Loop Mediated Isothermal Amplification: Principles and Applications in Plant Virology

Panno, Stefano; Matić, S.; Tiberini, Antonio; Caruso, A.; Bella, P.; Torta, Livio; Stassi, Raffaele; Davino, Salvatore

doi:10.3390/plants9040461

Cited by 156 publications

(125 citation statements)

References 189 publications

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“…Reverse transcriptase and RNase H are used to synthesize complementary ds DNA, and a T7 RNA polymerase to synthesize complementary RNA strands resulting in amplification. iv) Loop-mediated isothermal amplification (LAMP) is based on auto cycling and high DNA strand displacement activity mediated by Bst polymerase from Geobacillus stearothermophilus, under isothermal conditions at 60°C to 65°C (Panno et al, 2020). Recently, new tools for molecular diagnosis have been developed based on prokaryotic clustered regularly interspaced short palindromic repeats (CRISPR) immunity system, widely applied for genome editing (Chertow, 2018).…”

Section: Serological and Molecular Techniquesmentioning

confidence: 99%

Detection of Plant Viruses and Disease Management: Relevance of Genetic Diversity and Evolution

2020

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Plant viruses cause considerable economic losses and are a threat for sustainable agriculture. The frequent emergence of new viral diseases is mainly due to international trade, climate change, and the ability of viruses for rapid evolution. Disease control is based on two strategies: i) immunization (genetic resistance obtained by plant breeding, plant transformation, cross-protection, or others), and ii) prophylaxis to restrain virus dispersion (using quarantine, certification, removal of infected plants, control of natural vectors, or other procedures). Disease management relies strongly on a fast and accurate identification of the causal agent. For known viruses, diagnosis consists in assigning a virus infecting a plant sample to a group of viruses sharing common characteristics, which is usually referred to as species. However, the specificity of diagnosis can also reach higher taxonomic levels, as genus or family, or lower levels, as strain or variant. Diagnostic procedures must be optimized for accuracy by detecting the maximum number of members within the group (sensitivity as the true positive rate) and distinguishing them from outgroup viruses (specificity as the true negative rate). This requires information on the genetic relationships within-group and with members of other groups. The influence of the genetic diversity of virus populations in diagnosis and disease management is well documented, but information on how to integrate the genetic diversity in the detection methods is still scarce. Here we review the techniques used for plant virus diagnosis and disease control, including characteristics such as accuracy, detection level, multiplexing, quantification, portability, and designability. The effect of genetic diversity and evolution of plant viruses in the design and performance of some detection and disease control techniques are also discussed. High-throughput or next-generation sequencing provides broad-spectrum and accurate identification of viruses enabling multiplex detection, quantification, and the discovery of new viruses. Likely, this technique will be the future standard in diagnostics as its cost will be dropping and becoming more affordable.

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Section: Serological and Molecular Techniquesmentioning

confidence: 99%

Detection of Plant Viruses and Disease Management: Relevance of Genetic Diversity and Evolution

2020

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“…The detection of LAMP products was achieved using a laboratory real-time PCR instrument to measure fluorescence. Alternative detection methods such as agarose gel electrophoresis can be used 26 but these increase the risk of contamination through the opening of reaction tubes containing LAMP-amplified products 18 . LAMP reactions generate five to ten more amplicons than a standard PCR 27,28 .…”

Section: Discussionmentioning

confidence: 99%

“…Although the real-time detection used in our study avoided any risk of contamination associated with post-amplification processes there are several low-cost portable fluorescence-reading instruments (e.g. Genie II (Optigene Ltd., West Sussex, UK), Bioranger (Diagenetix, Inc., Honolulu, HI, USA) and Genelyzer FIII (Canon Medical Systems, Tochigi, Japan)) available in the market that can be used to measure the fluorescence signal and thereby reducing the cost of the assay 18,30,31 . These simple battery-powered instruments offer a major advantage of the LAMP assay in places experiencing repeated power failures as reactions could proceed even without electricity.…”

