Steffen Borrmann

Long-Term Treatment of Intestinal Helminths Increases Mite Skin-Test Reactivity in Gabonese Schoolchildren

BACKGROUND Several studies have shown an inverse association between helminth infections and atopy, but none have clearly established that the pathogens themselves, rather than other associated factors, cause the suppression of atopy. To show a direct link, prospective intervention studies are required. METHODS A randomized, controlled trial was performed to study whether repeated anthelminthic treatment results in increased allergic sensitivity to house dust mites (HDMs) in chronically infected children. The trial population consisted of 317 Gabonese schoolchildren with a high prevalence of intestinal helminths. Intervention consisted of treatment every 3 months with praziquantel and mebendazole and with placebo in the control group. Follow-up lasted 30 months: at 6-month intervals, skin-test sensitivity to mites, helminth infection status, and levels of total IgE were determined. RESULTS Treatment resulted in a significant increase in the rate of developing skin sensitivity to HDMs (hazard ratio, 2.51; 95% confidence interval, 1.85-3.41), which was mediated, in part, by reductions in Ascaris and/or Trichuris infections. Levels of total IgE were reduced, but this did not mediate the effect of treatment on skin-test reactivity. CONCLUSIONS Anthelminthic treatment of chronically infected children results in increased atopic reactivity, which indicates that helminths directly suppress allergic reactions.

Amodiaquine-artesunate versus amodiaquine for uncomplicated Plasmodium falciparum malaria in African children: a randomised, multicentre trial

BACKGROUND Increasing drug resistance limits the choice of efficacious chemotherapy against Plasmodium falciparum malaria in Africa. Amodiaquine still retains efficacy against P falciparum in many African countries. We assessed the safety, treatment efficacy, and effect on gametocyte carriage of adding artesunate to amodiaquine in three randomised trials in Kenya, Sénégal, and Gabon. METHODS We enrolled 941 children (400 in Kenya, 321 in Sénégal, and 220 in Gabon) who were 10 years or older and who had uncomplicated P falciparum malaria. Patients were randomly assigned amodiaquine (10 mg/kg per day for 3 days) plus artesunate (4 mg/kg per day for 3 days) or amodiaquine (as above) and placebo (for 3 days). The primary endpoints were parasitological cure rates at days 14 and 28. Analysis was by intention to treat and by an evaluability method. FINDINGS Both regimens were well tolerated. Six patients in the amodiaquine-artesunate group and five in the amodiaquine group developed early, drug-induced vomiting, necessitating alternative treatment. By intention-to-treat analysis, the day-14 cure rates for amodiaquine-artesunate versus amodiaquine were: 175/192 (91%) versus 140/188 (74%) in Kenya (D=16.7% [95% CI 9.3-24.1], p<0.0001), 148/160 (93%) versus 147/157 (94%) in Sénégal (-1.1% [-6.7 to 4.5], p=0.7), and 92/94 (98%) versus 86/96 (90%) in Gabon (8.3% [1.5-15.1], p=0.02). The corresponding rates for day 28 were: 123/180 (68%) versus 75/183 (41%) in Kenya (27.3% [17.5-37.2], p<0.0001), 130/159 (82%) versus 123/156 (79%) in Sénégal (2.9% [-5.9 to 11.7], p=0.5), and 80/94 (85%) versus 70/98 (71%) in Gabon (13.7% [2.2-25.2], p=0.02). Similar rates were obtained by evaluability analysis. INTERPRETATION The combination of artesunate and amodiaquine improved treatment efficacy in Gabon and Kenya, and was equivalent in Sénégal. Amodiaquine-artesunate is a potential combination for use in Africa. Further investigations to assess the potential effect on the evolution of drug resistance, disease transmission, and safety of amodiaquine-artesunate are warranted.

