We assessed Plasmodium falciparum mdr1 (Pfmdr1) gene polymorphisms and copy numbers as well as P. falciparum Ca 2؉ ATPase (PfATPase6) gene polymorphisms in 90 Nigerian children presenting with uncomplicated falciparum malaria and enrolled in a study of the efficacy of artemether-lumefantrine (AL). The nested PCR-restriction fragment length polymorphism and the quantitative real-time PCR methodologies were used to determine the alleles of the Pfmdr1 and PfATPase6 genes and the Pfmdr1 copy number variation, respectively, in patients samples collected prior to treatment and at the reoccurrence of parasites during a 42-day follow-up. The Pfmdr1 haplotype 86N-184F-1246D was significantly associated (P < 0.00001) with treatment failures and was selected for among posttreatment samples obtained from patients with newly acquired or recrudescing infections (P < 0.00001; 2 ‫؍‬ 36.5) and in gametocytes (log rank statistic ‫؍‬ 5; P ‫؍‬ 0.0253) after treatment with AL. All pre-and posttreatment samples as well as gametocytes harbored a single copy of the Pfmdr1 gene and the wild-type allele (L89) at codon 89 of the PfATPase6 gene. These findings suggest that polymorphisms in the Pfmdr1 gene are under AL selection pressure. Pfmdr1 polymorphisms may result in reduction in the therapeutic efficacy of this newly adopted combination treatment for uncomplicated falciparum malaria in Saharan countries of Africa.

Computed tomography was performed on 14 unconscious Kenyan children recovering from cerebral malaria (seven ofwhom had another scan 12-120 days later) to elucidate the cause of intracranial hypertension and neurological sequelae. Brain swelling, defined as a loss of cerebrospinal fluid spaces, was documented in six children, while a further two had conspicuously small ventricles only. There was severe intracranial hypertension in the two children with definite brain swelling in whom intracranial pressure was monitored. There was no evidence of acute hydrocephalus or vasogenic oedema. Four children with brain swelling also had widespread low density areas suggestive of ischaemic damage. The patterns of damage were not uniform but were consistent with a critical reduction in cerebral perfusion pressure (which was documented in the two in whom this was monitored), hypoglycaemia, or status epilepticus. All four had serious neurological sequelae. These data suggest that brain injury in cerebral malaria may be due in part to secondary systemic and intracranial factors as well as to the direct effect of intravascular sequestration.

This study investigated the association between Plasmodium falciparum chloroquine resistance transporter (pfcrt) T76 and P. falciparum multidrug resistance gene 1 (pfmdr1) Y86 alleles and in vivo amodiaquine (AQ) resistance, as well as the clearance of parasites harboring these two alleles in children treated with AQ in southwest Nigeria. One hundred one children with acute uncomplicated P. falciparum malaria infections were treated with the standard dosage of AQ and followed-up for 28 days. Blood samples were collected on filter paper samples at enrollment and during follow-up for identification of parasite genotypes and pfcrt and pfmdr1 mutations using polymerase chain reaction and restriction fragment length polymorphism approaches. Parasitologic assessment of response to treatment showed that 87% and 13% (RI) of patients were cured and failed treatment, respectively. Although infections in patients were polyclonal (as determined by merozoite surface protein 2 genotyping), the presence of both mutants pfcrtT76 and pfmdr1Y86 alleles in parasites is associated with in vivo AQ resistance (odds ratio = 7.58, 95% confidence interval = 1.58-36.25, P = 0.006) and is selected by the drug in children who failed AQ treatment. Treatment failure with the combination of mutant pfcrtT76 and pfmdr1Y86 alleles as well as the ability of patients to clear these resistant parasites is dependent on age, suggesting a critical role of host immunity in clearing AQ-resistant P. falciparum. The combination of mutant pfcrtT76 and pfmdr1Y86 alleles may be useful markers for monitoring the development and spread of AQ resistance, when combining this drug with other antimalarials for treatment of malaria in Africa.

