Mapping the Spread of Malaria Drug Resistance

Tim Anderson discusses a new study of molecular variation in alleles at the dihydropteroate synthase locus, which underlies resistance to sulfadoxine, in over 5,000 parasites from 50 locations

HIV vaccine trial safety and retention among 18-20 year olds in the HVTN 503/Phambili study support the inclusion of adolescents in future trials

Worldwide, many adolescents, especially women, acquire HIV before age 18. Yet to date, no HIV vaccine trials have enrolled adolescents. Reasons for excluding adolescents from these trials include regulations protecting vulnerable subjects and concerns regarding informed consent, social harms, adverse events, and loss to follow-up

Varying efficacy of artesunate+amodiaquine and artesunate+sulphadoxine-pyrimethamine for the treatment of uncomplicated falciparum malaria in the Democratic Republic of Congo: a report of two in-vivo studies

BACKGROUND: Very few data on anti-malarial efficacy are available from the Democratic Republic of Congo (DRC). DRC changed its anti-malarial treatment policy to amodiaquine (AQ) and artesunate (AS) in 2005. METHODS: The results of two in vivo efficacy studies, which tested AQ and sulphadoxine-pyrimethamine (SP) monotherapies and AS+SP and AS+AQ combinations in Boende (Equatorial province), and AS+SP, AS+AQ and SP in Kabalo (Katanga province), between 2003 and 2004 are presented. The methodology followed the WHO 2003 protocol for assessing the efficacy of anti-malarials in areas of high transmission. RESULTS: Out of 394 included patients in Boende, the failure rates on day 28 after PCR-genotyping adjustment of AS+SP and AS+AQ were estimated as 24.6% [95% CI: 16.6-35.5] and 15.1% [95% CI: 8.6-25.7], respectively. For the monotherapies, failure rates were 35.9% [95% CI: 27.0-46.7] for SP and 18.3% [95% CI: 11.6-28.1] for AQ. Out of 207 patients enrolled in Kabalo, the failure rate on day 28 after PCR-genotyping adjustment was 0 [1-sided 95% CI: 5.8] for AS+SP and AS+AQ [1-sided 95% CI: 6.2]. It was 19.6% [95% CI: 11.4-32.7] for SP monotherapy. CONCLUSION: The finding of varying efficacy of the same combinations at two sites in one country highlights one difficulty of implementing a uniform national treatment policy in a large country. The poor efficacy of AS+AQ in Boende should alert the national programme to foci of resistance and emphasizes the need for systems for the prospective monitoring of treatment efficacy at sentinel sites in the country

Protecting the malaria drug arsenal: halting the rise and spread of amodiaquine resistance by monitoring the PfCRT SVMNT type

The loss of chloroquine due to selection and spread of drug resistant Plasmodium falciparum parasites has greatly impacted malaria control, especially in highly endemic areas of Africa. Since chloroquine removal a decade ago, the guidelines to treat falciparum malaria suggest combination therapies, preferentially with an artemisinin derivative. One of the recommended partner drugs is amodiaquine, a pro-drug that relies on its active metabolite monodesethylamodiaquine, and is still effective in areas of Africa, but not in regions of South America. Genetic studies on P. falciparum parasites have shown that different pfcrt mutant haplotypes are linked to distinct levels of chloroquine and amodiaquine responses. The pfcrt haplotype SVMNT (termed after the amino acids from codon positions 72-76) is stably present in several areas where amodiaquine was introduced and widely used. Parasites with this haplotype are highly resistant to monodesethylamodiaquine and also resistant to chloroquine. The presence of this haplotype in Africa was found for the first time in 2004 in Tanzania and a role for amodiaquine in the selection of this haplotype was suggested. This commentary discusses the finding of a second site in Africa with high incidence of this haplotype. The >50% SVMNT haplotype prevalence in Angola represents a threat to the rise and spread of amodiaquine resistance. It is paramount to monitor pfcrt haplotypes in every country currently using amodiaquine and to re-evaluate current combination therapies in areas where SVMNT type parasites are prevalent

