Nak-Won Choi


In comparison with whole organism vaccines, subunit vaccines may be safer for immunization but may lack sufficient immunogenicity to provide complete immunity to the pathogen. To resolve this problem, bacterial and plant enterotoxin B subunit adjuvants containing a variety of receptor-binding properties were used to enhance the immunogenicity of rotavirus subunit vaccines. Enterotoxin B subunit adjuvants were employed to enhance protection against virus infection. Pentameric cholera toxin B subunit (CTB), shiga toxin-1 B subunit (STB) and monomeric ricin toxin B subunit (RTB) molecules were genetically linked to a 90 amino acid peptide from the simian rotavirus (SA11) nonstructural protein NSP4 or to the rotavirus outer capsid protein VP7. Lectin binding assays were used to demonstrate receptor binding activity of the ligand-antigen fusion proteins synthesized in E. coli. Mice co-fed RTB and NSP490 protein or heat denatured NSP490::RTB fusion protein generated higher serum anti-NSP490 IgG titers than mice inoculated with the NSPT90 antigen alone. Serum anti-NSP490 IgG and intestinal anti-NSP490 IgA titers were the highest in mice immunized with native NSP490::RTB or STB::NSP490 fusion proteins. Serum anti-NSPT90 IgG isotype analysis and secreted cytokine assays from splenocytes isolated from mice immunized with NSP490::RTB and STB::NSP490 fusion proteins generated elevated IFN-γ and IL-4 levels and increased IgG2a antibody titers confirming fusion protein stimulation of a dominant Th1 cell mediated immune response. Diarrhea in SA11 rotavirus challenged suckling neonates from dams immunized with NSP490::RTB and STB::NSP490 fusion proteins was dramatically reduced both in severity and duration in comparison with rotavirus challenged neonates from unimmunized mice, demonstrating the protective efficacy of ligand-antigen fusion proteins synthesized in bacteria.

The feasibility of using transformed plants for production of correct posttranslational modification of adjuvanted subunit rotavirus antigens was assessed. Immunoblot analyses and lectin-receptor binding enzyme-linked immunosorbent assays showed that CTB::NSP490, CTB::VP7 and VP7::RTB fusion proteins of expected molecular mass were synthesized in transformed potato leaf and tuber tissues. The production of adjuvant ligand-rotavirus antigen fusion proteins in both bacteria and plant tissues demonstrates the feasibility of using prokaryotic and eukaryotic systems for production of adjuvanted viral antigens for generating enhanced immunity against infectious rotavirus disease.

LLU Discipline





Graduate School

First Advisor

William H. R. Langridge

Second Advisor

Penelope J. Duerksen-Hughes

Third Advisor

E. Clifford Herrmann

Fourth Advisor

James D. Kettering

Fifth Advisor

Thomas A. Linkhart

Degree Name

Doctor of Philosophy (Medical Science)

Degree Level


Year Degree Awarded


Date (Title Page)




Library of Congress/MESH Subject Headings

Vaccines, Subunit -- immunology; Enterotoxins -- therapeutic use; Rotavirus vaccines; Rotavirus infections; Lectins -- therapeutic use; Adjuvants, Immunologic.



Page Count

xiv; 262

Digital Format


Digital Publisher

Loma Linda University Libraries

Usage Rights

This title appears here courtesy of the author, who has granted Loma Linda University a limited, non-exclusive right to make this publication available to the public. The author retains all other copyrights.


Loma Linda University Electronic Theses and Dissertations

Collection Website


Loma Linda University. Del E. Webb Memorial Library. University Archives