DETECTION OF ESCHERICHIA COLI O157:H7 WITH GOLD NANOPARTICLES SANDWICH HYBRIDIZATION METHOD

DETECTION OF ESCHERICHIA COLI O157:H7 WITH GOLD NANOPARTICLES SANDWICH HYBRIDIZATION METHOD

By Jingjing Shen

Thesis Advisor: Dr. Vivian C.H. Wu

An Abstract of the Thesis Presented

in Partial Fulfillment of the Requirements for the

Degree of Master of Science

(in Food Science and Human Nutrition)

August, 2010

The epidemiology of foodborne pathogen related diseases is becoming recognized as new pathogens emerge and known pathogens increase in prevalence or become associated with new food vehicles. For example, Escherichia coli O157:H7 infection is a major cause of hemolytic uremic syndrome, which is the most common cause of acute kidney failure in children in the United States. Protecting the public from microbial contamination of the food supply has become a top priority of the U.S. Food and Drug Administration (FDA). E. coliO157:H7 has been a major foodborne pathogen associated with numerous cases of fatal foodborne diseases. Therefore, there is a great need to develop an efficient method to detect this bacterium.

Our research aims at developing a highly efficient, sensitive and applicable protocol for rapid detection of E. coli O157:H7 in food samples through the observation of color change. The detection method is based on aggregation of DNA-functionalized gold nanoparticles due to the reactions among DNAs. The aggregation is controlled by different conditions such as temperature and salt concentration of the reaction. In this study, optimizations were conducted on experimental conditions so as to achieve good sensitivity. Detection results can be read via observing color change of reaction solution by naked eyes.

In conclusion, we have developed a highly sensitive method for optical detection of E. coli O157:H7 gene eaeA based on AuNPs sandwich hybridization. The developed assay is easy to operate and needs little experimental instruments. It showed good sensitivity for detection of E. coli O157:H7 in pure culture, blueberries, ground beef and spinach samples. Experimental results suggest that the assay has great potential for further applications in detection of E. coli O157:H7 as well as other organisms or the entire community of environmental and food microorganisms by selectively controlling of experimental conditions. The work lays the groundwork for commercialization of diagnostic kit for rapid detection in food samples that have been contaminated by pathogenic organisms.