Detecting wMel Wolbachia in field-collected Aedes aegypti mosquitoes using loop-mediated isothermal amplification (LAMP)

Goncalves, Daniela da Silva; Hooker, David J.; Dong, Yi; Baran, Nathan; Kyrylos, Peter; Iturbe‐Ormaetxe, Iñaki; Simmons, Cameron P.; O’Neill, Scott L.

doi:10.1186/s13071-019-3666-6

Cited by 29 publications

(27 citation statements)

References 37 publications

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“…These results are consistent with other published results that rely on colorimetric LAMP reactions, rather than on probe-based detection. 21 In fact, for many published assays, color changes must be read within a narrow window of time in order to minimize spurious conclusions, a consideration that does not scale well for diagnostic screening, especially at point-of-care or as an early part of a clinical diagnostics pipeline.…”

Section: Resultsmentioning

confidence: 99%

High-surety isothermal amplification and detection of SARS-CoV-2, including with crude enzymes

Bhadra

Lakhotia

et al. 2020

Preprint

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ABSTRACTIsothermal nucleic acid amplification tests (iNAT), such as loop-mediated isothermal amplification (LAMP), are good alternatives to polymerase chain reaction (PCR)-based amplification assays, especially for point-of-care and low resource use, in part because they can be carried out with relatively simple instrumentation. However, iNATs can generate spurious amplicons, especially in the absence of target sequences, resulting in false positive results. This is especially true if signals are based on non-sequence-specific probes, such as intercalating dyes or pH changes. In addition, pathogens often prove to be moving, evolving targets, and can accumulate mutations that will lead to inefficient primer binding and thus false negative results. Internally redundant assays targeting different regions of the target sequence can help to reduce such false negatives. Here we describe rapid conversion of three previously described SARS-CoV-2 LAMP assays that relied on non-sequence-specific readout into assays that can be visually read using sequence-specific fluorogenic oligonucleotide strand exchange (OSD) probes. We evaluate one-pot operation of both individual and multiplex LAMP-OSD assays and demonstrate detection of SARS-CoV-2 virions in crude human saliva.

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Section: Resultsmentioning

confidence: 99%

High-surety isothermal amplification and detection of SARS-CoV-2, including with crude enzymes

Bhadra

Lakhotia

et al. 2020

Preprint

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“…We recently developed a method for visual detection of LAMP amplification (9) using pH-sensitive dyes to exploit the change of pH that resulting from proton accumulation due to dNTP incorporation. This method has been used in a large scale field survey of Wolbachia-containing mosquitos (10), Grapevine red blotch virus without DNA extraction (11), testing urine samples for Zika virus (12) and even amplification detection on the International Space Station (13). The breadth of application highlights the applicability of visual detection methods to provide an advantage in simplicity and portability for enabling new, rapid diagnostics.…”

Section: Introductionmentioning

confidence: 99%

Rapid Molecular Detection of SARS-CoV-2 (COVID-19) Virus RNA Using Colorimetric LAMP

Zhang

Odiwuor

Xiong

et al. 2020

Preprint

359

477

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The ability to detect an infectious agent in a widespread epidemic is crucial to the success of quarantine efforts in addition to sensitive and accurate screening of potential cases of infection from patients in a clinical setting. Enabling testing outside of sophisticated laboratories broadens the scope of control and surveillance efforts, but also requires robust and simple methods that can be used without expensive instrumentation. Here we report a method to identify SARS-CoV-2 (COVID-19) virus RNA from purified RNA or cell lysis using loop-mediated isothermal amplification (LAMP) using a visual, colorimetric detection. This test was additionally verified using RNA samples purified from respiratory swabs collected from COVID-19 patients in Wuhan, China with equivalent performance to a commercial RT-qPCR test while requiring only heating and visual inspection. This simple and sensitive method provides an opportunity to facilitate virus detection in the field without a requirement for complex diagnostic infrastructure.. CC-BY 4.0 International license It is made available under a author/funder, who has granted medRxiv a license to display the preprint in perpetuity.is the (which was not peer-reviewed) The copyright holder for this preprint .

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“…The assays may be used as standalone tests, or as an initial screen followed by confirmatory laboratory testing using alternative diagnostic methods under more controlled conditions. Whilst we developed assays for mosquito species identification, LAMP technology has been applied to detection of arboviruses [23] and Wolbachia pipientis [22,24] in Ae. aegypti.…”

Section: Discussionmentioning

confidence: 99%

A LAMP-based colorimetric assay to expedite field surveillance of the invasive mosquito species Aedes aegypti and Aedes albopictus

et al. 2020

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Background Yellow fever, dengue, chikungunya and Zika viruses are responsible for considerable morbidity and mortality in humans. Aedes aegypti and Aedes albopictus are the most important mosquito vectors involved in their transmission. Accurate identification of these species is essential for the implementation of control programs to limit arbovirus transmission, during suspected detections at ports of first entry, to delimit incursions or during presence/absence surveillance programs in regions vulnerable to invasion. We developed and evaluated simple and rapid colorimetric isothermal tests to detect these two mosquito species based on loop-mediated isothermal amplification (LAMP) targeting the ribosomal RNA internal transcribed spacer 1 (ITS1). Methodology/Principal findings Samples were prepared by homogenizing and heating at 99 o C for 10 min before an aliquot was added to the LAMP reaction. After 40 min incubation at 65 o C, a colour change indicated a positive result. The tests were 100% sensitive and species-specific, and demonstrated a limit of detection comparable with PCR-based detection (TaqMan chemistry). The LAMP assays were able to detect target species for various life stages tested (adult, 1 st instar larva, 4 th instar larva and pupa), and body components, such as legs, wings and pupal exuviae. Importantly, the LAMP assays could detect Ae. aegypti DNA in mosquitoes stored in Biogents Sentinel traps deployed in the field for 14 d. A single 1 st instar Ae. aegypti larva could also be detected in a pool of 1,000 non-target 1 st instar Aedes notoscriptus, thus expediting processing of ovitrap collections obtained during presence/absence surveys. A simple syringe-sponge protocol facilitated the concentration and collection of larvae from the ovitrap water post-hatch.

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Detecting wMel Wolbachia in field-collected Aedes aegypti mosquitoes using loop-mediated isothermal amplification (LAMP)

Cited by 29 publications

References 37 publications

High-surety isothermal amplification and detection of SARS-CoV-2, including with crude enzymes

High-surety isothermal amplification and detection of SARS-CoV-2, including with crude enzymes

Rapid Molecular Detection of SARS-CoV-2 (COVID-19) Virus RNA Using Colorimetric LAMP

A LAMP-based colorimetric assay to expedite field surveillance of the invasive mosquito species Aedes aegypti and Aedes albopictus

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