An Update on Molecular Diagnostics for COVID-19

Islam, Khursheed Ul; Iqbal, Jawed

doi:10.3389/fcimb.2020.560616

Cited by 116 publications

(102 citation statements)

References 74 publications

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“…rRT-PCR detects RNA in NP swab specimens in the first week, even before the onset of initial symptoms [16]. [17][18][19][20][21][22][23][24][25][26].…”

Section: Results Interpretationmentioning

confidence: 99%

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Considerations in Real-time Reverse Transcription Polymerase Chain Reaction (rRT-PCR) for the Detection of SARS-CoV-2 from Nasopharyngeal Swabs

Sekar

Menezes

Shivappa

et al. 2021

JPRI

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Coronavirus Disease 2019 (COVID-19) was first reported in December 2019, in the City of Wuhan, China. Within the span of a few weeks, the disease had spread to other regions of China and eventually to different parts of the world. COVID 19 has affected 221 countries and territories around the world, with a total of 121,290,697 positive cases and 2,682,554 deaths as on March 17, 2021. Accurate disease diagnosis (for the SARS-Cov-2 virus and variants), coupled to patient isolation are currently critical strategies in restricting disease spread. Due to lack of time during this pandemic the diagnostics assays were not adequately validated. Infected individuals at times could potentially be missed by real-time reverse transcription polymerase chain reaction (rRT-PCR) for SARS-CoV-2 tests due to incorrect/inefficient sampling procedure, low limit of detection and epidemiology of the virus. rRT-PCR test results should be interpreted in conjunction with clinical examination and Computed Tomography (CT), particularly in suspected symptomatic individuals or those with epidemiological history of contact with known COVID-19 cases. Considering the above-mentioned constraints, the current scenario demands rapid and point-of-care tests for detection of SARS-CoV-2 in remote locations. To date, there is no reliable commercially available antigen detection kit. The infected subjects reveal low levels of antibodies against SARS-CoV-2 through the early period of infection. In addition, techniques such as, Digital RT-PCR technology and isothermal RNA amplification with electrochemical biosensors are some of the new technologies currently being developed to provide sensitive and specific SARS-Cov-2 antigen detection. The newly reported variant, SARS-CoV-2 VUI 202012/01 may not influence diagnostic outcomes as worldwide most PCR assays use two or more (including RdRp/ E/ N) reliable gene targets, besides S gene.

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“…rRT-PCR detects RNA in NP swab specimens in the first week, even before the onset of initial symptoms [16]. [17][18][19][20][21][22][23][24][25][26].…”

Section: Results Interpretationmentioning

confidence: 99%

“…A "positive" PCR result need not necessarily signify the presence of a viable virus. Viral RNA is, in some cases, detected even beyond week 6 by rRT-PCR [18][19][20][21][22]27]. Some centres have tried pooled samples (up to 30 samples) for to reduce the cost of testing for SARS-CoV-2 in asymptomatic people.…”

Section: Results Interpretationmentioning

confidence: 99%

Considerations in Real-time Reverse Transcription Polymerase Chain Reaction (rRT-PCR) for the Detection of SARS-CoV-2 from Nasopharyngeal Swabs

Sekar

Menezes

Shivappa

et al. 2021

JPRI

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“…After they are activated, Cas cleaves the nearby single-stranded DNA (ssDNA)/RNA molecules non-specifically, a feature called collateral cleavage or trans-cleavage ( Kumar et al, 2020 ; Wang M et al, 2020 ). There have been several reports on diagnostics and detection of SARS-CoV-2 using CRISPR/Cas-based therapeutic tools ( Chen et al, 2018 ; Ali et al, 2020 ; Broughton et al, 2020 ; Ding et al, 2020 ; Islam and Iqbal, 2020 ; Javalkote et al, 2020 ; Wang M et al, 2020 ; Wang XJ et al, 2020b ).…”

Section: Wide Use Of Crispr/cas In Clinical and Preclinical Researcmentioning

confidence: 99%

CRISPR/Cas: a Nobel Prize award-winning precise genome editing technology for gene therapy and crop improvement

