Inhibitory antibodies specific for the 19-kilodalton fragment of merozoite surface protein 1 do not correlate with delayed appearance of infection with Plasmodium falciparum in semi-immune individuals in Vietnam.

Inhibitory antibodies specific for the 19-kDa fragment of merozoite surface protein 1 (MSP1(19)) are a significant component of inhibitory responses in individuals immune to malaria. Nevertheless, conflicting results have been obtained in determining whether this antibody specificity correlates with protection in residents of areas where malaria is endemic. In this study, we examined sera collected from a population of semi-immune individuals living in an area of Vietnam with meso-endemicity during a 6-month period. We used two Plasmodium falciparum parasite lines that express either endogenous MSP1(19) or the homologous region from Plasmodium yoelii to measure the MSP1(19)-specific inhibitory activity. We showed that (i) the level of MSP1(19)-specific inhibitory antibodies was not associated with a delay in P. falciparum infection, (ii) MSP1(19)-specific inhibitory antibodies declined significantly during the convalescent period after infection, and (iii) there was no significant correlation between the MSP1(19)-specific inhibitory antibodies and the total antibodies measured by enzyme-linked immunosorbent assay. These results have implications for understanding naturally acquired immunity to malaria and for the development and evaluation of MSP1(19)-based vaccines.

Associations between responses to the rhoptry-associated membrane antigen of Plasmodium falciparum and immunity to malaria infection.

Rhoptry proteins participate in the invasion of red blood cells by merozoites during the malaria parasite's asexual-stage cycle. Interference with the rhoptry protein function has been shown to prevent invasion, and three rhoptry proteins have been suggested as potential components of a vaccine against malaria. Rhoptry-associated membrane antigen (RAMA) is a 170-kDa protein of Plasmodium falciparum which is processed to a 60-kDa mature form in the rhoptries. p60/RAMA is discharged from rhoptries of free merozoites and binds to the red-cell membrane before being internalized to form part of the parasitophorous vacuole of the newly developing ring. We examined the range of anti-RAMA responses in individuals living in an area of endemicity for malaria and determined its association with clinical immunity. RAMA is immunogenic during infections, and at least three epitopes within RAMA are recognized by hyperimmune sera in immunoblots. Sera from individuals living in a region of Vietnam where malaria is endemic possessed strong antibody responses toward two C-terminal regions of RAMA. Cytophilic antibody isotypes (immunoglobulin G1 [IgG1] and IgG3) predominated in humoral responses to both C-terminal epitopes. Acute episodes of P. falciparum infection result in significant boosting of levels of antibody to an epitope at the extreme C terminus of RAMA that harbors the red-cell-binding domain. Immunity to P. falciparum infection was linked to elevated levels of IgG3 responses to this functional domain of RAMA, suggesting that the region may contain a protective epitope useful for inclusion in a multiepitope vaccine against malaria.

Naturally acquired antibody responses to the components of the Plasmodium falciparum merozoite surface protein 1 complex.

Merozoite surface protein 6 (MSP6) and 7 (MSP7) of Plasmodium falciparum are peripheral membrane proteins whose cleaved products, MSP636, MSP722 and MSP719, are found on the merozoite surface as components of a non-covalently bound complex which also contains four polypeptides derived from merozoite surface protein 1 (MSP1). We have expressed both the precursor regions and the processed mature products of MSP6 and MSP7 in Escherichia coli and showed that these recombinant proteins react with human immune sera. In a set of sera collected from individuals living in malaria-endemic areas of Southern-central Vietnam, antibodies to the mature polypeptides of MSP636 and MSP722 were detected in 50.6 and 85.6% of the serum samples, whereas antibodies to the precursor regions of MSP6 and MSP7 were detected in only 12.1 and 42.5% of the serum samples, respectively. The predominant subclass of anti-MSP6 antibodies was IgG1, whereas the predominant subclass of anti-MSP7 antibodies was IgG3. In the same set of serum samples, the antibody responses to MSP119 are predominantly IgGI, whereas antibodies to merozoite surface protein 4 (MSP4) are mainly IgG3. This data is consistent with the proposition that, during malaria infection, variable proteins induce responses that are predominantly of the IgG3 isotype, and conserved proteins induce responses that are predominantly IgG1. The antibodies to MSP6, MSP7 and MSP119 all decreased at the time of infection, but increased during the convalescent period. No correlation was observed between the antibodies at the commencement of the study and absence of parasitaemia during surveillance in this population.

