RESUMO
Coronavirus disease 2019 (COVID-19) is a novel respiratory illness caused by SARS-CoV-2. Viral entry is mediated through viral spike protein and host ACE2 enzyme interaction. Most cases are mild; severe disease often involves cytokine storm and organ failure. Therapeutics including antivirals, immunomodulators, and vaccines are in development. To view this SnapShot, open or download the PDF.
Assuntos
Betacoronavirus/fisiologia , Infecções por Coronavirus/patologia , Pneumonia Viral/patologia , Animais , Betacoronavirus/classificação , Betacoronavirus/genética , COVID-19 , Teste para COVID-19 , Vacinas contra COVID-19 , Técnicas de Laboratório Clínico/métodos , Infecções por Coronavirus/diagnóstico , Infecções por Coronavirus/tratamento farmacológico , Infecções por Coronavirus/imunologia , Infecções por Coronavirus/prevenção & controle , Infecções por Coronavirus/terapia , Infecções por Coronavirus/transmissão , Humanos , Pandemias , Pneumonia Viral/imunologia , Pneumonia Viral/terapia , Pneumonia Viral/transmissão , SARS-CoV-2 , Vacinas Virais/imunologia , Tratamento Farmacológico da COVID-19RESUMO
The SARS-CoV-2 virus emerged in December 2019 and has caused a worldwide pandemic due to the lack of any pre-existing immunity. Accurate serology testing is urgently needed to help diagnose infection, determine past exposure of populations and assess the response to a future vaccine. The landscape of antibody responses to SARS-CoV-2 is unknown. In this study, we utilized the luciferase immunoprecipitation system to assess the antibody responses to 15 different SARS-CoV-2 antigens in patients with COVID-19. We identified new targets of the immune response to SARS-CoV-2 and show that nucleocapsid, open reading frame (ORF)8 and ORF3b elicit the strongest specific antibody responses. ORF8 and ORF3b antibodies, taken together as a cluster of points, identified 96.5% of COVID-19 samples at early and late time points of disease with 99.5% specificity. Our findings could be used to develop second-generation diagnostic tests to improve serological assays for COVID-19 and are important in understanding pathogenicity.
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Anticorpos Antivirais/sangue , Betacoronavirus/imunologia , Técnicas de Laboratório Clínico/métodos , Infecções por Coronavirus/diagnóstico , Pneumonia Viral/diagnóstico , Proteínas Virais/imunologia , Adulto , Idoso , Anticorpos Antivirais/imunologia , Antígenos Virais/imunologia , COVID-19 , Teste para COVID-19 , Infecções por Coronavirus/sangue , Infecções por Coronavirus/imunologia , Infecções por Coronavirus/virologia , Feminino , Hong Kong , Humanos , Masculino , Pessoa de Meia-Idade , Pandemias , Pneumonia Viral/sangue , Pneumonia Viral/imunologia , Pneumonia Viral/virologia , SARS-CoV-2 , Sensibilidade e Especificidade , Fatores de TempoRESUMO
We conducted a serological study to define correlates of immunity against SARS-CoV-2. Compared to those with mild coronavirus disease 2019 (COVID-19) cases, individuals with severe disease exhibited elevated virus-neutralizing titers and antibodies against the nucleocapsid (N) and the receptor binding domain (RBD) of the spike protein. Age and sex played lesser roles. All cases, including asymptomatic individuals, seroconverted by 2 weeks after PCR confirmation. Spike RBD and S2 and neutralizing antibodies remained detectable through 5-7 months after onset, whereas α-N titers diminished. Testing 5,882 members of the local community revealed only 1 sample with seroreactivity to both RBD and S2 that lacked neutralizing antibodies. This fidelity could not be achieved with either RBD or S2 alone. Thus, inclusion of multiple independent assays improved the accuracy of antibody tests in low-seroprevalence communities and revealed differences in antibody kinetics depending on the antigen. We conclude that neutralizing antibodies are stably produced for at least 5-7 months after SARS-CoV-2 infection.
