A Rapid, Highly Sensitive and Open-Access SARS-CoV-2 Detection Assay for Laboratory and Home Testing

Kellner, Max J.; Ross, James J.; Schnabl, Jakob; Dekens, Marcus P. S.; Matl, Martin; Heinen, Robert; Grishkovskaya, Irina; Bauer, Benedikt; Stadlmann, Johannes; Menéndez‐Arias, Luis; Straw, Andrew; Fritsche-Polanz, Robert; Traugott, Marianna; Seitz, Tamara; Zoufaly, Alexander; Födinger, Manuela; Wenisch, Christoph; Zuber, Johannes; Pauli, Andrea; Brennecke, Julius

doi:10.3389/fmolb.2022.801309

Cited by 37 publications

(38 citation statements)

References 52 publications

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“…The high acceptability of self-collected gargle as reported by some studies suggested that it may also potentially help to abate the testing hesitancy that is not uncommonly encountered in mandatory testing scenarios [ 35 , 47 , 77 ]. Our findings provide an evidence-based justification for COVID-19 testing approaches in national and international guidelines, and support the recommendation of gargling as an inexpensive and easy sampling approach for routine repeated testing in various community settings [ 78 ], including primary and secondary schools, universities [ 77 , 79 , 80 ], nursing homes [ 81 ], daycare facilities for children aged 1–7 years [ 67 ], children's hospitals [ 82 ], homes [ 83 ] and in other specific high-risk populations having medically indicated needs for repeated testing [ 78 ].…”

Section: Discussionmentioning

confidence: 66%

Performance of saline and water gargling for SARS-CoV-2 reverse transcriptase PCR testing: a systematic review and meta-analysis

Tsang

Cowling

et al. 2022

Eur Respir Rev

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The performance of gargling for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) reverse transcriptase (RT)-PCR testing has not been previously reviewed. This review systematically assessed the performance of saline and water gargling for SARS-CoV-2 RT-PCR testing in the settings of diagnosing and monitoring viral shedding.We included original studies comparing the performance of gargling and (oropharyngeal–)nasopharyngeal swabs for SARS-CoV-2 RT-PCR testing. Studies conducted in either suspected individuals or confirmed cases were included and analysed separately. The sensitivity, specificity, positive predictive value (PPV) and negative predictive value (NPV) were examined using random-effects models.Gargles achieved a high overall sensitivity (91%), specificity (97%), PPV (95%) and NPV (91%) for SARS-CoV-2 RT-PCR testing. Studies using saline gargle and water gargle have an overall sensitivity of 97% and 86%, respectively. The sensitivity values were largely maintained for saline and water gargling on stratified analysis, for both diagnosis (96% and 92%) and viral shedding monitoring (98% and 78%). A higher sensitivity was also reported by studies using sterile saline (100%), a smaller amount of gargling solution (92% versus 87%) and a longer gargling duration (95% versus 86%).Our results supported the use of gargling as a sampling approach for SARS-CoV-2 RT-PCR testing, which achieved a high sensitivity for both diagnosis and viral shedding monitoring purposes. Further investigation on the comparative performance of different gargling mediums is needed to draw a definitive conclusion.

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Section: Discussionmentioning

confidence: 66%

Performance of saline and water gargling for SARS-CoV-2 reverse transcriptase PCR testing: a systematic review and meta-analysis

Tsang

Cowling

et al. 2022

Eur Respir Rev

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“…For the implementation of the RT-LAMP assay we used NEB enzymes and a colorimetric-based readout, however, royalty-free self-purified Bst-LF polymerase and the HIV-RT have been also reported to work very well for RT-LAMP for SARS-CoV-2 (23). Using self-produced Bst-LF polymerase and HIV-RT preparations we found that slight modifications of the sample analysis procedures (using a fluorescent readout, e.g.…”

