Characterization of Plasmodium infections and genetic diversity of Plasmodium malariae msp-1 among asymptomatic individuals in southern Benin

Abstract Plasmodium malariae is less widespread than P. falciparum but can cause prolonged and persistent infections. This study aimed to characterize asymptomatic Plasmodium infections and assess the genetic diversity of the merozoite surface protein 1 ( msp-1 ) gene of P. malariae in southern Benin. Blood samples from 484 asymptomatic participants were analysed using rapid diagnostic tests (RDTs), microscopy, and nested PCR. Samples negative by nested PCR were subsequently analysed by nested qPCR for additional detection of Plasmodium species. P. malariae -positive samples were subjected to msp-1 genotyping and Sanger sequencing. Microscopy and RDTs detected Plasmodium infections in 17% and 20% of participants, respectively, whereas nested PCR detected infections in 38% (182/484; P < 0.0001). Among the 302 nested-PCR-negative samples, nested qPCR identified an additional 36 infections (12%). Nested PCR detected P. malariae in 6.6% (32/484) of participants, with nested qPCR identifying an additional 25 P. malariae infections (8.3%). The nucleotide diversity (π) of the successfully sequenced P. malariae msp-1 regions was 0.00602. These findings demonstrate a substantial burden of asymptomatic and sub-microscopic Plasmodium infections in southern Benin, including P. malariae infections missed by conventional diagnostic methods. The sequenced msp-1 regions showed relatively limited nucleotide diversity, providing baseline information on the genetic variation of P. malariae in southern Benin.