scholarly journals Evaluation of seven different rapid methods for nucleic acid detection of SARS-COV-2 virus

2021 ◽  
Author(s):  
Sally Mahmoud ◽  
Esra Ibrahim ◽  
Subhashini Ganesan ◽  
Bhagyashree Thakre ◽  
Juliet Teddy ◽  
...  
Keyword(s):  
Nucleic Acid ◽  
Gold Standard ◽  
Large Scale ◽  
Virus Detection ◽  
Detection Methods ◽  
Nucleic Acid Detection ◽  
Test Accuracy ◽  
Rt Pcr ◽  
Rapid Methods ◽  
Kappa Value

Background In the current COVID-19 pandemic there is mass screening of SARS-CoV-2 happening round the world due to the extensive spread of the infections. There is a high demand for rapid diagnostic tests to expedite identification of cases and to facilitate early isolation and control spread. Hence this study evaluates seven different rapid nucleic acid detection assays that are commercially available for SARS- CoV- 2 virus detection. Methods Nasopharyngeal samples were collected from 4859 participants and were tested for SARS-CoV-2 virus by the gold standard RT-PCR method along with one of these seven rapid methods of detection. Evaluation of the rapid nucleic acid detection assays was done by comparing the results of these rapid methods with the gold standard RT-qPCR results for SARS-COV-2 detection. Results AQ-TOP had the highest sensitivity (98%) and strong kappa value of 0.943 followed by Genechecker and Abbot ID NOW. The POCKIT (ii RT-PCR) assay had the highest test accuracy of 99.29% followed by Genechecker and Cobas Liat. Atila iAMP showed the highest percentage of invalid reports (35.5%) followed by AQ-TOP with 6% and POCKIT with 3.7% of invalid reports. Conclusion Genechecker system, Abbott ID NOW and Cobas Liat, were found to have best performance and agreement when compared to the standard RT-PCR for COVID-19 detection. With further research, these rapid tests have the potential to be employed in large scale screening of COVID-19.

scholarly journals Virus Detection: A Review of the Current and Emerging Molecular and Immunological Methods

2021 ◽  
Vol 8 ◽  
Author(s):  
A. Cassedy ◽  
A. Parle-McDermott ◽  
R. O’Kennedy
Keyword(s):  
Nucleic Acid ◽  
Virus Detection ◽  
High Sensitivity ◽  
High Volume ◽  
Detection Methods ◽  
Immunological Methods ◽  
Nucleic Acid Detection ◽  
Viral Genomes ◽  
Diagnostic Measures ◽  
Point Of Use

Viruses are ubiquitous in the environment. While many impart no deleterious effects on their hosts, several are major pathogens. This risk of pathogenicity, alongside the fact that many viruses can rapidly mutate highlights the need for suitable, rapid diagnostic measures. This review provides a critical analysis of widely used methods and examines their advantages and limitations. Currently, nucleic-acid detection and immunoassay methods are among the most popular means for quickly identifying viral infection directly from source. Nucleic acid-based detection generally offers high sensitivity, but can be time-consuming, costly, and require trained staff. The use of isothermal-based amplification systems for detection could aid in the reduction of results turnaround and equipment-associated costs, making them appealing for point-of-use applications, or when high volume/fast turnaround testing is required. Alternatively, immunoassays offer robustness and reduced costs. Furthermore, some immunoassay formats, such as those using lateral-flow technology, can generate results very rapidly. However, immunoassays typically cannot achieve comparable sensitivity to nucleic acid-based detection methods. Alongside these methods, the application of next-generation sequencing can provide highly specific results. In addition, the ability to sequence large numbers of viral genomes would provide researchers with enhanced information and assist in tracing infections.

scholarly journals Assessment of the Diagnostic Ability of Four Detection Methods Using Three Sample Types of COVID-19 Patients

2021 ◽  
Vol 11 ◽  
Author(s):  
Fei Yu ◽  
Guoliang Xie ◽  
Shufa Zheng ◽  
Dongsheng Han ◽  
Jiaqi Bao ◽  
...  
Keyword(s):  
Nucleic Acid ◽  
Reverse Transcription ◽  
Detection Methods ◽  
Nucleic Acid Detection ◽  
Sample Type ◽  
Rt Pcr ◽  
Chain Reactions ◽  
Polymerase Chain Reactions ◽  
Oropharyngeal Swab ◽  
And Control

