EVALUATION OF SYMPTOMATIC AND ASYMPTOMATIC P. FALCIPARUM WITH SPECIES-SPECIFIC LAMP ASSAY AND TRANSLATION TO THE POINT- OF-CARE

• Published in: International journal of infectious diseases Vol. 130; p. S133
• Main Authors: Malpartida-Cardenas, K., Moser, N., Ansah, F., Prah, D. Ahu, Awandare, G., Cunnington, A., Baum, J., Rodriguez-Manzano, J., Georgiou, P.
• Format: Journal Article
• Language: English
• Published: Elsevier Ltd 01.05.2023; Elsevier
• ISSN: 1201-9712; 1878-3511
• DOI: 10.1016/j.ijid.2023.04.328

Human malaria is a life-threatening parasitic disease with high impact in the Sub-Saharan African region, where 95% of global cases occurred in 2020. In particular, asymptomatic malaria provides a reservoir for transmission which is currently missed by rapid diagnostics. Loop-mediated isothermal amplification (LAMP) assay was designed for the specific detection of P. falciparum (LAMP-PfK13) using Primer Explorer 5.0 and manual optimisation based on sequences retrieved from PlasmoDB. A total of 70 samples were obtained from subjects who presented symptomatic (n=8) and asymptomatic (n=47) P. falciparum malaria at Obom (Ghana), including uninfected subjects (n=15). Samples were tested with LAMP, TaqMan qPCR, microscopy, ELISA and an in-house Lab-on-Chip platform. The LAMP-PfK13 assay had a turnaround time below 20 min, no cross- reactivity, and analytical sensitivity of 10 copies per reaction. Samples were previously quantified using the Plasmodium pan-genus and P. falciparum TaqMan qPCR. Symptomatic cases were detected with the LAMP-PfK13 assay, with a sensitivity of 100% (95% CI 63.1 - 100%) and specificity of 100% (95% CI 78.2 - 100%). Detection of asymptomatic malaria cases was achieved in 100% of samples from asymptomatic subjects with parasite loads above 80 gDNA copies per μL. Within the asymptomatic group, the LAMP assay had a sensitivity of 80.77% (95% CI 60.7 - 93.5%) and a specificity of 100% (95% CI 78.2 - 100%) when testing samples with submicroscopic parasitaemia (n=26); this demonstrates the superior sensitivity over microscopy. Extracted DNA from 18 samples was tested in Ghana using our Lab-on-Chip platform (Lacewing) obtaining comparable results (p-value > 0.05) against a conventional fluorescence-based instrument. Detection of low-level parasitaemia in symptomatic and asymptomatic carriers is required for epidemiological research and the identification of hot spots. The developed LAMP assay in combination with Lacewing could be used for digital diagnostics at the point-of-care, enabling rapid testing and surveillance.