Auxotrophs of Plasmodium falciparum dependent on p-aminobenzoic acid for growth.

Abstract

The isolation of auxotrophic strains of a parasite offers new opportunities for studying parasitology. We have isolated cloned lines of Plasmodium faklparum that, unlike the parent line from which they were derived, rely on exogenous p-aminobenzoic acid (PABA) for growth. Isolation involved random mutagenesis of a cloned line of P. falciparum and subsequent selection of PABA-dependent parasites. Both parent and PABA-dependent clones were analyzed for PABA uptake and synthesis. Each clone takes up comparable amounts of PABA from the medium. The parent line, clone 3D7, can synthesize PABA de novo, whereas the PABA-dependent clones cannot. The requirement ofexogenous PABA for growth by the auxotrophic strains coupled with their inability to synthesize PABA indicates that normal parasite growth can be completely supported by either synthesis or salvage. This work further clarifies the relationship between the availability of PABA and success of the parasite, an issue of debate from classic studies showing reduced parasite load in individuals on milk-fed diets.

Cytotoxic Effect of Curcumin on Malaria Parasite Plasmodium falciparum : Inhibition of Histone Acetylation and Generation of Reactive Oxygen Species

“…A stock solution of curcumin (Sigma, St. Louis, MO) was made in dimethyl sulfoxide (DMSO) at 100 mM. The effect of curcumin on parasite in vitro growth was tested using a standard [ 3 H]hypoxanthine labeling method (11,30) with serial dilutions of the drug to final concentrations of 5 to 60 M. Briefly, ring-stage parasite culture was diluted to 1% parasitemia and 5% hematocrit in a complete low-hypoxanthine medium (5 mg/liter hypoxanthine) and seeded in triplicate in 96-well flat-bottom plates. Curcumin or DMSO was added to 200 l of culture and incubated at 37°C for 24 h. Subsequently, 100 l supernatant was replaced with an equal volume of complete low-hypoxanthine medium containing 1 Ci of [ 3 H]hypoxanthine (Amersham, Piscataway, NJ).…”

The Fight Against Drug-Resistant Malaria: Novel Plasmodial Targets and Antimalarial Drugs

“…The Shikimate pathway for aromatic biosynthesis produces a key intermediate, para-aminobenzoate (pABA) which is used for the folate biosynthesis in P. falciparum [81,82]. The Shikimate pathway is present in plants, algae, bacteria, and fungi for chorismate production and has recently been identified in several apicomplexan parasites [83].…”

Metabolic dependency of chorismate in Plasmodium falciparum

“…Our first step was to systematically assess the development of both asexual and sexual stages of P. falciparum in a defined medium in the absence of an exogenous supply of chorismate end-products. It has been previously reported that asexual stages of P. falciparum can grow in pABA-and folate-deficient medium [22][23][24][25] , which forces parasites to rely on their own biosynthesis using the shikimate pathway. Therefore, in order to assess the metabolic dependency of chorismate for ubiquinone biosynthesis, we sought to standardize and characterize P. falciparum asexual growth and sexual development in a defined media, here referred to as RPMI Minimal Medium (MM), lacking aromatic metabolites (pABA, folic acid (FA), tryptophan, phenylalanine and tyrosine) that are found in the standard RPMI medium (CM) ( Table S1).…”