Doctoral Dissertations

Date of Award

3-1986

Degree Type

Dissertation

Degree Name

Doctor of Philosophy

Major

Microbiology

Major Professor

Thomas C. Montie

Committee Members

Moore, Bemis, Rouse, Legendre

Abstract

The role of motility as a virulence factor in Pseudomonas aeruginosa burn wound sepsis was examined using mutants deficient in the Fla or Mot phenotype. Differences in physiological profiles, antibiograms, 0 antigen types, growth rates, and proteolytic activity were not observed in the Fla" and Mot" mutants, providing evidence for the similarity of these strains. Animal susceptibility studies showed that Fla" and Mot" mutants are significantly less virulent than the wildtype, parent strains by subcutaneous, topical, and intraperitoneal routes of challenge. Tissue colonization experiments revealed a characteristic, rapidly systemic infection in burned mice challenged with wildtype organisms. Nonmotile cells similarly proliferated in the burn wound, but the characteristic bacteremia and systemic invasion was markedly absent. The infection remained localized in the skin wound and the mice survived. Non-motile bacteria appeared in the liver following i.p. challenge at much higher levels than motile cells, suggesting a difference in the rate of clearance from the peritoneal cavity.

Significant protection against burn wound sepsis was achieved with flagellar antisera. Cross-protection experiments revealed that protection was not only H antigen dependent but specific for the flagella antigen type. Antiserum raised against a-type flagellin would only protect against homologous bacterial challenge and not against b-type flagellated strains, and vice versa. The specificity of flagellar antisera was also demonstrated in cross-agglutination assays, motility inhibition, Western blot analyses, and an ELISA. Reactivity was not detected against somatic antigens.

Motile bacteria colonized the burn wounds of passively-protected mice to similar levels as seen in non-protected animals, but remained localized and did not cause systemic infection, a pattern remarkably similar to infections with mutants deficient in motility. Animals rendered neutropenic were not protected with flagellar antisera, indicating an important role for phagocytic cells in protection. Microscopic studies revealed that motility is directly inhibited by a specific action of IgG antibody with the flagellum. Antiflagellar antibody is hypothesized as exerting its protective capacity by (1) inhibiting the motility of invading bacteria by a cross-linking, agglutination reaction and (2) acting as an opsonin, targeting immobilized cells for phagocytosis.

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