scholarly journals Clinical Evaluation of Three Sample-to-Answer Platforms for Detection of SARS-CoV-2

2020 ◽  
Vol 58 (8) ◽  
Author(s):  
Wei Zhen ◽  
Elizabeth Smith ◽  
Ryhana Manji ◽  
Deborah Schron ◽  
Gregory J. Berry
Keyword(s):  
Severe Acute Respiratory Syndrome ◽  
Drug Administration ◽  
Clinical Evaluation ◽  
Limit Of Detection ◽  
Clinical Performance ◽  
Molecular Diagnostic ◽  
Analytical Sensitivity ◽  
Reference Standard ◽  
Slight Improvement ◽  
Percent Agreement

ABSTRACT Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has now spread across the globe. As part of the worldwide response, many molecular diagnostic platforms have been granted emergency use authorization (EUA) by the Food and Drug Administration (FDA) to identify SARS-CoV-2 positive patients. Our objective was to evaluate three sample-to-answer molecular diagnostic platforms (Cepheid Xpert Xpress SARS-CoV-2 [Xpert Xpress], Abbott ID NOW COVID-19 [ID NOW], and GenMark ePlex SARS-CoV-2 Test [ePlex]) to determine analytical sensitivity, clinical performance, and workflow for the detection of SARS-CoV-2 in nasopharyngeal swabs from 108 symptomatic patients. We found that Xpert Xpress had the lowest limit of detection (100% detection at 100 copies/ml), followed by ePlex (100% detection at 1,000 copies/ml), and ID NOW (20,000 copies/ml). Xpert Xpress also had highest positive percent agreement (PPA) compared to our reference standard (98.3%) followed by ePlex (91.4%) and ID NOW (87.7%). All three assays showed 100% negative percent agreement (NPA). In the workflow analysis, ID NOW produced the lowest time to result per specimen (∼17 min) compared to Xpert Xpress (∼46 min) and ePlex (∼1.5 h), but what ID NOW gained in rapid results, it lost in analytical and clinical performance. ePlex had the longest time to results and showed a slight improvement in PPA over ID NOW. Information about the clinical and analytical performance of these assays, as well as workflow, will be critical in making informed and timely decisions on testing platforms.

scholarly journals Analytical and Clinical Comparison of Three Nucleic Acid Amplification Tests for SARS-CoV-2 Detection

2020 ◽  
Vol 58 (9) ◽  
Author(s):  
Elizabeth Smith ◽  
Wei Zhen ◽  
Ryhana Manji ◽  
Deborah Schron ◽  
Scott Duong ◽  
...  
Keyword(s):  
Nucleic Acid ◽  
Severe Acute Respiratory Syndrome ◽  
Drug Administration ◽  
Clinical Performance ◽  
Absolute Quantification ◽  
Nucleic Acid Amplification ◽  
Clinical Correlation ◽  
Molecular Assays ◽  
Percent Agreement ◽  
Rna Transcript

ABSTRACT Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) was first identified in December 2019 and has quickly become a worldwide pandemic. In response, many diagnostic manufacturers have developed molecular assays for SARS-CoV-2 under the Food and Drug Administration (FDA) Emergency Use Authorization (EUA) pathway. This study compared three of these assays, the Hologic Panther Fusion SARS-CoV-2 assay (Fusion), the Hologic Aptima SARS-CoV-2 assay (Aptima), and the BioFire Defense COVID-19 test (BioFire), to determine analytical and clinical performance as well as workflow. All three assays showed similar limits of detection (LODs) using inactivated virus, with 100% detection, ranging from 500 to 1,000 genome equivalents/ml, whereas use of a quantified RNA transcript standard showed the same trend but had values ranging from 62.5 to 125 copies/ml, confirming variability in absolute quantification of reference standards. The clinical correlation found that the Fusion and BioFire assays had a positive percent agreement (PPA) of 98.7%, followed by the Aptima assay at 94.7%, compared to the consensus result. All three assays exhibited 100% negative percent agreement (NPA). Analysis of discordant results revealed that all four samples missed by the Aptima assay had cycle threshold (Ct) values of >37 by the Fusion assay. In conclusion, while all three assays showed similar relative LODs, we showed differences in absolute LODs depending on which standard was employed. In addition, the Fusion and BioFire assays showed better clinical performance, while the Aptima assay showed a modest decrease in overall PPA. These findings should be kept in mind when making platform testing decisions.

scholarly journals Analytical and Clinical Comparison of Three Nucleic Acid Amplification Tests for SARS-CoV-2 Detection

