Boosting of transmission-blocking immunity during natural Plasmodium vivax infections in humans depends upon frequent reinfection.

The infectivity to mosquitoes of 31 acute Plasmodium vivax patients was measured by permitting mosquitoes to feed directly on the patients. The infectivity of these patients correlated closely with titers of antibodies in their serum as measured by indirect immunofluorescence against air-dried female gametes of P. vivax. Infectivity by direct feeding was also closely parallel to the transmission-blocking activity of the sera of patients as measured by the suppression of infectivity of parasitized blood by autologous serum relative to normal (nonmalarial) human serum when fed to mosquitoes through a membrane. These results are consistent with serum antibodies in human P. vivax infections as major factors determining the infectivity of an infected individual to mosquitoes. It was further noted that individuals having a second attack of P. vivax within less than 4 months were considerably less infectious to mosquitoes than first-attack patients were. This "boosting" of transmission-blocking immunity was much less if longer intervals intervened between attacks. We discuss the immunological implications and possible epidemiological significance of this short-term boosting of transmission-blocking immunity by successive P. vivax infections.     

Plasmodium falciparum: natural and experimental transmission-blocking immunity

Fc-dependent regulation of humoral immune responses was investigated by immunization of BALB/c mice with immune complexes. These complexes were composed of DNP- and PC-conjugated KLH or Ficoll, and monoclonal T15 idiotype-positive anti-PC antibodies of different isotypes but indistinguishable V-region properties. Since the response to DNP was analyzed, effects due to masking of antigenic determinants by anti-PC antibodies are excluded. The responses to free and complexed antigens showed significant differences in the proportions of DNP-specific IgM, IgG1, IgG2 and IgG3 antibodies. Complexes containing the T-dependent carrier KLH elicited serum antibodies to DNP with significantly decreased IgM levels, irrespective of the isotype in the complex. Under these conditions, however, DNP-specific IgG classes were augmented to various extents, depending on the isotype in the complex. In the case of the T-independent carrier Ficoll, only complexes with IgM or IgG3 suppressed the IgM response. Moreover, immunization with complexes composed of IgM or IgG2a led to a significant decrease of DNP-specific IgG1. In contrast to changes induced by antibodies bound to the T-dependent antigen, immunization with T-independent complexes did not enhance the production of any of the immunoglobulin isotypes.     

Antibody response of humans to the circumsporozoite protein of Plasmodium vivax.

We studied the interaction of sera from residents of an area in northern Peru where vivax malaria is endemic with four recombinant DNA-derived circumsporozoite (CS) proteins of Plasmodium vivax. The antigens used in the enzyme-linked immunosorbent assay included one Escherichia coli-produced and three Saccharomyces cerevisiae-produced recombinant proteins. Three of the proteins (NS1(81)V20, Vivax-1, and Vivax-2) contain the entire central repeat region of the P. vivax CS protein, and one protein (Vivax-3) contains only two repeat sequences. Vivax-1, Vivax-2, and Vivax-3 contain different lengths of sequences flanking the repeats. A higher percentage of the sera had antibodies to Vivax-2 and Vivax-3, the two proteins containing the longest nonrepeat sequences, than to NS1(81)V20 or Vivax-1. Children less than 5 years of age did not have immunoglobulin G antibodies to NS1(81)V20; however, they had antibodies to Vivax-1, Vivax-2, and Vivax-3. The finding that individuals living in a malaria-endemic area produce antibodies to peptides containing nonrepeat regions of the CS protein emphasizes the need to characterize the immune response to these regions in naturally exposed and experimentally immunized humans.     

Evidence and Implications of Mortality Associated with Acute Plasmodium vivax Malaria

Vivax malaria threatens patients despite relatively low-grade parasitemias in peripheral blood. The tenet of death as a rare outcome, derived from antiquated and flawed clinical classifications, disregarded key clinical evidence, including (i) high rates of mortality in neurosyphilis patients treated with vivax malaria; (ii) significant mortality from zones of endemicity; and (iii) the physiological threat inherent in repeated, very severe paroxysms in any patient, healthy or otherwise. The very well-documented course of this infection, with the exception of parasitemia, carries all of the attributes of “perniciousness” historically linked to falciparum malaria, including severe disease and fatal outcomes. A systematic analysis of the parasite biomass in severely ill patients that includes blood, marrow, and spleen may ultimately explain this historic misunderstanding. Regardless of how this parasite is pernicious, recent data demonstrate that the infection comes with a significant burden of morbidity and associated mortality. The extraordinary burden of malaria is not heavily weighted upon any single continent by a single species of parasite—it is a complex problem for the entire endemic world, and both species are of fundamental importance. Humanity must rally substantial resources, intellect, and energy to counter this daunting but profound threat.     

