Development of an infrared-functionalized microbalance sensor for Cyclospora cayetanensis detection and differentiation

Cyclospora cayetanensis (Cc) is a parasite which causes diarrheal illness in humans worldwide and is spread through contaminated food and water. Current procedures for Cc detection are expensive and time consuming. A detection system for Cc which is simple, fast, low-cost, and can be used in the field is needed. This proposal will test and develop a novel detection system for Cc by pairing infrared microscopy with cantilever-based microsensor technology. The sensing system will initially be developed using commercially available parasites and final testing stages will use oocysts of Cc. To determine if the sensing system has the sensitivity needed for testing produce and water samples, it will be compared to methods currently used for Cc detection. This project represents the first step toward producing a new tool which can be used by growers, processors, researchers, and testing laboratories to detect and quantify Cc quickly and cost-effectively. Such a tool could significantly improve our understanding of Cc risk and risk factor contributors and be used by growers, producers, and regulators to improve the safety of the fresh produce available to consumers.

Cyclospora cayetanensis (Cc) is a prevalent worldwide intestinal protozoan parasite of humans which is spread through contaminated food and water. Testing of food and water samples to understand the risk factors for Cc contamination is needed to limit transmission, develop control strategies, and improve food safety. Currently, microscopy and molecular techniques are used to detect Cc but are time intensive and require extensive sample preparation and personnel expertise. A method for Cc detection which is simple, fast, and low-cost with the potential for scalable implementation in the field would greatly enhance food safety. To build toward this goal, we will test and develop a sensing system which pairs infrared microscopy with cantilever-based microsensor technology to detect Cc. The initial design phase will employ commercially available protozoan parasites before testing the system on Cc oocysts from human samples. Comparisons between the sensitivity and specificity of the sensing system, microscopy, and PCR will be made to provide quantifiable measures of success of the platform. This project will provide the foundational data needed to further develop the sensing platform into a tool which can be used by growers, processors, Cc researchers, and testing laboratories to detect and quantify Cc quickly and cost-effectively. Such a tool could lead to significant improvements in understanding Cc risk and risk factor contributors, which can be used by growers, producers, and regulators to mitigate transmission risk and improve the safety of the fresh produce available to consumers.

1. Determine the physiochemical signatures of model organisms using a cantilever-based microbalance sensing system. 

2. Test the ability of the cantilever-based microbalance sensing system to accurately distinguish between the physiochemical signatures of multiple model organisms. 

3. Determine the physiochemical signature of Cyclospora on the cantilever-based microbalance sensing system. 

4. Test the ability of the cantilever-based microbalance sensing system to accurately distinguish the physiochemical signatures of Cyclospora from other protozoan parasites.

Through this year-long proof-of-concept project, we have developed a new sensing platform for the characterization of single parasite cells, providing information about their mass, their composition, and their fluorescence properties. The data collected to date suggests that the sensor produces signals in agreement with what is measured with conventional tools in cases that were selected to work on both platforms. This provides confidence that the signal recorded with the new platform on systems that are beyond the limit of detection of conventional tools can be trusted. Thus, an infrared‐functionalized microbalance sensor seems promising as a potential tool to improve C. cayetanensis detection and differentiation. Delivering the samples to the sensor was the most challenging aspect of the project, indicating that moving toward a microfluidic flow cell solution is necessary for a more comprehensive study on this topic. However, our results showing the different morphology, stiffness, and composition of the single parasite forms suggest that this will be a viable solution. While the best application of the technology may be in the identification of new contaminants, or detection of minute quantities of a sample, rather than in the continuous screening of large amounts of water, the strides made in the project are expected to steer new ideas of applications of these types of sensors for the detection of C. cayetanensis and related pathogens.