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Separation and Comparison of Proteins From Virulent and Nonvirulent Strains of the Fish Pathogen Flavobacterium psychrophilum, Using a 2-D Electrophoretic Approach

e, but distinct proteins at lower molecular weight did not resolve as well or were less concentrated in the ATCC strain than the CSF strain. Two major proteins (p15 and p53) are represented in the virulent (CSF) strain but do not appear in the ATCC strain, while at least one protein (p37) appears to be differentially expressed in the ATCC strain.


Conclusions
The techniques outlined here demonstrate the effectiveness of sequential extraction in obtaining bacterial proteins from different compartments of the cell. Combined with 2-D PAGE techniques, the ReadyPrep sequential extraction kit allowed cell-associated and cytoplasmic proteins to be effectively separated. In addition to the development of techniques to separate different bacterial protein components, we have successfully separated proteins from both virulent and nonvirulent strains of F. psychrophilum by 2-D PAGE. The use of the PROTEAN IEF cell allowed rapid throughput of samples and exceptional control over focusing conditions. The separation of distinct proteins using these methods has allowed the comparison of potential virulence factors between two bacterial isolates grown under identical conditions. Further characterization is needed and will focus on comparison of differentially expressed proteins to active proteases (potential virulence components) and immunodominant bacterial antigens. Protein sequencing and eventual gene isolation may lead to the development of an efficacious recombinant vaccine for aquaculture, or may allow screening of the many F. psychrophilum isolates for potential vaccine candidates. This approach has the potential to substantially advance our current understanding of
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