To study the pathogenesis of fatal cerebral malaria, we conducted autopsies in 31 children with this clinical diagnosis. We found that 23% of the children had actually died from other causes. The remaining patients had parasites sequestered in cerebral capillaries, and 75% of those had additional intra- and perivascular pathology. Retinopathy was the only clinical sign distinguishing malarial from nonmalarial coma. These data have implications for treating malaria patients, designing clinical trials and assessing malaria-specific disease associations.

Whereas most nontyphoidal Salmonella (NTS) are associated with gastroenteritis, there has been a dramatic increase in reports of NTS-associated invasive disease in sub-Saharan Africa. Salmonella enterica serovar Typhimurium isolates are responsible for a significant proportion of the reported invasive NTS in this region. Multilocus sequence analysis of invasive S. Typhimurium from Malawi and Kenya identified a dominant type, designated ST313, which currently is rarely reported outside of Africa. Whole-genome sequencing of a multiple drug resistant (MDR) ST313 NTS isolate, D23580, identified a distinct prophage repertoire and a composite genetic element encoding MDR genes located on a virulence-associated plasmid. Further, there was evidence of genome degradation, including pseudogene formation and chromosomal deletions, when compared with other S. Typhimurium genome sequences. Some of this genome degradation involved genes previously implicated in virulence of S. Typhimurium or genes for which the orthologs in S. Typhi are either pseudogenes or are absent. Genome analysis of other epidemic ST313 isolates from Malawi and Kenya provided evidence for microevolution and clonal replacement in the field.

Molecular assays for monitoring sulfadoxine-pyrimethamine-resistant Plasmodium falciparum have not been implemented because of the genetic and statistical complexity of the parasite mutations that confer resistance and their relation to treatment outcomes. This study analyzed pretreatment dihydrofolate reductase (DHFR) and dihydropteroate synthase (DHPS) genotypes and treatment outcomes in a double-blind, placebo-controlled trial of sulfadoxine-pyrimethamine and chlorproguanil-dapsone treatment for uncomplicated P. falciparum malaria. Multiple logistic regression was used to identify mutations that were predictive of treatment failure and to identify interactions and confounding factors. Infections caused by parasites with 3 DHFR mutations and 2 DHPS mutations (the "quintuple mutant") were associated with sulfadoxine-pyrimethamine treatment failure but not with chlorproguanil-dapsone treatment failure. The presence of a single DHFR mutation (Arg-59) with a single DHPS mutation (Glu-540) accurately predicted the presence of the quintuple mutant. If this model is validated in other populations, it will finally be possible to use molecular markers for surveillance of antifolate-resistant P. falciparum malaria in Africa.

To investigate the relationship of TNFα levels to Plasmodium falciparum (PF) infection, plasma TNFα concentrations were measured in Pakistani adults and children with mild, severe, cerebral and chronic falciparum malaria and healthy (control) subjects. The initial geometric mean plasma concentrations of TNFα in adult patients with severe malaria (187.6 pg/mL) were significantly higher than mild malaria patients (87.1 pg/mL, P < 0.001). TNFα levels were not correlated to parasite density, cerebral malaria, young age, hypoglycemia or fatal outcome; however, they were associated with severe anemia, and hepatic and kidney dysfunction. TNFα levels were not significantly increased in 16 patients with hyperparasitemia and were significantly elevated (P < 0.02) in chronic malaria patients as compared to control subjects. TNFα levels were elevated independently in patients with anemia, hypoglycemia (P < 0.001, P < 0.05), and hepatic and kidney (P < 0.001 each) dysfunction. In this study, high TNFα levels were associated with several manifestations of severe malaria and were not specific to cerebral malaria and hyperparasitemia.

There are multiple causes of severe anemia in Malawian preschool children, but folate and iron deficiencies are not prominent among them. Even in the presence of malaria parasites, additional or alternative causes of severe anemia should be considered.

