Development and Characterization of Nanocomposite Materials for the Detection of Pore-Forming Toxins

Motivation. Despite stringent quality control and manufacturing procedures in the U.S. food industry, food-related disease remains a major problem with an estimated 76 million cases each year. Many pathogenic bacteria such as Listeria monocytogenes, Bacillus cereus, and Clostridium perfringens produce and release pore-forming toxins (PFTs) that play a significant role in the mechanism of the human illness. These bacteria and their toxins represent a serious threat to the safety of our food supply and are therefore an important detection target for new biosensors. New biosensors alone, however, are not sufficient. Rather new, robust, inexpensive, fast, and selective biosensors are necessary to overcome the practical and economic hurdles within the food supply system and to make a direct impact on human health and food biosecurity. Biological Inspiration. Biology holds much inspiration for the design of advanced biosensor materials. Living cells are far superior detectors of their chemical and biological environment than are most synthetic biosensor materials. Whole cell biosensors harness this capability but are impractical in many applications because of the fragility and growth requirements of living cells. On the other extreme, single biomolecule-based biosensors, can be made robust, but lack the biological functionality that leads to advanced performance. We suggest that more optimal engineered materials are likely to reside between these two extremes. Through the recreation of relevant biological functionality in nonliving, multi-component, hybrid materials, we propose to develop superior biosensors that rival cells in their detection capabilities, but are robust and compatible with engineered systems. Objectives. We propose the development of a nanocomposite material in the form of immobilized liposomes as \"artificial cells\" that can capture the functional essence of whole cell assays in a nonliving hybrid (inorganic / organic / biological) system. We will directly apply these materials to the development of robust, inexpensive, selective and fast biosensors for the detection of pore-forming toxins (PFTs) from foodborne pathogens. The overall objective of this work is to define and quantify the fundamental interactions that govern the performance of immobilized artificial cells or liposomes as a biosensor material for the detection of PFTs. We will use listeriolysin O (LLO) from Listeria monocytogenes as a model protein. The outcome of this work will enable the rational design of an optimized platform of nanocomposite materials for the selective detection of the broad class of pathogenic pore-forming toxins.