Din Syafruddin

TLR4 polymorphisms, infectious diseases, and evolutionary pressure during migration of modern humans.

Infectious diseases exert a constant evolutionary pressure on the genetic makeup of our innate immune system. Polymorphisms in Toll-like receptor 4 (TLR4) have been related to susceptibility to Gram-negative infections and septic shock. Here we show that two polymorphisms of TLR4, Asp299Gly and Thr399Ile, have unique distributions in populations from Africa, Asia, and Europe. Genetic and functional studies are compatible with a model in which the nonsynonymous polymorphism Asp299Gly has evolved as a protective allele against malaria, explaining its high prevalence in subSaharan Africa. However, the same allele could have been disadvantageous after migration of modern humans into Eurasia, putatively because of increased susceptibility to severe bacterial infections. In contrast, the Asp299Gly allele, when present in cosegregation with Thr399Ile to form the Asp299Gly/Thr399Ile haplotype, shows selective neutrality. Polymorphisms in TLR4 exemplify how the interaction between our innate immune system and the infectious pressures in particular environments may have shaped the genetic variations and function of our immune system during the out-of-Africa migration of modern humans.

A Worldwide Map of Plasmodium falciparum K13-Propeller Polymorphisms.

BACKGROUND Recent gains in reducing the global burden of malaria are threatened by the emergence of Plasmodium falciparum resistance to artemisinins. The discovery that mutations in portions of a P. falciparum gene encoding kelch (K13)-propeller domains are the major determinant of resistance has provided opportunities for monitoring such resistance on a global scale. METHODS We analyzed the K13-propeller sequence polymorphism in 14,037 samples collected in 59 countries in which malaria is endemic. Most of the samples (84.5%) were obtained from patients who were treated at sentinel sites used for nationwide surveillance of antimalarial resistance. We evaluated the emergence and dissemination of mutations by haplotyping neighboring loci. RESULTS We identified 108 nonsynonymous K13 mutations, which showed marked geographic disparity in their frequency and distribution. In Asia, 36.5% of the K13 mutations were distributed within two areas--one in Cambodia, Vietnam, and Laos and the other in western Thailand, Myanmar, and China--with no overlap. In Africa, we observed a broad array of rare nonsynonymous mutations that were not associated with delayed parasite clearance. The gene-edited Dd2 transgenic line with the A578S mutation, which expresses the most frequently observed African allele, was found to be susceptible to artemisinin in vitro on a ring-stage survival assay. CONCLUSIONS No evidence of artemisinin resistance was found outside Southeast Asia and China, where resistance-associated K13 mutations were confined. The common African A578S allele was not associated with clinical or in vitro resistance to artemisinin, and many African mutations appear to be neutral. (Funded by Institut Pasteur Paris and others.).

Malaria in pregnancy in the Asia-Pacific region

Most pregnant women at risk of for infection with Plasmodium vivax live in the Asia-Pacific region. However, malaria in pregnancy is not recognised as a priority by many governments, policy makers, and donors in this region. Robust data for the true burden of malaria throughout pregnancy are scarce. Nevertheless, when women have little immunity, each infection is potentially fatal to the mother, fetus, or both. WHO recommendations for the control of malaria in pregnancy are largely based on the situation in Africa, but strategies in the Asia-Pacific region are complicated by heterogeneous transmission settings, coexistence of multidrug-resistant Plasmodium falciparum and Plasmodium vivax parasites, and different vectors. Most knowledge of the epidemiology, effect, treatment, and prevention of malaria in pregnancy in the Asia-Pacific region comes from India, Papua New Guinea, and Thailand. Improved estimates of the morbidity and mortality of malaria in pregnancy are urgently needed. When malaria in pregnancy cannot be prevented, accurate diagnosis and prompt treatment are needed to avert dangerous symptomatic disease and to reduce effects on fetuses.

Increased serum hepcidin and alterations in blood iron parameters associated with asymptomatic P. falciparum and P. vivax malaria

Background Asymptomatic Plasmodium spp. infections and anemia are highly prevalent conditions in tropical regions. We studied whether asymptomatic parasitemia induces hepcidin- and/or cytokine-mediated iron maldistribution and anemia. Design and Methods A group of 1197 Indonesian schoolchildren, aged 5–15 years, were screened by microscopy for the presence of parasitemia. Concentrations of hemoglobin, serum hepcidin and parameters of iron status and inflammation were determined at baseline and 4 weeks after antimalarial treatment. Results Asymptomatic P. falciparum and P. vivax parasitemia were detected in 73 (6.1%) and 18 (1.5%) children, respectively, of whom 84% and 83% had a C-reactive protein concentration below 5 mg/L. Children with P. falciparum or P. vivax parasitemia had significantly lower hemoglobin concentrations than 17 aparasitemic controls (12.6 and 12.2 g/dL versus 14.4 g/dL; P<0.01), together with significantly higher serum hepcidin concentrations (5.2 and 5.6 nM versus 3.1 nM; P<0.05). The latter was associated with signs of iron maldistribution with higher ferritin concentrations and lower values of serum iron concentration, transferrin saturation and erythrocyte mean cell volume. Concentrations of growth differentiation factor 15 were similar across groups. Antimalarial treatment partly reversed these abnormalities and led to a significant increase in hemoglobin concentration. Conclusions Asymptomatic malarial parasitemia is associated with increased hepcidin concentrations and anemia, in the absence of a manifest acute phase response. Prolonged iron maldistribution may be an underestimated cause of anemia. Screening for parasitemia should be performed before starting iron supplementation, as iron therapy may be less effective and even hazardous in these circumstances.

