Invasive amoebiasis, a spectrum of diseases caused by the enteric protozoan parasite Entamoeba histolytica, constitutes a major health problem mainly in tropical and subtropical countries with poor sanitary conditions. The different forms of the disease are characterized by massive tissue lesions. Amoeba-induced tissue destruction requires an intimate contact between E. histolytica trophozoites and host cells. This contact is predominantly mediated by a galactose-inhibitable lectin located on the surface of the amoebae. Therefore, the lectin is considered a prime candidate for the development of a vaccine to prevent amoebiasis. This communication reports on recent developments in characterizing the structure and function of the E. histolytica surface lectin and its use as a subunit vaccine.

Plasmodium vivax is the most widely distributed human malaria with an estimate of 35 million cases per year. The deduced amino acid sequence comparisons of the Merozoite Surface Protein 1 (MSP1) from several plasmodial species, including that of P. vivax (PvMSP1), revealed the existence of highly conserved blocks and polymorphic blocks. We had previously shown that sequences within conserved blocks from the N-terminal region of the PvMSP1 were poorly immunogenic in natural human infections. These results suggest that these regions code for important and unknown structural and/or functional features and thus they could potentially be tested as a sub-unit PvMSP1 vaccine. In the present study, a battery of monoclonal antibodies (Mabs) was produced against the N-terminal region of the PvMSP1 in an attempt to determine whether these N-terminal ICBs contained all the epitopes exposed on the native molecule. The results suggest that the most terminal ICB2 and ICB3 blocks are not exposed on the surface of the PvMSP1 native molecule and clearly eliminate the possibility of considering the N-terminal domains as unique components of a sub-unit PvMSP1 vaccine candidate.

The protozoan parasite Toxoplasma gondii comprises three clonal lineages that are associated with the clinical outcome in infected individuals. Whereas group C strains are mainly found in animals, group A and B strains are associated with human disease (Howe and Sibley, 1995). An increased level of transcripts of the tachyzoite-specifically expressed gene SAG1 could be identified in group A T. gondii strains compared to group B strains. Since SAG1-mediated host-cell invasion seems to be important for parasite replication, the observed higher replication rate in group A T. gondii strains might explain the association with clinically overt symptoms at the acute stage in patients who are infected with this group of parasite strains. The presence of external stress factors, such as interferon-gamma (IFN-gamma)-mediated nitric oxide (NO) formation has been identified to stabilize the cyst stage, most likely by activation of promoter(s) which drive the expression of genes encoding bradyzoite-specific antigens. Reactivation of chronic toxoplasmosis thus might occur in the absence of external stress factors, as has been observed in AIDS patients with decreases levels of IFN-gamma. Since group B T. gondii strains might form more cysts in infected individuals due to an increased potential to convert into bradyzoites, reactivation with resulting toxoplasmic encephalitis could be a more common event in those AIDS patients who were infected with persistent cysts of this group of parasite strains.

The protozoan parasite Toxoplasma gondii provides a model system for studying invasion by intracellular parasites belonging to the phylum Apicomplexa. Taking advantage of the versatility of T. gondii for genetic and cell biological studies, we have shown that parasite motility and cell invasion are powered by an actin-myosin based motor in the parasite. Unlike bacterial cell uptake, parasite invasion does not involve significant alterations in the host cell cytoskeleton. Instead, invasion is an active process of penetration into the host cell by the parasite. The force for cell penetration is provided by a unique form of substrate-dependent motility termed gliding. Gliding motility is characterized by the rearward capping of surface membrane proteins that propels the parasite forward in a helical spiral. Both actin and myosin are localized beneath the plasma membrane in the parasite where they presumably combine to produce the force necessary for motility. During cell invasion, the rearward capping of cell surface receptors envelopes the parasite in a unique vacuole derived from the host cell plasma membrane. This system offers insights into force generation and motility in a simple organism that is also an important human pathogen.

Like Malaria sporozoites and the circumsporozoite protein, remnants of lipoproteins are rapidly cleared from the circulation and enter hepatocytes. Here we review the evidence that the same set of liver heparan sulfate proteoglycans are the initial binding sites of malaria sporozoites and the lipoprotein remnants.