Assuntos
Betacoronavirus/imunologia , Técnicas de Laboratório Clínico/métodos , Infecções por Coronavirus/epidemiologia , Infecções por Coronavirus/imunologia , Imunidade Humoral , Pneumonia Viral/epidemiologia , Pneumonia Viral/imunologia , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Anticorpos Neutralizantes/sangue , Anticorpos Antivirais/sangue , Arizona/epidemiologia , Betacoronavirus/isolamento & purificação , COVID-19 , Teste para COVID-19 , Infecções por Coronavirus/sangue , Infecções por Coronavirus/diagnóstico , Proteínas do Nucleocapsídeo de Coronavírus , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Proteínas do Nucleocapsídeo/imunologia , Pandemias , Fosfoproteínas , Pneumonia Viral/sangue , Pneumonia Viral/diagnóstico , Prevalência , Domínios e Motivos de Interação entre Proteínas , SARS-CoV-2 , Estudos Soroepidemiológicos , Glicoproteína da Espícula de Coronavírus/química , Glicoproteína da Espícula de Coronavírus/imunologia , Adulto JovemAssuntos
Angioedema/imunologia , Betacoronavirus/imunologia , Infecções por Coronavirus/imunologia , Síndrome da Liberação de Citocina/imunologia , Citocinas/metabolismo , Pneumonia Viral/imunologia , Angioedema/sangue , Angioedema/patologia , Angioedema/virologia , Enzima de Conversão de Angiotensina 2 , Anti-Inflamatórios/farmacologia , Anti-Inflamatórios/uso terapêutico , Antivirais/uso terapêutico , Biomarcadores/sangue , COVID-19 , Teste para COVID-19 , Técnicas de Laboratório Clínico/métodos , Congressos como Assunto , Infecções por Coronavirus/sangue , Infecções por Coronavirus/diagnóstico , Infecções por Coronavirus/tratamento farmacológico , Infecções por Coronavirus/epidemiologia , Síndrome da Liberação de Citocina/sangue , Síndrome da Liberação de Citocina/virologia , Citocinas/antagonistas & inibidores , Citocinas/sangue , Citocinas/imunologia , Humanos , Internet , Sistema Calicreína-Cinina/efeitos dos fármacos , Sistema Calicreína-Cinina/imunologia , Pandemias , Peptidil Dipeptidase A/metabolismo , Pneumonia Viral/sangue , Pneumonia Viral/diagnóstico , Pneumonia Viral/epidemiologia , Alvéolos Pulmonares/imunologia , Alvéolos Pulmonares/patologia , SARS-CoV-2 , Índice de Gravidade de Doença , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/imunologia , Glicoproteína da Espícula de Coronavírus/imunologia , Glicoproteína da Espícula de Coronavírus/metabolismo , Fatores de Tempo , Tempo para o Tratamento , Tratamento Farmacológico da COVID-19RESUMO
Throughout the SARS-CoV-2 pandemic, limited diagnostic capacities prevented sentinel testing, demonstrating the need for novel testing infrastructures. Here, we describe the setup of a cost-effective platform that can be employed in a high-throughput manner, which allows surveillance testing as an acute pandemic control and preparedness tool, exemplified by SARS-CoV-2 diagnostics in an academic environment. The strategy involves self-sampling based on gargling saline, pseudonymized sample handling, automated RNA extraction, and viral RNA detection using a semiquantitative multiplexed colorimetric reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay with an analytical sensitivity comparable with RT-qPCR. We provide standard operating procedures and an integrated software solution for all workflows, including sample logistics, analysis by colorimetry or sequencing, and communication of results. We evaluated factors affecting the viral load and the stability of gargling samples as well as the diagnostic sensitivity of the RT-LAMP assay. In parallel, we estimated the economic costs of setting up and running the test station. We performed > 35,000 tests, with an average turnover time of < 6 h from sample arrival to result announcement. Altogether, our work provides a blueprint for fast, sensitive, scalable, cost- and labor-efficient RT-LAMP diagnostics, which is independent of potentially limiting clinical diagnostics supply chains.
