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Volunteers immunized under chloroquine chemoprophylaxis with Plasmodium falciparum sporozoites (CPS) develop complete, longlasting protection against homologous sporozoite challenge. Chloroquine affects neither sporozoites nor liver-stages, but kills only asexual forms in erythrocytes once released from the liver into the circulation. Consequently, CPS immunization exposes the host to antigens from both preerythrocytic and blood stages, and induced immunity might target either of these stages. We therefore explored the life cycle stage specificity of CPS-induced protection. Twenty-five malaria-naïve volunteers were enrolled in a clinical trial, 15 of whom received CPS immunization. Five immunized subjects and five controls received a sporozoite challenge by mosquito bites, whereas nine immunized and five control subjects received an i.v. challenge with P. falciparum-infected erythrocytes. The latter approach completely bypasses preerythrocytic stages, enabling a direct comparison of protection against either life cycle stage. CPS-immunized subjects (13 of 14) developed anticircumsporozoite antibodies, whereas only one volunteer generated minimal titers against typical blood-stage antigens. IgG from CPS-immunized volunteers did not inhibit asexual blood-stage growth in vitro. All CPSimmunized subjects (5 of 5) were protected against sporozoite challenge. In contrast, nine of nine CPS-immunized subjects developed parasitemia after blood-stage challenge, with identical prepatent periods and blood-stage multiplication rates compared with controls. Intravenously challenged CPS-immunized subjects showed earlier fever and increased plasma concentrations of inflammatory markers D-dimer, IFN-γ, and monokine induced by IFN-γ than i.v. challenged controls. The complete lack of protection against blood-stage challenge indicates that CPS-induced protection is mediated by immunity against preerythrocytic stages. However, evidence is presented for immune recognition of P. falciparuminfected erythrocytes, suggesting memory responses unable to generate functional immunity.

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Apical membrane antigen 1 from Plasmodium is a leading malaria vaccine candidate. The protein is essential for host-cell invasion, but its molecular function is unknown. The crystal structure of the three domains comprising the ectoplasmic region of the antigen from P. vivax, solved at 1.8 angstrom resolution, shows that domains I and II belong to the PAN motif, which defines a superfamily of protein folds implicated in receptor binding. We also mapped the epitope of an invasion-inhibitory monoclonal antibody specific for the P. falciparum ortholog and modeled this to the structure. The location of the epitope and current knowledge on structure-function correlations for PAN domains together suggest a receptor-binding role during invasion in which domain II plays a critical part. These results are likely to aid vaccine and drug design.

P. berghei ANKA parasites deficient in schizont membrane-associated cytoadherence protein reveal a beneficial role for CD36-mediated tissue sequestration in aiding parasite growth.

Plasmodium falciparum apical membrane antigen 1 (PfAMA1), a candidate malaria vaccine, is polymorphic. This polymorphism is believed to be generated predominantly under immune selection pressure and, as a result, may compromise attempts at vaccination. Alignment of 355 PfAMA1 sequences shows that around 10% of the 622 amino acid residues can vary between alleles and that linkages between polymorphic residues occur. Using this analysis, we have designed three diversity-covering (DiCo) PfAMA1 sequences that take account of these linkages and, when taken together, on average incorporate 97% of amino acid variability observed. For each of the three DiCo sequences, a synthetic gene was constructed and used to transform the methylotrophic yeast Pichia pastoris, allowing recombinant expression. All three DiCo proteins were reactive with the reduction-sensitive monoclonal antibody 4G2, suggesting the DiCo sequences had conformations similar to those of naturally occurring PfAMA1. Rabbits were immunized with FVO strain PfAMA1 or with the DiCo proteins either individually or as a mixture. Antibody titers and the ability to inhibit parasite growth in vitro were determined. Animals immunized with the DiCo mix performed similarly to animals immunized with FVO AMA1 when measured against FCR3 strain parasites but outperformed animals immunized with FVO AMA1 when assessed against other strains. The levels of growth inhibition (ϳ70%) induced by the mix of three DiCo proteins were comparable for FVO, 3D7, and HB3, suggesting that a considerable degree of diversity in AMA1 is adequately covered. This suggests that vaccines based upon the DiCo mix approach provide a broader functional immunity than immunization with a single allele.

BackgroundContinuous high global tuberculosis (TB) mortality rates and variable vaccine efficacy of Mycobacterium bovis Bacille Calmette-Guérin (BCG) motivate the search for better vaccine regimes. Relevant models are required to downselect the most promising vaccines entering clinical efficacy testing and to identify correlates of protection.Methods and FindingsHere, we evaluated immunogenicity and protection against Mycobacterium tuberculosis in rhesus monkeys with two novel strategies: BCG boosted by modified vaccinia virus Ankara expressing antigen 85A (MVA.85A), and attenuated M. tuberculosis with a disrupted phoP gene (SO2) as a single-dose vaccine. Both strategies were well tolerated, and immunogenic as evidenced by induction of specific IFNγ responses. Antigen 85A-specific IFNγ secretion was specifically increased by MVA.85A boosting. Importantly, both MVA.85A and SO2 treatment significantly reduced pathology and chest X-ray scores upon infectious challenge with M. tuberculosis Erdman strain. MVA.85A and SO2 treatment also showed reduced average lung bacterial counts (1.0 and 1.2 log respectively, compared with 0.4 log for BCG) and significant protective effect by reduction in C-reactive protein levels, body weight loss, and decrease of erythrocyte-associated hematologic parameters (MCV, MCH, Hb, Ht) as markers of inflammatory infection, all relative to non-vaccinated controls. Lymphocyte stimulation revealed Ag85A-induced IFNγ levels post-infection as the strongest immunocorrelate for protection (spearman's rho: −0.60).ConclusionsBoth the BCG/MVA.85A prime-boost regime and the novel live attenuated, phoP deficient TB vaccine candidate SO2 showed significant protective efficacy by various parameters in rhesus macaques. Considering the phylogenetic relationship between macaque and man and the similarity in manifestations of TB disease, these data support further development of these primary and combination TB vaccine candidates.

Cytokines are key players in numerous inflammatory processes. Demonstration of a heritable component in the variation of cytokine production would indicate that simultaneous occurrence of conditions might be caused by a heritable inflammatory characteristic. We applied an extended twin study approach to assess heritability estimates of interleukin (IL)-1b, IL-1ra, IL-10, IL-6, and TNF-a production capacity after ex vivo stimulation with lipopolysaccharide. Cytokine production capacity was assessed in 42 monozygotic pairs, 52 dizygotic pairs, one trizygotic triplet, 33 single twins, and 83 additional siblings. Heritability estimates were derived from variance decomposition models using maximum likelihood estimation. For all cytokines, over 50% of the variance was genetically determined. IL-1ra and TNF-a had the lowest heritability estimate of 53%.Estimates for IL-6 and IL-10 were 57 and 62%, respectively. IL-1b had the highest estimate of 86%. We conclude that the production of cytokines is under tight genetic control.