2020 ◽  
Author(s):  
Elizabeth Smith ◽  
Wei Zhen ◽  
Ryhana Manji ◽  
Deborah Schron ◽  
Scott Duong ◽  
...  
Keyword(s):  
Nucleic Acid ◽  
Severe Acute Respiratory Syndrome ◽  
Limit Of Detection ◽  
Clinical Performance ◽  
Absolute Quantification ◽  
Nucleic Acid Amplification ◽  
Clinical Correlation ◽  
Molecular Assays ◽  
Percent Agreement ◽  
Rna Transcript

AbstractSevere Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) was first identified in December 2019 and has quickly become a worldwide pandemic. In response, many diagnostic manufacturers have developed molecular assays for SARS-CoV-2 under the Food and Drug Administration (FDA) Emergency Use Authorization (EUA) pathway. This study compared three of these assays: the Hologic Panther Fusion SARS-CoV-2 assay (Fusion), the Hologic Aptima SARS-CoV-2 assay (Aptima) and the BioFire Diagnostics COVID-19 test (BioFire), to determine analytical and clinical performance, as well as workflow. All three assays showed a similar limit of detection (LOD) using inactivated virus, with 100% detection ranging from 500-1,000 genome equivalents/ml, whereas use of a quantified RNA transcript standard showed the same trend, but had values ranging from 62.5 to 125 copies/ml, confirming variability in absolute quantification of reference standards. The clinical correlation found that the Fusion and BioFire assays had a positive percent agreement (PPA) of 98.7%, followed by the Aptima assay at 94.7% when compared to the consensus result. All three assays exhibited 100% negative percent agreement (NPA). Analysis of discordant results revealed that all four samples missed by the Aptima assay had Ct values >37 on the Fusion assay.In conclusion, while all three assays showed similar relative LODs, we showed differences in absolute LODs depending on which standard was employed. In addition, the Fusion and BioFire assays showed better clinical performance, while the Aptima assay showed a modest decrease in overall PPA. These findings should be kept in mind when making platform testing decisions.

scholarly journals Analytical and Clinical Evaluation of “AccuPower SARS-CoV-2 Multiplex RT-PCR kit (Bioneer, South Korea)” and “Allplex 2019-nCoV Assay (Seegene, South Korea)” for SARS-CoV-2 RT-PCR Diagnosis: Korean CDC EUA as a Quality Control Proxy for Developing Countries

2021 ◽  
Vol 11 ◽  
Author(s):  
Byron Freire-Paspuel ◽  
Miguel Angel Garcia-Bereguiain
Keyword(s):  
Developing Countries ◽  
Quality Control ◽  
South Korea ◽  
Clinical Evaluation ◽  
Limit Of Detection ◽  
Clinical Performance ◽  
High Sensitivity ◽  
Analytical Sensitivity ◽  
Rt Pcr ◽  
Pcr Diagnosis

BackgroundMultiple RT-qPCR kits are available in the market for SARS-CoV-2 diagnosis, some of them with Emergency Use Authorization (EUA) by FDA or their country of origin agency, but many of them lack of proper clinical evaluation.ObjectiveWe evaluated the clinical performance of two Korean SARS-CoV-2 RT-PCR kits available in South America, AccuPower SARS-CoV-2 Multiplex RT-PCR kit (Bioneer, South Korea) and Allplex 2019-nCoV Assay (Seegene, South Korea), for RT-qPCR SARS-CoV-2 diagnosis using the CDC protocol as a gold standard.ResultsWe found strong differences among both kits clinical performance and analytical sensitivity; while the Allplex 2019-nCoV Assay has sensitivity of 96.5% and an estimated limit of detection of 4,000 copies/ml, the AccuPower SARS-CoV-2 Multiplex RT-PCR kit has a sensitivity of 75.5% and limit of detection estimated to be bigger than 20,000 copies/ml.ConclusionsAccuPower SARS-CoV-2 Multiplex RT-PCR kit and Allplex 2019-nCoV Assay are both made in South Korea but EUA by Korean CDC was only granted to the later. Our results support that Korean CDC EUA should be considered as a quality control proxy for Korean SARS-CoV-2 RT-PCR kits prior to importation by developing countries to guarantee high sensitivity diagnosis.

