Sulfadoxine-Pyrimethamine Resistance in the Rodent Malaria Parasite Plasmodium chabaudi

Abstract

We have studied resistance to sulfadoxine-pyrimethamine (S/P) in the rodent malaria parasite Plasmodium chabaudi. A stable S/P-resistant mutant, AS(50S/P), was selected by drug treatment of a clone, AS(PYR), already resistant to pyrimethamine. The sequences of the P. chabaudi dhfr and dhps genes were obtained and found to be identical in AS(50S/P) and AS(PYR), showing that resistance to S/P in AS(50S/P) was not due to additional mutations in either gene. AS(50S/P) was crossed with a drug-sensitive clone, AJ, and 16 independent recombinant progeny were obtained. These clones were phenotyped for their susceptibility to S/P and to sulfadoxine and pyrimethamine separately. Pyrimethamine resistance was invariably associated with S/P resistance, but no correlation was found between resistance to S/P and resistance to sulfadoxine. Quantitative trait locus analysis of the progeny with 31 chromosome-specific markers showed that mutant P. chabaudi dhfr, or one or more genes closely linked to it, was a major determinant of S/P resistance. In addition, the inheritance of genes on chromosomes 5 and 13 from the sensitive parent appeared to contribute to the level of resistance observed. These results demonstrate that the S/P resistance of the AS(50S/P) mutant of P. chabaudi does not involve mutation in dhps and is not due simply to a combination of two genes determining resistance to pyrimethamine and sulfadoxine separately.

Whole genome re-sequencing identifies a mutation in an ABC transporter (mdr2) in a Plasmodium chabaudi clone with altered susceptibility to antifolate drugs

“…These markers (numbers 19 and 37; Hayton et al, 2002) map to g6pdh (PCHAS_132080) and fructose-bisphosphate aldolase (PCHAS_131180), respectively (Carlton et al, 1998). The K392Q MDR2 mutation maps between these two markers (Supplementary Fig.…”

“…Multi-drug resistant protein 2 ( mdr2 ), fructose-bisphosphate aldolase ( aldoI ) and glucose-6-phosphate dehydrogenase ( g6pdh ) are indicated. Numerical values next to gene acronyms indicate the level of linkage (mean difference in log parasitaemia) of the parental allele with sulphadoxine (SDX) response, as calculated by Hayton et al (2002). The asterisk indicates that linkage is statistically significant ( P  < 0.05).Supplementary Table S1All mutations identified uniquely in the Plasmodium chabaudi clone AS-50S/P following whole genome re-sequencing analysis.…”

“…As also reported in P. falciparum (Eastham and Rieckmann, 1983), the acquisition of PYR resistance in P. chabaudi is accompanied by an increase in susceptibility to SDX (Jacobs, 1964; Hayton et al, 2002). Subsequently, a single-step exposure of the AS-PYR clone to a high dose of S/P selected a mutant clone (AS-50S/P) which is more resistant to both S/P and SDX (Hayton et al, 2002) than the AS-PYR progenitor (Fig. 1).…”

A high-coverage artificial chromosome library for the genome-wide screening of drug-resistance genes in malaria parasites

“…This P. chabaudi strain evolved in the laboratory from the drug-sensitive P. chabaudi AS-sens strain and is resistant to both pyrimethamine and sulphadoxine (Hayton et al 2002;Culleton et al 2005). The genomic DNA of P. chabaudi AS(50S/P) was partially digested with Sau3AI, and then DNA fragments ranging from 10-50 kb were purified.…”

“…The parasites containing the integrated drug-resistant cassette were cloned by a limiting dilution method and used in the study. The pyrimethamine-resistant P. chabaudi AS(50S/P) line was previously generated (Hayton et al 2002) and obtained from MR4 (http://www.mr4.org/).…”