Differential serodiagnosis for cystic and alveolar echinococcosis using fractions of Echinococcus granulosus cyst fluid (antigen B) and E. multilocularis protoscolex (EM18).
Echinococcus granulosus cyst fluid and E. multilocularis protoscolex extract were fractionated by a single step of preparative isoelectric focusing, resulting in an antigen B-rich fraction (8-kD) and an Em18-rich fraction, respectively. The usefulness of both fractions for differential serodiagnosis of cystic (CE) and alveolar (AE) echinococcosis was evaluated by a large-scale immunoblot analysis on a battery of 354 serum samples. These included 66 from AE patients originating from four different endemic areas, 173 from CE patients originating from seven different endemic areas, 71 from patients with other parasitic diseases, 15 from patients with hepatomas, and 29 from healthy individuals. In an immunoblot with the antigen B-rich fraction, 92% (158 of 173) of the CE sera as well as 79% (52 of 66) of the AE sera reacted with the 8-kD subunit. No cross-reactivity occurred with any sera from patients with cysticercosis, other parasitic diseases, or with hepatomas, or from healthy controls. In an immunoblot with the Em18-rich fraction, all but two sera from AE patients (64 of 66, 97%) recognized Em18, and only nine of 34 CE sera from China reacted with it. All other (139) CE sera from six other countries were negative as were all (115) other non-echinococcosis sera. These findings indicate that antigen B (8-kD) is not species-specific for E. granulosus but is genus-specific for Echinococcus, and that the Em18 antigen is a reliable serologic marker for species-specific differentiation of AE from CE. (+info)
Development of a serologic assay to detect Taenia solium taeniasis.
We developed a serologic assay to identify adult Taenia solium tapeworm carriers using excretory/secretory (TSES) antigens collected from in vitro cultured T. solium tapeworms. To identify taeniasis-specific antigens we used an immunoblot assay with serum samples from T. solium tapeworm carriers and cysticercosis patients. Antigens were identified that reacted with antibodies present in serum samples from taeniasis cases and not with those from cysticercosis patients. Using serum samples collected from persons with confirmed T. solium tapeworm infections, the test was determined to be 95% (69 of 73) sensitive. Serum samples (n = 193) from persons with other parasitic infections, including T. saginata tapeworm infections, do not contain cross-reacting antibodies to TSES, indicating that the assay is 100% specific. These data suggest that the immunoblot assay using TSES antigens can be used to identify persons with current or recent T. solium tapeworm infections and provides a new, important tool for epidemiologic purposes, including control and prevention strategies. (+info)
Granulomatous inflammatory response to recombinant filarial proteins of Brugia species.
The lymphatic inflammatory response in Brugia-infected jirds peaks early during primary infections and then decreases in severity as judged by the numbers of lymph thrombi present within these vessels. Antigen-specific hypersensitivity reactions in these animals was measured by a pulmonary granulomatous inflammatory response (PGRN) induced by somatic adult worm antigen (SAWA)-coated beads, and by cellular proliferative responses of renal lymph node cells. The kinetics of these responses temporally correspond to lymphatic lesion formation. The importance of any single antigen to the induction of this inflammatory response has not been elucidated. In this study, the PGRN was used to measure the cellular immune response to four recombinant filarial proteins during the course of a primary B. pahangi infection. These proteins were BpL4, glycoprotein (glutathione peroxidase) gp29, heat shock protein (hsp) 70, and filarial chitinase. All were fusion proteins of maltose-binding protein (MBP). Control beads included those coated with diethanolamine (DEA), SAWA, or MBP. The measurements of PRGN were made at 14, 28, 56, and > 150 days postinfection (PI) in infected jirds, in jirds sensitized with SAWA, and in uninfected jirds. The secretory homolog of glutathione peroxidase gp29 was the only recombinant protein tested that induced a significantly greater PGRN (P < 0.05) than controls. This was seen at 28 days PI. These observations indicate that gp29 may be part of the worm antigen complex that induces an early inflammatory response, a response similar to that observed with SAWA. These studies indicate that this approach is useful in investigating the functional ability of specific proteins in the induction and down-regulation of immune-mediated inflammatory responses elicited by filarial parasites. Absence of a granulomatous response to the other recombinant proteins used may be related to the nature and sensitivity of the assay used or the character of recombinant proteins tested. (+info)
Inhibitory effect of artemether on proteinase of Schistosoma japonicum.
AIM: To study the effect of artemether (Art) on the thio proteinase ("hemoglobinase", Hem) of Schistosoma japonicum. METHODS: Hem was extracted from S japonicum adults. The inhibitory effect of Art on the activity of Hem to degrade human hemoglobin (Hgb) was examined with UV-photometer at 280 nm, SDS-PAGE and scanning at 600 nm on a chromoscanner. RESULTS: Human Hgb was degraded at pH 4.0 by the Hem. The activities of Hem preincubated at 37 degrees C with Art 0.14, 1.4, and 14 mmol.L-1, were inhibited by 30.2%, 39.8%, and 45.0%, respectively. CONCLUSION: Art possesses an inhibitory effect to Hem of S japonicum. (+info)
Interleukin-10 and antigen-presenting cells actively suppress Th1 cells in BALB/c mice infected with the filarial parasite Brugia pahangi.