No Evidence of Association between HIV-1 and Malaria in Populations with Low HIV-1 Prevalence

The geographic overlap between HIV-1 and malaria has generated much interest in their potential interactions. A variety of studies have evidenced a complex HIV-malaria interaction within individuals and populations that may have dramatic effects, but the causes and implications of this co-infection at the population level are still unclear. In a previous publication, we showed that the prevalence of malaria caused by the parasite Plasmodium falciparum is associated with HIV infection in eastern sub-Saharan Africa. To complement our knowledge of the HIV-malaria co-infection, the objective of this work was to assess the relationship between malaria and HIV prevalence in the western region of sub-Saharan Africa.Population-based cross-sectional data were obtained from the HIV/AIDS Demographic and Health Surveys conducted in Burkina Faso, Ghana, Guinea, Mali, Liberia and Cameroon, and the malaria atlas project. Using generalized linear mixed models, we assessed the relationship between HIV-1 and Plasmodium falciparum parasite rate (PfPR) adjusting for important socio-economic and biological cofactors. We found no evidence that individuals living in areas with stable malaria transmission (PfPR>0.46) have higher odds of being HIV-positive than individuals who live in areas with PfPR≤0.46 in western sub-Saharan Africa (estimated odds ratio 1.14, 95% confidence interval 0.86-1.50). In contrast, the results suggested that PfPR was associated with being infected with HIV in Cameroon (estimated odds ratio 1.56, 95% confidence interval 1.23-2.00).Contrary to our previous research on eastern sub-Saharan Africa, this study did not identify an association between PfPR and infection with HIV in western sub-Saharan Africa, which suggests that malaria might not play an important role in the spread of HIV in populations where the HIV prevalence is low. Our work highlights the importance of understanding the epidemiologic effect of co-infection and the relevant factors involved in this relationship for the implementation of effective control strategies

Efficacy of antimalarial treatment in Guinea: in vivo study of two artemisinin combination therapies in Dabola and molecular markers of resistance to sulphadoxine-pyrimethamine in N'Zérékoré

BACKGROUND: In the last five years, countries have been faced with changing their malaria treatment policy to an artemisinin-based combination therapy (ACT), many with no national data on which to base their decision. This is particularly true for a number of West African countries, including Guinea, where these studies were performed. Two studies were conducted in 2004/2005 in programmes supported by Medecins Sans Frontieres, when chloroquine was still national policy, but artesunate (AS)/sulphadoxine-pyrimethamine (SP) had been used in refugee camps for two years. METHODS: In Dabola (central Guinea), 220 children aged 6-59 months with falciparum malaria were randomized to receive either AS/amodiaquine (AQ) or AS/SP. In vivo efficacy was assessed following the 2003 World Health Organization guidelines. In a refugee camp in Laine (south of Guinea), where an in vivo study was not feasible due to the unstable context, a molecular genotyping study in 160 patients assessed the prevalence of mutations in the dihydrofolate reductase (dhfr) (codons 108, 51, 59) and dihydropteroate synthase (dhps) (codons 436, 437, 540) genes of Plasmodium falciparum, which have been associated with resistance to pyrimethamine and sulphadoxine, respectively. RESULTS: In Dabola, after 28 days of follow-up, Polymerase Chain Reaction (PCR)-adjusted failure rates were 1.0% (95%CI 0-5.3) for AS/AQ and 1.0% (95%CI 0-5.5) for AS/SP. In the refugee camp in Laine, the molecular genotyping study found three dhfr mutations in 85.6% (95%CI 79.2-90.7) patients and quintuple dhfr/dhps mutations in 9.6% (95%CI 5.2-15.9). CONCLUSION: Both AS/AQ and AS/SP are highly efficacious in Dabola, whereas there is molecular evidence of established SP resistance in Laine. This supports the choice of the national programme of Guinea to adopt AS/AQ as first line antimalarial treatment. The results highlight the difficulties faced by control programmes, which have gone through the upheaval of implementing ACTs, but cannot predict how long their therapeutic life will be, especially in countries which have chosen drugs also available as monotherapies

