长效和短效NO发生剂体内干预疟原虫红内期重要侵袭分子转录水平的观察

为探讨NO对疟原虫红内期侵袭相关分子MSP-1、AMA-1和RhopH complex转录水平的影响。通过雌性BALB／c小鼠腹腔感染1×10^6致死型约氏疟原虫P．yoelii 17XL,体内给予NO长效（NOC18）和短效（NOC5）发生剂进行干预后,纯化疟原虫成熟裂殖体,提取总RNA,通过Real—time PCR相对定量方法检测MSP-1、AMA—1和RhopH complex的转录水平。结果显示和正常感染组相比,NOC5处理后疟原虫侵入的关键分子MSP—1、AMA-1和RhopH complex的转录水平明显下降;而NOC18处理则未见这一现象。本研究结果提示NO抑制疟原虫侵袭关键分子的转录水平,进而可能下调疟原虫相应蛋白的表达,从而影响疟原虫的侵入过程。     

Determinants of distribution and prevalence of avian malaria in blue tit populations across Europe: separating host and parasite effects

Although avian malarial parasites are globally distributed, the factors that affect the geographical distribution and local prevalence of different parasite lineages across host populations or species are still poorly understood. Based on the intense screening of avian malarial parasites in nine European blue tit populations, we studied whether distribution ranges as well as local adaptation, host specialization and phylogenetic relationships can determine the observed prevalences within populations. We found that prevalence differed consistently between parasite lineages and host populations, indicating that the transmission success of parasites is lineage specific but is partly shaped by locality-specific effects. We also found that the lineage-specific estimate of prevalence was related to the distribution range of parasites: lineages found in more host populations were generally more prevalent within these populations. Additionally, parasites with high prevalence that were also widely distributed among blue tit populations were also found to infect more host species. These findings suggest that parasites reaching high local prevalence can also realize wide distribution at a global scale that can have further consequences for host specialization. Although phylogenetic relationships among parasites did not predict prevalence, we detected a close match between a tree based on the geographic distance of the host populations and the parasite phylogenetic tree, implying that neighbouring host populations shared a related parasite fauna.     

The fitness of drug-resistant malaria parasites in a rodent model: multiplicity of infection

Malaria infections normally consist of more than one clonally replicating lineage. Within-host interactions between sensitive and resistant parasites can have profound effects on the evolution of drug resistance. Here, using the Plasmodium chabaudi mouse malaria model, we ask whether the costs and benefits of resistance are affected by the number of co-infecting strains competing with a resistant clone. We found strong competitive suppression of resistant parasites in untreated infections and marked competitive release following treatment. The magnitude of competitive suppression depended on competitor identity. However, there was no overall effect of the diversity of susceptible parasites on the extent of competitive suppression or release. If these findings generalize, then transmission intensity will impact on resistance evolution because of its effect on the frequency of mixed infections, not because of its effect on the distribution of clones per host. This would greatly simplify the computational problems of adequately capturing within-host ecology in models of drug resistance evolution in malaria.     

Structural basis for the functional and inhibitory mechanisms of β-hydroxyacyl-acyl carrier protein dehydratase (FabZ) of Plasmodium falciparum

The β-hydroxyacyl-acyl carrier protein dehydratase of Plasmodium falciparum (PfFabZ) catalyzes the third and important reaction of the fatty acid elongation cycle. The crystal structure of PfFabZ is available in hexameric (active) and dimeric (inactive) forms. However, PfFabZ has not been crystallized with any bound inhibitors until now. We have designed a new condition to crystallize PfFabZ with its inhibitors bound in the active site, and determined the crystal structures of four of these complexes. This is the first report on any FabZ enzyme with active site inhibitors that interact directly with the catalytic residues. Inhibitor binding not only stabilized the substrate binding loop but also revealed that the substrate binding tunnel has an overall shape of “U”. In the crystal structures, residue Phe169 located in the middle of the tunnel was found to be in two different conformations, open and closed. Thus, Phe169, merely by changing its side chain conformation, appears to be controlling the length of the tunnel to make it suitable for accommodating longer substrates. The volume of the substrate binding tunnel is determined by the sequence as well as by the conformation of the substrate binding loop region and varies between organisms for accommodating fatty acids of different chain lengths. This report on the crystal structures of the complexes of PfFabZ provides the structural basis of the inhibitory mechanism of the enzyme that could be used to improve the potency of inhibitors against an important component of fatty acid synthesis common to many infectious organisms.     

