Malaria merozoite surface protein 1 (MSP1) is cleaved in an essential step during erythrocyte invasion. The responses of children to natural malaria infection included antibodies that inhibit this cleavage and others that block the binding of these inhibitory antibodies. There was no correlation between the titer of the antibody to the 19-kDa fragment of MSP1 and its inhibitory activity. These findings have implications for the design of MSP1-based vaccines.

Thrombocytopaenia, or platelet aggregation, is a serious complication of African trypanosomiasis. The biochemical basis is not clearly known. Proteases are known potent inducers of blood coagulation and platelet aggregation, and unknown factors released by Trypanosoma brucei have been shown to induce platelet aggregation. In attempts to define the biochemical mechanisms involved in thrombocytopaenia we purified and characterised a major proteolytic enzyme released extracellularly by T. brucei. Actively motile trypanosomes released proteins into the medium (phosphate saline/glucose, pH 8.0) in which the organisms were incubated in vitro. The M(r) of the released polypeptides ranged from 15 to > 200 kDa, amongst which are proteases. One of the major protein bands, a 250 kDa protease, was purified to homogeneity by ammonium sulfate precipitation followed by diethylaminoethyl (DEAE)-cellulose chromatography and Sephacryl S-300 gel filtration. The protease migrated as a single band of 63 kDa upon electrophoresis in both denaturing and non-denaturing gel co-polymerised with gelatin. The enzyme was strongly active against Z-ARR-AFC peptide substrate, with a pH optimum of 7.0. The proteolytic activity was enhanced by dithiothreitol and inhibited by E-64, leupeptin, TPCK and antipain. The released proteolytic enzyme is putatively identified as a cathepsin B-like cysteine protease.

BackgroundPlasmodium falciparum EBA175 and PfRh2 belong to two main families involved in parasite invasion, and both are potential vaccine candidates. Current knowledge is limited regarding which target antigens and subclasses of antibodies are actually important for protection, and how naturally acquired immunity is achieved.MethodsRepeated blood samples were collected from individuals in Nigeria over a period of almost one year. ELISA was used to analyse subclasses of IgG responses.ResultsFor both EBA175 (region III-V) and (a fragment of) PfRh2, the dominant antibody responses consisted of IgG1 and IgG3 followed by IgG2, while for PfRh2 there was also a relatively prominent response for IgG4. High levels of IgG1, IgG2 and IgG3 for EBA175 and total IgG for PfRh2 correlated significantly with a lower parasitaemia during the study period. Children with HbAS had higher levels of some subclasses compared to children with HbAA, while in adults the pattern was the opposite. The half-lives of IgG2 and IgG4 against EBA175 were clearly shorter than those for IgG1 and IgG3.ConclusionEBA175 and PfRh2 are potential targets for protective antibodies since both correlated with lower parasitaemia. The shorter half-lives for IgG2 and IgG4 might explain why these subclasses are often considered less important in protection against malaria. Triggering the right subclass responses could be of critical importance in a successful vaccine. Further studies are needed to evaluate the role of haemoglobin polymorphisms and their malaria protective effects in this process.

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Understanding the functional characteristics of naturally acquired antibodies against P. falciparum merozoite antigens is crucial for determining the protective functions of antibodies. Affinity (measured as kd) of naturally acquired antibodies against two key targets of acquired immunity, EBA175 and PfRh2, was determined using Surface Plasmon Resonance (SPR) in a longitudinal survey in Nigeria. A majority of the participants, 79% and 67%, maintained stable antibody affinities to EBA175 and PfRh2, respectively, over time. In about 10% of the individuals, there was a reciprocal interaction with a reduction over time in antibody affinity for PfRh2 and an increase for EBA175. In general, PfRh2 elicited antibodies with higher affinity compared to EBA175. Individuals with higher exposure to malaria produced antibodies with higher affinity to both antigens. Younger individuals (5–15 years) produced comparable or higher affinity antibodies than adults (>15 years) against EBA175, but not for PfRh2. Correlation between total IgG (ELISA) and affinity varied between individuals, but PfRh2 elicited antibodies with a higher correlation in a majority of the participants. There was also a correlation between antibody inhibition of erythrocyte invasion by merozoites and PfRh2 affinity. This work gives new insights into the generation and maintenance of antibody affinity over time.