput Nucleotide Sequencing/methods , RNA, Viral/isolation & purification , Real-Time Polymerase Chain Reaction/methods , Betacoronavirus/genetics , High-Throughput Nucleotide Sequencing/instrumentation , High-Throughput Nucleotide Sequencing/standards , Humans , RNA, Viral/genetics , Real-Time Polymerase Chain Reaction/instrumentation , Real-Time Polymerase Chain Reaction/standards , Reproducibility of Results , SARS-CoV-2 , Sensitivity and Specificity , Time Factors
11.
Front Cell Infect Microbiol ; 10: 356, 2020.
Article in English | MEDLINE | ID: covidwho-642075

ABSTRACT

CDC and WHO guidelines for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) diagnosis only recommend synthetic fiber swabs for nasopharyngeal (NP) sampling. We show that cotton-tipped plastic swabs do not inhibit PCR and have equivalent performance to rayon swabs. Cotton-tipped plastic swabs are massively produced worldwide and would prevent swab supply shortages under the current high SARS-CoV-2 testing demands, particularly in developing countries.


Subject(s)
Betacoronavirus/isolation & purification , Clinical Laboratory Techniques/instrumentation , Coronavirus Infections/diagnosis , Diagnostic Equipment/supply & distribution , Disposable Equipment/supply & distribution , Pneumonia, Viral/diagnosis , Real-Time Polymerase Chain Reaction/instrumentation , Betacoronavirus/genetics , COVID-19 , COVID-19 Testing , Cellulose/supply & distribution , Clinical Laboratory Techniques/methods , Coronavirus Infections/virology , Cotton Fiber/supply & distribution , Humans , Nasopharynx , Pandemics , Plastics/supply & distribution , Pneumonia, Viral/virology , RNA, Viral/analysis , Real-Time Polymerase Chain Reaction/methods , SARS-CoV-2 , Specimen Handling/instrumentation , Specimen Handling/methods
12.
Euro Surveill ; 25(24)2020 06.
Article in English | MEDLINE | ID: covidwho-605372

ABSTRACT

Containment strategies and clinical management of coronavirus disease (COVID-19) patients during the current pandemic depend on reliable diagnostic PCR assays for the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Here, we compare 11 different RT-PCR test systems used in seven diagnostic laboratories in Germany in March 2020. While most assays performed well, we identified detection problems in a commonly used assay that may have resulted in false-negative test results during the first weeks of the pandemic.


Subject(s)
Betacoronavirus/genetics , Clinical Laboratory Techniques/methods , Coronavirus Infections/diagnosis , Diagnostic Equipment , Pneumonia, Viral/diagnosis , COVID-19 , COVID-19 Testing , COVID-19 Vaccines , Clinical Laboratory Techniques/instrumentation , Feces/virology , Germany , Humans , Laboratories , Multiplex Polymerase Chain Reaction/instrumentation , Multiplex Polymerase Chain Reaction/methods , Pandemics , Real-Time Polymerase Chain Reaction/instrumentation , Real-Time Polymerase Chain Reaction/methods , Reverse Transcriptase Polymerase Chain Reaction/instrumentation , Reverse Transcriptase Polymerase Chain Reaction/methods , SARS-CoV-2 , Sensitivity and Specificity
13.
PLoS One ; 15(6): e0234682, 2020.
Article in English | MEDLINE | ID: covidwho-595220

