scholarly journals Improving RT-LAMP Detection of SARS-CoV-2 RNA through Primer Set Selection and Combination

2021 ◽  
Author(s):  
Nathan Tanner ◽  
Yinhua Zhang
Keyword(s):  
Nucleic Acid ◽  
Diagnostic Method ◽  
High Sensitivity ◽  
Sensitivity Enhancement ◽  
Molecular Diagnostic ◽  
Nucleic Acid Testing ◽  
Lamp Primer ◽  
Body Of Knowledge ◽  
Primer Sets ◽  
Molecular Diagnostic Method

Reverse transcription loop-mediated isothermal amplification (RT-LAMP) has emerged as a viable molecular diagnostic method to expand the breadth and reach of nucleic acid testing, particularly for SARS-CoV-2 detection and surveillance. While rapidly growing in prominence, RT-LAMP remains a relatively new method compared to the standard RT-qPCR, and contribution to our body of knowledge on designing LAMP primer sets and assays can have significant impact on its utility and adoption. Here we evaluate 18 LAMP primer sets for SARS-CoV-2, comparing speed and sensitivity with different LAMP formulations and conditions across more than 5,000 RT-LAMP reactions and identifying several primer sets with similar high sensitivity for different SARS-CoV-2 gene targets. Significantly we observe a consistent sensitivity enhancement by combining primer sets for different targets, confirming and building on earlier work to create a simple, general approach to building better and more sensitive RT-LAMP assays.

scholarly journals Why cannot a β-lactamase gene be detected using an efficient molecular diagnostic method?

2016 ◽  
Vol 32 (5) ◽  
Author(s):  
Sang Hee Lee ◽  
Kwang Seung Park ◽  
Jung Hun Lee ◽  
Moonhee Park ◽  
Asad Mustafa Karim
Keyword(s):  
Diagnostic Method ◽  
Molecular Diagnostic ◽  
Lactamase Gene ◽  
Molecular Diagnostic Method

A rapid molecular diagnostic method for spinal muscular atrophy

2020 ◽  
pp. 1-4
Author(s):  
Kai-Chen Wang ◽  
Chiao-Yuan Fang ◽  
Chi-Chang Chang ◽  
Chien-Kuan Chiang ◽  
Yi-Wen Chen
Keyword(s):  
Spinal Muscular Atrophy ◽  
Diagnostic Method ◽  
Muscular Atrophy ◽  
Molecular Diagnostic ◽  
Molecular Diagnostic Method

scholarly journals MO37-1 Identification and evaluation of a serum microRNA molecular diagnostic method for colorectal polyp patients

2021 ◽  
Vol 32 ◽  
pp. S322
Author(s):  
Lui Ng ◽  
Ryan W.Y. Sin ◽  
Oswens S.H. Lo ◽  
Carlos K.H. Wong ◽  
Cindy L.K. Lam ◽  
...  
Keyword(s):  
Diagnostic Method ◽  
Colorectal Polyp ◽  
Molecular Diagnostic ◽  
Serum Microrna ◽  
Molecular Diagnostic Method

scholarly journals Nucleic Acid Testing of SARS-CoV-2

2021 ◽  
Vol 22 (11) ◽  
pp. 6150
Author(s):  
Hee-Min Yoo ◽  
Il-Hwan Kim ◽  
Seil Kim
Keyword(s):  
Polymerase Chain Reaction ◽  
Nucleic Acid ◽  
Reverse Transcription ◽  
Diagnostic Methods ◽  
Detection Methods ◽  
Molecular Diagnostic ◽  
Nucleic Acid Testing ◽  
Rt Pcr ◽  
Chain Reaction ◽  
Polymerase Chain

The coronavirus disease 2019 (COVID-19) has caused a large global outbreak. It is accordingly important to develop accurate and rapid diagnostic methods. The polymerase chain reaction (PCR)-based method including reverse transcription-polymerase chain reaction (RT-PCR) is the most widely used assay for the detection of SARS-CoV-2 RNA. Along with the RT-PCR method, digital PCR has emerged as a powerful tool to quantify nucleic acid of the virus with high accuracy and sensitivity. Non-PCR based techniques such as reverse transcription loop-mediated isothermal amplification (RT-LAMP) and reverse transcription recombinase polymerase amplification (RT-RPA) are considered to be rapid and simple nucleic acid detection methods and were reviewed in this paper. Non-conventional molecular diagnostic methods including next-generation sequencing (NGS), CRISPR-based assays and nanotechnology are improving the accuracy and sensitivity of COVID-19 diagnosis. In this review, we also focus on standardization of SARS-CoV-2 nucleic acid testing and the activity of the National Metrology Institutes (NMIs) and highlight resources such as reference materials (RM) that provide the values of specified properties. Finally, we summarize the useful resources for convenient COVID-19 molecular diagnostics.