Epitope-specific humoral immunity to Plasmodium vivax Duffy binding protein

Erythrocyte invasion by Plasmodium vivax is completely dependent on binding to the Duffy blood group antigen by the parasite Duffy binding protein (DBP). The receptor-binding domain of this protein lies within a cysteine-rich region referred to as region II (DBPII). To examine whether antibody responses to DBP correlate with age-acquired immunity to P. vivax, antibodies to recombinant DBP (rDBP) were measured in 551 individuals residing in a village endemic for P. vivax in Papua New Guinea, and linear epitopes mapped in the critical binding region of DBPII. Antibody levels to rDBP(II) increased with age. Four dominant linear epitopes were identified, and the number of linear epitopes recognized by semi-immune individuals increased with age, suggesting greater recognition with repeated infection. Some individuals had antibodies to rDBP(II) but not to the linear epitopes, indicating the presence of conformational epitopes. This occurred in younger individuals or subjects acutely infected for the first time with P. vivax, indicating that repeated infection is required for recognition of linear epitopes. All four dominant B-cell epitopes contained polymorphic residues, three of which showed variant-specific serologic responses in over 10% of subjects examined. In conclusion, these results demonstrate age-dependent and variant-specific antibody responses to DBPII and implicate this molecule in partial acquired immunity to P. vivax in populations in endemic areas.     

Two antigens on zygotes and ookinetes of Plasmodium yoelii and Plasmodium berghei that are distinct targets of transmission-blocking immunity.

We have developed transmission-blocking monoclonal antibodies (MAbs) against Plasmodium yoelii 21-kDa (Pys21) and 28-kDa (Pys25) ookinete surface proteins. These MAbs block infectivity of P. yoelii to Anopheles stephensi. One MAb, 14, cross-reacted by Western blotting with a 28-kDa surface protein (Pbs25) of P. berghei ookinetes and blocked oocyst development, as assayed by direct mosquito feeds on passively immunized P. berghei-infected mice. In total, we have identified two ookinete surface proteins in P. yoelii, one of which is also present in P. berghei. The transmission-blocking activity of the anti-Pys25 MAb 4 was complete and more potent than that of the anti-Pys21 MAb 2. Moreover, Fab fragments of MAb 4 had transmission-blocking activity in mice. In comparison, Fab fragments of MAb 2 did not have detectable transmission-blocking effect, although F(ab')2 did. Furthermore, MAb 2 and MAb 4 appeared to block the in vitro formation and development of zygotes as well.     

Evaluation of the OptiMAL Test for Rapid Diagnosis of Plasmodium vivax and Plasmodium falciparum Malaria

The development of rapid and specific diagnostic tests to identify individuals infected with malaria is of paramount importance in efforts to control the severe public health impact of this disease. This study evaluated the ability of a newly developed rapid malaria diagnostic test, OptiMAL (Flow Inc., Portland, Oreg.), to detect Plasmodium vivax and Plasmodium falciparum malaria during an outbreak in Honduras. OptiMAL is a rapid (10-min) malaria detection test which utilizes a dipstick coated with monoclonal antibodies against the intracellular metabolic enzyme parasite lactate dehydrogenase (pLDH). Differentiation of malaria parasites is based on antigenic differences between the pLDH isoforms. Since pLDH is produced only by live Plasmodium parasites, this test has the ability to differentiate live from dead organisms. Results from the OptiMAL test were compared to those obtained by reading 100 fields of traditional Giemsa-stained thick-smear blood films. Whole-blood samples were obtained from 202 patients suspected of having malaria. A total of 96 samples (48%) were positive by blood films, while 91 (45%) were positive by the OptiMAL test. The blood films indicated that 82% (79 of 96) of the patients were positive for P. vivax and 18% (17 of 96) were infected with P. falciparum. The OptiMAL test showed that 81% (74 of 91) were positive for P. vivax and 19% (17 of 91) were positive for P. falciparum. These results demonstrated that the OptiMAL test had sensitivities of 94 and 88% and specificities of 100 and 99%, respectively, when compared to traditional blood films for the detection of P. vivax and P. falciparum malaria. Blood samples not identified by OptiMAL as malaria positive normally contained parasites at concentrations of less than 100/μl of blood. Samples found to contain P. falciparum were further tested by two other commercially available rapid malaria diagnostic tests, ParaSight-F (Becton Dickinson, Cockeysville, Md.) and ICT Malaria P.f. (ICT Diagnostics, Sydney, Australia), both of which detect only P. falciparum. Only 11 of the 17 (65%) P. falciparum-positive blood samples were identified by the ICT and ParaSight-F tests. Thus, OptiMAL correctly identified P. falciparum malaria parasites in patient blood samples more often than did the other two commercially available diagnostic tests and showed an excellent correlation with traditional blood films in the identification of both P. vivax malaria and P. falciparum malaria. We conclude that the OptiMAL test is an effective tool for the rapid diagnosis of malaria.     