ABSTRACT

Novel Corona virus/Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2 or 2019-nCoV), and the subsequent disease caused by the virus (coronavirus disease 2019 or COVID-19), is an emerging global health concern that requires a rapid diagnostic test. Quantitative reverse transcription PCR (qRT-PCR) is currently the standard for SARS-CoV-2 detection; however, Reverse Transcription Loop-Mediated Isothermal Amplification (RT-LAMP) may allow for faster and cheaper field based testing at point-of-risk. The objective of this study was to develop a rapid screening diagnostic test that could be completed in 30-45 minutes. Simulated patient samples were generated by spiking serum, urine, saliva, oropharyngeal swabs, and nasopharyngeal swabs with a portion of the SARS-CoV-2 nucleic sequence. RNA isolated from nasopharyngeal swabs collected from actual COVID-19 patients was also tested. The samples were tested using RT-LAMP as well as by conventional qRT-PCR. Specificity of the RT-LAMP was evaluated by also testing against other related coronaviruses. RT-LAMP specifically detected SARS-CoV-2 in both simulated patient samples and clinical specimens. This test was performed in 30-45 minutes. This approach could be used for monitoring of exposed individuals or potentially aid with screening efforts in the field and potential ports of entry.


Subject(s)
Betacoronavirus/isolation & purification , Coronavirus Infections/diagnosis , Molecular Diagnostic Techniques/methods , Nucleic Acid Amplification Techniques/methods , Pneumonia, Viral/diagnosis , Point-of-Care Testing , Real-Time Polymerase Chain Reaction/methods , Betacoronavirus/genetics , COVID-19 , Coronavirus Infections/virology , DNA Primers , Humans , Molecular Diagnostic Techniques/economics , Molecular Diagnostic Techniques/instrumentation , Nucleic Acid Amplification Techniques/economics , Nucleic Acid Amplification Techniques/instrumentation , Pandemics , Pneumonia, Viral/virology , Real-Time Polymerase Chain Reaction/economics , Real-Time Polymerase Chain Reaction/instrumentation , SARS-CoV-2 , Sensitivity and Specificity , Time Factors
14.
Clin Microbiol Infect ; 26(8): 1076-1081, 2020 Aug.
Article in English | MEDLINE | ID: covidwho-547614

ABSTRACT

OBJECTIVE: To evaluate the performance of an ultrafast single-tube nucleic acid isothermal amplification detection assay for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) RNA using clinical samples from multiple centres. METHODS: A reverse transcription recombinase-aided amplification (RT-RAA) assay for SARS-CoV-2 was conducted within 15 minutes at 39°C with portable instruments after addition of extracted RNA. The clinical performance of RT-RAA assay was evaluated using 947 clinical samples from five institutions in four regions of China; approved commercial fluorescence quantitative real-time PCR (qRT-PCR) kits were used for parallel detection. The sensitivity and specificity of RT-RAA were compared and analysed. RESULTS: The RT-RAA test results of 926 samples were consistent with those of qRT-PCR (330 were positive, 596 negative); 21 results were inconsistent. The sensitivity and specificity of RT-RAA was 97.63% (330/338, 95% confidence interval (CI) 95.21 to 98.90) and 97.87% (596/609, 95% CI 96.28 to 98.81) respectively. The positive and negative predictive values were 96.21% (330/343, 95% CI 93.45 to 97.88) and 98.68% (596/604, 95% CI 97.30 to 99.38) respectively. The total coincidence rate was 97.78% (926/947, 95% CI 96.80 to 98.70), and the kappa was 0.952 (p < 0.05). CONCLUSIONS: With comparable sensitivity and specificity to the commercial qRT-PCR kits, RT-RAA assay for SARS-CoV-2 exhibited the distinctive advantages of simplicity and rapidity in terms of operation and turnaround time.


Subject(s)
Betacoronavirus/genetics , Coronavirus Infections/diagnosis , Pneumonia, Viral/diagnosis , RNA, Viral/genetics , Real-Time Polymerase Chain Reaction/instrumentation , Adolescent , Adult , Aged , Aged, 80 and over , COVID-19 , Child , China , Diagnostic Tests, Routine/instrumentation , Female , Humans , Male , Middle Aged , Pandemics , SARS-CoV-2 , Sensitivity and Specificity , Temperature , Young Adult
15.
RNA ; 26(7): 771-783, 2020 07.
Article in English | MEDLINE | ID: covidwho-154676

