Designer DNA Nanostructure Based Biosensing for Rapid COVID19 Detection and Monitoring using Saliva Sample

ABSTRACTA Novel Saliva-Based Aptamer Detection Assay for SARS-CoV-2 Infection(RFA-OD-20-021 STTR Application)Automated, rapid diagnostics with little sample collection and preparation are needed to identifyand trace affected persons in times when hyper-infectious pathogens cause pandemics. Frequent,low cost and highly scalable testing is the only way to gain visibility on the magnitude of thepandemic and ultimately control the spread of the disease. We propose the development of aunique system that can cheaply and readily detect SARS-CoV-2 in saliva samples. Thedevelopment of a system that uses saliva present an opportunity to readily test patients using asample that is easily collected and harbors high concentration of viral particles.The SARS-CoV-2 pandemic has predominantly affected individuals with pre-existing conditionssuch as clotting disorders, diabetes, hypertension or other chronic diseases. Patients with thesepre-existing conditions who then are infected have exacerbated symptoms and complications thatcan lead to death. For example, many patients that have succumbed to SARS-CoV-2 infectionhave developed blood clots that have impaired pulmonary or cardiac function and ultimatelycardiac failure. A rapid diagnostic using easily collected samples (e.g. saliva) would allow forinfections to be identified sooner, therapies to be administered quicker, treatment to be monitored,and ultimately leading to fewer individuals that succumb to the infection.We outline a novel DNA Star biosensing approach based on the fact that viruses, such as SARS-CoV-2, express unique spatial patterns of antigens on their surfaces, facilitating multivalentbinding to host cells for infection. These configurations of epitopes drive the high sensitivity andspecificity of our assay. Based on this naturally occurring binding mechanism, we developed arational design approach producing pattern matching designer DNA architecture for viral sensing.A proof-of-concept Dengue virus (DENV) rapid diagnostics was developed to demonstrate itspower: DENV surface antigens present the most complex geometric pattern among all knownpathogens, a DNA star linked 10-aptamers nanostructure that offers polyvalent, spatial DENV-epitope pattern matching interactions has provided high DENV-binding avidity and specificity,increasing affinity by ~1,000× compared to the conventional aptamer approach which relies onmonovalent aptamer-epitope interactions. Our POCT diagnostics detected intact DENV virions inpatient samples with PCR equivalent sensitivity in <2 mins at a cost <$0.15.Current RT-PCR molecular test are suited to large, centralized laboratories, and difficult to scalefor rapid testing of samples and delivery of results to clinicians and patients. Immunoassay testshave lower sensitivity, and patients need to develop a response to the virus in order to detect theantibody response. Our "DNA star" biosensor-based rapid diagnostics will provide theinfrastructure for real time SARS-CoV-2 diagnostics that is easy to use (instrument-free), faster(sample to results in minutes) and cost effective (~$3 per test).