We have found that infection with the large extracellular parasite S. mansoni leads to the development of a type 1 CD8+ T cell response. While there are many poorly understood aspects of this immune response, our working hypothesis is that it functions primarily to regulate the parasite egg-antigen induced Th2 response, which itself is responsible for circumoval granunuloma formation. This view of the activity of CD8+ cells mirrors Bloom and colleagues' postulate that type 2 CD8+ cells function to regulate Th1 responses. Since it is well recognized that Th1 and Th2 cells can cross regulate each other, why should a type 1 CD8+ rather than a Th1 response be used for the regulation of the Th2 response during schistosomiasis? The answer to this may in part lie in the apparent dependence of the type 1 CD8+ cells on IL-4. Because of this, there is little likelihood for the over-production of IFN-gamma (a potentially dangerous proinflammatory cytokine) and "suppression" is provided only when needed. Th1 cells have no such dependence on IL-4 for IFN-gamma production. Current work in the laboratory is directed towards testing the various hypotheses put forward here.

The course of infection with Leishmania parasites is determined by the type of the developing CD4+ T cell immune response. Macrophages and Langerhans cells/dendritic cells play a decisive role in the interaction between the parasites and the host's immune system because they serve as host cells, as accessory cells that present parasite antigen, deliver costimulatory signals and secrete cytokines modulating the T cell activity and as effector cells eliminating the microorganisms. Therefore, we put particular emphasis on characterizing the role of these cells in cutaneous leishmaniasis and the factors regulating their activities. Our results show that (1) expression of the chemokine monocyte chemoattractant protein 1 (MCP-1) is associated with macrophage infiltration into the lesion and stimulation of leishmanicidal activity, (2) Langerhans cells are required for the transport of Leishmania from the infected skin to the draining lymph node and initiation of the specific T cell immune response in the early phase of infection, (3) lymph node dendritic cells containing persistent parasites may be involved in the maintenance of specific immunity, (4) Langerhans cells are able to present L. major LPG to T cells and (5) treatment of mice with antigen-pulsed Langerhans cells induces protective immunity against cutaneous leishmaniasis.

One of the most prominent functions of nitric oxide (NO) is its participation in antimicrobial and antiviral defense. This paper summarizes the evidence for this function and compiles the infectious agents which are currently thought to be controlled via high out-put generation of NO as it occurs in activated macrophages and other cells expressing the inducible isoform of NO-synthase (iNOS, NOS-2). Several less appreciated forms of interaction between NO and microbes will also be reviewed, including the role of NO as an immunosuppressive or tissue-destructive molecule during the course of infections, the regulation of microbial antioxidant systems by host cell-derived NO, the contribution of NO to parasite stage conversion, the induction or suppression of macrophage iNOS by microbial products, and the existence of endogenous NO synthase pathways in certain bacteria and parasites.

The genus Mycobacteria consists of over 50 species that include two of the best-known human pathogens, M. tuberculosis and M. leprae, the causes of tuberculosis (TB) and leprosy, respectively. Whereas the spread of leprosy currently appears to be under control, there are presently about 30 million active cases of TB worldwide, with an alarming increase in the number of multidrug resistant case of M. tuberculosis. As strategies for antibiotic intervention against TB become more limited, it is imperative to develop new therapeutic approaches against this oppressive disease. One promising avenue is to characterize the host genes and gene products which regulate resistance to mycobacterial infections. In the mouse, resistance and susceptibility to intracellular growth of Mycobacteria in macrophages is controlled by the Bcg (Nramp1) gene, which has now been cloned and shown to encode a macrophage transmembrane protein with a putative transporter function. Sequencing of Nramp1 revealed that susceptibility to infection is associates with a single, nonconservative glycine to aspartic acid substitution at position 169 (G169D). Although the intracellular location of the Nramp1 protein in macrophages has not yet been determined, a phagosomal site has been postulated. Consistent with the proposed role of Nramp1 in macrophage activation, recent studies of the Nramp1 promoter region have revealed consensus sequences associated with responsiveness to IFN-gamma and LPS. Finally, a total of 11 polymorphisms have been identified within the human NRAMP1 gene which are being used to test for linkage of NRAMP1 alleles with human susceptibility to TB and leprosy.

