The relationship between zinc intake and growth in children aged 1-8 years: a systematic review and meta-analysis

BACKGROUND/OBJECTIVES: It is estimated that zinc deficiency affects 17% of the world's population, and because of periods of rapid growth children are at an increased risk of deficiency, which may lead to stunting. This paper presents a systematic review and meta-analysis of the randomised controlled trials (RCTs) that assess zinc intake and growth in children aged 1–8 years. This review is part of a larger systematic review by the European Micronutrient Recommendations Aligned Network of Excellence that aims to harmonise the approach to setting micronutrient requirements for optimal health in European populations (www.eurreca.org). SUBJECT/METHODS: Searches were performed of literature published up to and including December 2013 using MEDLINE, Embase and the Cochrane Library databases. Included studies were RCTs in apparently healthy child populations aged from 1 to 8 years that supplied zinc supplements either as capsules or as part of a fortified meal. Pooled meta-analyses were performed when appropriate. RESULTS: Nine studies met the inclusion criteria. We found no significant effect of zinc supplementation of between 2 weeks and 12 months duration on weight gain, height for age, weight for age, length for age, weight for height (WHZ) or WHZ scores in children aged 1–8 years. CONCLUSIONS: Many of the children in the included studies were already stunted and may have been suffering from multiple micronutrient deficiencies, and therefore zinc supplementation alone may have only a limited effect on growth

Persistence of Protective Immunity to Malaria Induced by DNA Priming and Poxvirus Boosting: Characterization of Effector and Memory CD8+-T-Cell Populations

The persistence of immunity to malaria induced in mice by a heterologous DNA priming and poxvirus boosting regimen was characterized. Mice were immunized by priming with DNA vaccine plasmids encoding the Plasmodium yoelii circumsporozoite protein (PyCSP) and murine granulocyte-macrophage colony-stimulating factor and boosting with recombinant vaccinia encoding PyCSP. BALB/c mice immunized with either high-dose (100 µg of p PyCSP plus 30 µg of pGM-CSF) or low-dose (1 µg of p PyCSP plus 1 µg of pGM-CSF DNA) priming were protected against challenge with 50 P. yoelii sporozoites. Protection 2 weeks after immunization was 70 to 100%, persisted at this level for at least 20 weeks, and declined to 30 to 40% by 28 weeks. Eight of eight mice protected at 20 weeks were still protected when rechallenged at 40 weeks. The antigen (Ag)-specific effector CD8+-T-cell population present 2 weeks after boosting had ex vivo Ag-specific cytolytic activity, expressed both gamma interferon (IFN-{gamma}) and tumor necrosis factor alpha, and constituted 12 to 20% of splenic CD8+ T cells. In contrast, the memory CD8+-Ag-specific-cell population at 28 weeks lacked cytolytic activity and constituted only 6% of splenic CD8+ T cells, but at the single-cell level it produced significantly higher levels of IFN-{gamma} than the effectors. High levels of Ag- or parasite-specific antibodies present 2 weeks after boosting had declined three- to sevenfold by 28 weeks. Low-dose priming was similarly immunogenic and as protective as high-dose priming against a 50-, but not a 250-, sporozoite challenge. These results demonstrate that a heterologous priming and boosting vaccination can provide lasting protection against malaria in this model system

Protection from Experimental Cerebral Malaria with a Single Dose of Radiation-Attenuated, Blood-Stage Plasmodium berghei Parasites

BACKGROUND: Whole malaria parasites are highly effective in inducing immunity against malaria. Due to the limited success of subunit based vaccines in clinical studies, there has been a renewed interest in whole parasite-based malaria vaccines. Apart from attenuated sporozoites, there have also been efforts to use live asexual stage parasites as vaccine immunogens. METHODOLOGY AND RESULTS: We used radiation exposure to attenuate the highly virulent asexual blood stages of the murine malaria parasite P. berghei to a non-replicable, avirulent form. We tested the ability of the attenuated blood stage parasites to induce immunity to parasitemia and the symptoms of severe malaria disease. Depending on the mouse genetic background, a single high dose immunization without adjuvant protected mice from parasitemia and severe disease (CD1 mice) or from experimental cerebral malaria (ECM) (C57BL/6 mice). A low dose immunization did not protect against parasitemia or severe disease in either model after one or two immunizations. The protection from ECM was associated with a parasite specific antibody response and also with a lower level of splenic parasite-specific IFN-γ production, which is a mediator of ECM pathology in C57BL/6 mice. Surprisingly, there was no difference in the sequestration of CD8+ T cells and CD45+ CD11b+ macrophages in the brains of immunized, ECM-protected mice. CONCLUSIONS: This report further demonstrates the effectiveness of a whole parasite blood-stage vaccine in inducing immunity to malaria and explicitly demonstrates its effectiveness against ECM, the most pathogenic consequence of malaria infection. This experimental model will be important to explore the formulation of whole parasite blood-stage vaccines against malaria and to investigate the immune mechanisms that mediate protection against parasitemia and cerebral malaria