scholarly journals Effectiveness of CRISPR based tools as alternative methods for diagnosing COVID-19

2021 ◽  
Vol 8 (3) ◽  
pp. 150-152
Author(s):  
Shefali Mehta ◽  
Aditi Mehta
Keyword(s):  
Diagnostic Method ◽  
Diagnostic Methods ◽  
Alternative Methods ◽  
Molecular Diagnostic ◽  
Epigenome Editing ◽  
Adaptive Immune ◽  
Efficiency And Effectiveness ◽  
Associated Proteins ◽  
Rapid Pace ◽  
Molecular Diagnostic Method

The Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)-Cas (CRISPR – Associated Proteins) systems are prokaryotic adaptive immune mechanisms that are used for cleaving the invading nucleic acids in nature. Due to this reason, the CRISPR-based tools are used in numerous applications like genome and transcriptome engineering, gene therapy, epigenome editing and many others. In addition, the issue of errors in these tests is also increasing at a very rapid pace, due to which their reliability and efficiency and effectiveness are getting severely hampered. Therefore, it may not be wrong to say that there is a still a lot more room for improvement and development to enable, widespread, rapid, and scalable testing of the COVID-19 virus. In this regard, the RT-qPCR tests can be used as a routine molecular diagnostic method for detecting any COVID-19 symptoms. However, there are various limitations that mar the use and implementation of the RT-qPCR tests. But a few of these shortcomings can be overcome by using alternative molecular diagnostic methods like the CRISPR based tests.

scholarly journals Infectious Diseases Society of America Guidelines on the Diagnosis of Coronavirus Disease 2019

2020 ◽  
Author(s):  
Kimberly E Hanson ◽  
Angela M Caliendo ◽  
Cesar A Arias ◽  
Janet A Englund ◽  
Mark J Lee ◽  
...  
Keyword(s):  
Nucleic Acid ◽  
Infectious Diseases ◽  
Universal Access ◽  
Nucleic Acid Amplification ◽  
Evaluation Methodology ◽  
Molecular Diagnostic ◽  
Evidence Based ◽  
Care Hospital ◽  
Nucleic Acid Testing ◽  
Asymptomatic Individuals

Abstract IDSA Disclaimer As of the time of this publication, updates have been made to IDSA’s Guidelines on the Diagnosis of COVID-19. For the most updated version of these guidelines, please go to http://www.idsociety.org/covid19guidelines. Background Accurate molecular diagnostic tests are necessary for confirming a diagnosis of coronavirus disease 2019 (COVID-19). Direct detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) nucleic acids in respiratory tract specimens informs patient, healthcare institution, and public health–level decision-making. The number of available SARS-CoV-2 nucleic acid detection tests is rapidly increasing, as is the COVID-19 diagnostic literature. Thus, the Infectious Diseases Society of America (IDSA) recognized a significant need for frequently updated systematic reviews of the literature to inform evidence-based best practice guidance. Objective The IDSA’s goal was to develop an evidence-based diagnostic guidelines to assist clinicians, clinical laboratorians, patients, and policy makers in decisions related to the optimal use of SARS-CoV-2 nucleic acid amplification tests. In addition, the society provides a conceptual framework for understanding molecular diagnostic test performance, discusses the nuance of test result interpretation in a variety of practice settings, and highlights important unmet research needs in the COVID-19 diagnostic testing arena. Methods IDSA convened a multidisciplinary panel of infectious diseases clinicians, clinical microbiologists, and experts in systematic literature review to identify and prioritize clinical questions and outcomes related to the use of SARS-CoV-2 molecular diagnostics. Grading of Recommendations Assessment, Development and Evaluation methodology was used to assess the certainty of evidence and make testing recommendations. Results The panel agreed on 15 diagnostic recommendations. Conclusions Universal access to accurate SARS-CoV-2 nucleic acid testing is critical for patient care, hospital infection prevention, and the public response to the COVID-19 pandemic. Information on the clinical performance of available tests is rapidly emerging, but the quality of evidence of the current literature is considered low to very low. Recognizing these limitations, the IDSA panel weighed available diagnostic evidence and recommends nucleic acid testing for all symptomatic individuals suspected of having COVID-19. In addition, testing is recommended for asymptomatic individuals who have had a known or suspected contact with a COVID-19 case. Testing asymptomatic individuals without known exposure is suggested when the results will impact isolation/quarantine/personal protective equipment usage decisions, dictate eligibility for surgery, or inform administration of immunosuppressive therapy. Ultimately, prioritization of testing will depend on institution-specific resources and the needs of different patient populations.

