High residual chloroquine blood levels in African children with severe malaria seeking healthcare.

Despite widespread resistance, chloroquine remains widely used in West Africa, particularly in home treatment. We examined chloroquine blood levels on admission to a referral hospital with respect to the manifestation of severe malaria in 290 Ghanaian children. Of the patients, 78% exhibited chloroquine concentrations (subtherapeutic, 35%; therapeutic, 37%; supratherapeutic, 6%) and 11% died. Most parasites (78%) carried the pfcrt-T76 chloroquine resistance mutation. High drug concentrations correlated with reduced parasitaemia but also with selection of resistant parasites, lower respiratory and heart rates, increased plasma lactate levels and impaired consciousness. Geometric mean chloroquine concentrations tended to be higher in children who died than in survivors (1.135 vs. 778nmol/l; P=0.09). Supratherapeutic drug levels (>5000nmol/l) were associated with fatal outcome (odds ratio 8.6; 95% CI 1.4-51.7). Residual chloroquine concentrations were found to be abundant in children with severe malaria and to be associated with alterations in the clinical manifestation of the disease and its case fatality. This may result from toxic effects of the drug and/or reflect preceding overtreatment in children with acute life-threatening disease. In areas of intense chloroquine resistance and frequent pre-treatment, additional administration of chloroquine at hospital admission is not only ineffective but may even further endanger patients.

Age-dependent effect of plasma nitric oxide on parasite density in Ghanaian children with severe malaria.

Nitric oxide (NO) has toxic properties against Plasmodium falciparum. While high blood levels have been associated with protection against severe malarial disease, they may also contribute to the pathophysiology of cerebral malaria and severe anaemia. Promoter variants in the inducible nitric oxide synthase (iNOS) gene have been shown to influence NO concentrations and disease manifestation. However, findings are conflicting. We examined associations of plasma NO metabolites (NOx) with symptoms of severe malaria, particularly malarial anaemia and cerebral malaria, and with iNOS promoter variants. In 210 Ghanaian children with severe malaria, we measured plasma nitrite, nitrate, and S-nitrosothiol, and genotyped the iNOS promoter variants -954G-->C, -1173C-->T, and the -2.5 kb (CCTTT)(n) microsatellite. NOx levels decreased with age. In young children (<24 months), high NOx was associated with reduced parasite density. This was not seen in patients of 24-48 months of age and reversed in older children. Subgroup analysis revealed that in children with severe anaemia but without cerebral involvement (prostration, impaired consciousness, convulsions), high NOx levels correlated with low parasitaemia (P = 0.02). In these children, elevated NOx levels were also associated with the iNOS-954C-->T/(CCTTT)(8) haplotype (P = 0.03). No association between NOx or iNOS genotypes and cerebral malaria was observed. Our findings suggest that in young children with severe malaria NOx reduces parasitaemia. This effect wanes at higher ages and may reflect a predominance of unspecific immune responses to infection in early childhood. This finding may have importance for the understanding of associations between iNOS variants and severe malaria in regions of differing disease manifestation.

iNOS promoter variants and severe malaria in Ghanaian children.

Nitric oxide is an important mediator in the host defence against Plasmodium falciparum malaria. It has antiparasitic effects in vitro. However, its role in clinical disease remains controversial. Polymorphisms in the inducible nitric oxide synthase promoter (iNOS; -954G-->C, -1173C-->T, -2.6 kb CCTTT(n) microsatellite) may influence susceptibility to and severity of malaria. We tested this hypothesis in a case-control study among Ghanaian children with severe malaria (SM) and asymptomatic parasitaemia, respectively, and in healthy controls. In this study, the respective frequencies of iNOS-954G-->C and -1173C-->T did not differ between groups but > or =13 microsatellite copies were associated with SM. -954G-->C and -1173C-->T were in linkage disequilibrium with CCTTT(8) and CCTTT(13), respectively. -954G-->C/CCTTT(8) protected against hyperparasitaemia whereas -1173C-->T/CCTTT(13) increased fatality. These findings suggest that iNOS promoter haplotypes rather than single nucleotide polymorphisms are associated with malaria in Ghanaian children.

Manifestation and outcome of severe malaria in children in northern Ghana.

The symptoms of severe malaria and their contribution to mortality were assessed in 290 children in northern Ghana. Common symptoms were severe anemia (55%), prostration (33%), respiratory distress (23%), convulsions (20%), and impaired consciousness (19%). Age influenced this pattern. The fatality rate was 11.2%. In multivariate analysis, circulatory collapse, impaired consciousness, hypoglycemia, and malnutrition independently predicted death. Children with severe malaria by the current World Health Organization (WHO) classification, but not by the previous one (1990), showed relatively mild clinical manifestations and a low case fatality rate (3.2%). In hospitalized children with severe malaria in northern Ghana, severe anemia is the leading manifestation, but itself does not contribute to mortality. In this region, malnutrition and circulatory collapse were important predictors of fatal malaria. The current WHO criteria serve well in identifying life-threatening disease, but also include rather mild cases that may complicate the allocation of immediate care in settings with limited resources.

Alpha(+)-thalassemia protects African children from severe malaria.

The high frequency of alpha(+)-thalassemia in malaria-endemic regions may reflect natural selection due to protection from potentially fatal severe malaria. In Africa, bearing 90% of global malaria morbidity and mortality, this has not yet been observed. We tested this hypothesis in an unmatched case-control study among 301 Ghanaian children with severe malaria and 2107 controls (62% parasitemic). In control children, alpha(+)-thalassemia affected neither prevalence nor density of Plasmodium falciparum. However, heterozygous alpha(+)-thalassemia was observed in 32.6% of controls but in only 26.2% of cases (odds ratio [OR], 0.74; 95% confidence interval [CI], 0.56-0.98). Protection against severe malaria was found to be pronounced comparing severe malaria patients with parasitemic controls (adjusted OR in children < 5 years of age, 0.52; 95% CI, 0.34-0.78) and to wane with age. No protective effect was discernible for homozygous children. Our findings provide evidence for natural selection of alpha(+)-thalassemia in Africa due to protection from severe malaria.