Comparison of three antigen detection methods for diagnosis and therapeutic monitoring of malaria: a field study from southern Vietnam.

OBJECTIVES: To compare the sensitivity, specificity and post-treatment persistence of three commonly used rapid antigen detection methods. METHOD: We studied 252 Vietnamese patients aged from 4 to 60 years, 157 with falciparum and 95 with vivax malaria and 160 healthy volunteers. An initial blood sample was taken for microscopy, and OptiMAL, immunochromatographic test (ICT) malaria P.f./P.v. and Paracheck-Pf tests. Patients with falciparum malaria were treated with an artesunate-based combination regimen and those with vivax malaria received chloroquine. Eighty-seven patients with falciparum malaria who were initially positive for one of the antigen tests and who remained blood smear-negative underwent follow-up testing over 28 days. RESULTS: Paracheck-Pf was the most sensitive test for Plasmodium falciparum (95.8% vs. 82.6% for ICT malaria P.f./P.v. and 49.7% for OptiMAL). Specificities were all 100%. For vivax malaria, OptiMAL performed better than ICT malaria P.f./P.v. (sensitivities 73.7% and 20.0%, respectively), with 100% specificity in both cases. All tests had low sensitivities (< or = 75.0%) at parasitaemias < 1000/microl regardless of malaria species. During follow-up, Paracheck-Pf remained positive in the greatest proportion of patients, especially at higher parasitaemias (> 10,000/microl). Residual OptiMAL positivity occurred only in a relatively small proportion of patients (< 10%) with parasitaemias > 10,000/microl during the first 2 weeks after treatment. CONCLUSIONS: Although microscopy remains the gold standard for malaria diagnosis, Paracheck-Pf may prove a useful adjunctive test in uncomplicated falciparum malaria in southern Vietnam. OptiMAL had the lowest sensitivity for P. falciparum but it might have a use in the diagnosis of vivax malaria and perhaps to monitor efficacy of treatment for falciparum malaria where microscopy is unavailable.

The effect of 10 alpha-trifluoromethylhydroartemisinin on Plasmodium berghei infection and its toxicity in experimental animals.

The antimalarial activity of 10 alpha-trifluoromethylhydroartemisinin (TFMHA) was compared to that of dihydroartemisinin (DHA) in the Plamodium berghei mouse model. Treatment with TFMHA in mice infected with a P. berghei chloroquine-sensitive strain at 25 mg/kg for 3, 5, and 7 d, or DHA at the same dose for 7 d showed the parasite was eliminated from the host within 2.6 d. The radical cure and survival rates of these mice up to 60 d after infection were 90-100%. In mice infected with the P. berghei chloroquine-resistant strain, TFMHA used at 25 mg/kg/day for 3, 5, or 7 d reduced parasitaemia within 2 d. The radical cure and survival rates of these animals up to 60 d after infection were 30, 60, and 90% for the 3 treatment durations respectively. In contrast, DHA only had an inhibitory effect on the growth of the parasite within the first few days of treatment and could not eliminate the parasite even after 7 d of treatment. There was a 100% relapse and all mice died within 28 d after infection. The acute toxicity of TFMHA as determined by the median lethal dose (LD50) in mice treated orally was 820 mg/kg. In rabbits, TFMHA given orally at 20 mg/kg once daily for 28 successive days had no effect on the bodyweight, haematological, biochemical, histopathological and electrocardiogram parameters. The results showed that TFMHA is an effective antimalarial drug with a low level of toxicity.