STTR Phase I: A Novel Iothermal CRISPR-Cas12a Platform for the Detection of Viruses

The broader impact/commercial potential of this Small Business Technology Transfer (STTR) Phase I project is the creation of a reliable, highly sensitive, cost-effective, and easy-to-implement congenital cytomegalovirus (cCMV) screening that can be run with other newborn screening tests. cCMV affects about 25,000 babies born in the U.S. each year with moderate to severe long-term impairment (hearing loss and delayed neurological development) being diagnosed in almost 25% of infected children with an overall mortality of 0.5%. Efforts to address cCMV are complicated by the fact that 78-92% of cases show no symptoms at birth and despite its prevalence, cCMV is not universally screened largely due to limitations in current diagnostic workflows. Early detection and treatment within the first 72 hours of life greatly reduces lifelong disabilities and healthcare costs by approximately $1.3M per case. By providing a cost-effective, rapid, and scalable diagnostic tool that meets clinical needs, this solution will enable universal newborn screening for cCMV of the 3.6M live births that happen in the U.S. every year. Future extensions will adapt the platform to detect additional pathogens, broadening its public health impact through detecting infectious agents such as SARS-CoV-2, Epstein-Barr Virus, Herpes Simplex Virus type 1, and Human Herpesvirus 6B. This Small Business Technology Transfer (STTR) Phase I project aims to develop a one-step molecular diagnostic that employs Clustered Regularly Interspaced Short Palindromic Repeats-associated protein 12a (CRISPR-Cas12a) to identify cCMV directly from newborn dried blood spots. Current CMV Polymerase Chain Reaction (PCR) based diagnostics suffer from significant limitations, including low sensitivity (~73%), long turnaround times (24–48 hours), and reliance on off-site processing. This project aims to improve CMV diagnostics by offering a cost-effective, highly sensitive (>90%) solution with a rapid turnaround time of 40 minutes-2 hours. This project will streamline the testing workflow through a simplified, single-reaction “One-Pot” assay compatible with point-of-care settings, achieving results in under 30 minutes at ambient temperature. Additionally, all infrastructure for the assay is already present in clinical settings. Phase I will establish the feasibility of this platform as a universal screening tool for cCMV by addressing two primary areas of technical risk: system performance and workflow suitability. Objectives include to: 1) optimize the diagnostic to improve sensitivity and specificity to commercially sufficient levels and 2) simplify workflow by creating a single-reaction assay. This aims to prove the value of the platform as a whole and support expansion into other infectious agent detection spaces. This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.