Emergence of a<i>dhfr</i>Mutation Conferring High‐Level Drug Resistance in<i>Plasmodium falciparum</i>Populations from Southwest Uganda

Antifolate drugs have an important role in the treatment of malaria. Polymorphisms in the genes encoding the dihydrofolate reductase and dihydropteroate synthetase enzymes cause resistance to the antifol and sulfa drugs, respectively. Rwanda has the highest levels of antimalarial drug resistance in Africa. We correlated the efficacy of chlorproguanil-dapsone plus artesunate (CPG-DDS؉A) and amodiaquine plus sulfadoxinepyrimethamine (AQ؉SP) in children with uncomplicated malaria caused by Plasmodium falciparum parasites with pfdhfr and pfdhps mutations, which are known to confer reduced drug susceptibility, in two areas of Rwanda. In the eastern province, where the cure rates were low, over 75% of isolates had three or more pfdhfr mutations and two or three pfdhps mutations and 11% had the pfdhfr 164-Leu polymorphism. In the western province, where the cure rates were significantly higher (P < 0.001), the prevalence of multiple resistance mutations was lower and the pfdhfr I164L polymorphism was not found. The risk of treatment failure following the administration of AQ؉SP more than doubled for each additional pfdhfr resistance mutation (odds ratio [OR] ‫؍‬ 2.4; 95% confidence interval [CI] ‫؍‬ 1.01 to 5.55; P ‫؍‬ 0.048) and each pfdhps mutation (OR ‫؍‬ 2.1; 95% CI ‫؍‬ 1.21 to 3.54; P ‫؍‬ 0.008). The risk of failure following CPG-DDS؉A treatment was 2.2 times higher (95% CI ‫؍‬ 1.34 to 3.7) for each additional pfdhfr mutation, whereas there was no association with mutations in the pfdhps gene (P ‫؍‬ 0.13). The pfdhfr 164-Leu polymorphism is prevalent in eastern Rwanda. Antimalarial treatments with currently available antifol-sulfa combinations are no longer effective in Rwanda because of high-level resistance.

Section: Discussionsupporting

confidence: 77%

mentioning

confidence: 99%

Molecular Correlates of High-Level Antifolate Resistance in Rwandan Children with Plasmodium falciparum Malaria

Karema

Imwong

Fanello

et al. 2010

“…This single pfdhfr mutation was previously detected in the south of Madagascar in 2006 (17). Although it is rare in Africa (12,16), it is obvious that this single pfdhfr 164L allele, which has not been described before, represents a serious threat that may lead to the emergence of a mutant with quadruple mutations, which will render SP ineffective. The mutant with quadruple mutations could arise locally by recombination between mutant parasites with triple mutations and parasites with the single 164L mutation (especially in areas with high rates of transmission, such as the southeast, where the prevalence of the both genotypes is high), or by stepwise Of the pfdhps point mutations related to sulfadoxine resistance, only A437G was prevalent in both countries, confirming previous findings of studies performed in the Comoros Islands (23).…”

Section: Discussionmentioning

confidence: 84%

Plasmodium falciparumDrug Resistance in Madagascar: Facing the Spread of Unusualpfdhfrandpfmdr-1Haplotypes and the Decrease of Dihydroartemisinin Susceptibility

Andriantsoanirina

Ratsimbasoa

Bouchier

et al. 2009

The aim of this study was to provide the first comprehensive spatiotemporal picture of Plasmodium falciparum resistance in various geographic areas in Madagascar. Additional data about the antimalarial resistance in the neighboring islands of the Comoros archipelago were also collected. We assessed the prevalence of pfcrt, pfmdr-1, pfdhfr, and pfdhps mutations and the pfmdr-1 gene copy number in 1,596 P. falciparum isolates collected in 26 health centers ( In recent decades, the emergence and subsequent spread of Plasmodium falciparum chloroquine (CQ)-and sulfadoxinepyrimethamine (SP)-resistant parasites across areas where malaria is endemic have been a challenge to malaria control programs (41, 44). Substantial advances toward gaining an understanding of the genetic basis of antimalarial drug resistance have been made (14). Molecular evolutionary studies have concluded that the CQ-resistant P. falciparum chloroquine resistance transporter (pfcrt) and high-level pyrimethamine-resistant dihydrofolate (pfdhfr) alleles have emerged in a limited number of independent foci, from which they have rapidly spread in the local vicinity and have then invaded areas continent-wide and transferred between continents (1, 36). These lessons of the past have, first, stimulated changes in antimalarial treatment policies by introducing combinations of drugs that act on different targets and, second, resulted in the implementation of effective monitoring systems to detect as early as possible the emergence of resistant parasites on the basis of the assessment of the therapeutic efficacies of antimalarials (25, 46), determination of the decreased sensitivity of the parasites to drugs in vitro (4), and the detection of an increasing prevalence of molecular markers related to drug resistance (24).According to data published from 2002 to 2006, the epidemiological features of P. falciparum CQ and SP resistance differ considerably between Madagascar and the Comoros Islands, two countries located close to each other in the southwestern Indian Ocean (43). In vitro CQ resistance was moderate in Madagascar (29,33,45), although the level of therapeutic efficacy was declining. During that time, the rate of CQ resistance was high in the Comoros Islands (22,23,30). Likewise, pyrimethamine resistance was absent in Madagascar (28, 32) but was present at high levels in the Comoros Islands (23). The most recent in vivo data obtained on the basis of the WHO 28-day follow-up protocol, conducted in 2006 and 2007 at multiple sites, have confirmed that resistance to all antimalarials except CQ in Madagascar remains rare. Indeed, the prevalence of the clinical failure of treatment with amodiaquine, SP and the combination artesunate and amodiaquine was Ͻ5%, while the rate of failure of treatment with CQ was 44% (19). However, the recent demonstration of the introduction of multidrug-resistant P. falciparum parasites into Madagascar from the Comoros Islands (18) and the emergence of the uncommon dihydrofolate reductase I164L genotype in P. falciparum parasi...