Assuntos
COVID-19 , SARS-CoV-2 , Humanos , SARS-CoV-2/genética , COVID-19/diagnóstico , COVID-19/epidemiologia , Teste para COVID-19 , Técnicas de Laboratório Clínico/métodos , Pandemias/prevenção & controle , Sensibilidade e Especificidade , RNA Viral/genéticaRESUMO
The early prediction of deterioration could have an important role in supporting healthcare professionals, as an estimated 11% of deaths in hospital follow a failure to promptly recognize and treat deteriorating patients1. To achieve this goal requires predictions of patient risk that are continuously updated and accurate, and delivered at an individual level with sufficient context and enough time to act. Here we develop a deep learning approach for the continuous risk prediction of future deterioration in patients, building on recent work that models adverse events from electronic health records2-17 and using acute kidney injury-a common and potentially life-threatening condition18-as an exemplar. Our model was developed on a large, longitudinal dataset of electronic health records that cover diverse clinical environments, comprising 703,782 adult patients across 172 inpatient and 1,062 outpatient sites. Our model predicts 55.8% of all inpatient episodes of acute kidney injury, and 90.2% of all acute kidney injuries that required subsequent administration of dialysis, with a lead time of up to 48 h and a ratio of 2 false alerts for every true alert. In addition to predicting future acute kidney injury, our model provides confidence assessments and a list of the clinical features that are most salient to each prediction, alongside predicted future trajectories for clinically relevant blood tests9. Although the recognition and prompt treatment of acute kidney injury is known to be challenging, our approach may offer opportunities for identifying patients at risk within a time window that enables early treatment.
Assuntos
Injúria Renal Aguda/diagnóstico , Técnicas de Laboratório Clínico/métodos , Injúria Renal Aguda/complicações , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Simulação por Computador , Conjuntos de Dados como Assunto , Reações Falso-Positivas , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Doença Pulmonar Obstrutiva Crônica/complicações , Curva ROC , Medição de Risco , Incerteza , Adulto JovemRESUMO
Powassan virus is a tick-borne flavivirus that can cause severe neuroinvasive disease, with areas of endemicity in the Northeast and Midwest United States, Canada, and Russia. Diagnosis is challenging and relies on a high index of suspicion and choosing the right test based on duration of infection and the patient's immune status. This review covers laboratory testing for Powassan virus, including historical considerations, modern options, and methods being developed in the research space.
Assuntos
Vírus da Encefalite Transmitidos por Carrapatos , Encefalite Transmitida por Carrapatos , Humanos , Encefalite Transmitida por Carrapatos/diagnóstico , Encefalite Transmitida por Carrapatos/epidemiologia , Encefalite Transmitida por Carrapatos/virologia , Vírus da Encefalite Transmitidos por Carrapatos/isolamento & purificação , Técnicas de Laboratório Clínico/métodos , História do Século XXI , História do Século XX , Animais , Canadá/epidemiologia , Anticorpos Antivirais/sangueRESUMO
We studied the development of the severe acute respiratory syndrome-related coronavirus (SARS-CoV-2) pandemic in southern Finland in 2020 and evaluated the performance of two surrogate immunoassays for the detection of neutralizing antibodies (NAbs). The data set consisted of 12 000 retrospectively collected samples from pregnant women in their first trimester throughout 2020. All the samples were initially screened for immunoglobulin G (IgG) with SARS-CoV-2 spike antibody assay (EIM-S1, Euroimmun) followed by confirmation with nucleocapsid antibody assay (Architect SARS-CoV-2, Abbott). Samples that were reactive (positive or borderline) with both assays were subjected to testing with commercial surrogate immunoassays of NeutraLISA (EIM) and cPassTM (GenScript Biotech Corporation) by using pseudoneutralization assay (PNAbA) as a golden standard. No seropositive cases were detected between January and March. Between April and December, IgG (EIM-S1 and Abbott positive) and NAb (PNAbA positive) seroprevalences were between 0.4% and 1.4%. NeutraLISA showed 90% and cPass 55% concordant results with PNAbA among PNAbA negative samples and 49% and 92% among PNAbA positive samples giving NeutraLISA better specificity but lower sensitivity than cPass. To conclude, seroprevalence in pregnant women reflected that of the general population but the variability of the performance of serological protocols needs to be taken into account in inter-study comparison.