Pathogenic Roles of CD14, Galectin-3, and OX40 during Experimental Cerebral Malaria in Mice

An in-depth knowledge of the host molecules and biological pathways that contribute towards the pathogenesis of cerebral malaria would help guide the development of novel prognostics and therapeutics. Genome-wide transcriptional profiling of the brain tissue during experimental cerebral malaria (ECM ) caused by Plasmodium berghei ANKA parasites in mice, a well established surrogate of human cerebral malaria, has been useful in predicting the functional classes of genes involved and pathways altered during the course of disease. To further understand the contribution of individual genes to the pathogenesis of ECM, we examined the biological relevance of three molecules – CD14, galectin-3, and OX40 that were previously shown to be overexpressed during ECM. We find that CD14 plays a predominant role in the induction of ECM and regulation of parasite density; deletion of the CD14 gene not only prevented the onset of disease in a majority of susceptible mice (only 21% of CD14-deficient compared to 80% of wildtype mice developed ECM, p<0.0004) but also had an ameliorating effect on parasitemia (a 2 fold reduction during the cerebral phase). Furthermore, deletion of the galectin-3 gene in susceptible C57BL/6 mice resulted in partial protection from ECM (47% of galectin-3-deficient versus 93% of wildtype mice developed ECM, p<0.0073). Subsequent adherence assays suggest that galectin-3 induced pathogenesis of ECM is not mediated by the recognition and binding of galectin-3 to P. berghei ANKA parasites. A previous study of ECM has demonstrated that brain infiltrating T cells are strongly activated and are CD44+CD62L− differentiated memory T cells [1]. We find that OX40, a marker of both T cell activation and memory, is selectively upregulated in the brain during ECM and its distribution among CD4+ and CD8+ T cells accumulated in the brain vasculature is approximately equal

Determination of the radiation dose necessary to attenuate blood-stage <i>Pb-A</i> parasites.

<p>Virulent <i>Pb-A</i> parasitized RBC (pRBC) were exposed to increasing doses of gamma irradiation (10, 20, 40, 60, 80, 100 kilorads). 10⁷ irradiated pRBC or non-irradiated control pRBC (Ctrl) were injected intravenously (iv) into groups of naïve CD1 mice and parasitemia was monitored by blood smear. The graph shows the percentage of mice without patent parasitemia in each group over time beginning with day 2. Data are pooled from 3 experiments. Groups of mice infected with control pRBC or with pRBC exposed to 10, 20, 40, or 60 krad irradiation all developed patent parasitemia by day 7 (n = 4 mice each group). Accordingly, the curves describing these groups approach 0% by day 7. 70% (10/14) of mice infected with 80 krad irradiated pRBC remained free from patent blood-stage disease by day 14 (open circles, dashed line). 100% (10/10) of the mice injected with 100 krad irradiated pRBC remained free from blood stage infection under the period of observation (open squares, solid line). 100 krad irradiation was chosen as the attenuating dose in subsequent experiments.</p

Protection from parasitemia and severe disease in CD1 mice after immunization with irradiated blood-stage <i>Pb-A</i>.

<p><b>A</b>) Protection in CD1 mice after a single immunization with 10⁷ irradiation attenuated <i>Pb-A</i> blood-stage parasites (IrrPb). 10³ or 10⁷ IrrPb were injected iv into groups mice. On day 24 after injection, immunized and naïve mice were challenged with 10⁴ virulent <i>Pb-A</i> pRBC and parasitemia was monitored by blood smear (mean +/− SD) Daggers indicate mice that were euthanized or that died during the experiment. All naïve mice (4/4, filled circles) and 10³ IrrPb immunized mice (5/5, open triangles) displayed uncontrolled parasite growth and succumbed to acute blood-stage infection by day 12 after challenge. In contrast, mice immunized with 10⁷ IrrPb (4/5, open squares) controlled parasite growth after challenge and managed blood parasitemia down to low or undetectable levels. <b>B</b>) Protection in CD1 mice after a two immunizations with 10⁷ IrrPb. Groups of mice were immunized with 10³ or 10⁷ IrrPb, and then given a boost immunization 24 days later with an equal number of IrrPb (Boost). 24 days after the final immunization, immunized and naïve mice were challenged with 10⁴ virulent <i>Pb-A</i> pRBC. In mice immunized with 2 doses of 10³ IrrPb (n = 5 mice, open triangles), parasitemia levels continued to increase after challenge, similar to naïve mice (n = 5 mice, filled circles). In contrast, when mice were immunized with 2 doses of 10⁷ IrrPb (n = 5 mice, open squares), blood parasitemia declined over time to low or undetectable levels.</p

Protection from experimental cerebral malaria (ECM) in C57BL/6 mice after immunization with IrrPb.

