Lab-on-a-Chip Designs for Airborne Spore Detection: Towards the Forecasting of Sclerotinia Stem Rot of Canola
Creators: Duarte Riveros, Pedro
Publisher: name: University of Alberta Library
Year: 2022

<p>Sclerotinia stem rot (SSR), caused by the necrotrophic fungal pathogen Sclerotinia sclerotiorum, is one of the most devastating diseases affecting crops. More than 400 plant species around the globe are affected by this fungus. In canola, one of Canada’s most important crops, yield losses due to SSR can be as high as 50%. Although chemical control with fungicides is currently the most common tool for the management of SSR, the routine application without prior information about the risk of the disease development is also financially inefficient. The early prognosis of an outbreak is critical to avoid the severe economic losses caused by SSR and can be achieved by the detection of a small number of S. sclerotiorum airborne ascospores, one of the main agents of infection in stem rot. However, the current lack of simple and effective methods to detect fungal airborne pathogens has hindered the development of an accurate early warning system.<br>
In this thesis, we explored the design and development of lab-on-a-chip devices for the detection of S. sclerotiorum ascospores, aiming at their future integration with spore-trap samplers into an effective SSR forecasting system. Our first design is based on a Coulter counter approach, which consists of a microfluidic chip capable of quantifying single ascospores flowing in a microchannel. The target ascospores are injected into the device and selectively captured by dielectrophoresis, while other particles in the sample are flushed away to the outlet drain of the device. Subsequently, the target ascospores are released into the flow stream of the device and are detected when flowing through a constriction employing dynamic impedimetric sensing. Experimental results indicated a 0.3% change in the impedance signal produced by individual ascospores, which were detected using a benchtop potentiostat.<br>
In our second approach, we developed a microfluidic device that contains a nano-thick aluminum electrode structure integrated with a picoliter well array for dielectrophoresis-driven capture of ascospores and on-chip quantitative detection employing static impedimetric sensing. Based on experimental results, we demonstrated a highly efficient ascospore trapping rate of more than 90% with an effective impedimetric sensing method that allowed the ascospore quantification of each column in the array and achieved a sensitivity of 2%/ascospore at 5 kHz and 1.6%/ascospore at 20 kHz, enabling single ascospore detection.<br>
Finally, the dielectric properties of S. sclerotiorum ascospores were determined experimentally employing a microfluidic platform containing interdigitated aluminum microelectrodes. The dielectric properties of ascospores are of major importance for the development of dielectrophoretic filters, as it provides information about the dielectrophoretic response of the ascospores without the need for long iterative testing. The dielectric properties of ascospores were determined in media of different conductivities and they were modeled using a realistic ellipsoidal double-shell model based on the multi-shell theory. To validate the methodology and analysis, the dielectric properties of human embryonic kidney (HEK-293) cells were also determined and compared to values reported in the literature.<br>
We envision that the devices proposed in this thesis will contribute to the development of a low-cost, miniaturized, and automated platform technology that can be integrated into an infectious plant disease forecasting tool for canola crop protection.</p>