Cryptosporidiosis, caused by the apicomplexan parasite Cryptosporidium parvum, has become a well recognized diarrheal disease of immunodeficient humans and other mammals throughout the world. Specific therapy and immunoprophylaxis are currently unavailable, but passive immunization with C. parvum-specific monoclonal antibodies (mAbs) has demonstrated efficacy in immunocompromised hosts. In particular, mAbs eliciting the circumsporozoite precipitate (CSP)-like reaction protected against C. parvum infection. The circumsporozoite-like antigen (CSL), an ∼1,300 kDa apical glycoprotein of sporozoites and merozoites, is the molecular species mechanistically bound by mAbs having the ability to elicit the CSP-like reaction. These findings indicated that CSL has a functional role in sporozoite infectivity. In the present study, a quantitative in vitro sporozoite infectivity assay was developed to evaluate neutralizing activity of mAbs. 3E2, a mAb which elicited the CSP-like reaction, demonstrated the greatest level of neutralizing activity against C. parvum infections. Here, I report that CSL has properties consistent with being a sporozoite ligand for epithelial cells. For these studies, CSL was isolated from sporozoites by isoelectric focusing (IEF). The 1,200-1,400 kDa Mᵣ region containing CSL in SDS-PAGE of sporozoites was comprised of 31 species when analyzed by two-dimensional electrophoresis. Eight species were present in IEF-isolated CSL. CSL specifically bound to Caco-2 cells in a dose-dependent, saturable, and self-displaceable manner with high affinity. CSL bound to Caco-2 cells inhibited the attachment and invasion of sporozoites in a dose-dependent manner. Characterization of the epitope recognized by 3E2 in a dot immunoblot indicated a β-glucose component. Sporozoites having undergone the CSP-like reaction after incubation with CSL-reactive 3E2, did not attach to or invade Caco-2 cells. These findings indicated that at least 1 of 8 CSL species isolated by IEF was a sporozoite ligand. The Caco-2 cell receptor for CSL was comprised of 16, 51, and 85 kDa molecules. Further, sporozoites incubated with isolated Caco-2 receptors failed to attach to and invade Caco-2 cells. Finally, CSL bound distinctly to receptors present on calf ileum. Based on these findings, I concluded that CSL ligand function is amenable to targeted disruption by CSL-reactive mAbs and that it is a rational target for immunization against cryptosporidiosis.
| Identifer | oai:union.ndltd.org:arizona.edu/oai:arizona.openrepository.com:10150/282744 |
| Date | January 1998 |
| Creators | Langer, Rebecca Christine, 1972- |
| Contributors | Riggs, Michael W. |
| Publisher | The University of Arizona. |
| Source Sets | University of Arizona |
| Language | en_US |
| Detected Language | English |
| Type | text, Dissertation-Reproduction (electronic) |
| Rights | Copyright © is held by the author. Digital access to this material is made possible by the University Libraries, University of Arizona. Further transmission, reproduction or presentation (such as public display or performance) of protected items is prohibited except with permission of the author. |
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