Recognition of variant Rifin antigens by human antibodies induced during natural Plasmodium falciparum infections

Antibodies from individuals living in areas where malaria is endemic are known to react with parasite-derived erythrocyte surface proteins. The major immunogenic and clonally variant surface antigen described to date is Plasmodium falciparum erythrocyte membrane protein 1 (PfEMP-1), which is encoded by members of the multicopy var gene family. We report here that rifin proteins (RIF proteins), belonging to the largest known family of variable infected erythrocyte surface-expressed proteins, are also naturally immunogenic. Recombinant RIF proteins were used to analyze the antibody responses of individuals living in an area of intense malaria transmission. Elevated anti-rifin antibody levels were detected in the majority of the adult population tested, whereas the prevalence of such antibodies was much lower in malaria-exposed children. Despite the high degree of diversity between rif sequences and the high gene copy number, it appears that P. falciparum infections can induce antibodies that cross-react with several variant rifin molecules in many parasite isolates in a given community, and the immune response is most likely to be stable over time in a hyperendemic area. The protein was localized by fluorescence microscopy on the membrane of ring and young trophozoite-infected erythrocytes with antibodies from human immune sera with specificities for recombinant RIF protein.     

Cost-Benefit Analysis of Tafenoquine for Radical Cure of Plasmodium vivax Malaria in Korea

BackgroundPlasmodium vivax malaria has a persistent liver stage that causes relapse, and introducing tafenoquine to suppress relapse could aid in disease eradication. Therefore, we assessed the impact of tafenoquine introduction on P. vivax malaria incidence and performed a cost-benefit analysis from the payer’s perspective.MethodsWe expanded the previously developed P. vivax malaria dynamic transmission model and calibrated it to weekly civilian malaria incidences in 2014–2018. Primaquine and tafenoquine scenarios were considered by assuming different relapse probabilities, and relapse and total P. vivax malaria cases were predicted over the next decade for each scenario. We then estimated the number of cases prevented by replacing primaquine with tafenoquine. The cost and benefit of introducing tafenoquine were obtained using medical expenditure from a nationwide database, and a cost-benefit analysis was conducted. A probabilistic sensitivity analysis was performed to assess the economic feasibility robustness of tafenoquine introduction under uncertainties of model parameters, costs, and benefits.ResultsUnder 0.04 primaquine relapse probability, the introduction of tafenoquine with relapse probability of 0.01 prevented 129 (12.27%) and 35 (77.78%) total and relapse cases, respectively, over the next decade. However, under the same relapse probability as primaquine, introducing tafenoquine had no additional preventative effect. The 14-day primaquine treatment cost was $3.71. The tafenoquine and the glucose-6-phosphate dehydrogenase rapid diagnostic testing cost $57.37 and $7.76, totaling $65.13. The average medical expenditure per malaria patient was estimated at $1444.79. The cost-benefit analysis results provided an incremental benefit-cost ratio (IBCR) from 0 to 3.21 as the tafenoquine relapse probability decreased from 0.04 to 0.01. The probabilistic sensitivity analysis showed an IBCR > 1, indicating that tafenoquine is beneficial, with a probability of 69.1%.ConclusionTafenoquine could reduce P. vivax malaria incidence and medical costs and bring greater benefits than primaquine.     