Here we describe a novel methodology for the investigation of the intracellular pH of P. falciparum. This method is based on a fluorescent dye with pH-dependent spectral properties, which can be monitored using a digital imaging system. This non-invasive method allows the cytoplasmic pH of single, living P. falciparum parasites to be measured while still within the host erythrocyte. It was found that schizonts from the P. falciparum clone D10 have a cytoplasmic pH of 7.18 to 7.23, differing slightly on the buffering system used. The pH of uninfected erythrocytes is 7.10 +/- 0.05. This method offers an opportunity to study the parasite's physiology and define transport mechanisms essential for parasite growth.

The cell surface of Leishmania parasites is coated by a highly unusual glycocalyx which varies markedly during the parasite life cycle. The predominant molecule on the extracellular promastigote (sandfly) stage is a complex lipophosphoglycan (LPG), which together with a number of GPI-anchored proteins and a family of low molecular weight glycoinositolphospholipids (GIPLs), forms a morphologically distinct protective coat over the plasma membrane. The structure of the LPG has been shown to vary in different species and during promastigote development in the sandfly. This polymorphism is thought to be important in allowing Leishmania parasites to colonize a range of insect hosts, and in facilitating the regulated migration of promastigotes along the sandfly alimentary canal. Stage-specific changes in LPG are also involved in preadapting promastigotes to life in the mammalian host. This complex glycocalyx coat is absent from the amastigote stage that proliferates in the phagolysosomes of mammalian macrophages, as the expression of both the LPG and GPI-anchored proteins is massively down-regulated. Instead, the plasma membrane of amastigotes is coated by a densely packed layer of parasite-derived GIPLs and host-derived glycosphingolipids. We propose that the down-regulation of the promastigote macromolecules and the acquisition of host glycolipids by amastigotes represents an important strategy to avoid detection by specific and non-specific components of the immune system.

The intracellular protozoan parasite Toxoplasma gondii is an important opportunistic pathogen in animals and man. In parallel to its clinical significance, T. gondii is also receiving considerable attention as an attractive model organism for intracellular parasitism. Regulation of gene expression at various levels underlies the intricate interplay between the parasite and its host cell, as well as the interconversions between different life-stages. In this article we will discuss some of what is currently known about gene organization and gene regulation in T. gondii as well as some of the tools available to dissect the parasite at a molecular level.

The wide occurrence of molecular rearrangements associated with expression of specific members within multigene families led us to investigate whether this also happens during antigenic variation of malaria parasites. We have investigated here the Pf60 multigene family restriction patterns of four distinct variants of the Plasmodium falciparum Palo Alto line propagated in Saimiri monkeys. The O and its cloned Oc variant both express the O serotype, while the R variant (derived from O parasites) and the Vc variant (derived from Oc parasites) express distinct serotypes. We show that a specific modification of the restriction pattern could be associated with antigenic switching in this line. The DNA of the variants which expressed the O serotype (O and Oc) had a specific 5.5 kb Hind III/Eco RI restriction fragment which was absent from the R or Vc parasite DNA, whereas both R and Vc DNA presented a 3.5 kb Hind III/Eco RI restriction fragment, which was absent from the O and Oc parasites. These results indicate that both expression and silencing of the O serotype were associated with specific restriction patterns, suggesting that some molecular rearrangement or some modification of the DNA might control expression of the variant surface antigen in malaria parasites.

African trypanosomes can escape destruction by the immune system of their mammalian host by antigenic variation of the trypanosome surface coat. This coat is mainly composed of a single protein species, the Variant Surface Glycoprotein or VSG. The genes for VSGs are expressed in a polycistronic telomeric expression site together with at least eight expression site-associated genes (ESAGs). Trypanosomes may switch coat either by replacing the VSG gene in the active expression site by a different one, or by activating another expression site with concomitant silencing of the previously active one. Here we review our present knowledge of antigenic variation in Trypanosome brucei. We focus on four questions: How do trypanosomes switch from one VSG gene expression site to another one? What is the role of the novel base J in silencing expression sites? What is the functional significance of the antigenic variation of the heterodimeric transferrin receptor encoded by two ESAG genes? Why do trypanosomes have multiple expression sites at all?