Reduce Parasite Infection Research Articles

Genetic variation in resistance, but not tolerance, to a protozoan parasite in the monarch butterfly

Natural selection should strongly favour hosts that can protect themselves against parasites. Most studies on animals so far have focused on resistance, a series of mechanisms through which hosts prevent infection, reduce parasite growth or clear infection. However, animals may instead evolve tolerance, a defence mechanism by which hosts do not reduce parasite infection or growth, but instead alleviate the negative fitness consequences of such infection and growth. Here, we studied genetic variation in resistance and tolerance in the monarch butterfly (Danaus plexippus) to its naturally occurring protozoan parasite, Ophryocystis elektroscirrha. We exposed 560 monarch larvae of 19 different family lines to one of five different parasite inoculation doses (0, 1, 5, 10 and 100 infective spores) to create a range of parasite loads in infected butterflies. We then used two proxies of host fitness (adult lifespan and body mass) to quantify: (i) qualitative resistance (the ability to prevent infection; also known as avoidance or anti-infection resistance); (ii) quantitative resistance (the ability to limit parasite growth upon infection; also known as control or anti-growth resistance); and (iii) tolerance (the ability to maintain fitness with increasing parasite infection intensity). We found significant differences among host families in qualitative and quantitative resistance, indicating genetic variation in resistance. However, we found no genetic variation in tolerance. This may indicate that all butterflies in our studied population have evolved maximum tolerance, as predicted by some theoretical models.

Apical Surface Expression of Aspartic Protease Plasmepsin 4, a Potential Transmission-blocking Target of the Plasmodium Ookinete

To invade its definitive host, the mosquito, the malaria parasite must cross the midgut peritrophic matrix that is composed of chitin cross-linked by chitin-binding proteins and then develop into an oocyst on the midgut basal lamina. Previous evidence indicates that Plasmodium ookinete-secreted chitinase is important in midgut invasion. The mechanistic role of other ookinete-secreted enzymes in midgut invasion has not been previously examined. De novo mass spectrometry sequencing of a protein obtained by benzamidine affinity column of Plasmodium gallinaceum ookinete axenic culture supernatant demonstrated the presence of an ookinete-secreted plasmepsin, an aspartic protease previously only known to be present in the digestive vacuole of asexual stage malaria parasites. This plasmepsin, the ortholog of Plasmodium falciparum plasmepsin 4, was designated PgPM4. PgPM4 and PgCHT2 (the P. gallinaceum ortholog of P. falciparum chitinase PfCHT1) are both localized on the ookinete apical surface, and both are present in micronemes. Aspartic protease inhibitors (peptidomimetic and natural product), calpain inhibitors, and anti-PgPM4 monoclonal antibodies significantly reduced parasite infectivity for mosquitoes. These results suggest that plasmepsin 4, previously known only to function in the digestive vacuole of asexual blood stage Plasmodium, plays a role in how the ookinete interacts with the mosquito midgut interactions as it becomes an oocyst. These data are the first to delineate a role for an aspartic protease in mediating Plasmodium invasion of the mosquito and demonstrate the potential for plasmepsin 4 as a malaria transmission-blocking vaccine target.

Spatial Variation of Haemosporidian Parasite Infection in African Rainforest Bird Species

Spatial heterogeneity influences the distribution, prevalence, and diversity of haemosporidian parasites. Previous studies have found complex patterns of prevalence with respect to habitat characteristics and parasite genotype, and their interactions, but there is little information regarding how parasitemia intensity and the prevalence of co-infections may vary in space. Here, using both molecular methods and microscopy, we report an analysis of the variation of parasitemia intensity and co-infections of avian haemosporidian parasites ( Plasmodium and Haemoproteus species) in 2 common African birds species, the yellow-whiskered greenbul ( Andropadus latirostris ) and the olive sunbird ( Cyanomitra olivacea ), at 3 sites with distinct habitat characteristics in Ghana. First, we found an interaction between the site and host species for the prevalence of Plasmodium spp. and Haemoproteus spp. For the olive sunbird, the prevalence of Plasmodium spp., as well as the number of individuals with co-infections, varied significantly among the sites, but these measures remained constant for the yellow-whiskered greenbul. In addition, yellow-whiskered greenbuls infected with Haemoproteus spp. were found only at 1 site. Furthermore, for both bird species, the parasitemia intensity of Plasmodium spp. varied significantly among the 3 sites, but with opposing trends. These results suggest that spatial heterogeneity differently affects haemosporidian infection parameters in these vertebrate-hosts. Environmental conditions here can either favor or reduce parasite infection. We discuss the implications of these discrepancies for conservation and ecological studies of infectious diseases in natural populations.