Analysis of Plasmodium falciparum diversity in natural infections by deep sequencing

Malaria elimination strategies require surveillance of the parasite population for genetic changes that demand a public health response, such as new forms of drug resistance. Here we describe methods for the large-scale analysis of genetic variation in Plasmodium falciparum by deep sequencing of parasite DNA obtained from the blood of patients with malaria, either directly or after short-term culture. Analysis of 86,158 exonic single nucleotide polymorphisms that passed genotyping quality control in 227 samples from Africa, Asia and Oceania provides genome-wide estimates of allele frequency distribution, population structure and linkage disequilibrium. By comparing the genetic diversity of individual infections with that of the local parasite population, we derive a metric of within-host diversity that is related to the level of inbreeding in the population. An open-access web application has been established for the exploration of regional differences in allele frequency and of highly differentiated loci in the P. falciparum genome.

Artemether-lumefantrine treatment of uncomplicated Plasmodium falciparum malaria: a systematic review and meta-analysis of day 7 lumefantrine concentrations and therapeutic response using individual patient data

Achieving adequate antimalarial drug exposure is essential for curing malaria. Day 7 blood or plasma lumefantrine concentrations provide a simple measure of drug exposure that correlates well with artemether-lumefantrine efficacy. However, the ‘therapeutic’ day 7 lumefantrine concentration threshold needs to be defined better, particularly for important patient and parasite sub-populations. The WorldWide Antimalarial Resistance Network (WWARN) conducted a large pooled analysis of individual pharmacokinetic-pharmacodynamic data from patients treated with artemether-lumefantrine for uncomplicated Plasmodium falciparum malaria, to define therapeutic day 7 lumefantrine concentrations and identify patient factors that substantially alter these concentrations. A systematic review of PubMed, Embase, Google Scholar, ClinicalTrials.gov and conference proceedings identified all relevant studies. Risk of bias in individual studies was evaluated based on study design, methodology and missing data. Of 31 studies identified through a systematic review, 26 studies were shared with WWARN and 21 studies with 2,787 patients were included. Recrudescence was associated with low day 7 lumefantrine concentrations (HR 1.59 (95 % CI 1.36 to 1.85) per halving of day 7 concentrations) and high baseline parasitemia (HR 1.87 (95 % CI 1.22 to 2.87) per 10-fold increase). Adjusted for mg/kg dose, day 7 concentrations were lowest in very young children (<3 years), among whom underweight-for-age children had 23 % (95 % CI −1 to 41 %) lower concentrations than adequately nourished children of the same age and 53 % (95 % CI 37 to 65 %) lower concentrations than adults. Day 7 lumefantrine concentrations were 44 % (95 % CI 38 to 49 %) lower following unsupervised treatment. The highest risk of recrudescence was observed in areas of emerging artemisinin resistance and very low transmission intensity. For all other populations studied, day 7 concentrations ≥200 ng/ml were associated with >98 % cure rates (if parasitemia <135,000/μL). Current artemether-lumefantrine dosing recommendations achieve day 7 lumefantrine concentrations ≥200 ng/ml and high cure rates in most uncomplicated malaria patients. Three groups are at increased risk of treatment failure: very young children (particularly those underweight-for-age); patients with high parasitemias; and patients in very low transmission intensity areas with emerging parasite resistance. In these groups, adherence and treatment response should be monitored closely. Higher, more frequent, or prolonged dosage regimens should now be evaluated in very young children, particularly if malnourished, and in patients with hyperparasitemia.

Fosmidomycin for malaria

Safe and effective antimalarial drugs with new methods of action are urgently needed. Fosmidomycin inhibits the synthesis of isoprenoid by Plasmodium falciparum, and suppresses the growth of multidrug-resistant strains in vitro. Our aim was to assess the efficacy and tolerability of fosmidomycin in adults with malaria in Gabon. We administered the drug for 5, 4, or 3 days (1.2 g every 8 h), in nine, eight, and ten evaluable patients, respectively. All treatment regimens were well tolerated. Cure rates by day 14 were 89% (eight of nine), 88% (seven of eight), and 60% (six of ten), for treatment durations of 5, 4, and 3 days, respectively. These data suggest that fosmidomycin is a safe and effective treatment for uncomplicated malaria if given for 4 days or more.

