Plasmodium falciparum Sexual Commitment Rate Variation among Clinical Isolates and Diverse Laboratory-Adapted Lines.

Asexual blood-stage malaria parasites must produce sexual progeny to infect mosquitoes. It is important to understand the scope and causes of intraspecific variation in sexual commitment rates, particularly for the major human parasite P. falciparum. First, two alternative assay methods of measuring sexual commitment were compared to test a genetically modified P. falciparum line with elevated commitment rates inducible by overexpression of GDV1. The methods yielded correlated measurements with higher sensitivity and precision being achieved by one employing detection of the early gametocyte differentiation marker Pfs16. Thus, this was used to survey a diverse range of parasite lines and test each in multiple biological replicate assays in a serum-free medium supplemented with Albumax. There were differences among six recent clinical isolates from Ghana in their mean rates of sexual commitment per cycle, ranging from 3.3% to 12.2%. Among 13 diverse long-term laboratory-adapted lines, mean sexual commitment rates for most ranged from 4.7% to 13.4%, a few had lower rates with means from 0.3 to 1.6%, and one with a nonfunctional ap2-g gene always showed zero commitment. Among a subset of lines tested for the effects of exogenous choline to suppress commitment, there were significant differences. As expected, there was no effect in a line that had lost the gdv1 gene and that had generally low commitment, whereas the others showed quantitatively variable but significant responses to choline, suggesting potential trait variation. The results indicated the value of performing multiple replicate assays for understanding the variation of this key reproductive trait that likely affects transmission. IMPORTANCE Only sexual-stage malaria parasites are transmitted from human blood to mosquitoes. Thus, it is vital to understand variations in sexual commitment rates because these may be modifiable or susceptible to blocking. Two different methods of commitment rate measurement were first compared, demonstrating higher sensitivity and precision by the detection of an early differentiation marker, which was subsequently used to survey diverse lines. Clinical isolates from Ghana showed significant variation in mean per-cycle commitment rates and variation among biological replicates. Laboratory-adapted lines of diverse origins had a wider range with most being within the range observed for the clinical isolates, while a minority consistently had lower or zero rates. There was quantitative variation in the effects when adding choline to suppress commitment, indicating differing responsiveness of parasites to this environmental modification. Performing multiple assay replicates and comparisons of diverse isolates was important to understand this trait and its potential effects on transmission.

Improving malaria control in West Africa: interruption of transmission as a paradigm shift.

With the paradigm shift from the reduction of morbidity and mortality to the interruption of transmission, the focus of malaria control broadens from symptomatic infections in children ≤5 years of age to include asymptomatic infections in older children and adults. In addition, as control efforts intensify and the number of interventions increases, there will be decreases in prevalence, incidence and transmission with additional decreases in morbidity and mortality. Expected secondary consequences of these changes include upward shifts in the peak ages for infection (parasitemia) and disease, increases in the ages for acquisition of antiparasite humoral and cellular immune responses and increases in false-negative blood smears and rapid diagnostic tests. Strategies to monitor these changes must include: (1) studies of the entire population (that are not restricted to children ≤5 or ≤10 years of age), (2) study sites in both cities and rural areas (because of increasing urbanization across sub-Saharan Africa) and (3) innovative strategies for surveillance as the prevalence of infection decreases and the frequency of false-negative smears and rapid diagnostic tests increases.

Sahel, savana, riverine and urban malaria in West Africa: Similar control policies with different outcomes.

The study sites for the West African ICEMR are in three countries (The Gambia, Senegal, Mali) and are located within 750 km of each other. In addition, the National Malaria Control Programmes of these countries have virtually identical policies: (1) Artemisinin Combination Therapies (ACTs) for the treatment of symptomatic Plasmodium falciparum infection, (2) Long-Lasting Insecticide-treated bed Nets (LLINs) to reduce the Entomololgic Inoculation Rate (EIR), and (3) sulfadoxine-pyrimethamine for the Intermittent Preventive Treatment of malaria during pregnancy (IPTp). However, the prevalence of P. falciparum malaria and the status of malaria control vary markedly across the four sites with differences in the duration of the transmission season (from 4-5 to 10-11 months), the intensity of transmission (with EIRs from unmeasurably low to 4-5 per person per month), multiplicity of infection (from a mean of 1.0 to means of 2-5) and the status of malaria control (from areas which have virtually no control to areas that are at the threshold of malaria elimination). The most important priority is the need to obtain comparable data on the population-based prevalence, incidence and transmission of malaria before new candidate interventions or combinations of interventions are introduced for malaria control.