Generation of reactive oxygen species by blood monocytes in human Plasmodium falciparum and P. vivax infections

Generation of reactive oxygen radicals by peripheral blood monocytes was measured by luminol-dependent chemiluminescence in 23 P. vivax- and 7 P. falciparum-infected patients. The chemiluminescence index (CLI) was not found to be significantly higher in P. vivax-infected cases than in healthy controls. But in patients with P. falciparum infection, the CLI was significantly higher compared to controls as well as to P. vivax-infected patients. In two severe and complicated P. falciparum-infected cases, CLI was found to be higher than in mild cases. As immunosuppression is more marked in falciparum malaria than in vivax cases, the role of oxygen radical generation in immunopathology and causation of immunosuppression in falciparum malaria needs further investigation.     

Individual Plasmodium vivax msp1 variants within polyclonal P. vivax infections display different propensities for relapse

Using a newly developed Plasmodium vivax merozoite surface protein 1 gene (Pvmsp1) heteroduplex tracking assay, we genotyped 107 P. vivax infections in individuals from Cambodia, 45 of whom developed recurrent parasitemia within 42 days. The majority of isolates were polyclonal, but recurrent parasitemias displayed fewer variants compared to initial parasitemias. Two Pvmsp1 gene variants occurred more frequently in the initial genotypes of those who developed recurrent parasitemia, representing the first time P. vivax variants associated with a higher risk of relapse have been described.     

Diversity and natural selection in Plasmodium vivax Duffy binding protein gene

The Plasmodium vivax Duffy binding protein (DBP) binds to the Duffy blood group antigen on the surface of erythrocytes and is essential for invasion. Natural immunity develops to this protein making it an important vaccine candidate. Genetic diversity within and between populations was compared in 100 dbp sequences from isolates obtained from Papua New Guinea, Colombia, and South Korea. The cysteine-rich region II, that contains the binding domain, has the highest diversity compared to the rest of the dbp gene and appears to be under strong selective pressure based on statistical tests comparing rates of non-synonymous (K(n)) to synonymous mutations (K(s)) among P. vivax isolates and to those of closely related species. By contrast, meiotic recombination was not found to be significant for maintaining genetic diversity. A comparison of the patterns of nucleotide diversity within dbpII to that of genes encoding homologous erythrocyte binding proteins of Plasmodium knowlesi predict critical binding residues juxtaposed to polymorphic B- and T-cell epitopes. Phylogenic analysis and measurement of nucleotide diversity between and within the different geographic populations support emergence of distinct allelic families suggestive of divergent selection of alleles between populations. Development of a P. vivax DBP-based vaccine must take into account regions of high diversity within the molecule and alleles that show distinct geographic differences.     

Potential Immune Mechanisms Associated with Anemia in Plasmodium vivax Malaria: a Puzzling Question

The pathogenesis of malaria is complex, generating a broad spectrum of clinical manifestations. One of the major complications and concerns in malaria is anemia, which is responsible for considerable morbidity in the developing world, especially in children and pregnant women. Despite its enormous health importance, the immunological mechanisms involved in malaria-induced anemia remain incompletely understood. Plasmodium vivax, one of the causative agents of human malaria, is known to induce a strong inflammatory response with a robust production of immune effectors, including cytokines and antibodies. Therefore, it is possible that the extent of the immune response not only may facilitate the parasite killing but also may provoke severe illness, including anemia. In this review, we consider potential immune effectors and their possible involvement in generating this clinical outcome during P. vivax infections.     

Protective immunity induced in Aotus monkeys by recombinant SERA proteins of Plasmodium falciparum.

We describe the vaccination of Panamanian monkeys (Aotus sp.) with two recombinant blood stage antigens that each contain a portion of the N-terminal region of the SERA (serine repeat antigen) protein of the malaria parasite Plasmodium falciparum. We immunized with either a 262-amino-acid SERA fragment (SERA I) that contains amino acids 24 to 285 of the 989-amino-acid protein or a 483-amino-acid SERA fragment (SERA N) that contains amino acids 24 to 506 as part of a fusion protein with human gamma interferon. The recombinant proteins were shown to stimulate protective immunity when administered with complete and incomplete Freund adjuvant. Four of six immunized monkeys challenged by intravenous inoculation with blood stage P. falciparum developed parasitemias that were reduced by at least 1,000-fold. Two of six immunized monkeys developed parasitemias which were comparable to the lowest parasitemia in one of four controls and were 50- to 1,000-fold lower than in the other three controls.     

