Rapid and in-situ detection of fecal indicator bacteria in water using simple DNA extraction and portable loop-mediated isothermal amplification (LAMP) PCR methods

Lee, Seunguk; Khoo, Valerie Si Ling; Medriano, Carl Angelo; Lee, Taewoo; Park, Sung-Yong; Bae, Sung Woo

doi:10.1016/j.watres.2019.05.049

Cited by 62 publications

(49 citation statements)

References 23 publications

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“…As an alternative, researchers have been developing assays based on naked-eye observation 26,31 with end-point colorimetric LAMP being the most popular assay [32][33][34][35][36][37] . The above can employ simple means of heating e.g., a water bath, a thermos etc.…”

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confidence: 99%

“…The above can employ simple means of heating e.g., a water bath, a thermos etc. or portable and/or battery operated instruments for isothermal amplification and eye-observation, all suitable for applications in resource-limited areas 34,38 . Downsides of the naked-eye detection are the need of a "trained eye" for results interpretation, especially at low numbers of the target, and the qualitative nature of the results 26 .…”

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confidence: 99%

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Portable real-time colorimetric LAMP-device for rapid quantitative detection of nucleic acids in crude samples

Papadakis

Pantazis

Fikas

et al. 2020

Preprint

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Most methods applied to nucleic acids’ detection at the point-of-care require either expensive and mostly bench-top instruments or simpler inexpensive systems providing qualitative results. Truly decentralized approaches to reliable, quantitative and affordable diagnostics are still missing. Here, we report the development of real-time quantitative colorimetric LAMP based on a portable and cost-effective device, the use of which requires minimal training. Main advantages of the method are the rapid analysis time (<30min); quantification over a large dynamic range (9 log units); ability to work with crude samples (saliva, tissue); demonstrated low detection limit (1-10 copies); smartphone-operation and fast prototyping (3D-printing). The system’s broad detection capability is demonstrated during infectious diseases-testing for COVID-19 and pharmacogenetics for BRAF V600E mutation testing. Validation studies showed 97.4% and 100% agreement with qRT-PCR for SARS-CoV-2 RNA detection extracted from positive and negative patients’ samples (89), respectively; and 100% agreement with ddPCR and Sanger sequencing for BRAF V600E mutation detection from 12 clinical biopsy samples. The new methodology provides a needed solution for affordable healthcare at the point-of-care, with emphasis on global diagnostics.

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confidence: 99%

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confidence: 99%

Portable real-time colorimetric LAMP-device for rapid quantitative detection of nucleic acids in crude samples

Papadakis

Pantazis

Fikas

et al. 2020

Preprint

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“…The reason could be the presence of PCR inhibitors in the samples or a contaminating background which influences one method more than the other. The research group of Lee et al came to similar conclusions, whereby LAMP was more tolerant to PCR inhibitors than conventional PCR and therefore yielded better results [50].…”

Section: Discussionmentioning

confidence: 82%

Development and Validation of a Loop-Mediated Isothermal Amplification (LAMP) Assay for Rapid Detection of Glaesserella (Haemophilus) parasuis

et al. 2020

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Glaesserella parasuis is a fastidious pathogen that colonizes the respiratory tract of pigs and can lead to considerable economic losses in pig production. Therefore, a rapid detection assay for the pathogen, preferably applicable in the field, is important. In the current study, we developed a new and improved detection method using loop-mediated isothermal amplification (LAMP). This assay, which targets the infB gene, was tested on a collection of 60 field isolates of G. parasuis comprising 14 different serovars. In addition, 63 isolates from seven different closely related species of the family Pasteurellaceae, including A. indolicus, A. porcinus, and A. minor, and a species frequently found in the respiratory tract of pigs were used for exclusivity experiments. This assay showed an analytical specificity of 100% (both inclusivity and exclusivity) and an analytical sensitivity of 10 fg/µL. In further steps, 36 clinical samples were tested with the LAMP assay. An agreement of 77.1 (95% CI: 59.9, 89.6) and 91.4% (95% CI: 75.9, 98.2) to the culture-based and PCR results was achieved. The mean limit of detection for the spiked bronchoalveolar lavage fluid was 2.58 × 102 CFU/mL. A colorimetric assay with visual detection by the naked eye was tested to provide an alternative method in the field and showed the same sensitivity as the fluorescence-based LAMP assay. Overall, the optimized LAMP assay represents a fast and reliable method and is suitable for detecting G. parasuis in the laboratory environment or in the field.

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“…Water from the Sargasso Sea, a region with known low nutrient content [ 22 ], was filtered through a 0.2 µm filter to remove organic matter prior to use, as the presence of residual particulate matter has been shown to affect the accuracy and precision of colorimetric detection schemes [ 23 ]. Of note, the use of a simple syringe filter for this step means that in-field applications will be able to use similar filters without compromising widespread applicability of this method [ 24 ].…”

Section: Methodsmentioning

confidence: 99%

A Paper-Based Device for Ultrasensitive, Colorimetric Phosphate Detection in Seawater

Racicot

Mako

Olivelli

et al. 2020

Sensors

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High concentrations of certain nutrients, including phosphate, are known to lead to undesired algal growth and low dissolved oxygen levels, creating deadly conditions for organisms in marine ecosystems. The rapid and robust detection of these nutrients using a colorimetric, paper-based system that can be applied on-site is of high interest to individuals monitoring marine environments and others affected by marine ecosystem health. Several techniques for detecting phosphate have been reported previously, yet these techniques often suffer from high detection limits, reagent instability, and the need of the user to handle toxic reagents. In order to develop improved phosphate detection methods, the commonly used molybdenum blue reagents were incorporated into a paper-based, colorimetric detection system. This system benefited from improved stabilization of the molybdenum blue reagent as well as minimal user contact with toxic reagents. The colorimetric readout from the paper-based devices was analyzed and quantified using RGB analyses (via ImageJ), and resulted in the detection of phosphate at detection limits between 1.3 and 2.8 ppm in various aqueous media, including real seawater.

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Rapid and in-situ detection of fecal indicator bacteria in water using simple DNA extraction and portable loop-mediated isothermal amplification (LAMP) PCR methods

Cited by 62 publications

References 23 publications

Portable real-time colorimetric LAMP-device for rapid quantitative detection of nucleic acids in crude samples

Portable real-time colorimetric LAMP-device for rapid quantitative detection of nucleic acids in crude samples

Development and Validation of a Loop-Mediated Isothermal Amplification (LAMP) Assay for Rapid Detection of Glaesserella (Haemophilus) parasuis

A Paper-Based Device for Ultrasensitive, Colorimetric Phosphate Detection in Seawater

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