Functional and genetic evidence that the Mal/TIRAP allele variant 180L has been selected by providing protection against septic shock.

Adequate responses by our innate immune system toward invading pathogens were of vital importance for surviving infections, especially before the antibiotic era. Recently, a polymorphism in Mal (Ser180Leu, TIRAP rs8177374), an important adaptor protein downstream of the Toll-like receptor (TLR) 2 and 4 pathways, has been described to provide protection against a broad range of infectious pathogens. We assessed the functional effects of this polymorphism in human experimental endotoxemia, and we demonstrate that individuals bearing the TIRAP 180L allele display an increased, innate immune response to TLR4 and TLR2 ligands, but not to TLR9 stimulation. This phenotype has been related to an increased resistance to infection. However, an overshoot in the release of proinflammatory cytokines by TIRAP 180L homozygous individuals suggests a scenario of balanced evolution. We have also investigated the worldwide distribution of the Ser180Leu polymorphism in 14 populations around the globe to correlate the genetic makeup of TIRAP with the local infectious pressures. Based on the immunological, clinical, and genetic data, we propose that this mutation might have been selected in West Eurasia during the early settlement of this region after the out-of-Africa migration of modern Homo sapiens. This combination of functional and genetic data provides unique insights to our understanding of the pathogenesis of sepsis.

Mutations in the cytochrome b gene of Plasmodium berghei conferring resistance to atovaquone.

The molecular lesions which underlie the resistance of the malaria parasites to atovaquone, a coenzyme Q analogue, were investigated. Resistant clones of Plasmodium berghei ANKA strain were isolated following prolonged propagation in mice in the presence of increasing doses of the drug, and their cytochrome b gene sequenced. Three mutations were detected, T–C substitution at nt 431, G–A at nt 399 and G–T at nt 850, resulting in amino acid changes in the putative cytochrome b product at residues 133, 144 and 284. The V284F amino acid change is in the sixth transmembrane helix of the protein and was observed in all resistant clones. An additional M133I or L144S amino acid change within the Qo site at an extramembranous amphipathic helix significantly increases the resistance to atovaquone. Our results (a) provide evidence that the antimalarial activity of atovaquone indeed involves an interaction with the cytochrome b; (b) define atovaquone as an inhibitor of the ubiquinol oxidase activity of the cytochrome bc1 complex; and (c) define amino acid residues in the mammalian cytochrome b which might be critical in determining its relative resistance to atovaquone.

Tracing the Tiger: Population Genetics Provides Valuable Insights into the Aedes (Stegomyia) albopictus Invasion of the Australasian Region

Background The range of the Asian tiger mosquito Aedes albopictus is expanding globally, raising the threat of emerging and re-emerging arbovirus transmission risks including dengue and chikungunya. Its detection in Papua New Guineas (PNG) southern Fly River coastal region in 1988 and 1992 placed it 150 km from mainland Australia. However, it was not until 12 years later that it appeared on the Torres Strait Islands. We hypothesized that the extant PNG population expanded into the Torres Straits as an indirect effect of drought-proofing the southern Fly River coastal villages in response to El Nino-driven climate variability in the region (via the rollout of rainwater tanks and water storage containers). Methodology/Principal Findings Examination of the mosquitos mitochondrial DNA cytochrome oxidase I (COI) sequences and 13 novel nuclear microsatellites revealed evidence of substantial intermixing between PNGs southern Fly region and Torres Strait Island populations essentially compromising any island eradication attempts due to potential of reintroduction. However, two genetically distinct populations were identified in this region comprising the historically extant PNG populations and the exotic introduced population. Both COI sequence data and microsatellites showed the introduced population to have genetic affinities to populations from Timor Leste and Jakarta in the Indonesian region. Conclusions/Significance The Ae. albopictus invasion into the Australian region was not a range expansion out of PNG as suspected, but founded by other, genetically distinct population(s), with strong genetic affinities to populations sampled from the Indonesian region. We now suspect that the introduction of Ae. albopictus into the Australian region was driven by widespread illegal fishing activity originating from the Indonesian region during this period. Human sea traffic is apparently shuttling this mosquito between islands in the Torres Strait and the southern PNG mainland and this extensive movement may well compromise Ae. albopictus eradication attempts in this region.