Plasmodium vivax induced myocarditis: A rare case report

Malaria is one of the commonest parasitic disease in the tropics since ages. However the plasmodium still continues to give surprises to all of us. In the similar context we report a case of Plasmodium vivax induced myocarditis in a 20 year old male and review the literature related to this rare entitiy.     

约氏疟原虫Pys25、Pys21抗原的表达及其传播阻断免疫的研究

目的 :观察约氏疟原虫有性发育阶段虫体抗原Pys2 5、Pys2 1的表达及其单克隆抗体的传播阻断效应。方法 :分别用抗Pys2 5和抗Pys2 1单抗 (McAb4和McAb10 )被动免疫感染约氏疟原虫的小鼠 ,通过直接蚊喂养试验 ,观察抗Pys2 5McAb4和抗Pys2 1McAb10对疟原虫在蚊体内发育过程的影响 ;应用间接免疫荧光和Westernblotting试验 ,检测疟原虫有性发育阶段虫体抗原Pys2 5和Pys2 1表达的动态变化。结果 :与抗Pys2 1McAb10比较 ,抗Pys2 5McAb4具有更完全的传播阻断效应。Pys2 5和Pys2 1抗原蛋白存在于从配子体到动合子的整个发育过程中 ,但Pys2 5在合子表面的表达显著早于Pys2 1。结论 :约氏疟原虫Pys2 5和Pys2 1均是传播阻断免疫的靶抗原 ;抗Pys2 5McAb4具有更强的传播阻断活性与Pys2 5在合子表面的早期表达有关。     

Plasmodium vivax malaria vaccine development.

Plasmodium vivax represents the most widespread malaria parasite worldwide. Although it does not result in as high a mortality rate as P. falciparum, it inflicts debilitating morbidity and consequent economic impact in endemic communities. In addition, the relapsing behavior of this malaria parasite and the recent resistance to anti-malarials contribute to making its control more difficult. Although the biology of P. vivax is different from that of P. falciparum and the human immune response to this parasite species has been rather poorly studied, significant progress is being made to develop a P. vivax-specific vaccine based on the information and experience gained in the search for a P. falciparum vaccine. We have devoted great effort to antigenically characterize the P. vivax CS protein and to test its immunogenicity using the Aotus monkey model. Together with other groups we are also assessing the immunogenicity and protective efficacy of the asexual blood stage vaccine candidates MSP-1 and DBP in the monkey model, as well as the immunogenicity of Pvs25 and Pvs28 ookinete surface proteins. The transmission-blocking efficacy of the responses induced by these latter antigens is being assessed using Anopheles albimanus mosquitoes. The current status of these vaccine candidates and other antigens currently being studied is described.     

Phagocytosis does not play a major role in naturally acquired transmission-blocking immunity to Plasmodium falciparum malaria

Phagocytosis of Plasmodium falciparum sexual stages in vitro and within the mosquito midgut was assayed in order to assess its role in transmission-blocking immunity to malaria. Both monocytes/macrophages (MM) and polymorphonuclear neutrophils (PMN) phagocytosed malarial gametes in vitro, but levels of phagocytosis were low. Intraerythrocytic gametocytes were not susceptible to phagocytosis. In vitro phagocytosis was positively correlated with levels of antibodies against the gamete surface proteins Pfs230 and Pfs48/45. Immunoglobulin G (IgG) subclass analysis revealed that phagocytosis was correlated with levels of antigamete IgG1. In vivo membrane-feeding experiments were performed in the presence of both pooled and individual malaria immune sera. The phagocytic process proceeded less efficiently in vivo than in vitro, which may be related to the lower ambient temperature (26 degrees C, compared with 37 degrees C). Finally, although we found a correlation between the ability of a serum to promote phagocytosis in vitro and the presence of antibodies against transmission-blocking target antigens, we were unable to demonstrate a role for MM- or PMN-mediated phagocytosis in reduction of infectivity of the malarial parasite to mosquitoes.