Section: Discussionmentioning

confidence: 99%

“…In RT-LAMP, this can be replaced by using fluorescence dyes to detect the produced DNA, or dyes to detect a chemical change associated with a successful amplification, e.g. a pH drop (22) or the formation of a magnesium pyrophosphate precipitate (23). In addition, since fluorescence and color readout can constitute bottlenecks, we and others developed DNA barcoding strategies to permit pooled analysis of many samples using next-generation sequencing (termed LAMP-sequencing) (24)(25)(26), providing the possibility of a massively scaled up sample throughput.…”

Section: Introductionmentioning

confidence: 99%

Scalable RT-LAMP-based SARS-CoV-2 testing for infection surveillance with applications in pandemic control

Lou

Meurer

Ovchinnikova

et al. 2022

Preprint

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Throughout the current SARS-CoV-2 pandemic, limited diagnostic testing capacity prevented sentinel testing of the population, demonstrating the need for novel testing strategies and infrastructures. Here, we describe the set-up of an alternative testing platform, which allows scalable surveillance testing as an acute pandemic response tool and for pandemic preparedness purposes, exemplified by SARS-CoV-2 diagnostics in an academic environment. The testing strategy involves self-sampling based on gargling saline, pseudonymized sample handling, automated 96-well plate-based RNA extraction, and viral RNA detection using a semi-quantitative multiplexed colorimetric reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay with an analytical sensitivity comparable to RT-quantitative polymerase chain reaction (RT-qPCR). We provide standard operating procedures and an integrated software solution for all workflows, including sample logistics, LAMP assay analysis by colorimetry or by sequencing (LAMP-seq), and communication of results to participants and the health authorities. Using large sample sets including longitudinal sample series we evaluated factors affecting the viral load and the stability of gargling samples as well as the diagnostic sensitivity of the RT-LAMP assay. We performed >35,000 tests during the pandemic, with an average turnover time of fewer than 6 hours from sample arrival at the test station to result announcement. Altogether, our work provides a blueprint for fast, sensitive, scalable, cost- and labor-efficient RT-LAMP diagnostics. As RT-LAMP-based testing requires advanced, but non-specialized laboratory equipment, it is independent of potentially limiting clinical diagnostics supply chains.

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“…Relatively easy to implement; however, this strategy requires a heating source and optimization of the heating conditions. [10,13,23,[29][30][31][32][33][34]37,40,41] Nasopharyngeal swabs RT-PCR Thermal shock of the sample at 95 • C for 5 min followed by 4 • C for 10 min.…”

Section: Sample Type Naat/detection Methods Sample Preparation Strategymentioning

confidence: 99%

Strategies That Facilitate Extraction-Free SARS-CoV-2 Nucleic Acid Amplification Tests

Delgado-Diaz¹,

Sakthivel²,

Nguyen³

et al. 2022

Viruses

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The COVID-19 pandemic has resulted in an unprecedented global demand for in vitro diagnostic reagents. Supply shortages and hoarding have impacted testing capacity which has led to inefficient COVID-19 case identification and transmission control, predominantly in developing countries. Traditionally, RNA extraction is a prerequisite for conducting SARS-CoV-2 nucleic acid amplification tests (NAAT); however, simplified methods of sample processing have been successful at bypassing typical nucleic acid extraction steps, enabling extraction-free SARS-CoV-2 NAAT workflows. These methods involve chemical and physical approaches that are inexpensive and easily accessible alternatives to overcome extraction kit supply shortages, while offering acceptable test performance. Here we provide an overview of three main sample preparation strategies that have been shown to facilitate extraction-free SARS-CoV-2 NAATs.

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A Rapid, Highly Sensitive and Open-Access SARS-CoV-2 Detection Assay for Laboratory and Home Testing

Cited by 37 publications

References 52 publications

Performance of saline and water gargling for SARS-CoV-2 reverse transcriptase PCR testing: a systematic review and meta-analysis

Performance of saline and water gargling for SARS-CoV-2 reverse transcriptase PCR testing: a systematic review and meta-analysis

Scalable RT-LAMP-based SARS-CoV-2 testing for infection surveillance with applications in pandemic control

Strategies That Facilitate Extraction-Free SARS-CoV-2 Nucleic Acid Amplification Tests

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