Optimizing and Evaluating a Novel High Throughput Lamp Assay for Detection of Low Intensity Plasmodium Falciparum Infections

Abstract

The prevalence of malaria has been on the decline globally and optimism for the effective control of malaria has been growing. Many countries are making the shift from simply bringing down the number of new malaria cases to aiming for the elimination of malaria altogether. Supporting this bold ambition is the World Health Organization (WHO) which has guidelines for both how this goal may be attained as well as for how countries can obtain WHO certification of the elimination of malaria. Achieving the elimination milestone requires identification and treatment of all parasite carriers, both symptomatic and asymptomatic. There is therefore need to develop improved high throughput diagnostics, with capacity to detect low parasitaemia malaria infections. This would enable early and comprehensive identification and treatment of all asymptomatic cases especially in traditionally non-endemic areas that play a major role in malaria transmission. The aim of this project was to optimize and evaluate a Novel high–throughput loop mediated isothemal amplification (HT LAMP) assay for detection of low intensity P. falciparum infections. The assay was modified to enhance throughput and optimized by inclusion of DMSO to increase sensitivity and specificity. An experimental study design was used to evaluate 134 dried blood spot (DBS) samples from children in Busia county, a malaria endemic area in Western Kenya. Nested PCR was used as the gold standard method for detection of low parasitaemia infections that are undetectable by conventional light microscopy. The results indicated that HT-LAMP has a sensitivity of 93.5%, specificity of 100 %, a positive predictive value of 100% and a negative predictive value of 96.7%. HT-LAMP exhibited a strong agreement with nested PCR (k= 0.950). HT LAMP therefore showed superior diagnostic capacity just like that of Nested PCR and thus it can be used in field surveillance to complement malaria elimination interventions. Further optimizations should also be explored to make a multiplex HT-LAMP assay that can detect other Plasmodium species.