Comparison of Warmstart colorimetric LAMP using heat-treated samples with two Plasmodium-detection methods in Ekiti state, southwestern Nigeria

Abstract

Background: Malaria remains a deadly disease responsible for numerous deaths worldwide, especially in tropi-cal developing countries. Widely used methods for diagnosis include microscopy, which is laborious, lengthy, and error-prone, and rapid diagnostic tests (RDT), which have recently faced limitations on accuracy. Molecular methods like the polymerase chain reaction (PCR) and loop-mediated isothermal amplification (LAMP) have emerged as accurate, sensitive, and rapid ways of detecting the disease. While PCR requires expensive equip-ment, LAMP is less expensive to perform. However, contemporary malaria LAMP still requires DNA extraction kits from patient samples. DNA extraction by heating can be used to lower the cost, time, and level of training required from end-users to perform LAMP in low-resource settings. Methods: Blood samples were screened for Plasmodium parasites using microscopy, followed by a commonly used RDT kit. Heat treatment was used for quick DNA extraction from the blood samples, and LAMP primer sets were then used together with Warmstart colorimetric LAMP reagents for isothermal DNA amplification. Agarose gel electrophoresis was used to confirm that the colorimetric observations in LAMP reactions were due to successful DNA amplification. The data obtained were analysed in GraphPad Prism and subjected to chi-square tests with 95% confidence level. Results: Warmstart colorimetric LAMP using heat-treated blood samples achieved successful DNA amplifica-tion and detected Plasmodium parasites in 43.3% of samples, compared to 40% and 30% Plasmodium parasite detection yielded by antigen RDT kit and microscopy respectively. However, the difference between the percent-ages obtained using the different methods was not statistically significant. LAMP approach further enabled the identification/discrimination of different Plasmodium species. Conclusion: The Warmstart LAMP using heat-treated samples can be used as a rapid, cheaper, and easier-to-perform molecular method for the detection and species-level differentiation of malaria parasites.