scholarly journals Use of a NAT-based assay to improve the surveillance system and prevent transfusion-transmitted malaria in blood banks

2020 ◽  
Vol 19 (1) ◽  
Author(s):  
Daniele Rocha ◽  
Gisely Cardoso de Melo ◽  
José Marcelo Hipólito Carneiro ◽  
Marisa Ribeiro ◽  
Sthefanie Ribeiro ◽  
...  
Keyword(s):  
Nucleic Acid ◽  
Blood Donation ◽  
High Sensitivity ◽  
Nucleic Acid Amplification ◽  
Molecular Diagnostic ◽  
Hiv Rna ◽  
Immunodeficiency Virus ◽  
Blood Banks ◽  
Wide Availability

Abstract Background Malaria can be transmitted by blood transfusion through donations collected from asymptomatic donors. Transfusion-transmitted malaria (TTM) poses a great risk to blood services worldwide. A good screening tool for Plasmodium spp. detection in blood banks must have a high sensitivity for prevention of TTM. However, in Brazilian blood banks, screening for malaria still relies on microscopy. Methods In Brazil, screening for human immunodeficiency virus type 1 (HIV), RNA/DNA for hepatitis C (HCV) and hepatitis B (HBV) viruses is mandatory for every blood donation and uses nucleic acid amplification testing (NAT). The aim of this study was to evaluate the inclusion of an assay for malaria to identify Plasmodium sp. from total nucleic acid (TNA; DNA/RNA) by targeting the 18S rRNA gene of the parasite. Results Considering the limitations of microscopy and the wide availability of the Brazilian NAT platform in the screening of blood units for HIV, HCV, and HBV, a molecular diagnostic tool was validated for detection of Plasmodium sp. in blood banks; a pilot study showed that using this novel NAT assay could reduce the risk of TTM. Conclusion The prototype HIV/HCV/HBV/malaria NAT assay was effective in detecting infected candidate donors and has good prospects to be applied in routine screening for preventing TTM.

scholarly journals Development and Evaluation of a Molecular Diagnostic Method for Rapid Detection of Histoplasma capsulatum var. farciminosum, the Causative Agent of Epizootic Lymphangitis, in Equine Clinical Samples

2016 ◽  
Vol 54 (12) ◽  
pp. 2990-2999
Author(s):  
C. E. Scantlebury ◽  
G. L. Pinchbeck ◽  
P. Loughnane ◽  
N. Aklilu ◽  
T. Ashine ◽  
...  
Keyword(s):  
Causative Agent ◽  
Nested Pcr ◽  
Diagnostic Method ◽  
Histoplasma Capsulatum ◽  
Clinical Signs ◽  
Molecular Diagnostic ◽  
Clinical Samples ◽  
Blood Samples ◽  
Content Type ◽  
Molecular Diagnostic Method

Histoplasma capsulatumvar.farciminosum, the causative agent of epizootic lymphangitis (EZL), is endemic in parts of Africa. Diagnosis based on clinical signs and microscopy lacks specificity and is a barrier to further understanding this neglected disease. Here, a nested PCR method targeting the internal transcribed spacer (ITS) region of the rRNA operon was validated for application to equine clinical samples. Twenty-nine horses with signs of EZL from different climatic regions of Ethiopia were clinically examined. Blood samples and aspirates of pus from cutaneous nodules were taken, along with blood from a further 20 horses with no cutaneous EZL lesions. Among the 29 horses with suspected cases of EZL,H. capsulatumvar.farciminosumwas confirmed by extraction of DNA from pus and blood samples from 25 and 17 horses, respectively. Positive PCR results were also obtained with heat-inactivated pus (24 horses) and blood (23 horses) spotted onto Whatman FTA cards. Two positive results were obtained among blood samples from 20 horses that did not exhibit clinical signs of EZL. These are the first reports of the direct detection ofH. capsulatumvar.farciminosumin equine blood and at high frequency among horses exhibiting cutaneous lesions. The nested PCR outperformed conventional microscopic diagnosis, as characteristic yeast cells could be observed only in 14 pus samples. The presence ofH. capsulatumvar.farciminosumDNA was confirmed by sequencing the cloned PCR products, and while alignment of the ITS amplicons showed very little sequence variation, there was preliminary single nucleotide polymorphism-based evidence for the existence of two subgroups ofH. capsulatumvar.farciminosum. This molecular diagnostic method now permits investigation of the epidemiology of EZL.

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