Analytical and clinical validation of a novel amplicon-based NGS assay for the evaluation of circulating tumor DNA in metastatic colorectal cancer patients

2019 ◽  
Vol 57 (10) ◽  
pp. 1501-1510 ◽  
Author(s):  
Beili Wang ◽  
Shengchao Wu ◽  
Fei Huang ◽  
Minna Shen ◽  
Huiqin Jiang ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Metastatic Colorectal Cancer ◽  
Limit Of Detection ◽  
Clinical Performance ◽  
Circulating Tumor Dna ◽  
Concordance Rate ◽  
Tissue Samples ◽  
Percent Agreement ◽  
Pre Treatment ◽  
Tumor Dna

Abstract Background Evaluating the tumor RAS/BRAF status is important for treatment selection and prognosis assessment in metastatic colorectal cancer (mCRC) patients. Correction of artifacts from library preparation and sequencing is essential for accurately analyzing circulating tumor DNA (ctDNA) mutations. Here, we assessed the analytical and clinical performance of a novel amplicon-based next-generation sequencing (NGS) assay, Firefly™, which employs a concatemer-based error correction strategy. Methods Firefly assay targeting KRAS/NRAS/BRAF/PIK3CA was evaluated using cell-free DNA (cfDNA) reference standards and cfDNA samples from 184 mCRC patients. Plasma results were compared to the mutation status determined by ARMS-based PCR from matched tissue. Samples with a mutation abundance below the limit of detection (LOD) were retested again by droplet digital polymerase chain reaction (ddPCR) or NGS. Results The Firefly assay demonstrated superior sensitivity and specificity with a 98.89% detection rate at an allele frequency (AF) of 0.2% for 20 ng cfDNA. Generally, 40.76% and 48.37% of the patients were reported to be positive by NGS of plasma cfDNA and ARMS of FFPE tissue, respectively. The concordance rate between the two platforms was 80.11%. In the pre-treatment cohort, the concordance rate between plasma and tissue was 93.33%, based on the 17 common exons that Firefly™ and ARMS genotyped, and the positive percent agreement (PPA) and negative percent agreement (NPA) for KRAS/NRAS/BRAF/PIK3CA were 100% and 99.60%, respectively. Conclusions Total plasma cfDNA detected by Firefly offers a viable complement for mutation profiling in CRC patients, given the high agreement with matched tumor samples. Together, these data demonstrate that Firefly could be routinely applied for clinical applications in mCRC patients.

scholarly journals Design and Validation of Transcription-Mediated-Amplification Nucleic Acid Amplification Tests forMycoplasma genitalium

2019 ◽  
Vol 57 (8) ◽  
Author(s):  
Brett Kirkconnell ◽  
Barbara Weinbaum ◽  
Katherine Santos ◽  
Trisha Le Nguyen ◽  
Bryan Vinluan ◽  
...  
Keyword(s):  
Clinical Performance ◽  
Mycoplasma Genitalium ◽  
Vaginal Swab ◽  
Nucleic Acid Amplification ◽  
Clinical Validation ◽  
Analytical Sensitivity ◽  
Reference Standard ◽  
Content Type ◽  
Sensitivity Specificity

ABSTRACTMycoplasma genitaliumis a sexually transmitted bacterium linked to adverse sexual and reproductive health outcomes in women and men.M. genitaliumis difficult to culture, and in the absence of validated amplified molecular methods for diagnosis of infection, there is no reference standard available for use as a comparator for the validation of newM. genitaliumdiagnostic tests. We evaluated the analytical and clinical performance of three transcription-mediated amplification (TMA) tests forM. genitalium, each targeting unique rRNA sequences, for use as a composite comparator for clinical validation of the AptimaMycoplasma genitalium(AMG) assay, anin vitrodiagnostic (IVD) TMA test that targets 16 s rRNA ofM. genitalium. Analytical sensitivity, specificity, and strain inclusivity of all four TMA tests were determined using nine laboratory strains ofM. genitaliumand 56 nontarget bacteria, protozoa, and viruses. Analytical sensitivity of the tests forM. genitaliumranged from 0.017 to 0.040 genome equivalents/ml. None of the nontarget organisms evaluated cross-reacted with any test. A composite comparator reference standard consisting of the 3 alternate (Alt) TMA tests was used to evaluate the clinical performance of the AMG assay by testing residual vaginal swab, female urine, and male urine specimens obtained from 1,400 adult subjects from three U.S. clinical sites. Using this reference standard to establish infected specimen status, the sensitivity, specificity, and overall agreement of the AMG IVD assay were 100%, 99.9%, and 99.9%, respectively. These results demonstrate the utility of molecular composite reference standard methodology for the clinical validation of future IVD tests for this organism.

scholarly journals Verification and Validation of SARS-CoV-2 Assay Performance on the Abbott m2000 and Alinity m Systems

