The development and application of a LAMP assay for visualized detection of Chinese olive anthracnose

AbstractTilletia controversa causing dwarf bunt of wheat is a quarantine pathogen in several countries. Therefore, its specific detection is of great phytosanitary importance. Genomic regions routinely used for phylogenetic inferences lack suitable polymorphisms for the development of species-specific markers. We therefore compared 21 genomes of six Tilletia species to identify DNA regions that were unique and conserved in all T. controversa isolates and had no or limited homology to other Tilletia species. A loop-mediated isothermal amplification (LAMP) assay for T. controversa was developed based on one of these DNA regions. The specificity of the assay was verified using 223 fungal samples comprising 43 fungal species including 11 Tilletia species, in particular 39 specimens of T. controversa, 92 of T. caries and 40 of T. laevis, respectively. The assay specifically amplified genomic DNA of T. controversa from pure cultures and teliospores. Only Tilletia trabutii generated false positive signals. The detection limit of the LAMP assay was 5 pg of genomic DNA per reaction. A test performance study that included five laboratories in Germany resulted in 100% sensitivity and 97.7% specificity of the assay. Genomic regions, specific to common bunt (Tilletia caries and Tilletia laevis together) are also provided.

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Development and validation of a one-step reverse transcription loop-mediated isothermal amplification (RT-LAMP) for rapid detection of ZIKV in patient samples from Brazil

Scientific Reports ◽

10.1038/s41598-021-83371-1 ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Severino Jefferson Ribeiro da Silva ◽

Keith Pardee ◽

Udeni B. R. Balasuriya ◽

Lindomar Pena

Keyword(s):

Reverse Transcription ◽

Rapid Detection ◽

Quantitative Polymerase Chain Reaction ◽

Rna Extraction ◽

Lamp Assay ◽

Isothermal Amplification ◽

Serum Samples ◽

Low Resource ◽

Loop Mediated Isothermal Amplification ◽

One Step

AbstractWe have previously developed and validated a one-step assay based on reverse transcription loop-mediated isothermal amplification (RT-LAMP) for rapid detection of the Zika virus (ZIKV) from mosquito samples. Patient diagnosis of ZIKV is currently carried out in centralized laboratories using the reverse transcription-quantitative polymerase chain reaction (RT-qPCR), which, while the gold standard molecular method, has several drawbacks for use in remote and low-resource settings, such as high cost and the need of specialized equipment. Point-of-care (POC) diagnostic platforms have the potential to overcome these limitations, especially in low-resource countries where ZIKV is endemic. With this in mind, here we optimized and validated our RT-LAMP assay for rapid detection of ZIKV from patient samples. We found that the assay detected ZIKV from diverse sample types (serum, urine, saliva, and semen) in as little as 20 min, without RNA extraction. The RT-LAMP assay was highly specific and up to 100 times more sensitive than RT-qPCR. We then validated the assay using 100 patient serum samples collected from suspected cases of arbovirus infection in the state of Pernambuco, which was at the epicenter of the last Zika epidemic. Analysis of the results, in comparison to RT-qPCR, found that the ZIKV RT-LAMP assay provided sensitivity of 100%, specificity of 93.75%, and an overall accuracy of 95.00%. Taken together, the RT-LAMP assay provides a straightforward and inexpensive alternative for the diagnosis of ZIKV from patients and has the potential to increase diagnostic capacity in ZIKV-affected areas, particularly in low and middle-income countries.

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Visualized Detection of Aldehyde Dehydrogenase 2 Gene Polymorphism By Serial Invasive Reaction Coupled with Gold Nanoparticle Probe Assembling

Chinese Journal of Analytical Chemistry ◽

10.1016/s1872-2040(20)60072-x ◽

2021 ◽

Vol 49 (1) ◽

pp. 42-49

Author(s):

Nan SHENG ◽

Hui-Jie YUE ◽

Shu-Yun PANG ◽

Xie-Min QI ◽

Yan-Jie ZHANG ◽

...