“…One of the limitations of our study is that we were not able to genotype the Pfdhfr I164L mutation, which has been associated with high SP resistance (26,41). Similarly, we did not get a full haplotype on all of the SNPs due to poor-quality DNA in a few samples.…”

Section: Discussionmentioning

confidence: 84%

Prevalence of Plasmodium falciparum Resistance Markers to Sulfadoxine-Pyrimethamine among Pregnant Women Receiving Intermittent Preventive Treatment for Malaria in Uganda

Mbonye

Birungi

Yanow

et al. 2015

The aim of this study was to assess the prevalence of mutations in Plasmodium falciparum dihydrofolate reductase (Pfdhfr) and dihydropteroate synthase (Pfdhps) genes among pregnant women using sulfadoxine-pyrimethamine (SP) as an intermittent preventive treatment (IPTp). A molecular epidemiological study of P. falciparum parasite resistance markers to SP was conducted from August 2010 to February 2012 in Mukono district in central Uganda. DNA was extracted from 413 P. falciparum-positive samples. Real-time PCR, followed by melting curve analysis, was used to characterize point mutations in the Pfdhfr and Pfdhps genes that are associated with SP resistance. The prevalence of the single-nucleotide mutations in Pfdhfr at codons 51I, 59R, and 108N and in Pfdhps at codons 437G and 540E was high (>98%), reaching 100% fixation after one dose of SP, while the prevalence of 581G was 3.3% at baseline, reaching 12.5% after one dose of SP. At baseline, the prevalence of Pfdhfr and Pfdhps quintuple mutations was 89%, whereas the sextuple mutations (including 581G) were not prevalent (3.9%), reaching 16.7% after one dose of SP. However, the numbers of infections at follow-up visits were small, and hence there was insufficient statistical power to test whether there was a true rise in the prevalence of this allele. The overall high frequency of Pfdhfr and Pfdhps quintuple mutations throughout pregnancy excluded further analyses of possible associations between certain haplotypes and the risk of lower birth weight and anemia. However, women infected with P. falciparum had 1.3-g/dl-lower hemoglobin levels (P ‫؍‬ 0.001) and delivered babies with a 400-g-lower birth weight (P ‫؍‬ 0.001) compared to nonparasitemic women. Despite this, 44 women who were P. falciparum positive at baseline became negative after one or two doses of SP (i.e., 50.5%), implying that SP-IPTp still has some efficacy. P. falciparum resistance markers to SP are high in this population, whereas P. falciparum infection was associated with poor birth outcomes. Malaria in pregnancy is a major public health problem in areas where malaria is endemic (1, 2). The current policy in Uganda for treating malaria in pregnancy in the first trimester is to give quinine (10 mg/kg). During the second and third trimesters, the following artemesinin-based combination therapy is recommended: Coartem (artemether [20 mg] and lumefantrine [120 mg]) at four tablets twice a day for 3 days. For malaria prevention, the policy recommendation is at least two doses of sulfadoxinepyrimethamine (SP) as an intermittent preventive treatment of malaria in pregnancy (IPTp) (3). Recently, the World Health Organization (WHO) issued new guidelines on IPTp for countries in areas of moderate to high transmission: IPTp with SP is now recommended for all pregnant women at each scheduled antenatal care (ANC) visit, and each SP dose should be given at least monthly (4). This recommendation is based on evidence that three or more doses of SP as IPTp are more beneficial in reducing the risk of low-birth-w...