Assuntos
COVID-19 , Gravidez , Humanos , Feminino , COVID-19/diagnóstico , COVID-19/epidemiologia , SARS-CoV-2 , Gestantes , Estudos Retrospectivos , Estudos Soroepidemiológicos , Finlândia/epidemiologia , Teste para COVID-19 , Técnicas de Laboratório Clínico/métodos , Sensibilidade e Especificidade , Anticorpos Neutralizantes , Anticorpos Antivirais , Imunoglobulina GRESUMO
The COVID-19 pandemic brought to light the impact of a widespread disease on various aspects of human relationships, communities, and economies. One notable consequence was the increased demand for diagnostic kits, laboratory reagents, and personal health equipment. This surge in testing capacity worldwide led to shortages in the supply of essential items, including RNA extraction kits, which are crucial for detecting COVID-19 infections. To address this scarcity, researchers have proposed alternative and cost-effective strategies for RNA extraction, utilizing both chemical and physical solutions and extraction-free methods. These approaches aim to alleviate the challenges associated with the overwhelming number of tests being conducted in laboratories. The purpose of this review is intends to provide a comprehensive summary of the various kit-free RNA extraction methods available for COVID-19 diagnosis during the pandemic.
Assuntos
COVID-19 , Humanos , COVID-19/diagnóstico , Teste para COVID-19 , Pandemias , SARS-CoV-2/genética , Técnicas de Laboratório Clínico/métodos , RNA Viral/genética , Sensibilidade e EspecificidadeRESUMO
OBJECTIVES: According to ISO 15189:2022, analytical performance specifications (APS) should relate to intended clinical use and impact on patient care. Therefore, we aimed to develop a web application for laboratory professionals to calculate APS based on a simulation of the impact of measurement uncertainty (MU) on the outcome using the chosen decision limits, agreement thresholds, and data of the population of interest. METHODS: We developed the "APS Calculator" allowing users to upload and select data of concern, specify decision limits and agreement thresholds, and conduct simulations to determine APS for MU. The simulation involved categorizing original measurand concentrations, generating measured (simulated) results by introducing different degrees of MU, and recategorizing measured concentrations based on clinical decision limits and acceptable clinical misclassification rates. The agreements between original and simulated result categories were assessed, and values that met or exceeded user-specified agreement thresholds that set goals for the between-category agreement were considered acceptable. The application generates contour plots of agreement rates and corresponding MU values. We tested the application using National Health and Nutrition Examination Survey data, with decision limits from relevant guidelines. RESULTS: We determined APS for MU of six measurands (blood total hemoglobin, plasma fasting glucose, serum total and high-density lipoprotein cholesterol, triglycerides, and total folate) to demonstrate the potential of the application to generate APS. CONCLUSIONS: The developed data-driven web application offers a flexible tool for laboratory professionals to calculate APS for MU using their chosen decision limits and agreement thresholds, and the data of the population of interest.
Assuntos
Técnicas de Laboratório Clínico , Laboratórios , Humanos , Incerteza , Técnicas de Laboratório Clínico/métodos , Inquéritos Nutricionais , JejumRESUMO
An outbreak of monkeypox (Mpox) was reported in more than 40 countries in early 2022. Accurate diagnosis of Mpox can be challenging, but history, clinical findings, and laboratory diagnosis can establish the diagnosis. The pre-analytic phase of testing includes collecting, storing, and transporting specimens. It is advised to swab the lesion site with virus transport medium (VTM) containing Dacron or polyester flock swabs from two different sites. Blood, urine, and semen samples may also be used. Timely sampling is necessary to obtain a sufficient amount of virus or antibodies. The analytical phase of infectious disease control involves diagnostic tools to determine the presence of the virus. While polymerase chain reaction (PCR) is the gold standard for detecting Mpox, genome sequencing is for identifying new or modified viruses. As a complement to these methods, isothermal amplification methods have been designed. ELISA assays are also available for the determination of antibodies. Electron microscopy is another effective diagnostic method for tissue identification of the virus. Wastewater fingerprinting provides some of the most effective diagnostic methods for virus identification at the community level. The advantages and disadvantages of these methods are further discussed. Post-analytic phase requires proper interpretation of test results and the preparation of accurate patient reports that include relevant medical history, clinical guidelines, and recommendations for follow-up testing or treatment.