Nitric oxide is involved in the upregulation of IFN-γ and IL-10 mRNA expression by CD8⁺ T cells during the blood stages of P. chabaudi AS infection in CBA/Ca mice

Nitric oxide (NO) is involved in the clearance of several types of bacteria, viruses and parasites. Although the roles of NO and CD8⁺ T cells in the immune response to malaria have been extensively studied, their actual contributions during the blood stages of malaria infection remain unclear.In this work, we corroborate that serum NO levels are not associated with the in vivo elimination of the blood stages of Plasmodium chabaudi AS. In addition, we show that CD8⁺ T cells exhibit increased apoptosis and up regulate the expression of TNF-α mRNA on day 4 post-infection and IFN-γ and IL-10 mRNA on day 11 post-infection. Interestingly, only the levels of IFN-γ and IL-10 expression are affected when iNOS is inhibited with aminoguanidine (AG), suggesting that NO could be involved in the activation of CD8⁺ T cells during the blood stages of plasmodium infection.     

Antimalarial activity of endoperoxide compound 6-(1,2,6,7-tetraoxaspiro[7.11]nonadec-4-yl)hexan-1-ol

Plasmodium falciparum, the major causative parasite for the disease, has acquired resistance to most of the antimalarial drugs used today, presenting an immediate need for new antimalarial drugs. Here, we report the in vitro and in vivo antimalarial activities of 6-(1,2,6,7-tetraoxaspiro[7.11]nonadec-4-yl)hexan-1-ol (N-251) against P. falciparum and Plasmodium berghei parasites. The N-251 showed high antimalarial potencies both in the in vitro and the in vivo tests (EC₅₀ 2.3 × 10⁻⁸ M; ED₅₀ 15 mg/kg (per oral)). The potencies were similar to that of artemisinin in vitro and greater than artemisinin's activity in vivo (p.o.). In addition, N-251 has little toxicity: a single oral administration at 2000 mg/kg to a rat gave no health problems to it. Administration of N-251 to mice bearing 1% of parasitemia (per oral 68 mg/kg, 3 times a day for 3 consecutive days) resulted in a dramatic decrease in the parasitemia: all the 5 mice given N-251 were cured without any recurrence, with no diarrhea or weight loss occurring in the 60 days of experiment. N-251 deserves more extensive clinical evaluation, desirably including future trials in the human.     

微生物介导的恶性疟原虫PfMSP-1疫苗的研制现状

恶性疟原虫引起的恶性疟是一种严重危害人类健康的寄生虫病,采用疫苗防治该病是当前研究的热点领域之一。PfMSP-1抗原是一种有效的疫苗候选分子,鼠伤寒沙门菌、卡介苗、酵母菌、根癌农杆菌、嗜热四膜菌、腺病毒、牛痘病毒和杆状病毒等微生物经过改造后均可成为有希望的疫苗载体。本文综述了重组鼠伤寒沙门菌（rSt-PfMSP-1和rSt-PfMSP-119）、重组BCG疫苗（rBCG-PfMSP-1c和rBCG-PfMSP-119）、重组酵母菌（rPp-PfMSP-1）、重组根癌农杆菌（rAt-PfMSP-142、rAt-PfMSP4/5和rAt-Pf38）、重组嗜热四膜菌（rTt-PfMSP-119）、重组腺病毒（rAd5-PfMSP-142和rChAd63-PfMSP-1）、重组牛痘病毒（rVV-PfMSP-1和rVV-PfHGFSP）以及重组杆状病毒（rBDES-PfMSP-119）的构建及其免疫机制的研制现状。     

