This proposed project intends to evaluate the immunostimulating activity of saponins extracted from plant species endogenous to Kazakstan; to utilize these saponins with the Kazaskstany co-investigator’s unique MESK detergent to produce an antigen delivery system based on immunostimulating complexes (ISCOMs) that contain natural and recombinant coccidia (genus; Eimeria) protozoan parasite antigens; and to determine the potential of ISCOM delivery in the development of a subunit vaccine against these coccidial parasites of poultry. Comparisons will be made with ISCOMs produced with the commercially available glycoside Quil A to determine if the ISCOMs produced by the Kazakstany saponins have equal or greater immunostimulating activity. The project will combine the expertise of scientists from the Institute of Microbiology and Virology in Almaty, Kazakstan, the Scientific Agriculture Research Institute, Zhambil Region, Kazakstan and the Beltsville Agricultural Research Center, Beltsville, MD to extract and purify saponins obtained from Kasakastany plants for ISCOMs formulations, and then develop and evaluate an ISCOM vaccine delivery system for use in immunizing chickens. The coccidial protozoan parasites are estimated to cost the poultry industry world-wide over $1 billion annually in feed loss and lower bird performance, and control of this parasite has centered on the use of anticoccidial compounds mixed in the feed. Extensive use of these compounds has resulted in the development of drug resistance by the parasites and subsequent loss in disease control. The cost involved to discover and clear a new anticoccidial compound for use in food animals has become so cost prohibitive that it is now estimated that no new compounds will be developed for at least 10 years. This potential for loss of anticcoccidial control has caused increased concern in the poultry industry, and stimulated interest by the industry for development of successful anticoccidial vaccines to help insure continued bird productivity. To date, protein subunit vaccines prepared with isolated natural or recombinant coccidial antigens have shown limited efficacy with chickens in eliciting a protective immune response against coccidial challenge infections, and have required use of adjuvants or repeated boost immunizations to consistently elicit only partial immunity. Use of ISCOMs as an oral or intranasal immunization vector for vaccination with natural or recombinant coccidial antigens represents a new direction in antigen delivery that could increase elicited protective immune response, and may also have potential marketability in vaccine development for future use in the animal health industry. ISCOMs are stable particles made up of plant saponins and lipids arranged into multimolecular structures that contain entrapped antigens, and have been shown in bacterial and viral models to give enhanced immune responses by various delivery routes at both the humoral and cellular level. These unique abilities make ISCOMs excellent candidates for use in the poultry industry with parasite, viral and bacterial vaccines. The objectives listed below for this project are designed to evaluate the development and potential for use of ISCOMs as an effective delivery system for the immunization of commercially produced poultry.
Project Objectives:
1. Isolation, gel filtration chromotography and HPLC purification of saponins from local Kazakstany plants, electron microscopy evaluation of ISCOM formation, and determination of the toxicity of isolated saponins in animal models.
2. Production of isolated natural and recombinant coccidia antigens with the study of their physical-chemical (ie. Molecular structure and resultant hydrophilic and hydrophobic characteristics) and immuno-chemical (ie. Immunoblotting and antibody-antigen binding) properties.
3. Organization of immunostimulating complexes (ISCOMs) from Kasakastany saponins or commercial glycoside Quil A containing natural or recombinant coccidia antigens by use of dialysis technology based on use of MESK detergent.
4. Determination of humoral immune response after subcutaneous, intranasal and oral immunization with Kazakstany saponins or glycoside Quil A ISCOMs containing coccidia antigens in mouse and chicken models. Use of mouse model to determine cellular immune response to ISCOMs immunization by the same vaccination routes.
5. Evaluation of degree of elicited protective immune response in chickens vaccinated with selected ISCOMs preparations to coccidial challenge in caged battery trials.
6. Confirmation of use of selected ISCOM preparations with highest immunogenetic activity in vaccinated broiler chickens challenged with coccidia in 7-week grow-out floor-pen studies.