Assuntos
Mpox , Humanos , Mpox/diagnóstico , Mpox/virologia , Mpox/epidemiologia , Monkeypox virus/genética , Monkeypox virus/isolamento & purificação , Ensaio de Imunoadsorção Enzimática/métodos , Técnicas de Diagnóstico Molecular/métodos , Técnicas de Amplificação de Ácido Nucleico/métodos , Reação em Cadeia da Polimerase/métodos , Manejo de Espécimes/métodos , Técnicas de Laboratório Clínico/métodosRESUMO
The diagnosis of systemic vasculitis (SV) is a major clinical challenge due to the very different forms of presentation and requires an interdisciplinary approach. Targeted laboratory diagnostics support making the diagnosis, differential diagnosis and classification and are also a key component in the detection of active organ manifestations and treatment complications. The basic laboratory tests include the erythrocyte sedimentation rate (ESR), Creactive protein (CRP), blood count, serum creatinine, urinalysis, specific autoantibodies, complement, immunoglobulins, cryoglobulins and hepatitis B and C serology. Antineutrophil cytoplasmic autoantibodies (ANCA), antiglomerular basement membrane antibodies (anti-GBM antibodies) and anti-C1q antibodies are valuable laboratory markers for the diagnosis of the various forms of small vessel vasculitis. There are no specific laboratory tests for the diagnosis of medium and large vessel vasculitis. Despite advances in our understanding of the pathogenesis of vasculitis, no biomarkers have yet been identified that can be reliably used to guide treatment or that are useful in distinguishing vasculitis from other inflammatory diseases such as infections or treatment complications.
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Anticorpos Anticitoplasma de Neutrófilos , Biomarcadores , Vasculite , Humanos , Biomarcadores/sangue , Vasculite/diagnóstico , Vasculite/sangue , Vasculite/imunologia , Anticorpos Anticitoplasma de Neutrófilos/sangue , Anticorpos Anticitoplasma de Neutrófilos/imunologia , Autoanticorpos/sangue , Técnicas de Laboratório Clínico/métodos , Diagnóstico DiferencialRESUMO
BACKGROUND: Testing is critical for detecting SARS-CoV-2 infection, but the best sampling method remains unclear. OBJECTIVES: To determine whether nasopharyngeal swab (NPS), oropharyngeal swab (OPS) or saliva specimen collection has the highest detection rate for SARS-CoV-2 molecular testing. METHODS: We conducted a randomised clinical trial at two COVID-19 outpatient test centres where NPS, OPS and saliva specimens were collected by healthcare workers in different orders for reverse transcriptase PCR testing. The SARS-CoV-2 detection rate was calculated as the number positive by a specific sampling method divided by the number in which any of the three sampling methods was positive. As secondary outcomes, test-related discomfort was measured with an 11-point numeric scale and cost-effectiveness was calculated. RESULTS: Among 23 102 adults completing the trial, 381 (1.65%) were SARS-CoV-2 positive. The SARS-CoV-2 detection rate was higher for OPSs, 78.7% (95% CI 74.3 to 82.7), compared with NPSs, 72.7% (95% CI 67.9 to 77.1) (p=0.049) and compared with saliva sampling, 61.9% (95% CI 56.9 to 66.8) (p<0.001). The discomfort score was highest for NPSs, at 5.76 (SD, 2.52), followed by OPSs, at 3.16 (SD 3.16) and saliva samples, at 1.03 (SD 18.8), p<0.001 between all measurements. Saliva specimens were associated with the lowest cost, and the incremental costs per detected SARS-CoV-2 infection for NPSs and OPSs were US$3258 and US$1832, respectively. CONCLUSIONS: OPSs were associated with higher SARS-CoV-2 detection and lower test-related discomfort than NPSs for SARS-CoV-2 testing. Saliva sampling had the lowest SARS-CoV-2 detection but was the least costly strategy for mass testing. TRIAL REGISTRATION NUMBER: NCT04715607.
