Development of a point-of-care test to detect SARS-CoV-2 from saliva which combines a simple RNA extraction method with colorimetric reverse transcription loop-mediated isothermal amplification detection

Yamazaki, Wataru; Matsumura, Yasufumi; Thongchankaew-Seo, Uraiwan; Yamazaki, Yasuko; Nagao, Miki

doi:10.1016/j.jcv.2021.104760

Cited by 49 publications

(36 citation statements)

References 15 publications

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“…One example is the COVID-19 RT-LAMP test developed by Yamazaki et al . which digests saliva with a semi-alkaline protease for 15 min followed by inactivation at 95 °C for 5 min 35 . The SalivaDirect COVID-19 test uses a similar processing method although it utilizes RT-PCR and is therefore not a point-of-care test 33 .…”

Section: Discussionmentioning

confidence: 99%

A rapid near-patient detection system for SARS-CoV-2 using saliva

Toppings

Mohon

Lee

et al. 2021

Sci Rep

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The highly infectious nature of SARS-CoV-2 necessitates the use of widespread testing to control the spread of the virus. Presently, the standard molecular testing method (reverse transcriptase-polymerase chain reaction, RT-PCR) is restricted to the laboratory, time-consuming, and costly. This increases the turnaround time for getting test results. This study sought to develop a rapid, near-patient saliva-based test for COVID-19 (Saliva-Dry LAMP) with similar accuracy to that of standard RT-PCR tests. A lyophilized dual-target reverse transcription-loop-mediated isothermal amplification (RT-LAMP) test with fluorometric detection by the naked eye was developed. The assay relies on dry reagents that are room temperature stable. A device containing a centrifuge, heat block, and blue LED light system was manufactured to reduce the cost of performing the assay. This test has a limit of detection of 1 copy/µL and achieved a positive percent agreement of 100% [95% CI 88.43% to 100.0%] and a negative percent agreement of 96.7% [95% CI 82.78–99.92%] relative to a reference standard test. Saliva-Dry LAMP can be completed in 105 min. Precision, cross-reactivity, and interfering substances analysis met international regulatory standards. The combination of ease of sample collection, dry reagents, visual detection, low capital equipment cost, and excellent analytical sensitivity make Saliva-Dry LAMP particularly useful for resource-limited settings.

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Section: Discussionmentioning

confidence: 99%

A rapid near-patient detection system for SARS-CoV-2 using saliva

Toppings

Mohon

Lee

et al. 2021

Sci Rep

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“…Recently, an investigation into a series of 44 people in Japan suspected of having COVID-19, using a buccal sample yielded a sensitivity of 82.6% over 23 definite COVID-19 patients. Results were obtained in 45 min [ 15 ]. Further investigations using the same commercially available isothermal amplification technology that we evaluated here (ID NOWTM, Abbott, Scarborough, ME, USA) showed 94% positive percent agreement (90/96 nasopharyngeal swabs) in one study [ 20 ] and 91% positive percent agreement (30/33 nasopharyngeal swabs) in another study [ 13 ].…”

Section: Discussionmentioning

confidence: 99%

“…Among this emerging technology, loop-mediated amplification (LAMP) and Nicking enzyme-assisted reaction (NEAR) have been used in point of care, far from central laboratories to identify both symptomatic and asymptomatic individuals [ 14 ]. Furthermore, isothermal amplification applied to saliva samples has also proved promising on a limited series of COVID-19 patients [ 15 ]. Moreover, it was recently demonstrated that saliva collection with a roll cotton improved significantly the molecular detection of SARS-CoV-2 compared to nasopharyngeal swab (NPS) specimens [ 16 ].…”

Section: Introductionmentioning

confidence: 99%

Rapid Isothermal Amplification for the Buccal Detection SARS-CoV-2 in the Context of Out-Patient COVID-19 Screening

Bouam

Vincent

Glass

et al. 2021

JCM

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A commercially available isothermal amplification of SARS-CoV-2 RNA was applied to self-collected saliva samples using dry dental cotton rolls, which were held in the mouth for two minutes. Of 212 tests, isothermal amplification yielded three (0.14%) invalid results, 120 (56.6%) positive results and 89 (42%) negative results. Compared to reference RT-PCR assays routinely performed simultaneously on nasopharyngeal swabs, excluding the three invalid isothermal amplification assays and one RT-PCR invalid assay, these figures indicated that 119/123 (96.7%) samples were positive in both methods and 85/85 samples were negative in both methods. Four positive buccal swabs which were missed by the isothermal amplification, exhibited Ct values of 26–34 in reference RT-PCR assays. Positive isothermal amplification detection was achieved in less than 10 min. Supervision of the self-sampling procedure was key to achieve these performances. These data support the proposal to use the protocol reported in this paper, including supervised buccal self-sampling, to screen people suspected of having COVID-19 at the point of care.

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“…Various methods are available to extract RNA from saliva, such as methods using phenol and guanidinium isothiocyanate, or commercially available silica membrane spin columns or magnetic bead-based RNA isolation kits (63). Other molecular diagnostic methods, such as reverse transcription-loop-mediated isothermal amplification (RT-LAMP), have also been reported as useful for diagnosing COVID-19 in settings of point-of care testing (64)(65)(66). Rapid and extraction-free detection of SARS-CoV-2 from saliva by colorimetric RT-LAMP is a simple, sensitive, rapid, and cost-effective approach with broad potential to expand diagnostic testing for the virus causing COVID-19 (67,68).…”

Section: Obstacles To Sars-cov-2 Rna Extraction From Saliva and New Applicationsmentioning

confidence: 99%

One-Year Update on Salivary Diagnostic of COVID-19

Caixeta

Oliveira

Cardoso-Sousa

et al. 2021

Front. Public Health

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Background: Coronavirus disease 2019 (COVID-19) is a global health problem, which is challenging healthcare worldwide. In this critical review, we discussed the advantages and limitations in the implementation of salivary diagnostic platforms of COVID-19. The diagnostic test of COVID-19 by invasive nasopharyngeal collection is uncomfortable for patients and requires specialized training of healthcare professionals in order to obtain an appropriate collection of samples. Additionally, these professionals are in close contact with infected patients or suspected cases of COVID-19, leading to an increased contamination risk for frontline healthcare workers. Although there is a colossal demand for novel diagnostic platforms with non-invasive and self-collection samples of COVID-19, the implementation of the salivary platforms has not been implemented for extensive scale testing. Up to date, several cross-section and clinical trial studies published in the last 12 months support the potential of detecting SARS-CoV-2 RNA in saliva as a biomarker for COVID-19, providing a self-collection, non-invasive, safe, and comfortable procedure. Therefore, the salivary diagnosis is suitable to protect healthcare professionals and other frontline workers and may encourage patients to get tested due to its advantages over the current invasive methods. The detection of SARS-CoV-2 in saliva was substantial also in patients with a negative nasopharyngeal swab, indicating the presence of false negative results. Furthermore, we expect that salivary diagnostic devices for COVID-19 will continue to be used with austerity without excluding traditional gold standard specimens to detect SARS-CoV-2.

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Development of a point-of-care test to detect SARS-CoV-2 from saliva which combines a simple RNA extraction method with colorimetric reverse transcription loop-mediated isothermal amplification detection

Cited by 49 publications

References 15 publications

A rapid near-patient detection system for SARS-CoV-2 using saliva

A rapid near-patient detection system for SARS-CoV-2 using saliva

Rapid Isothermal Amplification for the Buccal Detection SARS-CoV-2 in the Context of Out-Patient COVID-19 Screening

One-Year Update on Salivary Diagnostic of COVID-19

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