scholarly journals Evaluation of reverse transcriptase loop-mediated isothermal amplification (RT-LAMP) assay for detection of infectious hematopoietic necrosis virus (IHNV)

2012 ◽  
Vol 25 (3) ◽  
pp. 257-262
Author(s):  
Wi-Sik Kim ◽  
Chan-Hyeok Jeon ◽  
Jeong-Ho Kim ◽  
Myung-Joo Oh
Keyword(s):  
Reverse Transcriptase ◽  
Lamp Assay ◽  
Infectious Hematopoietic Necrosis Virus ◽  
Isothermal Amplification ◽  
Necrosis Virus ◽  
Loop Mediated Isothermal Amplification ◽  
Infectious Hematopoietic Necrosis

Comparison of Reverse Transcriptase PCR, Reverse Transcriptase Loop-Mediated Isothermal Amplification, and Culture-Based Assays for Salmonella Detection from Pork Processing Environments

2011 ◽  
Vol 74 (2) ◽  
pp. 294-301 ◽  
Author(s):  
CHAYAPA TECHATHUVANAN ◽  
FRANCES ANN DRAUGHON ◽  
DORIS HELEN D'SOUZA
Keyword(s):  
Reverse Transcriptase ◽  
Salmonella Typhimurium ◽  
Real Time ◽  
Lamp Assay ◽  
Isothermal Amplification ◽  
Water Bath ◽  
Detection Sensitivity ◽  
Rt Pcr ◽  
Reverse Transcriptase Pcr ◽  
Loop Mediated Isothermal Amplification

Novel rapid Salmonella detection assays without the need for sophisticated equipment or labor remain in high demand. Real-time reverse transcriptase PCR (RT-PCR) assays, though rapid and sensitive, require expensive thermocyclers, while a novel RT loop-mediated isothermal amplification (RT-LAMP) method requires only a simple water bath. Our objective was to compare the detection sensitivity of Salmonella Typhimurium from the pork processing environment by RT-LAMP, RT-PCR, and culture-based assays. Carcass and surface swabs and carcass rinses were obtained from a local processing plant. Autoclaved carcass rinses (500 ml) were spiked with Salmonella Typhimurium and filtered. Filters were placed in stomacher bags containing tetrathionate broth (TTB) and analyzed with or without 10-h enrichment at 37°C. Natural swabs were stomached with buffered peptone water, and natural carcass rinses were filtered, preenriched, and further enriched in TTB. Serially-diluted enriched samples were enumerated by spread plating on xylose lysine Tergitol 4 agar. RNA was extracted from 5 ml of enriched TTB with TRIzol. RT-LAMP assay using previously described invA primers was conducted at 62°C for 90 min in a water bath with visual detection and by gel electrophoresis. SYBR Green I–based-real-time RT-PCR was carried out with invA primers followed by melt temperature analysis. The results of RT-LAMP detection for spiked carcass rinses were comparable to those of RT-PCR and cultural plating, with detection limits of 1 log CFU/ml, although they were obtained significantly faster, within 24 h including preenrichment and enrichment. RT-LAMP showed 4 of 12 rinse samples positive, while RT-PCR showed 1 of 12 rinse samples positive. For swabs, 6 of 27 samples positive by RT-LAMP and 5 of 27 by RT-PCR were obtained. This 1-day RT-LAMP assay shows promise for routine Salmonella screening by the pork industry.

scholarly journals A semi-automated, isolation-free, high-throughput SARS-CoV-2 reverse transcriptase (RT) loop-mediated isothermal amplification (LAMP) test

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Jonas Schmidt ◽  
Sandro Berghaus ◽  
Frithjof Blessing ◽  
Folker Wenzel ◽  
Holger Herbeck ◽  
...  
Keyword(s):  
Reverse Transcriptase ◽  
Mass Screening ◽  
High Throughput ◽  
Clinical Laboratory ◽  
Limit Of Detection ◽  
Lamp Assay ◽  
Isothermal Amplification ◽  
Rt Pcr ◽  
Loop Mediated Isothermal Amplification ◽  
Clinical Diagnostic