Assuntos
COVID-19 , SARS-CoV-2 , Adulto , Humanos , COVID-19/diagnóstico , Teste para COVID-19 , Saliva , Técnicas de Laboratório Clínico/métodos , Nasofaringe , Manejo de Espécimes/métodosRESUMO
The emergence of a novel coronavirus, namely, SARS-CoV-2, necessitated the use of rapid, accurate diagnostics to quickly diagnose COVID-19. This need has increased with the emergence of new variants and continued waves of COVID-19 cases. The ID NOW COVID-19 assay is a rapid nucleic acid amplification test (NAAT) that is used by hospitals, urgent care facilities, medical clinics, and public health laboratories for rapid molecular SARS-CoV-2 testing at the point of care. The District of Columbia Department of Forensic Sciences Public Health Laboratory Division (DC DFS PHL) implemented ID NOW COVID-19 testing in nontraditional laboratory settings, including a mobile testing unit, health clinic, and emergency department, to assist with rapid identification and isolation for populations at high risk of SARS-CoV-2 transmission in the District of Columbia. The DC DFS PHL provided these nontraditional laboratories with safety risk assessment, assay training, competency assessment, and quality control monitoring as parts of a comprehensive quality management system (QMS). We assessed the accuracy of the ID NOW COVID-19 assay when operated in the context of these trainings and systems. This was done by comparing results from 9,518 paired tests, and strong agreement (κ = 0.88, OPA = 98.3%) was found between the ID NOW COVID-19 assay and laboratory-based NAATs. These findings indicate that the ID NOW COVID-19 assay can be used to detect SARS-CoV-2 in nontraditional laboratory settings when used within the context of a comprehensive QMS.
Assuntos
COVID-19 , Humanos , COVID-19/diagnóstico , Teste para COVID-19 , SARS-CoV-2/genética , Sistemas Automatizados de Assistência Junto ao Leito , Técnicas de Laboratório Clínico/métodos , Laboratórios , Sensibilidade e Especificidade , Técnicas de Amplificação de Ácido Nucleico/métodosRESUMO
BACKGROUND: COVID-19 diagnosis lies on the detection of SARS-CoV-2 on nasopharyngeal specimens by RT-PCR. The Xpert-Xpress SARS-CoV-2 assay provides results in less than one hour from specimen reception, which makes it suitable for clinical/epidemiological circumstances that require faster responses. The analysis of a COVID-19 outbreak suspected in the neonatology ward from our institution showed that the Ct values obtained for the targeted genes in the Xpert assay were markedly different within each specimen (N Ct value > 20 cycles above the E Ct value). RESULTS: We identified the mutation C29200T in the N gene as responsible for an impairment in the N gene amplification by performing whole genome sequencing of the specimens involved in the outbreak (Omicron variant). Subsequently, a retrospective analysis of all specimens sequenced in our institution allowed us to identify the same SNP as responsible for similar impairments in another 12 cases (42% of the total cases reported in the literature). Finally, we found that the same SNP emerged in five different lineages independently, throughout almost all the COVID-19 pandemic. CONCLUSIONS: We demonstrated for the first time the impact of this SNP on the Xpert assay, when harbored by new Omicron variants. We extend our observation period throughout almost all the COVID-19 pandemic, offering the most updated observations of this phenomenon, including sequences from the seventh pandemic wave, until now absent in the reports related to this issue. Continuous monitoring of emerging SNPs that could affect the performance of the most commonly used diagnostic tests, is required to redesign the tests to restore their correct performance.