In Vitro Activities of Piperaquine, Lumefantrine, and Dihydroartemisinin in Kenyan Plasmodium falciparum Isolates and Polymorphisms in pfcrt and pfmdr1

ABSTRACT We have analyzed the in vitro chemosensitivity profiles of 115 Kenyan isolates for chloroquine (CQ), piperaquine, lumefantrine (LM), and dihydroartemisinin in association with polymorphisms in pfcrt at codon 76 and pfmdr1 at codon 86, as well as with variations of the copy number of pfmdr1. The median drug concentrations that inhibit 50% of parasite growth (IC50s) were 41 nM (interquartile range [IQR], 18 to 73 nM), 50 nM (IQR, 29 to 96 nM), 32 nM (IQR, 17 to 46 nM), and 2 nM (IQR, 1 to 3 nM) for CQ, LM, piperaquine, and dihydroartemisinin, respectively. The activity of CQ correlated inversely with that of LM (r2 = −0.26; P = 0.02). Interestingly, parasites for which LM IC50s were higher were wild type for pfcrt-76 and pfmdr1-86. All isolates had one pfmdr1 copy. Thus, the decrease in LM activity is associated with the selection of wild-type pfcrt-76 and pfmdr1-86 parasites, a feature that accounts for the inverse relationship between CQ and LM. Therefore, the use of LM-artemether is likely to lead to the selection of more CQ-susceptible parasites.

Efficacy and safety of artemether-lumefantrine dispersible tablets compared with crushed commercial tablets in African infants and children with uncomplicated malaria: a randomised, single-blind, multicentre trial

BACKGROUND Combination treatments, preferably containing an artemisinin derivative, are recommended to improve efficacy and prevent Plasmodium falciparum drug resistance. Our aim was to show non-inferiority of a new dispersible formulation of artemether-lumefantrine to the conventional crushed tablet in the treatment of young children with uncomplicated malaria. METHODS We did a randomised non-inferiority study on children weighing 5-35 kg with uncomplicated P falciparum malaria in Benin, Kenya, Mali, Mozambique, and Tanzania. The primary outcome measure was PCR-corrected 28-day parasitological cure rate. We aimed to show non-inferiority (with a margin of -5%) of dispersible versus crushed tablet. We constructed an asymptotic one-sided 97.5% CI on the difference in cure rates. A computer-generated randomisation list was kept centrally and investigators were unaware of the study medication administered. We used a modified intention-to-treat analysis. This trial is registered with ClinicalTrials.gov, number NCT00386763. FINDINGS 899 children aged 12 years or younger were randomly assigned to either dispersible (n=447) or crushed tablets (n=452). More than 85% of patients in each treatment group completed the study. 812 children qualified for the modified intention-to-treat analysis (n=403 vs n=409). The PCR-corrected day-28 cure rate was 97.8% (95% CI 96.3-99.2) in the group on dispersible formulation and 98.5% (97.4-99.7) in the group on crushed formulation. The lower bound of the one-sided 97.5% CI was -2.7%. The most common drug-related adverse event was vomiting (n=33 [7%] and n=42 [9%], respectively). No signs of ototoxicity or relevant cardiotoxicity were seen. INTERPRETATION A six-dose regimen of artemether-lumefantrine with the new dispersible formulation is as efficacious as the currently used crushed tablet in infants and children, and has a similar safety profile.

The Prevalence of Parasite Infestation and House Dust Mite Sensitization in Gabonese Schoolchildren

