Fluorescent Plasmodium berghei sporozoites and pre-erythrocytic stages: a new tool to study mosquito and mammalian host interactions with malaria parasites

To track malaria parasites for biological studies within the mosquito and m ammalian hosts, we constructed a stably transformed clonal line of Plasmodi um berghei, PbFluspo, in which sporogonic and pre-erythrocytic liver-stage parasites are autonomously fluorescent. A cassette containing the structura l gene for the FACS-adapted green fluorescent protein mutant 2 (GFPmut2), e xpressed from the 5' and 3' flanking sequences of the circumsporozoite (CS) protein gene, was integrated and expressed at the endogenous CS locus. Rec ombinant parasites, which bear a wild-type copy of CS, generated highly flu orescent oocysts and sporozoites that invaded mosquito salivary glands and were transmitted normally to rodent hosts. The parasites infected cultured hepatocytes in vitro, where they developed into fluorescent pre-erythrocyti c forms. Mammalian cells infected by these parasites can be separated from non-infected cells by fluorescence activated cell sorter (FACS) analysis. T hese fluorescent insect and mammalian stages of P. berghei should be useful for phenotypic studies in their respective hosts, as well as for identific ation of new genes expressed in these parasite stages.