Prevention of the recurrence of anaemia in Gambian children following discharge from hospital.

BACKGROUND: In malaria endemic countries, children who have experienced an episode of severe anaemia are at increased risk of a recurrence of anaemia. There is a need to find ways of protecting these at risk children from malaria and chemoprevention offers a potential way of achieving this objective. METHODS: During the 2003 and 2004 malaria transmission seasons, 1200 Gambian children with moderate or severe anaemia (Hb concentration <7 g/dL) were randomised to receive either monthly sulfadoxine-pyrimethamine (SP) or placebo until the end of the malaria transmission season in which they were enrolled, in a double-blind trial. All study subjects were treated with oral iron for 28 days and morbidity was monitored through surveillance at health centres. The primary endpoint was the proportion of children with moderate or severe anaemia at the end of the transmission season. Secondary endpoints included the incidence of clinical episodes of malaria during the surveillance period, outpatient attendances, the prevalence of parasitaemia and splenomegaly, nutritional status at the end of the malaria transmission season and compliance with the treatment regimen. RESULTS: The proportions of children with a Hb concentration of <7 g/dL at the end of the malaria transmission season were similar in the two study groups, 14/464 (3.0%) in children who received at least one dose of SP and 16/471 (3.4%) in those who received placebo, prevalence ratio 0.89 (0.44,1.8) P = 0.742. The protective efficacy of SP against episodes of clinical malaria was 53% (95% CI 37%, 65%). Treatment with SP was safe and well tolerated; no serious adverse events related to SP administration were observed. Mortality following discharge from hospital was low among children who received SP or placebo (6 in the SP group and 9 in the placebo group respectively). CONCLUSIONS: Intermittent treatment with SP did not reduce the proportion of previously anaemic children with moderate or severe anaemia at the end of the malaria season, although it prevented malaria. The combination of appropriate antimalarial treatment plus one month of iron supplementation and good access to healthcare during follow-up proved effective in restoring haemoglobin to an acceptable level in the Gambian setting. TRIAL REGISTRATION: ClinicalTrials.gov NCT00131716.

Oral activated charcoal prevents experimental cerebral malaria in mice and in a randomized controlled clinical trial in man did not interfere with the pharmacokinetics of parenteral artesunate.

BACKGROUND: Safe, cheap and effective adjunct therapies preventing the development of, or reducing the mortality from, severe malaria could have considerable and rapid public health impact. Oral activated charcoal (oAC) is a safe and well tolerated treatment for acute poisoning, more recently shown to have significant immunomodulatory effects in man. In preparation for possible efficacy trials in human malaria, we sought to determine whether oAC would i) reduce mortality due to experimental cerebral malaria (ECM) in mice, ii) modulate immune and inflammatory responses associated with ECM, and iii) affect the pharmacokinetics of parenteral artesunate in human volunteers. METHODS/PRINCIPAL FINDINGS: We found that oAC provided significant protection against P. berghei ANKA-induced ECM, increasing overall survival time compared to untreated mice (p<0.0001; hazard ratio 16.4; 95% CI 6.73 to 40.1). Protection from ECM by oAC was associated with reduced numbers of splenic TNF(+) CD4(+) T cells and multifunctional IFNgamma(+)TNF(+) CD4(+) and CD8(+) T cells. Furthermore, we identified a whole blood gene expression signature (68 genes) associated with protection from ECM. To evaluate whether oAC might affect current best available anti-malarial treatment, we conducted a randomized controlled open label trial in 52 human volunteers (ISRCTN NR. 64793756), administering artesunate (AS) in the presence or absence of oAC. We demonstrated that co-administration of oAC was safe and well-tolerated. In the 26 subjects further analyzed, we found no interference with the pharmacokinetics of parenteral AS or its pharmacologically active metabolite dihydroartemisinin. CONCLUSIONS/SIGNIFICANCE: oAC protects against ECM in mice, and does not interfere with the pharmacokinetics of parenteral artesunate. If future studies succeed in establishing the efficacy of oAC in human malaria, then the characteristics of being inexpensive, well-tolerated at high doses and requiring no sophisticated storage would make oAC a relevant candidate for adjunct therapy to reduce mortality from severe malaria, or for immediate treatment of suspected severe malaria in a rural setting. TRIAL REGISTRATION: Controlled-Trials.com ISRCTN64793756.