Graduate Thesis Or Dissertation
 

Chromatophores as cell-based biosensors for the detection of chemically and biologically toxic substances

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https://ir.library.oregonstate.edu/concern/graduate_thesis_or_dissertations/3x816r69f

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  • Cell-based biosensors are function-based detectors that use the physiological response of a living cell to sense biologically stimulating agents. This emerging technology extends the application of current detection methods by reporting on the toxicity of a sample and the potential to cause disease. Previously, Betta splendens erythrophores have been described as a method to detect toxic agents such as pesticides, chemicals, purified bacterial toxins and food-associated bacteria. The first objective of this study was to examine the B. splendens erythrophore response to Gram-negative food-associated bacteria by investigating the response to Salmonella typhimurium. Erythrophores aggregated in the presence of S. typhimurium in a growth-phase-dependent manner and this response was distinct from the media control. Additionally, it was found that erythrophore aggregation was dependent on the bacterial cell and erythrophores were not responsive to secreted bacterial products found in the culture supernatant. Transposon mutagenesis of S. typhimurium ATCC 700720 resulted in an interruption of the promoter region of operon prgHIJK. This insertion inactivated the expression of the SPI-1 T3SS structural proteins. The mutation resulted in a delayed erythrophore aggregation, implying that the SPI-1 T3SS is an important component contributing to erythrophore aggregation. The second objective of this study was to characterize the Oncorhynchus tshawytscha melanophore in terms of its responsiveness to aggregative and dispersive controls. It was hypothesized that the pigment response was biologically conserved between O. tshawytscha melanophores and B. splendens erythrophores. The data presented support this hypothesis and highlight potential applications. Melanophores aggregated in response to mercuric chloride and sodium arsenite but dispersed in the presence of ammonia. Additionally, melanophores aggregated in response to salmonid bacterial pathogens Aeromonas salmonicida, Yersinia ruckeri and Flavobacterium psychrophilum. Melanophores were capable of differentiating between healthy and diseased fish tissues. These results warrant a continued investigation into the full potential of the O. tshawytscha melanophore system. In conclusion, this study investigated two cell-based biosensor models; B. splendens erythrophores and O. tshawytscha melanophores. The data presented suggests that these models can be applied for the detection of foodborne bacterial pathogens and for the detection of chemical and bacterial contaminants pertinent to the aquaculture industry.
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