Nationwide Surveillance of Pfhrp2 Exon 2 Diversity in Plasmodium falciparum Circulating in Symptomatic Malaria Patients Living in Ghana

Abstract

Reports of increasing false-negative HRP2-based rapid diagnostic test results across Africa require constant monitoring of factors associated with these false-negative outcomes, as failure of this diagnostic tool will have severe consequences on malaria treatment and control programs. This study characterized the extent of genetic diversity in the Plasmodium falciparum histidine-rich protein 2 (Pfhrp2) gene in P. falciparum isolates from symptomatic malaria patients across the regions of Ghana. Exon 2 of Pfhrp2 was amplified from gDNA using polymerase chain reaction. All Pfhrp2-negative samples were subjected to Pf18S rRNA and Pfmsp2 gene amplifications. The amplified Pfhrp2 exon 2 fragments from clonal samples were sent for commercial Sanger sequencing. The type and number of PfHRP2 repeats, classified based on repeat types previously reported, were estimated from the sequence data and compared among geographical regions. About 81% (2,333/ 2,890) of the original microscopy positive dried blood spot (DBS) samples were available and used in this study. The Pfhrp2 exon 2 amplification was successful in 98.5% (2,297/2,333) of the tested samples, with band size ranging from 400 bp to 1,050 bp. A total of 13 out of the 24 previously reported repeat types were identified among the samples, with three samples lacking both type 2 and type 7 repeat motifs. This study suggested that the genetic diversity of Pfhrp2 exon 2 identified in P. falciparum circulating in symptomatic malaria patients in Ghana is unlikely to influence the sensitivity and specificity of HRP2 RDT-based diagnosis.