Assuntos
COVID-19 , SARS-CoV-2 , Humanos , SARS-CoV-2/genética , COVID-19/diagnóstico , Teste para COVID-19 , Pandemias , Técnicas de Laboratório Clínico/métodos , Estudos Retrospectivos , Sensibilidade e Especificidade , MutaçãoRESUMO
During the SARS-CoV-2 pandemic, rapid and sensitive detection of SARS-CoV-2 has been of high importance for outbreak control. Reverse transcriptase polymerase chain reaction (RT-PCR) is the current gold standard, however, the procedures require an equipped laboratory setting and personnel, which have been regularly overburdened during the pandemic. This often resulted in long waiting times for patients. In contrast, reverse transcriptase loop-mediated isothermal amplification (RT-LAMP) is a simple, cost-efficient, and fast procedure, allowing for rapid and remote detection of SARS-CoV-2. In the current study, we performed a clinical evaluation of a new point-of-care test system based on LAMP-technology for SARS-CoV-2 detection, providing a result within 25 min (1copy™ COVID-19 MDx Kit Professional system). We tested 112 paired nasopharyngeal swabs, collected in the COVID-19 Ghent University Hospital test center, using the 1copy™ COVID-19 MDx Kit Professional system, and RT-PCR as the reference method. The test system was found to have a clinical sensitivity of 93.24% (69/74) (95% confidence interval [CI]: 84.93%-97.77%) and specificity of 97.37% (37/38) (95% CI: 86.19%-99.93%). Due to its easy smartphone operation and ready-to-use reagents, it ought to be easily applied in for instance general practices, pharmacies, nursing homes, schools, and companies. This would facilitate an efficient SARS-CoV-2 outbreak control and quarantine policy, as diagnosis can occur sooner in a near-patient setting.
Assuntos
COVID-19 , SARS-CoV-2 , Humanos , SARS-CoV-2/genética , COVID-19/diagnóstico , Sistemas Automatizados de Assistência Junto ao Leito , Teste para COVID-19 , Smartphone , Técnicas de Laboratório Clínico/métodos , Sensibilidade e Especificidade , Técnicas de Diagnóstico Molecular/métodos , Técnicas de Amplificação de Ácido Nucleico/métodos , RNA Viral/genéticaRESUMO
BACKGROUND: The COVID-19 pandemic highlighted the importance of diagnostic testing against curbing the spread of SARS-CoV-2. The urgent need and scale for diagnostic tools resulted in manufacturers of SARS-CoV-2 assays receiving emergency authorization that lacked robust analytical or clinical evaluation. As it is highly likely that testing for SARS-CoV-2 will continue to play a central role in public health, the performance characteristics of assays should be evaluated to ensure reliable diagnostic outcomes are achieved. METHODS: VALCOR or "VALidation of SARS-CORona Virus-2 assays" is a study protocol designed to set up a framework for test validation of SARS-CoV-2 virus assays. Using clinical samples collated from VALCOR, the performance of Aptima SARS-CoV-2 assay was assessed against a standard comparator assay. Diagnostic test parameters such as sensitivity, specificity and overall per cent agreement were calculated for the clinical performance of Aptima SARS-CoV-2 assay. RESULTS: A total of 180 clinical samples were tested with an addition of 40 diluted clinical specimens to determine the limit of detection. When compared to the standard comparator assay Aptima had a sensitivity of 100.0% [95% CI 95.9-100.0] and specificity of 96.7% [95% CI 90.8-99.3]. The overall percent agreement was 98.3% with an excellent Cohen's coefficient of κ = 0.967 [95% CI 0.929-1.000]. For the limit of detection, Aptima was able to detect all of the diluted clinical samples. CONCLUSION: In conclusion. validation of Aptima SARS-CoV-2 assay using clinical samples collated through the VALCOR protocol showed excellent test performance. Additionally, Aptima demonstrated high analytical sensitivity by detecting all diluted clinical samples corresponding to a low limit of detection.
Assuntos
COVID-19 , SARS-CoV-2 , Humanos , Técnicas de Laboratório Clínico/métodos , Teste para COVID-19 , Técnicas de Diagnóstico Molecular/métodos , Pandemias , Sensibilidade e EspecificidadeRESUMO
The ongoing COVID-19 pandemic has created an unprecedented need for rapid diagnostic testing. The World Health Organization (WHO) recommends a standard assay that includes an RNA extraction step from a nasopharyngeal (NP) swab followed by reverse transcription-quantitative polymerase chain reaction (RT-qPCR) to detect the purified SARS-CoV-2 RNA. The current global shortage of RNA extraction kits has caused a severe bottleneck to COVID-19 testing. The goal of this study was to determine whether SARS-CoV-2 RNA could be detected from NP samples via a direct RT-qPCR assay that omits the RNA extraction step altogether. The direct RT-qPCR approach correctly identified 92% of a reference set of blinded NP samples (n = 155) demonstrated to be positive for SARS-CoV-2 RNA by traditional clinical diagnostic RT-qPCR that included an RNA extraction. Importantly, the direct method had sufficient sensitivity to reliably detect those patients with viral loads that correlate with the presence of infectious virus. Thus, this strategy has the potential to ease supply choke points to substantially expand COVID-19 testing and screening capacity and should be applicable throughout the world.