Brant

Budak

et al. 2021

J. Zhejiang Univ. Sci. B

122

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Since it was first recognized in bacteria and archaea as a mechanism for innate viral immunity in the early 2010s, clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein (Cas) has rapidly been developed into a robust, multifunctional genome editing tool with many uses. Following the discovery of the initial CRISPR/Cas-based system, the technology has been advanced to facilitate a multitude of different functions. These include development as a base editor, prime editor, epigenetic editor, and CRISPR interference (CRISPRi) and CRISPR activator (CRISPRa) gene regulators. It can also be used for chromatin and RNA targeting and imaging. Its applications have proved revolutionary across numerous biological fields, especially in biomedical and agricultural improvement. As a diagnostic tool, CRISPR has been developed to aid the detection and screening of both human and plant diseases, and has even been applied during the current coronavirus disease 2019 (COVID-19) pandemic. CRISPR/Cas is also being trialed as a new form of gene therapy for treating various human diseases, including cancers, and has aided drug development. In terms of agricultural breeding, precise targeting of biological pathways via CRISPR/Cas has been key to regulating molecular biosynthesis and allowing modification of proteins, starch, oil, and other functional components for crop improvement. Adding to this, CRISPR/Cas has been shown capable of significantly enhancing both plant tolerance to environmental stresses and overall crop yield via the targeting of various agronomically important gene regulators. Looking to the future, increasing the efficiency and precision of CRISPR/Cas delivery systems and limiting off-target activity are two major challenges for wider application of the technology. This review provides an in-depth overview of current CRISPR development, including the advantages and disadvantages of the technology, recent applications, and future considerations.

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“…Recently Feng Zang group at Massachusetts Institute of Technology, combined clustered regularly interspaced short palindromic repeats and CRISPR Associated protein (CRISPR Cas) system with isothermal amplification methods to develop diagnostic assays for infectious diseases called SHERLOCK an acronym for Specific High-sensitivity Enzymatic Reporter Unlocking ( 20 ). This has prompted researchers to develop several CRISPR-Cas based assays for COVID-19 diagnosis including SHERLOCK, one-pot visual SARS-CoV-2 detection system (opvCRISPR) ( 21 , 22 ), All-In-One Dual CRISPR Cas12a (AIODCRISPR) ( 23 ), Cas13-based, Rugged, equitable, scalable testing (CREST) ( 24 ) and Fn-Cas9 based enzymatic readout for nucleotide detection and nucleobase identification (FELUDA) ( 25 , 26 ). As the positive reaction for these assays can be visualized on a lateral flow paper dip stick, they can be used as point-of-care molecular diagnostic assays ( 20 ).…”

Section: Molecular Diagnosticsmentioning

confidence: 99%

Laboratory diagnosis of COVID-19: current status and challenges

Mathew¹,

Vyas²,

Khare³

et al. 2021

IJM

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The magnitude and pace of global affliction caused by Coronavirus Disease-19 (COVID-19) is unprecedented in the recent past. From starting in a busy seafood market in the Chinese city of Wuhan, the virus has spread across the globe in less than a year, infecting over 76 million people and causing death of close to 1.7 million individuals worldwide. As no specific antiviral treatment is currently available, the major strategy in containing the pandemic is focused on early diagnosis and prompt isolation of the infected individuals. Several diagnostic modalities have emerged within a relatively short period, which can be broadly classified into molecular and immunological assays. While the former category is centered around real-time PCR, which is currently considered the gold standard of diagnosis, the latter aims to detect viral antigens or antibodies specific to the viral antigens and is yet to be recommended as a stand-alone diagnostic tool. This review aims to provide an update on the different diagnostic modalities that are currently being used in diagnostic laboratories across the world as well as the upcoming methods and challenges associated with each of them. In a rapidly evolving diagnostic landscape with several testing platforms going through various phases of development and/or regulatory clearance, it is prudent that the clinical community familiarizes itself with the nuances of different testing modalities currently being employed for this condition.

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An Update on Molecular Diagnostics for COVID-19

Cited by 116 publications

References 74 publications

Considerations in Real-time Reverse Transcription Polymerase Chain Reaction (rRT-PCR) for the Detection of SARS-CoV-2 from Nasopharyngeal Swabs

Considerations in Real-time Reverse Transcription Polymerase Chain Reaction (rRT-PCR) for the Detection of SARS-CoV-2 from Nasopharyngeal Swabs

CRISPR/Cas: a Nobel Prize award-winning precise genome editing technology for gene therapy and crop improvement

Laboratory diagnosis of COVID-19: current status and challenges

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