Background: Diversity in Plasmodium falciparum poses a major threat to malaria control and elimination interventions. This study utilized 12 polymorphic microsatellite (MS) markers and the Msp2 marker to examine diversity, multiplicity of infection (MOI) as well as the population structure of parasites circulating in two sites separated by about 92 km and with varying malaria transmission intensities within the Greater Accra Region of Ghana. Methods: The diversity and MOI of P. falciparum parasites in 160 non-symptomatic volunteers living in Obom (high malaria transmission intensity) and Asutsuare (low malaria transmission intensity) aged between 8 and 60 years was determined using Msp2 genotyping and microsatellite analysis. Results: The prevalence of asymptomatic P. falciparum carriers as well as the parasite density of infections was significantly higher in Obom than in Asutsuare. Samples from Asutsuare and Obom were 100% and 65% clonal, respectively, by Msp2 genotyping but decreased to 50% and 5%, respectively, when determined by MS analysis. The genetic composition of parasites from Obom and Asutsuare were highly distinct, with parasites from Obom being more diverse than those from Asutsuare. Conclusion: Plasmodium falciparum parasites circulating in Obom are genetically more diverse and distinct from those circulating in Asutsuare. The MOI in samples from both Obom and Asutsuare increased when assessed by MS analysis relative to MSP2 genotyping. The TA40 and TA87 loci are useful markers for estimating MOI in high and low parasite prevalence settings.

Background Asymptomatic malaria infections can serve as potential reservoirs for malaria transmission. The density of parasites contained in these infections range from microscopic to submicroscopic densities, making the accurate detection of asymptomatic parasite carriage highly dependent on the sensitivity of the tools used for the diagnosis. This study sought to evaluate the sensitivities of a variety of molecular and serological diagnostic tools at determining the prevalence of asymptomatic Plasmodium falciparum parasite infections in two communities with varying malaria parasite prevalence. Methods Whole blood was collected from 194 afebrile participants aged between 6 and 70 years old living in a high (Obom) and a low (Asutsuare) malaria transmission setting of Ghana. Thick and thin blood smears, HRP2 based malaria rapid diagnostic test (RDT) and filter paper dried blood spots (DBS) were prepared from each blood sample. Genomic DNA was extracted from the remaining blood and used in Plasmodium specific photo-induced electron transfer polymerase chain reaction (PET-PCR) and Nested PCR, whilst the HRP2 antigen content of the DBS was estimated using a bead immunoassay. A comparison of malaria parasite prevalence as determined by each method was performed. Results Parasite prevalence in the high transmission site of Obom was estimated at 71.4%, 61.9%, 60%, 37.8% and 19.1% by Nested PCR, the HRP2 bead assay, PET-PCR, HRP2-RDT and microscopy respectively. Parasite prevalence in the low transmission site of Asutsuare was estimated at 50.1%, 11.2%, 5.6%, 0% and 2.2% by Nested PCR, the HRP2 bead assay, PET-PCR, RDT and microscopy, respectively. The diagnostic performance of Nested PCR, PET-PCR and the HRP2 bead assay was similar in Obom but in Asutsuare, Nested PCR had a significantly higher sensitivity than PET-PCR and the HRP2 bead assay, which had similar sensitivity. Conclusions Nested PCR exhibited the highest sensitivity by identifying the highest prevalence of asymptomatic P. falciparum in both the high and low parasite prevalence settings. However, parasite prevalence estimated by the HRP2 bead assay and PET-PCR had the highest level of inter-rater agreement relative to all the other tools tested and have the advantage of requiring fewer processing steps relative to Nested PCR and producing quantitative results.