Section: Discussionmentioning

confidence: 99%

“…In this study the reaction time of the LAMP assay was less than 40 min compared to > 90 min required for the PCR plus the time required for gel electrophoresis. Previous reports have described the use of LAMP in conjunction with crude plant extracts avoiding total DNA extractions 19,32 , reducing costs 18 , shortening the processing time and facilitating the use of the method in low-resource settings 32 . In contrast to LAMP, PCR needs more stringent conditions such as expensive equipment to perform thermal cycling steps at higher temperatures and highly specialised personnel.…”

Section: Discussionmentioning

confidence: 99%

“…In many African countries including Zambia, where there is inadequate laboratory equipment, isothermal amplification systems such as loop-mediated isothermal amplification (LAMP) offer a useful alternative. Lately, numerous LAMP assays have led to the development of several phytopathological diagnostic protocols for the early detection of many diseases caused by viruses, however, none has been developed for the detection of MSV 18 .…”

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confidence: 99%

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A quick and sensitive diagnostic tool for detection of Maize streak virus

Tembo

Adediji

Bouvaine

et al. 2020

Sci Rep

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Maize streak virus disease (MSVD), caused by Maize streak virus (MSV; genus Mastrevirus), is one of the most severe and widespread viral diseases that adversely reduces maize yield and threatens food security in Africa. An effective control and management of MSVD requires robust and sensitive diagnostic tests capable of rapid detection of MSV. In this study, a loop-mediated isothermal amplification (LAMP) assay was designed for the specific detection of MSV. This test has shown to be highly specific and reproducible and able to detect MSV in as little as 10 fg/µl of purified genomic DNA obtained from a MSV-infected maize plant, a sensitivity 105 times higher to that obtained with polymerase chain reaction (PCR) in current general use. The high degree of sequence identity between Zambian and other African MSV isolates indicate that this LAMP assay can be used for detecting MSV in maize samples from any region in Africa. Furthermore, this assay can be adopted in minimally equipped laboratories and with potential use in plant clinic laboratories across Africa strengthening diagnostic capacity in countries dealing with MSD.

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The next‐generation coronavirus diagnostic techniques with particular emphasis on the SARS‐CoV‐2

Hemida

2021

Journal of Medical Virology

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The potential zoonotic coronaviruses (SARS-CoV, MERS-CoV, and SARS-CoV-2) are of global health concerns. Early diagnosis is the milestone in their mitigation, control, and eradication. Many diagnostic techniques are showing great success and have many advantages, such as the rapid turnover of the results, high accuracy, and high specificity and sensitivity. However, some of these techniques have several pitfalls if samples were not collected, processed, and transported in the standard ways and if these techniques were not practiced with extreme caution and precision. This may lead to false-negative/positive results. This may affect the downstream management of the affected cases. These techniques require regular finetuning, upgrading, and optimization. The continuous evolution of new strains and viruses belong to the coronaviruses is hampering the success of many classical techniques. There are urgent needs for next generations of coronaviruses diagnostic assays that overcome these pitfalls. This new generation of diagnostic tests should be able to do simultaneous, multiplex, and high-throughput detection of various coronavirus in one reaction. Furthermore, the development of novel assays and techniques that enable the in situ detection of the virus on the environmental samples, especially air, water, and surfaces, should be given considerable attention in the future. These approaches will have a substantial positive impact on the mitigation and eradication of coronaviruses, including the current SARS-CoV-2 pandemic.

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Loop Mediated Isothermal Amplification: Principles and Applications in Plant Virology

Cited by 156 publications

References 189 publications

Detection of Plant Viruses and Disease Management: Relevance of Genetic Diversity and Evolution

Detection of Plant Viruses and Disease Management: Relevance of Genetic Diversity and Evolution

A quick and sensitive diagnostic tool for detection of Maize streak virus

The next‐generation coronavirus diagnostic techniques with particular emphasis on the SARS‐CoV‐2

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