Abstract

We propose to improve the sensitivity and reduce the cost of detecting pathogenic organisms carried by ticks and mosquitoes in Detroit by developing a new technology platform, ddLAMP molecular amplification. Current antigen- and PCR-based techniques to detect Lyme disease spirochetes in ticks and West Nile virus in mosquitoes are less sensitive than desired, likely resulting in numerous false negatives. We propose to develop ddLAMP, a faster isothermal amplification technology than PCR, and implement it on a microfluidics platform, designed and built in Wayne State's Nanofabrication facility. We will interface with ongoing tick and mosquito collection by the Detroit Health Department in Palmer Park and elsewhere in the City of Detroit to apply the new ddLAMP platform to sensitive detection of these disease organisms. This project will synergize with development of a bacteria-focused ddLAMP assay system that we are developing (see support letter from the Great Lakes Water Authority, which will fund that work) and will also complement a recent award from the Sharon L. Ram Aquatic Science Fund to Adrian Vasquez to study mosquitoes in the diets of water mites in Palmer Park. Innovations created by this project include the demonstration of ddLAMP capabilities on the BioRad digital droplet system and its much faster "time to droplet counting" than ddPCR. Furthermore, transformation of the ddLAMP process to a microfluidics platform will create further innovations including more than double the droplets generated by BioRad per sample in less time, an innovative cell phone-based imaging system, and application of recently developed GPU accelerated algorithms for rapid image analysis, leading to Poisson statistical analysis of the droplet results. As a result we expect to achieve detection results faster and less expensively than current methods. The ddLAMP platform will also be applicable to detection of other pathogens (Eastern equine encephalitis, Zika virus, SARS-CoV-2, etc) and to detection of diet sources of these organisms (e.g. measurement of how much human and animal [and which animal] blood may be present in ticks or mosquitoes), which may be useful in assessment of human health risk.