The effect of primaquine on gametocyte development and clearance in the treatment of uncomplicated falciparum malaria with dihydroartemisinin-piperaquine in South Sumatra, Western Indonesia: an open label randomized controlled trial

BACKGROUND Artemisinin-based combination therapy is very effective in clearing asexual stages of malaria and reduces gametocytemia, but may not affect mature gametocytes. Primaquine is the only commercially available drug that eliminates mature gametocytes. METHODS We conducted a 2-arm, open-label, randomized, controlled trial to evaluate the efficacy of single-dose primaquine (0.75 mg/kg) following treatment with dihydroartemisinin-piperaquine (DHP) on Plasmodium falciparum gametocytemia, in Indonesia. Patients aged ≥5 years with uncomplicated falciparum malaria, normal glucose-6-phosphate dehydrogenase enzyme levels, and hemoglobin levels ≥8 g/dL were assigned by computerized-generating sequence to a standard 3-day course of DHP alone (n = 178) or DHP combined with a single dose of primaquine on day 3 (n = 171). Patients were seen on days 1, 2, 3, and 7 and then weekly for 42 days to assess the presence of gametocytes and asexual parasites by microscopy. Survival analysis was stratified by the presence of gametocytes on day 3. RESULTS DHP prevented development of gametocytes in 277 patients without gametocytes on day 3. In the gametocytemic patients (n = 72), primaquine was associated with faster gametocyte clearance (hazard ratio = 2.42 [95% confidence interval, 1.39-4.19], P = .002) and reduced gametocyte densities (P = .018). The day 42 cure rate of asexual stages in the DHP + primaquine and DHP-only arms were: polymerase chain reaction (PCR) unadjusted, 98.7% vs 99.4%, respectively; PCR adjusted, 100% for both. Primaquine was well tolerated. CONCLUSIONS Addition of single-dose 0.75 mg/kg primaquine shortens the infectivity period of DHP-treated patients and should be considered in low-transmission regions that aim to control and ultimately eliminate falciparum malaria. Clinical Trials Registration. NCT01392014.

Seasonal prevalence of malaria in West Sumba district, Indonesia.

BackgroundAccurate information about the burden of malaria infection at the district or provincial level is required both to plan and assess local malaria control efforts. Although many studies of malaria epidemiology, immunology, and drug resistance have been conducted at many sites in Indonesia, there is little published literature describing malaria prevalence at the district, provincial, or national level.MethodsTwo stage cluster sampling malaria prevalence surveys were conducted in the wet season and dry season across West Sumba, Nusa Tenggara Province, Indonesia.ResultsEight thousand eight hundred seventy samples were collected from 45 sub-villages in the surveys. The overall prevalence of malaria infection in the West Sumba District was 6.83% (95% CI, 4.40, 9.26) in the wet season and 4.95% (95% CI, 3.01, 6.90) in the dry. In the wet season Plasmodium falciparum accounted for 70% of infections; in the dry season P. falciparum and Plasmodium vivax were present in equal proportion. Malaria prevalence varied substantially across the district; prevalences in individual sub-villages ranged from 0–34%. The greatest malaria prevalence was in children and teenagers; the geometric mean parasitaemia in infected individuals decreased with age. Malaria infection was clearly associated with decreased haemoglobin concentration in children under 10 years of age, but it is not clear whether this association is causal.ConclusionMalaria is hypoendemic to mesoendemic in West Sumba, Indonesia. The age distribution of parasitaemia suggests that transmission has been stable enough to induce some clinical immunity. These prevalence data will aid the design of future malaria control efforts and will serve as a baseline against which the results of current and future control efforts can be assessed.

Thrombocytopenia in early malaria is associated with GP1b shedding in absence of systemic platelet activation and consumptive coagulopathy.

Thrombocytopenia develops early in malaria, but the underlying mechanisms remain incompletely understood. We studied the aetiology of malaria‐associated thrombocytopenia in volunteers experimentally infected with Plasmodium falciparum malaria, in Indonesian malaria patients and in ex vivo studies. In experimental human malaria, the decrease in platelet counts was associated with a concurrent rise in young platelets (immature platelet fraction) and thrombopoietin. D‐dimer concentrations were moderately elevated without a prolongation in the activated partial thromboplastin time or decrease in fibrinogen. There was no increase in expression of the platelet surface markers CD62P, PAC‐1 and CD63 and in plasma concentrations of the platelet factors P‐selectin, CXCR4, CXCL7, RANTES and CD40L. In contrast, concentrations of soluble glycoprotein‐1b (sGP1b), the external domain of the platelet receptor for von Willebrand factor (VWF), increased early. Indonesian malaria patients also had elevated concentrations of sGP1b, which correlated with VWF concentrations. Finally, incubation of platelets with parasitized erythrocytes in vitro failed to induce platelet aggregation or activation. We concluded that neither compromised platelet production nor platelet activation or consumptive coagulopathy were responsible for the early thrombocytopenia in malaria. We hypothesize that the increase in sGP1b concentrations results from VWF‐mediated GP1b shedding; a process that may prevent excessive adhesion of platelets and parasitized erythrocytes.