The feasibility of a malaria vaccine is supported by the fact that children in endemic areas develop naturally acquired immunity to disease. Development of disease immunity is characterized by a decrease in the frequency and severity of disease episodes over several years despite almost continuous infection 1 , suggesting that immunity may develop through the acquisition of a repertoire of specific, protective antibodies directed against polymorphic target antigens 1-3 . Plasmodium falciparum erythrocyte membrane protein 1 (PfEMP1) is a potentially important family of target antigens, because these proteins are inserted into the red cell surface and are prominently exposed 4-6 and because they are highly polymorphic and undergo clonal antigenic variation 7,8,18 , a mechanism of immune evasion maintained by a large family of var genes [9][10][11] . In a large prospective study of Kenyan children, we have used the fact that anti-PfEMP1 antibodies agglutinate infected erythrocytes in a variant-specific manner 10,[12][13][14][15][16] , to show that the PfEMP1 variants expressed during episodes of clinical malaria were less likely to be recognized by the corresponding child's own preexisting antibody response than by that of children of the same age from the same community. In contrast, a heterologous parasite isolate was just as likely to be recognized. The apparent selective pressure exerted by established anti-PfEMP1 antibodies on infecting parasites supports the idea that such responses provide variant-specific protection against disease.To study protection by anti-PfEMP1 antibodies it was necessary to focus on a group of children who were at various stages of developing a full repertoire of anti-PfEMP1 responses. As the period over which the antibody repertoire develops will vary with local rate of exposure, it was important first to determine the age range to study. To this end, 200 serum samples (40 from each of five age classes: 1-2, 3-4, 5-6, 7-8 and 9-12 years) were taken from children in March 1993. Each was tested for its ability to agglutinate four parasites (C10, A4, W1008 and W1027). As shown in Fig. 1a, the prevalence of antibody responses to all isolates rose between 1 and 5 years of age. This agrees closely with the age range over which naturally acquired immunity to malaria develops in this part of Kenya (data not shown). As expected, responses to the different isolates were independent and specific, since the number of individuals in each age group that recognized between zero and four isolates (Fig. 1b) has a diagonal structure that fits very closely that predicted by a binomial distribution (not shown).© 1998 Nature Publishing Group Correspondence should be addressed to P.C.B.; pbull@africaonline.co.ke.. Defining the role of parasite antigens in naturally acquired immunity has been problematic. The ubiquity of asymptomatic infection in those age groups susceptible to disease combined with the difficulty of determining the immune status of any individual means that relating any particular...

The variant surface antigens expressed on Plasmodium falciparum–infected erythrocytes are potentially important targets of immunity to malaria and are encoded, at least in part, by a family of var genes, about 60 of which are present within every parasite genome. Here we use semi-conserved regions within short var gene sequence “tags” to make direct comparisons of var gene expression in 12 clinical parasite isolates from Kenyan children. A total of 1,746 var clones were sequenced from genomic and cDNA and assigned to one of six sequence groups using specific sequence features. The results show the following. (1) The relative numbers of genomic clones falling in each of the sequence groups was similar between parasite isolates and corresponded well with the numbers of genes found in the genome of a single, fully sequenced parasite isolate. In contrast, the relative numbers of cDNA clones falling in each group varied considerably between isolates. (2) Expression of sequences belonging to a relatively conserved group was negatively associated with the repertoire of variant surface antigen antibodies carried by the infected child at the time of disease, whereas expression of sequences belonging to another group was associated with the parasite “rosetting” phenotype, a well established virulence determinant. Our results suggest that information on the state of the host–parasite relationship in vivo can be provided by measurements of the differential expression of different var groups, and need only be defined by short stretches of sequence data.

The present data are useful in that they confirm the mechanisms by which HbAS confers protection against malaria and shed light on the relationships between HbAS, malaria, and other childhood diseases.

The variant surface antigens (VSAs) of Plasmodium falciparum-infected red blood cells are potentially important targets of naturally acquired immunity to malaria. Natural infections induce agglutinating antibodies specific to the VSA variants expressed by the infecting parasites. Previously, when different parasite isolates were tested against a panel of heterologous plasma from Kenyan children, the proportion of plasma that agglutinated the parasites (the agglutination frequency [AF]) was highly variable among isolates, suggesting the existence of rare and prevalent variants. Here, the AF of 115 isolates from Kenyan children were compared. The results show that the AF of isolates causing severe malaria were significantly higher than those of isolates causing mild malaria; and AF decreased significantly with the increasing age of the infected child. We propose that parasites causing severe disease tend to express a subset of VSA variants that are preferentially associated with infections of children with low immunity.