Molecular epidemiology of drug-resistant malaria in western Kenya highlands

<p>Abstract</p> <p>Background</p> <p>Since the late 1980s a series of malaria epidemics has occurred in western Kenya highlands. Among the possible factors that may contribute to the highland malaria epidemics, parasite resistance to antimalarials has not been well investigated.</p> <p>Methods</p> <p>Using parasites from highland and lowland areas of western Kenya, we examined key mutations associated with <it>Plasmodium falciparum </it>resistance to sulfadoxine – pyrimethamine and chloroquine, including dihydrofolate reductase (<it>pfdhfr</it>) and dihydropteroate synthetase (<it>pfdhps</it>), chloroquine resistance transporter gene (<it>pfcrt</it>), and multi-drug resistance gene 1 (<it>pfmdr1</it>).</p> <p>Results</p> <p>We found that >70% of samples harbored 76T <it>pfcrt </it>mutations and over 80% of samples harbored quintuple mutations (51I/59R/108N <it>pfdhfr </it>and 437G/540E <it>pfdhps</it>) in both highland and lowland samples. Further, we did not detect significant difference in the frequencies of these mutations between symptomatic and asymptomatic malaria volunteers, and between highland and lowland samples.</p> <p>Conclusion</p> <p>These findings suggest that drug resistance of malaria parasites in the highlands could be contributed by the mutations and their high frequencies as found in the lowland. The results are discussed in terms of the role of drug resistance as a driving force for malaria outbreaks in the highlands.</p

Drug resistance to sulphadoxine-pyrimethamine in Plasmodium falciparum malaria in Mlimba, Tanzania

BACKGROUND: Sulphadoxine-pyrimethamine (SP) has been and is currently used for treatment of uncomplicated Plasmodium falciparum malaria in many African countries. Nevertheless, the response of parasites to SP treatment has shown significant variation between individuals. METHODS: The genes for dihydrofolate reductase (dhfr) and dihydropteroate synthase (dhps) were used as markers, to investigate parasite resistance to SP in 141 children aged less than 5 years. Parasite DNA was extracted by Chelex method from blood samples collected and preserved on filter papers. Subsequently, polymerase chain reaction (PCR) and restriction fragment length polymorphism (PCR-RFLP) were applied to detect the SP resistance-associated point mutations on dhfr and dhps. Commonly reported point mutations at codons 51, 59, 108 and 164 in the dhfr and codons 437, 540 and 581 in the dhps domains were examined. RESULTS: Children infected with parasites harbouring a range of single to quintuple dhfr/dhps mutations were erratically cured with SP. However, the quintuple dhfr/dhps mutant genotypes were mostly associated with treatment failures. High proportion of SP resistance-associated point mutations was detected in this study but the adequate clinical response (89.4%) observed clinically at day 14 of follow up reflects the role of semi-immunity protection and parasite clearance in the population. CONCLUSION: In monitoring drug resistance to SP, concurrent studies on possible confounding factors pertaining to development of resistance in falciparum malaria should be considered. The SP resistance potential detected in this study, cautions on its useful therapeutic life as an interim first-line drug against malaria in Tanzania and other malaria-endemic countries

Molecular markers of anti-malarial drug resistance in Lahj Governorate, Yemen: baseline data and implications

