Paper Device Combining CRISPR/Cas12a and Reverse-Transcription Loop-Mediated Isothermal Amplification for SARS-CoV-2 Detection in Wastewater

Cao, Haorui; Mao, Kang; Fang, Ran; Xu, Pengqi; Zhao, Yirong; Zhang, Xiangyan; Zhou, Hourong; Yang, Zhugen; Zhang, Hua; Jiang, Guibin

doi:10.1021/acs.est.2c04727

Cited by 53 publications

(39 citation statements)

References 39 publications

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“…Ramírez-Chavarría et al combined an electrochemical sensor with RT-LAMP for the detection of SARS-CoV-2 in wastewater samples and showed that the sensor was able to specifically quantify RT-LAMP amplicons at a level below 2.5 × 10 –6 ng/μL, showing high reproducibility (Ramírez-Chavarría et al 2022 ). Cao et al developed a portable paper-based device based on CRISPR/Cas12a and reverse transcription LAMP for semi-quantitative analysis of SARS-CoV-2 in wastewater, and the device achieved semi-quantitative analysis from 0 to 310 copies/mL with good sensitivity and specificity, which is a method that could be a promising approach for wastewater monitoring (Cao et al 2022 ).…”

Section: Resultsmentioning

confidence: 99%

SARS-CoV-2 surveillance in medical and industrial wastewater—a global perspective: a narrative review

Yang

Guo

Møhlenberg

et al. 2023

Environ Sci Pollut Res

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The novel coronavirus SARS-CoV-2 has spread at an unprecedented rate since late 2019, leading to the global COVID-19 pandemic. During the pandemic, being able to detect SARS-CoV-2 in human populations with high coverage quickly is a huge challenge. As SARS-CoV-2 is excreted in human excreta and thus exposed to the aqueous environment through sewers, the goal is to develop an ideal, non-invasive, cost-effective epidemiological method for detecting SARS-CoV-2. Wastewater surveillance has gained widespread interest and is increasingly being investigated as an effective early warning tool for monitoring the spread and evolution of the virus. This review emphasizes important findings on SARS-CoV-2 wastewater-based epidemiology (WBE) in different continents and techniques used to detect SARS-CoV-2 in wastewater during the period 2020–2022. The results show that WBE is a valuable population-level method for monitoring SARS-CoV-2 and is a valuable early warning alert. It can assist policymakers in formulating relevant policies to avoid the negative impacts of early or delayed action. Such strategy can also help avoid unnecessary wastage of medical resources, rationalize vaccine distribution, assist early detection, and contain large-scale outbreaks.

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Section: Resultsmentioning

confidence: 99%

SARS-CoV-2 surveillance in medical and industrial wastewater—a global perspective: a narrative review

Yang

Guo

Møhlenberg

et al. 2023

Environ Sci Pollut Res

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“…This is the first report of a LAMP-CRISPR detection system on paper. Previously all LAMP-CRISPR reactions have been carried out in tubes or in combinations of tubes plus lateral flow devices [8], [30].…”

Section: Sensitivity Studiesmentioning

confidence: 99%

A paper based microfluidic platform combining LAMP-CRISPR/Cas12a for fluorometric detection of nucleic acids

Sen¹,

Morris²,

Priye

et al. 2023

Preprint

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Nucleic acid isothermal amplification methods are advantageous for point-of-care (POC) diagnostics because of their precision, sensitivity, and low power requirements. Loop-mediated isothermal amplification (LAMP) is an established method, renowned for its nucleic acid amplification efficiency and robust amplification of semi purified target nucleic acids. However, LAMP may be prone to non-specific amplification causing false positives and therefore fails to replace PCR as the gold standard method in clinical testing. We show that LAMP combined with clustered regularly interspaced short palindromic repeats (CRISPR) technology is an effective alternative for overcoming the limitations of LAMP alone. Nucleic acids are first isothermally pre-amplified to enrich for targets, then specific amplification detection signals are generated by sequences of RNA guided recognition of amplicons. We are the first to demonstrate a paper based microfluidic system for detecting pathogen nucleic acids in samples by combining the power of LAMP and CRISPR technology. We show that although LAMP may produce non-specific amplification, the possibility of detecting a false positive can be eliminated by combining LAMP with CRISPR based detection. We demonstrate that a paper based microfluidic platform has the potential to compete with the conventional Nucleic acid testing (NAT) technologies not only in terms of robustness but also in terms of cost and complexity.

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“…Many µPADs and µTAS have been developed to detect pathogens from a wide range of matrices, including food sources, wastewater, drinking water, and human serum and blood amongst others, and have been used for a variety of applications such as food and drinking water quality control, environmental monitoring, and clinical analysis. [46][47][48][49][50][51][52] However, the detection mechanism typically employed by these devices is often based on DNA amplification or protein detection that are not suitable biomarkers for cell viability, and therefore cannot be used for live pathogen sensing.…”

Section: Microfluidic Platforms For Detection Of Live Microorganismsmentioning

confidence: 99%

Microfluidics for Rapid Detection of Live Pathogens

Rossi

Coulon

et al. 2023

Adv Funct Materials

Self Cite

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Rapid, sensitive, and selective detection of live pathogens remains a key priority for quality control and risk assessment. While conventional methods often require complicated workflows, costly reagents, lab equipment, and are time-consuming, rendering them inadequate for field testing and lowresource settings. Increased attention has been drawn to developing alternative low-cost and rapid methods to detect on-site live pathogens in different environmental matrices. Among them, microfluidic devices that integrate various laboratory functions in a miniaturized manner have proven to be a promising tool for the rapid and sensitive detection of pathogens. Herein, the development of microfluidic devices specifically designed for the detection of live pathogens is discussed along a concise summary of novel microfluidics systems recently developed, contrasted to conventional methods regarding assay time, the limit of detection, and target organisms. These include a variety of micro total analysis systems (µTAS) and micro fluidic paper-based analytical devices (µPADs) in combination with molecular methods and traditional live cell detection techniques, such as cell culture, DNA intercalating dyes, resazurin, and immobilized bioreceptors (e.g., aptamers and capture antibodies). Furthermore, insights on the future perspectives of microfluidics for live pathogen detection with a highlight on the rapid and low-cost method development for field testing are provided.

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Paper Device Combining CRISPR/Cas12a and Reverse-Transcription Loop-Mediated Isothermal Amplification for SARS-CoV-2 Detection in Wastewater

Cited by 53 publications

References 39 publications

SARS-CoV-2 surveillance in medical and industrial wastewater—a global perspective: a narrative review

SARS-CoV-2 surveillance in medical and industrial wastewater—a global perspective: a narrative review

A paper based microfluidic platform combining LAMP-CRISPR/Cas12a for fluorometric detection of nucleic acids

Microfluidics for Rapid Detection of Live Pathogens

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