Yersinia ruckeri is the bacterial agent causing enteric redmouth disease (ERM) in rainbow trout leading to economic losses in intensive aquaculture. There are two main serovars and several minor groups based on O-antigens. The first goal of this thesis was to examine the difference between serotypes of Y. ruckeri in the course of infection in fish by applying in vivo bioluminescent imaging in rainbow trout (Oncorhynchus mykiss) and coho salmon (Oncorhynchus kisutch). In infection trials, the bioluminescent strains were infective, but the bioluminescent signal was not detected in fish infected with bacterial loads of 107 colony forming units per gram of kidney tissue. Skin and scales and the kidney blocked the luminescent signal emitted from the bacteria. The second goal of this thesis was to identify genetic markers which correlate with traditional O-antigen serotyping reactions. Using the sequences of genes which are part of the lipopolysaccharide biosynthetic pathway, oligonucleotide primers were designed to be complimentary to a fragment of wzx, the O-antigen flippase, and to wzy, the O-antigen polymerase, of serotype 1 Y. ruckeri strain RS 11. When these primers were used in polymerase chain reaction, an 1183 bp fragment of wzx and a 755 bp fragment of wzy were seen with DNA from 8 serovar 1 and 9 serovar 1a strains and not from other serovars identified by rabbit anti-sera agglutination. Southern blotting suggested there was little homology between serovar 1 wzx and wzy, and the same genes of the remaining serovars if present.

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This thesis project serves to fill experimental gaps needed to advance the goal of performing pre-clinical trials using an orthotopic rat glioblastoma model to evaluate the efficacy of high-frequency electroporation (H-FIRE) and QUAD-CTX tumor receptor-targeted cytotoxic conjugate therapies, individually and in combination, in selectively and thoroughly treating glioblastoma multiforme. In order to achieve this, an appropriate model must be developed and characterized. I have transduced F98 rat glioma cells to express red-shifted firefly luciferase, which will facilitate longitudinal tumor monitoring in vivo through bioluminescent imaging. I have characterized their response to H-FIRE relative to DI TNC1 rat astrocytes. I have demonstrated the presence of the molecular targets of QUAD in F98 cells. The in vitro characterization of this model has enabled preclinical studies of this promising glioblastoma therapy in an immunocompetent rat model, an important step before advancing ultimately to clinical human trials. / Master of Science / Treating glioblastoma multiforme (GBM), a form of cancer found in the brain, has not been very successful; patients rarely live two years following diagnosis, and there have been no major breakthrough advances in treatment to improve this outlook for decades. We have been working on two treatments which we hope to combine. The first is high-frequency electroporation (H-FIRE), which uses electrical pulses to kill GBM cells while leaving healthy cells alive and blood vessels intact. The second is QUAD-CTX, which combines a toxin with two types of protein that attach to other proteins that are more common on the surface of GBM cells than healthy cells. We have shown these to be effective at disproportionately killing human GBM cells growing in a lab setting. Before H-FIRE and QUAD-CTX may be tested on humans, we need to show them to be effective in an animal model, specifically rats. I have chosen rat glioma cells that will behave similarly to human GBM and a rat species that will not have an immune response to them. I have made these cells bioluminescent so that we may monitor the tumors as they grow and respond to our treatments. I have also shown that QUAD-CTX kills these rat glioma cells, as does H-FIRE. Because of this work, we are ready to begin testing these two treatments in rats.

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