BackgroundViral nucleic acid detection is considered the gold standard for the diagnosis of coronavirus disease 2019 (COVID-19), which is caused by SARS-CoV-2 infection. However, unsuitable sample types and laboratory detection kits/methods lead to misdiagnosis, which delays the prevention and control of the pandemic.MethodsWe compared four nucleic acid detection methods [two kinds of reverse transcription polymerase chain reactions (RT-PCR A: ORF1ab and N testing; RT-PCRB: only ORF1ab testing), reverse transcription recombinase aided amplification (RT-RAA) and droplet digital RT-PCR (dd-RT-PCR)] using 404 samples of 72 hospitalized COVID-19 patients, including oropharyngeal swab (OPS), nasopharyngeal swabs (NPS) and saliva after deep cough, to evaluate the best sample type and method for SARS-CoV-2 detection.ResultsAmong the four methods, dd-RT-PCR exhibited the highest positivity rate (93.0%), followed by RT-PCR B (91.2%) and RT-RAA (91.2%), while the positivity rate of RT-PCR A was only 71.9%. The viral load in OPS [24.90 copies/test (IQR 15.58-129.85)] was significantly lower than that in saliva [292.30 copies/test (IQR 20.20-8628.55)] and NPS [274.40 copies/test (IQR 33.10-2836.45)]. In addition, if OPS samples were tested alone by RT-PCR A, only 21.4% of the COVID-19 patients would be considered positive. The accuracy of all methods reached nearly 100% when saliva and NPS samples from the same patient were tested simultaneously.ConclusionsSARS-CoV-2 nucleic acid detection methods should be fully evaluated before use. High-positivity rate methods such as RT-RAA and dd-RT-PCR should be considered when possible. Furthermore, saliva after deep cough and NPS can greatly improve the accuracy of the diagnosis, and testing OPS alone is not recommended.

scholarly journals Molecular Diagnosis in Differentiating Active and Inactive Forms of Hepatitis B Virus Carriers

2018 ◽  
Vol 26 (10) ◽  
pp. 41-45
Author(s):  
Salah Tofik Jalal Balaky ◽  
Saeed Ghulam Hussain ◽  
Amer Ali Khaleel ◽  
Furat Tahseen Sabeer ◽  
Ahang Hasan Mawlood
Keyword(s):  
Nucleic Acid ◽  
Hepatitis B ◽  
Cross Sectional Study ◽  
Hbv Dna ◽  
Hbv Infection ◽  
Detection Methods ◽  
Rt Pcr ◽  
Cross Sectional ◽  
Blood Samples ◽  
Acid Test

Background & objectives: Introducing a nucleic acid test program is aimed to diagnose and reduces the risk of viral infection or transmission. DNA assay for HBV can detect infection in the windows period, chronic occult infection and can discriminate between active and inactive HBV infection. This cross-sectional study designed to diagnose, analyze HBV infection and to differentiate active from inactive infection based on viral DNA detection. Methods: Blood samples were collected from 256 patients previously diagnosed on the clinical ground as hepatitis B seropositive in Erbil Central Lab. The viral nucleic acid quantitative assessment was done for the collected samples using RT-PCR. Q-square was performed for statistical analysis. Results: Out of 256 collected blood samples 93 (36.3%) showed HBV-DNA positive titers above 50 IU/ml. Among positive subjects, 67 (72.04%) was categorized as inactive carriers (˂ 2000-20.000 IU/ml HBV-DNA titers). Conclusions: The data produced from this study confirmed the importance of the RT-PCR technique in sensitivity and reliability as a superior diagnostics tool specifically in differentiating active from inactive HBV carriers.

scholarly journals Rapid Detection of SARS-CoV-2 Using Reverse transcription RT-LAMP method

