Genotyping of Plasmodium falciparum gametocytes by reverse transcriptase polymerase chain reaction

Abstract

A molecular assay has been developed for the specific detection and genetic characterisation of Plasmodium falciparum gametocytes in the blood of malaria infected individuals. The assay is based on the reverse transcription and polymerase chain reaction (RT-PCR) amplification of the messenger RNA of gene pfg377, a sexual-stage specific transcript abundantly produced in maturing gametocytes. The gene contains four regions of repetitive sequences, of which region 3 was shown to be the most polymorphic in laboratory clones and field isolates of the parasite. Analysis of samples of malaria infected blood by RT-PCR specific for region 3 has enabled identification of multiple gametocyte-producing clones within single infections. The assay is able to detect gametocytes below the threshold of microscopic detection, and is highly specific for its gametocyte targets also in the presence of a vast excess of asexual forms. Copyright (C) 2000 Elsevier Science B.V.