Background: Allergic diseases seem less prevalent in communities in less developed parts of the world, where parasite infections are highly prevalent. Altogether not much is known about the association between chronic infections with tissue and blood-dwelling parasites and atopy. Methods: In an area in Gabon endemic for blood and tissue parasites, 520 schoolchildren were parasitologically examined and skin prick-tested for a set of common environmental aeroallergens. Levels of allergen-specific IgE and polyclonal IgE were measured. Results: In schoolchildren schistosome and filarial infections increased with age, whereas malaria was more prevalent in younger children. In contrast to allergen sensitization that increased with age, skin test reactivity tended to decline. The number of children with mite-specific IgE antibodies (47%) by far exceeded the number responding to skin prick testing (11%). Mite sensitization was found to be the highest in children infected with schistosomes and/or filariae whereas skin test reactivity was lowest. The multiple logistic regression showed that the risk of a positive skin test was 8-fold higher with increasing levels of mite-specific IgE but was reduced by 72% when infected with blood stage helminths. Conclusions: Chronic blood and tissue parasite infections that are often capable of modulating immune responses in the host are negatively associated with skin test reactivity in a sensitized population.

Chloroquine resistance before and after its withdrawal in Kenya

BackgroundThe spread of resistance to chloroquine (CQ) led to its withdrawal from use in most countries in sub-Saharan Africa in the 1990s. In Malawi, this withdrawal was followed by a rapid reduction in the frequency of resistance to the point where the drug is now considered to be effective once again, just nine years after its withdrawal. In this report, the polymorphisms of markers associated with CQ-resistance against Plasmodium falciparum isolates from coastal Kenya (Kilifi) were investigated, from 1993, prior to the withdrawal of CQ, to 2006, seven years after its withdrawal. Changes to those that occurred in the dihydrofolate reductase gene (dhfr) that confers resistance to the replacement drug, pyrimethamine/sulphadoxine were also compared.MethodsMutations associated with CQ resistance, at codons 76 of pfcrt, at 86 of pfmdr1, and at codons 51, 59 and 164 of dhfr were analysed using PCR-restriction enzyme methods. In total, 406, 240 and 323 isolates were genotyped for pfcrt-76, pfmdr1-86 and dhfr, respectively.ResultsFrom 1993 to 2006, the frequency of the pfcrt-76 mutant significantly decreased from around 95% to 60%, while the frequency of pfmdr1-86 did not decline, remaining around 75%. Though the frequency of dhfr mutants was already high (around 80%) at the start of the study, this frequency increased to above 95% during the study period. Mutation at codon 164 of dhf r was analysed in 2006 samples, and none of them had this mutation.ConclusionIn accord with the study in Malawi, a reduction in resistance to CQ following official withdrawal in 1999 was found, but unlike Malawi, the decline of resistance to CQ in Kilifi was much slower. It is estimated that, at current rates of decline, it will take 13 more years for the clinical efficacy of CQ to be restored in Kilifi. In addition, CQ resistance was declining before the drugs official withdrawal, suggesting that, prior to the official ban, the use of CQ had decreased, probably due to its poor clinical effectiveness.

Population Genetic Analysis of Plasmodium falciparum Parasites Using a Customized Illumina GoldenGate Genotyping Assay

The diversity in the Plasmodium falciparum genome can be used to explore parasite population dynamics, with practical applications to malaria control. The ability to identify the geographic origin and trace the migratory patterns of parasites with clinically important phenotypes such as drug resistance is particularly relevant. With increasing single-nucleotide polymorphism (SNP) discovery from ongoing Plasmodium genome sequencing projects, a demand for high SNP and sample throughput genotyping platforms for large-scale population genetic studies is required. Low parasitaemias and multiple clone infections present a number of challenges to genotyping P. falciparum. We addressed some of these issues using a custom 384-SNP Illumina GoldenGate assay on P. falciparum DNA from laboratory clones (long-term cultured adapted parasite clones), short-term cultured parasite isolates and clinical (non-cultured isolates) samples from East and West Africa, Southeast Asia and Oceania. Eighty percent of the SNPs (n = 306) produced reliable genotype calls on samples containing as little as 2 ng of total genomic DNA and on whole genome amplified DNA. Analysis of artificial mixtures of laboratory clones demonstrated high genotype calling specificity and moderate sensitivity to call minor frequency alleles. Clear resolution of geographically distinct populations was demonstrated using Principal Components Analysis (PCA), and global patterns of population genetic diversity were consistent with previous reports. These results validate the utility of the platform in performing population genetic studies of P. falciparum.