We report a genome-wide association (GWA) study of severe malaria in The Gambia. The initial GWA scan included 2,500 children genotyped on the Affymetrix 500K GeneChip, and a replication study included 3,400 children. We used this to examine the performance of GWA methods in Africa. We found considerable population stratification, and also that signals of association at known malaria resistance loci were greatly attenuated owing to weak linkage disequilibrium (LD). To investigate possible solutions to the problem of low LD, we focused on the HbS locus, sequencing this region of the genome in 62 Gambian individuals and then using these data to conduct multipoint imputation in the GWA samples. This increased the signal of association, from P = 4 × 10 −7 to P = 4 × 10 −14 , with the peak of the signal located precisely at the HbS causal variant. Our findings provide proof of principle that fine-resolution multipoint imputation, based on population-specific sequencing data, can substantially boost authentic GWA signals and enable fine mapping of causal variants in African populations.The malaria parasite Plasmodium falciparum kills on the order of a million African children each year 1 , and this is a small fraction of the number of infected individuals in the population [1][2][3] . In communities where everyone is repeatedly infected with P. falciparum, host genetic factors account for ~25% of the risk of severe malaria, that is, life-threatening forms of the disease 3 . The strongest known determinant of risk, hemoglobin S (HbS), accounts for 2% of the total variation, implying that only a small fraction of genetic resistance factors have so far been discovered 3 . Identifying the genetic basis of protective immunity against severe malaria may provide important insights for vaccine development.Here we examine the possibility of approaching this problem by genome-wide association (GWA) analysis. There are many unsolved methodological questions about how to conduct an effective GWA study in Africa 4 . High levels of ethnic diversity may result in false-positive associations owing to population structure. Variations in haplotype structure between different ethnic groups may reduce power to detect GWA signals, particularly when data are amalgamated across multiple study sites. Low LD implies the need for denser genotyping arrays than are currently available: a crude estimate is that an African GWA study with 1.5 million SNPs would have approximately the same statistical power as a European study with Jallow et al.Page 2Nat Genet. Author manuscript; available in PMC 2010 September 21. NIH-PA Author ManuscriptNIH-PA Author Manuscript NIH-PA Author Manuscript 0.6 million SNPs5, but this is based on HapMap data from a single ethnic group and a larger number of SNPs may be needed to achieve adequate power across different ethnic groups.We carried out an initial GWA study in Gambian children that explores these methodological questions. Genotyping of ~500,000 SNPs was conducted on 1,060 cases of severe malaria and 1...

Nontyphoidal strains of Salmonella (NTS) are a common cause of bacteremia among African children. Cellmediated immune responses control intracellular infection, but they do not protect against extracellular growth of NTS in the blood. We investigated whether antibody protects against NTS bacteremia in Malawian children, because we found this condition mainly occurs before 2 years of age, with relative sparing of infants younger than 4 months old. Sera from all healthy Malawian children tested aged more than 16 months contained anti-Salmonella antibody and successfully killed NTS. Killing was mediated by complement membrane attack complex and not augmented in the presence of blood leukocytes. Sera from most healthy children less than 16 months old lacked NTS-specific antibody, and sera lacking antibody did not kill NTS despite normal complement function. Addition of Salmonella-specific antibody, but not mannose-binding lectin, enabled NTS killing. All NTS strains tested had long-chain lipopolysaccharide and the rck gene, features that resist direct complement-mediated killing. Disruption of lipopolysaccharide biosynthesis enabled killing of NTS by serum lacking Salmonella-specific antibody. We conclude that Salmonella-specific antibody that overcomes the complement resistance of NTS develops by 2 years of life in Malawian children. This finding and the age-incidence of NTS bacteremia suggest that antibody protects against NTS bacteremia and support the development of vaccines against NTS that induce protective antibody.

BACKGROUND Case fatality rates among African children with cerebral malaria remain in the range of 15 to 25%. The key pathogenetic processes and causes of death are unknown, but a combination of clinical observations and pathological findings suggests that increased brain volume leading to raised intracranial pressure may play a role. Magnetic resonance imaging (MRI) became available in Malawi in 2009, and we used it to investigate the role of brain swelling in the pathogenesis of fatal cerebral malaria in African children. METHODS We enrolled children who met a stringent definition of cerebral malaria (one that included the presence of retinopathy), characterized them in detail clinically, and obtained MRI scans on admission and daily thereafter while coma persisted. RESULTS Of 348 children admitted with cerebral malaria (as defined by the World Health Organization), 168 met the inclusion criteria, underwent all investigations, and were included in the analysis. A total of 25 children (15%) died, 21 of whom (84%) had evidence of severe brain swelling on MRI at admission. In contrast, evidence of severe brain swelling was seen on MRI in 39 of 143 survivors (27%). Serial MRI scans showed evidence of decreasing brain volume in the survivors who had had brain swelling initially. CONCLUSIONS Increased brain volume was seen in children who died from cerebral malaria but was uncommon in those who did not die from the disease, a finding that suggests that raised intracranial pressure may contribute to a fatal outcome. The natural history indicates that increased intracranial pressure is transient in survivors. (Funded by the National Institutes of Health and Wellcome Trust U.K.)