2021 ◽  
Author(s):  
Julie W. Hirschhorn ◽  
April Kegl ◽  
Tanisha Dickerson ◽  
W. Bailey Glen ◽  
Gang Xu ◽  
...  
Keyword(s):  
Limit Of Detection ◽  
Clinical Performance ◽  
Cycle Number ◽  
Percent Agreement ◽  
Assay Performance ◽  
Diagnostic Panel ◽  
Highly Correlated ◽  
Clinical Verification ◽  
Bridging Study ◽  
Bridging Studies

We verified the analytical performance of the Abbott RealTime SARS-CoV-2 assay on the m2000 system and compared its clinical performance to the CDC 2019-nCoV Real-Time PCR Diagnostic Panel and the ThermoFisher TaqPath RT-PCR COVID-19 kit. We also performed a bridging study comparing the RealTime SARS-CoV-2 assay with the new Abbott Alinity m SARS-CoV-2 assay. A number of standards, reference materials, and commercially available controls were used for the analytical verification to confirm the limit of detection, linearity, and reproducibility. We used nasopharyngeal (NP) swab specimens collected in saline for the clinical verification and bridging studies. Overall, we found 91.2% positive percent agreement (PPA) (95% CI 76.2 to 98.14%) and a 100% negative percent agreement (NPA) (95% CI 97.97 to 100%) between the results of the RealTime SARS-CoV-2 and CDC tests with 217 NP specimens (P=0.13). We found a PPA of 100% (95% CI 90.26 to 100%) and a NPA of 95.15% (95% CI 83.47 to 99.4%) between the results of the RealTime and TaqPath tests with 77 NP specimens (P=0.24). Finally, we tested 203 NP swab specimens for SARS-CoV-2 on the m2000 on the Alinity m systems. The PPA and NPA were 92.2% (95% CI, 85.3 to 96.59%) and 92% (95% PI, 84.8 to 96.5%), respectively (P=0.4). Although cycle number (Cn) values obtained for the concordant positive samples were highly correlated (R2, 0.95), the Cn values were on average 14.14 higher on the Alinity m system due to the unread cycles with the RealTime SARS-CoV-2 assay.

scholarly journals Comparative analytical evaluation of four centralized platforms for the detection of M. tuberculosis complex and resistance to rifampicin and isoniazid

2020 ◽  
Author(s):  
Margaretha de Vos ◽  
Lesley Scott ◽  
Anura David ◽  
Andre Trollip ◽  
Harald Hoffmann ◽  
...  
Keyword(s):  
Drug Resistance ◽  
Limit Of Detection ◽  
Clinical Performance ◽  
Analytical Data ◽  
Multidrug Resistant ◽  
Resistance Mechanisms ◽  
Analytical Sensitivity ◽  
Isoniazid Resistance ◽  
Mdr Tb ◽  
Resistance Detection

Failure to rapidly identify drug-resistant tuberculosis (TB) increases the risk of patient mismanagement, the amplification of drug resistance and ongoing transmission. We generated comparative analytical data for four automated assays for detection of TB and multidrug-resistant (MDR) TB: Abbott RealTime MTB and MTB RIF/INH (Abbott), Hain Lifescience FluoroType® MTBDR (Hain), BD MAX™ MDR-TB (BD) and Roche cobas® MTB and MTB-RIF/INH (Roche). We included Xpert MTB/RIF (Xpert) and GenoType MTBDRplus as comparators for TB and drug resistance detection, respectively. We assessed analytical sensitivity for the detection of Mycobacterium tuberculosis complex using inactivated strains (M. tuberculosis H37Rv and M. bovis) spiked into TB-negative sputa and computed the 95% limit of detection (LOD95). We assessed the accuracy for rifampicin and isoniazid resistance detection using well characterized M. tuberculosis strains with high-confidence mutations accounting for >85% of first-line resistance mechanisms globally. For H37Rv and M. bovis, respectively, we measured LOD95 values of 3,781 and 2,926 (Xpert); 322 and 2,182 (Abbott); 826 and 4,301 (BD); 10,398 and 23,139 (Hain); 2,416 and 2,136 (Roche) genomes/mL. Assays targeting multi-copy genes or targets (Abbott, BD and Roche) showed increased analytical sensitivity compared to Xpert. Quantification of the panel by quantitative real-time PCR prevents the determination of absolute values and results reported here can only be interpreted for comparison purposes. All assays showed accuracy comparable to Genotype MTBDRplus for the detection of rifampicin and isoniazid resistance. The data from this analytical study suggest that the assays may have similar clinical performance to WHO-recommended molecular TB and MDR-TB assays.

