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Abstract: Animal schistosomiasis caused by Schistosoma bovis is a veterinary problem in many areas of the world. It affects a large number of animals and causes important economic losses in livestock production. The 22.6kDa antigen is a tegumental protein of unknown function, restricted to schistosomes. In S. bovis it has been identified in the tegument and in an excretion–secretion extract, consisting of several, non-glycosylated isoforms that are recognised by the sera of animals infected with S. bovis. The aims of the present work were to clone, sequence, express and characterize at molecular level the S. bovis 22.6 antigen (Sb22.6), as well as to assess the usefulness of the corresponding recombinant protein as a diagnostic antigen in ELISA tests for the detection of free-range cattle farms infested with S. bovis. Immunolocalization studies revealed that Sb22.6 is expressed in the tegument and some internal tissues of the adult worms, but it is not exposed on the surface of the adult worms and schistosomula. The reactivity of the recombinant Sb22.6 (rSb22.6) in ELISA against antibodies in sera from S. bovis experimentally infected hamsters and sera from free-range cattle from a S. bovis endemic area showed that the recombinant protein and the soluble extract of adult worms (SbC) exhibited a similar diagnostic performance. In addition, rSb22.6 did not show cross-reactions with antibodies against Fasciola hepatica, also a frequent trematode parasite in cattle. The rSb22.6 antigen can be readily produced in large amounts and in a highly reproducible fashion, avoiding the types of problem that arise upon using crude extracts such as the SbC. In conclusion, this protein represents a promising epidemiological tool for the surveillance of S. bovis and may help to implement control measures in the areas and farms were the parasite is present. [Copyright &y& Elsevier]

Abstract: Annexins belong to an evolutionarily conserved multigene family of proteins expressed throughout the animal and plant kingdoms. Although they are soluble cytosolic proteins that lack signal sequences, they have also been detected in extracellular fluids and have been associated with cell surface membranes, where they could be involved in anti-haemostatic and anti-inflammatory functions. Schistosome annexins have been identified on the parasite''s tegument surface and excretory/secretory products, but their functions are still unknown. Here we report the cloning, sequencing, in silico analysis, and functional characterization of a Schistosoma bovis annexin. The predicted protein has typical annexin secondary and tertiary structures. Bioassays with the recombinant protein revealed that the protein is biologically active in vitro, showing fibrinolytic and anticoagulant properties. Finally, the expression of the native protein on the tegument surface of S. bovis schistosomula and adult worms is demonstrated, revealing the possibility of exposure to the host''s immune system and thus offering a potential vaccine target for the control of schistosomiasis in ruminants. [Copyright &y& Elsevier]

Abstract: Schistosoma bovis is a ruminant parasite able to survive prolonged periods in the vasculature of its host without either being cleared by the host defensive systems or inducing thrombotic or coagulation disturbances. This suggests that the parasite modulates both the immune and haemostatic host responses. Previous studies have shown that host plasminogen binds to the surface of S. bovis adult worms, and that a tegument extract from S. bovis fixes and activates host plasminogen, generating plasmin, which in turn could both inhibit blood clotting and dissolve clots. Enolase has been identified among the tegumental proteins that bind plasminogen. The aim of the present study is to determine the physiological role of the enolase found in the tegument of S. bovis adult worms as regards plasminogen-binding and activation, and to confirm its surface exposure on the parasite. The study included the cloning and sequencing of S. bovis enolase cDNA, collection of the corresponding recombinant protein and evaluation of its plasminogen-binding and activation activity, and an exploration of the expression and localization of native enolase in adult worms and lung schistosomulae. Here we show that S. bovis male adult worms express enolase on their tegumental surface and that this protein binds host plasminogen and increases its activation in the presence of host tissue plasminogen activator (t-PA). This suggests that the surface-associated enolase found here is a physiological receptor of plasminogen that plays a role in the activation of the host fibrinolytic system, most probably to avoid blood clot formation on the worm''s surface. [Copyright &y& Elsevier]

Abstract: Schistosoma bovis is a parasite of wild and domestic ruminants that is broadly distributed throughout many tropical and temperate regions of the old world. S. bovis causes severe health problems and significant economic losses in livestock, but in contrast to human schistosomes, S. bovis has been little investigated at a molecular level. Since schistosome glycans and glycoproteins can play important roles in the host–parasite interplay, the aims of the present work were: (i) to characterize the glycans expressed by adult S. bovis worms on their excreted/secreted (ES) and tegumental (TG) glycoproteins and (ii) to identify their carrier protein backbones by mass spectrometry. Using a panel of lectins and monoclonal and polyclonal anti-glycan antibodies, we observed: (i) the absence of sialic acid in S. bovis; (ii) the presence of complex-type N-glycans and LDN antennae on ES glycoproteins; (iii) the presence of glycans containing the Fucα1-2Galβ motif in many TG glycoproteins, and (iv) the presence of glycans containing the Fucα1-3GlcNAc motif on many ES and TG glycoproteins but, simultaneously, the absence of the F-LDN(-F) glycans from both the ES and TG glycoproteins. Interestingly, we also found the LewisX and LewisY antigens co-expressed on several TG isoforms of ATP:guanidino kinase and glyceraldehyde-3-phosphate dehydrogenase. Finally, by ELISA we observed the presence of antibodies against LewisX, LewisY and F-LDN(-F) in the sera of sheep experimentally infected with S. bovis. [Copyright &y& Elsevier]

The percutaneous administration of cercariae is the usual method for experimental infections with Schistosoma bovis. These procedures are laborious and have important inconveniences when working with a large number of animals, especially if they are ruminants. In the present study, the efficiency of the oral, intramuscular and subcutaneous routes are evaluated by comparison with the percutaneous route in experimental infections with S. bovis. The infections developed in hamsters and sheep were evaluated taking as a basis the parasite burden, the concentrations of eggs in tissues and the levels of anti-Schistosoma antibodies. The oral infection failed in both hamsters and sheep. The administration of the cercariae by the intramuscular route was effective in sheep, developing infections of intensity similar to that of the infections acquired percutaneously. In hamsters, on the contrary, although all the animals developed the infection, they were very little intense. The injection of the cercariae by the subcutaneous route induces acceptable infections in hamsters and can also be an alternative route to percutaneous exposure. The levels of the anti-Schistosoma bovis antibodies detected in hamster and sheep were proportional to the number of worms present, which shows that the humoral response is a good indicator of the intensity of the infections. It can be concluded that the intramuscular route is a good alternative to the percutaneous route for experimental infections of sheep with S. bovis. Likewise, the subcutaneous route can also substitute, with some advantages, the percutaneous infections in hamsters. [Copyright &y& Elsevier]