2020 ◽  
Author(s):  
Weihua Yang ◽  
Xiaofei Dang ◽  
Qingxi Wang ◽  
Mingjie Xu ◽  
Qianqian Zhao ◽  
...  
Keyword(s):  
Nucleic Acid ◽  
Reverse Transcription ◽  
Virus Disease ◽  
Lamp Assay ◽  
Nucleic Acid Amplification ◽  
N Gene ◽  
Nucleic Acid Detection ◽  
Lamp Method ◽  
Rt Pcr ◽  
Rapid Amplification

AbstractCorona Virus Disease 2019 (COVID-19) is a recently emerged life-threatening disease caused by SARS-CoV-2. Real-time fluorescent PCR (RT-PCR) is the clinical standard for SARS-CoV-2 nucleic acid detection. To detect SARS-CoV-2 early and control the disease spreading on time, a faster and more convenient method for SARS-CoV-2 nucleic acid detecting, RT-LAMP method (reverse transcription loop-mediated isothermal amplification) was developed. RNA reverse transcription and nucleic acid amplification were performed in one step at 63 °C isothermal conditions, and the results can be obtained within 30 minutes. ORF1ab gene, E gene and N gene were detected at the same time. ORF1ab gene was very specific and N gene was very sensitivity, so they can guarantee both sensitivity and specificity for SARS-CoV-2. The sensitivity of RT-LAMP assay is similar to RT-PCR, and specificity was 99% as detecting 208 clinical specimens. The RT-LAMP assay reported here has the advantages of rapid amplification, simple operation, and easy detection, which is useful for the rapid and reliable clinical diagnosis of SARS-CoV-2.

scholarly journals First molecular and serological detection of Epizootic Hemorrhagic Disease virus in white tailed deer ( Odocoileus virginianus ) from Tamaulipas, Mexico

2019 ◽  
Vol 71 (1) ◽  
pp. 77-85
Author(s):  
J.O. Merino ◽  
NI. De la Cruz ◽  
G. Galvan ◽  
A.P. De León ◽  
J. Burnes
Keyword(s):  
Nucleic Acid ◽  
Nucleic Acid Detection ◽  
Hemorrhagic Disease ◽  
Rt Pcr ◽  
Biting Midges ◽  
Epizootic Hemorrhagic Disease ◽  
Two Samples ◽  
Mexico Border ◽  
Transboundary Disease ◽  
The U.S

ABSTRACT Epizootic hemorrhagic disease viruses (EHDV) are dsRNA arboviruses transmitted by biting midges of the genus Culicoides that cause disease in domestic and wild ruminants. Epizootic hemorrhagic disease (EHD) is considered the most important infectious disease of white tailed deer (WTD) in North America, some studies in Northeast Mexico reported EHDV-seropositive WTD and EHDV-infected Culicoides vectors. The increasing population of WTD that share habitat with livestock in Northeast México highlights the importance of EHD for the livestock industry in the transboundary region with the U.S. One hundred and twenty two samples from WTD in Tamaulipas state, Mexico were tested by ELISA and RT-PCR for EHDV antibodies and nucleic acid, respectively. Twelve animals were seropositive to ELISA and eleven animals were positive by RT-PCR. This is the first report of EHDV nucleic acid detection in WTD from Mexico. It is hypothesized that applying the transboundary disease approach to interdisciplinary research will help fill knowledge gaps, which could help develop countermeasures to mitigate the threat of EHDV infection in wildlife and livestock along the U.S.-Mexico border.

Development of a Molecular Method for the Detection of Enteric Viruses in Oysters

1995 ◽  
Vol 58 (12) ◽  
pp. 1357-1362 ◽  
Author(s):  
LEE-ANN JAYKUS ◽  
RICARDO DE LEON ◽  
MARK D. SOBSEY
Keyword(s):  
Cell Culture ◽  
Nucleic Acid ◽  
Hepatitis A Virus ◽  
Enteric Virus ◽  
Nucleic Acid Amplification ◽  
Detection Methods ◽  
Ammonium Bromide ◽  
Rt Pcr ◽  
Initial Inoculum ◽  
Ctab Precipitation