scholarly journals Clinical Validation of Automated and Rapid mariPOC SARS-CoV-2 Antigen Test

2021 ◽  
Author(s):  
Juha M. Koskinen ◽  
Petri Antikainen ◽  
Kristina Hotakainen ◽  
Anu Haveri ◽  
Niina Ikonen ◽  
...  
Keyword(s):  
Sensitivity And Specificity ◽  
Limit Of Detection ◽  
Clinical Performance ◽  
High Capacity ◽  
Nasopharyngeal Swab ◽  
Analytical Sensitivity ◽  
Clinical Sensitivity ◽  
Qrt Pcr ◽  
Antigen Tests ◽  
Novel Coronavirus

ABSTRACTNovel SARS coronavirus causing COVID-19 was recognized in late 2019. Diagnostics was quickly ramped up worldwide based on the detection of viral RNA. Based on the scientific knowledge for pre-existing coronaviruses, it was expected that the RNA of this novel coronavirus will be detected at significant rates from symptomatic and asymptomatic individuals due to existence of non-infectious RNA. To increase the efficacy of diagnostics, surveillance, screening and pandemic control, rapid methods, such as antigen tests, are needed for decentralized testing and to assess infectiousness. The objectives were to verify analytical sensitivity and specificity, and assess the clinical sensitivity, specificity and usability of a novel automated mariPOC SARS-CoV-2 test based on sophisticated optical laser technology detecting viral structure proteins. Analytical performance was verified using bacterial and viral preparations. Clinical performance of the test was evaluated against qRT-PCR in a retrospective study with nasopharyngeal swab specimens (N=211) collected from symptomatic patients suspected of acute SARS-CoV-2 infections. Sensitivity and specificity of the mariPOC test were 92.3% (12/13) and 100.0% (198/198), respectively. The test’s limit of detection was 22 PFU/test and it had no cross-reactions with the tested respiratory microbes. Our study shows that the mariPOC can detect infectious individuals already in 20 minutes while clinical sensitivity close to qRT-PCR is achieved in two hours or less. The test targets conserved epitopes of SARS-CoV-2 nucleoprotein, making it robust against strain variations. The new test is a promising and versatile tool for syndromic testing of symptomatic cases and for high capacity infection control screening.

scholarly journals Performance Evaluation of the SAMBA II SARS-CoV-2 Test for Point-of-Care Detection of SARS-CoV-2

2020 ◽  
Vol 59 (1) ◽  
pp. e01262-20 ◽  
Author(s):  
Sonny M. Assennato ◽  
Allyson V. Ritchie ◽  
Cesar Nadala ◽  
Neha Goel ◽  
Cuijuan Tie ◽  
...  
Keyword(s):  
Public Health ◽  
Infection Control ◽  
Patient Management ◽  
Clinical Performance ◽  
Point Of Care ◽  
Turnaround Time ◽  
Control Measures ◽  
Nucleic Acid Amplification ◽  
Analytical Sensitivity ◽  
Percent Agreement

ABSTRACTNucleic acid amplification for the detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) RNA in respiratory samples is the standard method for diagnosis. The majority of this testing is centralized and therefore has turnaround times of several days. Point-of-care (POC) testing with rapid turnaround times would allow more effective triage in settings where patient management and infection control decisions need to be made rapidly. The inclusivity and specificity of the Simple AMplification-Based Assay (SAMBA) II SARS-CoV-2 test were determined by both in silico analyses of the primers and probes and wet testing. The SAMBA II SARS-CoV-2 test was evaluated for performance characteristics. Clinical performance was evaluated in residual combined throat/nose swabs and compared to that of the Public Health England real-time PCR assay targeting the RdRp gene. The SAMBA II SARS-CoV-2 test has an analytical sensitivity of 250 copies/ml for detecting two regions of the genome (open reading frame 1ab [ORF1ab] and nucleocapsid protein [N]). The clinical performance was evaluated in 172 residual combined nose/throat swabs provided by the Clinical Microbiology and Public Health Laboratory, Addenbrooke’s Hospital, Cambridge (CMPHL), which showed an estimated positive percent agreement of 98.9% (95% confidence interval [CI], 93.83 to 99.97) and negative percent agreement of 96.4% (95% CI, 89.92 to 99.26) compared to testing by the CMPHL. The data show that the SAMBA II SARS-CoV-2 test performs equivalently to the centralized testing methods, but with a shorter turnaround time of 86 to 101 min. Point-of-care tests such as SAMBA should enable rapid patient management and effective implementation of infection control measures.

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