Keyword(s):

Gene Polymorphism ◽

Gold Nanoparticle ◽

Aldehyde Dehydrogenase ◽

Aldehyde Dehydrogenase 2 ◽

Aldehyde Dehydrogenase 2 Gene ◽

Visualized Detection

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A novel RdRp-based colorimetric RT-LAMP assay for rapid and sensitive detection of SARS-CoV-2 in clinical and sewage samples from Pakistan

Virus Research ◽

10.1016/j.virusres.2021.198484 ◽

2021 ◽

pp. 198484

Author(s):

Muhammad Farhan Ul Haque ◽

Syeda Sadia Bukhari ◽

Rabia Ejaz ◽

Faheem Uz Zaman ◽

Kamalalayam Rajan Sreejith ◽

...

Keyword(s):

Lamp Assay ◽

Sensitive Detection

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630. A 5-mRNA host response whole-blood classifier trained using patients with non-COVID-19 viral infections accurately predicts severity of COVID-19

Open Forum Infectious Diseases ◽

10.1093/ofid/ofaa439.824 ◽

2020 ◽

Vol 7 (Supplement_1) ◽

pp. S375-S376

Author(s):

ljubomir Buturovic ◽

Purvesh Khatri ◽

Benjamin Tang ◽

Kevin Lai ◽

Win Sen Kuan ◽

...

Keyword(s):

Logistic Regression ◽

Respiratory Failure ◽

Viral Infections ◽

Lamp Assay ◽

Training Data ◽

Diagnostic Tools ◽

Severe Respiratory Failure ◽

Proof Of Concept ◽

Risk Of Death ◽

Qpcr Platform

Abstract Background While major progress has been made to establish diagnostic tools for the diagnosis of SARS-CoV-2 infection, determining the severity of COVID-19 remains an unmet medical need. With limited hospital resources, gauging severity would allow for some patients to safely recover in home quarantine while ensuring sicker patients get needed care. We discovered a 5 host mRNA-based classifier for the severity of influenza and other acute viral infections and validated the classifier in COVID-19 patients from Greece. Methods We used training data (N=705) from 21 retrospective clinical studies of influenza and other viral illnesses. Five host mRNAs from a preselected panel were applied to train a logistic regression classifier for predicting 30-day mortality in influenza and other viral illnesses. We then applied this classifier, with fixed weights, to an independent cohort of subjects with confirmed COVID-19 from Athens, Greece (N=71) using NanoString nCounter. Finally, we developed a proof-of-concept rapid, isothermal qRT-LAMP assay for the 5-mRNA host signature using the QuantStudio 6 qPCR platform. Results In 71 patients with COVID-19, the 5 mRNA classifier had an AUROC of 0.88 (95% CI 0.80-0.97) for identifying patients with severe respiratory failure and/or 30-day mortality (Figure 1). Applying a preset cutoff based on training data, the 5-mRNA classifier had 100% sensitivity and 46% specificity for identifying mortality, and 88% sensitivity and 68% specificity for identifying severe respiratory failure. Finally, our proof-of-concept qRT-LAMP assay showed high correlation with the reference NanoString 5-mRNA classifier (r=0.95). Figure 1. Validation of the 5-mRNA classifier in the COVID-19 cohort. (A) Expression of the 5 genes used in the logistic regression model in patients with (red) and without (blue) mortality. (B) The 5-mRNA classifier accurately distinguishes non-severe and severe patients with COVID-19 as well as those at risk of death. Conclusion Our 5-mRNA classifier demonstrated very high accuracy for the prediction of COVID-19 severity and could assist in the rapid, point-of-impact assessment of patients with confirmed COVID-19 to determine level of care thereby improving patient management and healthcare burden. Disclosures ljubomir Buturovic, PhD, Inflammatix Inc. (Employee, Shareholder) Purvesh Khatri, PhD, Inflammatix Inc. (Shareholder) Oliver Liesenfeld, MD, Inflammatix Inc. (Employee, Shareholder) James Wacker, n/a, Inflammatix Inc. (Employee, Shareholder) Uros Midic, PhD, Inflammatix Inc. (Employee, Shareholder) Roland Luethy, PhD, Inflammatix Inc. (Employee, Shareholder) David C. Rawling, PhD, Inflammatix Inc. (Employee, Shareholder) Timothy Sweeney, MD, Inflammatix, Inc. (Employee)

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