<p>Abstract</p> <p>Background</p> <p>This is an investigation of anti-malarial molecular markers coupled with a therapeutic efficacy test of chloroquine (CQ) against falciparum malaria in an area of unstable malaria in Lahj Governorate, Yemen. The study was aimed at assessment of therapeutic response to CQ and elucidation of baseline information on molecular markers for <it>Plasmodium falciparum </it>resistance against CQ and sulphadoxine/pyrimethamine (SP).</p> <p>Methods</p> <p>Between 2002 and 2003 the field test was conducted according to the standard WHO protocol to evaluate the therapeutic efficacy of CQ in 124 patients with falciparum malaria in an endemic area in Lahj Governorate in Yemen. Blood samples collected during this study were analysed for <it>P. falciparum </it>chloroquine resistance transporter gene (<it>pfcrt</it>)-76 polymorphisms, mutation <it>pfcrt-</it>S163R and the antifolate resistance-associated mutations dihydrofolate reductase (<it>dhfr</it>)-C59R and dihydropteroate synthase (<it>dhps</it>)-K540E. Direct DNA sequencing of the <it>pfcrt </it>gene from three representative field samples was carried out after DNA amplification of the 13 exons of the <it>pfcrt </it>gene.</p> <p>Results</p> <p>Treatment failure was detected in 61% of the 122 cases that completed the 14-day follow-up. The prevalence of mutant <it>pfcrt </it>T76 was 98% in 112 amplified pre-treatment samples. The presence of <it>pfcrt </it>T76 was poorly predictive of <it>in vivo </it>CQ resistance (PPV = 61.8%, 95% CI = 52.7-70.9). The prevalence of <it>dhfr </it>Arg-59 mutation in 99 amplified samples was 5%, while the <it>dhps </it>Glu-540 was not detected in any of 119 amplified samples. Sequencing the <it>pfcrt </it>gene confirmed that Yemeni CQ resistant <it>P. falciparum </it>carry the old world (Asian and African) CQ resistant haplotype CVIETSESI at positions 72,73,74,75,76,220,271, 326 and 371.</p> <p>Conclusion</p> <p>This is the first study to report baseline information on the characteristics and implications of anti-malarial drug resistance markers in Yemen. It is also the first report of the haplotype associated with CQR <it>P. falciparum </it>parasites from Yemen. Mutant <it>pfcrt</it>T76 is highly prevalent but it is a poor predictor of treatment failure in the study population. The prevalence of mutation <it>dhfr</it>Arg59 is suggestive of emerging resistance to SP, which is currently a component of the recommended combination treatment of falciparum malaria in Yemen. More studies on these markers are recommended for surveillance of resistance in the study area.</p

Submicroscopic Gametocytes and the Transmission of Antifolate-Resistant Plasmodium falciparum in Western Kenya

BACKGROUND: Single nucleotide polymorphisms (SNPs) in the dhfr and dhps genes are associated with sulphadoxine-pyrimethamine (SP) treatment failure and gametocyte carriage. This may result in enhanced transmission of mutant malaria parasites, as previously shown for chloroquine resistant parasites. In the present study, we determine the association between parasite mutations, submicroscopic P. falciparum gametocytemia and malaria transmission to mosquitoes. METHODOLOGY/PRINCIPAL FINDINGS: Samples from children treated with SP alone or in combination with artesunate (AS) or amodiaquine were genotyped for SNPs in the dhfr and dhps genes. Gametocytemia was determined by microscopy and Pfs25 RNA-based quantitative nucleic acid sequence-based amplification (Pfs25 QT-NASBA). Transmission was determined by membrane-feeding assays. We observed no wild type infections, 66.5% (127/191) of the infections expressed mutations at all three dhfr codons prior to treatment. The presence of all three mutations was not related to higher Pfs25 QT-NASBA gametocyte prevalence or density during follow-up, compared to double mutant infections. The proportion of infected mosquitoes or oocyst burden was also not related to the number of mutations. Addition of AS to SP reduced gametocytemia and malaria transmission during follow-up. CONCLUSIONS/SIGNIFICANCE: In our study population where all infections had at least a double mutation in the dhfr gene, additional mutations were not related to increased submicroscopic gametocytemia or enhanced malaria transmission. The absence of wild-type infections is likely to have reduced our power to detect differences. Our data further support the use of ACT to reduce the transmission of drug-resistant malaria parasites