Abstract:
Background: Reverse-transcription loop-mediated isothermal amplification (RT-LAMP) with colorimetric readout is a rapid, robust, and cost-effective one-step amplification assay that we previously trialled for the identification of SARS-CoV-2 in nasopharyngeal swabs in four countries. Here, we expanded our assessment of RT-LAMP for SARS-CoV-2 detection to several other African countries and evaluated its operational performance with crude saliva as a pragmatic approach for outbreak surveillance and response in resource-limited settings.
Methods: We conducted a multicountry diagnostic accuracy study of RT-LAMP for the detection of SARS-CoV-2 in different types of clinical samples. A preliminary study was conducted in Slovenia and Italy to establish the analytical performance (limit of detection) of RT-LAMP and optimise this assay before its deployment in Africa. Subsequently, we tested RT-LAMP with RNA extracted from nasopharyngeal swabs in seven countries in Africa (Angola, Burkina Faso, Côte d'Ivoire, Ethiopia, Senegal, Sudan, and Zimbabwe), and, in parallel, with crude saliva samples (ie, without RNA extraction) in an additional four countries (Cameroon, Ethiopia, Kenya, and Nigeria; paired nasopharyngeal swabs were collected at the same time). In both contexts, quantitative RT-PCR (RT-qPCR) with RNA extracted from nasopharyngeal swabs was used as the gold-standard benchmarking assay to evaluate performance. For RT-qPCR testing, each laboratory followed their own standard diagnostic procedure, whereas a standardised protocol was used for RT-LAMP. Saliva test standardisation was ensured through centralised reagent distribution. We calculated diagnostic parameters (sensitivity, specificity, and accuracy) using a 2 × 2 contingency table.
Findings: The preliminary study reported 87% sensitivity and 98% specificity for RT-LAMP. Between Sept 1, 2021, and June 30, 2022, we collected 2774 nasopharyngeal swabs and 577 crude saliva samples. For RNA extracted from nasopharyngeal swabs, the sensitivity and specificity of RT-LAMP for detection of SARS-CoV-2 (relative to the standard of diagnostics-ie, the RT-qPCR assay used in each participating laboratory) were 89% (95% CI 87-90) and 95% (93-96), respectively. Similarly, RT-LAMP tested on saliva without RNA extraction showed 80% (75-84) sensitivity and 99% (96-100) specificity (relative to the results obtained with the standard of diagnostics for RNA extracted from paired nasopharyngeal samples).
Interpretation: Colorimetric RT-LAMP is a reliable assay for SARS-CoV-2 detection in both extracted RNA and crude saliva samples. The demonstrably acceptable performance on crude saliva samples (without RNA extraction) underscores the scalability of this method for efficient outbreak surveillance in resource-limited settings.