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Globally, Cryptosporidium infection continues to be a significant health problem where it is recognized as an important cause of diarrhoea in both immunocompromised and immunocompetent people. In developing countries persistent diarrhoea is the leading cause of death in children younger than five years of age, where it accounts for 30 to 50 percent of those deaths. Encouragingly an increasing number of investigations in developing countries employ molecular tools, significantly improving the quality of epidemiological information. This improved Cryptosporidium monitoring, with appropriate molecular methods, in surface water, livestock, wildlife and humans, will increase current knowledge of infection and transmission patterns, and ultimately help to control Cryptosporidium via improved risk assessments in the future.

Gram-negative bacterial proteins which are exported from the cytosol to the external environment by the type V secretion system are also known as autotransporters. Once translocated to the periplasmic compartment by the sec-dependent general secretory pathway, their C-terminal domain forms a pore through which the N-terminal domain travels to the outer membrane without the need of other accessory proteins. MisL (protein of membrane insertion and secretion) is a protein of unknown function located in the pathogenicity island SPI-3 of Salmonella enterica and classified as an autotransporter due to its high homology to Escherichia coli AIDA-I. In the present work, the MisL C-terminal translocator domain was used to display the immunodominant B-cell epitope of the circumsporozoite protein (CSP) from Plasmodium falciparum on the surface of Salmonella enterica serovar Typhimurium (serovar Typhimurium SL3261) and serovar Typhi (serovar Typhi CVD 908). The MisL ␤ domain was predicted by alignment with AIDA-I, amplified from serovar Typhimurium SL3261, cloned in a plasmid fused to four repeats of the tetrapeptide NANP behind the Escherichia coli heat-labile enterotoxin B subunit signal peptide to ensure periplasmic traffic, and expressed under the control of the anaerobically inducible nirB promoter. The fusion protein was translocated to the outer membrane of both bacterial strains, although the foreign epitope was displayed more efficiently in serovar Typhimurium SL3261, which elicited a better specific antibody response in BALB/c mice. More importantly, antibodies were able to recognize the native CSP in P. falciparum sporozoites. These results confirm that MisL is indeed an autotransporter and that it can be used to express foreign immunogenic epitopes on the surface of gramnegative bacteria.

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Hexagonal phase (H II )-preferring lipids such as phosphatidate, cardiolipin, and phosphatidylserine form nonbilayer molecular arrangements in lipid bilayers. While their presence in biological membranes has not been established, in vitro studies suggest that alterations in membrane properties modify their function. In this study, antiphospholipid monoclonal antibodies were developed against nonbilayer structures. One of the monoclonal antibodies identifies nonplanar surfaces in liposomes and in membranes of cultured cells. These results are the first evidence that natural membranes maintain a fragile balance between bilayer and nonbilayer lipid arrangements. Therefore, these antibodies can be used to evaluate the role of H II -preferring lipids in the modulation of membrane activities. Our studies demonstrated that nonplanar surfaces are highly immunogenic. Although these structures are normally transient, their formation can be stabilized by temperature variations, drugs, antibiotics, apolar peptides, and divalent cations. Our studies demonstrated that abnormal exposure of nonbilayer arrangements may induce autoimmune responses as found in the antiphospholipid syndrome.

Protease secretion by Giardia duodenalis trophozoites upon interaction with epithelial cells and its association with the parasite adhesion was studied in co-cultures of parasites with IEC6 epithelial cell monolayers in the presence or absence of protease inhibitors. Proteolytic activity in supernatants from trophozoites was enhanced when they were co-cultured with IEC6 cells. This activity was strongly inhibited by pre-incubation of live trophozoites with E-64 and TPCK and a concomitant inhibition of parasite adhesion to IEC6 cells was observed. These data suggest that trophozoites secrete cysteine-type proteases that play a role in the adhesion of G. duodenalis to epithelial cells.Key words: Giardia duodenalis -protease secretion -adhesion Giardiasis caused by the flagellated, intestinal protozoan parasite Giardia duodenalis (syn. G. lamblia, G. intestinalis) manifests as acute and chronic diarrhoea in communities of both developing and developed countries. It is also a leading cause of defined waterborne diarrhoea worldwide (Adam 1991). Infection results from ingestion of cysts, most commonly from faecal contamination of water and food whereas community-wide outbreaks result from contaminated water supplies. After excystation in the upper small intestine, adhesion of trophozoites to epithelial intestinal cells represents the first step in the pathogenesis of the disease. The use of IEC6, an epithelial cell line derived from rat small intestine, is a well validated model in which the inhibitory effect of both actin inhibitors and chelating agents on the attachment of G. duodenalis trophozoites has been previously demonstrated (McCabe et al. 1991). It is also well recognized that the adhesion process is a multi-factorial event that involves a suction force of the adhesive disk (Knaippe 1990), a mechanical process related to contractile proteins of the adhesive disk and the ventrolateral flange (Chávez et al. 1995) and surface molecules that allow the binding of trophozoites to receptors to host epithelial cells (Pegado & Souza 1994). It has been postulated that proteases may be involved in this process. Several studies have demonstrated protease activity in total extracts of Giardia trophozoites or in parasite vacuoles (Feely & Dyer 1987, Hare et al. 1989) including the Portland 1 (P1) strain cultured in vitro, with the predominance of cysteine, serine, and aspartic protease activities (Williams & Coombs 1995). It has also been shown that cysteine proteases of P1 strain degraded protein substrates such as gelatin, collagen, albumin, and azocasein while serine proteases were mainly active against haemoglobin (Coradi & Guimarães 2006). In addition, the presence of cysteine protease activities in excretory/secretory (E/S) products of P1 strain has also been reported (Jiménez et al. 2000). These E/S products are associated with mucosal injuries observed in murine giardiasis (Jiménez et al. 2004). However the possible relation between protease secretion and the adhesion of G. duodenalis to epithelial cells has not b...

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The parasitic nematode Trichinella spiralis, an aetiological agent of the disease known as trichinellosis, infects wild and domestic animals through contaminated pig meat, which is the major source for Trichinella transmission. Prevention of this disease by interrupting parasite transmission includes vaccine development for livestock; however, major challenges to this strategy are the complexity of the T. spiralis life cycle, diversity of stage-specific antigens, immune-evasion strategies and the modulatory effect of host responses. Different approaches have been taken to induce protective immune responses by T. spiralis immunogens. These include the use of whole extracts or excretory-secretory antigens, as well as recombinant proteins or synthesized epitopes, using murine experimental models for trichinellosis. Here these schemes are reviewed and discussed, and new proposals envisioned to block the zoonotic transmission of this parasite.