Mutant/mutation
The mutant lacks expression of Orotate phosphoribosyltransferase (OPRTase) and expresses GFP under control of a male and RFP under control of a female gametocyte specific promoter..

Protein (function)
Orotate phosphoribosyltransferase (OPRTase) is the fifth enzyme in the de novo pyrimidine-synthesis pathway in the parasite, which lacks salvage pathways.
OPRTase catalyses the conversion of orotate and 5-phospho-d-ribosyl 1-pyrophosphate (PRPP) into orotidine 5′-monophosphate (OMP).

Phenotype
Phenotype analyses indicate that OPRTase is not essential for blood stage development, both for asexual stages as for gametocytes.

(Strongly) reduced gametocyte-, ookinete- and oocyst production. No formation of sporozoites.

Additional information
The oprt-  mutant parasites grew slowly (asexual cycle prolonged by approximately 4–5 hours compared to wt), were rapidly outgrown in a competition growth assay with wt parasites and  seem to invade very young reticulocytes.
Furthermore, the mutant was found to generate, on average, significantly fewer merozoites than wt parasites (17.5 ± 1.8) per schizont (counted after completion of asexual cycle)  and the asexual parasites also took longer to mature to schizonts. The mutant showed altered lethality in the C57/B6 mouse model as the mice infected with the mutants did not manifest the symptoms of experimental cerebral malaria (ECM) but died between days 14–20 as a result of severe anaemia and hyperparasitemia. Gametocytemia was significantly reduced but no change was seen inmale- female ratio. Exflagellation (the production of mature male gametes) was found to be severely affected. Consistent with the defects in male gametogenesis, very few ookinetes were formed in in vitro cultures.  Infectivity to the mosquito was significantly reduced  as seen by observing oocysts in infected mosquito midguts and salivary gland sporozoites and infection to naïve mice was found to be completely blocked.


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