CelTOS, a novel malarial protein that mediates transmission to mosquito and vertebrate hosts

The malarial parasite has two hosts in its life cycle, a vertebrate and a mosquito. We report here that malarial invasion into these hosts is mediated by a protein, designated cell-traversal protein for ookinetes and sporozoites (CelTOS), which is localized to micronemes that are organelles for parasite invasive motility. Targeted disruption of the CelTOS gene in Plasmodium berghei reduced parasite infectivity in the mosquito host approximately 200-fold. The disruption also reduced the sporozoite infectivity in the liver and almost abolished its cell-passage ability. Liver infectivity was restored in Kupffer cell-depleted rats, indicating that CelTOS is necessary for sporozoite passage from the circulatory system to hepatocytes through the liver sinusoidal cell layer. Electron microscopic analysis revealed that celtos-disrupted ookinetes invade the midgut epithelial cell by rupturing the cell membrane, but then fail to cross the cell, indicating that CelTOS is necessary for migration through the cytoplasm. These results suggest that conserved cell-passage mechanisms are used by both sporozoites and ookinetes to breach host cellular barriers. Elucidation of these mechanisms might lead to novel antimalarial strategies to block parasite's transmission.

Chitinase is crucial for parasite invasion of mosquito midgut

A set of two new papers provides compelling evidence supporting the crucial role of chitinase secreted by Plasmodium ookinetes for invasion of the mosquito midgut. Y-L. Tsai et al. [(2001) Infect. Immun. 69, 4048–4054,] disrupted the only identified P. falciparum chitinase gene PfCHT1 – this led to a significant impairment in the ability of affected parasites to form oocysts in the midguts of Anopheles freeborni. J. Dessens et al. [(2001) Infect. Immun. 69, 4041–4047] identified and characterized PbCHT1, the first chitinase gene from the rodent malaria parasite P. berghei. Targeted disruption of PbCHT1 also dramatically reduced parasite infectivity in A. stephensi mosquitoes. Interestingly, PbCHT1 null mutants had no residual ookinete-derived chitinase activity, suggesting that P. berghei ookinetes express only one chitinase gene. PbCHT1 is a structural orthologue of PfCHT1 and of the avian Plasmodium chitinase gene, PgCHT1. SHK

Knockout of the rodent malaria parasite chitinase pbCHT1 reduces infectivity to mosquitoes.

During mosquito transmission, malaria ookinetes must cross a chitin-containing structure known as the peritrophic matrix (PM), which surrounds the infected blood meal in the mosquito midgut. In turn, ookinetes produce multiple chitinase activities presumably aimed at disrupting this physical barrier to allow ookinete invasion of the midgut epithelium. Plasmodium chitinase activities are demonstrated targets for human and avian malaria transmission blockade with the chitinase inhibitor allosamidin. Here, we identify and characterize the first chitinase gene of a rodent malaria parasite, Plasmodium berghei. We show that the gene, named PbCHT1, is a structural ortholog of PgCHT1 of the avian malaria parasite Plasmodium gallinaceum and a paralog of PfCHT1 of the human malaria parasite Plasmodium falciparum. Targeted disruption of PbCHT1 reduced parasite infectivity in Anopheles stephensi mosquitoes by up to 90%. Reductions in infectivity were also observed in ookinete feeds-an artificial situation where midgut invasion occurs before PM formation-suggesting that PbCHT1 plays a role other than PM disruption. PbCHT1 null mutants had no residual ookinete-derived chitinase activity in vitro, suggesting that P. berghei ookinetes express only one chitinase gene. Moreover, PbCHT1 activity appeared insensitive to allosamidin inhibition, an observation that raises questions about the use of allosamidin and components like it as potential malaria transmission-blocking drugs. Taken together, these findings suggest a fundamental divergence among rodent, avian, and human malaria parasite chitinases, with implications for the evolution of Plasmodium-mosquito interactions.