Trials for the co-expression of the merozoite surface protein-1 and circumsporozoite protein genes of Plasmodium vivax

Merozoite surface protein-1 (MSP-1), a major asexual blood stage antigen, and circumsporozoite protein (CSP), a component of sporozoites that includes a Plasmodium vivax B-cell epitope, are strong candidates for use in a malaria vaccine. A chimeric recombinant gene containing portions of both msp-1 and csp from P. vivax separated by Pro-Gly linker motif was generated. The construct gene was named mlc (msp-1, linker, and csp). The MLC chimeric recombinant protein had a molecular weight of approximately 25 kDa when expressed in Escherichia coli, as determined with sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS–PAGE) analysis. The purified chimeric protein reacted with the sera of patients infected with P. vivax but not with the sera of uninfected patients according to western blot analysis. The chimeric protein reacted well with sera of malaria patients (109/115, 94.78%) as assessed with enzyme-linked immunosorbent assay (ELISA). BALB/c mice that were orally immunized with the MLC chimeric recombinant protein successfully produced antigen-specific antibodies. Additionally, levels of the Th1-associated cytokines IL-12(p40), TNF-α, and IFN-γ were significantly increased in the spleens of the BALB/c mice. Therefore, the E. coli-expressed MLC chimeric recombinant protein might be used as a valuable vaccine candidate for oral immunization against vivax malaria.     

Competing for blood: the ecology of parasite resource competition in human malaria–helminth co‐infections

Ecological theory suggests that co‐infecting parasite species can interact within hosts directly, via host immunity and/or via resource competition. In mice, competition for red blood cells (RBCs) between malaria and bloodsucking helminths can regulate malaria population dynamics, but the importance of RBC competition in human hosts was unknown. We analysed infection density (i.e. the concentration of parasites in infected hosts), from a 2‐year deworming study of over 4000 human subjects. After accounting for resource‐use differences among parasites, we find evidence of resource competition, priority effects and a competitive hierarchy within co‐infected individuals. For example reducing competition via deworming increased Plasmodium vivax densities 2.8‐fold, and this effect is limited to bloodsucking hookworms. Our ecological, resource‐based perspective sheds new light into decades of conflicting outcomes of malaria–helminth co‐infection studies with significant health and transmission consequences. Beyond blood, investigating within‐human resource competition may bring new insights for improving human health.     

A comparative study on the heparin‐binding proteomes of Toxoplasma gondii and Plasmodium falciparum

Toxoplasma gondii is an obligatory intracellular apicomplexan parasite which exploits host cell surface components in cell invasion and intracellular parasitization. Sulfated glycans such as heparin and heparan sulfate have been reported to inhibit cell invasion by T. gondii and other apicomplexan parasites such as Plasmodium falciparum. The aim of this study was to investigate the heparin‐binding proteome of T. gondii. The parasite‐derived components were affinity‐purified on the heparin moiety followed by MS fingerprinting of the proteins. The heparin‐binding proteins of T. gondii and P. falciparum were compared based on functionality and affinity to heparin. Among the proteins identified, the invasion‐related parasite ligands derived from tachyzoite/merozoite surface and the secretory organelles were prominent. However, the profiles of the proteins were different in terms of affinity to heparin. In T. gondii, the proteins with highest affinity to heparin were the intracellular components with functions of parasite development contrasted to that of P. falciparum, of which the rhoptry‐derived proteins were prominently identified. The profiling of the heparin‐binding proteins of the two apicomplexan parasites not only explained the mechanism of heparin‐mediated host cell invasion inhibition, but also, to a certain extent, revealed that the action of heparin on the parasite extended after endocytosis.