Detection of enteric virus contamination of shellfish is limited by current methodology, which is time-consuming, tedious, and lacking in sensitivity due to reliance on cell culture infectivity. Alternative detection methods based on nucleic acid amplification have been hampered by high sample volumes and the presence of enzymatic inhibitors. The goal of this study was to develop methods to purify and concentrate intact virions from oyster extracts to a volume and quality compatible with viral genomic nucleic acid amplification by reverse transcriptase-polymerase chain reaction (RT-PCR). Fifty-gram oyster samples were homogenized and processed by standard adsorption-elution precipitation methodology and then seeded with 105 PFU of poliovirus 1 (PV1) or hepatitis A virus (HAV). Seeded viruses were concentrated by fluorocarbon extraction, polyethylene glycol (PEG) precipitation, chloroform extraction, and cetyltrimethyl ammonium bromide (CTAB) precipitation to a volume of 100 μl with removal of RT-PCR inhibitors. Virus recovery after elution of PEG precipitates was 50% for PVI and IS to 20% for HAV as evaluted by cell culture infectivity. The CTAB precipitation step yielded a concentrated sample which was directly compatible with RT-PCR reactions and capable of detecting about 100 placque=forming units (PFU) of PVl or HAV. When 50-g oyster extracts were seeded and processed by the entire concentration and purification scheme, direct RT-PCR detection of viral genomic RNA was possible at initial inoculum levels of 104 PFU of HAV and 103 PFU of PV1, with recoveries of 1 to 5% of seeded viruses.

scholarly journals Development and detection of genetically modified crops

2020 ◽  
Vol 145 ◽  
pp. 01013
Author(s):  
Zhao Yu-jia ◽  
Fan Pei-lei ◽  
Liang Liang ◽  
Liu Yin-yin ◽  
Zhao Hai-bo ◽  
...  
Keyword(s):  
Nucleic Acid ◽  
Genetically Modified ◽  
Genetically Modified Crops ◽  
Detection Methods ◽  
Social Benefits ◽  
Nucleic Acid Detection ◽  
Genetically Modified Crop ◽  
The World ◽  
The Difference

Genetically modified crops (GMCs) have been known for the excellent qualities. The commercializing of GMCs has taken great economic and social benefits. However, the bio-security of GMCs was still an issue. To solve this problem, countries around the world were constantly strengthening regulations on planting, processing and detecting of GMCs. This paper reviewed the development of commercialization and detection of GMCs. The difference between protein and nucleic acid detection methods of genetically modified crop was further discussed. This paper will provide new insights for the application of genetically modified crops.

scholarly journals Occurrence of Stone Fruit Viruses in Plum Orchards in Latvia

2013 ◽  
Vol 67 (2) ◽  
pp. 116-123
Author(s):  
Alina Gospodaryk ◽  
Inga Moročko-Bičevska ◽  
Neda Pūpola ◽  
Anna Kāle
Keyword(s):  
Mosaic Virus ◽  
Large Scale ◽  
Ringspot Virus ◽  
Stone Fruit ◽  
Detection Methods ◽  
Plum Pox Virus ◽  
Rt Pcr ◽  
Detection Rates ◽  
Arabis Mosaic Virus ◽  
Das Elisa

To evaluate the occurrence of nine viruses infecting Prunus a large-scale survey and sampling in Latvian plum orchards was carried out. Occurrence of Apple mosaic virus (ApMV), Prune dwarf virus (PDV), Prunus necrotic ringspot virus (PNRSV), Apple chlorotic leaf spot virus (ACLSV), and Plum pox virus (PPV) was investigated by RT-PCR and DAS ELISA detection methods. The detection rates of both methods were compared. Screening of occurrence of Strawberry latent ringspot virus (SLRSV), Arabis mosaic virus (ArMV), Tomato ringspot virus (ToRSV) and Petunia asteroid mosaic virus (PeAMV) was performed by DAS-ELISA. In total, 38% of the tested trees by RT-PCR were infected at least with one of the analysed viruses. Among those 30.7% were infected with PNRSV and 16.4% with PDV, while ApMV, ACLSV and PPV were detected in few samples. The most widespread mixed infection was the combination of PDV+PNRSV. Observed symptoms characteristic for PPV were confirmed with RT-PCR and D strain was detected. Comparative analyses showed that detection rates by RT-PCR and DAS ELISA in plums depended on the particular virus tested. The results obtained in this study revealed that commonly grown plum cultivars in Latvia are infected with economically important stone fruit viruses and highlight the need to implement a programme to produce and propagate virus-free planting material.

