| Current detection methods for bacterial contamination rely on structure based detection of proteins and nucleic acids. While these methods are easy to use and reliable, they cannot evaluate the toxicity of a sample and the potential to cause disease. Previously, erythrophore cells derived from Betta splendens had been suggested as a method to detect toxic agents such as chemicals and toxins.;The work described here, investigated the potential of erythrophore cells as a detection method for the model food-associated pathogenic bacteria Salmonella enteritidis, Clostridium perfringens, Clostridium botulinum and Bacillus cereus. Erythrophore cell response to each bacterial pathogen was unique and could be distinguished from the bacterial growth medium. These results demonstrate that erythrophore cell response can be used to detect a set of pathogenic bacteria.;A second focus of this research effort was to test the hypothesis that erythrophore cells respond to pathogenic bacteria differently then nonpathogenic bacteria. To test this hypothesis, erythrophore cells were exposed to pathogenic and nonpathogenic Bacillus strains. Erythrophore cells failed to respond to Bacillus subtilis and a nonpathogenic Bacillus anthracis vaccine strain. Erythrophore cells responded to members of the B. cereus group including: B. cereus ATCC 10987, B. cereus UW85, B. cereus ATCC 14579, emetic B. cereus NCTC 11143, and Bacillus thuringiensis subsp. kurstaki BGSC 4D1.;A third focus of this research effort was to identify the bacterial virulence factor(s) responsible for inducing erythrophore cell response. Random chemical mutagenesis of B. cereus ATCC 49064 resulted in a single amino acid conversion of alanine to threonine at residue 289 of L2 toxin component of HBL. This mutation resulted in a delayed erythrophore cell response which was complemented with the wildtype hbl gene implementing the HBL toxin complex as a factor capable of inducing erythrophore cell response.;In conclusion, this study demonstrates that erythrophore cells can be optically monitored to detect several different bacterial pathogens. From this work, the utility of erythrophore cell response can be expanded from function based detection to include studying toxin activity as well as how pathogenic bacteria interact with eukaryotic cells. |