<p><b>A and B</b>) Protection from ECM after a single immunization of 10⁷ IrrPb. Data are pooled from two independent experiments. Groups of mice were immunized with 10³ or 10⁷ IrrPb and then challenged with 10⁴ virulent <i>Pb-A</i> pRBC on day 19 or 28 after immunization. To assess ECM, challenged mice were monitored for neurological symptoms, the ability to survive beyond day 10 (A), and blood parasitemia levels (B). <b>A</b>) 10% (1/10) of naïve mice (filled circles) and 12% (2/17) of mice immunized with 10³ IrrPb (open triangles) survived beyond day 10 after challenge. In contrast, 81% (17/21) of mice immunized with 10⁷ IrrPb (open squares) survived beyond day 10 (two-sided p = .0003, 10⁷ IrrPb group vs. naïve group, Fisher's exact test). <b>B</b>) In each group, mice that succumbed to ECM before day 10 had blood parasitemias less than 10%, while mice that survived beyond day 10 eventually developed hyperparasitemia. <b>C and D</b>) Protection from ECM after two immunizations with 10⁷ IrrPb. Mice were immunized with 10³ or 10⁷ IrrPb twice with a 28 day interval (Boost), challenged with 10⁴ virulent pRBC, and then monitored for survival (C) and blood parasitemia (D). <b>C</b>) 20% (1/5) of naïve mice (filled circles) and none (0/10) of the mice immunized twice with 10³ attenuated parasites (open triangles) survived beyond day 10 after challenge. In contrast, all (10/10) of the mice immunized with 10⁷ attenuated parasites (open squares) survived beyond day 10 (p = .0037, 10⁷ IrrPb group vs. naïve group, Fisher's exact test), and eventually developed hyperparasitemia (D).</p

Identification, Cloning, Expression, and Characterization of the Gene for Plasmodium knowlesi Surface Protein Containing an Altered Thrombospondin Repeat Domain

Proteins present on the surface of malaria parasites that participate in the process of invasion and adhesion to host cells are considered attractive vaccine targets. Aided by the availability of the partially completed genome sequence of the simian malaria parasite Plasmodium knowlesi, we have identified a 786-bp DNA sequence that encodes a 262-amino-acid-long protein, containing an altered version of the thrombospondin type I repeat domain (SPATR). Thrombospondin type 1 repeat domains participate in biologically diverse functions, such as cell attachment, mobility, proliferation, and extracellular protease activities. The SPATR from P. knowlesi (PkSPATR) shares 61% and 58% sequence identity with its Plasmodium falciparum and Plasmodium yoelii orthologs, respectively. By immunofluorescence analysis, we determined that PkSPATR is a multistage antigen that is expressed on the surface of P. knowlesi sporozoite and erythrocytic stage parasites. Recombinant PkSPATR produced in Escherichia coli binds to a human hepatoma cell line, HepG2, suggesting that PkSPATR is a parasite ligand that could be involved in sporozoite invasion of liver cells. Furthermore, recombinant PkSPATR reacted with pooled sera from P. knowlesi-infected rhesus monkeys, indicating that native PkSPATR is immunogenic during infection. Further efficacy evaluation studies in the P. knowlesi-rhesus monkey sporozoite challenge model will help to decide whether the SPATR molecule should be developed as a vaccine against human malarias

Spleen weights of mice immunized with IrrPb increase during a virulent infection.

<p>C57BL/6 mice were immunized once with 10⁷ IrrPb, challenged with virulent parasites, and then spleens were harvested on day 6 after challenge. Spleens from immunized mice after challenge were an average 0.17 grams (0.09, 0.24) larger than the spleens from naïve mice after challenge (two-sided p<.001, Two Way ANOVA, Bonferroni post test). Results (n = 8 mice in each group) are pooled from two independent experiments.</p

A Novel Chimeric Plasmodium vivax Circumsporozoite Protein Induces Biologically Functional Antibodies That Recognize both VK210 and VK247 Sporozoites

A successful vaccine against Plasmodium vivax malaria would significantly improve the health and quality of the lives of more than 1 billion people around the world. A subunit vaccine is the only option in the absence of long-term culture of P. vivax parasites. The circumsporozoite protein that covers the surface of Plasmodium sporozoites is one of the best-studied malarial antigens and the most promising vaccine in clinical trials. We report here the development of a novel “immunologically optimal” recombinant vaccine expressed in Escherichia coli that encodes a chimeric CS protein encompassing repeats from the two major alleles, VK210 and VK247. This molecule is widely recognized by sera from patients naturally exposed to P. vivax infection and induces a highly potent immune response in genetically disparate strains of mice. Antibodies from immunized animals recognize both VK210 and VK247 sporozoites. Furthermore, these antibodies appear to be protective in nature since they cause the agglutination of live sporozoites, an in vitro surrogate of sporozoite infectivity. These results strongly suggest that recombinant CS is biologically active and highly immunogenic across major histocompatibility complex strains and raises the prospect that in humans this vaccine may induce protective immune responses