BackgroundMalaria resistance by the sickle cell trait (genotype HbAS) has served as the prime example of genetic selection for over half a century. Nevertheless, the mechanism of this resistance remains the subject of considerable debate. While it probably involves innate factors such as the reduced ability of Plasmodium falciparum parasites to grow and multiply in HbAS erythrocytes, recent observations suggest that it might also involve the accelerated acquisition of malaria-specific immunity.Methods and FindingsWe studied the age-specific protection afforded by HbAS against clinical malaria in children living on the coast of Kenya. We found that protection increased with age from only 20% in the first 2 y of life to a maximum of 56% by the age of 10 y, returning thereafter to 30% in participants greater than 10 y old.ConclusionsOur observations suggest that malaria protection by HbAS involves the enhancement of not only innate but also of acquired immunity to the parasite. A better understanding of the underlying mechanisms might yield important insights into both these processes.

SummaryPlasmodium falciparum erythrocyte membrane protein 1 (PfEMP1) is a potentially important family of immune targets, encoded by an extremely diverse gene family called var. Understanding of the genetic organization of var genes is hampered by sequence mosaicism that results from a long history of nonhomologous recombination. Here we have used software designed to analyse social networks to visualize the relationships between large collections of short var sequences tags sampled from clinical parasite isolates. In this approach, two sequences are connected if they share one or more highly polymorphic sequence blocks. The results show that the majority of analysed sequences including several var-like sequences from the chimpanzee parasite Plasmodium reichenowi can be either directly or indirectly linked together in a single unbroken network. However, the network is highly structured and contains putative subgroups of recombining sequences. The major subgroup contains the previously described group A var genes, previously proposed to be genetically distinct. Another subgroup contains sequences found to be associated with rosetting, a parasite virulence phenotype. The mosaic structure of the sequences and their division into subgroups may reflect the conflicting problems of maximizing antigenic diversity and minimizing epitope sharing between variants while maintaining their host cell binding functions.

BackgroundThe α-thalassaemias are the commonest genetic disorders of humans. It is generally believed that this high frequency reflects selection through a survival advantage against death from malaria; nevertheless, the epidemiological description of the relationships between α-thalassaemia, malaria, and other common causes of child mortality remains incomplete.Methods and FindingsWe studied the α +-thalassaemia-specific incidence of malaria and other common childhood diseases in two cohorts of children living on the coast of Kenya. We found no associations between α +-thalassaemia and the prevalence of symptomless Plasmodium falciparum parasitaemia, the incidence of uncomplicated P. falciparum disease, or parasite densities during mild or severe malaria episodes. However, we found significant negative associations between α +-thalassaemia and the incidence rates of severe malaria and severe anaemia (haemoglobin concentration < 50 g/l). The strongest associations were for severe malaria anaemia (> 10,000 P. falciparum parasites/μl) and severe nonmalaria anaemia; the incidence rate ratios and 95% confidence intervals (CIs) for α +-thalassaemia heterozygotes and homozygotes combined compared to normal children were, for severe malaria anaemia, 0.33 (95% CI, 0.15,0.73; p = 0.006), and for severe nonmalaria anaemia, 0.26 (95% CI, 0.09,0.77; p = 0.015). ConclusionsOur observations suggest, first that selection for α +-thalassaemia might be mediated by a specific effect against severe anaemia, an observation that may lead to fresh insights into the aetiology of this important condition. Second, although α +-thalassaemia is strongly protective against severe and fatal malaria, its effects are not detectable at the level of any other malaria outcome; this result provides a cautionary example for studies aimed at testing malaria interventions or identifying new malaria-protective genes.

BackgroundThe reservoir of Plasmodium infection in humans has traditionally been defined by blood slide positivity. This study was designed to characterize the local reservoir of infection in relation to the diverse var genes that encode the major surface antigen of Plasmodium falciparum blood stages and underlie the parasite's ability to establish chronic infection and transmit from human to mosquito.Methodology/Principal FindingsWe investigated the molecular epidemiology of the var multigene family at local sites in Gabon, Senegal and Kenya which differ in parasite prevalence and transmission intensity. 1839 distinct var gene types were defined by sequencing DBLα domains in the three sites. Only 76 (4.1%) var types were found in more than one population indicating spatial heterogeneity in var types across the African continent. The majority of var types appeared only once in the population sample. Non-parametric statistical estimators predict in each population at minimum five to seven thousand distinct var types. Similar diversity of var types was seen in sites with different parasite prevalences.Conclusions/Significance Var population genomics provides new insights into the epidemiology of P. falciparum in Africa where malaria has never been conquered. In particular, we have described the extensive reservoir of infection in local African sites and discovered a unique var population structure that can facilitate superinfection through minimal overlap in var repertoires among parasite genomes. Our findings show that var typing as a molecular surveillance system defines the extent of genetic complexity in the reservoir of infection to complement measures of malaria prevalence. The observed small scale spatial diversity of var genes suggests that var genetics could greatly inform current malaria mapping approaches and predict complex malaria population dynamics due to the import of var types to areas where no widespread pre-existing immunity in the population exists.