Infection with the third-stage larvae (L3) of the filarial nematode Brugia results in a Th2-biased immune response in mice and humans. Previously we have shown that the production of interleukin 4 (IL-4) is critical for down-regulating polyclonal Th1 responses in L3-infected mice. However, the in vitro neutralization of IL-4 did not fully recover the defective polyclonal Th1 responses, nor did it result in the production of any antigen (Ag)-specific Th1 cytokines, suggesting that perhaps infection with L3 does not result in priming of Th1 cells in vivo. In this study, we analyzed the role of IL-10 and Ag-presenting cells (APCs) in the spleen as additional factors controlling the Th2 bias in infected mice. Our data show that IL-10 and APCs also contribute to the suppression of mitogen-driven Th1 responses of spleen cells from infected mice. In addition, the neutralization of IL-10 or the replacement of the resident APC population from spleen cell cultures resulted in the production of Ag-specific Th1 cytokines. Irradiated spleen cells from either L3-infected or uninfected mice were able to restore Ag-specific Th1 responses in vitro. Therefore, it appears that Brugia-reactive Th1 cells are primed following infection with L3, but are actively suppressed in vivo by a mechanism that involves IL-10 and the resident APC population, but not IL-4. These results indicate that a complex interplay of cytokines and cell populations underscores the Th2-polarized response in L3-infected mice. (+info)
A novel 62-kilodalton egg antigen from Schistosoma mansoni induces a potent CD4(+) T helper cell response in the C57BL/6 mouse.
In infection with Schistosoma mansoni, hepatic granuloma formation is mediated by CD4(+) T helper (Th) cells sensitized to schistosomal egg antigens. There is considerable variation among infected individuals with respect to both severity of disease and the T-cell response to egg antigens. In the BL/6 mouse, the egg granulomas are relatively small and the relevant sensitizing egg antigens are largely unknown. We investigated the CD4(+) Th cell response of infected BL/6 mice to egg antigens fractionated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and found a prominent lymphoproliferative response to be directed against a 62-kDa component. With the aid of a specific T-cell hybridoma, 4E6, the 62-kDa antigen was isolated; following partial digestion with endoproteinase Glu-C, an internal amino acid sequence was found to be identical with one present in the enzyme phosphoenolpyruvate carboxykinase (PEPCK) of the organisms Caenorhabditis elegans and Treponema pallidum and to differ by one residue from PEPCK of various other species. In CD4(+) Th cells from 7.5- 8.5-week-infected BL/6 mice, the purified 62-kDa molecule elicited a potent proliferative response which, based on cytokine analysis, was of a mixed Th-1 and Th-2 type. Our results reveal a novel egg antigen of particular prominence in the BL/6 mouse and suggest that the immune response in schistosomiasis is a product of sensitization to egg antigens that may vary considerably in immunogenicity from strain to strain. (+info)
Vaccination with cathepsin L proteinases and with leucine aminopeptidase induces high levels of protection against fascioliasis in sheep.
The potential of different parasite proteinases for use as vaccine candidates against fascioliasis in sheep was studied by vaccinating animals with the cathepsin L proteinases CL1 and CL2 and with leucine aminopeptidase (LAP) purified from adult flukes. In the first trial, sheep were immunized with CL1 or CL2 and the mean protection levels obtained were 33 and 34%, respectively. Furthermore, a significant reduction in egg output was observed in sheep vaccinated either with CL1 (71%) or with CL2 (81%). The second trial was performed to determine the protective potential of the two cathepsin L proteinases assayed together, as well as in combination with LAP, and of LAP alone. The combination of CL1 and CL2 induced higher levels of protection (60%) than those produced when these enzymes were administered separately. Those sheep that received the cocktail vaccine including CL1, CL2, and LAP were significantly protected (78%) against metacercarial challenge, but vaccination with LAP alone elicited the highest level of protection (89%). All vaccine preparations induced high immunoglobulin G titers which were boosted after the challenge infection, but no correlations between antibody titers and worm burdens were found. However, the sera of those animals vaccinated with LAP contained LAP-neutralizing antibodies. Reduced liver damage, as assessed by the level of the liver enzyme gamma-glutamyl transferase, was observed in the groups vaccinated with CL1, CL2, and LAP or with LAP alone. (+info)
Tolerization of mice to Schistosoma mansoni egg antigens causes elevated type 1 and diminished type 2 cytokine responses and increased mortality in acute infection.
The granuloma that surrounds the Schistosoma mansoni egg is the cause of pathology in murine schistosomiasis, and its formation is driven by egg Ag-stimulated type 1 and type 2 cytokines. To determine the role of egg-driven immune responses during schistosome infection we rendered CBA/Ca mice unresponsive to schistosome eggs by combined cyclophosphamide treatment and thymectomy. In the early acute stages of schistosome infection, egg-tolerized mice suffered high mortalities. Granuloma size and deposition of collagen in the liver were significantly reduced in egg-tolerized mice. Similarly, limited granuloma responses were detected in the intestines of these mice, and this was associated with a >90% reduction in egg excretion. Histologically, egg-tolerized mice had exacerbated hepatocyte damage, with extensive microvesicular steatosis. Elevated plasma transaminase levels confirmed the damage to hepatocytes. Infected egg-tolerized mice had impaired proliferation responses to egg Ag but intact responses to worm Ag. Tolerized mice had diminished Ab responses to egg Ag and had a type 1 cytokine isotype pattern to worm Ag, with elevated IgG2a and diminished IgG1 and IgE. Egg-tolerized mice failed to down-regulate type 1 cytokines that are normally elicited during early schistosome infection. Hepatic granuloma cells from egg-tolerized mice were also type 1 cytokine dominated, with elevated frequencies of Tc1/Th1 and reduced Tc2/Th2 cells. This study demonstrates that mice tolerized to schistosome eggs have elevated type 1 cytokine responses with diminished type 2 responses and reduced anti-egg Ab during schistosome infection, and these effects are detrimental to the host. (+info)