scholarly journals Impact of COVID-19 pre-test probability on positive predictive value of high cycle threshold SARS-CoV-2 real-time reverse transcription PCR test results

2021 ◽  
pp. 1-18
Author(s):  
Jonathan B. Gubbay ◽  
Heather Rilkoff ◽  
Heather L. Kristjanson ◽  
Jessica D. Forbes ◽  
Michelle Murti ◽  
...  
Keyword(s):  
Positive Predictive Value ◽  
Real Time ◽  
Reverse Transcription ◽  
Predictive Value ◽  
Large Scale ◽  
Rdrp Gene ◽  
Nucleic Acid Detection ◽  
Rt Pcr ◽  
Pcr Assay ◽  
Test Probability

Abstract Objectives Performance characteristics of SARS-CoV-2 nucleic acid detection assays are understudied within contexts of low pre-test probability, including screening asymptomatic persons without epidemiological links to confirmed cases, or asymptomatic surveillance testing. SARS-CoV-2 detection without symptoms may represent presymptomatic or asymptomatic infection, resolved infection with persistent RNA shedding, or a false positive test. This study assessed positive predictive value of SARS-CoV-2 real-time reverse transcription polymerase chain reaction (rRT-PCR) assays by retesting positive specimens from five pre-test probability groups ranging from high to low with an alternate assay. Methods A total of 122 rRT-PCR positive specimens collected from unique patients between March and July 2020 were retested using a laboratory-developed nested RT-PCR assay targeting the RNA-dependent RNA polymerase (RdRp) gene followed by Sanger sequencing. Results Significantly fewer (15.6%) positive results in the lowest pre-test probability group (facilities with institution-wide screening having ≤ 3 positive asymptomatic cases) were reproduced with the nested RdRp gene RT-PCR assay than in each of the four groups with higher pre-test probability (individual group range 50·0% to 85·0%). Conclusions Large-scale SARS-CoV-2 screening testing initiatives among low pre-test probability populations should be evaluated thoroughly prior to implementation given the risk of false positives and consequent potential for harm at the individual and population level.

scholarly journals Extraction-free rapid cycle RT-qPCR and extreme RT-PCR for SARS-CoV-2 virus detection

2021 ◽  
Vol 156 (Supplement_1) ◽  
pp. S139-S139
Author(s):  
J C Lownik ◽  
J S Farrar ◽  
G Way ◽  
R K Martin
Keyword(s):  
Supply Chain ◽  
Diagnostic Testing ◽  
Rna Extraction ◽  
Virus Detection ◽  
Detection Methods ◽  
Molecular Diagnostic ◽  
Sample Collection ◽  
Rt Pcr ◽  
Hydrolysis Probe ◽  
Time Requirements

Abstract Introduction/Objective Since the start of the coronavirus disease 2019 (COVID-19) pandemic, molecular diagnostic testing for detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has faced substantial supply chain shortages and noteworthy delays in result reporting after sample collection. Supply chain shortages have been most evident in reagents for RNA extraction and rapid diagnostic testing. In this study, we explored the kinetic limitations of extraction-free rapid cycle RT-qPCR for SARS-CoV-2 virus detection using the commercially available capillary based LightCycler. Methods/Case Report We optimized reverse transcription and PCR under extraction-free and rapid thermocycling conditions utilizing hydrolysis probe-based detection methods using a Roche LightCycler. Results (if a Case Study enter NA) This protocol improves detection speed while maintaining the sensitivity and specificity of hydrolysis probe-based detection. Percentage agreement between the developed assay and previously tested positive patient samples was 97.6% (n= 40/41) and negative patient samples was 100% (40/40). We further demonstrate that using purified RNA, SARS-CoV-2 testing using extreme RT-PCR and product verification by melting can be completed in less than 3 minutes. Conclusion We developed a protocol for sensitive and specific RT-qPCR of SARS-CoV-2 RNA from nasopharyngeal swabs in less than 20 minutes, with minimal hands-on time requirements. Overall, these studies provide a framework for increasing the speed of SARS-CoV-2 and other infectious disease testing.

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