AbstractShortages of reverse transcriptase (RT)-polymerase chain reaction (PCR) reagents and related equipment during the COVID-19 pandemic have demonstrated the need for alternative, high-throughput methods for severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2)-mass screening in clinical diagnostic laboratories. A robust, SARS-CoV-2 RT-loop-mediated isothermal amplification (RT-LAMP) assay with high-throughput and short turnaround times in a clinical laboratory setting was established and compared to two conventional RT-PCR protocols using 323 samples of individuals with suspected SARS-CoV-2 infection. Limit of detection (LoD) and reproducibility of the isolation-free SARS-CoV-2 RT-LAMP test were determined. An almost perfect agreement (Cohen’s kappa > 0.8) between the novel test and two classical RT-PCR protocols with no systematic difference (McNemar’s test, P > 0.05) was observed. Sensitivity and specificity were in the range of 89.5 to 100% and 96.2 to 100% dependent on the reaction condition and the RT-PCR method used as reference. The isolation-free RT-LAMP assay showed high reproducibility (Tt intra-run coefficient of variation [CV] = 0.4%, Tt inter-run CV = 2.1%) with a LoD of 95 SARS-CoV-2 genome copies per reaction. The established SARS-CoV-2 RT-LAMP assay is a flexible and efficient alternative to conventional RT-PCR protocols, suitable for SARS-CoV-2 mass screening using existing laboratory infrastructure in clinical diagnostic laboratories.

scholarly journals Reverse Transcriptase Loop Mediated Isothermal Amplification (RT-LAMP) for COVID-19 Diagnosis: A Systematic Review and Meta-Analysis

2021 ◽  
Author(s):  
Anita Dominique Subali ◽  
Lowilius Wiyono
Keyword(s):  
Systematic Review ◽  
Reverse Transcriptase ◽  
Quality Assessment ◽  
Meta Analysis ◽  
Lamp Assay ◽  
Isothermal Amplification ◽  
Diagnostic Methods ◽  
Clinical Samples ◽  
Rt Pcr ◽  
Loop Mediated Isothermal Amplification

ABSTRACTBackgroundCoronavirus Disease 2019 (COVID-19) has caused a severe outbreak and become a global public health priority. Rapid increment of infection number along with significant deaths have placed the virus as a serious threat to human health. Rapid, reliable, and simple diagnostic methods are critically essential for disease control. While Reverse Transcriptase Polymerase Chain Reaction (RT-PCR) is the current diagnostic gold standard, Reverse Transcriptase Loop-Mediated Isothermal Amplification (RT-LAMP) appears as a compelling alternative diagnostic test due to its more simplicity, shorter time to result, and lower cost. This study examined RT-LAMP application for rapid identification of SARS-CoV-2 infection compared to RT-PCR assay.MethodsA systematic review and meta-analysis (2020) was conducted in 6 scientific databases following the PRISMA Guideline. Original published studies on human clinical samples in English were included. Articles evaluated sensitivity and specificity of RT-LAMP relative to RT-PCR were considered eligible. Quality assessment of bias and applicability was examined based on QUADAS-2.ResultsA total of 351 studies were found based on the keywords and search queries. 14 eligible case control studies fitted the respective criteria. Quality assessment using QUADAS-2 indicated low risk bias in all included studies. All case studies, comprises 2,112 samples, had the cumulative sensitivity of 95.5% (CI 97.5%=90.8-97.9%) and cumulative specificity of 99.5% (CI 97.5%=97.7-99.9%).ConclusionRT-LAMP assay could be suggested as a reliable alternative COVID-19 diagnostic method with reduced cost and time compared to RT-PCR. RT-LAMP could potentially be utilized during the high-throughput and high-demand critical situations.

scholarly journals Universal reverse-transcriptase real-time PCR for infectious hematopoietic necrosis virus (IHNV)

2013 ◽  
Vol 106 (2) ◽  
pp. 103-115 ◽  
Author(s):  
MK Purcell ◽  
RL Thompson ◽  
KA Garver ◽  
LM Hawley ◽  
WN Batts ◽  
...  
Keyword(s):  
Reverse Transcriptase ◽  
Real Time ◽  
Real Time Pcr ◽  
Infectious Hematopoietic Necrosis Virus ◽  
Necrosis Virus ◽  
Infectious Hematopoietic Necrosis
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