Assuntos
Betacoronavirus/genética , Técnicas de Laboratório Clínico/métodos , Infecções por Coronavirus/diagnóstico , Pneumonia Viral/diagnóstico , RNA Viral/genética , Kit de Reagentes para Diagnóstico/normas , Reação em Cadeia da Polimerase Via Transcriptase Reversa/normas , Betacoronavirus/patogenicidade , COVID-19 , Teste para COVID-19 , Técnicas de Laboratório Clínico/normas , Infecções por Coronavirus/virologia , Primers do DNA/normas , Humanos , Nasofaringe/virologia , Pandemias , Pneumonia Viral/virologia , SARS-CoV-2 , Sensibilidade e Especificidade , Estados Unidos , Carga ViralRESUMO
The current quantitative reverse transcription PCR (RT-qPCR) assay recommended for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) testing in the United States requires analysis of 3 genomic targets per sample: 2 viral and 1 host. To simplify testing and reduce the volume of required reagents, we devised a multiplex RT-qPCR assay to detect SARS-CoV-2 in a single reaction. We used existing N1, N2, and RP primer and probe sets by the Centers for Disease Control and Prevention, but substituted fluorophores to allow multiplexing of the assay. The cycle threshold (Ct) values of our multiplex RT-qPCR were comparable to those obtained by the single assay adapted for research purposes. Low copy numbers (≥500 copies/reaction) of SARS-CoV-2 RNA were consistently detected by the multiplex RT-qPCR. Our novel multiplex RT-qPCR improves upon current single diagnostics by saving reagents, costs, time, and labor.
Assuntos
Betacoronavirus/genética , Técnicas de Laboratório Clínico/métodos , Infecções por Coronavirus/diagnóstico , Reação em Cadeia da Polimerase Multiplex/normas , Pneumonia Viral/diagnóstico , RNA Viral/genética , Kit de Reagentes para Diagnóstico/normas , Reação em Cadeia da Polimerase Via Transcriptase Reversa/normas , Betacoronavirus/patogenicidade , COVID-19 , Teste para COVID-19 , Estudos de Casos e Controles , Técnicas de Laboratório Clínico/normas , Infecções por Coronavirus/virologia , Primers do DNA/normas , Células HEK293 , Humanos , Limite de Detecção , Nasofaringe/virologia , Pandemias , Pneumonia Viral/virologia , SARS-CoV-2 , Estados UnidosRESUMO
COVID-19 is a highly infectious disease caused by the SARS-CoV-2 virus, which primarily affects the respiratory system and can lead to severe illness. The virus is extremely contagious, early and accurate diagnosis of SARS-CoV-2 is crucial to contain its spread, to provide prompt treatment, and to prevent complications. Currently, the reverse transcriptase polymerase chain reaction (RT-PCR) is considered to be the gold standard for detecting COVID-19 in its early stages. In addition, loop-mediated isothermal amplification (LMAP), clustering rule interval short palindromic repeats (CRISPR), colloidal gold immunochromatographic assay (GICA), computed tomography (CT), and electrochemical sensors are also common tests. However, these different methods vary greatly in terms of their detection efficiency, specificity, accuracy, sensitivity, cost, and throughput. Besides, most of the current detection methods are conducted in central hospitals and laboratories, which is a great challenge for remote and underdeveloped areas. Therefore, it is essential to review the advantages and disadvantages of different COVID-19 detection methods, as well as the technology that can enhance detection efficiency and improve detection quality in greater details.