Grant: Discovery Grants Program - Individual

It is estimated that there are 4 million cases of foodborne illness in Canada annually, resulting in more than 11,600 hospitalizations and 230 deaths every year. While food poisoning and foodborne illness can be caused by harmful chemicals, heavy metals, parasites, fungi, and viruses, the majority of food poisoning cases are due to bacterial pathogens, highlighting the importance for foodborne pathogenic bacteria detection and identification. However, the "ideal" technology that provides both rapid result turn-around and high sensitivity remains elusive as all prevalent foodborne pathogenic bacteria detection methods suffer from either lack of sufficient sensitivity or lengthy sample-to-result turn-around. The ultimate goal of the proposed research program is to develop a microchannel-based detection system that aims to reduce background noises, enhance target capturing efficiency and amplify detection signals to achieve rapid and sensitive detection of multiple foodborne pathogenic bacteria. To increase detection sensitivities, two different approaches are to be taken at the same time in this two-pronged design: a) reducing background noises and b) increasing detection signals. To reduce background noises, we propose to modify the inner walls of the PDMS microfluidic channels with PAMAM generation 7 (G7) that we have successfully used to render the PDMS surfaces nonfouling, thereby reducing detection background noises resulting from surface non-specific adsorptions. To increase detection signals, signaling rolling circle amplification (sRCA) will be employed to directly intensify detection signals by incorporating tandem repeat sequences that act as binding sites for fluorescent probes. In addition, capturing RCA (cRCA) will be used to enhance the target capturing efficiency, particularly at high flowrates, thus further intensifying the signals and increasing the sensitivities. The objectives of the proposal are: 1) Modifications of PDMS microfluidic channel by PAMAM dendrimers; 2) Optimizations of RCA reaction conditions; 3) Optimization of circular template spacer design to enhance the capturing performance of RCA generated multivalent system; 4) Control of flow patterns inside of the microfluidic channel; 5) Detection of target pathogenic bacteria and results validation. Novelty/Impact: This design is like no other. While certain components of the above-mentioned design have been reported in the literature, including in our own studies, the logical and innovative compilation of these components has not been previously reported. In the proposed study, formalin de-activated E.coli O157:H7 and Listeria monocytogenes - the two bacteria behind Walkerton and Maple Leaf Foods outbreaks, respectively - will be used as model bacteria for our studies. Every year, approximately 1 in 8 Canadians are affected by a foodborne illness, and the majority the illness is caused by foodborne bacterial pathogens.