Publication Date


Date of Final Oral Examination (Defense)


Type of Culminating Activity


Degree Title

Master of Science in Biology



Major Advisor

Juliette K.Tinker, Ph.D.


Kristen Mitchell, Ph.D.


Troy Rohn, Ph.D.


The development of adjuvants that can promote the delivery of purified subunit vaccines by mucosal routes, such as the nose or the mouth, is recognized as a top priority for vaccine research. The bacterial enterotoxins; cholera toxin (CT) and E.coli heat-labile toxin (LTI), have long been recognized as powerful adjuvants with the ability to stimulate specific immune responses to co-administered antigens when delivered to mucosal surfaces. Shiga toxin 1 (ST1) and pertussis toxin (PT) are structurally homologous bacterial toxins secreted by Escherichia coli 0157:H7 and Bordetella pertussis respectively. ST1 and PT also have reported adjuvant activity but it is less well characterized. The receptor-binding affinity and protein stability of these AB5-type toxins appear to be the basis for their unique immunomodulatory properties. However, the toxicity of these molecules is a limiting factor for use as adjuvants in human vaccines. The non-toxic B subunit of CT, as well as chimeric CTA2B molecules, have shown recent promise as novel mucosal vaccines. A2B chimeras of CT retain the capacity to introduce antigens into host cells and modulate the immune response, but toxic domains are replaced with a vaccine antigen of interest. This work reveals the construction of a number of plasmids for the expression of ST1A2B chimeras containing the Yersinia pestis bacterial antigen, LcrV and the West Nile virus domain III (DIII) antigen. Plasmids were also constructed for expression of the ST1 B subunit and this pentamer was purified from the E.coli periplasm. The ability of the ST1 A2/B chimeras and STB to stimulate antigen uptake and immune stimulation in vitro was assayed by fluorescence microscopy, metabolic dye assay, T-cell proliferation assay and cytokine ELISA using both macrophage and dendritic cells. Findings suggest that STB can induce antigen uptake and may stimulate more of a Th-2 type and anti-inflammatory response, similar to CTB. These studies will contribute to the development of these toxins as novel mucosal adjuvants.