Parasite genotyping by a polymerase chain reaction was used to distinguish recrudescent from newly acquired Plasmodium falciparum infections in 50 of 160 Nigerian children taking part in a chloroquine efficacy study in Ibadan, Nigeria. A finger prick blood sample was taken from each child before and after treatment to identify recrudescent parasites. By investigating allelic variation in three polymorphic antigen loci, merozoite surface protein-1 (MSP-1), MSP-2, and glutamate-rich protein (GLURP), we determined parasite diversity in the population and in the infected host. DNA from pretreatment and post-treatment samples from 47 of the 50 patients who failed therapy was successfully amplified by the PCR. The MSP-1, MSP-2, and GLURP genotypes in all samples showed extensive diversity, indicating polyclonal infections. The average number of clones per infection in pre-treatment sample was 2.5 with MSP-1, 4.9 with MSP-2, and 2 with GLURP. The extent of multiplicity decreased significantly (P = 0.016) in posttreatment samples. Multiplicity of infection and initial parasite density were not age dependent. Comparison of the variant alleles in pretreatment and post-treatment samples of each patient indicates that 26 of the 47 children had genuinely recrudescent disease. Conversely, post-treatment samples from five children showed completely new genotypes, indicating either a previously sequestered population of parasites or a newly acquired infection. Overall, this study has shown the diversity and complexity of P. falciparum population in Ibadan, Nigeria. The study has also shown the dynamics of P. falciparum infections in this population before and after chloroquine treatment in an area of high malaria transmission.

No abstract

No abstract

Chloroquine (CQ) resistance in Plasmodium falciparum has been associated with specific point mutations in the pfcrt and pfmdr-1 genes. In the present study, 30 children aged 1-12 years, who were all suffering from acute, uncomplicated, P. falciparum malaria in Ibadan, Nigeria, were evaluated to assess the association between these mutations and clinical outcome following treatment with CQ. The parasites, in blood samples collected pre-treatment and, in those who failed treatment, on the day symptoms re-occurred post-treatment, were genotyped using the polymorphic MSP1, MSP2 and GLURP loci and PCR-RFLP. The results showed that, pre-treatment, all 30 patients had polyclonal infections, the mean numbers of P. falciparum clones detected per infection being 2.6 with MSP1, 4.2 with MSP2 and 2.8 with GLURP. The T76 allele of pfcrt and the Y86 allele of pfmdr-1 were found in 53% and 40%, respectively, of the pre-treatment samples from the 15 patients who failed CQ treatment, but the Y1246 mutation in pfmdr-1 was never detected. Although the parasites from the two patients with high-grade (RIII) resistance to CQ had both of these point mutations, the presence of the T76 allele of pfcrt or the Y86 allele of pfmdr-1 (considered individually) could not be used to predict treatment outcome. However, a high frequency of clonal multiplicity may confound attempts to associate the point mutations in pfcrt or pfmdr-1 with clinical response to CQ. It remains unclear whether the present results represent the characteristics of the predominant parasite populations in the study area. Further studies are needed before the strength of the association between the point mutations identified as markers of drug resistance and clinical outcome can be accurately evaluated, in this and other regions of intense transmission.

BackgroundChlorproguanil−dapsone−artesunate (CDA) was developed as an affordable, simple, fixed-dose artemisinin-based combination therapy for use in Africa. This trial was a randomized parallel-group, double-blind, double-dummy study to compare CDA and artemether−lumefantrine (AL) efficacy in uncomplicated Plasmodium falciparum malaria and further define the CDA safety profile, particularly its hematological safety in glucose-6-phosphate dehydrogenase (G6PD) -deficient patients.Methods and FindingsThe trial was conducted at medical centers at 11 sites in five African countries between June 2006 and August 2007. 1372 patients (≥1 to <15 years old, median age 3 years) with acute uncomplicated P. falciparum malaria were randomized (2∶1) to receive CDA 2/2.5/4 mg/kg once daily for three days (N = 914) or six-doses of AL over three days (N = 458). Non-inferiority of CDA versus AL for efficacy was evaluated in the Day 28 per-protocol (PP) population using parasitological cure (polymerase chain reaction [PCR]-corrected). Cure rates were 94.1% (703/747) for CDA and 97.4% (369/379) for AL (treatment difference –3.3%, 95%CI –5.6, −0.9). CDA was non-inferior to AL, but there was simultaneous superiority of AL (upper 95%CI limit <0). Adequate clinical and parasitological response at Day 28 (uncorrected for reinfection) was 79% (604/765) with CDA and 83% (315/381) with AL. In patients with a G6PD-deficient genotype (94/603 [16%] hemizygous males, 22/598 [4%] homozygous females), CDA had the propensity to cause severe and clinically concerning hemoglobin decreases: the mean hemoglobin nadir was 75 g/L (95%CI 71, 79) at Day 7 versus 97 g/L (95%CI 91, 102) for AL. There were three deaths, unrelated to study medication (two with CDA, one with AL).ConclusionsAlthough parasitologically effective at Day 28, the hemolytic potential of CDA in G6PD-deficient patients makes it unsuitable for use in a public health setting in Africa.Trial RegistrationClinicalTrials.gov NCT00344006