Quantification of female and male Plasmodium falciparum gametocytes by reverse transcriptase quantitative PCR

•We developed RT-qPCRs to count female and male gametocytes of Plasmodium falciparum.•The Pfs25 RT-qPCR quantifies densities as low as 0.3 mature females/μl blood.•The Pfs230p RT-qPCR has a lower detection limit of 1.8 mature males/μl blood.

The transmission of malaria parasites depends on the presence of sexual stages (gametocytes) in the blood, making the ratio and densities of female and male gametocytes important determinants of parasite fitness. This manuscript describes the development of reverse transcriptase quantitative PCR (RT-qPCR) assays to separately quantify mature female and male gametocytes of the human malaria parasite Plasmodium falciparum, and reveals that Pfs25 mRNA is expressed only in female gametocytes. The female (Pfs25) and male (Pfs230p) gametocyte specific RT-qPCR assays have lower detection limits of 0.3 female and 1.8 male gametocytes per microlitre of blood, respectively, making them more sensitive than microscopy. Accurate quantification of the ratio and densities of female and male gametocytes will increase understanding of P. falciparum transmission and improve the evaluation of transmission blocking interventions.

Graphical abstractSex-specific RT-qPCRs quantify densities as low as 0.04 female and 0.3 male gametocytes/PCR reaction (corresponding to 0.3 female and 1.8 male gametocytes/(l blood).Download high-res image (159KB)Download full-size image