Publication Date

12-2017

Date of Final Oral Examination (Defense)

10-24-2017

Type of Culminating Activity

Dissertation

Degree Title

Doctor of Philosophy in Biomolecular Sciences

Department

Biology

Major Advisor

Juliette Tinker, Ph.D.

Advisor

Kenneth A. Cornell, Ph.D.

Advisor

Julie Thom Oxford, Ph.D.

Advisor

Mark A. McGuire, Ph.D.

Abstract

Staphylococcus aureus is an important pathogen causing chronic and invasive disease worldwide. This bacterium is a leading cause of community and hospital acquired infections in humans, and is also known to infect wild and domestic animals. Bovine mastitis, or inflammation of the udder, is one of the most economically relevant diseases of the dairy industry, with a high incidence worldwide. S. aureus is a major etiological agent causing this disease. S. aureus mastitis is highly contagious and difficult or impossible to treat. Management practices at dairy farms, that include good sanitation and antibiotic use, have been partially successful in reducing the occurrence of this disease, however, a complete prevention or elimination is still to be achieved. Despite efforts over more than two decades, an effective vaccine for S. aureus mastitis, that can protect against heterologous strains of this bacterium, is not yet available. These efforts however, have improved our understanding of the pathogenicity, virulence factor expression and immune responses to this bacterium. Studies have indicated that there is significant intraspecies variability, and an effective vaccine against S. aureus will require the incorporation of multiple conserved and relevant antigens. Additionally, the route of vaccine administration and use of adjuvants to aid in antigen delivery and enhancement of immune responses will also be critical.

S. aureus contains a broad array of virulence factors required for colonization and disease, including: adhesins, toxins, a polysaccharide capsule, enzymes and immune evasion molecules, that are required for adhesion, invasion and colonization. Some virulence factors are highly conserved and centrally important for bacterial survival and sustenance, making them good vaccine targets. The iron-regulated surface determinant A (IsdA) and clumping factor A (ClfA) are two such conserved S. aureus extracellular-matrix adhesins that are promising vaccine antigens. However, an effective S. aureus vaccine for the prevention or reduction of mastitis will need to induce mucosal and systemic, as well as cellular and humoral, immune responses to these antigens. Bacterial enterotoxins are well characterized vaccine adjuvants that act by enhancing the mucosal delivery of antigens and promoting both systemic and mucosal humoral responses. Cholera toxin (CT), from Vibrio cholerae is a gold-standard vaccine adjuvant that can promote both humoral and cellular immune responses to co-administered antigens when delivered to mucosal surfaces.

The work presented here is based upon the overarching hypothesis that a mucosal, enterotoxin-based vaccine containing multiple relevant antigens will protect cows against S. aureus mastitis. To construct this vaccine, three immediate aims were developed. First, to ensure the incorporation of relevant antigens, the variability, genetic conservation and immunogenicity of the IsdA protein during bovine infection was determined. Second, a cholera toxin adjuvant based vaccine containing IsdA and a second antigen, ClfA (IsdA-CTA2/B +ClfA-CTA2/B) was used to vaccinate cows to determine immunogenicity. Lastly, a new immunoproteomics approach was used to identify immunogenic antigens for future incorporation into a multivalent vaccine. The results from these studies, as presented in chapters 2-4, indicate that: 1) the IsdA adhesin is expressed and conserved in bovine strains of S. aureus, 2) IsdA-CTA2/B + ClfA-CTA2/B can stimulate significant immune responses in vaccinated animals after intranasal administration and 3) immunoproteomics using milk antibodies can be used to identify new potential S. aureus vaccine antigens. The studies presented here will contribute to the advancement and understanding of staphylococcal vaccines in general, and specifically to those that will prevent bovine mastitis.

DOI

https://doi.org/10.18122/B26Q7C

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