Trypanosoma cruzi: Cytokine effects on macrophage trypanocidal activity

Mouse macrophages infected with Trypanosoma cruzi in vitro may be activated to reduce parasite infection by interferon γ (IFN-γ). The addition of up to 10,000 units of IFN-γ however, does not result in a 100% reduction of intracellular parasites. We, therefore, investigated the possibility that macrophages require an additional signal or signals to completely clear T. cruzi infection. Because the combination of IFN-γ with lipopolysaccharide greatly enhanced macrophages ability to decrease the number of intracellular parasites, the interaction of IFN-γ with tumor necrosis factor (TNF) was examined. TNF alone and the combination of TNF with IFN-γ did not have a significant effect on reducing parasite numbers below that obtained with IFN-γ alone. This was also true for lymphotoxin, a lymphokine similar to TNF in structure and function. The effect of IFN-γ in combination with a cytokine-rich supernatant containing IL-2, IL-3, IL-4, IL-5, and IFN-γ on macrophage clearance of the parasite was also examined. The cytokine-rich supernatant alone had no effect on reducing parasite infection of the macrophages; indeed, in some experiments the addition of the supernatant resulted in an increase in the level of parasite infection. However, 1000 units of IFN-γ combined with the complex cytokine mixture caused a decrease in parasite infection of nearly 100% compared to that of control cultures treated with media alone. To determine which cytokine or cytokines in the supernatant were responsible for this synergistic activity, anti-cytokine antibodies were added to the supernatant prior to its addition with IFN-γ to the cultures. Anti-IL-4 was the only antibody found to inhibit the synergism of IFN-γ with the cytokine-rich supernatant. IL-4, however, did not significantly enhance the ability of IFN-γ to induce macrophage clearance of the parasite, and IL-4 alone caused a slight increase in parasite infection in vitro. These results further define the role that cytokines play in T. cruzi infection of macrophages in vitro and suggest that the interaction of cytokine networks within this system is complex.

Circumsporozoite proteins of human malaria parasites Plasmodium falciparum and Plasmodium vivax.

Monoclonal antibodies were raised against sporozoites of two species of malaria parasites, Plasmodium falciparum and Plasmodium vivax. The antibodies reacted with polypeptides (circumsporozoite proteins) that are uniformly distributed over the entire surface of sporozoites, as shown by indirect immunofluorescence and by the circumsporozoite precipitin reaction. The epitopes recognized by the monoclonal antibodies were expressed on sporozoites from different geographical isolates of the homologous species but were not detected on sporozoites of heterologous species nor on blood forms of the parasite. The monoclonal antibody to P. falciparum specifically immunoprecipitated two polypeptides of apparent 67,000 mol wt (Pf67) and 58,000 mol wt (Pf58) from extracts of [35S]methionine-labeled P. falciparum sporozoites. Similarly, the anti-P. vivax monoclonal immunoprecipitated two proteins of 51,000 mol wt (Pv51) and 45,000 mol wt (Pv45) from extracts of metabolically labeled P. vivax sporozoites. The extracts were also reacted with the serum of human volunteers successfully vaccinated with sporozoites of either P. vivax or P. falciparum. The patterns of immunoprecipitation were almost identical to those obtained with the corresponding monoclonal antibodies. The circumsporozoite proteins of P. falciparum and P. vivax play a role in immune protection. Incubation of the appropriate monoclonal antibody with viable sporozoites of the homologous species significantly reduced parasite infectivity, as determined by sporozoite neutralization assays carried out in splenectomized chimpanzees.