ABSTRACT

The current COVID-19 pandemic presents a serious public health crisis, and a better understanding of the scope and spread of the virus would be aided by more widespread testing. Nucleic-acid-based tests currently offer the most sensitive and early detection of COVID-19. However, the "gold standard" test pioneered by the U.S. Centers for Disease Control and Prevention takes several hours to complete and requires extensive human labor, materials such as RNA extraction kits that could become in short supply, and relatively scarce qPCR machines. It is clear that a huge effort needs to be made to scale up current COVID-19 testing by orders of magnitude. There is thus a pressing need to evaluate alternative protocols, reagents, and approaches to allow nucleic-acid testing to continue in the face of these potential shortages. There has been a tremendous explosion in the number of papers written within the first weeks of the pandemic evaluating potential advances, comparable reagents, and alternatives to the "gold-standard" CDC RT-PCR test. Here we present a collection of these recent advances in COVID-19 nucleic acid testing, including both peer-reviewed and preprint articles. Due to the rapid developments during this crisis, we have included as many publications as possible, but many of the cited sources have not yet been peer-reviewed, so we urge researchers to further validate results in their own laboratories. We hope that this review can urgently consolidate and disseminate information to aid researchers in designing and implementing optimized COVID-19 testing protocols to increase the availability, accuracy, and speed of widespread COVID-19 testing.


Subject(s)
Betacoronavirus/isolation & purification , Clinical Laboratory Techniques , Nucleic Acid Amplification Techniques/methods , Real-Time Polymerase Chain Reaction/methods , Betacoronavirus/genetics , COVID-19 Testing , CRISPR-Cas Systems , Centers for Disease Control and Prevention, U.S. , Clustered Regularly Interspaced Short Palindromic Repeats , Coronavirus Infections/diagnosis , Humans , Nasopharynx/virology , Nucleic Acid Amplification Techniques/instrumentation , Point-of-Care Systems , RNA, Viral/genetics , RNA, Viral/isolation & purification , Real-Time Polymerase Chain Reaction/instrumentation , SARS-CoV-2 , Time Factors , United States , Workflow
16.
J Clin Virol ; 128: 104390, 2020 07.
Article in English | MEDLINE | ID: covidwho-133461

ABSTRACT

BACKGROUND: The ongoing SARS-CoV-2 pandemic presents a unique challenge for diagnostic laboratories around the world. Automation of workflows in molecular diagnostics is instrumental for coping with the large number of tests ordered by clinicians, as well as providing fast-tracked rapid testing for highly urgent cases. In this study we evaluated a SARS-CoV-2 LDT for the NeuMoDx 96 system, a fully automated device performing extraction and real-time PCR. METHODS: A publicly available SARS-CoV-2 RT-PCR assay was adapted for the automated system. Analytical performance was evaluated using in-vitro transcribed RNA and clinical performance was compared to the cobas 6800-based reference assay within the lab. RESULTS: The Envelope (E) Gene-LDT displayed good analytical performance with an LoD of 95.55 cp/mL and no false positives during evaluation of cross-reactivity. A total of 176 patient samples were tested with both the E-Gene-LDT and the reference assay. Positive and negative agreement were 100 % and 99.2 % respectively. Invalid-rate was 6.3 %. CONCLUSION: The E-Gene-LDT showed analytical and clinical performance comparable to the cobas6800-based reference assay. Due to its random-access workflow concept and rapid time-to-result of about 80 min, the system is very well suited for providing fast-tracked SARS-CoV-2 diagnostics for urgent clinical samples in the hospital setting.


Subject(s)
Clinical Laboratory Techniques/methods , Pandemics , Reverse Transcriptase Polymerase Chain Reaction/methods , Betacoronavirus , COVID-19 , COVID-19 Testing , COVID-19 Vaccines , Clinical Laboratory Techniques/instrumentation , Coronavirus Infections/diagnosis , Hospitals , Humans , Pneumonia, Viral , Real-Time Polymerase Chain Reaction/instrumentation , Real-Time Polymerase Chain Reaction/methods , Reverse Transcriptase Polymerase Chain Reaction/instrumentation , SARS-CoV-2 , Time Factors , Workflow
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